硫化氢对肝纤维化大鼠肝脏Ⅰ、Ⅲ型胶原表达的影响

目的:探讨硫化氢(hydrogen sulfide,H2S)对肝纤维化大鼠肝脏Ⅰ型胶原(COL-Ⅰ)和Ⅲ型胶原(COL-Ⅲ)的影响.方法:选择硫氢化钠(sodium hydrosulfide,NaHS)作为H2S的供体,将32只♀SD大鼠分为3组:正常组(N组)8只、肝纤维化组(hepatic fibrosis,HF组)13只、NaHS干预组(S组)11只,采用四氯化碳复合因素法复制肝纤维化模型,S组自造模第6周始给予NaHS56μmol/(kg·d)腹腔注射12d,N组和HF组给予同等剂量的生理盐水腹腔注射.干预结束后,宰杀大鼠留取肝脏行肝组织病理切片HE染色评价肝纤维化分期,行Masson染色观察胶原纤维沉积情况,应用RT-PCR法检测肝脏中COL-Ⅰ、COL-ⅢmRNA表达,采用SP免疫组织化学法检测肝脏COL-Ⅰ、COL-Ⅲ表达.结果:HF组与N组相比,COL-Ⅰ、COL-Ⅲ及其mRNA表达升高(均P=0.000),与肝纤维化分期结果一致(P=0.000);S组与HF组相比,COL-Ⅰ和COL-Ⅲ表达降低(均P=0.000);COL-ⅠmRNA表达降低(P=0.009),同时COL-ⅢmRNA表达亦降低(P=0.003),肝纤维化分期下降(P=0.047).结论:H2S具有降低肝脏Ⅰ、Ⅲ型胶原表达的作用,能够延缓肝纤维化的发生发展.     

H_2S对低氧性肺动脉高压大鼠肺组织BaxmRNA、Bcl-2mRNA表达的影响

目的研究H2S对大鼠低氧性肺动脉高压及凋亡相关基因BaxmRNA、Bcl-2mRNA表达的影响,探讨其在低氧性肺动脉高压发病中的作用。方法将30只雄性Wistar大鼠随机分为三组:常氧对照组、低氧组及低氧+Na HS组(腹腔注射Na HS并进行低氧处理)。常压间歇低氧法建立大鼠低氧性肺动脉高压模型,右心导管法检测平均肺动脉压(m PAP);分别称量大鼠右心室(RV)和左心室加室间隔(LV+S)的质量,计算出右心室肥厚指数RV/(LV+S);HE及弹力纤维+VG染色光镜下观察肺血管形态及肺小动脉平滑肌结构,弹力纤维、肌纤维及胶原纤维增生情况;RT-PCR方法检测BaxmRNA、Bcl-2mRNA在肺组织中的表达。结果 1低氧各组较对照组体重明显减轻(P0.05),但低氧+Na HS组明显高于低氧组(P0.05),造模前各组大鼠体重无差别(P0.05)。2低氧各组较对照组m PAP明显升高(P0.05)、RV/(LV+S)明显增加(P0.05)。低氧+Na HS组较低氧组m PAP明显降低(P0.05),RV/(LV+S)明显减少(P0.05)。3光镜下观察,对照组肺动脉管壁较薄,薄厚均匀,管腔较大。内弹力膜呈平缓波浪状,管壁层次结构清晰;低氧组肺小动脉平滑肌增生,胶原纤维增多,管壁增厚,管腔狭窄。内弹力膜增厚,外膜胶原纤维增生、沉积,外弹力膜也增厚,内外弹力板间距离均增宽;中膜平滑肌细胞也增生明显,管壁增厚,管腔明显变小,管周水肿。低氧+Na HS组较低氧组上述表现有所缓减。4低氧各组较对照组肺组织BaxmRNA表达均显著降低,Bcl-2 mRNA表达均显著增加(P0.05)。低氧+Na HS组较低氧组肺组织BaxmRNA表达显著升高,Bcl-2mRNA表达明显降低(P0.05),低氧+Na HS组较对照组大鼠肺组织BaxmRNA、Bcl-2 mRNA表达无统计学差异(P0.05).结论 H2S可以调节低氧性肺动脉高压细胞大鼠肺组织凋亡相关基因BaxmRNA、Bcl-2mRNA的表达,缓解低氧性肺血管重建,对低氧性肺动脉高压的发病起到保护作用。     

