Forgot your password?
New user?
New Window Opens on the Secret Life of Microbes: Scientists Develop First Microbial Profiles of Ecosystems
CAS announces the release of SciFinder Scholar for Mac OS X
Press Releases
Press Releases
| Check out our New Publishers' Select for Free Articles |
BBA - Molecular Cell Research (v.1763, #10)
Thy-1, a versatile modulator of signaling affecting cellular adhesion, proliferation, survival, and cytokine/growth factor responses by Tanya A. Rege; James S. Hagood (pp. 991-999).
Thy-1 is a 25–37 kDa glycosylphosphatidylinositol (GPI)-anchored protein involved in T cell activation, neurite outgrowth, apoptosis, tumor suppression, wound healing, and fibrosis. To mediate these diverse effects, Thy-1 participates in multiple signaling cascades. In this review, we discuss Thy-1 signaling primarily in non-immunologic cell types, including neurons, mesangial cells, ovarian cancer cells, nasopharyngeal carcinoma cells, endothelial cells, and fibroblasts. We review the current literature regarding Thy-1 signaling via integrins, protein tyrosine kinases, and cytokines and growth factors; and the roles of these signaling pathways in cellular adhesion, apoptosis, cell proliferation, and cell adhesion and migration. We also discuss the role of Thy-1 localization to lipid rafts, and of the GPI anchor in Thy-1 signaling. Ongoing research on the mechanisms of Thy-1 signaling will add to our understanding of the diverse physiologic and pathologic processes in which Thy-1 plays a role.
Keywords: Abbreviations; COX; cyclooxygenase; FAK; focal adhesion kinase; Gadd 45 γ; DNA-damage-inducible protein 45 gamma; GAP; GTPase activating protein; GPI; glycosylphosphatidylinositol; LAT; linker for activation of T cells; MAPK; mitogen-activated protein kinase; NCAM; neural cell adhesion molecule; NGFI-B; nerve growth factor induced protein I-B; NPC; nasopharyngeal carcinoma; PDGF; platelet-derived growth factor; PG; prostaglandin; PI 3-kinase; phosphatidylinositol 3-kinase; PKC; protein kinase C; PTK; protein tyrosine kinase; SFK; Src family kinase; SMA; smooth muscle actin; STAT3; signal transducer and activator of transcription 3; TGF; transforming growth factor; TNF; tumor necrosis factor; TUNEL; TdT-mediated dUTP nick-end labelingThy-1; GPI anchor; Cell signaling; Integrins; Tyrosine kinases; Lipid rafts
Evaluation of IFN-γ effects on apoptosis and gene expression in neuroblastoma—Preclinical studies by Tanya M. Tekautz; Kejin Zhu; Jose Grenet; Deepak Kaushal; Vincent J. Kidd; Jill M. Lahti (pp. 1000-1010).
Loss of caspase-8 expression and resistance to cytotoxic agents occurs frequently in late stage neuroblastoma (NB). Interferon-γ (IFN-γ) induces caspase-8 in NB cells, sensitizing them to death receptor mediated apoptosis. This study characterizes the kinetics of this phenomenon and examines the effects of IFN-γ on global gene expression to determine whether IFN-γ responses are achievable at physiologically relevant doses and to define the biological effects of this cytokine. Here we examine the IFN-γ responses of 16 NB cell lines. A single <5-min exposure to IFN-γ (0.5 ng/ml) induced caspase-8 expression in all non-expressing cell lines and in 3/6 cell lines which already expressed high caspase-8. This increase in caspase-8 proteins was observed within 16 h and persisted for up to 9 days. Furthermore, IFN-γ pretreatment of NB cells increased doxorubicin-induced apoptosis nearly 3-fold. Microarray analysis was used to identify additional genes involved in proliferation, signaling and apoptosis whose expression was modulated via IFN-γ. Altered expression of these genes should further enhance the responsiveness of NB cells to chemotherapeutics. Thus, the use of IFN-γ to sensitize NB cells to cytotoxic agents represents an attractive therapeutic strategy and warrants further investigation.
