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Biochemical Pharmacology (v.79, #12)

Graphical Abstracts Contents (pp. e1-e4).

Editorial Advisory Board (pp. iii).

RLIP76: A versatile transporter and an emerging target for cancer therapy by Rit Vatsyayan; Poorna Chandra Rao Lelsani; Sanjay Awasthi; Sharad S. Singhal (pp. 1699-1705).

In the last few years, extensive research has been made to elucidate the functional significance of RLIP76. The resulting novel breakthroughs have helped us understand its transport and signaling functions. RLIP76 is a ubiquitously expressed, key stress-defensive, anti-apoptotic, multi-functional protein that transports glutathione-conjugates of electrophilic compounds, thus controlling the intracellular concentration of pro-apoptotic oxidized lipid byproducts and other xenobiotics such as chemotherapeutic agents. These properties place RLIP76 at a very important position in the hierarchy of the stress defense mechanism adopted by the cell. Selective over-expression of RLIP76 in malignant cells of diverse origin is one of the possible mechanisms by which these cells overcome chemotherapy and radiation induced oxidative damage. RLIP76 has also been shown to be an effective transporter of many conventional chemotherapeutic drugs. Such transport, if inhibited, can lead to increased cellular accumulation of drugs which in turn translates to enhanced drug sensitivity. Recent studies have shown that inhibition and/or depletion of RLIP76 by antibodies, siRNA, or antisense can lead to drastic and sustained regression of lung, kidney, melanoma, colon, and prostate cancer xenografts with no observed recurrence of tumors. All these findings converge on the fact that such inhibition/depletion of RLIP76 can be used clinically to terminate cancer growth and progression. In the present review, we will discuss the role of RLIP76 as a multi-drug transporter, its involvement in cancer, and the prospects of using RLIP76 inhibition as an emerging treatment for cancer.

Keywords: Abbreviations; RLIP76; Ral-interacting protein; GSH; glutathione; GS-E; glutathione-electrophile conjugate; DNP-SG; dinitrophenyl S-glutathione; DOX; doxorubicin; 4-HNE; 4-hydroxy-; t; -nonenal; POB1; partner of RLIP76; Cdc2; cell cycle regulatory protein CDK1; HLBEC; human lung bronchio-epithelial cells; HUVEC; human umbilical vein endothelial cell; HAVSMC; human aortic vascular smooth muscle cellsRLIP76; Cancer; Drug resistance; Radiation resistance; Xenografts; Chemotherapy

Progenitor cell-derived smooth muscle cells in vascular disease by Augusto Orlandi; Martin Bennett (pp. 1706-1713).

Accumulation of vascular smooth muscle cells (VSMCs) in the tunica intima plays a major role in the pathogenesis of atherosclerosis and restenosis following endovascular procedures. Arterial VSMCs are heterogeneous even in the normal vessel wall and display different phenotypes in physiological and pathological conditions. In the classical paradigm, vascular wall injury induces VSMC de-differentiation, proliferation and migration from the media into the intima in response to growth factors and proteolytic agents. Accordingly, VSMCs in atherosclerotic plaques and in restenosis display a de-differentiated or ‘synthetic’ phenotype compared to a ‘contractile’ phenotype in the normal media. In contrast, recent studies have identified bone marrow and peripheral blood-derived endothelial and VSMC progenitors that may contribute to intimal formation in atherosclerosis, after arterial injury and in transplant atherosclerosis. The precise frequency of these bone marrow-derived vascular precursor cells is controversial and their role is unknown. In addition, additional data support the presence of a resident progenitor cell subpopulation and its involvement in the response of the adult arterial wall to damage or ischemia. This review will examine the evidence for and the putative role of progenitor cell-derived VSMCs in arterial disease, a necessary prerequisite before deciding whether progenitor cells are therapeutic targets in vascular disease.

