Expression of focal adhesion kinase and α5andβ1integrins in carcinonas and its clinical significance

AIM：To detect the expression pattern of FAK(focal adhesion kinase)and integrinα5andβ1 subunits in different kinds of cancerous tissues and to study their correlation with clinicopathological data including tumor type,grade and lymph node status.METHODS:Using an immunohistochemical technique,we examined the expression of FAK and integrin and subunits in cancerous and noncancerous tissues obtained from75patients with gastric carcinomas,21colorectal carcinomas,16 hepatocellular carcinomas,20uterocervical carcinomas,and20breast carcinomas.RESULTS:The staining of FAK was stronger in cancerous than in noncancerous areas,Enhanced expression of FAKwas detected in poor-differentiated carcinoma of the stomach and colorectum.Tumors with lymph node metastases had more FAKprotein than those without metastases.In addition.the deeper the extent of tumor infiltration,the higher the FAKexpression.The expression of integrinα5andβ1subunits was lower in cancerous areas than in noncancerous areas,but it was higher in well-differentiated cancerous tissues than in poor differentiated tissues.The relationship between the expression of integrinα5andβ1subunits and infiltration or metastasis was not significant.Cancerous tissues with stronger FAK expression(++or+++)also had a higher expression of integrinα5andβ1subunits in the tumor and its unaffected margins.CONCLUSION:FAKis a better marker for carcinogenesis and the progression of cancer than integrinα5orβ1subunit,and it may be not only a transformation-linked enzyme but also a progression-linked enzyme.     

Tetrandrine inhibits activation of rat hepatic stellate cells in vitro via transforming growth factor-β signaling

AIM: To investigate the effect of various concentrations of tetrandrine on activation of quiescent rat hepatic stellate cells (HSCs) and transforming growth factor-β (TGF-β) signaling in vitro. METHODS: HSCs were isolated from rats by in situ perfusion of liver and 18% Nycodenz gradient centrifugugation, and primarily cultured on uncoated plastic plates for 24 h with DMEM containing 20% fetal bovine serum (FBS/DMEM) before the culture medium was substituted with 2% FBS/DMEM for another 24 h. Then, the HSCs were cultured in 2% FBS/DMEM with tetrandrine (0.25, 0.5, 1,2 mg/L, respectively). Cell morphological features were observed under an inverted microscope, smooth muscle-α-actin (α-SMA) was detected by immunocytochemistry and image analysis system, laminin (LN) and type Ⅲ procollagen (PCⅢ) in supernatants were determined by radioimmunoassay. TGF-β1 mRNA, Smad 7 mRNA and Smad 7 protein were analyzed with RT-PCR and Western blotting, respectively. RESULTS: Tetrandrine at the concentrations of 0.25-2 mg/L prevented morphological transformation of HSC from the quiescent state to the activated one, while α-SMA, LN and PCⅢ expressions were inhibited. As estimated by gray values, the expression of α-SMA in tetrandrine groups (0.25, 0.5, 1, 2 mg/L) was reduced from 21.3% to 42.2% (control: 0.67, tetrandrine groups: 0.82, 0.85, 0.96, or 0.96, respectively, which were statistically different from the control, P<0.01), and the difference was more significant in tetrandrine at 1 and 2 mg/L. The content of LN in supernatants was significantly decreased in tetrandrine groups to 58.5%, 69.1%, 65.8% or 60.0% that of the control respectively, and that of PCII] to 84.6%, 81.5%, 75.7% or 80.7% respectively (P<0.05 vs control), with no significant difference among tetrandrine groups. RTPCR showed that TGF-β1 mRNA expression was reduced by tetrandrine treatments from 56.56% to 87.90% in comparison with the control, while Smad 7 mRNA was increased 1.4-4.8 times. The TGF-β1 mRNA and Smad 7 mRNA expression was in a significant negative correlation (r = -0.755, P<0.01), and both were significantly correlated with α-SMA protein expression (r = -0.938, P<0.01; r = 0.938, P<0.01, respectively). The up-regulation of Smad 7 protein by tetrandrine (1 mg/L) was confirmed by Western blotting as well. CONCLUSION: Tetrandrine has a direct inhibiting effect on the activation of rat HSCs in culture. It up-regulates the expression of Smad 7 which in turn blocks TGF-β1 expression and signaling.     