Keywords: Interferon-γ; MYCN; Neuroblastoma; Caspase-8; Apoptosis; Microarray
Heterotypic complex formation between subunits of microtubule-associated proteins 1A and 1B is due to interaction of conserved domains by Rainer Noiges; Heike Stroissnigg; Alžbeta TranÄ?iková; Ilse Kalny; René Eichinger; Friedrich Propst (pp. 1011-1016).
The microtubule-associated proteins MAP1A and MAP1B are related but distinct multi-subunit protein complexes that consist of heavy and light chains. The predominant forms of these complexes are homotypic, i.e. they consist of a MAP1A heavy chain associated with MAP1A light chains or a MAP1B heavy chain associated with MAP1B light chains, respectively. In addition, MAP1A and MAP1B can exchange subunits and form heterotypic complexes consisting of a MAP1A heavy chain associated with MAP1B light chains which might play a role in a transition period of neuronal differentiation. Here we extend previous findings by confirming that heterotypic MAP1B heavy chain-MAP1A light chain complexes also exist in the developing murine brain. We show that these complexes form through interaction of homologous domains conserved in heavy and light chains of MAP1A and MAP1B. Likewise, conserved domains of the MAP1A and MAP1B light chains account for formation of light chain heterodimers. By yeast 2-hybrid analysis we located the light chain binding domain on the heavy chain to amino acids 211–508, thereby defining a new functional subdomain.
Keywords: Abbreviations; HRP; horse radish peroxidase; MAP; microtubule-associated protein; MH; MAP1 homology; HC; heavy chain; LC; light chain; SDS-PAGE; SDS-polyacrylamide gel electrophoresisMicrotubule-associated protein; Multimeric protein complex; Protein–protein interaction; Neuronal cytoskeleton; Brain development
Bafilomycin A1-sensitive pathway is required for the maturation of cystic fibrosis transmembrane conductance regulator by Tsukasa Okiyoneda; Akiko Niibori; Kazutsune Harada; Taijun Kohno; Yasuaki Hashimoto; Hiroyuki Kusuhara; Tappei Takada; Tsuyoshi Shuto; Mary Ann Suico; Yuichi Sugiyama; Hirofumi Kai (pp. 1017-1023).
Cystic fibrosis (CF) is the most common lethal genetic disease in Caucasians caused by the trafficking defects of CF transmembrane conductance regulator (CFTR), which is a cAMP-dependent Cl− channel at the plasma membrane. The trafficking pathway of CFTR is thought to be non-conventional because CFTR maturation is inhibited by the dysfunction of syntaxin 13, which is involved in protein recycling via endosomal pathway. In this study, to clarify whether the endosomal trafficking is required for CFTR maturation, we utilized a specific vacuolar H+-ATPase inhibitor, bafilomycin A1 (BafA1), which inhibits the protein trafficking from early endosome. Our data showed that low concentration of BafA1 (50nM) decreased the expression of mature CFTR but induced the accumulation of immature CFTR in the juxta-nuclear region containing an early endosome marker. Pulse-chase analysis showed that BafA1 inhibited the maturation of CFTR, but it slightly stabilized immature CFTR. These results indicate that BafA1-sensitive pathway is required for CFTR maturation and emphasize that endosomal trafficking pathway might be involved in the maturation of CFTR.
Keywords: Abbreviations; CF; cystic fibrosis; CFTR; CF transmembrane conductance regulator; ER; endoplasmic reticulum; BafA1; bafilomycin A1; BFA; brefeldin A; BHK; baby hamster kidney; CHO; Chinese hamster ovary; GFP; green fluorescence protein; ERGIC; ER-Golgi intermediate compartmentCFTR; Bafilomycin A1; Endosome; Maturation and trafficking
Stimulation of tPA-dependent provisional extracellular fibrin matrix degradation by human recombinant soluble melanotransferrin by Y. Bertrand; M. Demeule; G.-E. Rivard; R. Béliveau (pp. 1024-1030).