Keywords: Arterial wall; Atherosclerosis; Restenosis; Progenitors; Smooth muscle cells; Stem cellsAbbreviations; VSMCs; vascular smooth muscle cells, stem cell antigen-1 (sca-1); α-SMA; alpha smooth muscle cell actin; SMMHC; smooth muscle cell myosin heavy chain

Suppression of PMA-induced tumor cell invasion by dihydroartemisinin via inhibition of PKCα/Raf/MAPKs and NF-κB/AP-1-dependent mechanisms by Yong Pil Hwang; Hyo Jeong Yun; Hyung Gyun Kim; Eun Hee Han; Gye Won Lee; Hye Gwang Jeong (pp. 1714-1726).

Proposed signal transduction pathways by which PMA induced and DHA-inhibited invasion and migration of HT-1080 cells.Dihydroartemisinin (DHA), a semi-synthetic derivative of artemisinin, has recently been shown to possess antitumor activity in various cancer cells. However, the effects of DHA in preventing the invasion of cancer cells have not been studied. In the present study, we investigated the inhibitory effects of DHA on tumor invasion and migration and the possible mechanisms involved using human fibrosarcoma HT-1080 cells. DHA reduced PMA-induced activation of MMP-9 and MMP-2 and further inhibited cell invasion and migration. DHA suppressed PMA-enhanced expression of MMP-9 protein, mRNA, and transcriptional activity through suppressing NF-κB and AP-1 activation without changing the level of tissue inhibitor of metalloproteinase (TIMP)-1. DHA also reduced PMA-enhanced MMP-2 expression by suppressing membrane-type 1 MMP (MT1-MMP), but did not alter TIMP-2 levels. DHA-inhibited PMA-induced NF-κB and c-Jun nuclear translocation, which are upstream of PMA-induced MMP-9 expression and invasion. Furthermore, DHA strongly repressed the PMA-induced phosphorylation of Raf/ERK and JNK, which are dependent on the PKCα pathway. In conclusion, we demonstrated that the anti-invasive effects of DHA may occur through inhibition of PKCα/Raf/ERK and JNK phosphorylation and reduction of NF-κB and AP-1 activation, leading to down-regulation of MMP-9 expression. The data presented show that DHA is an effective anti-metastatic agent that functions by down-regulating MMP-9 gene expression.

Keywords: Dihydroartemisinin; Matrix metalloproteinases-9; PKCα; Invasion; Antitumor activity

Inhibition of calcium-independent phospholipase A₂ activates p38 MAPK signaling pathways during cytostasis in prostate cancer cells by Bin Sun; Xiaoling Zhang; Christopher Yonz; Brian S. Cummings (pp. 1727-1735).

Role of iPLA₂ in prostate cancer cell signaling and growth.The p38 mitogen-activated protein kinase (MAPK) signaling pathways activated during cytostasis induced by Ca²⁺-independent phospholipase A₂ (iPLA₂) inhibition in prostate cancer cells were investigated. iPLA₂ inhibition using siRNA, or the selective inhibitor bromoenol lactone (BEL) and it's enantiomers, decreased growth in LNCaP (p53 positive) and PC-3 (p53 negative) human prostate cancer cells. Decreased cell growth correlated to time- and concentration-dependent activation of the mitogen-activated protein kinase p38 in both cell lines. Inhibition of cytosolic iPLA₂β using S-BEL, induced significantly higher levels of P-p53, p53, p21 and P-p38 expression than inhibition of microsomal iPLA₂γ using R-BEL. Inhibition of p38 using SB202190 or SB203580 inhibited BEL-induced increases in P-p53 (ser15), p53 and p21, and altered the number of cells in G1 in LNCaP cells, and S-phase in PC-3 cells. BEL treatment also induced reactive species in PC-3 and LNCaP cells, which was partially reversed by pretreatment with N-acetyl-cysteine (NAC). NAC subsequently inhibited BEL-induced activation of p38 and p53 in LNCaP cells. In addition, treatment of cells with NAC partially reversed the effect of BEL on cell growth and preserved cell morphology. Collectively, these data demonstrate the novel findings that iPLA₂ inhibition activates p38 by inducing reactive species, and further suggest that this signaling kinase is involved in p53 activation, cell cycle arrest and cytostasis.

