周细胞在大鼠肺动脉高压时肺血管构型改建中的作用

应用电镜组织化学方法，观察野百合碱诱发大鼠肺动脉高压模型中周细胞的超微结构改变。发现正常对照组大鼠腺泡内无肌型肺动脉内皮下存在着周细胞层，后者与前者的胞浆突起紧密相连。在实验第16天和第24天实验组大鼠（T组）肺动脉压升高持续在4.19～5.79kPa时，可见无肌型肺动脉周细胞向平滑肌样细胞分化现象，细胞外基质也增多，使无肌型肺动脉壁出现肌层和增厚。因此认为，在慢性肺动脉高压时，周细胞向平滑肌样细胞分化和增生是腺泡内肺动脉构型重建的重要原因。     

An image analysis on pathological changes in pulmonary arteries in chronic obstructive pulmonary disease

Summary The changes in small pulmonary arteries of 15 patients with chronic obstructive pulmonary disease (COPD) were investigated by light and electron microscopy, image analysis etc. It was found that the structural changes in the pulmonary arteries of the patients with COPD were characterized by muscularization of non-muscular arterioles, media hypertrophy, longitudinal smooth muscle bundles in the intima and fibrosis in both the media and intima. In the course of time, these lesions resulted in thickening of the arterial wall and narrowing of the lumen. Clinically, the patients developed pulmonary hypertension causing cor pulmonale. Initial data on the structure of arterial wall at different segments were compared statistically. There was very significant difference between the COPD and control groups (P<0.001). By Fisher’s autoclassification (automatic pattern recognition), the rate recognized was correct in more than 90 % in the small arteries of less than 200 μm in diameter. It is suggested that these arteriolar changes are closely related to pulmonary hypertension. The image analysis showed that the ratios of MWA/MVA and MWT/MD were of great value in evaluating the degree of the changes in the arteries of the patients with COPD.     

血小板衍生生长因子-BB对血管内皮细胞、平滑肌细胞及成纤维细胞胶原蛋白合成的影响

目的:观察血小板衍生生长因子-BB对培养的人血管内皮细胞、平滑肌细胞和成纤维细胞胶原蛋白合成的影响.方法:采用培养的人血管内皮细胞、平滑肌细胞和成纤维细胞,应用3H-脯氨酸掺入方法,观察血小板衍生生长因子-BB对3种细胞胶原蛋白合成的影响.结果:血小板衍生生长因子-BB可促进处于静止状态的成纤维细胞和平滑肌细胞胶原蛋白合成,并呈现出明显的浓度依赖关系,成纤维细胞和平滑肌细胞胶原蛋白合成分别在30 ng/ml和40 ng/ml时达到高峰,血小板衍生生长因子-BB对血管内皮细胞胶原蛋白合成总量无明显促进作用.在30 ng/ml的血小板衍生生长因子-BB作用下,成纤维细胞和平滑肌细胞分别在36 h和48 h时胶原蛋白合成达到高峰.结论:血小板衍生生长因子-BB可明显促进培养的人血管平滑肌细胞和成纤维细胞的胶原蛋白的合成,对血管内皮细胞胶原蛋白合成总量无明显促进作用.     

Effects of DDPH on HECCM-induced Proliferation and Immunophenotypes of the Pulmonary Vascular Pericytes

Muscularization of non- muscular arterioles of in-tra- acinar pulmonary arteries is an important mor-phological feature of pulmonary vascular structuralremodeling ( PVSR ) during the development ofchronic hypoxic pulmonary hypertension[1] .In Chi-na,to explore the cell origin of muscularization ofnon- muscular arterioles,the cell culture of pul-monary vascular pericytes was firstestablished at ourlaboratory[2 ] .Resentstudiesin vitro[3] suggested thathypoxic endothelium cell conditioned medi…     