Tissue-type plasminogen activator (tPA) and its substrate plasminogen (Plg) are key components in the fibrinolytic system. We have recently demonstrated, that truncated human recombinant soluble melanotransferrin (sMTf) could stimulate the activation of Plg by urokinase plasminogen activator and inhibit angiogenesis. Since various angiogenesis inhibitors were shown to stimulate tPA-mediated plasminogen activation, we examined the effects of sMTf on tPA-dependent fibrinolysis. This study demonstrated that sMTf enhanced tPA-activation of Plg by 6-fold. sMTf also increased the release of [125I]-fibrin fragments by tPA-activated plasmin. Moreover, we observed that the interaction of sMTf with Plg provoked a change in the fibrin clot structure by cleaving the fibrin α and β chains. Overall, the present study shows that sMTf modulates tPA-dependent fibrinolysis by modifying the clot structure. These results also suggest that sMTf properties could involve enhanced dissolution of the provisional extracellular fibrin matrix.
Keywords: Fibrin matrix; Melanotransferrin; Plasminogen
Transcriptome analysis of fetal metatarsal long bones by microarray, as a model for endochondral bone formation by Rachael V. Sugars; Elerin Kärner; Ulrika Petersson; Bernhard Ganss; Mikael Wendel (pp. 1031-1039).
Endochondral bone formation is orchestrated by mesenchymal cell condensation to form cartilage anlagen, which act as a template for bone formation and eventual mineralization. The current study performed gene expression analysis to examine pre- and post-mineralization stages (E15 and E19) of endochondral bone formation, using fetal metatarsal long bones as a model. An extensive number of genes were differentially expressed, with 543 transcripts found to have at least 2-fold up-regulation and 742 with a greater than 2-fold down-regulation. A bioinformatics approach was adopted based on gene ontology groups, and this identified genes associated with the regulation of signaling and skeletal development, cartilage replacement by bone, and matrix degradation and turnover. Transcripts linked to skeletal patterning, including Hoxd genes 10–12, Gli2 and Noggin were considerably down-regulated at E19. Whereas genes associated with bone matrix formation and turnover, ACP5, MMP-13, bone sialoprotein, osteopontin, dentin matrix protein-1 and MMP-9 all were distinctly up-regulated at this later time point. This approach to studying the formation of the primary ossification center provides a unique picture of the developmental dynamics involved in the molecular and biochemical processes during this intricately regulated process.
Keywords: Microarray; Endochondral bone; Remodeling; Gene expression
Toxicity of enzymatic oxidation products of spermine to human melanoma cells (M14): Sensitization by heat and MDL 72527 by Enzo Agostinelli; Francesca Belli; Agnese Molinari; Maria Condello; Paola Palmigiani; Laura Dalla Vedova; Manuela Marra; Nikolaus Seiler; Giuseppe Arancia (pp. 1040-1050).
In situ formation of cytotoxic metabolites by an enzyme-catalyzed reaction is a recent approach in cancer chemotherapy. We demonstrate that multidrug resistant human melanoma cells (M14 ADR) are more sensitive than the corresponding wild type cells (M14 WT) to hydrogen peroxide and aldehydes, the products of bovine serum amine oxidase (BSAO)-catalyzed oxidation of spermine. Hydrogen peroxide was mainly responsible for the loss of cell viability. With about 20%, the aldehydes formed from spermine contribute also to cytotoxicity. Elevation of temperature from 37 °C to 42 °C decreased survival of both cell lines by about one log unit. Pre-treatment with N1,N4-bis(2,3-butadienyl)-1,4-butanediamine (MDL 72527), a lysosomotropic compound, sensitized cells to toxic spermine metabolites. MDL 72527 (at 300 μM) produced in M14 cells numerous cytoplasmic vacuoles which, however, disappeared by 24 h, even in the presence of the drug. Mitochondrial damage, as observed by transmission electron microscopy, correlated better with the cytotoxic effects of the treatment than vacuole formation. Since the release of lysosomal enzymes causes oxidative stress and apoptosis, we suggest that the lysosomotropic effect of MDL 72527 is the major reason for its sensitizing effect.
