Experimental Study of Salvia Miltiorrhiza on the Prevention of Restenosis after Angioplasty

Ｅｘｐｅｒｉｍｅｎｔａｌ　Ｓｔｕｄｙ　ｏｆ　Ｓａｌｖｉａ　Ｍｉｌｔｉｏｒｒｈｉｚａ　ｏｎ　ｔｈｅ　Ｐｒｅｖｅｎｔｉｏｎ　ｏｆ　Ｒｅｓｔｅｎｏｓｉｓ　ａｆｔｅｒ　ＡｎｇｉｏｐｌａｓｔｙＺＨＯＵＸｉａｏ－ｍｉｎｇ，ＬＵＺａｉ－ｙｉｎｇａｎｄＷＡＮＧＤａｏ...     

高浓度氧抑制兔动脉平滑肌细胞的增殖

目的　探讨缺氧和高氧环境对兔主动脉平滑肌细胞增殖的抑制作用 ,间接判断缺氧与冠脉再狭窄的关系和研究氧疗是否能用于 PTCA支架后冠脉再狭窄的防治 .方法　用含 10 0 m L· L- 1小牛血清的 RPMI16 40培养液体外培养新西兰白兔的主动脉平滑肌细胞至 3～ 7代 ,将含 5 0 m L· L- 1小牛血清的 RPMI16 40培养液与动脉平滑肌细胞的混悬液10 0 μL 加入 96孔培养板中培养 ,按照实验分组分别在 10 0 ,2 10 ,2 5 0 ,5 0 0和 75 0 m L· L- 1 O2 及纯氧环境中培养主动脉平滑肌细胞 48h.用 MTT法和 3H- Td R掺入法检测动脉平滑肌细胞增殖的数量 .结果　以 2 10 m L· L- 1 O2 环境下动脉平滑肌细胞增殖的数量为基准对照组 ,其他各组为实验组 .5 0 0 m L· L- 1 O2 至纯氧环境下动脉平滑肌细胞的增殖显著低于对照组 (P<0 .0 5 ) ,2 5 0 m L· L- 1 O2 与 2 10 m L·L- 1 O2环境下主动脉平滑肌细胞的增殖无显著差异 (P>0 .0 5 ) ,10 0m L· L- 1 O2 环境下动脉平滑肌细胞的增殖显著高于对照组(P<0 .0 5 ) .结论　动脉平滑肌细胞的增殖随着培养环境中氧浓度升高而降低 ,缺氧促进动脉平滑肌细胞的增殖 ,高氧对动脉平滑肌细胞增殖具有显著的抑制作用 ;氧疗可能有助于降低冠脉再狭窄率 ,并为高压氧治疗其他以动脉血管     

川芎嗪对抗原诱发致敏豚鼠气管平滑肌环的舒张作用

目的:观察川芎嗪对过敏原诱发的哮喘豚鼠离体气管平滑肌环收缩的影响及其相关机制。方法:采用卵蛋白致敏法建立豚鼠哮喘动物模型,每只豚鼠制备5～7个气管平滑肌环,悬挂于恒温浴槽中,并将其与力位移换能器相连来纪录气管环张力的变化,所有标本分为正常KH液和无钙KH液2大组。正常KH液组又分为卵蛋白组、10mmol/L川芎嗪+卵蛋白组、30mmol/L川芎嗪+卵蛋白组、100mmol/L川芎嗪+卵蛋白组、300mmol/L川芎嗪+卵蛋白组、thapsigargin(TSG)+卵蛋白组、TSG+川芎嗪(100mmol/L)+卵蛋白组。无钙KH液组分为卵蛋白组和川芎嗪(100mmol/L)+卵蛋白组。结果:①卵蛋白组平滑肌环收缩产生的张力为(59.7±8.5)×10-6N,分别预先加入30mmol/L、100mmol/L、300mmol/L的川芎嗪20min后,再加入卵蛋白,平滑肌环产生的张力分别降低至(44.1±5.5)×10-6N、(20.7±20.4)×10-6N、(10.0±1.1)×10-6N(P均<0.05),10mmol/L的川芎嗪对卵蛋白的作用无明显影响(P>0.05);②去除溶液中的Ca2+后,再加入卵蛋白,平滑肌环收缩产生的张力为(10.5±1.9)×10-6N,预先加入100mmol/L的川芎嗪后,卵蛋白诱导产生的平滑肌环的收缩力无明显改变(P>0.05);③预先加入TSG(10mmol/L)40min后,再加入卵蛋白,平滑肌环收缩产生的张力为(27.3±3.4)×10-6N,分别预先加入TSG和100mmol/L的川芎嗪后,再加入卵蛋白诱导,平滑肌环的收缩力减少至(6.5±1.1)×10-6N(P<0.05)。结论:①川芎嗪可能通过抑制细胞膜上的钙通道来抑制抗原引起的哮喘豚鼠离体气管平滑肌的收缩作用。②抗原可能通过细胞外的Ca2+内流和细胞内肌浆网对Ca2+的释放来促进哮喘豚鼠离体气管平滑肌的收缩。     

