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1.
AIM: To investigate the effect of Kangxian ruangan keli (KXR)on hepatic stellate cell (HSC) proliferation mediated by platelet-derived growth factor (PDGF) and the underlying mechanism.METHODS: In a serum-free culture system, HSCs were treated with a KXR preparation for 24 hours, followed by stimulation with PDGF-BB for 24 hours. Then the cells were incubated again in the medium containing KXR for 3 hours stimulated with PDGF-BB for 5 minutes, and collected. The proliferation of HSC was examined using an MTT assay and flow cytometry. Tyrosine phosphorylation was detected with Western blotting and visualized by the enhenced chemiluminescent (ECL) method.RESULTS: The OD values for the HSCs growing in the media without and with addition of PDGF were 0.17±0.06 and 0.82±0.05, respectively. The PDGF-induced increase was hindered remarkably by KXR preparation in a dosedependent manner. The reaction values for the systems with 5 mg/mL, 2.5 rng/mL and 1.25 mg/mL of KXR were 0.28±0.03,0.37±0.02 and 0.43±0.04, respectively. Mloreover, the percentages of S-phase cells in these KXR-containing culture systems were 10.95±1.35, 32.76±1.07 and 43.19±1.09,respectively, all of which were significantly lower than that in the culture free of KXR (68.24±2.72). In addition, the values for tyrosine-phosphorylated protein in HSCs treated with 5 mg/mL and 1.25 mg/mL of KXR were 0.1349±0.0072and 0.1658±0.0025, respectively, which were smaller than that in the cells treated only with PDGF-BB (0.1813±0.0117).CONCLUSION: Within the dose range used in the present study, KXR preparation shows an inhibitory effect on HSC proliferation induced by PDGF. The mechanism of this process may involve interference with tyrosine phosphorylation mediated by PDGF.  相似文献   

2.
目的:研究抗纤软肝颗粒(KXR)对血小板源生长因子(PDGF)诱导的肝星状细胞(HSC)增殖的影响.方法:采用无血清培养,不同浓度的KXR温育HSC 24 h后,PDGF-BB(10 ng/ml)刺激24 h,再加入上述浓度的KXR 3 h后,又加入PDGF-BB(10 ng/ml)作用5 min,然后收集细胞.采用细胞计数法及流式细胞仪测定细胞增殖及细胞周期.结果:无血清培养显示该方对于PDGF诱导的细胞增殖具有抑制作用,并呈剂量依赖性(P<0.01).流式细胞术分析提示KXR能抑制PDGF诱导的HSC DNA合成期及合成后期和分裂期DNA百分含量,阻断细胞由静止期/DNA合成前期向DNA合成期转化,呈剂量依赖性(5 mg/ml组作用最显著,P<0.01).结论:KXR能抑制PDGF诱导的HSC增殖.  相似文献   

3.
AIM: To investigate the role of nuclear factor-κB (NF-κB)inhibitor caffeic acid phenethy1 ester (CAPE) in the proliferation, collagen synthesis and apoptosis of hepatic stellate cells (HSCs) of rats. METHODS: The HSCs from rats were isolated and cultured in Dulbecco's Modified Eagle's Medium (DMEM) and treated with CAPE. The proliferation and collagen synthesis of HSCs were determined by 3H-TdR and 3H-proline incorporation respectively, and the expression of type Ⅰ, Ⅲ procollagen genes was further explored byin situ hybridization. Apoptosis cell indices (AIs) were examined using terminal deoxynucleotidyl transferase- mediated DIG-dUTP nick end labeling (TUNEL). RESULTS: Tn activated HSC in culture, CAPE significantly inhibited 3H-TdR and 3H-proline incorporation by HSCs at concentrations of 5 μmol/L and 10 μmol/L respectively. CAPE also reduced the type I procollagen gene expression (P<0.05)at higher concentration. Apoptosis of HSC was induced by CAPE and the AIs were time-and dose-dependently increased from 2.82+0.73 % to 7.66±1.25 % at 12 h (P<0.01) and from 3.15±0.88 % to 10.6L±2.88 % at 24 h (P<0.01). CONCLUSION: CAPE inhibits proliferation and collagen synthesis of HSC at lower concentration and induces HSC apoptosis at higher concentration.  相似文献   