Keywords: Abbreviations; BSA; bovine serum albumin; BSAO; bovine serum amine oxidase; CHO; Chinese hamster ovary; DX; doxorubicin; ADR; adriamicin resistent; IU; international unit; MDL 72527; N; 1; ,N; 4; -bis(2,3-butadienyl)-1,4-butanediamine; MDR; multidrug resistance; PBS; phosphate-buffered saline; ROS; reactive oxygen species; SD; standard deviationBovine serum amine oxidase; Melanoma cell; Spermine; MDL 72527; Hyperthermia
Differential regulation of lipopolysaccharide and Gram-positive bacteria induced cytokine and chemokine production in splenocytes by Gαi proteins by Hongkuan Fan; David L. Williams; Basilia Zingarelli; Kevin F. Breuel; Giuseppe Teti; George E. Tempel; Karsten Spicher; Guylain Boulay; Lutz Birnbaumer; Perry V. Halushka; James A. Cook (pp. 1051-1058).
Heterotrimeric Gi proteins play a role in lipopolysaccharide (LPS) and Staphylococcus aureus (SA) activated signaling leading to inflammatory mediator production. We hypothesized that genetic deletion of Gi proteins would alter cytokine and chemokine production induced by LPS and SA. LPS- and heat killed SA-induced cytokine and chemokine production in splenocytes from wild type (WT), Gαi2 (−/−) or Gαi1/3 (−/−) mice were investigated. LPS- or SA-induced production of TNFα, IL-6, IFNγ, IL-12, IL-17, GM-CSF, MIP-1α, MCP-1, MIG and IP-10 were significantly increased (1.2 to 33 fold, p<0.05) in splenocytes harvested from Gαi2(−/−) mice compared with WT mice. The effect of Gαi protein depletion was remarkably isoform specific. In splenocytes from Gαi1/3 (−/−) mice relative to WT mice, SA-induced IL-6, IFNγ, GM-CSF, and IP-10 levels were decreased (59% to 86%, p<0.05), whereas other LPS- or SA-stimulated cytokines and chemokines were not different relative to WT mice. LPS- and SA-induced production of KC were unchanged in both groups of the genetic deficient mice. Splenocytes from both Gαi2 (−/−) and Gαi1/3 (−/−) mice did not exhibit changes in TLR2 and TLR4 expression. Also analysis of splenic cellular composition by flow cytometry demonstrated an increase in splenic macrophages and reduced CD4 T cells in both Gαi2 (−/−) and Gαi1/3 (−/−) mice relative to WT mice. The disparate response of splenocytes from the Gαi2 (−/−) relative to Gαi1/3 (−/−) mice therefore cannot be attributed to major differences in spleen cellular composition. These data demonstrate that Gi2 and Gi1/3 proteins are both involved and differentially regulate splenocyte inflammatory cytokine and chemokine production in a highly Gi isoform specific manner in response to LPS and Gram-positive microbial stimuli.
Keywords: G; i; protein deficient mice; Endotoxin; Staphylococcus aureus; TLR signaling
Thymopentin (TP5), an immunomodulatory peptide, suppresses proliferation and induces differentiation in HL-60 cells by Ying-zhe Fan; Hui Chang; Ye Yu; Jing Liu; Li Zhao; Ding-jian Yang; Rui Wang (pp. 1059-1066).