Apoptosis of hepatoma cells SMMC—7721 induced by Ginkgo biloba seed polysaccharide

AIM：To study the apoptosis of hepatoma cells SMMC-7721induced by polysaccharide isolated from Ginkgo biloba seed.METHODS:Ginkgo biloba seed polysaccharide(GBSP)was isolated by ethanol fractionation of Ginkgo biloba seed and purified by Sephadex G-200 chromatography,The purity of GBSP was verified by reaction with iodine-potassium iodide and ninhydrin and confirmed by UV spectrophotometer,cellulose acetate membrane electrophoresis and Separose 4B gel filtration chromatography,The Scanning Electron Microscope(SEM)and Flow Cytometry(FCM)were used to examine the SMMC-7721 cells with and without GBSP treatment at 500mg/ml for 36h.RESULTS:GBSP product obtained was of high purity with the average molecular weight of 1.86×10^5.Quantitative analysis of SMMC-7721cells in vitro with FCM showed that the percentages of G2-Mcells without and with GBSP treatment were17.01±1.28%and 11.77±1.50%(P<0.05).the debris ratio of the cells were0.46±0.12%and 0.06±0.06%(P<0.01).and the apoptosis ration of cells was3.84±0.55%and9.13±1.48%(P<0.010respectively,Following GBSP treatment,microvilli of SMMC-7721cells appeared thinner and the number of spherical cells increased markedly,Most significantly,the apoptosis bodies were formed on and around the sppherical cells treated with GBSP.CONCLUSION:GBSP could potentially induce the apoptosis of SMMC-7721cells.     

Apoptosis of human gastric adenocarcinoma cells induced by β-ionone

AIM:To investigate the effect of β-ionone on the growth and apoptosis of gastric adenocarcinoma cell line SGC-7901.METHODS: Using M-IT, fluorescence dye (Hoechst-33258),transmission electron microscopy and the TUNEL assay,we examined growth and apoptosis of SGC-7901 cells treated with β-ionone at various concentrations (i.e. 25, 50, 100 and 200μmol/L) for 24h,48h.RESULTS:The growth of SGC-7901 cells was inhibited by β-ionone. Seven days after treatment with β-ionone at four concentrations, the inhibition rates were 12.04%, 30.59%,78.25% and 94.15%, respectively. The IC50 value of β-ionone for SGC-7901 cells was estimated to be 89μmol/L.The apoptotic morphology was demonstrated in SGC-7901 cells treated with β-ionone by Hoechst-33258 staining and electron microscopy. Apoptosis was also shown in β-iononetreated SGC-7901 cells by the TUNEL assay.CONCLUSION:β-ionone can inhibit cell proliferation and induce apoptosis of SGC-7901 cells.However, the mechanism needs to be further investigated.     

Role of adhesion molecules and dendritic cells in rat hepatic／renal ischemia-reperfusion injury and anti-adhesive intervention with anti-P-selectin lectin-EGF domain monoclonal antibody

