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Biochemical Pharmacology (v.83, #10)
Antioxidants for prostate cancer chemoprevention: Challenges and opportunities
by Dinesh Thapa; Rita Ghosh (pp. 1319-1330).
Extensive research has led to the firm conclusion that antioxidants protect cells from damage caused by oxidative stress and its associated pathological conditions including inflammation. It has also been established that inflammation is a precursor in neoplastic transformation of the prostate. Although, a vast body of experimental and clinical evidence shows efficacy of antioxidants as preventive strategies for prostate cancer, there is a lack of consistent agreement in outcomes especially from recent large-scale randomized clinical trials. Despite these concerns, our understanding of the preventive mechanisms as well as clinical efficacy and safety data indicate that novel antioxidant therapeutics still hold great promise for prostate cancer chemoprevention. We propose that for effective use of antioxidants for prostate cancer prevention, further high impact translational research is needed with special attention on selecting those patients who will benefit from such intervention. Therefore, it is important to validate predictive biomarkers from successful trials and combine this with knowledge of preclinical characterization of antioxidants (and combinations) that will eventually facilitate the development of ‘personalized prostate cancer chemoprevention’. In this review, we briefly describe some common and emerging antioxidants that have shown benefits in preclinical and clinical settings. Above all, we focus on summarizing the progress we made thus far in prostate cancer chemoprevention using antioxidants, the heightened interest and challenges in the future.
Keywords: Abbreviations; 8-oxo-dG; 8-oxo-2′-deoxyguanosine; ADT; androgen-deprivation therapy; AR; androgen receptor; ARE; antioxidant response element; ATBC; Alpha-Tocopherol, Beta-Carotene; BPH; benign prostatic hyperplasia; CARET; Carotene and Retinol Efficacy Trial; CRPC; castration-resistant prostate cancer; DRE; digital rectal exam; GPx; glutathione peroxidase; GSR; glutathione reductase; HGPIN; high-grade prostatic intraepithelial neoplasia; JPHC; Japan Public Health Center; NPC; Nutritional Prevention of Cancer; NQO; NADPH: quinone oxidoreductase; PCNA; proliferating cell nuclear antigen; PHS; Physicians’ Health Study; PIA; proliferative inflammatory atrophy; PIN; prostatic intraepithelial neoplasia; PKC; protein kinase C; PSA; prostate-specific antigen; RNS; reactive nitrogen species; ROS; reactive oxygen species; SELECT; Selenium and Vitamin E Cancer Prevention Trial; SOD; superoxide dismutase; SU.VI.MAX; supplementation en vitamines et minerauxantioxydants; SWOG; Southwest Oncology Group; TRAMP; transgenic adenocarcinoma of the mouse prostate; TrxR; thioredoxin reductaseAntioxidants; Chemoprevention; Prostate cancer; ROS; Oxidative stress
Antioxidants for prostate cancer chemoprevention: Challenges and opportunities
by Dinesh Thapa; Rita Ghosh (pp. 1319-1330).
Extensive research has led to the firm conclusion that antioxidants protect cells from damage caused by oxidative stress and its associated pathological conditions including inflammation. It has also been established that inflammation is a precursor in neoplastic transformation of the prostate. Although, a vast body of experimental and clinical evidence shows efficacy of antioxidants as preventive strategies for prostate cancer, there is a lack of consistent agreement in outcomes especially from recent large-scale randomized clinical trials. Despite these concerns, our understanding of the preventive mechanisms as well as clinical efficacy and safety data indicate that novel antioxidant therapeutics still hold great promise for prostate cancer chemoprevention. We propose that for effective use of antioxidants for prostate cancer prevention, further high impact translational research is needed with special attention on selecting those patients who will benefit from such intervention. Therefore, it is important to validate predictive biomarkers from successful trials and combine this with knowledge of preclinical characterization of antioxidants (and combinations) that will eventually facilitate the development of ‘personalized prostate cancer chemoprevention’. In this review, we briefly describe some common and emerging antioxidants that have shown benefits in preclinical and clinical settings. Above all, we focus on summarizing the progress we made thus far in prostate cancer chemoprevention using antioxidants, the heightened interest and challenges in the future.
Keywords: Abbreviations; 8-oxo-dG; 8-oxo-2′-deoxyguanosine; ADT; androgen-deprivation therapy; AR; androgen receptor; ARE; antioxidant response element; ATBC; Alpha-Tocopherol, Beta-Carotene; BPH; benign prostatic hyperplasia; CARET; Carotene and Retinol Efficacy Trial; CRPC; castration-resistant prostate cancer; DRE; digital rectal exam; GPx; glutathione peroxidase; GSR; glutathione reductase; HGPIN; high-grade prostatic intraepithelial neoplasia; JPHC; Japan Public Health Center; NPC; Nutritional Prevention of Cancer; NQO; NADPH: quinone oxidoreductase; PCNA; proliferating cell nuclear antigen; PHS; Physicians’ Health Study; PIA; proliferative inflammatory atrophy; PIN; prostatic intraepithelial neoplasia; PKC; protein kinase C; PSA; prostate-specific antigen; RNS; reactive nitrogen species; ROS; reactive oxygen species; SELECT; Selenium and Vitamin E Cancer Prevention Trial; SOD; superoxide dismutase; SU.VI.MAX; supplementation en vitamines et minerauxantioxydants; SWOG; Southwest Oncology Group; TRAMP; transgenic adenocarcinoma of the mouse prostate; TrxR; thioredoxin reductaseAntioxidants; Chemoprevention; Prostate cancer; ROS; Oxidative stress
Antioxidants for prostate cancer chemoprevention: Challenges and opportunities
by Dinesh Thapa; Rita Ghosh (pp. 1319-1330).
Extensive research has led to the firm conclusion that antioxidants protect cells from damage caused by oxidative stress and its associated pathological conditions including inflammation. It has also been established that inflammation is a precursor in neoplastic transformation of the prostate. Although, a vast body of experimental and clinical evidence shows efficacy of antioxidants as preventive strategies for prostate cancer, there is a lack of consistent agreement in outcomes especially from recent large-scale randomized clinical trials. Despite these concerns, our understanding of the preventive mechanisms as well as clinical efficacy and safety data indicate that novel antioxidant therapeutics still hold great promise for prostate cancer chemoprevention. We propose that for effective use of antioxidants for prostate cancer prevention, further high impact translational research is needed with special attention on selecting those patients who will benefit from such intervention. Therefore, it is important to validate predictive biomarkers from successful trials and combine this with knowledge of preclinical characterization of antioxidants (and combinations) that will eventually facilitate the development of ‘personalized prostate cancer chemoprevention’. In this review, we briefly describe some common and emerging antioxidants that have shown benefits in preclinical and clinical settings. Above all, we focus on summarizing the progress we made thus far in prostate cancer chemoprevention using antioxidants, the heightened interest and challenges in the future.
Keywords: Abbreviations; 8-oxo-dG; 8-oxo-2′-deoxyguanosine; ADT; androgen-deprivation therapy; AR; androgen receptor; ARE; antioxidant response element; ATBC; Alpha-Tocopherol, Beta-Carotene; BPH; benign prostatic hyperplasia; CARET; Carotene and Retinol Efficacy Trial; CRPC; castration-resistant prostate cancer; DRE; digital rectal exam; GPx; glutathione peroxidase; GSR; glutathione reductase; HGPIN; high-grade prostatic intraepithelial neoplasia; JPHC; Japan Public Health Center; NPC; Nutritional Prevention of Cancer; NQO; NADPH: quinone oxidoreductase; PCNA; proliferating cell nuclear antigen; PHS; Physicians’ Health Study; PIA; proliferative inflammatory atrophy; PIN; prostatic intraepithelial neoplasia; PKC; protein kinase C; PSA; prostate-specific antigen; RNS; reactive nitrogen species; ROS; reactive oxygen species; SELECT; Selenium and Vitamin E Cancer Prevention Trial; SOD; superoxide dismutase; SU.VI.MAX; supplementation en vitamines et minerauxantioxydants; SWOG; Southwest Oncology Group; TRAMP; transgenic adenocarcinoma of the mouse prostate; TrxR; thioredoxin reductaseAntioxidants; Chemoprevention; Prostate cancer; ROS; Oxidative stress
Dog bites man or man bites dog? The enigma of the amino acid conjugations
by Diren Beyoğlu; Robert L. Smith; Jeffrey R. Idle (pp. 1331-1339).
The proposition posed is that the value of amino acid conjugation to the organism is not, as in the traditional view, to use amino acids for the detoxication of aromatic acids. Rather, the converse is more likely, to use aromatic acids that originate from the diet and gut microbiota to assist in the regulation of body stores of amino acids, such as glycine, glutamate, and, in certain invertebrates, arginine, that are key neurotransmitters in the central nervous system (CNS). As such, the amino acid conjugations are not so much detoxication reactions, rather they are homeostatic and neuroregulatory processes. Experimental data have been culled in support of this hypothesis from a broad range of scientific and clinical literature. Such data include the low detoxication value of amino acid conjugations and the Janus nature of certain amino acids that are both neurotransmitters and apparent conjugating agents. Amino acid scavenging mechanisms in blood deplete brain amino acids. Amino acids glutamate and glycine when trafficked from brain are metabolized to conjugates of aromatic acids in hepatic mitochondria and then irreversibly excreted into urine. This process is used clinically to deplete excess nitrogen in cases of urea cycle enzymopathies through excretion of glycine or glutamine as their aromatic acid conjugates. Untoward effects of high-dose phenylacetic acid surround CNS toxicity. There appears to be a relationship between extent of glycine scavenging by benzoic acid and psychomotor function. Glycine and glutamine scavenging by conjugation with aromatic acids may have important psychosomatic consequences that link diet to health, wellbeing, and disease.
Keywords: Abbreviations; BA; benzoic acid; PAA; phenylacetic acid; HA; hippuric acid; GLY; glycine; GLN; glutamine; TAU; taurine; GLU; glutamic acid; EAA; excitatory amino acid; IAA; inhibitory amino acid; EAAT; excitatory amino acid transporter; ECF; extracellular fluid; CSF; cerebrospinal fluid; PAGLN; phenacetylglutamine; PAGLY; phenacetylglycine; PATAU; phenacetyltaurine; 4HBA; 4-hydroxybenzoic acid); 4HHA; 4-hydroxyhippuric acid; 2FA; 2-furoic acid; 2FGLY; 2-furoylglycine; 3IAA; 3-indolylacrylic acid; IAG; 3-indolylacryloylglycine; ECT; electroconvulsive therapy; NMDAR; N; -methyl-; d; -aspartate receptor; mGluR; metabotropic glutamate receptor; CNS; central nervous system; BBB; blood-brain barrier; HPLC; high-performance liquid chromatography; LPI; lysinuric protein intolerance; NaPBA; sodium phenylbutyric acid; GPB; glyceryltri(4-phenylbutyrate); 4NB; 4-nitrobenzoic acid; 4ABA; 4-aminobenzoic acid; ARG; arginineAmino acid conjugation; Neuroregulation; Amino acid scavenging; Hippuric acid; Phenacetylglutamine
Dog bites man or man bites dog? The enigma of the amino acid conjugations
by Diren Beyoğlu; Robert L. Smith; Jeffrey R. Idle (pp. 1331-1339).
The proposition posed is that the value of amino acid conjugation to the organism is not, as in the traditional view, to use amino acids for the detoxication of aromatic acids. Rather, the converse is more likely, to use aromatic acids that originate from the diet and gut microbiota to assist in the regulation of body stores of amino acids, such as glycine, glutamate, and, in certain invertebrates, arginine, that are key neurotransmitters in the central nervous system (CNS). As such, the amino acid conjugations are not so much detoxication reactions, rather they are homeostatic and neuroregulatory processes. Experimental data have been culled in support of this hypothesis from a broad range of scientific and clinical literature. Such data include the low detoxication value of amino acid conjugations and the Janus nature of certain amino acids that are both neurotransmitters and apparent conjugating agents. Amino acid scavenging mechanisms in blood deplete brain amino acids. Amino acids glutamate and glycine when trafficked from brain are metabolized to conjugates of aromatic acids in hepatic mitochondria and then irreversibly excreted into urine. This process is used clinically to deplete excess nitrogen in cases of urea cycle enzymopathies through excretion of glycine or glutamine as their aromatic acid conjugates. Untoward effects of high-dose phenylacetic acid surround CNS toxicity. There appears to be a relationship between extent of glycine scavenging by benzoic acid and psychomotor function. Glycine and glutamine scavenging by conjugation with aromatic acids may have important psychosomatic consequences that link diet to health, wellbeing, and disease.
