ATP6V1A deletion leads to impaired clearance of septic cardiac resident macrophages

Abstract

Objective To investigate the clearance capacity of cardiac resident macrophages in post-sepsis and its underlying mechanism. Methods A mouse model of sepsis was established using cecum perforation ligation. Thirty male C57BL/6 mice (8 weeks old, weighing 20~25 g) were randomly and equally divided into a sham operation group (sham group) and a model group (sepsis group). Immunofluorescence assay was employed to label the cardiomyocytes and macrophages to observe the apoptosis of cardiomyocytes and the phagocytosis by cardiac resident macrophages. Cardiac resident macrophages were extracted for transcriptomic sequencing to determine the functional changes of the cells after sepsis. Cardiac resident macrophage cell lines were established at the cellular level and served as the normal group (RAC group), and the RAC cells treated with LPS were subjected as the sepsis group (RAC+LPS group). Then the differences in the ability to clear apoptotic cardiomyocytes between the 2 groups were observed. Then DQ-BSA-RED lysosomal activity detection probe, Lyso-Sensor yellow/bule dye, ELISA, and Western blotting were applied to detect the lysosomal function of cardiac resident macrophages, activity and expression of important lysosomal hydrolases, changes in contents and related subunits of vacuolar-type adenosine triphosphatases (V-ATPase). Results Compared with the sham group, the sepsis group had larger number of apoptotic cardiomyocytes (P < 0.05) and increased phagocytosis of cardiomyocytes by cardiac macrophages (P < 0.05). The results of transcriptomic sequencing revealed a significant dysfunction of lysosome-associated functions of cardiac-resident macrophages after sepsis. In in vitro experiments, the RAC+LPS group had a reduced fragmentation capacity of apoptotic cardiomyocytes, reduction in the intensity of yellow fluorescence of lysosomes (P < 0.05), and decrease in lysosomal hydrolase activity (P < 0.05) when compared with the RAC group. In addition, LPS treatment significantly decreased the activity and expression of V-ATPase and its major subunit ATP6V1A in cardiac resident macrophages (P < 0.05). Conclusion Cardiac resident macrophages show reduced clearance of apoptotic cardiomyocytes after sepsis, which may be related to a decrease in the activity of ATP6V1A, an important subunit of its lysosomal V-ATPase, and reduced activity of lysosomal hydrolases. [Key words] cardiac dysfunction in sepsis , cardiac resident macrophages , lysosome, vacuolar-type adenosine triphosphatases ,

Keywords

• cardiac dysfunction in sepsis
• cardiac resident macrophages
• lysosome
• vacuolar-type adenosine triphosphatases