【作者】 翟凤国；

【导师】 王怀良；

【作者基本信息】 中国医科大学 ， 药理学， 2009， 博士

【摘要】 目的肺动脉高压(PAH)是以肺动脉压持续性、渐进性增高,导致右心室肥厚、右心衰竭并最终引起死亡为特征的一组疾病。肺动脉高压发病机制复杂,目前尚未明确,可行的治疗方法有限、昂贵并伴有明显的副作用。PAH具有复杂的病理改变,主要包括肺血管收缩性增强、肺血管构型重建、肺小血管内微血栓形成以及肺组织炎症反应。在肺动脉高压中,肺血管构型重建具有肺血管结构改变并伴有中膜肥厚的特征。这主要是由于肺动脉平滑肌细胞(PASMCs)增殖和凋亡失衡所致。已证明PASMCs增殖增加和/或PASMCs凋亡减少都可促进肺血管中膜肥厚和血管构型重建。因此,肺动脉高压更有效的治疗将直接并选择性的针对导致肺血管平滑肌细胞增殖增加和凋亡减少的机制。最近,以诱导细胞凋亡为基础来降低肺血管增厚的治疗方法受到关注并在动物实验中获得成功。因此,抑制肺动脉平滑肌细胞增殖和促进肺动脉平滑肌细胞凋亡可能成为人类肺动脉高压治疗的突破口。近年来,国内外学者研究发现钾通道也可通过调节PASMCs增殖和凋亡参与肺血管构型重建。PASMCs中有多种Kv通道表达,其中最重要的是Kv1.5。Kv1.5表达减少或活性降低不仅通过引发肺动脉平滑肌细胞膜去极化和增加胞浆钙离子浓度引起肺血管收缩,而且还通过抑制细胞皱缩和凋亡促进肺血管中膜肥厚。反之,Kv1.5表达增高或活性增强可增加肺动脉平滑肌细胞凋亡/增殖比率并进而抑制肺动脉高压。肺动脉高压患者和实验性肺动脉高压动物模型也显示Kv1.5表达下调,对细胞凋亡的抵抗作用进一步增强。肺动脉平滑肌细胞的增殖和凋亡被许多血管活性因子、生长因子和细胞因子所调控。其中,5-羟色胺(5-hydroxytryptamine,5-HT)是一个强有力的血管收缩因子和促有丝分裂因子,在PAH发病机制中具有重要作用。研究发现,肺动脉高压患者血浆中5-HT水平明显高于正常人,体外实验证明高浓度的5-HT能使人离体肺血管明显收缩。5-HT可经细胞膜上的5-羟色胺转运体(5-hydfoxytraptaminetransporter,5-HTT)介导进入肺动脉平滑肌细胞,并通过多种增殖信号转导途径引起PASMCs增殖肥大,导致肺血管构型重建。5-HT也可以抑制Kv1.5电流。此外,肺动脉高压患者肺动脉中5-HTT表达增加,暴露在低氧条件下的PASMCs 5-HTTmRNA表达也增加。越来越多的研究结果表明5-羟色胺和5-羟色胺转运体在肺动脉高压发病机制中起关键作用。选择性5-羟色胺再摄取抑制剂(SSRIs)包括氟西汀、舍曲林、帕罗西汀、西肽普兰等,是近年来临床常用的抗抑郁症药物。它们通过阻断突触前膜上的5-HTT,抑制5-HT再摄取,提高突触间隙5-HT浓度,从而发挥治疗作用。我们最近报道了氟西汀可在体外抑制5-HT介导的肺动脉平滑肌细胞增殖。也有研究表明氟西汀可抑制5-羟色胺对肺动脉平滑肌细胞Kv电流的影响并抑制肺血管收缩。此外,氟西汀也保护慢性低氧诱导的肺血管构型重建和肺动脉高压。氟西汀抗肺动脉高压的作用是否与它对Kv1.5通道及肺动脉平滑肌细胞增殖和凋亡的调控有关均不清楚。因此在本研究中,我们通过建立野百合碱(MCT)诱导的慢性大鼠肺动脉高压模型来观察肺动脉平滑肌细胞的增殖、凋亡和Kv1.5在MCT诱导的肺动脉高压中的作用,并探讨氟西汀抗肺动脉高压作用是否与这些因素有关。实验方法(一)建立肺动脉高压大鼠模型体重(160±10g)的雄性Wistar大鼠80只(中国医科大学实验动物中心,GradeⅡ,Certificate No.034),随机分成4组:对照组(control)、MCT组、MCT+氟西汀2mg/kg剂量组(MCT+F2)和MCT+氟西汀10mg/kg剂量组(MCT+F10)。MCT组、MCT+F2组和MCT+F10组大鼠一次性腹腔内注射MCT 60mg/kg,对照组注射同等剂量的溶剂。继而,MCT+F2组和MCT+F10组大鼠每天分别给予氟西汀2mg/kg和10mg/kg灌胃,MCT组和对照组大鼠每天给予等量溶剂黄耆胶灌胃。各组大鼠常规给予食物和水,保持温度(18℃～22℃)和湿度(50%～70%),饲养3周。(二)大鼠肺动脉压、体循环压和右心室肥厚指数检测给药21天后,大鼠用3%戊巴比妥(40mg/kg)经腹腔内注射麻醉,切开颈部皮肤,分离右侧颈外静脉和颈总动脉。将微型聚乙烯导管插入右颈总动脉测量体循环压(SAP),将另一聚乙烯导管自右颈外静脉插入肺动脉测量肺动脉压(PAP),由压力传感器连至多导生理记录仪记录数据。经以上检测后将大鼠过量麻醉处死,取出整个心脏,分离右心室(RV)和左心室+室间隔(LV+S),分别称量RV和LV+S的重量,RV/(LV+S)即为右心室肥厚指数(RVI)。(三)肺组织形态学检测处死大鼠后,取大鼠右肺下叶固定于4%的多聚甲醛,常规石蜡包埋。将5μm厚的大鼠肺动脉石蜡切片用HE染色后,光学显微镜检测,并对肺小动脉血管壁厚度进行统计学分析。每组取3只大鼠,每只大鼠取外径在50～200μm的20个肺肌型小动脉,测其内外径,用下面公式估算肺血管壁厚度:血管壁厚度%=(血管外径-血管内径)/血管外径×100%(四)肺动脉平滑肌细胞增殖和凋亡的检测用免疫组化染色法检测PCNA表达来观察肺动脉平滑肌细胞增殖情况;用TUNEL法检测肺动脉平滑肌细胞凋亡情况。每张切片随机计数10个视野,在光学显微镜(×400)下观察PCNA和TUNEL阳性细胞数。(五) Western Blot法检测各组大鼠肺动脉Bcl-2,Bcl-xl,Cleavedcaspase-3蛋白的表达v(六)RT-PCR法检测各组大鼠肺动脉Kv1.5 mRNA的表达(七) Western Blot法检测各组大鼠肺动脉Kv1.5蛋白的表达(八)统计学分析所有数据以mean±SD表示,采用单因素方差分析。二组间差异采用LSD法检验。P＜0.05时认为差异具有统计学意义。采用SPSS 13.0软件进行统计学分析。实验结果1、血流动力学指标检测与对照组相比,MCT组大鼠平均肺动脉压从17.8±1.2 mmHg明显升高到32.7±5.7 mmHg(P＜0.01);与MCT组相比,MCT+F10剂量组平均肺动脉压明显降低,平均肺动脉压为25.8±5.4 mmHg(P＜0.05)。MCT+F2剂量组平均肺动脉压比MCT组低,但没有达到统计学意义。平均体循环压在各组间也无明显差异。2、右心室肥厚指数检测与对照组相比,MCT组右心室肥厚指数由34.1%±2.1%明显增加到51.3%±8.8%(P＜0.01);氟西汀以剂量依赖性方式明显降低了MCT引起的右心室肥厚指数的增加,MCT+F2剂量组降至45.2%±5.3%(P＜0.05),MCT+F10剂量组降至43.4%±3.3%(P＜0.01)。