Keywords: Abbreviations; BA; benzoic acid; PAA; phenylacetic acid; HA; hippuric acid; GLY; glycine; GLN; glutamine; TAU; taurine; GLU; glutamic acid; EAA; excitatory amino acid; IAA; inhibitory amino acid; EAAT; excitatory amino acid transporter; ECF; extracellular fluid; CSF; cerebrospinal fluid; PAGLN; phenacetylglutamine; PAGLY; phenacetylglycine; PATAU; phenacetyltaurine; 4HBA; 4-hydroxybenzoic acid); 4HHA; 4-hydroxyhippuric acid; 2FA; 2-furoic acid; 2FGLY; 2-furoylglycine; 3IAA; 3-indolylacrylic acid; IAG; 3-indolylacryloylglycine; ECT; electroconvulsive therapy; NMDAR; N; -methyl-; d; -aspartate receptor; mGluR; metabotropic glutamate receptor; CNS; central nervous system; BBB; blood-brain barrier; HPLC; high-performance liquid chromatography; LPI; lysinuric protein intolerance; NaPBA; sodium phenylbutyric acid; GPB; glyceryltri(4-phenylbutyrate); 4NB; 4-nitrobenzoic acid; 4ABA; 4-aminobenzoic acid; ARG; arginineAmino acid conjugation; Neuroregulation; Amino acid scavenging; Hippuric acid; Phenacetylglutamine
Dog bites man or man bites dog? The enigma of the amino acid conjugations
by Diren Beyoğlu; Robert L. Smith; Jeffrey R. Idle (pp. 1331-1339).
The proposition posed is that the value of amino acid conjugation to the organism is not, as in the traditional view, to use amino acids for the detoxication of aromatic acids. Rather, the converse is more likely, to use aromatic acids that originate from the diet and gut microbiota to assist in the regulation of body stores of amino acids, such as glycine, glutamate, and, in certain invertebrates, arginine, that are key neurotransmitters in the central nervous system (CNS). As such, the amino acid conjugations are not so much detoxication reactions, rather they are homeostatic and neuroregulatory processes. Experimental data have been culled in support of this hypothesis from a broad range of scientific and clinical literature. Such data include the low detoxication value of amino acid conjugations and the Janus nature of certain amino acids that are both neurotransmitters and apparent conjugating agents. Amino acid scavenging mechanisms in blood deplete brain amino acids. Amino acids glutamate and glycine when trafficked from brain are metabolized to conjugates of aromatic acids in hepatic mitochondria and then irreversibly excreted into urine. This process is used clinically to deplete excess nitrogen in cases of urea cycle enzymopathies through excretion of glycine or glutamine as their aromatic acid conjugates. Untoward effects of high-dose phenylacetic acid surround CNS toxicity. There appears to be a relationship between extent of glycine scavenging by benzoic acid and psychomotor function. Glycine and glutamine scavenging by conjugation with aromatic acids may have important psychosomatic consequences that link diet to health, wellbeing, and disease.
Keywords: Abbreviations; BA; benzoic acid; PAA; phenylacetic acid; HA; hippuric acid; GLY; glycine; GLN; glutamine; TAU; taurine; GLU; glutamic acid; EAA; excitatory amino acid; IAA; inhibitory amino acid; EAAT; excitatory amino acid transporter; ECF; extracellular fluid; CSF; cerebrospinal fluid; PAGLN; phenacetylglutamine; PAGLY; phenacetylglycine; PATAU; phenacetyltaurine; 4HBA; 4-hydroxybenzoic acid); 4HHA; 4-hydroxyhippuric acid; 2FA; 2-furoic acid; 2FGLY; 2-furoylglycine; 3IAA; 3-indolylacrylic acid; IAG; 3-indolylacryloylglycine; ECT; electroconvulsive therapy; NMDAR; N; -methyl-; d; -aspartate receptor; mGluR; metabotropic glutamate receptor; CNS; central nervous system; BBB; blood-brain barrier; HPLC; high-performance liquid chromatography; LPI; lysinuric protein intolerance; NaPBA; sodium phenylbutyric acid; GPB; glyceryltri(4-phenylbutyrate); 4NB; 4-nitrobenzoic acid; 4ABA; 4-aminobenzoic acid; ARG; arginineAmino acid conjugation; Neuroregulation; Amino acid scavenging; Hippuric acid; Phenacetylglutamine
Inhibition of the PI3K-Akt signaling pathway disrupts ABCG2-rich extracellular vesicles and overcomes multidrug resistance in breast cancer cells
by Vicky Goler-Baron; Irina Sladkevich; Yehuda G. Assaraf (pp. 1340-1348).
Extracellular vesicles (EVs) formed between neighbor breast cancer cells mediate ABCG2-dependent multidrug resistance (MDR) via intravesicular drug sequestration. Inhibition of Akt signaling abolishes ABCG2 targeting, EVs biogenesis and overcomes MDR.We have recently shown that ABCG2-rich extracellular vesicles (EVs) form between neighbor breast cancer cells and actively concentrate various chemotherapeutics, resulting in multidrug resistance (MDR). Here we studied the signaling pathway regulating ABCG2 targeting to EVs as its inhibition would overcome MDR. The PI3K-Akt signaling pathway was possibly implicated in subcellular localization of ABCG2; we accordingly show here that pharmacological inhibition of Akt signaling results in gradual re-localization of ABCG2 from the EVs membrane to the cytoplasm. Cytoskeletal markers including β-actin and the tight junction protein ZO-1, along with the EVs markers ABCG2 and Ezrin–Radixin–Moesin revealed that this intracellular ABCG2 retention leads to gradual decrease in the size and number of EVs, resulting in EVs elimination and complete reversal of MDR. Inhibition of Akt signaling restored drug sensitivity to mitoxantrone and topotecan, bona fide ABCG2 transport substrates, hence being equivalent to MDR reversal achieved with the ABCG2 transport inhibitor Ko143. Remarkably, apart from loss of ABCG2 transport activity, treatment of MCF-7/MR cells with Ko143 resulted in cytoplasmic re-localization of ABCG2, similarly to the phenotype observed after Akt inhibition. We conclude that the PI3K-Akt signaling pathway is a key regulator of subcellular localization of ABCG2, EVs biogenesis and functional MDR. Furthermore, proper folding of ABCG2 and its targeting to the EVs membrane are crucial components of the biogenesis of EVs and their MDR function. We propose that Akt signaling inhibitors which disrupt ABCG2 targeting and EVs biogenesis may readily overcome MDR thus warranting in vivo studies with these promising drug combinations.
Keywords: Extracellular vesicles (EVs); PI3K-Akt signaling; Chemotherapy; ABC transporters; Overcoming multidrug resistance (MDR); Breast cancer
Inhibition of the PI3K-Akt signaling pathway disrupts ABCG2-rich extracellular vesicles and overcomes multidrug resistance in breast cancer cells
by Vicky Goler-Baron; Irina Sladkevich; Yehuda G. Assaraf (pp. 1340-1348).
Extracellular vesicles (EVs) formed between neighbor breast cancer cells mediate ABCG2-dependent multidrug resistance (MDR) via intravesicular drug sequestration. Inhibition of Akt signaling abolishes ABCG2 targeting, EVs biogenesis and overcomes MDR.We have recently shown that ABCG2-rich extracellular vesicles (EVs) form between neighbor breast cancer cells and actively concentrate various chemotherapeutics, resulting in multidrug resistance (MDR). Here we studied the signaling pathway regulating ABCG2 targeting to EVs as its inhibition would overcome MDR. The PI3K-Akt signaling pathway was possibly implicated in subcellular localization of ABCG2; we accordingly show here that pharmacological inhibition of Akt signaling results in gradual re-localization of ABCG2 from the EVs membrane to the cytoplasm. Cytoskeletal markers including β-actin and the tight junction protein ZO-1, along with the EVs markers ABCG2 and Ezrin–Radixin–Moesin revealed that this intracellular ABCG2 retention leads to gradual decrease in the size and number of EVs, resulting in EVs elimination and complete reversal of MDR. Inhibition of Akt signaling restored drug sensitivity to mitoxantrone and topotecan, bona fide ABCG2 transport substrates, hence being equivalent to MDR reversal achieved with the ABCG2 transport inhibitor Ko143. Remarkably, apart from loss of ABCG2 transport activity, treatment of MCF-7/MR cells with Ko143 resulted in cytoplasmic re-localization of ABCG2, similarly to the phenotype observed after Akt inhibition. We conclude that the PI3K-Akt signaling pathway is a key regulator of subcellular localization of ABCG2, EVs biogenesis and functional MDR. Furthermore, proper folding of ABCG2 and its targeting to the EVs membrane are crucial components of the biogenesis of EVs and their MDR function. We propose that Akt signaling inhibitors which disrupt ABCG2 targeting and EVs biogenesis may readily overcome MDR thus warranting in vivo studies with these promising drug combinations.
Keywords: Extracellular vesicles (EVs); PI3K-Akt signaling; Chemotherapy; ABC transporters; Overcoming multidrug resistance (MDR); Breast cancer
Inhibition of the PI3K-Akt signaling pathway disrupts ABCG2-rich extracellular vesicles and overcomes multidrug resistance in breast cancer cells
by Vicky Goler-Baron; Irina Sladkevich; Yehuda G. Assaraf (pp. 1340-1348).
Extracellular vesicles (EVs) formed between neighbor breast cancer cells mediate ABCG2-dependent multidrug resistance (MDR) via intravesicular drug sequestration. Inhibition of Akt signaling abolishes ABCG2 targeting, EVs biogenesis and overcomes MDR.We have recently shown that ABCG2-rich extracellular vesicles (EVs) form between neighbor breast cancer cells and actively concentrate various chemotherapeutics, resulting in multidrug resistance (MDR). Here we studied the signaling pathway regulating ABCG2 targeting to EVs as its inhibition would overcome MDR. The PI3K-Akt signaling pathway was possibly implicated in subcellular localization of ABCG2; we accordingly show here that pharmacological inhibition of Akt signaling results in gradual re-localization of ABCG2 from the EVs membrane to the cytoplasm. Cytoskeletal markers including β-actin and the tight junction protein ZO-1, along with the EVs markers ABCG2 and Ezrin–Radixin–Moesin revealed that this intracellular ABCG2 retention leads to gradual decrease in the size and number of EVs, resulting in EVs elimination and complete reversal of MDR. Inhibition of Akt signaling restored drug sensitivity to mitoxantrone and topotecan, bona fide ABCG2 transport substrates, hence being equivalent to MDR reversal achieved with the ABCG2 transport inhibitor Ko143. Remarkably, apart from loss of ABCG2 transport activity, treatment of MCF-7/MR cells with Ko143 resulted in cytoplasmic re-localization of ABCG2, similarly to the phenotype observed after Akt inhibition. We conclude that the PI3K-Akt signaling pathway is a key regulator of subcellular localization of ABCG2, EVs biogenesis and functional MDR. Furthermore, proper folding of ABCG2 and its targeting to the EVs membrane are crucial components of the biogenesis of EVs and their MDR function. We propose that Akt signaling inhibitors which disrupt ABCG2 targeting and EVs biogenesis may readily overcome MDR thus warranting in vivo studies with these promising drug combinations.
Keywords: Extracellular vesicles (EVs); PI3K-Akt signaling; Chemotherapy; ABC transporters; Overcoming multidrug resistance (MDR); Breast cancer
p38MAPK and ERK1/2 dictate cell death/survival response to different pro-oxidant stimuli via p53 and Nrf2 in neuroblastoma cells SH-SY5Y
by Giuseppe Filomeni; Sara Piccirillo; Giuseppe Rotilio; Maria R. Ciriolo (pp. 1349-1357).
Redox changes are often reported as causative of neoplastic transformation and chemoresistance, but are also exploited as clinical tools to selectively kill tumor cells. We previously demonstrated that gastrointestinal-derived tumor histotypes are resistant to ROS-based treatments by means of the redox activation of Nrf2, but highly sensitive to disulfide stressors triggering apoptosis via the redox induction of Trx1/p38MAPK/p53 signaling pathway.Here, we provide evidence that neuroblastoma SH-SY5Y has a complete opposite behavior, being sensitive to H2O2, but resistant to the glutathione (GSH)-oxidizing molecule diamide. Consistent with these observations, the apoptotic pathway activated upon H2O2 treatment relies upon Trx1 oxidation, and is mediated by the p38MAPK/p53 signaling axis. Pre-treatment with different antioxidants, pharmacological inhibitor of p38MAPK, or small interfering RNA against p53 rescue cell viability. On the contrary, cell survival to diamide relies upon redox activation of Nrf2, in a way independent on Keap1 oxidation, but responsive to ERK1/2 activation. Chemical inhibition of GSH neo-synthesis or ERK1/2 phosphorylation, as well as overexpression of the dominant-negative form of Nrf2 sensitizes cells to diamide toxicity. In the searching for the molecular determinant(s) unifying these phenomena, we found that SH-SY5Y cells show high GSH levels, but exhibit very low GPx activity. This feature allows to efficiently buffer disulfide stress, but leaves them being vulnerable to H2O2-mediated insult. The increase of GPx activity by means of selenium supplementation or GPx1 ectopic expression completely reverses death phenotype, indicating that the response of tumor cells to diverse oxidative stimuli deeply involves the entire GSH redox system.