Keywords: Abbreviations; ASA; ascorbic acid; ATR; ataxia telangiectasia and rad-3-related kinase; BEL; bromoenol lactone; CM-H2DCFDA; 5-(and-6)-chloromethyl-2,7-dichlorodihydrofluorescein diacetate acetyl ester; ER; endoplasmic reticulum; iPLA; ₂; Ca; ²⁺; -independent phospholipase A; ₂; MAPK; mitogen-activated protein kinases; mTOR; mammalian target of rapamycin; NAC; N-acetyl-cysteine; PBS; phosphate-buffered saline; PI; propidium iodide; RS; reactive speciesCa; ²⁺; -independent phospholipase A; ₂; Mitogen-activated protein kinases; p38; Reactive species; Prostate cancer

Mode of action of Nifurtimox and N-oxide-containing heterocycles against Trypanosoma cruzi: Is oxidative stress involved? by Mariana Boiani; Lucia Piacenza; Paola Hernández; Lucia Boiani; Hugo Cerecetto; Mercedes González; Ana Denicola (pp. 1736-1745).

No production of ROS/RNS was observed by Trypanosoma cruzi epimastigotes treated with several N-oxide-containing heterocycles and Nifurtimox at 25μM.Chagas disease is caused by the trypanosomatid parasite Trypanosoma cruzi and threatens millions of lives in South America. As other neglected diseases there is almost no research and development effort by the pharmaceutical industry and the treatment relies on two drugs, Nifurtimox and Benznidazole, discovered empirically more than three decades ago. Nifurtimox, a nitrofurane derivative, is believed to exert its biological activity through the bioreduction of the nitro-group to a nitro-anion radical which undergoes redox-cycling with molecular oxygen. This hypothesis is generally accepted, although arguments against it have been presented. In the present work we studied the ability of Nifurtimox and five N-oxide-containing heterocycles to induce oxidative stress in T. cruzi. N-Oxide-containing heterocycles represent a promising group of new trypanosomicidal agents and their mode of action is not completely elucidated. The results here obtained argue against the oxidative stress hypothesis almost for all the studied compounds, including Nifurtimox. A significant reduction in the level of parasitic low-molecular-weight thiols was observed after Nifurtimox treatment; however, it was not linked to the production of reactive oxidant species. Besides, redox-cycling is only observed at high Nifurtimox concentrations (>400μM), two orders of magnitude higher than the concentration required for anti-proliferative activity (5μM). Our results indicate that an increase in oxidative stress is not the main mechanism of action of Nifurtimox. Among the studied N-oxide-containing heterocycles, benzofuroxan derivatives strongly inhibited parasite dehydrogenase activity and affected mitochondrial membrane potential. The indazole derivative raised intracellular oxidants production, but it was the least effective as anti- T. cruzi.

Keywords: Nifurtimox; N; -Oxide-containing heterocycles; Trypanosoma cruzi; Oxidative stress

Different contribution of BH3-only proteins and caspases to doxorubicin-induced apoptosis in p53-deficient leukemia cells by Nuria López-Royuela; Patricia Pérez-Galán; Patricia Galán-Malo; Victor J. Yuste; Alberto Anel; Santos A. Susín; Javier Naval; Isabel Marzo (pp. 1746-1758).