3、肺组织形态学分析肺组织HE染色显示,对照组大鼠肺小动脉内皮细胞连续性好,细胞分布均匀,管壁薄,管腔大且厚薄一致;MCT组大鼠肺小动脉内皮细胞连续性破坏,细胞肿胀、变性、坏死和脱落,血管中膜平滑肌细胞明显增生,管壁厚度明显增加,管腔面积明显变小;与MCT组相比,MCT+F2和MCT+F10剂量组大鼠肺小动脉内皮细胞连续性较好、管壁厚度减小、管腔面积增大,且以MCT+F10组更为明显。与对照组相比,MCT组肺小动脉血管壁明显增厚,管壁厚度由26.9%±7.1%增加至50.5%±10.8%(P＜0.01);氟西汀剂量依赖性的降低了MCT引起的肺血管壁增厚,MCT+F2、MCT+F10剂量组管壁厚度分别由MCT组50.5%±10.8%降至46.0%±8.2%(P＜0.01)和36.6%±7.0%(P＜0.01)(图2,3)。这些结果表明MCT可诱发明显的肺血管构型重建,氟西汀明显抑制了MCT引起的肺血管构型重建。4、氟西汀抑制肺血管平滑肌细胞增殖与对照组相比,MCT组肺动脉中膜血管平滑肌细胞中PCNA阳性细胞百分比由3.7%±1.2%明显增加到15.7%±3.2%(P＜0.01)。与MCT组相比,MCT+F2剂量组PCNA阳性细胞百分比无明显改变,但MCT+F10剂量组明显抑制肺血管平滑肌细胞增殖,PCNA阳性细胞减少到8.2%±2.8%(P＜0.01)。5、氟西汀增强肺血管平滑肌细胞的凋亡与对照组相比,MCT组肺动脉中膜血管平滑肌细胞中TUNEL阳性细胞百分比由3.3%±1.3%略降低到2.9%±1.1%(P＜0.05)。氟西汀剂量依赖性地增强肺动脉平滑肌细胞凋亡,与MCT组相比,MCT+F2,MCT+F10剂量组TUNEL阳性细胞百分比分别增加至7.9%±1.9%(P＜0.01)和11.7%±2.5%(P＜0.01)。6、肺动脉Bcl-2,Bcl-xl和Cleaved caspase-3蛋白表达用Western blot方法检测肺动脉Bcl-2,Bcl-xl和Cleaved caspase-3蛋白表达变化。结果显示,与对照组相比,MCT组肺动脉Bcl-2和Bcl-xl蛋白表达明显增加,分别由1.22±0.10和1.86±0.25增加至2.89±0.82(P＜0.05)和2.91±0.57(P＜0.05);氟西汀剂量依赖性的降低这两种抗凋亡蛋白的表达,MCT+F2剂量组蛋白表达分别降低至2.65±0.97和2.32±0.45,而MCT+F10剂量组蛋白表达明显降低至1.55±0.60(P＜0.05vsMCT)和2.01±0.37(P＜0.05 vs MCT)。相反,MCT组肺动脉Cleaved caspase-3蛋白表达由对照组的1.49±0.10明显降低至0.68±0.18(P＜0.01),氟西汀剂量依赖性的增加Cleaved caspase-3蛋白的表达,MCT+F2和MCT+F10剂量组蛋白表达分别增加至1.20±0.27(P＜0.05 vs MCT)和1.33±0.24(P＜0.01 vs MCT)。7、肺动脉Kv1.5 mRNA和蛋白表达MCT组肺动脉Kv1.5 mRNA表达明显降低,由对照组的1.23±0.42降低至2.00±0.55(P＜0.05)。与MCT组相比,MCT+F2剂量组肺动脉Kv1.5 mRNA表达无明显改变,但MCT+F10剂量组Kv1.5 mRNA表达明显增加,mRNA表达增加至1.90±0.38(P＜0.05 vs MCT)。Western Blot蛋白分析表明,与对照组比较,MCT组肺动脉Kv1.5蛋白表达由1.23±0.22降低到0.70±0.06(P＜0.01);而氟西汀剂量依赖性地增加了Kv1.5的蛋白表达,MCT+F2,MCT+F10剂量组Kv1.5蛋白表达分别增加至1.02±0.14(P＜0.05 vs MCT)和1.09±0.14(P＜0.05 vs MCT)。结论1、氟西汀对MCT诱导的慢性肺动脉高压大鼠有明显的抑制肺血管构型重建、减轻右心室肥厚和降低肺动脉压作用。2、氟西汀通过抑制MCT诱导的肺动脉高压大鼠PASMCs增殖和促进PASMCs凋亡减轻肺小动脉中膜增厚,从而抑制肺血管构型重建,其机制与氟西汀抑制抗凋亡蛋白Bcl-2、Bcl-xl和增强促凋亡蛋白Cleaved caspase-3的表达有关。3、氟西汀抑制了MCT引起的Kv1.5 mRNA和蛋白表达的降低。推测氟西汀抗肺动脉高压作用与其增加Kv1.5 mRNA和蛋白表达,从而抑制肺血管收缩和肺血管构型重建有关。更多还原

【Abstract】 ObjectivePulmonary arterial hypertension（PAH） is characterized by a sustained and progressive elevation of pulmonary arterial pressure leading to right ventricular hypertrophy,right heart failure and ultimately to death.The cause remains unknown and available treatments are limited,expensive,and often associated with significant side effects.PAH has a complex pathobiology involving pulmonary vasoconstriction, vascular remodeling,inflammation and microthrombosis.The pulmonary vascular remodeling in PAH is characterized by changes in pulmonary vascular structure associated with medial hypertrophy,which is mainly caused by imbalanced proliferation and apoptosis in pulmonary artery smooth muscle cells（PASMCs）.It has also been demonstrated that either increased proliferation or decreased apoptosis of PASMCs contribute to pulmonary