卡托普利对缺氧诱导的肺动脉平滑肌细胞增殖和胶的合成的抑制作用

AIM: To study the effect of captopril (Cap) on hypoxia-induced proliferation and collagen synthesis in vascular smooth muscle cells (VSMC). METHODS: VSMC were isolated from rabbit pulmonary artery. Cultured VSMC were evaluated by incorporation of [3H]thymidine and [3H]proline, cell number, and intracellular calcium concentration ([Ca2+]i). RESULTS: Pretreatment of pulmonary VSMC with Cap 1 mumol.L-1 blocked hypoxia-induced increase in cell number and incorporation of [3H]proline and [3H]thymidine, which were decreased 25%, 21%, and 36%, respectively, as compared with hypoxic control. It also inhibited the increase of intracellular Ca2+ concentration under hypoxic condition. Addition of nifedipine inhibited hypoxia-stimulated increase in the collagen, DNA synthesis, and [Ca2+]i. Bay-K-8644 increased cell number (35%), DNA (55%), collagen synthesis (36%), and [Ca2+]i (33%) in pulmonary VSMC, that was completely abolished by Cap 1 mumol.L-1. CONCLUSION: Cap inhibited hypoxia-induced proliferation and collagen synthesis in VSMC.     

丝裂素活化蛋白激酶反义寡核苷酸对血管紧张素Ⅱ诱导的大鼠心肌？…

AIM: To investigate the inhibitory effect of down-regulating mitogen activated protein kinase (MAPK) on c-myc gene expression and further on cardiac fibroblast proliferation. METHODS: Cultured neonatal rat cardiac fibroblasts was pretreated with a phosphorothioate-protected 17-mer antisense MAPK oligodeoxynucleotide (ODN) directed against the initiation of translation sites of the p42 and p44 MAPK isoforms by liposomal transfection. A 17-mer sense and mismatch sequence MAPK ODN were used as controls. After liposomal transfecting, cells were exposed to angiotensin II (Ang II) 10 nmol.L-1 for 5 min and then harvested in lysis buffer. MAPK activity was measured by Western blot and P-81 phosphocellulose filter paper method by using [gamma-32P]ATP and myelin basic protein as substrates. c-myc mRNA expression stimulated by Ang II for 30 min was measured by Northern blot. DNA synthesis and collagen protein synthesis induced by Ang II for 24 h were measured by [3H]thymidine incorporation and [3H]Proline incorporation, respectively. RESULTS: Antisense ODN 0.2 mumol.L-1 reduced Ang II-induced MAPK activities by 72%, MAPK protein expression by 80%, and suppressed c-myc mRNA expression by 97%, respectively. [3H]thymidine incorporation and [3H]proline incorporation in Ang II-induced cardiac fibroblast were inhibited by 59% and 58%, respectively. CONCLUSION: A 17-mer MAPK antisense oligonucleotide directed againsts the initiation of translation sites of MAPK could specifically inhibit Ang II-stimulated cultured neonatal rat cardiac fibroblast proliferation through down-regulating MAPK activity and further depleting c-myc mRNA expression.     

血小板源生生长因子刺激血管平滑肌细胞增殖及其分子机制

目的：探讨血小板源生生长因子（PDGF－BB）刺激血管平滑肌细胞（VSMC）增殖及其分子机制。方法：用Western Blot法测定p44/p42 CCDPK活性。[^3H]脱氧胸腺嘧啶核苷酸掺入测定VSMC DNA合成。原位杂交检测c-myc mRNA的表达。结果：PDGFBB诱导的磷酸化CCDPK蛋白表达和[^3H]脱氧胸腺嘧啶核苷酸掺入呈浓度依赖性,此作用可被PTK抑制剂Genistein,外钙络合剂依他酸和MEK抑制剂PD 98059抑制。PDGF－BB刺激可引起c-myc mRNA的明显表达,此作用可被PD 98059抑制。结论：PDGF－BB通过激活p44/p42 CCDPK,上调c-myc mRNA的表达从而促进VSMC增殖,其作用是由PTK和Ca^2 介导的。     

