University of Technology Dresden, Department of Anatomy, Medical Clinic, Fetscherstr. 76, 01307 Dresden, Germany.
Int J Biochem Cell Biol. 2012 Mar;44(3):514-24. doi: 10.1016/j.biocel.2011.12.003. Epub 2011 Dec 14.
Changes in intracellular calcium concentration Ca(2+) are believed to influence the proliferation and differentiation of airway epithelial cells both in vivo and in vitro. In the present study, using mouse alveolar epithelial E10 cells, we demonstrated that the treatment of lung epithelial cells with BLM resulted in elevated intracellular Ca(2+) levels. BLM further increased P2rx7 mRNA expression and P2X7R protein levels, paralleled by increased PKC-β1 levels. BLM treatment or stimulation of the P2X7R with the P2X7R agonist BzATP induced translocation of PKC-β1 from the cytoplasm to the membrane. The expression of PKC-β1 was repressed by the P2X7R inhibitor oxATP, suggesting that PKC-β1 is downstream of P2X7R activation. Furthermore, cells exposed to BLM contained increased amounts of P2X7R and PKC-β1 in Cav-1 containing lipid raft fractions. The comparison of lung tissues from wild-type and P2rx7(-/-) mice revealed decreased protein and mRNA levels of PKC-β1 and CaM as well as decreased immunoreactivity for PKC-β1. The knockdown of P2X7R in alveolar epithelial cells resulted also in a loss of PKC-β1. These data suggest that the effect of P2X7R on expression of PKC-β1 detected in alveolar epithelial cells is also functioning in the animal model. Immunohistochemical evaluation of fibrotic lungs derived from a BLM-induced mouse model revealed a strong increase in PKC-β1 immunoreactivity. The present experiments demonstrated that the increased expression of P2X7R influences PKC-β1. We predict that increased Ca(2+) concentration stimulates PKC-β1, whereas the prerequisite for activating PKC-β1 after P2X7R increase remained to be determined. Our findings suggest that PKC-β1 is important in the pathogenesis of pulmonary fibrosis.
细胞内钙离子浓度 Ca(2+) 的变化被认为会影响体内和体外气道上皮细胞的增殖和分化。在本研究中，我们使用小鼠肺泡上皮 E10 细胞证明，博来霉素处理肺上皮细胞会导致细胞内 Ca(2+)水平升高。BLM 进一步增加了 P2rx7 mRNA 表达和 P2X7R 蛋白水平，同时增加了 PKC-β1 水平。BLM 处理或用 P2X7R 激动剂 BzATP 刺激 P2X7R 可诱导 PKC-β1 从细胞质向膜转位。P2X7R 抑制剂 oxATP 抑制 PKC-β1 的表达，表明 PKC-β1 是 P2X7R 激活的下游。此外，暴露于 BLM 的细胞在含有 Cav-1 的脂筏部分含有更多的 P2X7R 和 PKC-β1。野生型和 P2rx7(-/-) 小鼠肺组织的比较显示 PKC-β1 和 CaM 的蛋白和 mRNA 水平降低，以及 PKC-β1 的免疫反应性降低。肺泡上皮细胞中 P2X7R 的敲低也导致 PKC-β1 的丢失。这些数据表明，在肺泡上皮细胞中检测到的 P2X7R 对 PKC-β1 表达的影响在动物模型中也起作用。BLM 诱导的小鼠模型纤维化肺组织的免疫组织化学评价显示 PKC-β1 免疫反应性明显增加。本实验表明，P2X7R 的表达增加会影响 PKC-β1。我们预测，Ca(2+) 浓度的增加会刺激 PKC-β1，而 P2X7R 增加后激活 PKC-β1 的前提仍有待确定。我们的研究结果表明，PKC-β1 在肺纤维化的发病机制中很重要。