4.
The proliferation of hepatic stellate cells (HSCs) is a critical step in hepatic fibrogenesis. Platelet-derived growth factor (PDGF) is the most potent mitogen for HSCs. We investigated the role of nonphagocytic NAD(P)H oxidase-derived reactive oxygen species (ROS) in PDGF-induced HSC proliferation. The human HSC line, LI-90 cells, murine primary-cultured HSCs, and PDGF-BB were used in this study. We examined the mechanism of PDGF-BB-induced HSC proliferation in relation to the role of a ROS scavenger and diphenylene iodonium, an inhibitor of NAD(P)H oxidase. We also measured ROS production with the aid of chemiluminescence. We showed that PDGF-BB induced proliferation of HSCs through the intracellular production of ROS. We also demonstrated that HSCs expressed key components of nonphagocytic NAD(P)H oxidase (p22phox, gp91phox, p47phox, and p67phox) at both the messenger RNA and protein levels. Diphenylene iodonium suppressed PDGF-BB-induced ROS production and HSC proliferation. Coincubation of H2O2 and PDGF-BB restored the proliferation of HSCs that was inhibited by diphenylene iodonium pretreatment. Phosphorylation of the mitogen-activated protein kinase (MAPK) family constitutes a signal transduction pathway of cell proliferation. Our data demonstrate that NAD(P)H oxidase-derived ROS induce HSC proliferation mainly through the phosphorylation of p38 MAPK. Moreover, an in vivo hepatic fibrosis model also supported the critical role of NAD(P)H oxidase in the activation and proliferation of HSCs. In conclusion, NAD(P)H oxidase is expressed in HSCs and produces ROS via activation of NAD(P)H oxidase in response to PDGF-BB. ROS further induce HSC proliferation through the phosphorylation of p38 MAPK.  相似文献   

5.
BACKGROUND & AIMS: The Na+/H+ exchanger is the main intracellular pH regulator in hepatic stellate cells (HSCs), and its activity is increased by platelet-derived growth factor (PDGF). Amiloride, an Na+/H+ exchange inhibitor, reduces PDGF-induced HSC proliferation, suggesting that the Na+/H+ exchanger plays a role in regulating HSC proliferative response. The aim of this study was to characterize the intracellular pathways mediating activation of the Na+/H+ exchanger by PDGF in HSCs. METHODS: The activity of the Na+/H+ exchanger and HSC proliferation rate were evaluated under control condition and after incubation with PDGF in the absence or presence of specific inhibitors of the main intracellular pathways of signal transduction. Na+/H+ exchange protein expression was evaluated by means of Western blot. RESULTS: PDGF induced a significant increase in the activity of the Na+/H+ exchanger without modifying protein expression. Inhibition of the calcium/calmodulin- and protein kinase C-dependent pathways resulted in a significant inhibition of both Na+/H+ exchange activity and of PDGF-induced HSC proliferation. The involvement of the two pathways was confirmed by showing that incubation of HSCs with both phorbol-12-myristate-13-acetate, a potent protein kinase C activator, and thapsigargin, which increases intracellular calcium levels, significantly increased both the Na+/H+ exchanger activity and HSC proliferation rate. Inhibition of the protein kinase A pathway did not modify either PDGF-induced Na+/H+ exchange activation or PDGF-induced HSC proliferation. On the contrary, inhibition of the mitogen-activated protein kinase- and of phosphatidylinositol 3-kinase-dependent pathways significantly reduced PDGF-induced HSC proliferation without affecting the activity of the Na+/H+ exchanger. CONCLUSIONS: Activation of the Na+/H+ exchanger by PDGF in HSCs is mediated by calcium/calmodulin- and protein kinase C-dependent pathways. PDGF-induced HSC proliferation is mediated by Na+/H+ exchange-dependent and -independent pathways.  相似文献   