AIM: To investigate the role of P-selectin, intercellular adhesion molecule-1 (ICAM-1) and dendritic cells (DCs) in liver/kidney of rats with hepatic/renal ischemia-reperfusion injury and the preventive effect of anti-Pselectin lectin-EGF domain monoclonal antibody (anti-PsL-EGFmAb) on the injury. METHODS: Rat models of hepatic and renal ischemiareperfusion were established. The rats were then divided into two groups, one group treated with anti-PsL-EGFmAb (n = 20) and control treated with saline (n = 20). Both groups were subdivided into four groups according to reperfusion time (1, 3, 6 and 24 h). The sham-operated group (n = 5) served as a control group. DCs were observed by the microscopic image method, while P-selectin and ICAM-1 were analyzed by immunohistochemistry. RESULTS: P-selectin increased significantly in hepatic sinusoidal endothelial cells and renal tubular epithelial cells 1 h after ischemia-reperfusion, and the expression of ICAM-1 was up-regulated in hepatic sinusoid and renal vessels after 6 h. CD1a+CD80+DCs gradually increased in hepatic sinusoidal endothelium and renal tubules and interstitium 1 h after ischemia-reperfusion, and there was the most number of DCs in 24-h group. The localization of DCs was associated with rat hepatic/renal function. These changes became less significant in rats treated with anti-PsL-EGFmAb. CONCLUSION: DCs play an important role in immune pathogenesis of hepatic/renal ischemia-reperfusion injury. Anti-PsL-EGFmAb may regulate and inhibit local DC immigration and accumulation in liver/kidney.     

Contractile effects and intracellular Ca^2＋ signalling induced by emodin in circular smooth muscle cells of rat colon

AIM: To investigate whether emodin has any effects on circular smooth muscle cells of rat colon and to examine the mechanism underlying its effect. METHODS: Smooth muscle cells were isolated from the circular muscle layer of Wistar rat colon and the cell length was measured by computerized image micrometry. Intracellular Ca(2+) ([Ca(2+)]i) signalling was studied in smooth muscle cells using Ca(2+) indicator Fluo-3 AM on a laser-scanning confocal microscope. RESULTS: Emodin dose-dependently induced smooth muscle cells contraction. The contractile responses induced by emodin were inhibited by preincubation of the cells with ML-7 (an inhibitor of MLCK). Emodin caused a large, transient increase in [Ca(2+)]i followed by a sustained elevation in [Ca(2+)]i. The emodin -induced increase in [Ca(2+)]i was unaffected by nifedipine, a voltage-gated Ca(2+)-channel antagonist, and the sustained phase of the rising of [Ca(2+)]i was attenuated by extracellular Ca(2+) removal with EGTA solution. Inhibiting Ca(2+) release from ryanodine-sensitive intracellular stores by ryanodine reduced the peak increase in [Ca(2+)]i. Using heparin, an antagonist of IP(3)R, almost abolished the peak increase in [Ca(2+)]i. CONCLUSION: Emodin has a direct excitatory effect on circular smooth muscle cells in rat colon mediated via Ca(2+)/CaM dependent pathways. Furthermore, emodin-induced peak [Ca(2+)]i increase may be attributable to the Ca(2+) release from IP(3) sensitive stores, which further promote Ca(2+) release from ryanodine-sensitive stores through CICR mechanism. Additionally, Ca(2+) influx from extracellular medium contributes to the sustained increase in [Ca(2+)]i.     

Effect of ligand of peroxisome proliferator-activated receptor γ on the biological characters of hepatic stellate cells

AIM: To study the effect of rosiglitazone, which is a ligand of peroxisome proliferator-activated receptor gamma (PPARgamma), on the expression of PPARgamma in hepatic stellate cells (HSCs) and on the biological characteristics of HSCs. METHODS: The activated HSCs were divided into three groups: control group, 3 micromol/L rosiglitazone group, and 10 micromol/L rosiglitazone group. The expression of PPARgamma, alpha-smooth muscle actin (alpha-SMA), and type I and III collagen was detected by RT-PCR, Western blot and immunocytochemical staining, respectively. Cell proliferation was determined with methylthiazolyltetrazolium (MTT) colorimetric assay. Cell apoptosis was demonstrated with flow cytometry. RESULTS: The expression of PPARgamma at mRNA and protein level markedly increased in HSCs of 10 micromol/L rosiglitazone group (t value was 10.870 and 4.627 respectively, P<0.01 in both). The proliferation of HSCs in 10 micromol/L rosiglitazone group decreased significantly (t = 5.542, P<0.01), alpha-SMA expression level and type I collagen synthesis ability were also reduced vs controls (t value = 10.256 and 14.627 respectively, P<0.01 in both). The apoptotic rate of HSCs significantly increased in 10 micromol/L rosiglitazone group vs control (chi(2) = 16.682, P<0.01). CONCLUSION: By increasing expression of PPARgamma in activated HSCs, rosiglitazone, an agonist of PPARgamma, decreases alpha-SMA expression and type I collagen synthesis, inhibits cell proliferation, and induces cell apoptosis.     