vascular medial hypertrophy and vascular remodeling.More effective therapies for PAH will have to directly and selectively target the mechanisms leading to the pro-proliferative and anti-apoptotic environment in the vascular wall.Recently,"apoptosis-based therapeutic strategies" to reduce pulmonary vascular thickening gained attention and have been successful in experimental animals.Thus,inhibition of PASMCs growth and augmentation of PASMCs apoptosis could serve as therapeutic approaches for patients with PAH. However,the precise mechanisms involved in the regulation of PASMCs proliferation and apoptosis in PAH are still incompletely understood.It has been recently discovered that Kv channels also participate in vascular remodeling by regulating cell proliferation and apoptosis.From the variety of Kv channels expressed in PASMCs,special interest has been paid to Kvl.5.The decreased Kv channels expression and activity not only causes pulmonary vasoconstriction by inducing membrane depolarization and increases in cytoplasmic Ca²⁺ concentration([Ca²⁺]cyt) in PASMCs but also contributes to pulmonary vascular medial hypertrophy by inhibiting apoptotic cell shrinkage and apoptosis.However, up-regulation of Kvl.5 correlates with an increase in apoptosis/proliferation ratio and inhibition of PAH.The resistance to apoptosis is further enhanced by the selective downregulation of Kv channels that has been shown in human and animal models of PAH.Endogenous substances,e.g.vasoactive agonists,growth factors and cytokines, regulate PASMCs proliferation and apoptosis.Among these substances,serotonin （5-hydroxytryptamine,5-HT） is a potent vasoconstrictor and mitogen that has been implicated in the pathophysiology of PAH.Serotonin mediated proliferative signaling pathways via serotonin transporter（5-HTT） and serotonin receptors on PASMCs. Furthermore,the expression of 5-HTT in the pulmonary artery is increased in patients with pulmonary arterial hypertension.Exposure of PASMCs to hypoxia results in a rapid increase in the level of 5-HTT mRNA.It has been shown that 5-HT and 5-HTT play a central role in the pathogenesis of pulmonary artery smooth muscle cell proliferation,not only in experimental PAH but also in human PAH,whether idiopathic （iPAH） or associated with various diseases（aPAH）.Selective serotonin re-uptake inhibitors（SSRIs）,which are used clinically as antidepressants,such as fluoxetine,sertraline and paroxetine etc.,block serotonin internalization by inhibiting 5-HTT.We previously reported that fluoxetine decreased serotonin mediated proliferation of PASMCs in vitro.It has been shown that fluoxetine inhibited the effects of 5-HT on Kv currents in PASMCs and pulmonary vasoconstriction.Furthermore,fluoxetine also protected rats from both chronically hypoxic and