6.
AIM: Liver fibrosis is a common pathological process of chronic liver diseases. Activation of hepatic stellate cells (HSCs) is the key issue in the occurrence of liver fibrosis. In this study, we observed the inhibitory action of rat serum containing Biejiajian oral liquid (BOL), a decoction of turtle shell, on proliferation of rat HSCs, and to explore the anti-hepatofibrotic mechanisms of BOL. METHODS: A rat model of hepatic fibrosis was induced by subcutaneous injection of CCl(4). Serum containing low, medium and high dosages of BOL was prepared respectively. Normal and fibrotic HSCs were isolated and cultured. The effect of sera containing BOL on proliferation of HSCs was determined by (3)H-TdR incorporation. RESULTS: The inhibitory rate of normal rat HSC proliferation caused by 100 mL/mL sera containing medium and high dosages of BOL showed a remarkable difference as compared with that caused by colchicine (medium dosage group: 34.56+/-4.21% vs 29.12+/-2.85%, P<0.01; high dosage group: 37.82+/-1.32% vs 29.12+/-2.85%, P<0.01). The inhibitory rate of fibrotic rat HSC proliferation caused by 100 mL/L serum containing medium and high dosages of BOL showed a remarkable difference as compared with that caused by colchicine (medium dosage group: 51.31+/-3.14% vs 38.32+/-2.65%, P<0.01; high dosage group: 60.15+/-5.36% vs 38.32+/-2.65%, P<0.01). The inhibitory rate of normal rat HSC proliferation caused by 100 mL/L and 200 mL/L sera containing a medium dosage of BOL showed a significant difference as compared with that caused by 50 mL/L (100 mL/L group: 69.02+/-9.96% vs 50.82+/-9.28%, P<0.05; 200 mL/L group: 81.78+/-8.92% vs 50.82+/-9.28%, P<0.01). The inhibitory rate of fibrotic rat HSC proliferation caused by 100 mL/L and 200 mL/L sera containing a medium dosage of BOL showed a significant difference as compared with that caused by 50 mL/L (100 mL/L group: 72.19+/-10.96% vs 61.38+/-7.16%, P<0.05; 200 mL/L group: 87.16+/-8.54% vs 61.38+/-7.16%, P<0.01). CONCLUSION: Rat serum containing BOL can inhibit proliferation of rat HSCs, and the inhibition depends on the dosage and concentration of BOL. The inhibitory effect on HSC proliferation is one of the main anti-hepatofibrotic mechanisms of BOL.  相似文献   

7.
目的 观察内源性大麻素N-花生四烯酸氨基乙醇(AEA)及大麻素受体(CBR)2对肝星状细胞(HSC)增殖活化的影响,以探讨内源性大麻素及其受体系统在肝纤维化发展中的作用.方法 采用免疫荧光观察血小板衍生生长因子(PDGF)刺激前后HSC中CBR1和CBR2的表达.Western blot、PCR法观察不同浓度AEA及CBR2拮抗剂AM630对PDGF刺激下HSC增殖及活化的影响,同时用四甲基偶氮唑盐、流式细胞仪分析AEA对HSC活力及凋亡的影响.结果 HSC中CBR2的表达较CBR1高(F=116.797,P<0.01),且PDGF刺激后CBR2的表达明显增强(F=7.878,P<0.05).AEA可剂量依赖地抑制HSC的增殖,在浓度为10,20、50μmol/L时抑制率分别为7.12%±0.34%、12.52%±0.78%、80.13%±1.57%,差异有统计学意义(F=533.41,P<0.01);但对HSC凋亡的影响不明显.同时AEA可抑制HSC的活化指标α-平滑肌肌动蛋白、转化生长因子β1、Ⅰ型胶原、Ⅲ型胶原及基质金属蛋白酶抑制因子等的表达,但这种抑制作用在给予CBR2拮抗剂AM630后明显减弱,差异有统计学意义(P<0.05).结论 CBR2在AEA引起的HSC增殖及活化抑制中起关键作用,AEA和CBR2可望成为肝纤维治疗的新靶点.  相似文献   