Bcl-2 family proteins are key regulators of the intrinsic apoptotic pathway, either facilitating (Bax, Bak, BH3-only) or inhibiting (Bcl-2, Bcl-x_L, Mcl-1, A1) mitochondrial release of apoptogenic factors. The role of caspases in this process is a matter of controversy. We have analyzed the relative contribution of caspases and Bcl-2 family of proteins in the induction phase of apoptosis triggered by doxorubicin in two p53-deficient leukemia cell lines, Jurkat and U937. First, we have found that caspases are dispensable for the induction phase of doxorubicin-induced apoptosis in both cell lines but they are needed to speed up the execution phase in Jurkat cells, not expressing Bax. Thus, down-regulation of Bak expression by siRNA significantly prevented doxorubicin-induced apoptosis in Jurkat but not in U937 cells. Reduction of Mcl-1 protein levels with siRNA increased sensitivity to apoptosis in both cell lines. Moreover, our results indicate that the contribution of BH3-only proteins to apoptosis is cell line specific. In Jurkat cells simultaneous silencing of Bim and PUMA was necessary to reduce doxorubicin-induced apoptosis. In U937 cells silencing of Bim or Noxa reduced sensitivity to doxorubicin. Immunoprecipitation experiments discarded an interaction between Mcl-1 and Bak in both cell lines and underscored the role of Bim and PUMA as mediators of Bax/Bak activation.

Keywords: Abbreviations; AIF; apoptosis-inducing factor; Δ; Ψ; _m; mitochondrial transmembrane potential; DiOC; ₆; (3); 3,3′-dihexyloxa-carbocyanine iodide; PS; phosphatidylserineDoxorubicin; Caspases; Bcl-2 family proteins; BH3-only proteins; AIF

Modulation of stress genes expression profile by nitric oxide-releasing aspirin in Jurkat T leukemia cells by Niharika Nath; Mitali Chattopadhyay; Ravinder Kodela; Song Tian; Peter Vlismas; Daniel Boring; James A. Crowell; Khosrow Kashfi (pp. 1759-1771).

NO-donating aspirin (NO-ASA, para isomer) has been reported to exhibit strong growth inhibitory effect in Jurkat T-acute lymphoblastic leukemia (T-ALL) cells mediated in part by β-catenin degradation and caspase activation, but the mechanism(s) still remains unclear. In this study, DNA oligoarrays with 263 genes were used to examine the gene expression profiles relating to stress and drug metabolism, and characterize the stress responses at IC₅₀ and subIC₅₀ concentrations of p-NO-ASA (20 and 10μM, respectively) in Jurkat T cells. A total of 22 genes related to heat shock response, apoptosis signaling, detoxifiers and Phase II enzymes, and regulators of cell growth were altered in expression by array analysis based on the expression fold change criteria of ≥1.5-fold or ≤0.65-fold. Real time quantitative RT-PCR confirmed that 20μM p-NO-ASA strongly upregulated the mRNA levels of two heat shock genes HSPA1A (41.5±7.01-fold) and HSPA6 (100.4±8.11-fold), and FOS (16.2±3.2-fold), moderately upregulated HSPH1 (1.71±0.43-fold) , FMO4 (4.5±1.67-fold), CASP9 (1.77±0.03-fold), DDIT3 (5.6±0.51-fold), and downregulated NF- κB1 (0.54±0.01-fold) and CCND1 (0.69±0.06-fold). Protein levels of Hsp70, the product of HSPA1A, and fos were increased in p-NO-ASA-treated Jurkat T and HT-29 colon cancer cells in a dose-dependent manner. Silencing of Hsp70 enhanced the growth inhibitory effect of p-NO-ASA at low concentrations. The altered gene expression patterns by NO-ASA in Jurkat T cells suggest mechanisms for carcinogen metabolism, anti-proliferative activity and possible chemoprotective activity in T-ALL.

