TNF-α inhibits SCF, ghrelin, and substance P expressions through the NF-κB pathway activation in interstitial cells of Cajal

Ulcerative colitis is a chronic inflammatory disease of the colon where intestinal motility is disturbed. Interstitial cells of Cajal (ICC) are required to maintain normal intestinal motility. In the present study, we assessed the effect of tumor necrosis factor-alpha (TNF-α) on viability and apoptosis of ICC, as well as on the expression of stem cell factor (SCF), ghrelin, and substance P. ICC were derived from the small intestines of Swiss albino mice. Cell viability and apoptosis were measured using CCK-8 assay and flow cytometry, respectively. ELISA was used to measure the concentrations of IL-1β, IL-6, ghrelin, substance P, and endothelin-1. Quantitative RT-PCR was used to measure the expression of SCF. Western blotting was used to measure the expression of apoptosis-related proteins, interleukins, SCF, and NF-κB signaling pathway proteins. TNF-α induced inflammatory injury in ICC by decreasing cell viability and increasing apoptosis and levels of IL-1β and IL-6. TNF-α decreased the levels of SCF, ghrelin, and substance P, but had no effect on endothelin-1. TNF-α down-regulated expressions of SCF, ghrelin, and substance P by activating the NF-κB pathway in ICC. In conclusion, TNF-α down-regulated the expressions of SCF, ghrelin, and substance P via the activation of the NF-κB pathway in ICC.

溃疡性结肠炎（Ulcerative Colitis）是一种以肠道动力紊乱为特征的慢性结肠炎症性疾病。卡哈尔间质细胞（Interstitial Cells of Cajal, ICC）是维持正常肠道动力所必需的细胞群。本研究旨在评估肿瘤坏死因子-α（Tumor Necrosis Factor-alpha, TNF-α）对卡哈尔间质细胞（ICC）活力与凋亡的影响，以及其对干细胞因子（Stem Cell Factor, SCF）、饥饿素（ghrelin）和P物质（Substance P）表达的调控作用。本研究所用ICC均从瑞士白化小鼠的小肠组织中分离获取。分别采用CCK-8检测法与流式细胞术测定细胞活力与凋亡水平；采用酶联免疫吸附测定（Enzyme-Linked Immunosorbent Assay, ELISA）检测白细胞介素-1β（IL-1β）、白细胞介素-6（IL-6）、饥饿素、P物质与内皮素-1的浓度水平；采用定量逆转录聚合酶链反应（qRT-PCR）检测干细胞因子（SCF）的基因表达水平；采用蛋白质免疫印迹（Western Blotting）检测凋亡相关蛋白、白细胞介素、干细胞因子（SCF）以及核因子-κB（NF-κB）信号通路相关蛋白的表达水平。肿瘤坏死因子-α（TNF-α）可通过降低ICC活力、促进细胞凋亡并升高IL-1β与IL-6的表达水平，诱导ICC发生炎症损伤；TNF-α可下调SCF、饥饿素与P物质的表达水平，但对内皮素-1的表达无显著影响；TNF-α可通过激活ICC中的NF-κB信号通路，下调SCF、饥饿素与P物质的基因表达。综上，肿瘤坏死因子-α（TNF-α）可通过激活ICC中的NF-κB信号通路，下调SCF、饥饿素与P物质的表达。