8.
AIM:Hepatic stellate cell(HSC)plays a pivotal role in liver fibrosis and is considered as the therapeutic target for the treatment of hepatic fibrosis,Tyrosine protein kinase plays an important role in the proliferation,activation of HSC.The purpose of the study is to investigate the effects of the tyosine protein kinase inhibitor genistein on the proliferation and activation of cultured rat HSC.METHODS:Rat HSC were isolated from Wistar rats by in situ perfusion of collagenase and pronase and single-step density Nycodenz gradient,Culture-activated HSC were serum-starved and incubated with10^-9to10^-5mol/L concentration of genistein for 24,48or 72h,In PDGF-induced HSC proliferation,HSC were stimulated with10μg&#183;L^-1PDGF-BBfo15min,and thentreated with genistein for the same time.Cell proliferation was measured by MПassay and based on flow cytometric analysis of cell cycle.The a-smooth muscle actin(α-SMA)expression in HSC was studied with confocallaser microscopy and flow cytometry.c-fos,c-jun and cyclinD1expression in HSCwas also detected by flow cytometry.RESULTS:Genistein inhibited basal and PDGF-induced proliferation of HSCat the concentration of 10^-8to10^-5mol/L,and treatment with10^-7mol/L concentration of genistein for 48h inhibited the HSCproliferation significantly(the inhibition rate was 70.3%,P&lt;0.05).Immunofluorescence detected by confocal laser microscopy and flow cytometry showed that treatment with10^-7mol/L genistein for48h suppressed the expression of α-SMA significantly in HSC(the specific fluorescence intensity were60.2&#177;21.5vs35.3&#177;11.6and12.8&#177;10.4vs9.54&#177;6.39,respectively,bothP&lt;0.05).The intensity of c-fos,c-jun and cyclinD1 expression of HSCs treated with 10^-7mol/L genistein for 48h was also significantly decreased compared with the controls.CONCLUSION;Genistein influences proliferation of HSC,suppresses the expression of α-SMA in HSC and tinhibits the intensity of c-fos,c-jun and cyclinD1 expression of HCSs,Genistein has therapeutic potential against liver fibrosis.  相似文献   

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目的:探讨莪术提取物药物血清对PDGF诱导的肝星状细胞(HSC)内Ca2 、PI3-K的影响。方法:制备药物血清,不同浓度的莪术提取物药物血清温育肝星状细胞24小时后,PDGF-BB(10ng/ml)刺激24小时,再加入上述浓度的莪术提取物药物血清,3小时后,又加入PDGF-BB(10ng/ml)作用5分钟,然后收集细胞。采用钙指示剂Fura-2/Am测定细胞内Ca2 ,免疫印迹化学发光法检测PI3-K的表达。结果:不同浓度的莪术提取物药物血清与同浓度的生理盐水药物血清比较,能显著抑制PDGF诱导的胞内Ca2 浓度升高(P<0.01)。经10%莪术提取物药物血清处理的HSC,其PDGF诱导的PI3-K表达水平较10%生理盐水药物血清组显著降低(P<0.01)。结论:莪术提取物药物血清能抑制PDGF诱导的肝星状细胞内Ca2 内流及PI3-K的表达。  相似文献   