反义碱性成纤维细胞生长因子寡核苷酸对血管紧张素Ⅱ诱导培养的 …

AIM: To study the effect of antisense basic fibroblast growth factor (bFGF) oligonucleotides (ODN) transfection on the growth of cultured aortic smooth muscle cells (SMC) in spontaneously hypertensive rats (SHR). METHODS: Using cationic liposome-mediated method, antisense bFGF ODN were introduced into SMC, bFGF gene expression was detected by Northern blotting, cell hyperplasia was evaluated by [3H] thymidine incorporation and cell counting. RESULTS: Transfection of antisense bFGF ODN (5 mumol.L-1) almost completely inhibited enhanced bFGF mRNA expression and inhibited cell proliferation induced by angiotensin II (Ang 1 mumol.L-1). In basal state and Ang-stimulated state, [3H]thymidine incorporation was inhibited by 26.5% (P < 0.01) and 42.0% (P < 0.01) and cell number was inhibited by 17.3% (P < 0.01) and by 22.2% (P < 0.01), respectively. CONCLUSION: The transfection of antisense bFGF ODN into cultured SMC effectively suppressed bFGF mRNA expression and inhibited the SMC proliferation induced by Ang.     

Inhibitory effect of antisense oligodeoxynucleotide to p44／p42 MAPK on angiotensin II-induced hypertrophic response in cultured neonatal rat cardiac myocyte

AIM: To explore the inhibitory effect of antisense oligonucleotide (ODN) to mitogen activated protein kinase(MAPK) on cardiomyocyte hypertrophy induced by angiotensin Ⅱ (Ang Ⅱ). METHODS: A 17-mer phosphorothioate-protected antisense ODN directed against the initiation of translation sites of the p42 and p44 MAPK isoforms byliposomal transfection was applied to inhibit the translation of p44/p42 MAPK mRNA. The sense and random ODNs to p44/p42MAPK were used as sequence controls. Neonatal cardiac myocytes were exposed to Ang Ⅱ (10nmol/L) for 5 min and then harvested in lysis buffer for the measurement of the activity and the phosphorylated protein content of p44/p42MAPK that were tested by P-81 phosphocellulose filter paper method and Western blotting, respectively. The rate of protein synthesis by [^3H]leucine incorporation and the diameter of cell were measured after exposure to Ang Ⅱ for 24 h and 72 h, respectively. RESULTS: In cardiac myocyte Ang Ⅱ increased p44/p42MAPK activity and phosphorylated protein content by 140 % and 699 %, and also increased [^3H]leucine incorporation and cell diameter by 40 % and 27 %. c-fos and c-myc mRNAs were induced significantly after exposure to Ang Ⅱ. Antisense ODN to p44/p42MAPK (0.2 μmol/L) reduced Ang Ⅱ-induced MAPK activity by 30 %,and phophorylated MAPK protein expression by 59 % in cardiac myocyte, and inhibited c-fos and c-myc mRNA expression induced by Ang Ⅱ by 44 % and 43 %, respectively. The diameter and the rate of protein synthesis of cardiac myocyte induced by Ang Ⅱ were decreased by 16 % and 22 % after pretreatment with antisense ODN to p44/p42MAPK. CONCLUSION: Antisense ODN to p44/p42 MAPK inhibited the increase of rate of protein synthesis,and the augmentation of cell diameter and expression of c-fos and c-myc mRNA induced by Ang Ⅱ in culturedcardiac myocytes, p44/p42 MAPK played a critical role in the hypertrophic response induced by Ang Ⅱ in cultured neonatal rat cardiac myocytes.     

