5-ALA/Fe²⁺ attenuates testicular inflammatory injury and spermatogenic failure in non-obstructive azoospermia mice by suppressing the TP53-CASP3 apoptotic cascade.

Authors

Jiewen Zhu, Jing Zhai, Zhanxin Xu, Yifan Wang, Jinle Xie, Yangyun Wang, Jiangang Hou, Ke Xu

Published in

Frontiers in cell and developmental biology. Volume 14. Pages 1884004. Epub Jul 16, 2026.

Abstract

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.
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.
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.
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.

PMID:
42534744
Bibliographic data and abstract were imported from PubMed on 31 Jul 2026.

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