Septic serum mediates inflammatory injury in human umbilical vein endothelial cells via reactive oxygen species, mitogen activated protein kinases and nuclear factor‑κB

Abstract

Sepsis-induced blood vessel dysfunction is mainly caused by microvascular endothelial cell injury. However, the mechanism underlying sepsis-induced endothelial cell injury remains unclear. The present study hypothesized that sepsis-induced inflammatory injury of endothelial cells may be the first step of endothelial barrier dysfunction. Therefore, the present study aimed to uncover the mechanism underlying the inflammatory effects of sepsis. A rat model of cecal ligation and puncture-induced sepsis was established, and septic serum was collected. Subsequently, human umbilical vein endothelial cells (HUVECs) were treated with the isolated septic or normal serum. HUVEC viability was assessed using a Cell Count Kit-8 assay. Furthermore, transmission electron microscopy and reverse transcription-quantitative PCR (RT-qPCR) analysis were carried out to observe the cell morphology and determine the mRNA expression levels in septic serum-induced HUVECs. The protein expression levels were evaluated by western blot analysis, and the secretion of the inflammatory factors interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α was determined by ELISA. Additionally, reactive oxygen species (ROS) generation and nuclear factor (NF)-κB nuclear translocation were observed under a fluorescence microscope. The results of the present study demonstrated that HUVEC viability was significantly decreased following 12- or 24-h treatment with septic serum. In addition, chromatin condensation, mitochondrial vacuolization and endoplasmic reticulum degranulation were observed following treatment with septic serum. Furthermore, the secretion levels of IL-1β, IL-6 and TNF-α were increased in septic serum-stimulated HUVECs. Septic serum treatment also enhanced superoxide anion generation, promoted extra-cellular signal regulated kinase 1/2 (ERK1/2), N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38) phosphorylation, and increased NF-κB levels in the nuclei of HUVECs. Finally, pre-treatment of HUVECs with the antioxidant N-acetylcysteine, the ERK1/2 inhibitor PD98059, the p38 inhibitor SB203580, the JNK inhibitor SP610025 or the NF-κB inhibitor pyrrolidine dithiocarbamate restored the septic serum-induced IL-1β, IL-6 and TNF-α expression. In conclusion, the results of the current study suggested that the septic serum-induced endothelial cell injury may be mediated by increasing ROS generation, activation of mitogen-activated protein kinases and NF-κB translocation.

ADAM10-a “multitasker” in sepsis: focus on its posttranslational target

“…On the basis of their apparent molecular weights, both two ADAM10 forms, both proADAM10 and mature ADAM10 dimers were presence at the cell surface [ 27 ]. Septic serum mediated vascular endothelial injury by changing redox conditions, promoting ROS production and MAPK and NF-κB activity [ 98 ]. Increased intracellular Ca 2+ concentration allowing the ADAM10 prodomain to be available for cleavage by furin [ 92 ], as well as VEGF/Erk-induced shedding activity of ADAM10 [ 99 ], which provides the possibility that the ADAM10 prevalent in sepsis is also readily activated (Fig.…”