糖基化终产物对培养的大鼠主动脉平滑肌细胞增殖及胞浆游离钙含…

目的：研究糖基化终产物（ＡＧＥＰ）对主动脉平滑肌细胞增殖的影响及其与［Ｃａ＾２＋］ｉ的关系。方法：采用同位素掺入法分别测定ＤＮＡ和蛋白质合成；Ｆｕｒａ２－ＡＭ测定［Ｃａ＾２＋］ｉ。结果：ＡＧＥＰ以浓度、时间相关的方式促进［＾３Ｈ］ＴｄＲ与［＾Ｈ］Ｌｅｕ掺入细胞，随ＡＧＥＰ作用时间、糖化时间延长，掺入率增加明显，ＡＧＥＰ增加［Ｃａ＾２＋］ｉ，与时间、浓度相关，但随ＡＧＥＰ作用时间延长（４０分钟后）而     

Effects of histamine on protein and glycoprotein production of isolated pig gastric mucosal cells

The production of glycoprotein and protein by isolated pig gastric non-parietal cells was measured by incorporation of N-acetyl-[14C]-D-glucosamine [( 14C]GlcNAc) and [3H]-L-leucine [( 3H]Leu), respectively, into acid insoluble material (AIM). Histamine enhanced incorporation of the tracers into cellular and released AIM in a concentration-dependent manner. The H2 receptor antagonist ranitidine completely blocked the effects of histamine (100 mumol/l) on [3H]Leu incorporation into cellular and released AIM (IC50 37 and 32 mumol/l, respectively) but had no inhibitory effect on the 16,16-dimethylprostaglandin-E2 - and forskolin-stimulated tracer incorporation. The H1 receptor antagonist mepyramine did not inhibit the histamine effect. We conclude that histamine is a stimulant of protein, via H2 receptors, and glycoprotein production of isolated pig gastric non-parietal cells.     

丹参酮ⅡA对AngⅡ诱导的心肌肥大中c-fos、c-myc和c-jun mRNA表达的影响

目的在原代培养的新生大鼠心肌细胞上,观察丹参酮ⅡA(tanshinoneⅡ,TSN)对血管紧张素Ⅱ(angiotensinⅡA,AngⅡ)诱导的心肌细胞肥大的影响。方法实验用新生大鼠,差数贴壁法体外分离培养大鼠心肌细胞。用血管紧张素Ⅱ诱导心肌细胞肥大,丹参酮ⅡA和缬沙坦(valsartan)进行干预;采用相差显微镜测量细胞大小;测定心肌细胞3H-亮氨酸参入作为心肌细胞肥大的指标;用逆转录聚合酶链式反应(RT-PCR)检测心肌细胞原癌基因c-fos、c-myc和c-junmR-NA的表达;MTT法检测细胞活力。结果用MTT法检测发现,培养的心肌细胞中加入TSN(5~80μmol·L-1),24h后会产生微弱的细胞毒性,但心肌细胞单层在TSN存在的情况下可以继续同步收缩。在AngⅡ持续作用7d后,血管紧张素Ⅱ组心肌细胞直径增大(28·5±3·8)μm,与对照组(19·8±1·9)μm相比差异有显著性(P<0·05);TSN抑制AngⅡ介导的心肌细胞直径增大(21·3±2·5)μm,与AngⅡ组相比差异有显著性(P<0·05),AngⅡ作用24h后,心肌细胞合成速率AngⅡ组(1900±100)cpm较对照组(1205±129)cpm明显增加(P<0·01),TSN和valsartan本身对心肌细胞蛋白质的合成没有影响,但均能抑制AngⅡ刺激的心肌细胞蛋白质合成速率的增加(P<0·01)。在培养液中加入AngⅡ作用30min后,c-fos、c-myc和c-junmRNA的表达明显增强(P<0·01),预先加入TSN或valsartan作用30min,可阻断AngⅡ的作用(P<0·01),而TSN和valsartan本身对原癌基因c-fos、c-myc和c-junmRNA的表达无影响。结论TSN可以抑制AngⅡ诱导的心肌细胞肥大,这可能与其抑制了原癌基因c-fos、c-myc和c-jun的表达有关。