Preparation of monoclonal antibody against apoptosis—associated antigens of hepatoma cells by subtractive immunization

AIM：To elucidate the expression of the apoptosis-associated molecules in human primary hepatocellular carcinoma(HCC)cells,and prepare the monoclonal antibodies(mAb)against the apoptosis-associated antigens of HCC cells.METHODS:Human HCC cell line HCC-9204 cells were induced apoptosis with 60mL&#183;L^-1 ethanol for 6h and their morphological changes were obserevd by transmission electron microscope,The cell DNA fragmentations were detcted by Terminal Deoxynucleotidyl transferase-mediated dUTPnick end labeling(TUNEL)assay,and the cell DNA contents by flow cytometry.Ten mice were immunized with ethanol-induced apoptoticHCC-9204 cells with the method of subtractive immunization.while the other 10mice used as the control were immunized by the routin procedures.The tail blood of all the mice were prepared after the last immunization.and the produced antibodies were determined by the immunocytochemical ABC staining .The splenic cells of the mice whose tail blood sera-HCC-9204 cells serum reactions were most different between the apoptotic and the non-apoptotic were prepared an fused with the mouse myeloma cell lineSP2/0cells.The positive antibodies were selected by ELISA assay.The fusion rates of hybridona cells and the producing rates of antibodies were calculated.The fused cells that secreted candidate objective antibody were cloned continually with the of limited dilution method.and then selected and analyzed further by the immunocytochemical ABCstaining.The chromosomes of the cloned hybridoma cells that secreted objective mAb and the mAb immunoglobulin(Ig)subtpe of the prepared mAb were also determined.The molecular mass of the mAb associated antigen was analyzed by Western blot assay.RESULTS:HCC-9204 cells treated with60mL&#183;L^-1 ethanod for 6h,manifested obvious apoptotic morphological changes,the majority of the cells wereTUNEL-positive,and the sub-G1 apoptotic peak was evident.There were 2mice in the experimental group whose tail blood serum reacted strongly with the apoptotic HCC-9204 cells,but weakly with their non-apoptotic counterparts,In the fusion rates of hybridoma cells as well as the producing rates of the antibody deseribed above,there did not show significant difference between the experimental and the control group,but weakly with non-apoptoticHCC-9204.However,the total producing rate of antibodies in the experimental group wa significantly lower compared with the control(P&lt;0.01).and so was the producing rate of the Antibodies which racted strongly with both paoptotic and non-apoptotic HCC-9204cells(P&lt;0.01).After cloned continually for several times the cell that produce mAb which reacted strongly with the nuclei of ethanol-induced apoptotic HCC-9204cells,but very weakly with that of non-apoptotic cells was selected out.Chromosme analysis revealed that the selected cell was with the universal characteristics of the monoclonal hybridoma cells which secreted mAb,and the Ig subtype of the prepared mAb was IgG1,The molecular mass of this mAb associated antigen of was about75ku.CONCLUSION :Subtractiv immunization is a useful method to prpar the mAb against the apoptosis-associated antigens of cells,The expression of some molecules increases to some extent in HCC-9204cells in the process of apoptosis induced by low-concentration ethanol.The mAb that may be against ethanol-induced apoptosis-associated antigens of HCC cells was successfully prepared and primarily identified.     

