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Updated: Sep 4, 2025
Using Ex Vivo Upright Droplet Cultures of Whole Fetal Organs to Study Developmental Processes during Mouse Organogenesis
Published on: October 21, 2015
The changing landscape of immune cells in the fetal mouse testis
Samira Hosseini1,2, Sarah C Moody1,2, Daniela Fietz3
1Centre for Reproductive Health, Hudson Institute of Medical Research, 27-31 Wright St, Clayton, VIC, 3168, Australia.
Insights
Immune cells, particularly macrophages and neutrophils, play a crucial role in fetal testis development and remodeling. Their density, distribution, and interactions with germ cells change significantly from embryonic day 13.5 to birth.
Area of Science:
- Reproductive biology and immunology
- Developmental biology
- Cellular and molecular biology
Background:
- Fetal testis development involves significant cellular remodeling and immune cell infiltration.
- Macrophages are known to facilitate normal cord formation and removal of misplaced cells in early fetal testes.
- Understanding the dynamic roles of various immune cells during testis organogenesis is crucial for reproductive health.
Purpose of the Study:
- To investigate the density, distribution, and cellular contacts of macrophages and other immune cells in the developing mouse fetal testis.
- To characterize the changes in immune cell populations from embryonic day 13.5 to post-natal day 0.
- To explore the origins and functional significance of immune cells in fetal testis morphogenesis.
Main Methods:
- Immunofluorescence and immunohistochemistry were used to identify and localize macrophages (F4/80, CD206, MHCII), T cells (CD3), and granulocytes/neutrophils (Ly6G).
- RT-qPCR was employed to analyze the expression of immune marker mRNAs.
- Histological quantification and cellular interaction analysis were performed on C57BL6J mouse testes across different embryonic and post-natal stages.
Main Results:
- F4/80+ macrophages were the most abundant immune cells, decreasing from 90% at E13.5 to 65% at PND0, with distinct phenotypes suggesting different origins.
- Ly6G+ granulocytes/neutrophils were scarce early but increased significantly by PND0, comprising half of immune cells.
- Immune cell populations, including macrophages and neutrophils, were found in close proximity to germ cells within testis cords, with distinct macrophage subtypes identified at different developmental stages.
Conclusions:
- The study reveals dynamic changes in immune cell populations, particularly macrophages and neutrophils, during fetal testis development.
- Distinct macrophage populations with different origins and markers are present at various stages of testis organogenesis.
- These findings provide new insights into immune cell-testis interactions and their functional roles in fetal testis morphogenesis, opening avenues for future research.
Abstract:
Fetal testis growth involves cell influx and extensive remodeling. Immediately after sex determination in mouse, macrophages enable normal cord formation and removal of inappropriately positioned cells. This study provides new information about macrophages and other immune cells after cord formation in fetal testes, including their density, distribution, and close cellular contacts. C57BL6J mouse testes from embryonic day (E) 13.5 to birth (post-natal day 0; PND0), were examined using immunofluorescence, immunohistochemistry, and RT-qPCR to identify macrophages (F4/80, CD206, MHCII), T cells (CD3), granulocytes/neutrophils (Ly6G), and germ cells (DDX4). F4/80+ cells were the most abundant, comprising 90% of CD45+ cells at E13.5 and declining to 65% at PND0. Changes in size, shape, and markers (CD206 and MHCII) documented during this interval align with the understanding that F4/80+ cells have different origins during embryonic life. CD3+ cells and F4/80-/MHCII+ were absent to rare until PND0. Ly6G+ cells were scarce at E13.5 but increased robustly by PND0 to represent half of the CD45+ cells. These immunofluorescence data were in accord with transcript analysis, which showed that immune marker mRNAs increased with testis age. F4/80+ and Ly6G+ cells were frequently inside cords adjacent to germ cells at E13.5 and E15.5. F4/80+ cells were often in clusters next to other immune cells. Macrophages inside cords at E13.5 and E15.5 (F4/80Hi/CD206+) were different from macrophages at PND0 (F4/80Dim/CD206-), indicating that they have distinct origins. This histological quantification coupled with transcript information identifies new cellular interactions for immune cells in fetal testis morphogenesis, and highlights new avenues for studies of their functional significance.
