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Morphology and cellular differentiation markers in the human fetal testis
Yi Li, MD, Maya Overland, MD, Amber Derpinghaus, BS, Sena Aksel, MD, Mei Cao, MD, Nicholas Ladwig, MD, Gerald Cunha, MD, Laurence Baskin, MD.
UCSF, San Francisco, CA, USA.

BACKGROUND: A comprehensive immunohistochemical ontogeny of the developing human fetal testis has remained incomplete in the literature to date. Prior studies have been limited to specific markers or targets, or have not encompassed the broad range of development from fetal life into puberty.
METHODS: We collected human fetal testes from 8 to 21 weeks of fetal age, as well as postnatal human testes at minipuberty, pre-pubertal, and pubertal stages. Immunohistochemistry was performed with a comprehensive panel of antigens targeting germ cells, Leydig cells, Sertoli cells, peritubular myoid cells, and others.
RESULTS: Testicular cords developed from 8 to 14 weeks of fetal age, separating the testis into the interstitial and intracordal compartments. Macroscopic lightsheet microscopy reveals smooth muscle staining of testicular vessels and epididymal structures and Sertoli cell specific SOX9 staining of the spermatic cords (Figure 1). Fetal gonadocytes were localized within the cords and evaluated for Testis-Specific Protein Y, Octamer-binding transcription factor 4, Sal-like protein 4, and placental alkaline phosphatase expression. Fetal Sertoli cells were also localized in the cords and evaluated for SRY-box Transcription Factor 9, inhibin, and anti-Mullerian hormone expression. Fetal Leydig cells were present in the interstitium and stained for cytochrome p450c17 and calretinin, while interstitial peritubular myoid cells were examined using smooth muscle α-actin. Androgen receptor expression was localized close to the testicular medulla at 8 weeks and then around the testicular cords in the interstitium as they matured in structure (Figure 2). Postnatal staining showed that Testis-Specific Protein Y remained positive of male gonadocytes throughout adulthood. Anti-Mullerian hormone, SRY-box Transcription Factor 9, and Steroidogenic factor 1 are expressed by the postnatal Sertoli cells at all ages examined. Leydig cell markers cytochrome p450c17 and calretinin are expressed during mini-puberty and puberty, but not during the pre-pubertal period (Figure 2). Smooth muscle α-actin and androgen receptor were not expressed during mini-puberty or pre-puberty.
CONCLUSIONS: The ontogenic map of the human fetal and postnatal testicular structure and expression patterns described here will serve as a reference for future investigations into normal and abnormal testicular development.



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