Inhibition on the production of collagen type Ⅰ,Ⅲ of activated hepatic stellate cells by antisense TIMP-1 recombinant plasmid

AIM: To investigate the inhibition effects on the production of collagen type Ⅰ, Ⅲ secreted by activated rat hepatic stellate cells (rHSCs) by antisense tissue inhibitors of metalloproteinase 1 (TIMP-1) recombinant plasmid through elevating interstitial collagenase activity.METHODS: rHSCs were extracted from normal rat liver by pronase and collagenase digestion and purified by centrifugal elutriation, and were cultured on plastic dishes until they were activated to a myofibroblastic phenotype after 7-10 days. RT-Nest-PCR and gene recombinant techniques were used to construct the rat antisense TIMP1 recombinant plasmids which can express in eucaryotic cells. The recombinant plasmid and the pcDNA3 empty plasmid were transfected in rHSCs by Effectene (QIAGEN)separately. Cells were selected after growing in DMEM containing 400μg/ml G418 for 2-3 weeks. Expression of exogenous gene was assessed by Northern blot, and expression of TIMP-1 in rHSCs was determined by Northern blot and Western blot. We tested the interstitial collagenase activity with FITC-labled type I collagen as substrate.Ultimately, we quantified the type Ⅰ,Ⅲ collagen byWestern blot.RESULTS: The exogenous antisense TIMP-1 recombinant plasmid could be expressed in rHSCs well, which could block the expression of TIMP-1 greatly, the ratio of TIMP-1/GAPDH was 0.67, 2.41, and 2.97 separately at mRNA level (P&lt;0.05); the ratio of TIMP-1/β-actin was 0.31, 0.98 and 1.32 separately at protein level (P&lt;0.05), It might elevate active and latent interstitial collagenase activity,the collagenase activity was 0.3049, 0.1411 and 0.1196 respectively. (P&lt;0.05), which led to promotion the degradation of type Ⅰ, Ⅲ collagen, the ratio of collagen I/β-actin was 0.63, 1.78 and 1.92 separately (P&lt;0.05); and the ratio of collagen Ⅲ/β-actin was 0.59, 1.81 and 1.98separately (P&lt;0.05).CONCLUSION: These data shows that the antisense TIMP-1 recombinant plasmid has the inhibitory effects on the production of type Ⅰ,Ⅲ collagens secreted by activated rHSCs in vitro. It could be a novel method to reverse hepatic fibrosis in the future.     

Downregulation of matrix and β-PDGF receptor gene expression by anti-TGFβ antibody in rat hepatic stellate cells during experimental liver injury

Excess matrix in hepatic fibrosis results from both fibrogenic stimulation of stellate cells by TGFβ1 and cell proliferation due to induction of β-platelet derived growth factor receptor (β-PDGFR). In this paper, treatment of culture-activated rat stellate cells with anti-TGFβ inhibited collagen and fibronectin mRNA expression by 82 and 58%, respectively, versus control cells. In vivo, anti-TGFβ inhibited collagen I gene expression by 86% in stellate cells isolated from rats treated with CC14 compared with control antibody. In contrast to stellate cells, anti-TGFβ had no effect on collagen I gene expression in isolated sinusoidal endothelial cells. Anti-TGFβ administered in vivo to rats with liver injury also reduced expression of stellate cell β-PDGFR mRNA to that of control animals. Anti-TGFβ antibody had no effect on the histologic appearance of the tissue. These data support a role for TGFβ in stellate cell matrix expression and provide evidence for transmodulation of PDGF receptor by TGFβ in vivo. However, inhibition of TGFβ alone may not be adequate to attenuate severe hepatic injury and fibrosis.     