12.
In the present study, we analyzed the effect of conditioned media (CM) from bovine aortic endothelial cells exposed to laminar shear stress (SS) of 5 dyne/cm2 (SS5) or 15 dyne/cm2 (SS15) for 16 hours on smooth muscle cell (SMC) migration. In response to CM from bovine aortic endothelial cells exposed to SS5 (CMSS5) and SS15 (CMSS15), migration was 45 +/- 5.5 and 30 +/- 1.5 cells per field, respectively (P<0.05). Similar results were obtained with SS of 2 versus 20 dyne/cm2 and also when SS of 5 and 15 dyne/cm2 lasted 24 hours. Platelet-derived growth factor (PDGF)-AA levels in CMSS5 and CMSS15 were 9 +/- 7 and 18 +/- 5 ng/10(6) cells for 16 hours, respectively (P<0.05); PDGF-BB levels in CMSS5 and CMSS15 were 38 +/- 10 and 53 +/- 10 ng/10(6) cells for 16 hours, respectively (P<0.05). PDGF receptor alpha (PDGFRalpha) and PDGF receptor beta (PDGFRbeta) in SMCs were phosphorylated by CMSS15>CMSS5. In response to CMSS15, a neutralizing antibody against PDGF-AA enhanced SMC migration to a level comparable to that of CMSS5; in contrast, antibodies against PDGF-BB abolished SMC migration. Transfection of SMCs with a dominant-negative PDGFRalpha or PDGFRbeta increased or inhibited, respectively, SMC migration in response to CMSS15. Overexpression of wild-type PDGFRalpha inhibited SMC migration in response to CMSS5, CMSS15, or recombinant PDGF-BB (P<0.001). These results suggest that the ability of high SS to inhibit arterial wall thickening in vivo may be related to enhanced activation of PDGFRalpha in SMCs by PDGF isoforms secreted by the endothelium.  相似文献   

13.
Proliferation and activation of hepatic stellate cells (HSCs) are critical steps for the development of postnecrotic fibrosis in the liver. The present study aimed to reveal the inhibitory effect of the semisynthetic analogue of fumagillin TNP-470 on these events for its possible use as an antifibrogenic agent. Rat models of carbon tetrachloride (CCl(4))- and dimethylnitrosamine-induced hepatic fibrosis were used for an in vivo study. In both models, the fibrotic area was considerably decreased by concurrent repetitive subcutaneous injections of 30 mg/kg body weight of TNP-470. In CCl(4)-induced fibrosis, factor VIII-related antigen-positive blood vessels, desmin-, or alpha-smooth muscle actin (alphaSMA)-positive mesenchymal cells, bromodeoxyuridine (BrdU)-positive mesenchymal cells also decreased in number by treatment with TNP-470. In in vitro experiments, a supplement of 1,000 ng/mL TNP-470 suppressed BrdU incorporation and cyclins D1, D2, and E expression by cultured HSCs in the absence and/or presence of platelet-derived growth factor (PDGF). Expression of HSC activation markers, i.e., alphaSMA and PDGF receptor beta, was also suppressed. The present results indicate that TNP-470 inhibits HSC proliferation by blocking the cell-cycle transition from G1 to S and HSC activation, and, as the consequence, prevents the progression of hepatic fibrosis, probably being coupled with its antiangiogenic effect.  相似文献   

14.
[目的]观察肌细胞增强因子2A (Myocyte enhancer factor 2A、MEF2A)在肝星状细胞(Hepatic Stellate Cell、HSC)活化过程中的可能作用及四甲基吡嗪对其影响,进一步探讨四甲基吡嗪抗肝纤维化的作用机制.[方法]采用SD大鼠肝脏原位灌流消化法获得HSC,并原代及传代培养,分离的HSC在塑料细胞培养皿中培养后逐步活化,分别收集新分离(0 d)和经培养(1、2、3、4、5、6、7、8 d)的HSC,观察MEF2A与HSC活化指标α-平滑肌肌动蛋白(α-smooth muscle action、α-SMA)表达的关系;取传2~4代HSC,加四甲基吡嗪至终浓度50、100、150、200mg/L,分组培养48 h,观察四甲基吡嗪对HSC增殖抑制作用的同时,对MEF2A基因及蛋白表达的影响.[结果]①随着HSC培养逐步活化,细胞内MEF2A及α-SMA逐步增加,二者呈正相关(r=0.90,P<0.05),且MEF2ADNA结合活性同步增强.②随着四甲基吡嗪浓度增加,对HSC增殖的抑制作用增强,同时对MEF2A表达抑制作用逐渐增强.[结论]MEF2A可能参与了HSC活化过程,四甲基吡嗪抑制HSC增殖作用可能与有效抑制MEF2A表达有关.  相似文献   