monocrotaline（MCT）-induced pulmonary vascular remodeling and pulmonary arterial hypertension.However,it is not clear whether the beneficial effects of fluoxetine on pulmonary artery remodeling is relevant to its effects on Kv channel,PASMCs proliferation and apoptosis.Therefore,the present study was designed to investigate the roles of apoptosis and Kvl.5 channels in MCT-induced PAH, and if these factors are involved in the protective effect against PAH by fluoxetine.Methods1.Animal Models and Experimental DesignEighty male Wistar rats（160±10g） from Animal Resource Center,China Medical University（Certificate No:Liaoning 034） were randomly divided into four groups,i.e. control,MCT,MCT plus fluoxetine 2mg/kg（MCT + F2） and MCT plus fluoxetine 10 mg/kg（MCT + F10）.Rats in the MCT group and two fluoxetine-treated groups were treated with a single intraperitoneal injection of MCT,and rats in the control group were received an intraperitoneal injection of vehicle.Rats in MCT + F2 and MCT + F10 groups were further treated with intragastric administration of fluoxetine 2 mg/kg and 10 mg/kg once a day for 3 weeks,respectively.Meanwhile,rats in the control and MCT groups were treated with vehicle only.And then these rats were fed with solid food and water ad lib in an alternating 12 h light/dark cycle under controlled temperature（18-22℃） and humidity（50%-70%） for 3 weeks.2.Assessment of pulmonary arterial hypertensionAfter 3 weeks,rats in all groups were intraperitoneal anaesthetized with 3% pentobarbital sodium（40 mg/kg）.A polyethylene catheter was inserted into the right carotid artery to measure systemic arterial pressure（SAP）.Another polyvinyl PV-1 catheter was introduced into the right jugular vein and pushed through the right ventricle into the pulmonary artery for measurement of pulmonary arterial pressure （PAP）.After the measurements had been completed,rats were sacrificed with overdose of pentobarbital sodium.Heart tissue was separated as right ventricle（RV） and left ventricle plus septum（LV + S）,right ventricular index was calculated by RV/（LV + septum）.3.Morphometric AnalysisThe right lower lobes of the rat lungs were cut and fixed with formalin buffer. After paraffin embedding,5μm-thick lung sections were stained with hematoxyline-eosine（HE）.The morphologic alterations of the pulmonary arteries were observed under optical microscope.Three rats per group were studied and from each rat at least 2 separate lung sections were examined.Resistance pulmonary arteries（PAs）（50-200mm） chosen randomly from low power fields（magnification,×10） were analyzed（approximately 60 arteries/group;2-3 slides/rat;3 rats/group） by 2 blinded investigators.Pulmonary arteries remodeling was measured as percent medial thickness.Medial wall thickness of pulmonary arteries was calculated and expressed