肥大细胞对平滑肌源性泡沫细胞Ⅰ型胶原和Ⅲ型胶原表达的影响

目的　检测大鼠腹腔肥大细胞(mastcell,MC)对平滑肌源性泡沫细胞Ⅰ型胶原和Ⅲ型胶原表达的影响。方法　SD大鼠体重约2 0 0g ,雌雄不限,供采集MC和原代培养平滑肌细胞(smoothmusclecell,SMC)。采用梯度密度离心分离MC ,提取MC上清液(含MC释放的各种炎症介质)。用MC上清液和氧化型低密度脂蛋白(oxLDL)处理SMC ,实验分四组:对照组SMC在5mL含10 %小牛血清的DMEM中培养4 8h ;oxLDL组SMC在5mL含10 %小牛血清的DMEM中(含oxLDL ,终浓度为10 0mg L)培养4 8h ;MC上清液组SMC在5mL含10 %小牛血清的MC上清液(由含1×10 6 个MC的细胞悬液提取)中培养4 8h ;MC上清液+ox LDL组SMC在5mL含10 %小牛血清的MC上清液(由含1×10 6 个MC的细胞悬液提取)中(含oxLDL ,终浓度为10 0mg L)培养4 8h。采用油红O染色检测SMC泡沫化。RT PCR检测SMCⅠ型胶原和Ⅲ型胶原mRNA的表达,PCR引物序列Ⅰ型胶原为5′GACACTGAACCCTTTGTAATG 3′和5′GTGAAACTCCCGTCTGCT 3′,扩增片段长度为399bp ;Ⅲ型胶原引物序列为5′AGCGGAGAATACTGGGTT 3′和5′TGTAATGTTCTGGGAGGC 3′,扩增片段长度为2 88bp ;内参照采用βactin ,引物序列为5′GTGGGGCGCCCCAGGCACCA 3′和5′CTCCTTAATGTCACGCACGATTTC 3′,扩增片段长度为5 4 8bp。免疫细胞化学染色检测SMCⅠ型胶原和Ⅲ型胶原蛋白表达。结果　油红O染色显示MC上清液可加速SMC泡沫化。RT PCR结果发现,与对照组SMC相比,用oxLDL或MC上清液分别单独处理SMC 4 8h后,Ⅰ型胶原和Ⅲ型胶原的mRNA表达均降低,用oxLDL和MC上清液同时处理SMC 4 8h后,Ⅰ型胶原和Ⅲ型胶原的mRNA表达降低更明显。免疫细胞化学染色显示,对照组SMC细胞浆内有大量棕色颗粒,颗粒粗大而染色深,说明对照组Ⅰ型胶原和Ⅲ型胶原的蛋白表达为强阳性;用oxLDL或MC上清液分别单独处理SMC 8h后,细胞浆内棕色颗粒少而染色浅,说明Ⅰ型胶原和Ⅲ型胶原的蛋白表达均明显降低;用oxLDL和MC上清液同时处理SMC 4 8h后,Ⅰ型胶原和Ⅲ型胶原的蛋白表达降低更明显。结论　MC在体外抑制SMCⅠ型胶原和Ⅲ型胶原表达,并且能协同oxLDL对Ⅰ型胶原和Ⅲ型胶原表达的抑制作用。因此MC释放的炎症介质也许参与了动脉粥样斑块纤维帽的重构。     

