The role of NLRC4 inflammasome-mediated autophagy in a mouse model of sepsis-induced myocardial dysfunction

doi:10.12307/2023.472

Abstract

Abstract: BACKGROUND: Studies have found that inflammasome and autophagy play important roles in the immune function of patients with sepsis. However, the current research is only limited to exploring the characteristics of changes in a certain autophagy signaling pathway in sepsis, and the specific mechanism of autophagy regulation in sepsis-induced myocardial dysfunction has not been elucidated.
OBJECTIVE: To investigate the changes of NOD-like receptor family, pyrin domain-containing protein 4 (NLRC4) inflammasome and autophagy levels in a mouse model of sepsis-induced myocardial dysfunction, in order to provide a theoretical basis for the pathogenesis of sepsis-induced myocardial dysfunction.
METHODS: Forty-eight clean and healthy Kunming male mice were randomly divided into six groups, sham operation group (6, 12, 24 hours) and cecal ligation and puncture group (6, 12, 24 hours) with three groups each. In the cecal ligation and puncture group, the distal half of the cecum of the mice was ligated, and a 22-gauge needle was used for puncturing twice to squeeze out a little intestinal content. Except for ligation and perforation, other operations in the sham operation group were the same as those in the cecal ligation and perforation group. The general condition of the mice after operation was observed. Mouse cardiac function was measured by VINNO 7 6LAB ultrasound. Levels of tumor necrosis factor α and troponin T were detected by enzyme-linked immunosorbent assay. Pathological changes of the myocardium were observed by hematoxylin-eosin staining. Changes in myocardial mitochondria and autophagosome were observed under transmission electron microscope. Real-time quantitative polymerase chain reaction was used to detect the expression of NLRC4 mRNA in myocardial tissue, and western blot was used to detect the expression of inflammatory factors, LC3, Beclin-1, and NLRC4 protein in mouse myocardial tissue.
RESULTS AND CONCLUSION: Tumor necrosis factor α level, Beclin-1 level, and LC3-II/LC3-I ratio were significantly increased in mice at 6 hours after cecal ligation and puncture (P < 0.05). At 12 hours after cecal ligation and puncture, tumor necrosis factor α and troponin T levels in the peripheral blood of mice increased, left ventricular ejection fraction and left ventricular shortening rate decreased, and NLRC4 mRNA and protein levels, interleukin-1β level, interleukin-18 level, Beclin-1 level, and LC3-II/LC3-I ratio increased (P < 0.05). At 24 hours after cecal ligation and puncture, NLRC4 mRNA and protein levels increased progressively, interleukin-1β and interleukin-18 were highly expressed, and Beclin-1 level and LC3-II/LC3-I ratio significantly decreased (P < 0.05). Myocardial pathological changes detected by hematoxylin-eosin staining showed that the myocardial fibers were normal in the sham operation group and at 6 hours after cecal ligation and perforation, and there was no inflammatory cell infiltration. Myocardial fibroedema and inflammatory cell infiltration occurred at 12 hours after cecal ligation and perforation. With the prolongation of time, the arrangement of myocardial fibers was disordered, the infiltration of inflammatory cells was aggravated, and some myocardial fibers appeared to have degeneration and necrosis. Under the transmission electron microscope, in the cecal ligation and perforation group, myocardial mitochondria were swollen, and with the extension of time, the swelling was aggravated, vacuolar degeneration of mitochondria occurred, and the myofibrils were slightly dissolved. Autophagosomes appeared at 6 hours after cecal ligation and perforation and increased in number at 12 hours, but occasionally appeared at 24 hours. In conclusion, the mouse model of sepsis-induced myocardial dysfunction is successfully established at 12 hours after cecal ligation and perforation. Excessive activation of NLRC4 inflammasomes is involved in the pathogenesis of sepsis-induced myocardial dysfunction by down-regulating the expression of autophagy.

Key words: sepsis, sepsis-induced myocardial dysfunction, cecal ligation and perforation, immune disorder, NLRC4 inflammasome, autophagy

CLC Number:

Jiang Dajun, Jiang Wanwei, Zhou Ying, Tian Yong, Yang Guohui. The role of NLRC4 inflammasome-mediated autophagy in a mouse model of sepsis-induced myocardial dysfunction[J]. Chinese Journal of Tissue Engineering Research, 2023, 27(23): 3667-3673.

Figures/Tables 6

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