Keywords: Abbreviations; AMS; 4-acetamido-4-maleimidylstilbene-2,2-disulfonic acid; ARE; antioxidant response elements; BSO; buthionine sulfoximine; DMSO; dimethyl sulfoxide; DMTU; dimethylthiourea; DN Nrf2; dominant negative Nrf2 protein; DNP; 2,4-dinitrophenylhydrazine; ERK1/2; extracellular signal related kinase 1 and 2; GI-GPx; gastrointestinal isoform of glutathione peroxidase; GPx; glutathione peroxidase; GSH; reduced glutathione; GSHest; reduced glutathione ethyl ester; GSSG; glutathione disulfide; HO-1; heme oxygenase-1; JNK; c-Jun-NH; 2; -terminal kinase; Keap1; kelch-like ECH-associated protein 1; MAPK; mitogen activated protein kinase; NAC; N-cetylcysteine; Nrf2; nuclear erythroid factor 2 (NE-F2)-related factor 2; p62/SQSTM1; protein of 62; kDa sequestrosoma1; PARP; polyADP-ribose polymerase; ROS; reactive oxygen species; Trx1; thioredoxin 1p53; Nrf2; MAPK; Glutathione; Oxidative stress; Diamide; Glutathione peroxidase
p38MAPK and ERK1/2 dictate cell death/survival response to different pro-oxidant stimuli via p53 and Nrf2 in neuroblastoma cells SH-SY5Y
by Giuseppe Filomeni; Sara Piccirillo; Giuseppe Rotilio; Maria R. Ciriolo (pp. 1349-1357).
Redox changes are often reported as causative of neoplastic transformation and chemoresistance, but are also exploited as clinical tools to selectively kill tumor cells. We previously demonstrated that gastrointestinal-derived tumor histotypes are resistant to ROS-based treatments by means of the redox activation of Nrf2, but highly sensitive to disulfide stressors triggering apoptosis via the redox induction of Trx1/p38MAPK/p53 signaling pathway.Here, we provide evidence that neuroblastoma SH-SY5Y has a complete opposite behavior, being sensitive to H2O2, but resistant to the glutathione (GSH)-oxidizing molecule diamide. Consistent with these observations, the apoptotic pathway activated upon H2O2 treatment relies upon Trx1 oxidation, and is mediated by the p38MAPK/p53 signaling axis. Pre-treatment with different antioxidants, pharmacological inhibitor of p38MAPK, or small interfering RNA against p53 rescue cell viability. On the contrary, cell survival to diamide relies upon redox activation of Nrf2, in a way independent on Keap1 oxidation, but responsive to ERK1/2 activation. Chemical inhibition of GSH neo-synthesis or ERK1/2 phosphorylation, as well as overexpression of the dominant-negative form of Nrf2 sensitizes cells to diamide toxicity. In the searching for the molecular determinant(s) unifying these phenomena, we found that SH-SY5Y cells show high GSH levels, but exhibit very low GPx activity. This feature allows to efficiently buffer disulfide stress, but leaves them being vulnerable to H2O2-mediated insult. The increase of GPx activity by means of selenium supplementation or GPx1 ectopic expression completely reverses death phenotype, indicating that the response of tumor cells to diverse oxidative stimuli deeply involves the entire GSH redox system.
Keywords: Abbreviations; AMS; 4-acetamido-4-maleimidylstilbene-2,2-disulfonic acid; ARE; antioxidant response elements; BSO; buthionine sulfoximine; DMSO; dimethyl sulfoxide; DMTU; dimethylthiourea; DN Nrf2; dominant negative Nrf2 protein; DNP; 2,4-dinitrophenylhydrazine; ERK1/2; extracellular signal related kinase 1 and 2; GI-GPx; gastrointestinal isoform of glutathione peroxidase; GPx; glutathione peroxidase; GSH; reduced glutathione; GSHest; reduced glutathione ethyl ester; GSSG; glutathione disulfide; HO-1; heme oxygenase-1; JNK; c-Jun-NH; 2; -terminal kinase; Keap1; kelch-like ECH-associated protein 1; MAPK; mitogen activated protein kinase; NAC; N-cetylcysteine; Nrf2; nuclear erythroid factor 2 (NE-F2)-related factor 2; p62/SQSTM1; protein of 62; kDa sequestrosoma1; PARP; polyADP-ribose polymerase; ROS; reactive oxygen species; Trx1; thioredoxin 1p53; Nrf2; MAPK; Glutathione; Oxidative stress; Diamide; Glutathione peroxidase
p38MAPK and ERK1/2 dictate cell death/survival response to different pro-oxidant stimuli via p53 and Nrf2 in neuroblastoma cells SH-SY5Y
by Giuseppe Filomeni; Sara Piccirillo; Giuseppe Rotilio; Maria R. Ciriolo (pp. 1349-1357).
Redox changes are often reported as causative of neoplastic transformation and chemoresistance, but are also exploited as clinical tools to selectively kill tumor cells. We previously demonstrated that gastrointestinal-derived tumor histotypes are resistant to ROS-based treatments by means of the redox activation of Nrf2, but highly sensitive to disulfide stressors triggering apoptosis via the redox induction of Trx1/p38MAPK/p53 signaling pathway.Here, we provide evidence that neuroblastoma SH-SY5Y has a complete opposite behavior, being sensitive to H2O2, but resistant to the glutathione (GSH)-oxidizing molecule diamide. Consistent with these observations, the apoptotic pathway activated upon H2O2 treatment relies upon Trx1 oxidation, and is mediated by the p38MAPK/p53 signaling axis. Pre-treatment with different antioxidants, pharmacological inhibitor of p38MAPK, or small interfering RNA against p53 rescue cell viability. On the contrary, cell survival to diamide relies upon redox activation of Nrf2, in a way independent on Keap1 oxidation, but responsive to ERK1/2 activation. Chemical inhibition of GSH neo-synthesis or ERK1/2 phosphorylation, as well as overexpression of the dominant-negative form of Nrf2 sensitizes cells to diamide toxicity. In the searching for the molecular determinant(s) unifying these phenomena, we found that SH-SY5Y cells show high GSH levels, but exhibit very low GPx activity. This feature allows to efficiently buffer disulfide stress, but leaves them being vulnerable to H2O2-mediated insult. The increase of GPx activity by means of selenium supplementation or GPx1 ectopic expression completely reverses death phenotype, indicating that the response of tumor cells to diverse oxidative stimuli deeply involves the entire GSH redox system.
Keywords: Abbreviations; AMS; 4-acetamido-4-maleimidylstilbene-2,2-disulfonic acid; ARE; antioxidant response elements; BSO; buthionine sulfoximine; DMSO; dimethyl sulfoxide; DMTU; dimethylthiourea; DN Nrf2; dominant negative Nrf2 protein; DNP; 2,4-dinitrophenylhydrazine; ERK1/2; extracellular signal related kinase 1 and 2; GI-GPx; gastrointestinal isoform of glutathione peroxidase; GPx; glutathione peroxidase; GSH; reduced glutathione; GSHest; reduced glutathione ethyl ester; GSSG; glutathione disulfide; HO-1; heme oxygenase-1; JNK; c-Jun-NH; 2; -terminal kinase; Keap1; kelch-like ECH-associated protein 1; MAPK; mitogen activated protein kinase; NAC; N-cetylcysteine; Nrf2; nuclear erythroid factor 2 (NE-F2)-related factor 2; p62/SQSTM1; protein of 62; kDa sequestrosoma1; PARP; polyADP-ribose polymerase; ROS; reactive oxygen species; Trx1; thioredoxin 1p53; Nrf2; MAPK; Glutathione; Oxidative stress; Diamide; Glutathione peroxidase
Inhibition of pyrimidine and purine nucleoside phosphorylases by a 3,5-dichlorobenzoyl-substituted 2-deoxy-d-ribose-1-phosphate derivative
by Johan Vande Voorde; Maurizio Quintiliani; Christopher McGuigan; Sandra Liekens; Jan Balzarini (pp. 1358-1363).
The 3,5-dichlorobenzoyl-substituted 2-deoxy-d-ribose-1-phosphate derivative, designated Cf2891, was found to inhibit a variety of pyrimidine and purine nucleoside phosphorylases (NPs) with preference for uridine- and inosine-hydrolyzing enzymes [uridine phosphorylase (UP; EC 2.4.2.3), pyrimidine nucleoside phosphorylase (PyNP; EC 2.4.2.2) and purine nucleoside phosphorylase (PNP; EC 2.4.2.1)]. Kinetic analyses revealed that Cf2891 competes with inorganic phosphate (Pi) for binding to the NPs and, depending on the nature of the enzyme, acts as a competitive or non-competitive inhibitor with regard to the nucleoside binding site. Also, the compound prevents breakdown of pyrimidine analogues used in the treatment of viral infections and cancer. Since NPs are abundantly present in tumor tissue and may be overexpressed due to secondary bacterial infections in immunocompromised patients suffering viral infections, Cf2891 may serve as a lead molecule for the development of inhibitors to be used in nucleoside-based combination therapy.
Keywords: Nucleoside phosphorylase (NP); Purine nucleoside phosphorylase (PNP); Uridine phosphorylase (UP); Thymidine phosphorylase (TP); Pyrimidine nucleoside phosphorylase (PyNP); Mycoplasma hyorhinis
Inhibition of pyrimidine and purine nucleoside phosphorylases by a 3,5-dichlorobenzoyl-substituted 2-deoxy-d-ribose-1-phosphate derivative
by Johan Vande Voorde; Maurizio Quintiliani; Christopher McGuigan; Sandra Liekens; Jan Balzarini (pp. 1358-1363).
The 3,5-dichlorobenzoyl-substituted 2-deoxy-d-ribose-1-phosphate derivative, designated Cf2891, was found to inhibit a variety of pyrimidine and purine nucleoside phosphorylases (NPs) with preference for uridine- and inosine-hydrolyzing enzymes [uridine phosphorylase (UP; EC 2.4.2.3), pyrimidine nucleoside phosphorylase (PyNP; EC 2.4.2.2) and purine nucleoside phosphorylase (PNP; EC 2.4.2.1)]. Kinetic analyses revealed that Cf2891 competes with inorganic phosphate (Pi) for binding to the NPs and, depending on the nature of the enzyme, acts as a competitive or non-competitive inhibitor with regard to the nucleoside binding site. Also, the compound prevents breakdown of pyrimidine analogues used in the treatment of viral infections and cancer. Since NPs are abundantly present in tumor tissue and may be overexpressed due to secondary bacterial infections in immunocompromised patients suffering viral infections, Cf2891 may serve as a lead molecule for the development of inhibitors to be used in nucleoside-based combination therapy.
Keywords: Nucleoside phosphorylase (NP); Purine nucleoside phosphorylase (PNP); Uridine phosphorylase (UP); Thymidine phosphorylase (TP); Pyrimidine nucleoside phosphorylase (PyNP); Mycoplasma hyorhinis
Inhibition of pyrimidine and purine nucleoside phosphorylases by a 3,5-dichlorobenzoyl-substituted 2-deoxy-d-ribose-1-phosphate derivative
by Johan Vande Voorde; Maurizio Quintiliani; Christopher McGuigan; Sandra Liekens; Jan Balzarini (pp. 1358-1363).
The 3,5-dichlorobenzoyl-substituted 2-deoxy-d-ribose-1-phosphate derivative, designated Cf2891, was found to inhibit a variety of pyrimidine and purine nucleoside phosphorylases (NPs) with preference for uridine- and inosine-hydrolyzing enzymes [uridine phosphorylase (UP; EC 2.4.2.3), pyrimidine nucleoside phosphorylase (PyNP; EC 2.4.2.2) and purine nucleoside phosphorylase (PNP; EC 2.4.2.1)]. Kinetic analyses revealed that Cf2891 competes with inorganic phosphate (Pi) for binding to the NPs and, depending on the nature of the enzyme, acts as a competitive or non-competitive inhibitor with regard to the nucleoside binding site. Also, the compound prevents breakdown of pyrimidine analogues used in the treatment of viral infections and cancer. Since NPs are abundantly present in tumor tissue and may be overexpressed due to secondary bacterial infections in immunocompromised patients suffering viral infections, Cf2891 may serve as a lead molecule for the development of inhibitors to be used in nucleoside-based combination therapy.
Keywords: Nucleoside phosphorylase (NP); Purine nucleoside phosphorylase (PNP); Uridine phosphorylase (UP); Thymidine phosphorylase (TP); Pyrimidine nucleoside phosphorylase (PyNP); Mycoplasma hyorhinis
Prednisolone exerts exquisite inhibitory properties on platelet functions
by Elisabetta Liverani; Sreemoti Banerjee; Wayne Roberts; Khalid M. Naseem; Mauro Perretti (pp. 1364-1373).
