It has been well-established that T cells function in inducing immune regulation, immune responses and inflammation in the postnatal testis, but their function and frequency during fetal testis development is undocumented in mouse, rat and human. To study T cells during mammalian testis development, we explored their frequency and localisation in the testis and their contacts with other cells in C57BL6J mice at embryonic day (E) E13.5, E15.5, and at birth (PND0).
PFA-fixed OCT-embedded testes (n=4-7 mice/age) were completely sectioned (6 µM). Four central sections spaced >15 µM were examined using immunofluorescence to detect T cells (CD3), macrophages (F4/80), germ cells (DDX4) and cord basement membrane (laminin). Cell position was designated as in the section perimeter (between capsule and cords) or internal (inside cords or interstitium). Levels of transcripts encoding CD3, F4/80, CD45 (pan-immune cell marker) and DDX4 were measured by RT-qPCR (n=3).
Our study revealed a significant increase in the T cell number from E13.5 to PND0. T cells were absent in the E13.5 testis, rare at E15.5 (0-1/ section), and frequent at PND0 (15±5/ section). RT-qPCR results validated the immunofluorescence data, as Cd3 was mostly undetectable at E13.5, but increased 2.5-fold from E15.5 to PND0. T cells were detected in the perimeter area, interstitium, and cord perimeter positions. Moreover, the co-localization of T cells (one and/or two) and macrophages appearing as a cluster was a frequent observation at PND0 (2 to 4 clusters/ section).
This study documents for the first time the changing frequency and localisation of T cells during testis development in the fetal and newborn mice and identifies cellular contacts between T cells and macrophages. These results shed light on the presence of T cells and their interactions with macrophages during testis development.