Background Gut ischemia and hypoxia post severe burn leads to breakdown of intestinal epithelial barrier and enteric bacterial translocation (EBT), resulting in serious complications, such as systemic inflammatory response syndrome, sepsis and multiple organ failure. Cystic fibrosis transmembrane conductance regulator (CFTR) is known to be downregulated by hypoxia and modulate junctional complexes, which are crucial structures maintaining the intestinal barrier. This study aimed to investigate whether CFTR plays a role in both regulating the intestinal barrier and mediating EBT post severe burn, as well as the signaling pathways involved in these processes. Methods An in vitro Caco-2 cell model subjected to hypoxic injury and an in vivo mouse model with a 30% total body surface area full-thickness dermal burn were established. DF 508 mice (mice with F508del CFTR gene mutation) were used as an in vivo model to further demonstrate the role of CFTR in maintaining normal intestinal barrier function. QRT-PCR, western blot, ELISA, TER assay and immunofluorescence staining were used to detect the expression and localization of CFTR and tight junction proteins, as well as the function of tight junctions. Results Our data indicated that, in Caco-2 cells, the hypoxia condition significantly reduced CFTR expression; activated extracellular signal-regulated kinase and nuclear factor-κB signaling; elevated secretion of inflammatory factors (tumor necrosis factor-α, interleukin-1β and interleukin-8); downregulated zonula occludens-1, occludin and E-cadherin expression; decreased transepithelial electrical resistance values; and led to a cellular mislocation of ZO-1. More importantly, knockdown of CFTR caused similar alterations. The upregulation of inflammatory factors and downregulation of tight junction proteins (ZO-1 and occludin) induced by knockdown of CFTR could be reversed by specific extracellular signal-regulated kinase or nuclear factor-κB inhibition. In support of the in vitro data, exuberant secretion of pro-inflammatory mediators and EBT was observed in the intestine of severely burnt mice in vivo. EBT occurred in DF508 mice (mice with the F508del CFTR gene mutation), accompanied by augmented tumor necrosis factor-α, interleukin-1β and interleukin-8 levels in the ileum compared to wildtype mice. In addition, vitamin D3 was shown to protect the intestinal epithelial barrier from hypoxic injury. Conclusions Collectively, the present study illustrated that CFTR and downstream signaling were critical in modulating the intestinal epithelial junction and EBT post severe burn.

中文翻译：

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严重烧伤后肠道细菌易位的分子机制：囊性纤维化跨膜电导调节剂的作用

背景严重烧伤后肠道缺血缺氧导致肠上皮屏障破坏和肠道细菌易位（EBT），导致严重并发症，如全身炎症反应综合征、败血症和多器官衰竭。已知囊性纤维化跨膜电导调节剂 (CFTR) 会被缺氧下调并调节连接复合物，而连接复合物是维持肠道屏障的关键结构。本研究旨在调查 CFTR 是否在调节肠道屏障和介导严重烧伤后 EBT 以及参与这些过程的信号通路中发挥作用。方法建立体外缺氧损伤Caco-2细胞模型和30%体表面积全层皮肤烧伤小鼠体内模型。DF 508 小鼠（F508del CFTR 基因突变小鼠）被用作体内模型，以进一步证明 CFTR 在维持正常肠道屏障功能中的作用。采用QRT-PCR、western blot、ELISA、TER法和免疫荧光染色检测CFTR和紧密连接蛋白的表达和定位，以及紧密连接的功能。结果 我们的数据表明，在 Caco-2 细胞中，缺氧条件显着降低了 CFTR 的表达；激活的细胞外信号调节激酶和核因子-κB 信号传导；炎症因子分泌增加（肿瘤坏死因子-α、白细胞介素-1β和白细胞介素-8）；下调 zonula occludens-1、occludin 和 E-cadherin 表达；降低跨上皮电阻值；并导致 ZO-1 的细胞错位。更重要的是，CFTR 的击倒引起了类似的改变。CFTR 敲低诱导的炎症因子上调和紧密连接蛋白（ZO-1 和 occludin）下调可以通过特异性细胞外信号调节激酶或核因子-κB 抑制来逆转。为了支持体外数据，在体内严重烧伤的小鼠的肠道中观察到促炎介质和 EBT 的旺盛分泌。EBT 发生在 DF508 小鼠（具有 F508del CFTR 基因突变的小鼠）中，与野生型小鼠相比，回肠中的肿瘤坏死因子-α、白细胞介素 1β 和白细胞介素 8 水平升高。此外，维生素 D3 被证明可以保护肠上皮屏障免受缺氧损伤。结论 总的来说，