新型katp开放剂埃他卡林对原代培养人肺动脉平滑肌细胞pcna mrna表达的影响

目的:观察新型ATP敏感性钾通道(KATP)开放剂埃他卡林(IPT)对原代培养人肺动脉平滑肌细胞增殖细胞核抗原(PCNA)mRNA表达的影响。方法:分离3～4级人肺小动脉,按照组织贴块法原代培养人肺动脉平滑肌细胞,分成对照组、缺氧组、缺氧+埃他卡林或吡那地尔组及缺氧+埃他卡林或吡那地尔+格列本脲组,应用实时荧光定量逆转录聚合酶链反应(FQ-RT-PCR)技术检测细胞PCNA mRNA表达。结果:缺氧使人肺动脉平滑肌细胞PCNA mRNA表达增加(3.90±0.24)倍,与对照组比较差异有显著性(P<0.01);相同条件下,在缺氧的同时,分别在培养液中加入IPT 10-7、10-6、10-5 mol/L,PCNA mRNA表达分别下降(26.00±2.01)%、(55.33±4.04)%、(79.00±1.00)%,与缺氧组比较差异均有显著性(P均<0.01);加入IPT 10-5mol/L前30 min,预先加入格列本脲(GLI)10-5 mol/L,IPT的抑制作用抵消,与缺氧+IPT 10-5 mol/L组比较差异有显著性(P<0.01)。结论:IPT通过激活细胞KATP通道,浓度依赖性地抑制缺氧时原代培...     

An experimental study of using “tendon bridge” or “tendon tunnel” as a conduit for nerve regeneration

In our experiments, a “tendon bridge” or a “tendon tunnel” was used as a conduit to repair the defect of a peripheral nerve in rabbits. A gap of about 1 cm was made on the deep peroneal nerve in each animal. In group 1, the tendon of anterior tibial muscle was isolated near the severed nerve, and a segment of half cross section of tendon was removed in ladder-shaped reversed fashion. The proximal and distal nerve stump were sutured respectively to the tendon by 11/0 atraumatic sutures under surgical microscope. In group 2, the nerve stumps were sutured to the tendon of anterior tibial muscle without cutting the tendon, but the tendon was dissected free and turned to form a tunnel wrapping the gap in between the nerve stumps. The results showed that a gap of 1 cm of the deep peroneal nerve might be repaired by both methods, but the results of using “tendon tunnel” in group 2 were much better than those of using “tendon bridge” in group 1.     

Expression of Connexin43 in Rat Epithelial Cells and Fibroblasts

To explore the role of connexin43 (Cx43) in gap junctional intercellular communication (GJIC) and propagated sensation along meridians, the expression of Cx43 in the rat epithelial cellsand fibroblasts was studied both in vitro and in vivo. With the in vitro study, the rat epithelial cells and fibroblasts were cultured together, and the localization of Cx43 was detected by immunohistochemistry and indirect immunofluorescent cytochemistry and under confocal microscopy . And the expression of Cx43 on the surface of the cells was examined by flow cytometry. With the in vivo examination, 20 SD rats were randomized into control group (n=10) and electrical acupuncture group (EAgroup, n=10). EA ( 0.5-1.5 V, 4-16 Hz , 30 min) was applied to “Zusanli”acupoint for 30 min at rat‘s hind paw, the localization of Cx43 was immunohistochemically detected. The immunohistochemical staining and indirect immunfluoresce.nt cytochemistry showed that Cx43 was localized on the surface of the cells and in the cytoplasm. The relative expression level of Cx43 on the cellular membrane surfaces of the rat epithelial cells and fibroblasts, as determined by FACS, were 13.91 % and 29.53 % respectively. Our studied suggested that Cx43 might be involved in GJIC and propagated sensation along meridians.     