Inhibitory effect of iron on in vitro proliferation of smooth muscle cells

Background Iron is a biocorrodible metal that might be used in bioabsorbable stents.This study investigated the effects at the cellular and protein levels of soluble divalent iron （ferrous gluconate） and soluble trivalent iron （ferric chloride） on the proliferation of human aortic smooth muscle cell （HASMC） in vitro.Methods The water-soluble tetrazolium （WST-1） test was used to evaluate the effect of iron on proliferation of HASMC and Western blotting was used to measure the levels of signaling proteins involved in proliferative and apoptosis pathways.Results HASMC proliferation was inhibited in a concentration dependent manner after treatment with soluble divalent and trivalent iron at concentrations of 100-500 μmol/L.Western blotting analysis showed that the proliferating cell nuclear antigen （PCNA） expression following treatment with soluble divalent iron and trivalent iron at 100,300 and 500 μmol/L was reduced compared to the control.The PCNA expression decreased with increasing iron concentration and to a greater extent with the trivalent iron than with the divalent iron treatment group.The p53 expression was markedly increased in a concentration dependent manner in both iron treatment groups.Conclusion The soluble divalent iron and,to a greater degree trivalent iron,inhibited HASMC proliferation in a dosedependent manner,which may be attributed to reduction of PCNA expression and increase of p53 expression.     

三氧化二砷诱导血管平滑肌细胞凋亡的实验研究

目的：探讨三氧化二砷（Ａｓ２Ｏ３）诱导大鼠血管平滑肌细胞（ＶＳＭＣｓ）调亡的作用。方法：建立大鼠ＶＳＭＣｓ增生模型。用四甲基偶氮唑蓝还原反应（ＭＴＴ法）、透射电镜、琼脂糖凝胶电泳、流式细胞术等方法观察Ａｓ２Ｏ３诱导ＶＳＭＣｓ凋亡的作用。结果：经Ａｓ２Ｏ３作用后ＶＳＭＣｓ出现典型的凋亡变化,透射电镜下可风细胞核仁消失,染色质浓缩聚集于核膜下。ＤＡ琼脂糖凝胶电泳出现凋亡特征性梯形电泳条带。汉式细胞术检     

川芎嗪对脑组织的保护作用

目的 研究川芎嗪对脑损伤的保护作用。方法 采用小鼠断头法、亚硝酸钠（NaNO2）致死法、大气大腑中动脉栓塞法，通过观察断头喘息时间、亚硝酸钠致死时间、脑缺血行为评分、脑缺血面积，明确川芎嗪对脑损伤的保护作用。结果 川芎嗪20mg／kg、50mg／kg可明显延长断头小鼠的喘息时间及亚硝酸钠致死小鼠的生存时间；10mg／kg、20mg／kg可显著改善中动脉栓塞致脑缺血（MCAO）大鼠的行为障碍，降低脑缺血梗死面积。结论 川芎嗪对缺氧缺血导致的脑损伤具有明显的保护作用。     

