缺氧时肺动脉内皮细胞与肺动脉平滑肌细胞相互作用对细胞内 …

目的：观察了培养的小牛肺动脉内皮细胞（ＰＡＥＣ）和肺动脉平滑肌细胞（ＰＡＳＭ）于缺氧时对细胞内环核苷酸的影响。结果：ＰＡＥＣ和ＵＰＡＳＭ共培养２４ｈ，ＰＡＥＣ细胞内ｃＡＭＰ含量显著降低（Ｐ〈０．０１），而ＰＡＳＭ细胞内ｃＡＭＰ含量显著增加（Ｐ〈０．０１），二种细胞内ｃＧＭＰ含量显著增加（Ｐ〈０．０１）。缺氧对二种细胞内ｃＧＭＰ含量无显著影响，但能增加ＰＡＳＭ的ｃＧＭＰ含量（Ｐ〈０．０１），降低ＰＡ     

L-精氨酸和牛磺酸联合抗缺氧损伤及防治缺氧性肺动脉高压的研究

目的研究L-精氨酸、牛磺酸联合应用的抗缺氧损伤以及对缺氧性肺动脉高压(HPH)的防治效果。方法雄性W istar大鼠40只,制作缺氧性HPH模型,随机分为平原对照组(C)、单纯缺氧组(H)、缺氧 L-精氨酸组(Arg)、缺氧 牛磺酸组(Tau)、缺氧 牛磺酸 L-精氨酸组(TA),测定各组的肺动脉压(mPAP)、右心室肥大指数(RVH I)、血浆乳酸脱氢酶(LDH)、脂质过氧化产物丙二醛(MDA)、肺匀浆NO含量的变化。结果H组与C组相比,mPAP高出约25 mmHg(P<0.01),RVH I增加1.56倍(P<0.01),血浆LDH活性增高10.1倍(P<0.01),血浆MDA含量增加1.64(P<0.01),而肺匀浆NO含量降低68%(P<0.01)。各治疗组与H组相比,mPAP均显著降低5 mmHg(P<0.05),RVH I均显著降低20%(P<0.01),血浆LDH活性显著降低(P<0.01),其中TA组与Arg组比较有显著差异(P<0.01),血浆MDA含量显著降低(P<0.01),但各治疗组之间无显著性差异。肺匀浆NO含量在Tau组无显著性变化,在Arg和TA组显著回升(P<0.01)。结论L-精氨酸和牛磺酸具有抗缺氧损伤及防治肺动脉高压的效果,两者联用可进一步增加NO产生,减少乳酸脱氢酶的漏出,加强细胞保护作用。     

缺氧对肺动脉内皮细胞和平滑肌细胞5—羟色胺转载体基因表达 …

目的：探讨无氧（０％Ｏ２＋９５％Ｎ２＋５％ＣＯ２）和低氧（２．５￣３％＋９％Ｎ２＋５％ＣＯ２）对新生小牛肺动脉内皮细胞（ＰＡＥＣ）和肺动脉平滑肌细胞（ＰＡＳＭ）５－羟色胺转载体基因表达的影响。方法：应用细胞培养，核到分子杂交技术。结果：无氧组和低氧组（１．５ｈ、３ｈ、６ｈ）ＰＡＥＣ５－羟色胺转载体ｍＲＮＡ表达显著同于常氧组（Ｐ〈０．０５），而无氧和低氧１２ｈ至４８ｈ对ＰＡＥＣ５－羟色胺转载体ｍＲＮ     

西拉普利和牛磺酸对慢性缺氧肺组织细胞凋亡的影响

目的：初步观察西拉普利和牛磺酸对慢性缺氧肺组织细胞凋亡的影响。方法：采用原位末端民法。结果：缺氧模型及普利组细胞凋亡明显高于其他各组,而牛磺酸组、自然恢复组册与正常对照无差异。结论：西拉普利可能诱发细胞凋亡的作用,细胞凋亡的发生与否可能与肺动脉高压的形成与发展有关。     