糖尿病大鼠视网膜血管超微结构改变与核因子-κB的关系

目的:观察不同病程糖尿病大鼠视网膜血管超微结构的改变与核因子-κB(NF-κB)在视网膜中的表达情况,并研究两者间的关系,以探讨NF-κB在糖尿病视网膜病变(DR)发病机制中的作用。方法:SD大鼠48只,随机分为糖尿病组24只,对照组24只。糖尿病模型的建立采用链脲佐菌素(STZ)一次性腹腔注射。应用免疫组化SP三步法分别于1,3,5个月对大鼠视网膜的NF-κB表达情况进行观察,并用透射电镜进行超微结构的观察。结果:大鼠视网膜血管超微结构:对照组,微血管管壁完整,基底膜连续完整,内皮细胞、周细胞结构正常,异染色质分布均匀。糖尿病组,1个月时,微血管形态基本正常;3个月时内皮细胞明显肿胀、变形,周细胞异染色质边集、浓染,细胞器结构不清,基底膜增厚;5个月时,内皮细胞肿胀、变形更加明显,并有不同程度增生,血管腔明显变窄,周细胞内异染色质分布不均、边集、浓染,基底膜显著增厚。免疫组化结果:对照组,NF-κB仅表达于视网膜神经节细胞和内皮细胞胞质中,细胞核内未见其表达。糖尿病组,从1个月开始,可见视网膜各层细胞均有表达,并主要表达于细胞核内,随病程延长,表达进行性增高。结论:在糖尿病视网膜病变早期,视网膜微血管还未出现明显改变时,NF-κB已被激活,并且随着病程的延长,视网膜微血管超微结构明显改变,NF-κB的表达活性也不断增强。因此,NF-κB在DR的早期发生和后期增殖性发展中可能发挥着重要作用,对今后治疗DR也提供了新的依据。     

低氧刺激复合培养的平滑肌细胞表达Janus激酶

目的：研究低氧对复合培养的平滑肌细胞中Janus激酶（JAK）基因表达的作用。方法：鼠肺微血管内皮细胞与肺动脉平滑肌细胞复合培养，采用半定量RT－PCR技术检测常氧组、低氧2、6、12h组的JAK1、JAK2和JAK3基因的表达水平，并对PCR产物进行测序。结果：低氧复合培养2h组肺动脉平滑肌细胞中JAK1、JAK2、JAK3 mRNA表达水平升高，在低氧6h组达最高，与正常组比较均有明显差别。低氧12h组的表达量低于6h组，但高于正常组。RT－PCR产物直接测序证实扩增产物片断与Genebank中相应序列相同。结论：低氧增强复合培养的肺动脉平滑肌细胞中JAK基因的表达，而JAK基因在低氧所致的肺动脉收缩和低氧肺动脉高压的信号转导中可能发挥重要作用。     

微血管周细胞收缩功能的研究进展

周细胞定位在微血管壁外侧,是微血管的重要组成细胞之一,它在微血管的形成及局部血流调节中发挥重要作用。周细胞是类似平滑肌细胞的一类细胞,表达多种收缩蛋白,具有收缩性。周细胞收缩可调节微血管管径及血流,控制局部微血流的灌流量。近年,越来越多的研究表明周细胞的收缩功能与多种微血管疾病的病变过程有关,因此日益受到关注。对其收缩功能的进一步理解,可能为治疗微血管疾病提供新的方法。     

组织块法培养大鼠肠系膜小动脉的平滑肌细胞

利用贴块法成功地大量培养了大鼠肠系膜小动脉的平滑肌细胞。培养的细胞经光镜和免疫组化鉴定为平滑肌细胞。方法简便，经济，为阻力血管的细胞水平的研究提供了实验材料。     

组织工程化血管的种子细胞在体外培养扩增方法的实验性研究

目的为培养组织工程化血管的实验研究提供细胞来源.方法采用酶消化法和组织块法培养人脐静脉血管内皮细胞、人脐动脉血管平滑肌细胞及成纤维细胞,并进行细胞的传代、纯化、鉴定以及形态学观察.结果培养细胞符合血管内皮细胞、血管平滑肌细胞及成纤维细胞的各有形态特征.结论所培养的细胞有望作为体外构建组织工程化血管的应用.     

从超微结构探讨子宫平滑肌瘤的组织学发生

为进一步研究子宫平滑肌瘤的组织发生，本文对４例肌瘤组织，孕５、６、７、８个月胎儿子宫肌组织的腔周细胞及１例肌瘤患者子宫肌组织进行了电镜观察，结果是：①肌瘤患者子宫肌组织主要为典型的平滑肌细胞构成。②肌瘤组织中腔周细胞：腔内层为成熟或不成熟内皮细胞，其外可见无特征分化细胞、成熟和不成熟平滑肌样细胞、肌成纤维样细胞，以及成纤维样细胞。③胎儿肌组织中腔周细胞与肌瘤组织腔周细胞相类似，但主要由成熟和未成熟平滑肌细胞及无特征性分化细胞构成。     

