ATP6V1A deletion leads to impaired clearance of septic cardiac resident macrophages

Shunxin YANG; Yuanqun ZHOU; Xinming XIANG; Liangming LIU; Tao LI; Yi HU

doi:10.16016/j.2097-0927.202406060

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ATP6V1A deletion leads to impaired clearance of septic cardiac resident macrophages

Basic Medicine | 更新时间：2026-01-09

- ATP6V1A deletion leads to impaired clearance of septic cardiac resident macrophages
- Journal of Army Medical University Vol. 46, Issue 23, Pages: 2599-2607(2024)
- 作者机构：
  
  1.400042 重庆，陆军特色医学中心：麻醉科
  2.400042 重庆，野战外科研究部战伤休克与输血研究室，创伤与化学中毒全国重点实验室
- 作者简介：
- 基金信息：
  
  the General Program of National Natural Science Foundation of China(82270523)
- DOI：10.16016/j.2097-0927.202406060
  CLC： R322.11;R345;R631.1
- Received：18 June 2024，
  
  Revised：2024-10-16，
  
  Published：15 December 2024
- 稿件说明：
移动端阅览
Shunxin YANG, Yuanqun ZHOU, Xinming XIANG, et al. ATP6V1A deletion leads to impaired clearance of septic cardiac resident macrophages[J]. Journal of Army Medical University, 2024, 46(23): 2599-2607.
DOI：

Shunxin YANG, Yuanqun ZHOU, Xinming XIANG, et al. ATP6V1A deletion leads to impaired clearance of septic cardiac resident macrophages[J]. Journal of Army Medical University, 2024, 46(23): 2599-2607. DOI： 10.16016/j.2097-0927.202406060.

摘要

目的

探讨脓毒症后心脏驻留巨噬细胞的清除能力及其机制。

方法

利用盲肠穿孔结扎术建立脓毒症小鼠模型，将30只雄性C57BL/6小鼠(8周龄，体质量20~25 g)按随机数字抽样法分为假手术组(Sham组，

=15)和脓毒症模型组(Sepsis组，

=15)。使用免疫荧光技术标记心肌细胞与巨噬细胞观察小鼠心肌细胞凋亡及心脏驻留巨噬细胞对心肌细胞的吞噬情况。提取心脏驻留巨噬细胞进行转录组学测序，观察脓毒症后心脏驻留巨噬细胞的功能变化。在细胞水平建立心脏驻留巨噬细胞系，即正常组(RAC组)；用LPS处理RAC组细胞后，即脓毒症组(RAC+LPS组)。观察2组间对凋亡心肌细胞清除能力的差异；使用DQ-BSA-RED溶酶体活性检测探针、Lyso-Sensor yellow/bule染料

、ELISA以及蛋白质印迹实验法检测心脏驻留巨噬细胞溶酶体功能、溶酶体重要水解酶活性及蛋白表达、囊泡型腺苷三磷酸酶(vacuolar-type adenosine triphosphatases，V-ATPase)含量及其相关亚基的改变。

结果

与Sham组相比，Sepsis组小鼠心肌细胞凋亡数量增加(

＜0.05)，心脏巨噬细胞对心肌细胞的吞噬增加(

＜0.05)，转录组学结果分析发现脓毒症后心脏驻留巨噬细胞的溶酶体相关功能显著失调。体外实验结果显示，与RAC组相比，RAC+LPS组对凋亡心肌细胞的碎片化能力降低，溶酶体的黄色荧光强度明显减弱(

＜0.05)，溶酶体水解酶活性降低(

＜0.05)。此外，LPS处理显著降低了心脏驻留巨噬细胞V-ATPase及其主要亚基ATP6V1A的活性和表达(

＜0.05)。

结论

脓毒症后心脏驻留巨噬细胞对凋亡心肌细胞的清除能力降低，其机制可能与其溶酶体V-ATPase重要亚基ATP6V1A和溶酶体水解酶活性降低有关。

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 (

0.05) and increased phagocytosis of cardiomyocytes by cardiac macrophages (

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 (

0.05)

and decrease in lysosomal hydrolase activity (

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 (

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.

关键词

Keywords

references

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