We have previously reported presence of the glucocorticoid (GC) receptor (GR) alpha on blood platelets, and its ability to modulate platelet aggregation when activated by the synthetic GC prednisolone (Pred). In the present study we investigated the effects of Pred on broader aspects of platelet functions to unveil novel non-genomic actions on this cell type. Using whole blood assay we demonstrated that Pred was the only GC able to inhibit platelet aggregation and platelet–monocyte interactions. This latter effect was due to regulation of platelets, not monocytes. We next examined the effects of Pred on physiological actions of platelets, observing inhibition of platelet adhesion and spreading on collagen under static conditions. Moreover Pred inhibited thrombus formation under flow, suggesting potential important effects in haemostasis and thrombosis. Pred was unable to regulate platelet reactivity under conditions where the effects of platelet-derived ADP and TxA2 were blocked, suggesting that the GC targeted the activation-dependent component of the adhesion and aggregation response. The effects of Pred were not mediated through cyclic nucleotide signaling, but rather seemed to evolve around selective regulation of P2Y12 ADP receptor signaling, intimating a novel mode of action. This study details the actions of Pred on platelets unveiling novel properties which could be relevant for this GC in controlling unwanted vascular and thrombotic diseases.
Keywords: Platelet biology; Prednisolone; Glucocorticoid receptor; Adhesion; Aggregation; Cell-to-cell interaction
Prednisolone exerts exquisite inhibitory properties on platelet functions
by Elisabetta Liverani; Sreemoti Banerjee; Wayne Roberts; Khalid M. Naseem; Mauro Perretti (pp. 1364-1373).
We have previously reported presence of the glucocorticoid (GC) receptor (GR) alpha on blood platelets, and its ability to modulate platelet aggregation when activated by the synthetic GC prednisolone (Pred). In the present study we investigated the effects of Pred on broader aspects of platelet functions to unveil novel non-genomic actions on this cell type. Using whole blood assay we demonstrated that Pred was the only GC able to inhibit platelet aggregation and platelet–monocyte interactions. This latter effect was due to regulation of platelets, not monocytes. We next examined the effects of Pred on physiological actions of platelets, observing inhibition of platelet adhesion and spreading on collagen under static conditions. Moreover Pred inhibited thrombus formation under flow, suggesting potential important effects in haemostasis and thrombosis. Pred was unable to regulate platelet reactivity under conditions where the effects of platelet-derived ADP and TxA2 were blocked, suggesting that the GC targeted the activation-dependent component of the adhesion and aggregation response. The effects of Pred were not mediated through cyclic nucleotide signaling, but rather seemed to evolve around selective regulation of P2Y12 ADP receptor signaling, intimating a novel mode of action. This study details the actions of Pred on platelets unveiling novel properties which could be relevant for this GC in controlling unwanted vascular and thrombotic diseases.
Keywords: Platelet biology; Prednisolone; Glucocorticoid receptor; Adhesion; Aggregation; Cell-to-cell interaction
Prednisolone exerts exquisite inhibitory properties on platelet functions
by Elisabetta Liverani; Sreemoti Banerjee; Wayne Roberts; Khalid M. Naseem; Mauro Perretti (pp. 1364-1373).
We have previously reported presence of the glucocorticoid (GC) receptor (GR) alpha on blood platelets, and its ability to modulate platelet aggregation when activated by the synthetic GC prednisolone (Pred). In the present study we investigated the effects of Pred on broader aspects of platelet functions to unveil novel non-genomic actions on this cell type. Using whole blood assay we demonstrated that Pred was the only GC able to inhibit platelet aggregation and platelet–monocyte interactions. This latter effect was due to regulation of platelets, not monocytes. We next examined the effects of Pred on physiological actions of platelets, observing inhibition of platelet adhesion and spreading on collagen under static conditions. Moreover Pred inhibited thrombus formation under flow, suggesting potential important effects in haemostasis and thrombosis. Pred was unable to regulate platelet reactivity under conditions where the effects of platelet-derived ADP and TxA2 were blocked, suggesting that the GC targeted the activation-dependent component of the adhesion and aggregation response. The effects of Pred were not mediated through cyclic nucleotide signaling, but rather seemed to evolve around selective regulation of P2Y12 ADP receptor signaling, intimating a novel mode of action. This study details the actions of Pred on platelets unveiling novel properties which could be relevant for this GC in controlling unwanted vascular and thrombotic diseases.
Keywords: Platelet biology; Prednisolone; Glucocorticoid receptor; Adhesion; Aggregation; Cell-to-cell interaction
Inhibition of farnesyltransferase reduces angiogenesis by interrupting endothelial cell migration
by Guoyuan Peng; Yuan Ren; Xiaodong Sun; Jun Zhou; Dengwen Li (pp. 1374-1382).
Inhibitors of farnesyltransferase (FTI) have been developed for cancer treatment for more than a decade. Aside from being a therapeutic target in tumor cells, little is known about the role of farnesyltransferase (FTase) in other physiological processes. In this study, we revealed the involvement of FTase in angiogenesis and showed that FTI inhibited angiogenesis by directly acting on endothelial cells. Inhibition of FTase interrupted cell migration in vitro and in vivo. In addition, we found that FTase was important for cell polarization, cell spreading and pseudopodia formation. We also found that FTase interacted with microtubule end binding protein 1 (EB1) and that this interaction was critical for the localization of EB1 to microtubule tips. Our findings thus offer novel insight into the functions of FTase in endothelial cells and provide valuable information for the use of FTI in cancer therapy.
Keywords: Farnesyltransferase; Angiogenesis; Migration; Endothelial cell; Microtubule
Inhibition of farnesyltransferase reduces angiogenesis by interrupting endothelial cell migration
by Guoyuan Peng; Yuan Ren; Xiaodong Sun; Jun Zhou; Dengwen Li (pp. 1374-1382).
Inhibitors of farnesyltransferase (FTI) have been developed for cancer treatment for more than a decade. Aside from being a therapeutic target in tumor cells, little is known about the role of farnesyltransferase (FTase) in other physiological processes. In this study, we revealed the involvement of FTase in angiogenesis and showed that FTI inhibited angiogenesis by directly acting on endothelial cells. Inhibition of FTase interrupted cell migration in vitro and in vivo. In addition, we found that FTase was important for cell polarization, cell spreading and pseudopodia formation. We also found that FTase interacted with microtubule end binding protein 1 (EB1) and that this interaction was critical for the localization of EB1 to microtubule tips. Our findings thus offer novel insight into the functions of FTase in endothelial cells and provide valuable information for the use of FTI in cancer therapy.
Keywords: Farnesyltransferase; Angiogenesis; Migration; Endothelial cell; Microtubule
Inhibition of farnesyltransferase reduces angiogenesis by interrupting endothelial cell migration
by Guoyuan Peng; Yuan Ren; Xiaodong Sun; Jun Zhou; Dengwen Li (pp. 1374-1382).
Inhibitors of farnesyltransferase (FTI) have been developed for cancer treatment for more than a decade. Aside from being a therapeutic target in tumor cells, little is known about the role of farnesyltransferase (FTase) in other physiological processes. In this study, we revealed the involvement of FTase in angiogenesis and showed that FTI inhibited angiogenesis by directly acting on endothelial cells. Inhibition of FTase interrupted cell migration in vitro and in vivo. In addition, we found that FTase was important for cell polarization, cell spreading and pseudopodia formation. We also found that FTase interacted with microtubule end binding protein 1 (EB1) and that this interaction was critical for the localization of EB1 to microtubule tips. Our findings thus offer novel insight into the functions of FTase in endothelial cells and provide valuable information for the use of FTI in cancer therapy.
Keywords: Farnesyltransferase; Angiogenesis; Migration; Endothelial cell; Microtubule
Double-edged sword effect of biochanin to inhibit nuclear factor kappaB: Suppression of serine/threonine and tyrosine kinases
by Sunil Kumar Manna (pp. 1383-1392).
Several protein tyrosine kinase (PTK) inhibitors predominantly isoflavones, such as genistein, erbstatin, quercetin, daidzein, present in red clover, cabbage and alfalfa, show apoptotic effect against cancer cells. In this study I found that biochanin, a methoxy form of genistein, inhibits IL-8-mediated activation of nuclear transcription factor kappaB (NF-κB) and activator protein 1 (AP-1) more potently than genistein as shown in Jurkat T-cell line. Both biochanin and genistein potently inhibited activity of Lck and Syk, but biochanin specifically inhibited activity of IKK. Biochanin inhibited completely NF-κB activation induced by PMA, LPS, pervanadate (PV), or H2O2, but only partially that induced by TNFα. Genistein was unable to inhibit IL-8-induced IKK activity, but it blocked PV-induced IKK activity. Biochanin inhibited activation of NF-κB by TRAF6 completely, but by TRAF2 partially. In silico data suggested that biochanin interacted strongly with serine/threonine kinase than genistein, though both equally interacted with PTK. The data show that both biochanin and genistein are potent inhibitors of PTK, but biochanin is a potent inhibitor of serine/threonine kinase too. Formononetin, having hydroxyl methoxy group is less potent to inhibit IKK than biochanin. Biochanin inhibits NF-κB activation not only by blocking the upstream IKK, but also PTK that phosphorylate tyrosine residues of IκBα. Thus, the double-edged sword effect of inhibition of NF-κB via inhibition of both serine/threonine kinase and PTK by biochanin might show useful therapeutic value against activities of cells that lead to tumorigenesis and inflammation.
Keywords: Abbreviations; AP-1; activator protein 1; C3-toxin; Clostridium botulinum; C3 transferase; CE; cytoplasmic extracts; Cox2; cyclooxygenase 2; ICAM; intercellular adhesion molecule; NF-κB; nuclear transcription factor kappaB; IL; interleukin; IκB; inhibitory subunit of NF-κB; IκBα-DN; IκBα dominant negative; IKK; IκB kinase; Lck; lymphocyte-specific PTK (protein tyrosine kinase); LPS; lipopolysaccharide; NE; nuclear extracts; PV; pervanadate; RMSD; root mean square deviation; Syk; spleen tyrosine kinase; TNFα; tumor necrosis factor alpha; TRAF; TNF receptor-associated factorProtein tyrosine kinase; Ser/Thr kinase; Biochanin; NF-κB; IKK; PTK inhibitors
Double-edged sword effect of biochanin to inhibit nuclear factor kappaB: Suppression of serine/threonine and tyrosine kinases
by Sunil Kumar Manna (pp. 1383-1392).
Several protein tyrosine kinase (PTK) inhibitors predominantly isoflavones, such as genistein, erbstatin, quercetin, daidzein, present in red clover, cabbage and alfalfa, show apoptotic effect against cancer cells. In this study I found that biochanin, a methoxy form of genistein, inhibits IL-8-mediated activation of nuclear transcription factor kappaB (NF-κB) and activator protein 1 (AP-1) more potently than genistein as shown in Jurkat T-cell line. Both biochanin and genistein potently inhibited activity of Lck and Syk, but biochanin specifically inhibited activity of IKK. Biochanin inhibited completely NF-κB activation induced by PMA, LPS, pervanadate (PV), or H2O2, but only partially that induced by TNFα. Genistein was unable to inhibit IL-8-induced IKK activity, but it blocked PV-induced IKK activity. Biochanin inhibited activation of NF-κB by TRAF6 completely, but by TRAF2 partially. In silico data suggested that biochanin interacted strongly with serine/threonine kinase than genistein, though both equally interacted with PTK. The data show that both biochanin and genistein are potent inhibitors of PTK, but biochanin is a potent inhibitor of serine/threonine kinase too. Formononetin, having hydroxyl methoxy group is less potent to inhibit IKK than biochanin. Biochanin inhibits NF-κB activation not only by blocking the upstream IKK, but also PTK that phosphorylate tyrosine residues of IκBα. Thus, the double-edged sword effect of inhibition of NF-κB via inhibition of both serine/threonine kinase and PTK by biochanin might show useful therapeutic value against activities of cells that lead to tumorigenesis and inflammation.
Keywords: Abbreviations; AP-1; activator protein 1; C3-toxin; Clostridium botulinum; C3 transferase; CE; cytoplasmic extracts; Cox2; cyclooxygenase 2; ICAM; intercellular adhesion molecule; NF-κB; nuclear transcription factor kappaB; IL; interleukin; IκB; inhibitory subunit of NF-κB; IκBα-DN; IκBα dominant negative; IKK; IκB kinase; Lck; lymphocyte-specific PTK (protein tyrosine kinase); LPS; lipopolysaccharide; NE; nuclear extracts; PV; pervanadate; RMSD; root mean square deviation; Syk; spleen tyrosine kinase; TNFα; tumor necrosis factor alpha; TRAF; TNF receptor-associated factorProtein tyrosine kinase; Ser/Thr kinase; Biochanin; NF-κB; IKK; PTK inhibitors
Double-edged sword effect of biochanin to inhibit nuclear factor kappaB: Suppression of serine/threonine and tyrosine kinases
by Sunil Kumar Manna (pp. 1383-1392).