IL-22在支气管哮喘患者血清中的表达及其受体IL-22R1在人气道细胞中的表达

\[摘要\]目的检测IL-22在支气管哮喘患者血清中的表达,检测IL-22R1在人气道上皮细胞、气道平滑肌细胞、肺成纤维细胞中的表达,寻找IL-22作用的靶细胞。方法应用ELISA法检测36例支气管哮喘患者及20例正常对照者血清IL-22及IL-17的表达,同时检测36例患者肺通气功能,根据1秒率(FEV1/FVC)及FEV1占预计值的百分比将支气管哮喘患者分为支气管激发试验阳性组(17例)和支气管舒张试验阳性组(19例)。应用实时定量PCR检测人气道平滑肌细胞、人肺成纤维细胞、人气道上皮细胞IL-22R1 mRNA的表达,用免疫荧光以及蛋白质印迹法检测IL-22R1蛋白在上述3种细胞中的表达。结果支气管哮喘患者血清IL-22及IL-17水平与正常对照组相比差异无统计学意义,但支气管舒张试验阳性组患者血清IL-22和IL-17水平高于支气管激发试验阳性组(P<0.05)。IL-22R1 mRNA及蛋白在上述3种细胞均有表达。结论IL-22可能涉及支气管哮喘的发生发展过程,气道平滑肌细胞、肺成纤维细胞、人气道上皮细胞可能均是IL-22发挥作用的靶细胞。     

慢性缺氧所致肺血管结构改建机制的探讨

利用常压缺氧舱建立大鼠肺缺氧模型,通过ＨＥ、ＶＧ、免疫组化染色及电镜,观察慢性缺氧所致的肺血管结构改建。发现其结构改建主要表现为：①肺动脉外膜胶原纤维增生;②肺外周肌性小动脉中膜增厚;③肺泡内无肌性小动脉肌型化。并对慢性缺氧所致肺血管结构改建的机制进行了探讨。     

糖尿病大鼠脑内微血管细胞外信号调节激酶磷酸化的研究

目的研究链脲佐菌素诱导的Ⅰ型糖尿病大鼠脑膜、脑实质微血管的病理改变及其中细胞外信号调节激酶ERK1/2及其下游作用底物转录因子Elk-1的磷酸化状态,探讨糖尿病血管的发病机制。方法应用链脲佐菌素制备Ⅰ型糖尿病大鼠模型,HE染色观察制模后3d和2、4、8周脑膜、脑实质微血管病变,免疫组织化学观察高血糖状态下脑内微血管中p-ERK1/2及下游作用底物p-ELK1的表达情况。结果 p-ERK1/2、p-ELK1在各时间点正常对照组和糖尿病组大鼠脑膜、脑实质微血管中均有不同程度的表达。p-ERK1/2在糖尿病4、8周组脑膜、脑实质微血管中表达均明显上调;p-ELK1在糖尿病4、8周组脑膜、脑实质微血管内皮细胞中表达均明显的上调(P<0.05),分别与糖尿病3d、2周组及各期正常对照组比较差异有统计学意义(P<0.01)。糖尿病各组与正常对照组各组脑膜微血管平滑肌细胞中p-ELK1的表达差异无统计学意义(P>0.05)。结论在糖尿病4、8周组SD大鼠脑内微血管中存在ERK1/2信号转导通路的异常激活。     

改良组织块法培养SD大鼠阴茎海绵体平滑肌细胞

目的应用改良植块法体外培养大鼠阴茎海绵体平滑肌细胞。方法15只SD雄性大鼠,随机分成3组,每组5只,分别采用组织块法、酶消化法及改良组织块法分离大鼠阴茎海绵体平滑肌细胞,在含双抗及20%胎牛血清DMEM,37℃、5%CO2、95%空气的细胞培养箱静置中培养,相差显微镜观察细胞形态及扩增情况,用α-平滑肌肌动蛋白(a-SM-Actin)及结蛋白(desmin)免疫组织化学染色,鉴定细胞类型。结果α-平滑肌肌动蛋白在阴茎海绵体平滑肌细胞阳性率为96.3%,在成纤维细胞中阳性率为23.8%,结蛋白在阴茎海绵体平滑肌细胞阳性率为74.4%,在成纤维细胞中呈阴性。三组间结蛋白阳性细胞率具有显著差异,改良组织块法组结蛋白阳性细胞率高于组织块法和酶消化法组。结论结蛋白是鉴定海绵体平滑肌细胞的特异性指标,改良组织块法可获纯度更高、结构和功能良好的阴茎海绵体平滑肌细胞。     

