抑制腱糖蛋白C对新生大鼠缺氧性肺动脉高压的改善功能

Protective function of inhibition of tenascin C in neonatal rats with hypoxic pulmonary hypertension

目的研究抑制腱糖蛋白C(TNC)调控Toll样受体4(TLR4)/核因子-κB(NF-κB)对新生大鼠缺氧性肺动脉高压的影响。方法将新生大鼠分成对照组、模型组(新生大鼠缺氧性肺动脉高压)、sh-NC组(新生大鼠缺氧性肺动脉高压,shRNA control慢病毒载体处理)、sh-TNC组(新生大鼠缺氧性肺动脉高压,TNC shRNA慢病毒载体处理)、sh-TNC+Vector组(新生大鼠缺氧性肺动脉高压,阴性对照慢病表达载体、TNC shRNA慢病毒载体处理)、sh-TNC+TLR4组(新生大鼠缺氧性肺动脉高压,TLR4过表达慢病表达载体、TNC shRNA慢病毒载体处理)。以免疫组化法检测TNC、TLR4、p65、内皮素-1(ET-1)、诱导型一氧化氮合酶(iNOS)、B淋巴细胞瘤-2(Bcl-2)、Bcl-2相关X基因(Bax),以直接测压法检测平均右心室收缩压(RVSP),以苏木精-伊红(HE)染色分析中膜横截面积(MA)、中膜厚度(MT)。结果对照组、模型组、sh-NC组、sh-TNC组、sh-TNC+Vector组和sh-TNC+TLR4组大鼠处理10 d后TLR4 IOD值分别为0.12±0.02、0.19±0.02、0.19±0.01、0.12±0.01、0.13±0.02和0.18±0.02,RVSP分别为(19.17±1.16)、(25.65±1.59)、(26.57±2.11)、(20.43±0.77)、(19.18±1.90)和(21.08±1.03)mmHg,iNOS IOD值分别为2.29±0.19、4.64±0.12、4.52±0.08、3.43±0.37、3.42±0.18和4.29±0.21,MA值分别为(56.15±4.84)%、(69.48±2.72)%、(70.35±5.71)%、(58.26±2.43)%、(57.29±3.73)%和(66.34±2.20)%,Bax IOD值分别为0.12±0.01、0.22±0.02、0.23±0.01、0.16±0.02、0.17±0.01和0.22±0.03。模型组的上述指标与对照组比较,sh-TNC组的上述指标与sh-NC组比较,sh-TNC+TLR4组的上述指标与sh-TNC+Vector组比较,差异均有统计学意义(均P<0.05)。结论抑制TNC通过下调TLR4/NF-κB激活水平改善新生大鼠缺氧性肺动脉高压模型肺血管重塑,并减少肺血管内皮细胞凋亡。

Objective To study the effect of inhibiting tenascin C(TNC)on regulation of toll-like receptor-4(TLR4)/nuclear factor-κB(NF-κB)in neonatal rats with hypoxic pulmonary hypertension.Methods Neonatal rats were divided into control group,model group(hypoxic pulmonary hypertension neonatal rats),sh-NC group(hypoxic pulmonary hypertension neonatal rats,shRNA control lentiviral vector treatment),sh-TNC group(hypoxia pulmonary hypertension neonatal rats,treated with TNC shRNA lentiviral vector),sh-TNC+Ve cto r group(hypoxic pulmonary hypertension neonatal rats,treated with negative control chronic disease expression vector,TNC shRNA lentiviral vector),sh-TNC+TLR4 group(hypoxic pulmonary hypertension neonatal rats,treated with TLR4 overexpressed chronic disease expression vector,TNC shRNA lentiviral vector).Immunohistochemical detection of TNC,TLR4,p65,vascular endothelin-1(ET-1),inducible nitric oxide synthase(iNOS),Bcl-2 associated X protein(Bax),B cell leukemia iymphoma-2(Bcl-2),and direct manometry was used to measure mean right ventricular systolic pressure(RVSP),hematoxylin-eosin staining was used to analyze median cross-sectional area(MA),media thickness(MT).Results The TLR4 IOD after 10 days of treatment in control group,model group,sh-NC group,sh-TNC group,sh-TNC+Vector group and sh-TNC+TLR4 group were 0.12±0.02,0.19±0.02,0.19±0.01,0.12±0.01,0.13±0.02 and 0.18±0.02,respectively;RVSP were(19.17±1.16),(25.65±1.59),(26.57±2.11),(20.43±0.77),(19.18±1.90)and(21.08±1.03)mmHg,respectively;iNOS IOD values were 2.29±0.19,4.64±0.12,4.52±0.08,3.43±0.37,3.42±0.18 and 4.29±0.21,respectively;MA were(56.15±4.84)%,(69.48±2.72)%,(70.35±5.71)%,(58.26±2.43)%,(57.29±3.73)%and(66.34±2.20)%,respectively;Bax IOD values were 0.12±0.01,0.22±0.02,0.23±0.01,0.16±0.02,0.17±0.01 and 0.22±0.03,respectively.There were statistically significant differences of the above indexes between model group and control group,between sh-TNC group and sh-NC group,between sh-TNC+TLR4 group and sh-TNC+Vector group(all P<0.05).Conclusion Inhibition of TNC improves pulmonary vascular remodeling and reduces pulmonary vascular endothelial cell apoptosis in neonatal rats with hypoxic pulmonary hypertension by down-regulating the activation level of TLR4/NF-κB.