Keywords: NO-releasing aspirin; Stress; Heat shock protein 70; β-Catenin; COX-2; Anticancer

An RNA-directed nucleoside anti-metabolite, 1-(3-C-ethynyl-beta-d-ribo-pentofuranosyl)cytosine (ECyd), elicits antitumor effect via TP53-inducedGlycolysis and Apoptosis Regulator (TIGAR) downregulation by Vivian Wai Yan Lui; Cecilia Pik Yuk Lau; Crystal Sao Fong Cheung; Kakiu Ho; Margaret Heung Ling Ng; Suk Hang Cheng; Bo Hong; Sai-Wah Tsao; Chi Man Tsang; Kenny Ieng Kit Lei; Yasundo Yamasaki; Akira Mita; Anthony T.C. Chan (pp. 1772-1780).

1-(3-C-ethynyl-beta-d-ribo-pentofuranosyl)cytosine (ECyd) is a ribose-modified nucleoside analog of cytidine with potent anticancer activity in several cancers. The main antitumor mechanism of this promising RNA-directed nucleoside anti-metabolite is efficient blockade of RNA synthesis in cancer cells. Here, we examined the therapeutic potential of this RNA-directed anti-metabolite in in vitro models of nasopharyngeal cancer (NPC). In a panel of 6 NPC cell lines, ECyd effectively inhibited cellular proliferation at nM concentrations (IC₅₀:∼13–44nM). Moreover, cisplatin-resistant NPC cells were highly sensitive to ECyd (at nM concentration). The ECyd-mediated growth inhibition was associated with G₂/M cell cycle arrest, PARP cleavage (a hallmark of apoptosis) and Bcl-2 downregulation, indicating induction of apoptosis by ECyd in NPC cells. Unexpectedly, ECyd-induced significant downregulation of TIGAR, a newly described dual regulator of apoptosis and glycolysis. More importantly, this novel action of ECyd on TIGAR was accompanied by marked depletion of NADPH, the major reducing power critically required for cell proliferation and survival. We hypothesized that ECyd-induced TIGAR downregulation was crucially involved in the antitumor activity of ECyd. Indeed, overexpression of TIGAR was able to rescue NPC cells from ECyd-induced growth inhibition, demonstrating a novel mechanistic action of ECyd on TIGAR. We demonstrated for the first time that an RNA-directed nucleoside analog, ECyd, exerts its antitumor activity via downregulation of a novel regulator of apoptosis, TIGAR. Moreover, ECyd may represent a novel therapy for NPC.

Keywords: Abbreviations; ECyd; 1-(3-C-ethynyl-beta-; d; -ribo-pentofuranosyl)cytosine; NPC; nasopharyngeal carcinoma; LMP1; latent membrane protein 1; TIGAR; TP53; -induced glycolysis and apoptosis regulator; DRB; 5,6-dichloro-1-β-; d; -ribofuranosylbenzimidazole; ARC; 4-amino-6-hydrazino-7-beta-; d; -ribofuranosyl-7H-pyrrolo(2,3-d)-pyrimidine-5-carboxamide; EUrd; 1-(3-C-ethynyl-beta-; d; -ribopentofuranosyl)uracilECyd; Antitumor mechanism; TIGAR; NADPH; NPC

Remarkable interference with telomeric function by a G-quadruplex selective bisantrene regioisomer by Marco Folini; Claudia Pivetta; Giuseppe Zagotto; Cinzia De Marco; Manlio Palumbo; Nadia Zaffaroni; Claudia Sissi (pp. 1781-1790).