Impact of silencing MMP9 gene on the biological behaviors of trophoblasts

This study examined the effect of MMP9 gene on the biological behaviors of trophoblasts and explore the relation between MMP9 gene and the "superficial implantation of placenta". In vitro cultured trophoblasts (TEV-1 cells) were transfected with synthesized double-stranded MMP9 RNA (siRNA) by using lipofectamine2000TM technique and the expressions of MMP9 mRNA and protein and the growth and invasiveness of the TEV-1 cells were determined. Our results showed that siRNA transfection could significantly inhibit the expression of MMP9 gene in the TEV-1 cells and the growth and invasiveness of the TEV-1 cells transfected RNA was significantly reduced (P<0.01). We are led to conclude that silencing of MMP9 gene with siRNA can inhibit the growth and invasiveness of trophoblasts and increasing the expression of MMP9 might help prevent and treat preeclampsia.     

应用骨髓干细胞和自制胶原-壳聚糖构建组织工程心瓣膜

目的探讨应用骨髓基质干细胞(bone mesenchymal stem cells,BMSCs)诱导分化的内皮细胞为种子细胞,以胶原-壳聚糖复合物为支架材料,运用组织工程学的原理和方法体外构建组织工程心脏瓣膜(tissue engineering heart valve,TEHV)的可行性。方法兔主动脉壁体外分离、培养、传代平滑肌细胞、成纤维细胞,将培养的成纤维细胞、平滑肌细胞与兔骨髓基质干细胞诱导分化的内皮细胞按顺序种植在预湿处理的胶原-壳聚糖复合支架上,通过组织学及倒置显微镜、扫描电镜观察内皮细胞、成纤维细胞、平滑肌细胞在支架上生长、增殖和基质分泌情况。结果 HE染色示内皮细胞能够在胶原-壳聚糖复合支架黏附、伸展、增殖,在TEHV表面长成连续的细胞层;扫描电子显微镜示TEHV表面光滑,细胞生长良好,细胞层完整,且瓣膜表面的内皮细胞具有合成和分泌前列环素(PGI2)的功能。结论以兔主动脉壁体外分离、培养、传代的平滑肌细胞、成纤维细胞和BMSCs诱导分化的内皮细胞为种子细胞,依次种植在胶原-壳聚糖复合支架上可以构建TEHV,且内皮细胞能够分泌PGI2。     

成年鼠骨髓间质细胞在体外培养中分化为平滑肌细胞

目的　探讨小鼠骨髓间质细胞在体外经条件培养基诱导定向分化成平滑肌细胞的可行性 ,为研究血管损伤后新生内膜形成机制奠定基础。方法　采用骨髓干细胞培养基和贴壁法从骨髓中分离骨髓间质细胞 ,通过补加人血管平滑肌条件培养基诱导骨髓间质细胞分化成平滑肌细胞 ,采用形态学和免疫荧光染色分析分化后的细胞特异性标志物 ,并测定 1μmol/L血管紧张素Ⅱ对细胞胞液钙离子浓度及细胞收缩功能的影响 ,通过加入氯沙坦抑制血管紧张素Ⅱ受体验证血管紧张素Ⅱ特异性激活钙通道引发的细胞收缩。结果　骨髓间质细胞经诱导在体外可传代生长 ,稳定传代 6代后 ,分化的细胞形态呈梭形平滑肌样 ,融合后形成峰谷状排列。表达平滑肌特异性蛋白标志物α 肌动蛋白、肌钙结合蛋白和平滑肌肌球蛋白。分化后的细胞经血管紧张素Ⅱ刺激产生瞬间胞液钙离子转移并偶联细胞收缩反应 ,细胞 [Ca2 + ]i呈现瞬间增高变化 ,[Ca2 + ]i为 2 131nmol/L± 14 0nmol/L ,经用氯沙坦后 [Ca2 + ]i被阻断 ,为 4 0 2nmol/L± 31nmol/L。结论　骨髓间质细胞经体外诱导可分化为有收缩功能的平滑肌细胞 ,可用于平滑肌细胞分化和血管损伤后新生内膜形成起源等研究。