【摘要】：目的:探討蛋白酪氨酸激酶6(PTK6)對TNF-α誘導的人氣道上皮屏障功能減損的影響及相關(guān)機制。方法:體外培養(yǎng)人氣道上皮16HBE細胞,予TNF-α刺激,分別轉(zhuǎn)染PTK6 siRNA和recombined PTK6,以轉(zhuǎn)染Scramble siRNA和Empty vector為對照。四甲基偶氮唑鹽法(MTT)檢測細胞活性;跨皮細胞組抗儀檢測細胞跨皮細胞阻抗(TER);辣根過氧化物酶(HRP)流量法反應細胞通透性;RT-PCR檢測ZO-1、Occludin mRNA水平;Western blot檢測PTK6、ZO-1、Occludin、磷酸化細胞外信號調(diào)節(jié)激酶1/2(p-ERK1/2)、磷酸化c-Jun氨基端激酶1/2(p-JNK1/2)和磷酸化p38(p-p38)蛋白水平。結(jié)果:TNF-α刺激后,細胞ZO-1、Occludin轉(zhuǎn)錄及蛋白水平、TER值顯著降低,細胞通透性增高,同時PTK6、p-ERK1/2、p-JNK1/2和p-p38蛋白水平顯著增高(P0.05);上調(diào)PTK6使ZO-1、Occludin轉(zhuǎn)錄及蛋白水平和TER值進一步降低,細胞通透性和p-JNK1/2、p-p38蛋白水平也進一步增高(P0.05),但下調(diào)PTK6后上述指標呈相反變化(P0.05);上調(diào)或下調(diào)PTK6對p-ERK1/2無明顯影響。結(jié)論:下調(diào)PTK6可以降低JNK1/2、p38MAPK磷酸化水平,進而改善TNF-α誘導的氣道上皮屏障功能減損。
[Abstract]:Aim: to investigate the effect of protein tyrosine kinase 6 (PTK6) on TNF- 偽 induced impairment of human airway epithelial barrier function and its related mechanism. Methods: human airway epithelial 16HBE cells were cultured in vitro and stimulated by TNF- 偽. PTK6 siRNA and recombined PTK6, were transfected with Scramble siRNA and Empty vector as control. The cell activity was detected by (MTT) assay, the cell permeability was detected by (TER); horseradish peroxide (HRP) flow assay, and the level of ZO-1,Occludin mRNA was detected by RT-PCR. The levels of extracellular signal-regulated kinase 1 鈮，

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