蛋白激酶C对慢性低氧大鼠肺动脉胶原表达的调控及灯盏花素的干预作用

目的　探讨蛋白激酶C(PKC)对慢性低氧大鼠肺动脉胶原表达的调控作用及灯盏花素的影响。方法　将二级Sprague Dawley(SD)大鼠分为 :对照组 (A) ,低氧组 (B) ,低氧 +灯盏花素组 (C) ,低氧时间为 4周。采用透射电镜、放射活性测定法、免疫组化、原位杂交等方法综合进行评价。结果(1)B组肺动脉平均压 (mPAP)、右心室重量比 (RV/LV +S)显著高于A组 (P <0 0 1) ,C组mPAP、RV/LV +S显著低于B组 (P <0 0 1) ;(2 )电镜显示B组肺动脉胶原纤维较A组明显为多 ,C组较B组明显为少 ;(3)B组肺组织PKC总活性 (PKCt)、胞膜PKC活性 (PKCm)、胞浆PKC活性 (PKCc)及PKCm占PKCt的百分比显著高于A组 (P <0 0 1) ,C组PKCt、PKCm、PKCc及PKCm占PKCt的百分比显著低于B组 (P <0 0 5 ) ;(4)免疫组化显示B组肺细小动脉 (直径 10 0～ 2 0 0 μm)PKC含量 (平均吸光度A值 )显著高于A组 (P <0 0 1) ,C组较B组显著为低 (P <0 0 1) ;(5 )免疫组化和原位杂交显示B组肺细小动脉 (直径约 10 0～ 2 0 0 μm)Ⅰ型胶原及Ⅰ型前胶原mRNA平均A值较A组明显为高 (P <0 0 1) ,C组较B组为低 (P <0 0 1) ,Ⅲ型胶原及Ⅲ型前胶原mRNA平均A值各组间差异无显著性 (P >0 0 5 ) ;(6 )肺组织PKC活性和肺动脉管壁PKC的表达与肺动脉管壁Ⅰ型胶原mRNA和蛋白     

SB203580对肝纤维化大鼠肝脏Ⅰ、Ⅲ型胶原蛋白表达的影响

目的:探讨SB203580对肝纤维化大鼠肝脏Ⅰ、Ⅲ型胶原蛋白表达的影响.方法:将32只♀SD大鼠分为4组:正常组(N组)8只、肝纤维化组(HF组)8只、DMSO溶剂干预组(D组)8只、P38MAPK抑制剂SB203580干预组(SB组)8只.采用四氯化碳复合因素法复制肝纤维化模型,肝纤维化模型制作结束后,D组给予2‰DMSO溶液3 mL/(kg?d),SB组给予SB203580溶液10 mg/(kg?d),N组、HF组给予同等剂量0.9%生理盐水3 mL/(kg?d)腹腔注射,连续注射4 d.干预结束后,宰杀大鼠留取肝脏,行肝组织病理切片HE染色评价肝纤维化分期,行Masson染色观察胶原纤维沉积情况,采用SP免疫组织化学法检测肝脏Ⅰ、Ⅲ型胶原蛋白表达,应用逆转录PCR法检测肝脏中Ⅰ、Ⅲ型胶原mRNA表达.结果:正常对照组(N组)、肝纤维化组(H F组)、S B203580溶剂组(D M S O组)和P38MAPK通路特异性抑制剂组(SB203580组)的肝纤维化分期平均秩分别为4.50、22.50、24.00和15.00;SSS评分分别为2.750±0.707、15.875±0.835、16.000±0.926和11.625±0.9 1 6;Ⅰ型胶原显色指数分别为1.5 7 5±0.249、7.650±0.621、7.725±0.501和4.625±0.495;Ⅲ型胶原显色指数分别为2.375±0.518、4.025±0.446、4.075±0.544和3.375±0.167;Ⅰ型胶原mRNA表达分别为0.020±0.003、0.012±0.002、0.009±0.002和0.016±0.005;Ⅲ型胶原mRNA表达分别为0.412±0.772、0.773±0.137、0.799±0.116和0.572±0.862.HF组与N组相比,Ⅰ、Ⅲ型胶原及其mRNA表达升高(均P<0.001),与肝纤维化分期结果一致(P<0.001);D组与HF组无明显差异(均P>0.05);SB组与HF组相比,Ⅰ、Ⅲ型胶原表达降低(Ⅰ型胶原显色指数P<0.001,Ⅲ型胶原显色指数P=0.041);Ⅰ型胶原mRNA表达降低(P=0.005),同时Ⅲ型胶原mRNA表达亦降低(P=0.005),肝纤维化分期下降(P=0.015).结论:P38MAPK抑制剂SB203580阻断P38MAPK通路具有降低肝脏Ⅰ、Ⅲ型胶原表达的作用,能够延缓肝纤维化的发生发展.     