The use of small molecules able to induce and stabilize selected G-quadruplex arrangements can cause telomerase inhibition and telomere dysfunction in cancer cells, thus providing very selective therapeutic approaches. Effective stabilizers usually comprise a planar aromatic portion to grant effective stacking onto the G-quartet and positively charged side chains to exploit the highly negative charge density on the quadruplex grooves. Since the relative position of these two pharmacophoric moieties is expected to play an important role in DNA folding stabilization, we evaluated a series of anthracene derivatives substituted with one or two 4,5-dihydro-1 H-imidazol-2-yl-hydrazonic groups (the bisantrene side chain) at different positions of the aromatic system. Indeed, the various regioisomers showed distinct binding affinities for telomeric G-quadruplex, and the most effective was the 1,5 and 1,7 bis-substituted analogues. On turn, the 1,8 regioisomer was poorly effective. Interestingly, G-quadruplex binding is clearly related to telomerase inhibition in this class of compounds, thus confirming their ability to shift the nucleic acid conformational equilibrium upon binding and consequently produce interference with the telomere processing enzyme. Additionally, the 1,5 regioisomer was shown to inhibit telomerase activity at lower concentrations than those required to reduce tumor cell proliferation. Comparative analysis of drug effects in telomerase-positive and telomerase-negative cancer cells showed consistent cell growth impairment, as a consequence of activation of the senescence pathway, which was mainly attributable to anthracene-mediated telomere dysfunction.

Keywords: Alternative lengthening of telomeres; Bisantrene; G-quadruplex; Telomerase; Telomere

d-Serine modulates neurogenic relaxation in rat corpus cavernosum by Mehdi Ghasemi; Fatemeh Rezania; Jackie Lewin; Kevin P. Moore; Ali R. Mani (pp. 1791-1796).

d-Serine, an endogenous co-agonist for the N-methyl-d-aspartate (NMDA) receptor in mammals, is synthesized froml-serine by serine racemase. Although much attention has been focused on the role ofd-serine within the central nervous system, the physiological role ofd-serine in peripheral nerves such as corpus cavernosal nerves has not been investigated. The present study was aimed to study the expression, cellular localization and function of serine racemase/d-serine system in isolated rat corpus cavernosum. Reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis showed the expression of serine racemase in rat corpus cavernosum. Immunogold electron microscopy demonstrated the cellular localization of serine racemase in the cavernosal nerves’ membrane of the tissue. The organ bath studies on isolated rat corpus cavernosum showed thatd-serine increases the non-adrenergic non-cholinergic neurogenic relaxation of isolated rat corpus cavernosum in vitro. This effect ofd-serine was inhibited by a variety of NMDA receptor antagonists (ketamine, MK 801 and ifenprodil), suggesting that NMDA receptors are involved in the effects ofd-serine on the neurogenic relaxation of corporal tissue strips. These observations provide the first evidence for the role ofd-serine in modulating the neurogenic relaxation of rat corpus cavernosum, and may open new therapeutic avenues for the treatment of impotence.

Keywords: d; -Serine; Serine racemase; d; -Amino acid oxidase; Corpus cavernosum; Non-adrenergic non-cholinergic (NANC) relaxation; Rat

The intestinal antiinflammatory agent glycomacropeptide has immunomodulatory actions on rat splenocytes by Pilar Requena; Raquel González; Rocío López-Posadas; Ana Abadía-Molina; María Dolores Suárez; Antonio Zarzuelo; Fermín Sánchez de Medina; Olga Martínez-Augustin (pp. 1797-1804).

Bovine glycomacropeptide (GMP) is an immunologically active milk peptide that is a part of the normal human diet. GMP has therapeutic value in preclinical models of intestinal inflammation, and its mechanism may be related to effects on lymphocytes. This study focuses on the actions of GMP on rat splenocytes in vitro and in vivo. Bovine serum albumin and lactoferrin were used for comparative purposes. GMP (0.01–0.1mgmL⁻¹) enhanced Concanavalin A (ConA) evoked but not basal splenocyte proliferation. At 1mgmL⁻¹ GMP lost this effect but augmented basal TNF-α secretion and also iNOS and COX2 expression. IFN-γ, IL-2 and IL-17 were not affected by GMP in quiescent splenocytes, but IL-10 was augmented at all concentrations tested. On the other hand, GMP produced a marked inhibitory effect (70%) on IFN-γ secretion and to a lower extent (50%) also on TNF-α. GMP was shown to block STAT4 but not IκB-α phosphorylation. The Treg marker Foxp3 was markedly upregulated by GMP. Bovine serum albumin had some effects on splenocyte function which were of lower magnitude and not entirely coincidental, while lactoferrin had a strong antiproliferative effect, as expected, indicating a specific effect of GMP. When administered for 3 days to normal Wistar rats, GMP reproduced the Foxp3 induction effect observed previously in vitro. This was observed in splenocytes but not in thymocytes, and only when administered by the oral rather than the intraperitoneal route. Thus our results support the hypothesis that GMP may limit intestinal inflammation acting at least in part on lymphocytes.