Several protein tyrosine kinase (PTK) inhibitors predominantly isoflavones, such as genistein, erbstatin, quercetin, daidzein, present in red clover, cabbage and alfalfa, show apoptotic effect against cancer cells. In this study I found that biochanin, a methoxy form of genistein, inhibits IL-8-mediated activation of nuclear transcription factor kappaB (NF-κB) and activator protein 1 (AP-1) more potently than genistein as shown in Jurkat T-cell line. Both biochanin and genistein potently inhibited activity of Lck and Syk, but biochanin specifically inhibited activity of IKK. Biochanin inhibited completely NF-κB activation induced by PMA, LPS, pervanadate (PV), or H2O2, but only partially that induced by TNFα. Genistein was unable to inhibit IL-8-induced IKK activity, but it blocked PV-induced IKK activity. Biochanin inhibited activation of NF-κB by TRAF6 completely, but by TRAF2 partially. In silico data suggested that biochanin interacted strongly with serine/threonine kinase than genistein, though both equally interacted with PTK. The data show that both biochanin and genistein are potent inhibitors of PTK, but biochanin is a potent inhibitor of serine/threonine kinase too. Formononetin, having hydroxyl methoxy group is less potent to inhibit IKK than biochanin. Biochanin inhibits NF-κB activation not only by blocking the upstream IKK, but also PTK that phosphorylate tyrosine residues of IκBα. Thus, the double-edged sword effect of inhibition of NF-κB via inhibition of both serine/threonine kinase and PTK by biochanin might show useful therapeutic value against activities of cells that lead to tumorigenesis and inflammation.
Keywords: Abbreviations; AP-1; activator protein 1; C3-toxin; Clostridium botulinum; C3 transferase; CE; cytoplasmic extracts; Cox2; cyclooxygenase 2; ICAM; intercellular adhesion molecule; NF-κB; nuclear transcription factor kappaB; IL; interleukin; IκB; inhibitory subunit of NF-κB; IκBα-DN; IκBα dominant negative; IKK; IκB kinase; Lck; lymphocyte-specific PTK (protein tyrosine kinase); LPS; lipopolysaccharide; NE; nuclear extracts; PV; pervanadate; RMSD; root mean square deviation; Syk; spleen tyrosine kinase; TNFα; tumor necrosis factor alpha; TRAF; TNF receptor-associated factorProtein tyrosine kinase; Ser/Thr kinase; Biochanin; NF-κB; IKK; PTK inhibitors
Cannabinoid receptor trafficking in peripheral cells is dynamically regulated by a binary biochemical switch
by Jonas Kleyer; Simon Nicolussi; Peter Taylor; Deborah Simonelli; Evelyne Furger; Pascale Anderle; Jürg Gertsch (pp. 1393-1412).
The cannabinoid G protein-coupled receptors (GPCRs) CB1 and CB2 are expressed in different peripheral cells. Localization of GPCRs in the cell membrane determines signaling via G protein pathways. Here we show that unlike in transfected cells, CB receptors in cell lines and primary human cells are not internalized upon agonist interaction, but move between cytoplasm and cell membranes by ligand-independent trafficking mechanisms. Even though CB receptors are expressed in many cells of peripheral origin they are not always localized in the cell membrane and in most cancer cell lines the ratios between CB1 and CB2 receptor gene and surface expression vary significantly. In contrast, CB receptor cell surface expression in HL60 cells is subject to significant oscillations and CB2 receptors form oligomers and heterodimers with CB1 receptors, showing synchronized surface expression, localization and trafficking. We show that hydrogen peroxide and other nonspecific protein tyrosine phosphatase inhibitors (TPIs) such as phenylarsine oxide trigger both CB2 receptor internalization and externalization, depending on receptor localization. Phorbol ester-mediated internalization of CB receptors can be inhibited via this switch. In primary human immune cells hydrogen peroxide and other TPIs lead to a robust internalization of CB receptors in monocytes and an externalization in T cells. This study describes, for the first time, the dynamic nature of CB receptor trafficking in the context of a biochemical switch, which may have implications for studies on the cell-type specific effects of cannabinoids and our understanding of the regulation of CB receptor cell surface expression.
Keywords: Abbreviations; ATTC; American type culture collection; AMAM; anilino-monoindolylmaleimide; CB; 1; cannabinoid receptor type-1; CB; 2; cannabinoid receptor type-2; CDx; cluster of differentiation x; CHO; Chinese hamster ovary-K1; CM; cell membrane; CNR1; CB; 1; receptor gene; CNR2; CB; 2; receptor gene; DCA; dichloroacetate; DMEM; Dulbecco's modified eagle medium; DMSZ; German collection of microorganisms and cell cultures; DTSSP; 3,3′-dithiobis(sulfosuccinimidyl propionate); ECS; endocannabinoid system; ER; endoplasmic reticulum; FACS; fluorescence-activated cell sorting; FBS; fetal bovine serum; GRK; G protein receptor kinase; GPCR; G protein-coupled receptor; HEK; human embryonic kidney-293; H; 2; O; 2; hydrogen peroxide; MCSF; macrophage colony stimulating factor; MCD; methyl-β-cyclodextrin; PAO; phenylarsine oxide; PBMCs; peripheral blood mononuclear cells; PBS-T; PBS with 0.01% Tween twenty; PDI; protein disulfide isomerase; PFA; paraformaldehyde; PKC; protein kinase C; PMA; phrobol 12-myristate 13-acetate; PTK; protein tyrosine kinase; PTP; protein tyrosine phosphatase; RT; room temperature; SFM; serum-free media; TPI; protein tyrosine phosphatase inhibitor; TBS; tris-buffered salineCannabinoid receptors; GPCR heterodimerization; Endocannabinoid system; Immune cells; Receptor trafficking
Cannabinoid receptor trafficking in peripheral cells is dynamically regulated by a binary biochemical switch
by Jonas Kleyer; Simon Nicolussi; Peter Taylor; Deborah Simonelli; Evelyne Furger; Pascale Anderle; Jürg Gertsch (pp. 1393-1412).
The cannabinoid G protein-coupled receptors (GPCRs) CB1 and CB2 are expressed in different peripheral cells. Localization of GPCRs in the cell membrane determines signaling via G protein pathways. Here we show that unlike in transfected cells, CB receptors in cell lines and primary human cells are not internalized upon agonist interaction, but move between cytoplasm and cell membranes by ligand-independent trafficking mechanisms. Even though CB receptors are expressed in many cells of peripheral origin they are not always localized in the cell membrane and in most cancer cell lines the ratios between CB1 and CB2 receptor gene and surface expression vary significantly. In contrast, CB receptor cell surface expression in HL60 cells is subject to significant oscillations and CB2 receptors form oligomers and heterodimers with CB1 receptors, showing synchronized surface expression, localization and trafficking. We show that hydrogen peroxide and other nonspecific protein tyrosine phosphatase inhibitors (TPIs) such as phenylarsine oxide trigger both CB2 receptor internalization and externalization, depending on receptor localization. Phorbol ester-mediated internalization of CB receptors can be inhibited via this switch. In primary human immune cells hydrogen peroxide and other TPIs lead to a robust internalization of CB receptors in monocytes and an externalization in T cells. This study describes, for the first time, the dynamic nature of CB receptor trafficking in the context of a biochemical switch, which may have implications for studies on the cell-type specific effects of cannabinoids and our understanding of the regulation of CB receptor cell surface expression.
Keywords: Abbreviations; ATTC; American type culture collection; AMAM; anilino-monoindolylmaleimide; CB; 1; cannabinoid receptor type-1; CB; 2; cannabinoid receptor type-2; CDx; cluster of differentiation x; CHO; Chinese hamster ovary-K1; CM; cell membrane; CNR1; CB; 1; receptor gene; CNR2; CB; 2; receptor gene; DCA; dichloroacetate; DMEM; Dulbecco's modified eagle medium; DMSZ; German collection of microorganisms and cell cultures; DTSSP; 3,3′-dithiobis(sulfosuccinimidyl propionate); ECS; endocannabinoid system; ER; endoplasmic reticulum; FACS; fluorescence-activated cell sorting; FBS; fetal bovine serum; GRK; G protein receptor kinase; GPCR; G protein-coupled receptor; HEK; human embryonic kidney-293; H; 2; O; 2; hydrogen peroxide; MCSF; macrophage colony stimulating factor; MCD; methyl-β-cyclodextrin; PAO; phenylarsine oxide; PBMCs; peripheral blood mononuclear cells; PBS-T; PBS with 0.01% Tween twenty; PDI; protein disulfide isomerase; PFA; paraformaldehyde; PKC; protein kinase C; PMA; phrobol 12-myristate 13-acetate; PTK; protein tyrosine kinase; PTP; protein tyrosine phosphatase; RT; room temperature; SFM; serum-free media; TPI; protein tyrosine phosphatase inhibitor; TBS; tris-buffered salineCannabinoid receptors; GPCR heterodimerization; Endocannabinoid system; Immune cells; Receptor trafficking
Cannabinoid receptor trafficking in peripheral cells is dynamically regulated by a binary biochemical switch
by Jonas Kleyer; Simon Nicolussi; Peter Taylor; Deborah Simonelli; Evelyne Furger; Pascale Anderle; Jürg Gertsch (pp. 1393-1412).
The cannabinoid G protein-coupled receptors (GPCRs) CB1 and CB2 are expressed in different peripheral cells. Localization of GPCRs in the cell membrane determines signaling via G protein pathways. Here we show that unlike in transfected cells, CB receptors in cell lines and primary human cells are not internalized upon agonist interaction, but move between cytoplasm and cell membranes by ligand-independent trafficking mechanisms. Even though CB receptors are expressed in many cells of peripheral origin they are not always localized in the cell membrane and in most cancer cell lines the ratios between CB1 and CB2 receptor gene and surface expression vary significantly. In contrast, CB receptor cell surface expression in HL60 cells is subject to significant oscillations and CB2 receptors form oligomers and heterodimers with CB1 receptors, showing synchronized surface expression, localization and trafficking. We show that hydrogen peroxide and other nonspecific protein tyrosine phosphatase inhibitors (TPIs) such as phenylarsine oxide trigger both CB2 receptor internalization and externalization, depending on receptor localization. Phorbol ester-mediated internalization of CB receptors can be inhibited via this switch. In primary human immune cells hydrogen peroxide and other TPIs lead to a robust internalization of CB receptors in monocytes and an externalization in T cells. This study describes, for the first time, the dynamic nature of CB receptor trafficking in the context of a biochemical switch, which may have implications for studies on the cell-type specific effects of cannabinoids and our understanding of the regulation of CB receptor cell surface expression.
Keywords: Abbreviations; ATTC; American type culture collection; AMAM; anilino-monoindolylmaleimide; CB; 1; cannabinoid receptor type-1; CB; 2; cannabinoid receptor type-2; CDx; cluster of differentiation x; CHO; Chinese hamster ovary-K1; CM; cell membrane; CNR1; CB; 1; receptor gene; CNR2; CB; 2; receptor gene; DCA; dichloroacetate; DMEM; Dulbecco's modified eagle medium; DMSZ; German collection of microorganisms and cell cultures; DTSSP; 3,3′-dithiobis(sulfosuccinimidyl propionate); ECS; endocannabinoid system; ER; endoplasmic reticulum; FACS; fluorescence-activated cell sorting; FBS; fetal bovine serum; GRK; G protein receptor kinase; GPCR; G protein-coupled receptor; HEK; human embryonic kidney-293; H; 2; O; 2; hydrogen peroxide; MCSF; macrophage colony stimulating factor; MCD; methyl-β-cyclodextrin; PAO; phenylarsine oxide; PBMCs; peripheral blood mononuclear cells; PBS-T; PBS with 0.01% Tween twenty; PDI; protein disulfide isomerase; PFA; paraformaldehyde; PKC; protein kinase C; PMA; phrobol 12-myristate 13-acetate; PTK; protein tyrosine kinase; PTP; protein tyrosine phosphatase; RT; room temperature; SFM; serum-free media; TPI; protein tyrosine phosphatase inhibitor; TBS; tris-buffered salineCannabinoid receptors; GPCR heterodimerization; Endocannabinoid system; Immune cells; Receptor trafficking
The effects of a polyphenol present in olive oil, oleuropein aglycone, in an experimental model of spinal cord injury in mice
by Daniela Impellizzeri; Emanuela Esposito; Emanuela Mazzon; Irene Paterniti; Rosanna Di Paola; Placido Bramanti; Valeria Maria Morittu; Antonio Procopio; Enzo Perri; Domenico Britti; Salvatore Cuzzocrea (pp. 1413-1426).