硫化氢对低氧性肺动脉高压大鼠细胞因子的调节作用

目的探讨硫化氢对低氧性肺动脉高压大鼠细胞因子的调节作用。方法将Wistar大鼠22只随机分为3组。对照组(8只),低氧组(7只),低氧+硫氢化钠(NaHS)组(7只),测定3组大鼠肺动脉平均压(mPAP),观测肺血管结构变化,并用免疫组织化学方法研究α-平滑肌肌动蛋白(-αSM-actin)、弹力蛋白、转化生长因子β(TGF-β)和结缔组织生长因子(CTGF)在肺动脉平滑肌细胞的表达含量。结果低氧组的mPAP明显高于对照组和低氧+NaHS组;低氧组肌型动脉、部分肌型动脉百分比明显高于对照组和低氧+NaHS组,非肌型动脉百分比明显低于对照组和低氧+NaHS组;低氧组肺中、小型肺动脉平滑肌细胞-αSM-actin、弹力蛋白和TGF-β表达含量分别明显高于对照组和低氧+NaHS组;对照组、低氧+NaHS组以及低氧组肺中、小型肺动脉平滑肌细胞CTGF表达含量依次增高,但3组之间无显著差异。结论硫化氢可能通过调节低氧性肺动脉高压大鼠肺小血管肌性动脉TGF-β的表达,进而抑制细胞外基质成分之一的弹力蛋白合成,参与低氧性肺动脉高压的形成。     

L-精氨酸对低氧性肺动脉高压大鼠肺动脉平滑肌细胞凋亡的影响

目的 探讨L-精氨酸(L-Arg)对低氧性肺动脉高压大鼠不同节段肺动脉平滑肌细胞凋亡的影响。方法 将Wistar大鼠(n=19)随机分为对照组(n=7)、低氧组(n=6)及低氧 L-Arg组(n=6)。经右心导管法测定各组大鼠肺动脉压力和右室(RV)/左室 室间隔(LV S)比值,以分光光度法间接测定血浆一氧化氮(NO)含量,通过TUNEL法检测各组大鼠不同节段的肺动脉平滑肌细胞凋亡数目,并计算肺动脉平滑肌细胞凋亡数目与肺动脉平滑肌细胞数目比值。结果 低氧组大鼠肺动脉平均压(PAMP)显著高于对照组[(2.71±0.29)kPa vs(2.05±0.14)kPa,P<0.01],低氧 L-Arg组大鼠的PAMP显著低于低氧组[(2.23±0.18)kPa vs(2.71±0.29)kPa,P<0.05];低氧组大鼠RV/(LV S)比值显著高于对照组[(0.42±0.03)kPa vs(0.30±0.05)kPa,P<0.01],低氧 L-Arg组大鼠RV/(LV S)比值显著低于低氧组[(0.36±0.02)kPa vs(0.42±0.03)kPa,P<0.01];低氧组大鼠血浆NO含量明显低于对照组[(3.54±0.47)μmol/L vs(4.79±0.17)μmol/L,P<0.05],低氧 L-Arg组大鼠血浆NO含量显著高于低氧组[(5.21±0.26)μmol/L vs(3.54±0.47)μmol/L,P<0.01];低氧组大鼠与终末细支气管伴行的肺动脉和与呼吸细支气管伴行的肺动脉平滑肌细胞凋亡数目与平滑肌细胞数目比值明显低于对照组[(0.051±0.016     

虫草多糖脂质体对实验性大鼠肝纤维化Ⅰ、Ⅲ型前胶原基因表达的影响

目的:探讨虫草多糖脂质体(CPL)对CC14大鼠肝纤维化的预防作用及可能机制。方法:采用虫草多糖脂质体(CPL)预防CC14大鼠肝纤维化,运用免疫组和Northern杂交化检测肝组织Ⅰ、Ⅲ型前胶原蛋白及Ⅰ、Ⅲ型前胶原基因的表达。结果:CPL预防组肝脏中Ⅰ、Ⅲ型胶原蛋白较模型对照组明显减少(P值分别<0.01);CPL预防组大鼠肝脏组织Ⅰ、Ⅲ型前胶原mRNA表达量明显下降,与模型组大鼠比较差异有显著性(P值分别<0.05)。结论:虫草多糖脂质体可以预防大鼠肝纤维化,其抗纤维化作用是通过在转录水平抑制肝脏胶原合成而实现的。     

