紫杉醇对肺动脉高压大鼠肥大细胞-CCL2-巨噬细胞轴的影响

Effect of paclitaxel on mast cell-CCL2-macrophage axis in rats with pulmonary hypertension

目的评价紫杉醇对肺动脉高压大鼠肥大细胞-趋化因子配体2(CCL2)-巨噬细胞轴的影响。方法SPF级健康雄性SD大鼠30只,8~10周龄,体重180~220 g,采用随机数字表法分为3组(n=10):对照组(C组)、肺动脉高压组(PH组)和紫杉醇组(PTX组)。采用皮下注射野百合碱60 mg/kg的方法制备大鼠肺动脉高压模型,模型制备开始后25 d,PTX组经尾静脉注射紫杉醇2 mg/kg,1次/4 d,共4次。余2组注射等量生理盐水。模型制备后40 d,行平均肺动脉压(mPAP)测定。随后取心脏烘干后分别称重右心室(RV)和左心室加室间隔(LV+S),计算Fulton指数。取左下肺叶,HE染色测算肺血管中膜厚度比值,免疫组化法计数每个血管周围Tryptase^(+)、CD68^(+)和CD163^(+)总细胞数,并计算平均值,计数并计算血管Ki67阳性细胞数量百分比,测算肌化血管比例。ELISA法检测CCL2含量,Western blot法检测裂解caspase-3和Cyclin D1表达。结果与C组比较,PH组和PTX组mPAP、Fulton指数、血管中膜厚度比值和肌化血管比例升高,Tryptase^(+)、CD68^(+)和CD163^(+)细胞数量增加,Ki67^(+)细胞百分比升高,肺组织裂解caspase-3表达下调,PH组肺组织Cyclin D1表达上调(P<0.05),PTX组差异无统计学意义(P>0.05);与PH组比较,PTX组mPAP、Fulton指数、血管中膜厚度比值及肌化血管比例降低,Tryptase^(+)、CD68^(+)和CD163^(+)细胞数量减少,Ki67^(+)细胞百分比降低,肺组织CCL2和Cyclin D1表达下调,裂解caspase-3表达上调(P<0.05)。结论紫杉醇缓解大鼠肺动脉高压的机制与抑制肥大细胞-CCL2-巨噬细胞轴有关。

Objective To evaluate the effect of paclitaxel on the mast cell-CCL2-macrophage axis in rats with pulmonary hypertension.Methods Thirty SPF-grade healthy male Sprague-Dawley rats,aged 8-10 weeks,weighing 180-220 g,were divided into 3 groups(n=10 each)using a random number table method:control group(group C),pulmonary hypertension group(group PH),and paclitaxel group(group PTX).The model of pulmonary hypertension was established by subcutaneous injection of monocrotaline 60 mg/kg in rats.At 25 days after establishing the models,paclitaxel 2 mg/kg was injected via the tail vein once every four days,for 4 times in total in group PTX.The equal volume of normal saline was injected in the remaining 2 groups.The mean pulmonary artery pressure(mPAP)was performed at 40 days after establishing the model.The heart was removed and dried,the right ventricle(RV)and left ventricle plus ventricular septum(LV+S)was weighed,and the Fulton index[RV/(LV+S)]was calculated.The inferior lobe of left lung was taken,the ratio of media wall thickness of pulmonary vessels was calculated by HE staining,the number of Tryptase^(+),CD68^(+),CD163^(+),and Ki67^(+)cells was recorded by immunohistochemistry,the mean value was calculated,the percentage of Ki67-positive cells in blood vessels was recorded,and the proportion of muscularized blood vessels was calculated.The content of CCL2 was measured by enzyme-linked immunosorbent assay,and the expression of cleaved caspase-3 and Cyclin D1 was detected by Western blot.Results Compared with group C,the mPAP,Fulton index,ratio of media wall thickness,proportion of muscularized blood vessels,the number of Tryptase^(+),CD68^(+)and CD163^(+)cells and percentage of Ki67^(+)cells were significantly increased,and the expression of cleaved caspase-3 was down-regulated in PH and PTX groups(P<0.05),the expression of Cyclin D1 was significantly up-regulated in group PH(P<0.05),and no significant change was found in group PTX(P>0.05).Compared with group PH,the mPAP,Fulton index,ratio of media wall thickness,percentage of muscularized blood vessels,the number of Tryptase^(+),CD68^(+)and CD163^(+)cells and percentage of Ki67^(+)cells were significantly decreased,the expression of CCL2 and Cyclin D1 was down-regulated,and the expression of cleaved caspase-3 was up-regulated in group PTX(P<0.05).Conclusion The mechanism by which paclitaxel alleviates pulmonary hypertension is related to inhibiting the mast cell-CCL2-macrophage axis in rats.