本文關鍵詞：�；撬釋Υ笫蠓蝿用}平滑肌細胞凋亡的影響　出處：《哈爾濱商業(yè)大學》2011年碩士論文　論文類型：學位論文

  更多相關文章： 牛磺酸 肺動脈平滑肌細胞 細胞凋亡 線粒體途徑 死亡受體途徑

【摘要】：肺動脈高壓是指靜息時肺動脈平均壓3.33kPa(25mmHg)或運動時4kPa(30mmHg)者。肺動脈平滑肌細胞是肺血管的主要成分,肺動脈平滑肌細胞的增殖與凋亡失衡會引起肺血管官腔狹窄、閉塞,肺血管阻力增加,最終導致肺動脈高壓。肺動脈高壓的兩個重要的的病理生理改變,即肺動脈血管收縮和肺動脈血管的重塑。而由于重構主要是由于破壞細胞的動態(tài)平衡,主要包括抑制細胞的凋亡或促進細胞的增殖兩個方面。而且,肺動脈平滑肌細胞,作為肺動脈血管的主要構成成分,是肺動脈高壓血管重構的主要執(zhí)行者。因此研究調控PASMCs的增殖與凋亡能為預防和治療肺動脈高壓尋找治療的方法提供理論基礎。 細胞凋亡是細胞的一種基本生物學現象,在多細胞生物去除不需要的或異常的細胞中起著必要的作用。它在生物體的進化、內環(huán)境的穩(wěn)定以及多個系統的發(fā)育中起著重要的作用。細胞凋亡不僅是一種特殊的細胞死亡類型,而且具有重要的生物學意義及復雜的分子生物學機制。 �；撬�(Tau)幾乎存在于所有動物細胞中,大多以游離形式存在,它是機體一種內源性抗損傷物質,Tau具有很多生物學效應,例如保持內鈣穩(wěn)態(tài),穩(wěn)定細胞膜,清除體內自由基等。有研究表明Tau在某些細胞中也具有誘導細胞凋亡的作用。我們選用大鼠肺動脈平滑肌細胞為研究對象,觀察Tau對它的作用。 目的：觀察Tau對肺動脈平滑肌細胞(PASMCs)凋亡作用的影響,并探討其機制是否通過線粒體途徑和死亡受體途徑。 方法：取Wistar雄性大鼠肺動脈平滑肌細胞培養(yǎng)；分為正常組(control)、凋亡組(SD)、給藥組(Tau高、中、低劑量)。給藥前,按需要將所有組用低糖無血清培養(yǎng)基饑餓24h,使細胞處于相同生長狀態(tài),按需要給藥24h或48h后,MTT法測定96孔板細胞的生存率；用吖啶橙(AO)法,在熒光顯微鏡下觀察6孔板細胞核形態(tài)學變化；用線粒體膜電位試劑盒測定細胞膜電位的變化；用Western-blot法分別測定Bax、Bcl-2、Procaspase-9、Procaspase-3和Fas的蛋白表達變化。 結果：MTT測定細胞生存率發(fā)現,和正常組相比,加藥組生存率明顯低于正常組,并和凋亡組的趨勢相同,可以明顯看出呈劑量依賴性誘導肺動脈平滑肌細胞凋亡,且40mmol/L的Tau給藥量為最佳濃度。吖啶橙法三組結果可以看出,凋亡組和Tau給藥組的細胞核皺縮,形成凋亡小體。線粒體膜電位法測定,發(fā)現凋亡組和Tau組相對正常組,膜電位降低,圖中出現較多綠色熒光區(qū)域。(?)Vestern-blot法測定發(fā)現Tau升高Bax和Fas蛋白表達,降低Procaspase-3、Procaspase-9和Bcl-2的表達。 結論：Tau誘導大鼠PASMCs凋亡,其作用機制可能與調節(jié)線粒體途徑和死亡受體途徑有關。
[Abstract]:Pulmonary hypertension refers to resting pulmonary arterial pressure (25mmHg) or 3.33kPa 4kPa (30mmHg) during exercise. Pulmonary artery smooth muscle cells is the main component of pulmonary vascular, imbalance of proliferation and apoptosis of pulmonary artery smooth muscle cells can cause pulmonary vascular lumens stenosis, occlusion, increased pulmonary vascular resistance, resulting in pulmonary hypertension pulmonary hypertension. Two important pathophysiological changes, namely pulmonary vasoconstriction and pulmonary vascular remodeling. Because the reconstruction is mainly due to the dynamic balance of destruction of cells, including inhibition of cell apoptosis and promote cell proliferation in two aspects. Moreover, pulmonary artery smooth muscle cells, as the main form the pulmonary artery is the main component of high pressure to perform remodeling of the pulmonary arteries. Therefore study on the regulation of proliferation and apoptosis of PASMCs can provide the theory for the prevention and treatment of pulmonary arterial hypertension for treatment Basics.
Apoptosis is a basic biological phenomenon of cells, plays a necessary role in multicellular organisms to remove unwanted or abnormal cells. It is in the evolution of organisms, plays an important role in the stability of the environment and the development of multiple systems. Apoptosis is not only a kind of special types of cell death. It also has important biological significance and complex molecular mechanisms.
Taurine (Tau) exists in almost all animal cells, mostly in the free form, it is a endogenous anti injury substance, Tau has many biological effects, such as maintaining calcium homeostasis, cell membrane stability, scavenging free radicals in vivo. Studies have shown that Tau has induced apoptosis in certain cells. We use the rat pulmonary artery smooth muscle cells as the research object, to study the effect of Tau on it.
Objective: To observe the effect of Tau on the apoptosis of pulmonary artery smooth muscle cells (PASMCs) and to explore whether its mechanism is through mitochondrial pathway and death receptor pathway.
Methods: Wistar male rat pulmonary artery smooth muscle cells; divided into normal group (control group), apoptosis (SD), drug group (Tau high, low dose). Before administration, according to the needs of all groups with low sugar starvation in serum-free medium 24h, the cells in the same growth according to the state Administration of 24h or 48h, the survival rate was measured in 96 hole plate cell MTT; using acridine orange (AO), to observe the morphological changes of the cell nucleus in 6 hole plate under a fluorescence microscope; Determination of cell membrane potential changes in mitochondrial membrane potential kit; Bax were measured by Western-blot method, Bcl-2. Procaspase-9, the expression of Procaspase-3 and Fas protein.
Results: MTT determination of cell survival rate was found, compared with the normal group, the survival rate of treatment group was significantly lower than that in normal group, and apoptosis groups the same trend, evident dose dependently induced pulmonary artery smooth muscle cell apoptosis, 40mmol/L and Tau dosage was the best concentration of acridine orange method. The results of the three groups can be seen apoptosis group and Tau treatment group to cell shrinkage and formation of apoptotic bodies. Determination of mitochondrial membrane potential, apoptosis group and Tau group compared with normal group, the membrane potential decreased, more green fluorescent areas appear in the picture. (?) indicated that the increased expression of Tau, Bax and Fas Vestern-blot protein by the method of reduced expression of Procaspase-9 and Procaspase-3. Bcl-2.
Conclusion: Tau induces apoptosis of PASMCs in rats, and its mechanism may be related to the regulation of mitochondrial pathway and death receptor pathway.

【學位授予單位】：哈爾濱商業(yè)大學
【學位級別】：碩士
【學位授予年份】：2011
【分類號】：R363

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