as follows:index（%） =[（external diameter-internal diameter）/external diameter]x 100.4.Protein expression of Bcl-2,Bcl-xl and Cleaved caspase-3 were detected by Western Blot5.Expression of Kvl.5 mRNA was evaluated by reverse transcfiption-polymerase chain reaction（RT-PCR）6.Protein expression of Kv 1.5 was detected by Western blot7.Proliferative and apoptotic assessmentProliferating cells were evaluated by using proliferating cell nuclear antigen immunohistological staining;apoptotic cells were detected by using the terminal deoxynucleotidyl transferase（TdT）-mediated dUTP nick end-labeling（TUNEL） method according to the manufacture’s instruction of the in situ cell death detection kit. The number of PCNA- and TUNEL-positive cells in 10 fields for each section of small to moderate size pulmonary arteries was quantitatively evaluated as a percent of total SMCs at a magnification of 400×in a blind manner. 8.Statistical analysisAll data are expressed as mean±SD.Statistical analyses were performed by one-way ANOVA.A value of P＜0.05 was considered statistically significant.Results1.Hemodynamics assessmentCompared with the control group,mean PAP in the MCT group was significantly elevated from 17.8±1.2 to 32.7±5.7 mmHg（P＜0.01 vs control）,but was reduced from 32.7±5.7 to 25.8±5.4 mmHg by fluoxetine 10mg/kg（P＜0.05,vs MCT）.Yet, Mean SAP did not differ significantly in all groups.2.Right ventricular indexRVI（%） was significantly increased by MCT treatment from 34.1±2.1 to 51.3±8.8（P＜0.01 vs control） and decreased in MCT + F2 and MCT + F10 groups from 51.3±8.8 to 45.2±5.3（P＜0.05 vs MCT） and 43.4±3.3（P＜0.01,vs MCT）,respectively.3.Morphological analysis of pulmonary arteriesPulmonary arteries in the MCT group exhibited obvious medial wall thickening. The percentage of medial wall thickness of pulmonary arteries was significantly increased in the MCT group from 26.9±7.1 to 50.5±10.8（P＜0.01 vs control）. Compared with the MCT group,the percentage of medial wall thickness of pulmonary arteries in MCT + F2 and MCT + F10 groups were reduced from 50.5±10.8 to 46.0±8.2（P＜0.05 vs MCT） and 36.6±7.0（P＜0.01 vs MCT）,respectively.These results indicated that MCT significantly promotes pulmonary vascular remodeling,whereas fluoxetine attenuates the effects of MCT.4.Protein expression of Bcl-2,Bcl-xl and Cleaved caspase-3Protein expression of Bcl-2,Bcl-xl and Cleaved caspase-3 were measured by Western blot.Compared with the control group,the levels of Bcl-2,Bcl-xl in the MCT group were significantly increased from 1.22±0.10 and 1.86±0.25 to 2.89± 0.82 and 2.91±0.57（both P＜0.05 vs control）,respectively.Fluoxetine inhibited MCT induced increase of these anti-apoptotic genes in a dose dependent manner.In MCT + F2 group,these levels were decreased to 2.65±0.97 and 2.32±0.45, respectively,although