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• Testicular inflammation induced by aging is identified as a potential risk factor for late-onset hypogonadism. • TNFAIP3, an anti-inflammatory factor, demonstrates close involvement in various aging and inflammatory diseases. • Our study reveals TNFAIP3 promotes testosterone production and suppresses apoptosis in Leydig cells within the aging inflammatory microenvironment. The aging inflammatory microenvironment surrounding Leydig cells is linked to reduced testosterone levels in males. Tumor necrosis factor alpha-induced protein 3 (TNFAIP3) acts as a critical anti-inflammatory factor in various aging-related diseases. This study aims to investigate the protective effect of TNFAIP3 on testosterone production in Leydig cells under an aging inflammatory microenvironment. Bioinformatics analysis examined TNFAIP3 expression differences in aging rat testes and validated the findings in aging mouse testes. In vitro models of inflammation were established using two Leydig cell lines, with tumor necrosis factor alpha (TNF-α) as the inflammatory factor. Lentiviral transduction was utilized to manipulate TNFAIP3 expression in these cell lines. Transcriptomic sequencing identified differentially expressed genes in TNFAIP3-overexpressing cells. Bioinformatics analysis and validation experiments revealed increased inflammatory signaling and elevated TNFAIP3 expression in aging rat and mouse testes. TNFAIP3 knockdown worsened testosterone synthesis inhibition and apoptosis in cells, while TNFAIP3 overexpression reversed these effects. Transcriptome analysis identified alterations in the P38MAPK pathway following TNFAIP3 overexpression. TNFAIP3 knockdown enhanced TNF-induced P38MAPK signaling, whereas its overexpression attenuated this effect. TNFAIP3 was found to regulate testosterone synthesis by upregulating CEBPB expression. TNFAIP3 exhibits inhibitory effects on apoptosis and promotes testosterone production in Leydig cells. The protective influence of TNFAIP3 on Leydig cells within an inflammatory microenvironment is likely mediated through by inhibiting the P38MAPK pathway and upregulating CEBPB expression. [ABSTRACT FROM AUTHOR]

The mechanism by which spinal cord injury (SCI) induces neuronal death has not been thoroughly understood. Investigation on the molecular signal pathways involved in SCI-mediated neuronal apoptosis is important for development of new therapeutics for SCI. In the current study, we explore the role of heme oxygenase-1 (HO-1) in the modulation of mixed lineage kinase 3/mitogen-activated protein kinase kinase/cJUN N-terminal kinase 3 (MLK3/MKK7/JNK3) signaling, which is a pro-apoptotic pathway, after SCI. We found that MLK3/MKK7/JNK3 signaling was activated by SCI in a time-dependent manner, demonstrated by increase in activating phosphorylation of MLK3, MKK7, and JNK3. SCI also induced HO-1 expression. Administration of HO-1-expressing adeno-associated virus before SCI introduced expression of exogenous HO-1 in injured spinal cords. Exogenous HO-1 reduced phosphorylation of MLK3, MKK7, and JNK3. Consistent with its inhibitory effect on MLK3/MKK7/JNK3 signaling, exogenous HO-1 decreased SCI-induced neuronal apoptosis and improved neurological score. Further, we found that exogenous HO-1 inhibited expression of cell division cycle 42 (Cdc42), which is crucial for MLK3 activation. In vitro experiments indicated that Cdc42 was essential for neuronal apoptosis, while transduction of neurons with HO-1-expressing adeno-associated virus significantly reduced neuronal apoptosis to enhance neuronal survival. Therefore, our study disclosed a novel mechanism by which HO-1 exerted its neuroprotective efficacy. Our discovery might be valuable for developing a new therapeutic approach for SCI. [ABSTRACT FROM AUTHOR]