Germ cell quantification in human fetal and prepubertal testis tissues: a comparison of current methodologies

Area of Science:

• Reproductive Biology
• Developmental Biology
• Oncology

Background:

• Quantifying germ cells in human immature testicular tissue is vital for assessing chemotherapy's impact on fertility and optimizing fertility preservation strategies for prepubertal boys.
• Current quantification methods necessitate round tubules, which are scarce in immature human testicular tissue, particularly after in vitro culture.

Purpose of the Study:

• To evaluate an alternative germ cell quantification method that does not require round tubules.
• To compare the efficacy of this new method with established techniques in human fetal and prepubertal testicular tissues.

Main Methods:

• Human fetal and prepubertal testicular tissues were utilized, including samples exposed in vitro to chemotherapy agents (cisplatin, carboplatin) or controls.
• Immunofluorescence staining identified germ cells (AP2γ for gonocytes, MAGE-A4 for (pre)spermatogonia).
• Germ cells were quantified using tubular germ cell density (Method 1) and compared to three established methods requiring round tubules (S/T, fertility index, round tubular germ cell density).

Main Results:

• Tubular germ cell density (Method 1) demonstrated strong and significant correlations with all three established methods across both fetal (r = 0.752–0.863) and prepubertal (r = 0.802–0.914) tissues.
• Method 1 is effective regardless of tubule shape, potentially increasing the number of quantifiable germ cells.

Conclusions:

• The tubular germ cell density method provides reliable germ cell quantification in human immature testicular tissues, even when round tubules are limited.
• This method is valuable for fertility preservation research and clinical applications involving limited tissue samples or tissues with few round tubules.