Keywords: Glycomacropeptide; STAT4; Inflammatory bowel disease; Th1; Treg; Foxp3

Endogenous N-acyl-dopamines induce COX-2 expression in brain endothelial cells by stabilizing mRNA through a p38 dependent pathway by Carmen M. Navarrete; Moisés Pérez; Amaya García de Vinuesa; Juan A. Collado; Bernd L. Fiebich; Marco A. Calzado; Eduardo Muñoz (pp. 1805-1814).

Schematic diagram describing the effect of endogenous N-acyl-dopamines on COX-2 expression in brain endothelial cells.Cerebral microvascular endothelial cells play an active role in maintaining cerebral blood flow, microvascular tone and blood brain barrier (BBB) functions. Endogenous N-acyl-dopamines like N-arachidonoyl-dopamine (NADA) and N-oleoyl-dopamine (OLDA) have been recently identified as a new class of brain neurotransmitters sharing endocannabinoid and endovanilloid biological activities. Endocannabinoids are released in response to pathogenic insults and may play an important role in neuroprotection. In this study we demonstrate that NADA differentially regulates the release of PGE₂ and PGD₂ in the microvascular brain endothelial cell line, b.end5. We found that NADA activates a redox-sensitive p38 MAPK pathway that stabilizes COX-2 mRNA resulting in the accumulation of the COX-2 protein, which depends on the dopamine moiety of the molecule and that is independent of CB₁ and TRPV1 activation. In addition, NADA inhibits the expression of mPGES-1 and the release of PGE₂ and upregulates the expression of L-PGD synthase enhancing PGD₂ release. Hence, NADA and other molecules of the same family might be included in the group of lipid mediators that could prevent the BBB injury under inflammatory conditions and our findings provide new mechanistic insights into the anti-inflammatory activities of NADA in the central nervous system and its potential to design novel therapeutic strategies to manage neuroinflammatory diseases.

Keywords: N; -Arachidonoyl-dopamine; COX-2; p38 MAPK; mPGES-1; Endothelial cells

Falcarinol is a covalent cannabinoid CB₁ receptor antagonist and induces pro-allergic effects in skin by Marco Leonti; Laura Casu; Stefan Raduner; Filippo Cottiglia; Costantino Floris; Karl-Heinz Altmann; Jürg Gertsch (pp. 1815-1826).

The skin irritant polyyne falcarinol (panaxynol, carotatoxin) is found in carrots, parsley, celery, and in the medicinal plant Panax ginseng. In our ongoing search for new cannabinoid (CB) receptor ligands we have isolated falcarinol from the endemic Sardinian plant Seseli praecox.We show that falcarinol exhibits binding affinity to both human CB receptors but selectively alkylates the anandamide binding site in the CB₁ receptor ( K_i=594nM), acting as covalent inverse agonist in CB₁ receptor-transfected CHO cells. Given the inherent instability of purified falcarinol we repeatedly isolated this compound for biological characterization and one new polyyne was characterized. In human HaCaT keratinocytes falcarinol increased the expression of the pro-allergic chemokines IL-8 and CCL2/MCP-1 in a CB₁ receptor-dependent manner. Moreover, falcarinol inhibited the effects of anandamide on TNF-alpha stimulated keratinocytes. In vivo, falcarinol strongly aggravated histamine-induced oedema reactions in skin prick tests. Both effects were also obtained with the CB₁ receptor inverse agonist rimonabant, thus indicating the potential role of the CB₁ receptor in skin immunopharmacology. Our data suggest anti-allergic effects of anandamide and that falcarinol-associated dermatitis is due to antagonism of the CB₁ receptor in keratinocytes, leading to increased chemokine expression and aggravation of histamine action.