慢性缺氧对大鼠肺动脉平滑肌细胞胞内钙的影响及其机制研究

目的:研究慢性缺氧对大鼠肺动脉平滑肌细胞(PASMCs)胞内钙浓度(［Ca²⁺］_i)的影响及L-型钙通道和胞内钙库的作用,为缺氧性肺动脉高压(HPH)发病机制的进一步研究提供理论依据。 方法:复制大鼠缺氧性肺动脉高压动物模型，利用Fura－2/AM钙离子成像方法测定PASMCs在不同钙离子浓度细胞外液及L-型钙通道阻滞剂nifedipine和IP3R钙通道抑制剂肝素干预前后 ［Ca²⁺］_i变化。 结果:(1)缺氧＋含钙外液组PASMCs ［Ca²⁺］_i 显著高于对照＋含钙外液组（P<0.05）。缺氧＋含钙外液组PASMCs ［Ca²⁺］_i显著高于缺氧＋无钙外液组（P<0.05）。(2)缺氧nifedipine组PASMCs［Ca²⁺］_i在加药前后无显著差异（P>0.05）。（3）缺氧未干预组与缺氧肝素组PASMCs ［Ca²⁺］_i无明显差异（P>0.05）。 结论:慢性缺氧可使PASMCs的［Ca²⁺］_i增加。慢性缺氧引起［Ca²⁺］_i增加可能与细胞外钙内流有关，L-型钙通道和IP3R钙通道在调节［Ca²⁺］_i的过程中可能不独立发挥作用。     

缺氧对肺动脉平滑肌细胞增殖的影响及其传递途径

本文观察了低氧对离体培养成年牛肺动脉平滑肌细胞（ＰＡＳＭＣ）＾３Ｈ－ＴｄＲ参入的影响，并用异搏定、Ｈ７、ＰＭＡ和ＥＧＦ进行干预实验，对ＭＳＣ增殖中信息传递途径作了初探。结果显示：（１）缺氧２４ｈ，ＰＡＳＭＣ＾３Ｈ－ＴｄＲ参入显著增高。（２）异搏定及Ｈ７显著降低低氧Ｈ－ＳＭＣ＾３Ｈ－ＴｄＲ参入，而ＰＭＡ及ＥＧＦ显著增加ＨＳＭＣ＾３ＨＴｄＲ参入。（３）Ｈ－ＳＭＣ对ＰＭＡ和ＥＧＦ反应强于常氧Ｎ－ＳＭＣ；     

急性缺氧肺动脉平滑肌细胞线粒体活性氧变化的观察

 目的:观察急性缺氧时大鼠肺动脉平滑肌细胞（PASMCs）线粒体活性氧（ROS）的变化。方法:分离并培养大鼠PASMCs,常氧（35 ℃、5% CO2、21% O2、74% N2）和急性缺氧(35 ℃、5% CO2、1% O2、94% N2)条件下,利用分子探针chloromethyl dichlorodihydrofluorescein diacetate (CM-H2DCF/DA) 和RedoxSensor Red CC-1通过激光共聚焦显微镜来检测细胞内ROS的生成量;直接分离线粒体,用线粒体电子传递链（ETC）复合物抑制剂通过荧光分光光度计检测线粒体ROS及其生成的具体位点。结果:急性缺氧细胞内ROS生成明显增加,其中缺氧组H2O2较常氧组增加3.35倍 (P<0.01),而H2O2 及O-·2较常氧组增加1.61倍（P<0.01）。与缺氧组比,用线粒体ETC复合物I抑制剂MPP、复合物II抑制剂NPA和TTFA及复合物III前泛半醌位点抑制剂myxothiazol都能显著降低缺氧时PASMCs胞内ROS的生成量（分别降低60%、73%、75%和61%,P<0.01）;而复合物III后泛半醌位点抑制剂antimycin A及复合物IV 抑制剂NaN3对ROS的生成无明显影响（升高13%和9.1%,P>0.05）。 直接检测线粒体ROS与测定细胞内ROS结果一致。结论: 急性缺氧PASMCs线粒体ROS（主要是H2O2）的生成量明显增加;其生成位点主要是线粒体ETC复合物Ⅰ、Ⅱ及III前泛半醌位点,而与复合物III后泛半醌位点和IV位点无明显关系。     

缺氧对肺动脉平滑肌细胞钾通道Kv1.2、Kv1.6基因表达的影响

目的:研究不同缺氧时间对大鼠肺动脉平滑肌细胞(PASMC)不同亚型钾通道(Kv)mRNA和蛋白质表达的影响。方法:采用半定量RT-PCR和Western-blot方法对常氧和不同缺氧时间后PASMC上Kv1.2、Kv1.6mRNA和蛋白质的表达进行测定。结果:(1)PASMC在常氧和缺氧时均有Kv1.2、Kv1.6mRNA和蛋白质表达;(2)缺氧18h使Kv1.2mRNA和蛋白质表达增强,缺氧48h则使其表达减弱且低于常氧时的表达;(3)缺氧18h、48h对Kv1.6的mRNA和蛋白质表达无影响。结论:Kv1.6不是氧敏感的钾通道;作为氧感受器的Kv1.2,其mRNA和蛋白质表达随缺氧时间而改变。     