扶正化瘀方药对NIH／3T3成纤维细胞增殖及胶原表达的影响

目的：观察扶正化瘀方对成纤维细胞增殖、胶原生成及胶原mRNA表达的影响，探讨该方抗肝纤维化的作用机制。方法：经口给予大鼠扶正化瘀方药(丹参、桃仁及虫草菌丝等组成)后，制备药物血清，观察药物对NIH／3T3成纤维细胞增殖、胶原生成及Ⅰ型胶原mRNA表达的影响。细胞增殖采用^3H—TdR掺入试验，胶原酶法检测细胞内外的胶原生成率，免疫荧光细胞化学观察细胞Ⅰ、Ⅱ、Ⅳ型胶原的表达，Ⅰ型胶原mRNA表达采用斑点杂交法。结果：(1)10%药物血清对细胞增殖的抑制率达46．87%(P<0．05)；(2)药物血清能显著抑制细胞内(P<0．05)外(P<0．01)胶原生成率及细胞胶原的表达；(3)药物能显著降低细胞Ⅰ型胶原mRNA的表达量（P<0．01)。结论：抑制成纤维细胞增殖、胶原mRNA的表达及胶原的生成量是扶正化瘀方抗肝纤维化的主要作用机制之一。     

灯盏花素对低氧大鼠肺动脉压和肺小动脉Rho激酶的影响

目的 观察灯盏花素对低氧大鼠肺动脉压、肺小动脉Rho激酶ROCK Ⅰ和ROCK Ⅱ及Rho激酶mRNA的影响,探讨灯盏花素预防低氧性肺动脉高压的作用和机制.方法 将18只健康雄性SD大鼠分为健康组、低氧组和灯盏花素预防组.以常压低氧法复制肺动脉高压模型,以微导管法测定平均肺动脉压(mPAP).分离大鼠心脏,测量右心室(RV)及左心室加室间隔(LV+S)的重量,以RV/(LV+S)代表右心肥厚指数.应用图像分析技术测定肺小动脉管壁厚度占外径的百分比和管壁面积占总面积的百分比,反映肺血管重塑情况.应用免疫组化法测定肺小动脉Rho激酶蛋白的表达,原位杂交法测定肺小动脉Rho激酶mRNA的表达.结果 低氧组大鼠mPAP为(27.3±5.0)mm Hg(1 mm Hg=0.133 kPa),明显高于健康组的(16.0±0.6)mm Hg(t=6.74,P<0.05),灯盏花素预防组mPAP为(19.83±1.47)mm Hg,明显低于低氧组(t=4.28,P<0.05);低氧组RV/(LV+S)及肺小动脉厚度指数明显高于健康组(t=3.43,P<0.05),灯盏花素预防组低于低氧组(t=2.39,P<0.05);低氧组ROCK Ⅰ和ROCK Ⅱ免疫组织化学阳性染色(1.29±0.08和1.63±0.24)明显高于健康组(1.17±0.09和1.30±0.16),灯盏花素预防组(1.18±0.10和1.30±0.12)明显低于低氧组(t值分别为3.96,5.85,3.90,5.82,均P<0.05);低氧组ROCK Ⅰ mRNA和ROCK Ⅱ mRNA原位杂交阳性染色(分别为1.37±0.13和1.59±0.31)明显高于健康组(1.22±0.09和1.2±10.15),灯盏花素预防组(1.23±0.13和1.22±0.06)明显低于低氧组(t值分别为4.00,6.02,3.94,5.83,均P<0.05).结论 灯盏花素具有明显预防低氧性肺动脉高压和降低Rho激酶及Rho激酶mRNA的作用.     

Sodium hydrosulfide alleviates pulmonary artery collagen remodeling in rats with high pulmonary blood flow

This study aimed to explore the effect of sodium hydrosulfide (NaHS) on pulmonary artery collagen remodeling in rats with high pulmonary blood flow. Thirty-two Sprague-Dawley rats were randomly divided into a sham group, shunt group, sham + NaHS (an H₂S donor) group, and shunt + NaHS group. After 11 weeks of shunting, mean pulmonary artery pressure (MPAP), relative median area (RMA) of pulmonary arteries, H₂S concentration in lung tissues, plasma endothelin-1 (ET-1) levels, and ET-1 mRNA in lung tissues were investigated. Collagen I and collagen III were evaluated by immunohistochemistry. Hydroxyproline assay and Sirius-red staining were performed. Matrix metalloproteinase-13 (MMP-13), tissue inhibitor of metalloproteinase-1 (TIMP-1), and connective tissue growth factor (CTGF) were evaluated by immunohistochemistry. After 11 weeks of shunting, rats showed a significant pulmonary hypertension and pulmonary artery collagen remodeling in association with a decrease in lung tissue H₂S content. After NaHS treatment for 11 weeks, lung tissue H₂S content was increased, whereas MPAP was attenuated and RMA was reduced. Meanwhile, pulmonary artery collagen I and collagen III protein expressions of intra-acinar pulmonary arteries were inhibited, but MMP-13/TIMP-1 ratio was augmented with a decreased plasma ET-1 content and lung tissue ET-1mRNA and CTGF expressions. The downregulation of H₂S is involved in the development of pulmonary artery collagen remodeling induced by high pulmonary blood flow.     