Thymopentin (Arg–Lys–Asp–Val–Tyr, TP5) has shown immuno-regulatory activities in humans. In the present study, we investigated the effects of TP5 on the proliferation and differentiation of a human promyelocyte leukemia cell line, HL-60. It is noteworthy that TP5 displayed concentration-dependent inhibitory effects on the proliferation and colony formation of HL-60 cells. Furthermore, the decrease or even disappearance of AgNORs from nucleoli was observed in HL-60 cells after the treatment with TP5. The suppression induced by TP5 was accompanied by an accumulation of cell cycle in the G0/G1 phase. Moreover, TP5 significantly increased the NBT-reduction activity of HL-60 cells. Cytofluorometric and morphologic analysis indicated that TP5 had induced differentiation along the granulocytes lineage in HL-60 cells.d-tubocurarine (TUB) significantly antagonized the inhibitory effects induced by TP5, whereas atropine did not exhibit such effect. All the results indicated that TP5 was able to significantly inhibit proliferation and induce differentiation in HL-60 cells. Our observations also implied that TP5 not only acted as an immunomodulatory factor in cancer chemotherapy, but is also a potential chemotherapeutic agent in the human leukemia therapy.
Keywords: Thymopentin; HL-60; Differentiation; Proliferation inhibition; Chemotherapeutic agent
Site specific differential activation of ras/raf/ERK signaling in rabbit isoproterenol-induced left ventricular hypertrophy by Nari Kim; Hyunju Kim; Jae Boum Youm; Won Sun Park; Mohamad Warda; Jae-Hong Ko; Jin Han (pp. 1067-1075).
To understand better the mediating role of ras/raf/ERK signaling pathway in development of cardiac hypertrophy and cerebrovascular events in vivo, the molecular mechanism of the pathway in heart and cerebral arteries after isoproterenol (ISO) induced β-adrenergic receptor (βAR) stimulation was examined in rabbit as animal model. Compared with the heart, our findings indicate that ISO-stimulation results in increase in mRNA levels of ras, raf, and immediate-early genes in the cerebral arteries. Conversely, the ras and raf protein expression levels (determined by Western blot) and the ras-GTP level (determined by pull-down assay) in the heart, but not the cerebral arteries, are markedly elevated after treatment. In addition, despite constant ERK1/2 abundance, phosphorylated ERK ( pERK) activity was elevated at both sites with prominent effect on heart following stimulation. Opposing to the PKA and PKC, as upstream contributors in the pathway, which seem to be similarly affected at both sites following ISO-stimulation, the results imply that the downstream candidates ras and raf, as well as immediate-early genes, have different responses at both sites post-stimulation. The results provide an evidence of site-dependent differential response of ras/ raf/ERK pathway after cardiac hypertrophy-induced by ISO-stimulation. This varied response may account for underlying mechanisms of development of cardiac hypertrophy and cerebrovascular events in heart and cerebral arteries, respectively.
Keywords: Cardiac hypertrophy; β-adrenergic receptor; Cerebrovascular event; ras; /; raf; /MAPK pathway
CRM 1-mediated degradation and agonist-induced down-regulation of β-adrenergic receptor mRNAs by Ying Bai; Huafei Lu; Curtis A. Machida (pp. 1076-1089).
