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A Seminiferous Tubule Squash Technique for the Cytological Analysis of Spermatogenesis Using the Mouse Model
Published on: February 6, 2018
5-ALA/Fe2+ attenuates testicular inflammatory injury and spermatogenic failure in non-obstructive azoospermia mice by
Jiewen Zhu1,2, Jing Zhai3, Zhanxin Xu1,2
1Department of Urology, Shanghai Fifth People's Hospital, Fudan University, Shanghai, China.
Background:
Non-obstructive azoospermia (NOA) remains a major challenge in the management of male infertility, with currently limited effective therapeutic options available. The study investigates the therapeutic potential and underlying mechanism of 5-aminolevulinic acid with ferrous iron (5-ALA/Fe2+) for NOA.
Methods:
A busulfan-induced NOA mouse model was established and treated with low or high-dose 5-ALA/Fe2+. Therapeutic effects were assessed via testicular histopathology, hormone assays, and evaluation of apoptosis. The mechanism was systematically explored through integrated network pharmacology, proteomic analysis, and experimental validation using Western blot, qPCR, and flow cytometry.
Results:
5-ALA/Fe2+ administration significantly ameliorated testicular atrophy, restored spermatogenic architecture, and improved hormonal profiles in NOA mice. It notably upregulated Sertoli cell-specific markers (WT1, GATA4, SOX9) and suppressed testicular cell apoptosis. Integrated analysis identified the TP53-CASP3 axis as a key downstream node. Both bioinformatics predictions and experimental validation confirmed that 5-ALA/Fe2+ exerts its protective effect by inhibiting the TP53-CASP3 pathway, evidenced by reduced TP53 and cleaved Caspase-3 levels, decreased Bax/Bcl-2 ratio, and improved mitochondrial membrane potential and cell viability.
Conclusion:
Our findings demonstrate that 5-ALA/Fe2+ is a promising therapeutic agent for NOA, capable of mitigating testicular damage and restoring spermatogenic function. The protective effect is primarily mediated through the targeted suppression of the TP53-CASP3 apoptotic signaling pathway.