洛沙坦对蛋白激酶C在慢性缺氧大鼠模型肺动脉胶原表达作用的影响

目的　观察洛沙坦对蛋白激酶C(PKC)在慢性缺氧大鼠模型肺动脉胶原表达作用的影响。方法　将二级SD大鼠分为 3组 :A组 (正常对照组 )大鼠室内常规饲养。B组 (单纯缺氧 4周组 )大鼠置于常压低氧舱中 ,舱内充入氮气 ,使氧浓度维持在 (1 0 0± 0 5) % ,每天 8h ,每周 6d ,连续 4周 ;大鼠每天缺氧前用 2ml蒸馏水灌胃。C组 (洛沙坦干预组 )缺氧条件同B组 ,大鼠每天缺氧前用洛沙坦 (洛沙坦 50mg/kg溶于 2ml蒸馏水 )灌胃。采用透射电镜、放射活性测定法、免疫组化、原位杂交等方法观察 3组大鼠肺细小动脉超微结构、肺组织PKC活性、肺动脉管壁PKC免疫组化及Ⅰ、Ⅲ型胶原和Ⅰ、Ⅲ型前胶原基因表达的变化。结果　 (1 )B组大鼠平均肺动脉压、右心室重量比显著高于A组(P <0 0 1 ) ,C组显著低于B组 (P <0 0 1 )。 (2 )光镜下可见B组大鼠肺血管管壁厚度占血管外经的百分比、管壁面积占管总面积的百分比显著高于A组 (P <0 0 1 ) ,C组显著低于B组 (P <0 0 1 )。电镜下可见B组大鼠肺动脉胶原纤维较A组明显为多 ,C组较B组明显为少。 (3)B组大鼠肺组织细胞PKC总活性、胞膜PKC活性、胞质PKC活性及胞膜PKC活性占PKC总活性的百分比显著高于A组(P <0 0 1 ) ,C组上述指标均显著低于B组 (P <0 0 1 )。 (4)免疫组化显示 ,B     

L——精氨酸对缺氧大鼠肺动脉胶原含量的影响

目的：研究缺氧时肺动脉胶原含量的变化Ｌ－精氨酸（Ｌ－Ａｒｇ）对其影响。方法将Ｗｉｓｔａｒ大鼠随机分为对照组,缺氧组和缺氧＋Ｌ－精氨酸组,采用兔组织化学方法测定各组大鼠肺内肺动脉Ⅰ型胶原的含量。结果缺氧时肺内大、中、小型肺动脉中Ⅰ型胶原表达与对照组比较明显增加（ｑ值分别为８．６０,１６．００,１３．１４,Ｐ均〈０．０１）,而缺氧＋Ｌ－精氨酸组肺内大、中、小型肺动脉中Ⅰ型胶原表达与缺氧组相比明显降低（     

低氧性肺动脉高压形成中气体信号分子硫化氢对一氧化碳/血红素加氧酶途径的影响

目的研究新型气体信号分子硫化氢(H2S)在大鼠低氧性肺动脉高压形成中对一氧化碳(CO)/血红素加氧酶(HO1)体系的调节作用,以深入探讨H2S在大鼠低氧性肺动脉高压形成中的病理生理意义。方法将27只大鼠随机分为4组对照组(7只),低氧组(7只),低氧 硫氢化钠(NaHS)组(7只),低氧 炔丙基甘氨酸(PPG)组(6只)。低氧21d后分别测定肺动脉平均压、血浆H2S及CO含量,观察肺动脉平滑肌HO1蛋白及HO1mRNA表达。结果随着低氧性肺动脉高压的形成,血浆H2S的含量显著下降,低氧组[(196±22)μmol/L]与对照组[(294±26)μmol/L]比较差异有显著性(P<005);而CO含量、大、中、小各级肺动脉平滑肌HO1蛋白表达[对照组、低氧组分别为(0313±0020)μmol/L、(0348±0021)μmol/L,066±008、079±008,064±005、077±008,054±005、076±009]及其mRNA表达(对照组、低氧组分别为0573±0148、0813±0052,0532±0131、0831±0043,0473±0102、0819±0032)显著升高(P均<005);外源性给予H2S的供体后,低氧 NaHS组血浆H2S含量[(324±33)μmol/L]显著高于低氧组[(196±22)μmol/L,P<005],肺动脉压显著下降(P<005),且血浆CO含量[(0393±0032)μmol/L]、大、中、小各级肺动脉平滑肌HO1蛋白表达(088±004、089±005、089±006)及mRNA表达(0913±0022、0     

Effect of L-arginine on collagen of high flow-induced pulmonary arterial remodeling.