The β1-adrenergic receptor (β1-AR) mRNAs are post-transcriptionally regulated at the level of mRNA stability and undergo accelerated agonist-mediated degradation via interaction of its 3′ untranslated region (UTR) with RNA binding proteins, including the HuR nuclear protein. In a previous report [Kirigiti et al. (2001). Mol. Pharmacol. 60:1308–1324], we examined the agonist-mediated down-regulation of the rat β1-AR mRNAs, endogenously expressed in the rat C6 cell line and ectopically expressed in transfectant hamster DDT1MF2 and rat L6 cells. In this report, we determined that isoproterenol treatment of neonatal rat cortical neurons, an important cell type expressing β1-ARs in the brain, results in significant decreases in β1-AR mRNA stability, while treatment with leptomycin B, an inhibitor of the nuclear export receptor CRM 1, results in significant increases in β1-AR mRNA stability and nuclear retention. UV-crosslinking/immunoprecipitation and glycerol gradient fractionation analyses indicate that the β1-AR 3′ UTR recognize complexes composed of HuR and multiple proteins, including CRM 1. Cell-permeable peptides containing the leucine-rich nuclear export signal (NES) were used as inhibitors of CRM 1-mediated nuclear export. When DDT1MF2 transfectants were treated with isoproterenol and peptide inhibitors, only the co-addition of the NES inhibitor reversed the isoproterenol-induced reduction of β1-AR mRNA levels. Our results suggest that CRM 1-dependent NES-mediated mechanisms influence the degradation and agonist-mediated down-regulation of the β1-AR mRNAs.
Keywords: β; 1; -adrenergic receptor; CRM 1; mRNA degradation and stability; Agonist-induced down-regulation
ATM activation is accompanied with earlier stages of prostate tumorigenesis by Catherine Fan; Rebecca Quan; Xinchang Feng; Aubrey Gillis; Lizhi He; Edward D. Matsumoto; Sam Salama; Jean-Claude Cutz; Anil Kapoor; Damu Tang (pp. 1090-1097).
The ATM (ataxia telangiectasia mutated) kinase plays an essential role in maintaining genome integrity by coordinating cell cycle arrest, apoptosis, and DNA damage repair. Phosphorylation of ATM at serine 1981 (ATMpSer1981) by DNA damage activates ATM, which subsequently phosphorylates H2AX Ser139 (γH2AX), Chk2 Thr68 (Chk2pThr68), and p53 Ser15 (p53pSer15). To determine the role of the ATM pathway in prostate cancer tumorigenesis, we have analyzed 35 primary prostate cancer specimens for ATMpSer1981 (ATM activation), Chk2pThr68, γH2AX, and p53pSer15 by immunohistochemistry (IHC) in normal glands, prostatic intraepithelial neoplasias (PINs), and carcinomas. Increases in the intensities of ATMpSer1981, Chk2pThr68, and γH2AX and in the percentage of cells that are positive for ATMpSer1981, Chk2pThr68, or γH2AX were observed in PINs ( p<0.001) compared to normal prostatic glands and carcinoma. However, this pattern of immunostaining was not seen for p53pSer15. Thus, ATM and Chk2 are specifically activated in PINs. As PINs are generally regarded as precursors of prostatic carcinoma, our results suggest that ATM and Chk2 activation at earlier stages of prostate tumorigenesis suppresses tumor progression, with attenuation of ATM activation leading to cancer progression.
Keywords: ATM; Chk2; p53; γH2AX; Prostate tumorigenesis
An ITIM-like motif within the CCK2 receptor sequence required for interaction with SHP-2 and the activation of the AKT pathway by Sébastien Vatinel; Audrey Ferrand; Fréderic Lopez; Aline Kowalski-Chauvel; Jean-Pierre Estève; Daniel Fourmy; Marlène Dufresne; Catherine Seva (pp. 1098-1107).
SHP-2 is a tyrosine phosphatase which functions as a positive regulator downstream of RTKs, activating growth-stimulatory signalling pathways. To date, very few G protein-coupled receptors (GPCRs) have been shown to be connected to SHP-2 and very little is known about the positive role of SHP-2 in GPCR signalling. The CCK2 receptor (CCK2R), a GPCR, is now recognized to mediate mitogenic effects of gastrin on gastrointestinal cells. In the present study, we demonstrate the role of SHP-2 in the activation of the AKT pathway by the CCK2R in COS-7 cells transfected with the CCK2R and in a pancreatic cancer cell line expressing the endogenous receptor. Using surface plasmon resonance analysis, we identified a highly conserved ITIM motif, containing the tyrosine residue 438, located in the C-terminal intracellular tail of the CCK2R which directly interacts with the SHP-2 SH2 domains. The interaction was confirmed by pull down assays and co-immunoprecipitation of the receptor with SHP-2. This interaction was transiently increased following gastrin stimulation of the CCK2R and correlated with the tyrosine phosphorylation of SHP-2. Mutational analysis of the key ITIM residue 438 confirmed that the CCK2R ITIM sequence is required for interaction with SHP-2 and the activation of the AKT pathway.