15.
Activated hepatic stellate cells (HSCs) are a potential source of gelatinase A, which accumulates in fibrotic livers. Progelatinase A activation requires its binding to a complex of membrane-type matrix metalloproteinase (MT-MMP) and tissue inhibitor of metalloproteinases (TIMP)-2. These studies examine gelatinase A, MT1-MMP, and TIMP-2 synthesis by HSCs during activation in vitro and the potential role of gelatinase A in promoting HSC proliferation. Gelatinase A, MT1-MMP, and TIMP-2 messenger RNA (mRNA) were all upregulated in HSCs activated on plastic over 5 to 14 days. Gelatinase A expression was maximal at 7 days of culture, coinciding with the peak of HSC proliferation and the onset of procollagen I and alpha-smooth muscle actin (alpha-SMA) mRNA expression. Active forms of gelatinase A of 62 kd and 66 kd were secreted by activated HSCs and reached a maximum of 12.1% of total enzyme in 14-day culture supernatants. Treatment of HSCs with concanavalin A (con A) induced activation of MT1-MMP and enhanced secretion of activated gelatinase A, which reached a maximum of 44.4% of the total enzyme secreted into culture supernatants using 30 microgram/mL con A. [(14)C]-gelatin degradation assays confirmed the presence of gelatinolytic activity in activated HSC supernatants, which reached a maximum level at 7 days of culture. Antisense oligonucleotide inhibition of endogenous progelatinase A production, or the MMP inhibitor 1,10-phenanthroline inhibited (3)H-thymidine incorporation into HSC DNA by greater than 50%. We conclude that HSCs produce progelatinase A during activation in vitro and activate this enzyme coincident with MT1-MMP and TIMP-2 synthesis. Gelatinase A activity is required for maximal proliferation of HSCs in vitro suggesting this metalloproteinase is an autocrine proliferation factor for HSCs.  相似文献   

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Ma G  Mason DP  Young DB 《Hypertension》2000,35(4):948-951
The effect of potassium on the migration of vascular smooth muscle cells was analyzed in media made with extracellular potassium concentrations of 3, 4, 5, and 6 mmol/L. The migration of cultured porcine coronary artery cells was stimulated with platelet-derived growth factor (PDGF)-BB. In the first study, cells were exposed to PDGF-BB at concentrations of 0, 10, or 20 ng/mL for 5 hours with the use of a Boyden chamber. Cells were quiescent overnight in 0.5% fetal bovine serum in Dulbecco's modified Eagle's medium with an extracellular potassium concentration of 4 mmol/L. With increasing potassium concentration, migration was significantly inhibited (P<0. 02, 2-way ANOVA). In the cells exposed to 10 ng/mL PDGF-BB, migration ranged from 500+/-86% to 294+/-44% (value in wells with 0 ng/mL PDGF-BB and 4 mmol/L potassium concentration=100%) in medium containing 3 to 6 mmol/L extracellular potassium concentration (P<0. 03). Long-term potassium exposure was investigated in cells grown in 5% serum in Dulbecco's modified Eagle's medium with an extracellular potassium concentration of 3, 4, 5, or 6 mmol/L for 3 to 4 weeks. Migration was assessed with 0 or 20 ng/mL PDGF-BB. Migration was significantly inhibited by the elevation of extracellular potassium concentration (P<0.01, 2-way ANOVA). With 20 ng/mL PDGF-BB, the migration rates ranged from 152+/-11% in medium with 3 mmol/L potassium to 69+/-5% in 6 mmol/L potassium (P<0.01). Increases in extracellular potassium concentration within the physiological range significantly and directly inhibit vascular smooth muscle cell migration.  相似文献   