Several olive oil phenolic compounds, such us oleuropein have attracted considerable attention because of their antioxidant activity, anti-atherosclerotic and anti-inflammatory properties. The aim of this experimental study was to determine the effect of oleuropein aglycone, a hydrolysis product of oleuropein, in the inflammatory response, in particular in the secondary injury associated with the mouse model of spinal cord trauma. The injury was induced by application of vascular clips to the dura via a four-level T5–T8 laminectomy in mice. Oleuropein aglycone was administered in mice (100μg/kg, 40μg/kg, 20μg/kg, 10% ethanol, i.p.) 1h and 6h after the trauma. The treatment with oleuropein aglycone significantly decreased: (1) histological damage, (2) motor recovery, (3) nuclear factor (NF)-κB expression and IKB-α degradation, (4) protein kinase A (PKA) activity and expression, (5) pro-inflammatory cytokines production such as tumor necrosis factor (TNF-α) and interleukin-1β (IL-1β), 6) inducible nitric oxide synthase (iNOS) expression, (7) neutrophil infiltration, (8) lipid peroxidation, (9) nitrotyrosine and poly-ADP-ribose (PAR) formation, (10) glial cell-derived neurotrophic factor (GDNF) levels, (11) apoptosis (TUNEL staining, FAS ligand expression, Caspase 3, Bax and Bcl-2 expression). Thus, we propose that olive oil phenolic constituents such as oleuropein aglycone may be useful in the treatment of various inflammatory diseases.
Keywords: Spinal cord; Inflammation; Cytokines; Apoptosis; Oxidative stress; Olive oil
The effects of a polyphenol present in olive oil, oleuropein aglycone, in an experimental model of spinal cord injury in mice
by Daniela Impellizzeri; Emanuela Esposito; Emanuela Mazzon; Irene Paterniti; Rosanna Di Paola; Placido Bramanti; Valeria Maria Morittu; Antonio Procopio; Enzo Perri; Domenico Britti; Salvatore Cuzzocrea (pp. 1413-1426).
Several olive oil phenolic compounds, such us oleuropein have attracted considerable attention because of their antioxidant activity, anti-atherosclerotic and anti-inflammatory properties. The aim of this experimental study was to determine the effect of oleuropein aglycone, a hydrolysis product of oleuropein, in the inflammatory response, in particular in the secondary injury associated with the mouse model of spinal cord trauma. The injury was induced by application of vascular clips to the dura via a four-level T5–T8 laminectomy in mice. Oleuropein aglycone was administered in mice (100μg/kg, 40μg/kg, 20μg/kg, 10% ethanol, i.p.) 1h and 6h after the trauma. The treatment with oleuropein aglycone significantly decreased: (1) histological damage, (2) motor recovery, (3) nuclear factor (NF)-κB expression and IKB-α degradation, (4) protein kinase A (PKA) activity and expression, (5) pro-inflammatory cytokines production such as tumor necrosis factor (TNF-α) and interleukin-1β (IL-1β), 6) inducible nitric oxide synthase (iNOS) expression, (7) neutrophil infiltration, (8) lipid peroxidation, (9) nitrotyrosine and poly-ADP-ribose (PAR) formation, (10) glial cell-derived neurotrophic factor (GDNF) levels, (11) apoptosis (TUNEL staining, FAS ligand expression, Caspase 3, Bax and Bcl-2 expression). Thus, we propose that olive oil phenolic constituents such as oleuropein aglycone may be useful in the treatment of various inflammatory diseases.
Keywords: Spinal cord; Inflammation; Cytokines; Apoptosis; Oxidative stress; Olive oil
The effects of a polyphenol present in olive oil, oleuropein aglycone, in an experimental model of spinal cord injury in mice
by Daniela Impellizzeri; Emanuela Esposito; Emanuela Mazzon; Irene Paterniti; Rosanna Di Paola; Placido Bramanti; Valeria Maria Morittu; Antonio Procopio; Enzo Perri; Domenico Britti; Salvatore Cuzzocrea (pp. 1413-1426).
Several olive oil phenolic compounds, such us oleuropein have attracted considerable attention because of their antioxidant activity, anti-atherosclerotic and anti-inflammatory properties. The aim of this experimental study was to determine the effect of oleuropein aglycone, a hydrolysis product of oleuropein, in the inflammatory response, in particular in the secondary injury associated with the mouse model of spinal cord trauma. The injury was induced by application of vascular clips to the dura via a four-level T5–T8 laminectomy in mice. Oleuropein aglycone was administered in mice (100μg/kg, 40μg/kg, 20μg/kg, 10% ethanol, i.p.) 1h and 6h after the trauma. The treatment with oleuropein aglycone significantly decreased: (1) histological damage, (2) motor recovery, (3) nuclear factor (NF)-κB expression and IKB-α degradation, (4) protein kinase A (PKA) activity and expression, (5) pro-inflammatory cytokines production such as tumor necrosis factor (TNF-α) and interleukin-1β (IL-1β), 6) inducible nitric oxide synthase (iNOS) expression, (7) neutrophil infiltration, (8) lipid peroxidation, (9) nitrotyrosine and poly-ADP-ribose (PAR) formation, (10) glial cell-derived neurotrophic factor (GDNF) levels, (11) apoptosis (TUNEL staining, FAS ligand expression, Caspase 3, Bax and Bcl-2 expression). Thus, we propose that olive oil phenolic constituents such as oleuropein aglycone may be useful in the treatment of various inflammatory diseases.
Keywords: Spinal cord; Inflammation; Cytokines; Apoptosis; Oxidative stress; Olive oil
The prototypic pharmacogenetic drug debrisoquine is a substrate of the genetically polymorphic organic cation transporter OCT1
by Ali R. Saadatmand; Sina Tadjerpisheh; Jürgen Brockmöller; Mladen V. Tzvetkov (pp. 1427-1434).
Debrisoquine is a probe drug for in vivo phenotyping of human CYP2D6 metabolic activity. However, debrisoquine is positively charged under physiological conditions and it is unclear how it enters the hepatocytes to undergo CYP2D6 metabolism. We analysed whether debrisoquine is a substrate of the hepatic organic cation transporter OCT1 and whether drug–drug interactions at OCT1, or polymorphisms in OCT1 gene, affect debrisoquine uptake.Debrisoquine showed low carrier-independent membrane permeability ( Pe of 0.01×10−6cm/s in artificial PAMPA membranes) and strongly inhibited the uptake of the model OCT1 substrate MPP+ (IC50 of 6.2±0.8μM). Debrisoquine uptake was significantly increased in HEK293 cells overexpressing OCT1 compared to control cells. The OCT1-mediated uptake of debrisoquine followed Michaelis–Menten kinetics ( KM of 5.9±1.5μM and Vmax of 41.9±4.5pmol/min/mg protein) and was inhibited by known OCT1 inhibitors and by commonly used drugs. OCT1-mediated debrisoquine uptake was reduced or missing in cells expressing loss-of-function OCT1 isoforms. Deletion of Met420 or substitution of Arg61Cys or Gly401Ser reduced Vmax by 48, 63 and 91%, respectively, but did not affect the KM. The OCT1 isoforms carrying Cys88Arg or Gly465Arg substitutions completely lacked OCT1-mediated debrisoquine uptake.In conclusion, debrisoquine is a substrate of OCT1 and has the potential to be used as a phenotyping marker for OCT1 activity. Moreover, variations in debrisoquine metabolic phenotypes and their associations with diseases may be due not only to genetic variations CYP2D6, but also in OCT1.
Keywords: Organic cation transporter 1; OCT1; SLC22A1; Polymorphisms; Debrisoquine; Drug–drug interactions
The prototypic pharmacogenetic drug debrisoquine is a substrate of the genetically polymorphic organic cation transporter OCT1
by Ali R. Saadatmand; Sina Tadjerpisheh; Jürgen Brockmöller; Mladen V. Tzvetkov (pp. 1427-1434).
Debrisoquine is a probe drug for in vivo phenotyping of human CYP2D6 metabolic activity. However, debrisoquine is positively charged under physiological conditions and it is unclear how it enters the hepatocytes to undergo CYP2D6 metabolism. We analysed whether debrisoquine is a substrate of the hepatic organic cation transporter OCT1 and whether drug–drug interactions at OCT1, or polymorphisms in OCT1 gene, affect debrisoquine uptake.Debrisoquine showed low carrier-independent membrane permeability ( Pe of 0.01×10−6cm/s in artificial PAMPA membranes) and strongly inhibited the uptake of the model OCT1 substrate MPP+ (IC50 of 6.2±0.8μM). Debrisoquine uptake was significantly increased in HEK293 cells overexpressing OCT1 compared to control cells. The OCT1-mediated uptake of debrisoquine followed Michaelis–Menten kinetics ( KM of 5.9±1.5μM and Vmax of 41.9±4.5pmol/min/mg protein) and was inhibited by known OCT1 inhibitors and by commonly used drugs. OCT1-mediated debrisoquine uptake was reduced or missing in cells expressing loss-of-function OCT1 isoforms. Deletion of Met420 or substitution of Arg61Cys or Gly401Ser reduced Vmax by 48, 63 and 91%, respectively, but did not affect the KM. The OCT1 isoforms carrying Cys88Arg or Gly465Arg substitutions completely lacked OCT1-mediated debrisoquine uptake.In conclusion, debrisoquine is a substrate of OCT1 and has the potential to be used as a phenotyping marker for OCT1 activity. Moreover, variations in debrisoquine metabolic phenotypes and their associations with diseases may be due not only to genetic variations CYP2D6, but also in OCT1.
Keywords: Organic cation transporter 1; OCT1; SLC22A1; Polymorphisms; Debrisoquine; Drug–drug interactions
The prototypic pharmacogenetic drug debrisoquine is a substrate of the genetically polymorphic organic cation transporter OCT1
by Ali R. Saadatmand; Sina Tadjerpisheh; Jürgen Brockmöller; Mladen V. Tzvetkov (pp. 1427-1434).
Debrisoquine is a probe drug for in vivo phenotyping of human CYP2D6 metabolic activity. However, debrisoquine is positively charged under physiological conditions and it is unclear how it enters the hepatocytes to undergo CYP2D6 metabolism. We analysed whether debrisoquine is a substrate of the hepatic organic cation transporter OCT1 and whether drug–drug interactions at OCT1, or polymorphisms in OCT1 gene, affect debrisoquine uptake.Debrisoquine showed low carrier-independent membrane permeability ( Pe of 0.01×10−6cm/s in artificial PAMPA membranes) and strongly inhibited the uptake of the model OCT1 substrate MPP+ (IC50 of 6.2±0.8μM). Debrisoquine uptake was significantly increased in HEK293 cells overexpressing OCT1 compared to control cells. The OCT1-mediated uptake of debrisoquine followed Michaelis–Menten kinetics ( KM of 5.9±1.5μM and Vmax of 41.9±4.5pmol/min/mg protein) and was inhibited by known OCT1 inhibitors and by commonly used drugs. OCT1-mediated debrisoquine uptake was reduced or missing in cells expressing loss-of-function OCT1 isoforms. Deletion of Met420 or substitution of Arg61Cys or Gly401Ser reduced Vmax by 48, 63 and 91%, respectively, but did not affect the KM. The OCT1 isoforms carrying Cys88Arg or Gly465Arg substitutions completely lacked OCT1-mediated debrisoquine uptake.In conclusion, debrisoquine is a substrate of OCT1 and has the potential to be used as a phenotyping marker for OCT1 activity. Moreover, variations in debrisoquine metabolic phenotypes and their associations with diseases may be due not only to genetic variations CYP2D6, but also in OCT1.
Keywords: Organic cation transporter 1; OCT1; SLC22A1; Polymorphisms; Debrisoquine; Drug–drug interactions
Metabolomics reveals the metabolic map of procainamide in humans and mice
by Fei Li; Andrew D. Patterson; Kristopher W. Krausz; Bernhard Dick; Felix J. Frey; Frank J. Gonzalez; Jeffrey R. Idle (pp. 1435-1444).
Procainamide, a type I antiarrhythmic agent, is used to treat a variety of atrial and ventricular dysrhythmias. It was reported that long-term therapy with procainamide may cause lupus erythematosus in 25–30% of patients. Interestingly, procainamide does not induce lupus erythematosus in mouse models. To explore the differences in this side-effect of procainamide between humans and mouse models, metabolomic analysis using ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOFMS) was conducted on urine samples from procainamide-treated humans, CYP2D6-humanized mice, and wild-type mice. Thirteen urinary procainamide metabolites, including nine novel metabolites, derived from P450-dependent, FMO-dependent oxidations and acylation reactions, were identified and structurally elucidated. In vivo metabolism of procainamide in CYP2D6-humanized mice as well as in vitro incubations with microsomes and recombinant P450s suggested that human CYP2D6 plays a major role in procainamide metabolism. Significant differences in N-acylation and N-oxidation of the drug between humans and mice largely account for the interspecies differences in procainamide metabolism. Significant levels of the novel N-oxide metabolites produced by FMO1 and FMO3 in humans might be associated with the development of procainamide-induced systemic lupus erythematosus. Observations based on this metabolomic study offer clues to understanding procainamide-induced lupus in humans and the effect of P450s and FMOs on procainamide N-oxidation.