Keywords: Gastrin; CCK2 receptor; G protein-coupled receptor; Cell signalling; Protein interaction
Structural and functional consequences of c-N-Ras constitutively associated with intact mitochondria by Janice C. Wolfman; Sarah M. Planchon; Jinhui Liao; Alan Wolfman (pp. 1108-1124).
We demonstrate that both c-N-Ras and c-K(B)-Ras are constitutively associated with purified mitochondria. c-K(B)-Ras is associated with the mitochondrial outer membrane, and c-N-Ras is associated with both the outer membrane and inner mitochondrial compartments. The mitochondrial morphology is abnormal in both c-N-Ras negative and K-Ras negative cells. Normal mitochondrial morphology was restored by targeting N-Ras to both the inner and outer mitochondrial compartments, or by ectopically expressing c-K(B)-Ras. Impaired mitochondrial function can result in increased CHOP and NFκB activity, typical for a retrograde signaling response. Both are constitutively elevated in the N-Ras negative cells, but not in the K-Ras negative background, and are restored by c-N-Ras targeted exclusively to the inner mitochondrial compartment. Surprisingly, both targeting and the ability to functionally reduce retrograde transcriptional activity were found to be independent of c-N-Ras farnesylation. Overall, these data demonstrate for the first time a (1) farnesylation independent function for c-N-Ras and (2) that N-Ras within the inner mitochondrial compartment is an essential component of the retrograde signaling system between the mitochondria and nucleus.
Keywords: Ras; Retrograde signaling; Mitochondrial stress; Morphology; Transcriptional activity
The C-terminal CD47/IAP-binding domain of thrombospondin-1 prevents camptothecin- and doxorubicin-induced apoptosis in human thyroid carcinoma cells by G.M. Rath; C. Schneider; S. Dedieu; B. Rothhut; M. Soula-Rothhut; C. Ghoneim; B. Sid; H. Morjani; H. El Btaouri; L. Martiny (pp. 1125-1134).
Camptothecin and doxorubicin belong to a family of anticancer drugs that exert cytotoxic effects by triggering apoptosis in various cell types. However there have only been few investigations showing that matricellular proteins like thrombospondin-1 (TSP-1) could be involved in the underlying mechanism of this cytotoxicity. In this report, using Hoechst reagent staining, reactive oxygen species production and caspase-3 activity measurement, we determined that both camptothecin and doxorubicin induced apoptosis in human thyroid carcinoma cells (FTC-133). On the one hand, we demonstrated that camptothecin and doxorubicin inhibited TSP-1 expression mainly occurring at the transcriptional level. On the other hand, drug-induced apoptosis determined by western blot analysis for PARP cleavage and caspase-3 activity measurement, was significantly decreased in presence of exogenous TSP-1. In order to identify the sequence responsible for this effect, we used the CD47/IAP-binding peptide 4N1 (RFYVVMWK), derived from the C-terminal domain of TSP-1, and known to play a role in apoptosis. Thus, in presence of 4N1, camptothecin and doxorubicin-induced pro-apoptotic activity was considerably inhibited. These findings suggest that induction of apoptosis by camptothecin or doxorubicin in FTC-133 cells is greatly dependent by a down-regulation of TSP-1 expression and shed new light on a possible role for TSP-1 in drug resistance.
Keywords: 4N1; Camptothecin; Doxorubicin; Apoptosis; Thyroid carcinoma cell