BACKGROUND: Pulmonary hypertension (PH) is a common complication of congenital heart disease and pulmonary vascular structural remodeling due to the high pulmonary blood flow is considered as the key pathologic process. In the present study the effects of L-arginine on the collagen metabolism of pulmonary arteries in rats with high pulmonary blood flow-induced PH were investigated to elucidate its mechanism. METHODS AND RESULTS: The rat model of PH was established with an abdominal aorta and inferior vena cava shunt. L-arginine (1 g/kg per day) was given directly into the stomachs of the rats with the shunt (L-arginine+shunt group) and 11 weeks later, the pulmonary hemodynamics were studied. Collagen I and collagen III expressions were detected by immunohistochemical assay. The expressions of procollagen I mRNA, procollagen III mRNA, the tissue inhibitor of metalloproteinase-1 mRNA and the matrix metalloproteinase-1 mRNA were detected by in situ hybridization. The expressions of pulmonary artery collagen I, collagen III, procollagen I mRNA and procollagen III mRNA in the shunt rats were obviously elevated compared with the control rats (p<0.01). The positive signals were mainly located in the media and adventitia of median and small pulmonary arteries. The expressions of the pulmonary artery tissue inhibitor of metalloproteinase-1 mRNA, metalloproteinase-1 mRNA and the ratio of tissue inhibitor of metalloproteinase-1/metalloproteinase-1 were elevated in the shunt rats (p<0.01). However, the expressions of pulmonary artery collagen I, collagen III, procollagen I mRNA and procollagen III mRNA were significantly reduced in the shunt rats of the L-arginine group (p<0.01). L-arginine also downregulated the expressions of tissue inhibitor of metalloproteinase-1 mRNA and metalloproteinase-1 mRNA, as well as the ratio of tissue inhibitor of metalloproteinase-1/metalloproteinase-1 (p<0.05). CONCLUSIONS: L-arginine can reduce the synthesis of extracellular matrix-collagen and increase its degradation, thus having an important modulating effect on pulmonary vascular matrix remodeling induced by high pulmonary blood flow.     

慢性痛导致低氧性肺动脉功能异常

目的 研究慢性病理性神经痛对低氧性肺动脉高压发生发展的影响及可能机制。方法 30只SD大鼠随机分为5组:常氧组、慢性痛＋常氧组、低氧组、慢性痛+低氧组和慢性痛+低氧+Alda1（乙醛脱氢酶2特异性激动剂）组，每组6只。慢性痛采用经典的大鼠背根节慢性压迫(chronic compression of dorsal root ganglia,CCD)模型。造模成功后用间断性低压低氧法建立大鼠低氧性肺动脉高压模型，然后分离大鼠三级肺小动脉，检测不同组大鼠的肺血管环舒张、收缩功能变化，同时测定各组大鼠血液循环中4-羟基壬烯酸（4-hydroxy-2-nonenal,4-HNE）的水平和肺动脉上乙醛脱氢酶(Aldehyde dehydrogenase,ALDH)2的表达。结果 低氧导致肺小动脉的舒张功能减弱(P<0.05)，收缩功能亢进；但是在慢性痛状态下，低氧组大鼠肺小动脉舒张功能进一步减弱(P<0.01)，而肺小动脉收缩功能更加亢进(P<0.01)。与此同时，低氧促进大鼠血液中4-HNE的水平升高(P<0.01)，而慢性痛+低氧组大鼠循环血液中4-HNE的水平增高更为显著(P<0.01)，提示慢性痛+低氧组大鼠肺动脉舒缩、功能异常可能是由长期慢性神经痛产生大量的4-HNE造成的。使用ALDH2的激动剂Alda1后能够显著改善慢性痛和低氧导致的肺小动脉舒张、收缩功能异常，促进肺小动脉舒张(P<0.01)，抑制收缩(P<0.01)，而且Alda-1能够显著上调慢性痛和低氧降低的ALDH2的表达(P<0.01)，减少循环血中4-HNE的水平(P<0.01)。结论 慢性病理性神经痛促进循环中4-HNE的水平增高，加重肺小动脉舒张、收缩功能异常，降低ALDH2的表达，促进低氧性肺动脉高压的发生发展。慢性神经痛可能是促进、加重低氧性肺动脉高压发生发展的诱因之一。     