川芎嗪对顺铂耳毒性保护作用的实验研究

目的探讨活血化瘀中药川芎嗪是否对顺铂的耳毒性具有保护作用。方法将听力正常的30只豚鼠随机分为3组,每组10只。对照组：生理盐水3 ml/（kg.d）腹腔注射6天;顺铂组：顺铂3 mg/（kg.d）腹腔注射6天;顺铂加川芎嗪组：先腹腔注射川芎嗪140 mg/（kg.d）3天,从第4天起腹腔注射川芎嗪140 mg/（kg.d）后再注射顺铂3 mg/（kg.d）6天。给药前后通过听觉脑干诱发电位（ABR）检测豚鼠听功能的变化;然后在麻醉状态下断头取耳蜗,半数标本切片后用DNA末端转移酶介导的缺口末端标记法（TUNEL）检测耳蜗内细胞凋亡情况;半数标本进行耳蜗基底膜铺片观察毛细胞形态及酶学变化。结果顺铂组用药后ABR阈值升高,毛细胞受损,琥珀酸脱氢酶（SDH）活性减弱,有部分毛细胞凋亡,与对照组及川芎嗪组比较差异具有统计学意义（P〈0.05）。顺铂加川芎嗪组用药后ABR阈值升高,毛细胞受损较轻,凋亡细胞较少,与顺铂组比较差异有统计学意义（P〈0.05）。结论川芎嗪能降低顺铂的耳毒性。     

3-羟基苯甲醛抑制兔自体移植静脉桥内膜增生的研究

目的:研究3-羟基苯甲醛(3-HBA)对兔搭桥后静脉桥内膜增生的影响.方法:24只新西兰兔计算机随机均分实验组和对照组,建立颈外静脉移植到颈总动脉实验模型,实验组动物予以耳缘静脉注射3-HBA 100mg/(kg﹒d),对照组动物予以同剂量生理盐水.于术后的2周、4周采集各组移植血管,计算移植血管桥内膜厚度,Western Blot检测AKT蛋白活化情况.人大隐静脉平滑肌细胞进行体外培养,不同浓度3-HBA作用于平滑肌细胞,MTT法检测细胞增殖,Transwell小室法检测细胞迁移.结果:2周时,实验组静脉桥内膜厚度较之对照组稍有减小(76.57±6.76μm vs 87.32±6.89μm),p=0.123,差异无统计学意义;4周时实验组较对照组内膜厚度明显变薄(45.39±9.41μm vs 134.07±18.80μm),P<0.05.p-AKT在实验组中的表达明显降低,P<0.05.相比于对照组,实验组大隐静脉平滑肌细胞增殖与迁移能力均受到抑制,P<0.05.结论: 3-HBA能抑制实验用新西兰兔自体移植静脉桥内膜增生,可能与其抑制AKT的磷酸化从而抑制平滑肌细胞增殖迁移有关.     

Inhibitory effect of iron on in vitro proliferation of smooth muscle cells

at the cellular and protein levels of soluble divalent iron (ferrous gluconate) and soluble trivalent iron (ferric chloride) on the proliferation of human aortic smooth muscle cells (HASMC) in vitro.

Methods The WST-1 test was used to evaluate the effect of iron on proliferation of HASMC and Western blot was used to measure the levels of signaling proteins involved in proliferative and apoptosis pathways.

Results HASMC proliferation was inhibited in a concentration dependent manner after treatment with soluble divalent and trivalent iron at concentrations of 100-500μmol/L. Western blot analysis showed the proliferating cell nuclear antigen (PCNA) expression following treatment with soluble divalent iron and trivalent iron at 100, 300 and 500μmol/L was reduced compared to the control. The PCNA expression decreased with increasing iron concentration and to a greater extent with the trivalent iron than with the divalent iron treatment group. The p53 expression was markedly increased in a concentration dependent manner in both iron treatment groups.

Conclusions The soluble divalent iron and, to a greater degree trivalent iron, inhibited HASMC proliferation in a dose-dependent manner, which may be attributed to reduction of PCNA expression and increase of p53 expression.