Keywords: Procainamide; Systemic lupus erythematosus; Metabolomics; N; -Oxidation; Ultra-performance liquid chromatography; Time-of-flight mass spectrometry
Metabolomics reveals the metabolic map of procainamide in humans and mice
by Fei Li; Andrew D. Patterson; Kristopher W. Krausz; Bernhard Dick; Felix J. Frey; Frank J. Gonzalez; Jeffrey R. Idle (pp. 1435-1444).
Procainamide, a type I antiarrhythmic agent, is used to treat a variety of atrial and ventricular dysrhythmias. It was reported that long-term therapy with procainamide may cause lupus erythematosus in 25–30% of patients. Interestingly, procainamide does not induce lupus erythematosus in mouse models. To explore the differences in this side-effect of procainamide between humans and mouse models, metabolomic analysis using ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOFMS) was conducted on urine samples from procainamide-treated humans, CYP2D6-humanized mice, and wild-type mice. Thirteen urinary procainamide metabolites, including nine novel metabolites, derived from P450-dependent, FMO-dependent oxidations and acylation reactions, were identified and structurally elucidated. In vivo metabolism of procainamide in CYP2D6-humanized mice as well as in vitro incubations with microsomes and recombinant P450s suggested that human CYP2D6 plays a major role in procainamide metabolism. Significant differences in N-acylation and N-oxidation of the drug between humans and mice largely account for the interspecies differences in procainamide metabolism. Significant levels of the novel N-oxide metabolites produced by FMO1 and FMO3 in humans might be associated with the development of procainamide-induced systemic lupus erythematosus. Observations based on this metabolomic study offer clues to understanding procainamide-induced lupus in humans and the effect of P450s and FMOs on procainamide N-oxidation.
Keywords: Procainamide; Systemic lupus erythematosus; Metabolomics; N; -Oxidation; Ultra-performance liquid chromatography; Time-of-flight mass spectrometry
Metabolomics reveals the metabolic map of procainamide in humans and mice
by Fei Li; Andrew D. Patterson; Kristopher W. Krausz; Bernhard Dick; Felix J. Frey; Frank J. Gonzalez; Jeffrey R. Idle (pp. 1435-1444).
Procainamide, a type I antiarrhythmic agent, is used to treat a variety of atrial and ventricular dysrhythmias. It was reported that long-term therapy with procainamide may cause lupus erythematosus in 25–30% of patients. Interestingly, procainamide does not induce lupus erythematosus in mouse models. To explore the differences in this side-effect of procainamide between humans and mouse models, metabolomic analysis using ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOFMS) was conducted on urine samples from procainamide-treated humans, CYP2D6-humanized mice, and wild-type mice. Thirteen urinary procainamide metabolites, including nine novel metabolites, derived from P450-dependent, FMO-dependent oxidations and acylation reactions, were identified and structurally elucidated. In vivo metabolism of procainamide in CYP2D6-humanized mice as well as in vitro incubations with microsomes and recombinant P450s suggested that human CYP2D6 plays a major role in procainamide metabolism. Significant differences in N-acylation and N-oxidation of the drug between humans and mice largely account for the interspecies differences in procainamide metabolism. Significant levels of the novel N-oxide metabolites produced by FMO1 and FMO3 in humans might be associated with the development of procainamide-induced systemic lupus erythematosus. Observations based on this metabolomic study offer clues to understanding procainamide-induced lupus in humans and the effect of P450s and FMOs on procainamide N-oxidation.
Keywords: Procainamide; Systemic lupus erythematosus; Metabolomics; N; -Oxidation; Ultra-performance liquid chromatography; Time-of-flight mass spectrometry
Structure-dependent activation of NR4A2 (Nurr1) by 1,1-bis(3′-indolyl)-1-(aromatic)methane analogs in pancreatic cancer cells
by Xi Li; Syng-Ook Lee; Stephen Safe (pp. 1445-1455).
NR4A2 (Nurr1) is an orphan nuclear receptor with no known endogenous ligands and is highly expressed in many cancer cell lines including Panc1 and Panc28 pancreatic cancer cells. Structure-dependent activation of NR4A2 by a series of 1,1-bis(3′-indolyl)-1-(aromatic)methane (C-DIM) analogs was determined in pancreatic cancer cells transfected with yeast GAL4-Nurr1 chimeras and a UASx5-luc reporter gene or constructs containing response elements that bind NR4A2. Among 23 different structural analogs, phenyl groups containing p-substituted trifluoromethyl, t-butyl, cyano, bromo, iodo and trifluoromethoxy groups were the most active compounds in transactivation assay. The p-bromophenyl analog (DIM-C-pPhBr) was used as a model for structure–activity studies among a series of ortho-, meta- and para-bromophenyl isomers and the corresponding indole 2- and N-methyl analogs. Results show that NR4A2 activation was maximal with the p-bromophenyl analog and methylation of the indole NH group abrogated activity. Moreover, using GAL4-Nurr1 (full length) or GAL-Nurr1-A/B and GAL4-Nurr1-(C-F) chimeras expressing N- and C-terminal domains of Nurr1, respectively, DIM-C-pPhBr activated all three constructs and these responses were differentially affected by kinase inhibitors. DIM-C-pPhBr also modulated expression of several Nurr1-regulated genes in pancreatic cancer cells including vasoactive intestinal peptide (VIP), and the immunohistochemical and western blot analyses indicated that DIM-C-pPhBr activates nuclear NR4A2.
Keywords: DIM analogs; NR4A2/Nurr1; Structure–activity
Structure-dependent activation of NR4A2 (Nurr1) by 1,1-bis(3′-indolyl)-1-(aromatic)methane analogs in pancreatic cancer cells
by Xi Li; Syng-Ook Lee; Stephen Safe (pp. 1445-1455).
NR4A2 (Nurr1) is an orphan nuclear receptor with no known endogenous ligands and is highly expressed in many cancer cell lines including Panc1 and Panc28 pancreatic cancer cells. Structure-dependent activation of NR4A2 by a series of 1,1-bis(3′-indolyl)-1-(aromatic)methane (C-DIM) analogs was determined in pancreatic cancer cells transfected with yeast GAL4-Nurr1 chimeras and a UASx5-luc reporter gene or constructs containing response elements that bind NR4A2. Among 23 different structural analogs, phenyl groups containing p-substituted trifluoromethyl, t-butyl, cyano, bromo, iodo and trifluoromethoxy groups were the most active compounds in transactivation assay. The p-bromophenyl analog (DIM-C-pPhBr) was used as a model for structure–activity studies among a series of ortho-, meta- and para-bromophenyl isomers and the corresponding indole 2- and N-methyl analogs. Results show that NR4A2 activation was maximal with the p-bromophenyl analog and methylation of the indole NH group abrogated activity. Moreover, using GAL4-Nurr1 (full length) or GAL-Nurr1-A/B and GAL4-Nurr1-(C-F) chimeras expressing N- and C-terminal domains of Nurr1, respectively, DIM-C-pPhBr activated all three constructs and these responses were differentially affected by kinase inhibitors. DIM-C-pPhBr also modulated expression of several Nurr1-regulated genes in pancreatic cancer cells including vasoactive intestinal peptide (VIP), and the immunohistochemical and western blot analyses indicated that DIM-C-pPhBr activates nuclear NR4A2.
Keywords: DIM analogs; NR4A2/Nurr1; Structure–activity
Structure-dependent activation of NR4A2 (Nurr1) by 1,1-bis(3′-indolyl)-1-(aromatic)methane analogs in pancreatic cancer cells
by Xi Li; Syng-Ook Lee; Stephen Safe (pp. 1445-1455).
NR4A2 (Nurr1) is an orphan nuclear receptor with no known endogenous ligands and is highly expressed in many cancer cell lines including Panc1 and Panc28 pancreatic cancer cells. Structure-dependent activation of NR4A2 by a series of 1,1-bis(3′-indolyl)-1-(aromatic)methane (C-DIM) analogs was determined in pancreatic cancer cells transfected with yeast GAL4-Nurr1 chimeras and a UASx5-luc reporter gene or constructs containing response elements that bind NR4A2. Among 23 different structural analogs, phenyl groups containing p-substituted trifluoromethyl, t-butyl, cyano, bromo, iodo and trifluoromethoxy groups were the most active compounds in transactivation assay. The p-bromophenyl analog (DIM-C-pPhBr) was used as a model for structure–activity studies among a series of ortho-, meta- and para-bromophenyl isomers and the corresponding indole 2- and N-methyl analogs. Results show that NR4A2 activation was maximal with the p-bromophenyl analog and methylation of the indole NH group abrogated activity. Moreover, using GAL4-Nurr1 (full length) or GAL-Nurr1-A/B and GAL4-Nurr1-(C-F) chimeras expressing N- and C-terminal domains of Nurr1, respectively, DIM-C-pPhBr activated all three constructs and these responses were differentially affected by kinase inhibitors. DIM-C-pPhBr also modulated expression of several Nurr1-regulated genes in pancreatic cancer cells including vasoactive intestinal peptide (VIP), and the immunohistochemical and western blot analyses indicated that DIM-C-pPhBr activates nuclear NR4A2.
Keywords: DIM analogs; NR4A2/Nurr1; Structure–activity
Antitumor activity of a novel STAT3 inhibitor and redox modulator in non-small cell lung cancer cells
by Xiaoying Liu; Wei Guo; Shuhong Wu; Li Wang; Ji Wang; Bingbing Dai; Edward S. Kim; John V. Heymach; Michael Wang; Luc Girard; John Minna; Jack A. Roth; Stephen G. Swisher; Bingliang Fang (pp. 1456-1464).
NSC-743380 is a novel STAT3 inhibitor that suppresses the growth of several NCI-60 cancer cell lines derived from different tissues and induces regression of xenograft tumors in vivo at various doses. To evaluate the antitumor activity of NSC-743380 in lung cancer cells, we analyzed the susceptibility of 50 NSCLC cell lines to this compound using cell viability assay. About 32% (16 of 50) of these cell lines were highly susceptible to this compound, with a 50% inhibitory concentration (IC50) of <1μM. In mechanistic studies, the increased numbers of apoptotic cells as well as increased PARP cleavage showed that cytotoxic effects correlate with apoptosis induction. Treatment with NSC-743380 inhibited transcription factor STAT3 activation and induced ROS production in sensitive human lung cancer cell lines but not in resistant cells. Blocking ROS generation with the antioxidant NDGA dramatically abolished NSC-743380-induced growth suppression and apoptosis, but had minimal effect on NSC-743380-induced STAT3 inhibition, suggesting that STAT3 inhibition is not caused by ROS production. Interestingly, knockdown of STAT3 with use of shSTAT3 induced ROS generation and suppressed tumor cell growth. Moreover, scavenging ROS induced by STAT3 inhibition also diminished antitumor activity of STAT3 inhibition. In vivo administration of NSC-743380 suppressed tumor growth and p-STAT3 in lung tumors. Our results indicate that NSC-743380 is a potent anticancer agent for lung cancer and that its apoptotic effects in lung cancer cells are mediated by induction of ROS through STAT3 inhibition.
Keywords: Abbreviations; H; 2; DCFDA; 2′,7′-dichlorofluorescein diacetate; JNK; c-Jun N-terminal kinase; HBEC3KT; normal human bronchial epithelial; NDGA; nordihydroguaiaretic acid; PI; propidium iodide; PARP; poly(ADP-ribose) polymerase; ROS; reactive oxygen species; STAT3; signal transducer and activator of transcription 3; SRB; sulforhodamine BDrug development; Lung Cancer; STAT3; Reactive oxygen species
Antitumor activity of a novel STAT3 inhibitor and redox modulator in non-small cell lung cancer cells
by Xiaoying Liu; Wei Guo; Shuhong Wu; Li Wang; Ji Wang; Bingbing Dai; Edward S. Kim; John V. Heymach; Michael Wang; Luc Girard; John Minna; Jack A. Roth; Stephen G. Swisher; Bingliang Fang (pp. 1456-1464).
NSC-743380 is a novel STAT3 inhibitor that suppresses the growth of several NCI-60 cancer cell lines derived from different tissues and induces regression of xenograft tumors in vivo at various doses. To evaluate the antitumor activity of NSC-743380 in lung cancer cells, we analyzed the susceptibility of 50 NSCLC cell lines to this compound using cell viability assay. About 32% (16 of 50) of these cell lines were highly susceptible to this compound, with a 50% inhibitory concentration (IC50) of <1μM. In mechanistic studies, the increased numbers of apoptotic cells as well as increased PARP cleavage showed that cytotoxic effects correlate with apoptosis induction. Treatment with NSC-743380 inhibited transcription factor STAT3 activation and induced ROS production in sensitive human lung cancer cell lines but not in resistant cells. Blocking ROS generation with the antioxidant NDGA dramatically abolished NSC-743380-induced growth suppression and apoptosis, but had minimal effect on NSC-743380-induced STAT3 inhibition, suggesting that STAT3 inhibition is not caused by ROS production. Interestingly, knockdown of STAT3 with use of shSTAT3 induced ROS generation and suppressed tumor cell growth. Moreover, scavenging ROS induced by STAT3 inhibition also diminished antitumor activity of STAT3 inhibition. In vivo administration of NSC-743380 suppressed tumor growth and p-STAT3 in lung tumors. Our results indicate that NSC-743380 is a potent anticancer agent for lung cancer and that its apoptotic effects in lung cancer cells are mediated by induction of ROS through STAT3 inhibition.