低氧致肺动脉平滑肌细胞增殖及其作用机制初探

目的和方法：实验采用ＰＤＧＦ生物活性检测法，流式细胞术，「＾３Ｈ」－ＴｄＲ掺入以及细胞免疫线化等方法探讨了低氧状态下，血小板源生长因子（ＰＤＧＦ）及其受体与肺动产滑肌细胞（ＰＡＳＭＣ）增殖间的相互关系。结果：低氧可以增强ＰＤＧＦ的活性，上调ＰＧＦ受体促进ＰＡＳＭＣ增殖以ＰＤＧＦ作为刺激嘲一步促进低氧的促增殖作用。结论：低氧状态下，ＰＡＳＭＣ的ＰＤＧＦ及其受体进一步活化，从而增强了ＰＤＧＦ自分泌、旁     

慢性低氧性肺动脉高压大鼠肺组织P53表达增高

目的观察P53在低氧性肺动脉高压大鼠肺组织中的表达。方法采用减压低氧方法复制大鼠慢性低氧性肺动脉高压模型;取肺组织,常规SABC免疫组化法染色和Hoechst染色,观察P53表达的变化及细胞凋亡变化。结果P53在慢性肺动脉高压大鼠肺小动脉壁中有少量表达,其中低氧3周组>低氧2周组>正常组。Hoechst染色显示肺组织凋亡细胞增加,其变化趋势与P53的表达变化趋势相同。结论在慢性低氧性肺动脉高压形成过程中,肺小动脉壁发生了增殖和凋亡,程度随低氧时间的延长而增加,同时P53表达增多,提示P53参与了慢性低氧性肺动脉高压肺小血管重建的调控。     

牛磺酸对大鼠慢性缺氧性肺动脉高压的预防作用

目的：进一步研究牛磺酸对慢性缺氧性肺动脉高压的防治作用。方法：复制大鼠间断缺氧４周模型,采用透射电镜、放射免疫和生化技术研究牛磺酸对肺动脉高压的治疗效果。结果：缺氧４周大鼠肺动脉压力升高,右心肥大,血中ＥＴ－１、ＡＣＥ水平升高,ＮＯ－２／ＮＯ－３和ＳＯＤ水平降低;ＭＰＡＰ与ＥＴ－１呈正相关,与ＮＯ－２／ＮＯ－３呈负相关;牛磺酸可抑制缺氧大鼠ＥＴ－１和ＡＣＥ分泌,增加ＮＯ和ＳＯＤ水平,降低肺动脉压力;透射电镜观察表明牛磺酸能对抗缺氧大鼠的心肺组织损伤。结论：牛磺酸通过减轻缺氧所致细胞损伤,调节血管舒缩物质的平衡,对缺氧性肺高压有一定防治作用。     

肾上腺髓质素缓解大鼠低氧性肺动脉高压

目的探讨肾上腺髓质素(ADM)对大鼠低氧性肺动脉高压的防治作用及机制。方法雄性Wistar大鼠18只,分为对照组、低氧组和低氧 ADM组,每组6只。持续皮下注射ADM1-50后,测定平均肺动脉压(mPAP)、右心室肥大指数RV/(LV S)、肺小动脉病理及形态计量学和体循环平均压(mSBP),放免法测定肺动脉血浆ADM水平,原位杂交测定肺动脉ADMR mRNA的表达。结果①低氧组大鼠mPAP,RV/(LV S),管壁厚度与血管外径比值(MT%)及管壁面积与血管面积比值(MA%)均显著升高(P<0.01);ADM组显著缓解以上变化(P<0.01)。②低氧组与低氧 ADM组肺动脉血浆ADM浓度均高于对照组,且低氧 ADM组较低氧组ADM浓度低(P<0.05)。③低氧组与低氧 ADM组的ADMR mRNA表达较对照组增强(P<0.01)。结论持续皮下注射ADM对慢性低氧所致的肺动脉高压及肺血管重塑有预防和部分逆转作用。     

Cytoskeletal features of immature pulmonary vascular smooth muscle cells: the influence of pulmonary hypertension on normal development

