Effects of aging and macrophages on mice stem Leydig cell proliferation and differentiation in vitro

Jingjing Shao; Jiexia Wang; Xin Wen; Jiajia Xie; Fu Huang; Xiaoju Guan; Xinrui Hao; Ping Duan; Congde Chen; Haolin Chen

doi:10.3389/fendo.2023.1139281

Effects of aging and macrophages on mice stem Leydig cell proliferation and differentiation in vitro

Front Endocrinol (Lausanne). 2023 Mar 27:14:1139281. doi: 10.3389/fendo.2023.1139281. eCollection 2023.

Authors

Jingjing Shao^{1

2}, Jiexia Wang³, Xin Wen³, Jiajia Xie⁴, Fu Huang³, Xiaoju Guan², Xinrui Hao¹, Ping Duan³, Congde Chen², Haolin Chen^{1

2

3

4}

Affiliations

¹ Zhejiang Provincial Key Laboratory of Anesthesiology, Department of Anesthesiology, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, China.
² Key Laboratory of Children Genitourinary Diseases of Wenzhou City, Department of Pediatric Urology, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, China.
³ Department of Gynecology and Obstetrics, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, China.
⁴ Department of Pharmacology, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, China.

Abstract

Background: Testosterone plays a critical role in maintaining reproductive functions and well-beings of the males. Adult testicular Leydig cells (LCs) produce testosterone and are generated from stem Leydig cells (SLCs) during puberty through adulthood. In addition, macrophages are critical in the SLC regulatory niche for normal testicular function. Age-related reduction in serum testosterone contributes to a number of metabolic and quality-of-life changes in males, as well as age-related changes in immunological functions. How aging and testicular macrophages may affect SLC function is still unclear.

Methods: SLCs and macrophages were purified from adult and aged mice via FACS using CD51 as a marker protein. The sorted cells were first characterized and then co-cultured in vitro to examine how aging and macrophages may affect SLC proliferation and differentiation. To elucidate specific aging effects on both cell types, co-culture of sorted SLCs and macrophages were also carried out across two ages.

Results: CD51+ (weakly positive) and CD51++ (strongly positive) cells expressed typical SLC and macrophage markers, respectively. However, with aging, both cell types increased expression of multiple cytokine genes, such as IL-1b, IL-6 and IL-8. Moreover, old CD51+ SLCs reduced their proliferation and differentiation, with a more significant reduction in differentiation (2X) than proliferation (30%). Age matched CD51++ macrophages inhibited CD51+ SLC development, with a more significant reduction in old cells (60%) than young (40%). Crossed-age co-culture experiments indicated that the age of CD51+ SLCs plays a more significant role in determining age-related inhibitory effects. In LC lineage formation, CD51+ SLC had both reduced LC lineage markers and increased myoid cell lineage markers, suggesting an age-related lineage shift for SLCs.

Conclusion: The results suggest that aging affected both SLC function and their regulatory niche cell, macrophages.

Keywords: CD51; aging; macrophages; stem Leydig cells; testis; testosterone.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Aging
Animals
Cell Differentiation
Cell Proliferation
Macrophages / metabolism
Male
Mice
Sexual Maturation*
Testosterone* / metabolism

Substances

Testosterone

Grants and funding

This work was supported by grants from National Natural Science Foundation of China (91949123, HC and 82071626, PD), Natural Science Foundation of Zhejiang Province, CN (Z23H040003, HC and Q23H040007, XG) and Wenzhou Major Scientific and Technological Innovation Project, CN (ZY2019002, HC). These sources did not influence the study design, the collection, analysis or interpretation of data, or the writing of the paper.