     

牛磺酸对肺动脉内皮和肺动脉平滑肌细胞增殖的双向调节作用

目的探讨肺动脉高压发生发展的细胞机制和牛磺酸对缺氧性肺动脉高压的防治作用。方法体外培养新生雄性小牛肺动脉内皮细胞(PAECs)及肺动脉平滑肌细胞(PASMCs),用3H-TdR掺入法比较缺氧时PASMCs与PAECs的增殖情况,同时测定在培养液中加入牛磺酸后其增殖的改变。结果2.5%O2和<1%O2条件下培养6~24 h,PAECs的3H-TdR掺入量均显著低于常氧对照组(降低15%~48%,P<0.05);而PASMCs于缺氧12 h后3H-TdR掺入量均显著高于常氧对照组(增加19%~34%,P<0.05)。牛磺酸对细胞增殖的影响与浓度有关,高浓度(10~20 mmol/L)的牛磺酸抑制缺氧的内皮及平滑肌细胞的3H-TdR掺入,而低浓度的牛磺酸促进缺氧时PAECs的3H-TdR掺入(P<0.05),抑制缺氧时PASMCs的3H-TdR掺入(P<0.05)。结论缺氧抑制内皮细胞的增殖而促进平滑肌细胞的增殖,适当的牛磺酸可以对抗缺氧对PAECs和PASMs的作用,减弱缺氧对PAECs的增殖抑制作用,抑制缺氧的促PASMCs增殖作用,使之接近常氧水平。     

大鼠再狭窄模型的建立和内膜增生时相变化的观察

目的:为进一步探讨血管成形术后再狭窄的发生机制和血管损伤后内膜增生的时相变化。方法:取30只Sprague—Dawley大鼠行右颈总动脉球囊内膜剥脱术建立再狭窄动物模型，于第1，3，9和28，60天取实验动脉段和对侧正常动脉段行光镜、电镜观察及计算机图像分析。结果:实验发现在术后第3天内膜即明显增厚，并在第28天达到高峰，此后不再继续增加，同时通过电镜观察发现这种时相特征与平滑肌细胞的表型转化有关。结论:在大鼠再狭窄模型上的内膜增生与再狭窄发生过程类似，具有明显的时相特征，并与平滑肌细胞的表型转化有关。     

肝细胞生长因子对冠状动脉平滑肌细胞增殖迁移的影响

为观察肝细胞生长因子(HGF)对牛冠状动脉平滑肌细胞(BCASMC)增殖、迁移的影响,分离和培养BCASMC,采用四甲基偶氮唑蓝法观察HGF对BCASMC增殖的作用,倒置显微镜观察BCASMC的迁移。结果显示,对照组、HGF组BCASMC的D值相差显著(P<0.05),HGF组的BCASMC增殖率为45.2％,且迁移明显。提示HGF能促进BCASMC的增殖和迁移。     

Evidence of apoptotic smooth muscle cells in proliferative intima of injured arteries

Objective To investigate the occurrence and extent of apoptosis in the course of restenosis. Methods The experimental models of vessel narrowness and intima thickness were established in minipigs’ iliac arteries by balloon injury and specimens were retrieved on the 1st, 3rd, 6th,12th and 30th days for dynamic observation. Apoptotic smooth muscle cells (SMCs) were detected by terminal deoxynucleotidyl transferase- mediated dUTP nick- end labeling (TUNEL). Results Apoptotic SMCs occurred only in the thickened intima 12 days after injury accompanied with the proliferative SMCs, the percentage of apoptosis was 1. 94%±0. 42% on the 12th day and 1. 36%±0. 31% on the 30th day respectively. The low frequency of apoptosis compared with the proliferative SMCs was a feature in the restenotic pathology. Conclusions Apoptosis participates in the pathogenetic process of intimal thickening and its level was low compared with proliferation. The findings suggest that attempts to modulate apoptosis after vessel injury constitute a theoretical approach to the prevention of restenosis.     