there was no statistical significance compared with the MCT group.And in MCT±F10 group,these levels were significantly decreased to 1.55±0.60 and 2.01±0.37（both P＜0.05 vs MCT）,respectively.It was also found that Cleaved caspase-3 expression decreased in the MCT group from 1.49±0.10 to 0.68±0.18（P＜0.01 vs control）.And it was increased to 1.20±0.27（P＜0.05 vs MCT） and 1.33±0.24（P＜0.01 vs MCT） in MCT + F2 group and MCT + F10 group, respectively.5.Expression of Kv1.5 by RT-PCR and Western blotSemiquantitative RT-PCR analysis was utilized to detect changes in mRNA expression of Kv1.5 in pulmonary arteries of MCT-induced PAH rats.The PCR products of the expected sizes for Kv1.5 andβ-actin were 267bp and 612bp, respectively.MCT significantly decreased the expression of Kv1.5 mRNA levels compared with the control group（1.23±0.42 vs 2.00±0.55;P＜0.05）.Compared with the MCT group,the level of Kv1.5 mRNA was not obviously changed in MCT+ F2 group,yet it was increased significantly from 1.23±0.42 to 1.90±0.38 in MCT + F10 group（P＜0.05 vs MCT）.Results from Western blot analysis also demonstrated that the levels of relative Kv1.5 expression decreased in the MCT group compared with the control group（0.70±0.06 vs 1.23±0.22;P＜0.01）,and it was increased in MCT + F2 and MCT + F10 groups（1.02±0.14 vs 0.70±0.06,1.09±0.14 vs 0.70±0.06,respectively;P＜0.05 vs MCT）.6.Effect of fluoxetine treatment on PASMCs proliferationFluoxetine treatment suppressed proliferation and enhanced apoptosis of PASMCs in MCT-induced PAH rats.The percentage of PCNA-positive PASMCs in small to moderate sized pulmonary arteries was significantly increased in the MCT group compared with the control group（15.7±3.2%vs 3.7±1.2%;P＜0.01）.Then,it was decreased in MCT + F10 group（8.2±2.8%vs 15.7±3.2%;P＜0.01 vs MCT）.7.Effect of fluoxetine treatment on PASMCs apoptosisThe percentage of TUNEL-positive cells was slightly decreased in the MCT group compared with the control group（2.9±1.1%vs 3.3±1.3%;P＜0.05）,which was increased in MCT + F2 and MCT + F10 groups（7.9±1.9%vs 2.9±1.1%,11.7±2.5% vs 2.9±1.1%,respectively;P＜0.01 vs MCT）.Conclusion1.Fluoxetine protected against MCT-induced pulmonary arterial hypertension by decreasing pulmonary arterial pressure,reducing right ventricular index and inhibiting pulmonary artery remodeling.2.Fluoxetine protected against vascular remodeling by suppressing PASMCs proliferation and inducing PASMCs apoptosis which might be related to decreasing expression of Bcl-2 and Bcl-xl protein,increasing expression of Cleaved caspase-3 protein.3.Fluoxetine enhanced expression of Kvl.5 protein and mRNA levels in pulmonary arteries of MCT-induced PAH rats.These findings suggested that fluoxetine protected against MCT-induced PAH,in which up-regulating Kvl.5 channels was considered to be involved.更多还原