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AIM:To investigate the effects of IH764-3 on HSC apoptosis,and the expression of caspase-3 protein in SHC apoptotic process.METHODS:HSCs were cultured in medium with different IH764-3 doses(10μg&#183;mL^-1,20μg&#183;mL^-1,30μg&#183;mL^-1,40μg&#183;mL^-1)and without IH 764-3,and HSC proliferation was quantitatively measured by 3H-thymidine incorporation,The morphological changes of HSCs were observed with transmission electron microscope after exposure to the dose of 40μg&#183;mL^-1 of IH 764-3 for 48 hr.The apoptosis rates were detected by annexin v/PI and TdT-mediated dUTP nick end labeling (TUNEL),The expression of caspase-3 protein was determined by flow cytometry.RESULTS:(1) HSC proliferation rates induced with different IH764-3 doses(10μg&#183;mL^-1,20μg&#183;mL^-1,30μg&#183;mL^-1m,40μg&#183;mL^-1)Were significantly reduced compared with that of the control group(P&lt;0.01).(2)With the doses above,IH764-3 dose-dependently produced HSC apoptosis rate of 6.7%(9.4%),9.3%(21.6%),15.1%(27.2%).and 19.0%(28.4%)respectively,by annexin V and PI-labeled flow cytometry assay (or TUNEL),while it was only 2.3%(6.7%),in the control.(3) The expression of caspase-3 protein in IH 764-3 groups was significantly higher than that of the control(P&lt;0.05).CONCLUSION:Within the dose range used in present study IH 764-3 can inhibit HSC proliferation,as well as enhance HSC apopptosis.Furthermore,IH764-3 can significantly increase the caspase-3 prtein expression.  相似文献   

20.
黄芪注射液对肝纤维化抑制作用的实验研究   总被引:16,自引:0,他引:16  
目的探讨黄芪注射液对大鼠肝星状细胞(HSC)和肝纤维化的作用。方法体外细胞实验: 用不同浓度黄芪注射液(0、25、50、100、200、400 mg/ml)作用HSC不同时间(24、48、72h)后,采用四甲基偶氮唑盐法检测其活化增殖;流式细胞术检测HSC增殖周期;溴乙锭/吖啶橙荧光染色和流式细胞术检测HSC凋亡。动物实验:用40%四氯化碳和5%乙醇制备大鼠肝纤维化动物模型,实验分为正常对照组、模型组和黄芪注射液组。黄芪注射液组和模型组在模型制备的同时分别给予黄芪注射液(800 mg·kg-1·d-1)和等渗盐水腹腔注射,第8周时测定血清透明质酸(HA),层黏连蛋白(LN)水平及肝组织中超氧化物歧化酶(SOD)活性,丙二醛(MDA)含量,免疫组织化学方法观察肝组织LN的表达,苏木素-伊红、苦味酸-酸性品红染色观察肝组织病理改变。结果在体外细胞实验中,与0 mg/ml组比较,黄芪注射液其它浓度组明显抑制了HSC增殖,并呈剂量和时间依赖性;HSC增殖周期被抑制在G2-M期;黄芪注射液各浓度组荧光染色法和流式细胞术均未检测到HSC凋亡。在体内实验中,血清HA、LN含量:模型组分别为(114.3±25.6)μg/L和(78.8±11.7)μg/L,黄芪注射液组分别为(85.6±37.3)μg/L和(66.8±17.6)μg/L,P <0.05;肝组织SOD活性黄芪注射液组为(75.9±5.9)NU/mg,模型组为(49.6±5.7)NU/mg,P< 0.01;而MDA含量黄芪注射液组为(2.4±0.2)μmol/g,模型组为(3.7±0.4)μmol/g,P<0.01。显微镜下黄芪注射液组肝纤维化程度明显轻于模型组,免疫组织化学结果黄芪注射液组肝组织LN表达明显减少。结论黄芪注射液可延缓肝纤维化的发生,其机制除可直接抑制HSC增殖外,还有抗氧化、抗脂质过氧化、减少LN产生,防止肝窦毛细血管化等作用。  相似文献   

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