Using an immunohistochemical technique, the development of the cytoskeletal proteins desmin, vimentin, and actin (using alpha isotype and non-isotype specific antibodies) was assessed using a semi-quantitative grading system in the pulmonary vascular smooth muscle of nine normal pigs and 19 normal humans at different ages, and in 13 children with pulmonary hypertensive congenital heart disease. In the normal of both species, immunostaining for vimentin decreased after birth and then increased gradually while immunostaining for desmin and alpha actin increased steadily with age. In pulmonary hypertension, immunostaining for alpha actin and vimentin showed an accelerated increase at between 2 and 8 months. Also, the media showed regional differences in immunostaining which preceded the development of intimal proliferation. The inner media showed less immunoreactivity for all cytoskeletal proteins studied than did the outer media. Within areas of intimal proliferation many cells were immunonegative. These results suggest that the cytoskeletal features of medial smooth muscle cells are remodelled in the normal infant; that this process is altered from at least 2 months in the pulmonary hypertensive infant; and that the smooth muscle cells immediately beneath the internal elastic lamina are remodelled before migrating to form intimal proliferation. Changes in cytoskeletal composition can be related to the previously described postnatal maturation of pulmonary vascular smooth muscle cells.     

Ca2+ and ion channels in hypoxia-mediated pulmonary hypertension

Alveolar hypoxia, a consequence of many lung diseases, can have adverse effects on the pulmonary vasculature. The changes that occur in the pulmonary circulation with exposure to chronic hypoxia include reductions in the diameter of the pulmonary arteries due to structural remodeling of the vasculature. Although the structural and functional changes that occur in the development of pulmonary hypertension have been well investigated, less is known about the cellular and molecular mechanisms of this process. This review will discuss the role of several potassium and calcium channels in hypoxic pulmonary vasoconstriction, both in elevating calcium influx into pulmonary artery smooth muscle cells (PASMCs). In addition to other signal transduction pathways, Ca²⁺ signaling in PASMCs plays an important role in the development and progression of pulmonary hypertension due to its central roles in vasoconstriction and vascular remodeling. This review will focus on the effect of chronic hypoxia on ion channels and the potential pathogenic role of Ca²⁺ signaling and regulation in the progression of pulmonary hypertension.     

Remodelling of the pulmonary arteries during recovery from pulmonary hypertension induced by neonatal hypoxia

Little is understood of the mechanisms involved in reducing pulmonary arterial wall thickness on recovery from pulmonary hypertension and the present study sought to clarify the events that occur. Piglets were exposed to hypobaric hypoxia for 3 days, either from birth or from 3 days of age, and others were exposed for 11 days starting at 3 days. All recovered in room air for up to 6 days. Using light and electron microscopy, the pulmonary artery wall thickness, the relative contribution of smooth muscle and matrix, smooth muscle cell replication, and apoptosis were assessed after hypoxic exposure and during recovery from hypoxic exposure. In elastic arteries, after 6 days' recovery in room air, a reduction in wall thickness to normal was associated with a similar reduction in proportional area of smooth muscle cells and matrix (p < 0.05), increased apoptosis (p < 0.05), and an abnormally low replication rate (p < 0.05). In peripheral muscular arteries, an increase in external diameter, and wall thinning on recovery, was achieved by smooth muscle cell remodelling and a reduction in cell replication (p < 0.05). Apoptosis did not contribute. Thus, different mechanisms are involved in recovery from hypoxia-induced pulmonary hypertension in elastic and muscular pulmonary arteries. Recovery is slower in animals exposed from birth rather than from 3 days of age.     

低氧抑制猪肺动脉平滑肌细胞MMP-2和MMP-9的表达

目的探讨低氧对猪肺动脉平滑肌细胞（PASMC）分泌基质金属蛋白酶（MMPs）的影响。方法采用酶谱法测定PASMC培养基中MMP-2和MMP-9的酶活性，免疫印迹法检测培养基中MMP-2和MMP-9的蛋白表达，免疫组化法测定细胞原位MMP-2和MMP-9的蛋白表达，RT-PCR法检测mRNA的表达。结果低氧后PASMC分泌的MMP-2酶活性、细胞内外蛋白表达量、mRNA表达量均下降；MMP-9酶活性、细胞外蛋白表达量下降，而细胞内蛋白表达无变化。结论低氧可抑制PASMC分泌MMP-2和MMP-9的酶活性，其机制可能是低氧影响PASMC中MMP-2基因的转录、影响MMP-9蛋白表达后的分泌与活化，导致MMP-2和MMP-9酶活性的改变。