三七总皂甙诱导大鼠肝星状细胞凋亡的研究

三七及其主要有效成分三七总皂甙（PNS）具有抗肝纤维化作用。采用体外培养的传代大鼠肝星状细胞（HSC），利用有关凋亡检测技术，观察PNS对LISC增殖、调亡，胶原和层黏连蛋白（LN）合成的影响，从细胞水平探讨PNS抗肝纤维化机制。     

Apoptosis of hepatic stellate cells in carbon tetrachloride induced acute liver injury of the rat: analysis of isolated hepatic stellate cells

BACKGROUND/AIMS: Analysis of isolated hepatic stellate cells (HSCs) from the injured liver may provide direct information on HSC apoptosis. However, it has not been established whether apoptotic HSCs would be isolated using the usual density gradient centrifugation method. The aim of this study was to observe the serial pattern of proliferation and apoptosis in isolated HSCs in comparison with that of liver tissue sections in CCl4 induced acute liver injury. METHODS: Male Sprague-Dawley rats were treated with a single intraperitoneal injection of carbon tetrachloride (CCl4) and were killed at various time points after the treatment. RESULTS: HSC proliferation showed a maximal increase at 32 h after CCl4 injection. Apoptosis of HSC, examined by quantitative analysis of annexin-V-fluorescein isothiocyanate (FITC)staining, showed the maximal increase at 64 h. Apoptosis of HSC in liver tissue sections examined by counting desmin and Tdt-mediated-dUTP biotin nick end labeling (TUNEL) double staining cells, peaked at 64 h. The number of TUNEL positive HSCs in liver tissue sections correlated significantly with annexin-V-FITC binding of isolated HSC. CONCLUSIONS: Studying apoptosis using apoptotic HSCs isolated by a usual density gradient centrifugation method from injured tissue sections would be feasible since it correlated with in vivo apoptosis of HSC.     

雷帕霉素诱导大鼠肝星状细胞凋亡的研究

目的观察雷帕霉素(RAPA)对大鼠肝星状细胞系rHSCs-99增殖、凋亡的影响。方法将rHSCs-99分成4组:对照组(A组),RAPA 50 nmol/L组(B组),RAPA 100 nmol/L组(C组),RAPA 150 nmol/L组(D组)。RAPA作用72 h后,分别用MTT法检测rHSCs-99增殖,ELISA法检测I型胶原表达,流式细胞仪Annexin-V FICT/PI法检测细胞凋亡情况,Wright-Giemsa染色法观察细胞形态学改变,细胞免疫组化法检测Fas、P53与Bcl-2的表达变化。结果 RAPA作用后,rHSCs-99增殖受到抑制,I型胶原含量降低,凋亡率增高,Fas、P53表达增多,Bcl-2表达减少。结论 RAPA能抑制rHSCs-99的增殖及I型胶原合成。促进rHSCs-99凋亡,其机制可能是通过开启Fas/FasL凋亡途径及上调凋亡相关基因P53、下调Bcl-2的表达来实现。     

体外RNA干扰下调黏着斑激酶表达对HSC-T6细胞黏附与迁移的影响

目的 探讨特异性阻断黏着斑激酶(FAK)表达对纤维连接蛋白刺激的肝星状细胞(HSC-T6)黏附与迁移的影响.方法 构建靶向FAK的RNA干扰重组体,在阳离子聚合物介导下转染大鼠肝星状细胞系HSC-T6,筛选出可高效抑制FAK表达的重组质粒;荧光实时定量PCR和Western blot检测FAK基因敲除效果;甲苯胺蓝染色法检测细胞黏附,划痕修复实验和改良的Boyden双腔系统检测细胞迁移.多组间均数差异性比较采用单因素方差分析.结果 成功构建并筛选出可高效抑制FAK的质粒表达载体.质粒转染后,FAK mRNA和蛋白表达分别下降了76.82%和72.53%,同时,p-FAK(Tyr397)蛋白表达下降了62.71% FAK表达下调可明显抑制HSC-T6细胞黏附,抑制率约58.69%;FAK基因沉默可显著抑制纤维连接蛋白诱导的HSC迁移,使细胞迁移距离降低了58.27%,跨膜迁移细胞数减少了83.70%. 结论 RNA干扰技术可选择性下调HSC中FAK的表达,并可显著抑制HSC-T6的黏附和迁移.     

