Department of Anesthesiology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.
J Trauma Acute Care Surg. 2012 Jul;73(1):67-72. doi: 10.1097/TA.0b013e318256a0fe.
Gap junctions regulate proper kidney function by facilitating intercellular communication, vascular conduction, and tubular purinergic signaling. However, no clear relationship has been described between gap-junction function and acute kidney injury induced by the endotoxin lipopolysaccharide (LPS).
Normal rat kidney epithelial cells (NRK52E cells) were seeded at high and low densities to promote or impede gap-junction formation, respectively, and establish distinctive levels of intercellular communication in culture. Cells were then challenged with LPS at various concentrations (10-1,000 ng/mL). LPS-induced formation and function of gap junctions were assessed by measuring changes in cell proliferation and colony-forming rates, fluorescent dye transmission to adjacent cells, expression levels of connexin43, and repositioning of confluent cells in response to the gap junction inhibitor oleamide or agonist retinoic acid.
The cell proliferation rate and colony-forming rate of high- and low-density NRK52E cells were decreased upon LPS challenge, in a dose-dependent manner. The colony-forming rate of confluent high-density cells was significantly lower than that of low-density cells. Oleamide treatment raised the LPS-induced colony-forming rate of high-density cells, whereas retinoic acid decreased the rate. Neither oleamide nor retinoic acid significantly affected the LPS-induced colony-forming rate of low-density cells. Fluorescence transmission of high-density cells was reduced by LPS challenge, in a dose-dependent manner, but inclusion of retinoic acid increased the LPS-induced transmission of fluorescence. LPS challenge of either high- or low-density NRK52E cells resulted in down-regulated connexin43 expression.
Gap-junction function plays an important role in concentration-dependent cytotoxic effect of LPS on normal rat kidney cells in vitro.
缝隙连接通过促进细胞间通讯、血管传导和管状嘌呤能信号传导来调节正常的肾脏功能。然而，在由内毒素脂多糖 (LPS) 引起的急性肾损伤与缝隙连接功能之间尚未描述明确的关系。
将正常大鼠肾上皮细胞 (NRK52E 细胞) 分别以高和低密度接种，以促进或阻碍缝隙连接的形成，并在培养中建立不同水平的细胞间通讯。然后用不同浓度 (10-1,000ng/mL) 的 LPS 对细胞进行刺激。通过测量细胞增殖和集落形成率的变化、荧光染料向相邻细胞的传递、连接蛋白 43 的表达水平以及缝隙连接抑制剂油酰胺或激动剂维甲酸对细胞重新定位来评估 LPS 诱导的缝隙连接的形成和功能。
LPS 刺激后，高和低密度 NRK52E 细胞的细胞增殖率和集落形成率呈剂量依赖性下降。高密细胞的集落形成率明显低于低密度细胞。油酰胺处理可提高高密细胞 LPS 诱导的集落形成率，而维甲酸则降低该率。油酰胺或维甲酸均未显著影响低密度细胞 LPS 诱导的集落形成率。LPS 刺激以剂量依赖性方式降低高密细胞的荧光传递，但维甲酸可增加 LPS 诱导的荧光传递。LPS 刺激高密或低密度 NRK52E 细胞均可导致连接蛋白 43 的表达下调。
缝隙连接功能在 LPS 对体外正常大鼠肾细胞的浓度依赖性细胞毒性作用中起重要作用。