结缔组织生长因子在吸烟者及慢性阻塞性肺疾病患者肺血管中的表达及其与肺血管重塑的关系

目的探讨吸烟者及慢性阻塞性肺疾病（COPD）患者的肺血管重塑中结缔组织生长因子（CTGF）的表达及意义。方法取24份（非吸烟对照组、吸烟组、吸烟伴COPD组,每组8例）手术切除的肺组织,HE染色观察肺血管重塑,天狼猩红染色检测胶原增殖,免疫组化观察CTGF在肺动脉的表达,RT-PCR检测肺动脉CTGF mRNA表达。结果（1）肺动脉管壁面积／管总面积（WA％）,非吸烟对照组为（28．4±4．7）％,吸烟组为（46．3±3．5）％,吸烟伴COPD组为（55．5±3．9）％（P〈0．01）。HE染色可见,吸烟组、吸烟伴COPD组肺动脉管壁B月显增厚。（2）肺动脉壁胶原厚度,非吸烟对照组为（6．4±1．6）μm,吸烟组为（15．9±2．4）μm,吸烟伴COPD组为（16．4±2．3）μm（P〈0．01）。天狼猩红染色可见,吸烟组、吸烟伴COPD组肺动脉管壁胶原明显增多。（3）CTGF mRNA表达量,非吸烟对照组为0．095±0．015,吸烟组为0．396±0．167,吸烟伴COPD组为0．501±0．177（P〈0．01）。（4）CTGF蛋白表达量,非吸烟对照组为0．085±0．011,吸烟组为0．245±0．095,吸烟伴COPD组为0．303±0．191（P〈0．01）。（5）相关分析：CTGF mRNA及蛋白表达量与WA％呈正相关（r分别为0．915、0．919,P〈0．01）。结论单纯吸烟者即有肺血管重塑,吸烟伴COPD者的肺血管重塑程度更重,CTGF可能在此过程中起重要作用。     

Birth associated changes in pulmonary arterial connective tissue gene expression in the normal and hypertensive lung

OBJECTIVES: To determine the temporal and spatial expression of the connective tissue precursors, procollagen and tropoelastin mRNA in normal and pulmonary hypertensive porcine pulmonary arteries from birth onwards. METHODS: Using in situ hybridisation, connective tissue gene expression for procollagen alpha1(I) and alpha1(III) and tropoelastin was studied in intrapulmonary arteries from normal piglets, 5 min-16 weeks, and from piglets made pulmonary hypertensive by exposure to hypobaric hypoxia for 3 days, from birth, 3 or 14 days of age. In addition, Type III pN-procollagen, tropoelastin and collagen I and III were studied by immunohistochemistry. Quantitative or semi-quantitative techniques were applied to both in situ and immunohistochemical studies. RESULTS: Procollagen alpha1(I) and alpha1(III) mRNA expression increased rapidly in the media and adventitia between birth and 3 days of age (P<0.05). The increase was transient and the number of cells expressing procollagen mRNA decreased to the low newborn number after 6 days of age. Type III pN-procollagen immunostaining was greatest in newborn elastic and muscular arteries and then decreased. Collagen I and III increased mainly after 6 days of age. In animals exposed to chronic hypobaric hypoxia from birth, the increase in procollagens I and III mRNA was prevented. Exposure to hypoxia from 3 or 14 days led to little change in either gene expression or in procollagen and mature collagen from the normal. Tropoelastin gene expression was high at birth in the endothelium and media for the first 6 days, and then decreased. Normally, tropoelastin decreased in the media and increased in the adventitia after 16 days of age. Hypoxia had no effect on the mRNA but led to increased tropoelastin. CONCLUSION: We demonstrated marked, rapid changes in temporal and cell specific connective tissue gene expression in normal pulmonary arteries immediately after birth as the vasculature remodels. Each gene appeared to have its own timetable of expression and responded differently to hypoxia-induced hypertension.