Hepatic immune tolerance induced by hepatic stellate cells

The liver, which is a metabolic organ, plays a pivotal role in tolerance induction. Hepatic stellate cells(Hp SCs), which are unique non-parenchymal cells, exert potent immunoregulatory activity during cotransplantation with allogeneic islets effectively protecting the islet allografts from rejection. Multiple mechanisms participate in the immune tolerance induced by Hp SCs, including the marked expansion of myeloid-derived suppressor cells(MDSCs), attenuation of effector T cell functions and augmentation of regulatory T cells. Hp SC conditioned MDSC-based immunotherapy has been conducted in mice with autoimmune disease and the results show that this technique may be promising. This article demonstrates how Hp SCs orchestrate both innate immunity and adaptive immunity to build a negative network that leads to immune tolerance.     

细胞外信号调节激酶在精-甘-天冬-丝氨酸四肽诱导肝星状细胞凋亡中的作用

目的 探讨细胞外信号调节激酶(ERK)在精-甘-天冬丝氨酸(RGDS)四肽诱导肝星状细胞(HSC)凋亡中的作用。方法 将培养的HSC细胞分为6组：①空白对照组，②纤维连接蛋白(FN)组，③FN RGDS(25mg／L)组，④FN RGDs(50mg／L)组，⑤FN RGDS(100mg／L)组，⑥FN 精-甘-谷-丝氨酸(RGES，100mgL)组。采用3H-胸腺嘧啶核苷(3H-TdR)掺入法测定HSC增生；透射电镜和末端脱氧核苷酸转移酶介导的脱氧三磷酸尿苷缺口末端标记(TUNEL)方法测定HSC凋亡；甲苯胺兰染色法测定细胞粘附率；分别应用Western印迹和逆转录-聚合酶链反应(RT-PCR)技术测定ERK蛋白及其mRNA表达。结果 ①25、50、100mg／L浓度的RGDS四肽呈剂量依赖性抑制HSC增生并诱导HSC凋亡(P＜0．05)。②RGDS四肽作用于HSC 2h，HSC粘附率下降(P＜0．01)。③在RGDS四肽于预组，HSC的ERK及其mRNA表达下调。结论 RGDS四肽可抑制HSCs增生并诱导其凋亡；RGDS四肽诱导HSCs凋亡的效应与其抗粘附作用、抑制ERK蛋白和ERKI mRNA表达有关。     

硝普钠诱导肝星状细胞HSC-T6凋亡及其机制

目的:探讨一氧化氮(nitric oxide,NO)供体硝普钠(sodium nitroprusside,SNP)在诱导肝星状细胞(hepatic stellate cell.HSC-T6)凋亡中的作用及其机制.方法:应用流式细胞仪和Hoechst 33258染色法检测HSC凋亡:激光扫描共聚焦显微镜检测荧光标记NF-kB p65的核转位:Real-time PCR方法检测基质金属蛋白酶抑制因子-1(tissue inhibitors of matrix metalloproteinase-1,TIMP-1)、Ⅰ型前胶原(Procollagen Ⅰ)、抗凋亡蛋白基因(growth arrest and DNA damage-inducible protein.GADD4513)mRNA表达.结果:SNP组HSC凋亡率较对照组显著增加(20.78%±5.91% vs 3.25%±1.26%,P=0.031),Hoeehst 33258染色法显示SNP组HSC细胞核呈致密浓染块状或颗粒状的荧光,提示细胞出现凋亡:SNP抑制TNF介导活化的HSC NF-kB p65核转位;随着其剂量不断增加,TIMP-1,Procollagen Ⅰ,GADD45β mRNA表达在随之减少(P<0.05).结论:SNP能诱导HSC凋亡,减少TIMP-1,Procollagen Ⅰ mRNA表达,其机制可能与通过抑制NF-kB活性,减少抗凋亡蛋白基因GADD45β mRNA表达有关.