Keywords: Falcarinol; Skin irritation; CB1 cannabinoid receptor; Anandamide; Endocannabinoid system

The dopamine receptor D2 agonist bromocriptine inhibits glucose-stimulated insulin secretion by direct activation of the α2-adrenergic receptors in beta cells by J.E. de Leeuw van Weenen; E.T. Parlevliet; P. Maechler; L.M. Havekes; J.A. Romijn; D.M. Ouwens; H. Pijl; B. Guigas (pp. 1827-1836).

Treatment with the dopamine receptor D2 (DRD2) agonist bromocriptine improves metabolic features in obese patients with type 2 diabetes by a still unknown mechanism. In the present study, we investigated the acute effect of bromocriptine and its underlying mechanism(s) on insulin secretion both in vivo and in vitro. For this purpose, C57Bl6/J mice were subjected to an intraperitoneal glucose tolerance test (ipGTT) and a hyperglycemic (HG) clamp 60min after a single injection of bromocriptine or placebo. The effects of bromocriptine on glucose-stimulated insulin secretion (GSIS), cell membrane potential and intracellular cAMP levels were also determined in INS-1E beta cells. We report here that bromocriptine increased glucose levels during ipGTT in vivo, an effect associated with a dose-dependent decrease in GSIS. During the HG clamp, bromocriptine reduced both first-phase and second-phase insulin response. This inhibitory effect was also observed in INS-1E beta cells, in which therapeutic concentrations of bromocriptine (0.5–50nM) decreased GSIS. Mechanistically, neither cellular energy state nor cell membrane depolarization was affected by bromocriptine whereas intracellular cAMP levels were significantly reduced, suggesting involvement of G-protein-coupled receptors. Surprisingly, the DRD2 antagonist domperidone did not counteract the effect of bromocriptine on GSIS, whereas yohimbine, an antagonist of the α2-adrenergic receptors, completely abolished bromocriptine-induced inhibition of GSIS. In conclusion, acute administration of bromocriptine inhibits GSIS by a DRD2-independent mechanism involving direct activation of the pancreatic α2-adrenergic receptors. We suggest that treatment with bromocriptine promotes beta cells rest, thereby preventing long-lasting hypersecretion of insulin and subsequent beta cell failure.

Keywords: Abbreviations; α2-AR; α2-adrenergic receptors; AMPK; AMP-activated protein kinase; Ca; ²⁺; calcium; CREB; cAMP response element binding; DRD2; dopamine receptor D2; ERK1/2; extracellular signal-regulated kinases 1/2; GSIS; glucose-stimulated insulin secretion; HG clamp; hyperglycemic clamp; IBMX; 3-isobutyl-1-methylxanthine; ipGTT; intraperitoneal glucose tolerance test; K; _ATP; -channel; ATP-sensitive potassium channel; KRBH buffer; Krebs–Ringer bicarbonate HEPES buffer; PKA; cAMP-dependent protein kinase ABromocriptine; Insulin secretion; Mice; INS-1E cells; Dopamine receptor; α2-Adrenergic receptor

Corrigendum to “Heteronemin, a spongean sesterterpene, inhibits TNFα-induced NF-κB activation through proteasome inhibition and induces apoptotic cell death” [Biochem. Pharmacol. 79 (2010) 610–622] by Marc Schumacher; Claudia Cerella; Serge Eifes; Sébastien Chateauvieux; Franck Morceau; Marcel Jaspars; Mario Dicato; Marc Diederich (pp. 1837-1837).

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Biochemical Pharmacology (v.79, #12)

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Biochemical Pharmacology (v.79, #12)