Keywords: Abbreviations; H; 2; DCFDA; 2′,7′-dichlorofluorescein diacetate; JNK; c-Jun N-terminal kinase; HBEC3KT; normal human bronchial epithelial; NDGA; nordihydroguaiaretic acid; PI; propidium iodide; PARP; poly(ADP-ribose) polymerase; ROS; reactive oxygen species; STAT3; signal transducer and activator of transcription 3; SRB; sulforhodamine BDrug development; Lung Cancer; STAT3; Reactive oxygen species
Antitumor activity of a novel STAT3 inhibitor and redox modulator in non-small cell lung cancer cells
by Xiaoying Liu; Wei Guo; Shuhong Wu; Li Wang; Ji Wang; Bingbing Dai; Edward S. Kim; John V. Heymach; Michael Wang; Luc Girard; John Minna; Jack A. Roth; Stephen G. Swisher; Bingliang Fang (pp. 1456-1464).
NSC-743380 is a novel STAT3 inhibitor that suppresses the growth of several NCI-60 cancer cell lines derived from different tissues and induces regression of xenograft tumors in vivo at various doses. To evaluate the antitumor activity of NSC-743380 in lung cancer cells, we analyzed the susceptibility of 50 NSCLC cell lines to this compound using cell viability assay. About 32% (16 of 50) of these cell lines were highly susceptible to this compound, with a 50% inhibitory concentration (IC50) of <1μM. In mechanistic studies, the increased numbers of apoptotic cells as well as increased PARP cleavage showed that cytotoxic effects correlate with apoptosis induction. Treatment with NSC-743380 inhibited transcription factor STAT3 activation and induced ROS production in sensitive human lung cancer cell lines but not in resistant cells. Blocking ROS generation with the antioxidant NDGA dramatically abolished NSC-743380-induced growth suppression and apoptosis, but had minimal effect on NSC-743380-induced STAT3 inhibition, suggesting that STAT3 inhibition is not caused by ROS production. Interestingly, knockdown of STAT3 with use of shSTAT3 induced ROS generation and suppressed tumor cell growth. Moreover, scavenging ROS induced by STAT3 inhibition also diminished antitumor activity of STAT3 inhibition. In vivo administration of NSC-743380 suppressed tumor growth and p-STAT3 in lung tumors. Our results indicate that NSC-743380 is a potent anticancer agent for lung cancer and that its apoptotic effects in lung cancer cells are mediated by induction of ROS through STAT3 inhibition.
Keywords: Abbreviations; H; 2; DCFDA; 2′,7′-dichlorofluorescein diacetate; JNK; c-Jun N-terminal kinase; HBEC3KT; normal human bronchial epithelial; NDGA; nordihydroguaiaretic acid; PI; propidium iodide; PARP; poly(ADP-ribose) polymerase; ROS; reactive oxygen species; STAT3; signal transducer and activator of transcription 3; SRB; sulforhodamine BDrug development; Lung Cancer; STAT3; Reactive oxygen species
The low intestinal and hepatic toxicity of hydrolyzed fumonisin B1 correlates with its inability to alter the metabolism of sphingolipids
by Bertrand Grenier; Ana-Paula F.L. Bracarense; Heidi Elisabeth Schwartz; Catherine Trumel; Anne-Marie Cossalter; Gerd Schatzmayr; Martine Kolf-Clauw; Wulf-Dieter Moll; Isabelle P. Oswald (pp. 1465-1473).
Fumonisins are mycotoxins frequently found as natural contaminants in maize, where they are produced by the plant pathogen Fusarium verticillioides. They are toxic to animals and exert their effects through mechanisms involving disruption of sphingolipid metabolism. Fumonisin B1 (FB1) is the predominant fumonisin in this family. FB1 is converted to its hydrolyzed analogs HFB1, by alkaline cooking (nixtamalization) or through enzymatic degradation. The toxicity of HFB1 is poorly documented especially at the intestinal level. The objectives of this study were to compare the toxicity of HFB1 and FB1 and to assess the ability of these toxins to disrupt sphingolipids biosynthesis. HFB1 was obtained by a deesterification of FB1 with a carboxylesterase. Piglets, animals highly sensitive to FB1, were exposed by gavage for 2 weeks to 2.8μmol FB1 or HFB1/kg body weight/day. FB1 induced hepatotoxicity as indicated by the lesion score, the level of several biochemical analytes and the expression of inflammatory cytokines. Similarly, FB1 impaired the morphology of the different segments of the small intestine, reduced villi height and modified intestinal cytokine expression. By contrast, HFB1 did not trigger hepatotoxicity, did not impair intestinal morphology and slightly modified the intestinal immune response. This low toxicity of HFB1 correlates with a weak alteration of the sphinganine/sphingosine ratio in the liver and in the plasma. Taken together, these data demonstrate that HFB1 does not cause intestinal or hepatic toxicity in the sensitive pig model and only slightly disrupts sphingolipids metabolism. This finding suggests that conversion to HFB1 could be a good strategy to reduce FB1 exposure.
Keywords: Abbreviations; AP; 1; aminopentol; b.w.; body weight; CerS; ceramide Synthase; FB; 1; FB; 2; FB; 3; fumonisin B; 1; B; 2; B; 3; GGT; Gamma-glutamyl transferase; HE; hematoxylin-eosin; HFB; 1; HFB; 2; HFB; 3; hydrolyzed fumonisin B; 1; B; 2; B; 3; HPLC-FLD; high performance liquid chromatography with postcolumn fluorescence derivatization; IFN; interferon; IL; interleukin; IPEC-1; intestinal porcine epithelial cell line; LC–MS; liquid chromatography–mass spectrometry; MHC-II; major histocompatibility complex class II; NOAEL; no observable adverse effect level; OPA; ortho-phthalaldehyde; ppm; parts per million (or mg/kg of feed); RPL32; ribosomal protein L32; Sa; sphinganine; SEM; standard error of mean; SI; small intestine; So; sphingosine; TCA; tricarballylic acids; TNF; tumor necrosis factorFumonisin; Hydrolyzed fumonisin; Sphingoid bases; Liver; Digestive tract
The low intestinal and hepatic toxicity of hydrolyzed fumonisin B1 correlates with its inability to alter the metabolism of sphingolipids
by Bertrand Grenier; Ana-Paula F.L. Bracarense; Heidi Elisabeth Schwartz; Catherine Trumel; Anne-Marie Cossalter; Gerd Schatzmayr; Martine Kolf-Clauw; Wulf-Dieter Moll; Isabelle P. Oswald (pp. 1465-1473).
Fumonisins are mycotoxins frequently found as natural contaminants in maize, where they are produced by the plant pathogen Fusarium verticillioides. They are toxic to animals and exert their effects through mechanisms involving disruption of sphingolipid metabolism. Fumonisin B1 (FB1) is the predominant fumonisin in this family. FB1 is converted to its hydrolyzed analogs HFB1, by alkaline cooking (nixtamalization) or through enzymatic degradation. The toxicity of HFB1 is poorly documented especially at the intestinal level. The objectives of this study were to compare the toxicity of HFB1 and FB1 and to assess the ability of these toxins to disrupt sphingolipids biosynthesis. HFB1 was obtained by a deesterification of FB1 with a carboxylesterase. Piglets, animals highly sensitive to FB1, were exposed by gavage for 2 weeks to 2.8μmol FB1 or HFB1/kg body weight/day. FB1 induced hepatotoxicity as indicated by the lesion score, the level of several biochemical analytes and the expression of inflammatory cytokines. Similarly, FB1 impaired the morphology of the different segments of the small intestine, reduced villi height and modified intestinal cytokine expression. By contrast, HFB1 did not trigger hepatotoxicity, did not impair intestinal morphology and slightly modified the intestinal immune response. This low toxicity of HFB1 correlates with a weak alteration of the sphinganine/sphingosine ratio in the liver and in the plasma. Taken together, these data demonstrate that HFB1 does not cause intestinal or hepatic toxicity in the sensitive pig model and only slightly disrupts sphingolipids metabolism. This finding suggests that conversion to HFB1 could be a good strategy to reduce FB1 exposure.
Keywords: Abbreviations; AP; 1; aminopentol; b.w.; body weight; CerS; ceramide Synthase; FB; 1; FB; 2; FB; 3; fumonisin B; 1; B; 2; B; 3; GGT; Gamma-glutamyl transferase; HE; hematoxylin-eosin; HFB; 1; HFB; 2; HFB; 3; hydrolyzed fumonisin B; 1; B; 2; B; 3; HPLC-FLD; high performance liquid chromatography with postcolumn fluorescence derivatization; IFN; interferon; IL; interleukin; IPEC-1; intestinal porcine epithelial cell line; LC–MS; liquid chromatography–mass spectrometry; MHC-II; major histocompatibility complex class II; NOAEL; no observable adverse effect level; OPA; ortho-phthalaldehyde; ppm; parts per million (or mg/kg of feed); RPL32; ribosomal protein L32; Sa; sphinganine; SEM; standard error of mean; SI; small intestine; So; sphingosine; TCA; tricarballylic acids; TNF; tumor necrosis factorFumonisin; Hydrolyzed fumonisin; Sphingoid bases; Liver; Digestive tract
The low intestinal and hepatic toxicity of hydrolyzed fumonisin B1 correlates with its inability to alter the metabolism of sphingolipids
by Bertrand Grenier; Ana-Paula F.L. Bracarense; Heidi Elisabeth Schwartz; Catherine Trumel; Anne-Marie Cossalter; Gerd Schatzmayr; Martine Kolf-Clauw; Wulf-Dieter Moll; Isabelle P. Oswald (pp. 1465-1473).
Fumonisins are mycotoxins frequently found as natural contaminants in maize, where they are produced by the plant pathogen Fusarium verticillioides. They are toxic to animals and exert their effects through mechanisms involving disruption of sphingolipid metabolism. Fumonisin B1 (FB1) is the predominant fumonisin in this family. FB1 is converted to its hydrolyzed analogs HFB1, by alkaline cooking (nixtamalization) or through enzymatic degradation. The toxicity of HFB1 is poorly documented especially at the intestinal level. The objectives of this study were to compare the toxicity of HFB1 and FB1 and to assess the ability of these toxins to disrupt sphingolipids biosynthesis. HFB1 was obtained by a deesterification of FB1 with a carboxylesterase. Piglets, animals highly sensitive to FB1, were exposed by gavage for 2 weeks to 2.8μmol FB1 or HFB1/kg body weight/day. FB1 induced hepatotoxicity as indicated by the lesion score, the level of several biochemical analytes and the expression of inflammatory cytokines. Similarly, FB1 impaired the morphology of the different segments of the small intestine, reduced villi height and modified intestinal cytokine expression. By contrast, HFB1 did not trigger hepatotoxicity, did not impair intestinal morphology and slightly modified the intestinal immune response. This low toxicity of HFB1 correlates with a weak alteration of the sphinganine/sphingosine ratio in the liver and in the plasma. Taken together, these data demonstrate that HFB1 does not cause intestinal or hepatic toxicity in the sensitive pig model and only slightly disrupts sphingolipids metabolism. This finding suggests that conversion to HFB1 could be a good strategy to reduce FB1 exposure.
Keywords: Abbreviations; AP; 1; aminopentol; b.w.; body weight; CerS; ceramide Synthase; FB; 1; FB; 2; FB; 3; fumonisin B; 1; B; 2; B; 3; GGT; Gamma-glutamyl transferase; HE; hematoxylin-eosin; HFB; 1; HFB; 2; HFB; 3; hydrolyzed fumonisin B; 1; B; 2; B; 3; HPLC-FLD; high performance liquid chromatography with postcolumn fluorescence derivatization; IFN; interferon; IL; interleukin; IPEC-1; intestinal porcine epithelial cell line; LC–MS; liquid chromatography–mass spectrometry; MHC-II; major histocompatibility complex class II; NOAEL; no observable adverse effect level; OPA; ortho-phthalaldehyde; ppm; parts per million (or mg/kg of feed); RPL32; ribosomal protein L32; Sa; sphinganine; SEM; standard error of mean; SI; small intestine; So; sphingosine; TCA; tricarballylic acids; TNF; tumor necrosis factorFumonisin; Hydrolyzed fumonisin; Sphingoid bases; Liver; Digestive tract
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