菠(艹棱)子平喘作用的实验研究

作者通过试验,初次证实,菠(艹棱)子能解除由组胺引起的豚鼠离体支气管平滑肌痉挛,有解痉平喘作用。其中以911(0.25ml/ml浓度)作用最显著。     

The Effect of Ligustrazine on Peritoneal Transport in Peritoneal dialysis

In order to investigate the effect of ligustrazine (Lig)i.p.on peritoneal permeability in peritoneal dialysis and its side effects,creatinine was given intravenously and continuously to maintain the high plasma creatinine level.All the rabbits were divided into three groups:normal control group (goup A),group B treated with 0.12% Lig and group C treated with 0.24% Lig.The peritoneal dialysis of all rabbits lasted 2h.The plasma and dialysate levels of glucose,protein and creatining were observed immediate,30min,60min,90min,120min after dialysis.Creastinine dialysate/plasma ratio (D/P),protein D/P ratio,grucose D/Do at different time points after dialysis and creatinine mass transfer area coefficient (MTAC) at 120min were calculated.The structures of peritoneum were observed under optical microscope and electron microscope after continuously intraperitoneal injection of Lig for 14 days.The results showed that the 90-min and 12-min creatinine D/P ratios in the group C were higher than in the group A.The 120-min creatinine MATC in the group C was higher than in the group A.The rabbits treated with Lig did not show significant structure changes of peritoneum and signs of peritoneal irritation.It was suggested that Lig could increase mass transfer ability of peritoneum without significant side effects.     

亮氨酸脑啡肽对脂多糖促血管平滑肌细胞增殖效应的抑制作用

观察亮氨酸脑啡肽和脂多糖对血平滑肌细胞增殖的影响，并观察两者之间的相互作用及阿片受体阻滞剂纳洛酮的影响，方法：实验以离体培养的ＳＤ大鼠胸主动脉ＶＳＭＣ为模型。     

壳多糖对兔主动脉平滑肌细胞和内皮细胞增殖的影响

目的：研究壳多糖对体外培养的兔主动脉平滑肌细胞和内皮细胞生长的影响。方法：将不同质量浓度（０．２,２,２０,２００,２０００μｇ／ｍｌ）的壳多糖溶液分别加入兔主动脉平滑肌细胞和内皮细胞培养液中,培养１２,２４,４８ｈ后用ＭＴＴ经色法测定细胞数。结果：平滑肌细胞数随着壳多糖质量浓度和作用时间的增加而减少;内皮细胞数随着其质量浓度和作用时间的增加,在一定范围内相应增加。结论：壳多糖能抑制兔主动脉平滑肌     

Experimental study of Hirudo andSalvia miltiorrhiza on smooth muscle cell hyperplasia following balloon angioplasty

Percutaneoustransluminalcoronaryangioplasty(PTCA)isanimprotantmethodtotreatcoronaryarterystenosisandmyocardialischemia.ButtheapplyingofPTCAwererestrictedbythecausingofrestenosisfollowingangioplasty(').Sofartherearenoeffectivewaytopreventrestenosis.TOprobetheeffectoftraditionalChinesemedicineonrestenosis,theHirudoandSalviamiltiorrhiza(SM)werestudiedinthisexperiment,Theeffectsofthetwomedicinesonhyperplasiaofvascularsmoothmusclecells(VSMCs)wereobserved.'GrantssupportedbyBeijingNaturalSci…     

反义TR和p21双基因共表达对人主动脉平滑肌细胞增殖与凋亡的作用

目的观察携带反义凝血酶受体(ATR)或/和p21的单、双基因重组腺病毒伴随病毒(rAAV)载体对人主动脉平滑肌细胞(ASMC)增殖与凋亡的影响。方法以携带ATR或/和p21的单、双基因rAAV感染人ASMC。半定量RT-PCR检测各基因的整合与表达;MTT法测定感染不同时间点的细胞存活率;流式细胞术检测细胞周期与凋亡细胞数目的变化;吖啶橙/溴化乙锭(AO/EB)染色检测凋亡细胞的比率。结果单、双基因在ASMC中均获得整合,且双基因发生了共表达;病毒感染4d时,ATR组、p21组和AP双基因组的细胞存活率与对照组相比分别降低了16.7%、21.6%和29.4%;三组的G0/G1期细胞数分别为(61.8±2.9)%、(82.5±4.0)%和(80.4±6.1)%;凋亡细胞数为(4.8±0.5)%、(5.7±0.1)%和(9.2±0.9)%;AO/EB染色显示的凋亡细胞比率分别为:对照组(1.5±0.8)%、ATR组(7.2±3.3)%、p21组(10.7±5.6)%、AP组(18.3±2.7)%。结论(1)双基因共表达对抑制细胞增殖和诱导凋亡的作用均较单基因具有较强的生物学效应。(2)为再狭窄的基因治疗提示了更优化的途径。