RPS14 as an aging-related biomarker for early-onset alterations of testicular senescence-associated genes in spermatogenic dysfunction at childbearing age.

RPS14 was identified as an upregulated testicular senescence-associated gene (SAG) in testes of older men.

• Two single-cell RNA sequencing (scRNA-seq) datasets, three bulk microarray datasets, and testicular tissue from older mice, older males, and patients at childbearing age were employed.
• RPS14 was identified among testicular SAGs through comparison of aging and spermatogenic dysfunction expression profiles.
• The study used scRNA-seq analyses and immunofluorescence to characterize RPS14 expression and distribution in testicular tissue.

RPS14 was significantly upregulated in testes of young patients with spermatogenic dysfunction compared to those with full spermatogenesis.

• Patients studied were at childbearing age, representing a young cohort with spermatogenic dysfunction.
• The upregulation mirrored the pattern observed in older men's testes, suggesting early-onset senescence-like transcriptional changes.
• The finding was supported by both bulk microarray datasets and scRNA-seq analyses.

RPS14 showed significant correlation with Johnsen scores and follicle-stimulating hormone (FSH) levels in patients with spermatogenic dysfunction.

• Johnsen scores are a histological grading system used to assess spermatogenesis quality in testicular biopsies.
• FSH levels are a clinical endocrine marker of spermatogenic failure.
• The correlation with both histological and hormonal parameters suggests RPS14 reflects the degree of spermatogenic impairment.

RPS14 demonstrated potential predictive value for outcomes of sperm retrieval surgery.

• Sperm retrieval surgery is a clinical procedure used in azoospermic patients to obtain sperm for assisted reproduction.
• RPS14 expression levels were associated with sperm retrieval success, suggesting a diagnostic application.
• This finding positions RPS14 as a potential preoperative biomarker to guide surgical decision-making in male infertility.

RPS14 was found to be significantly correlated with testicular mast cells and involved in the testicular immune microenvironment.

• The analysis revealed a deep involvement of RPS14 in the testicular immune microenvironment.
• Significant correlation between RPS14 expression and testicular mast cell presence was identified.
• The immune microenvironment correlation suggests RPS14 may play a role in inflammation-related aspects of spermatogenic dysfunction.

scRNA-seq analyses and immunofluorescence demonstrated a partially similar expression pattern and distribution of RPS14 in testes of both older males and young males with spermatogenic dysfunction.

• The partially overlapping expression patterns between aging and spermatogenic dysfunction support the concept of early-onset testicular senescence.
• Both scRNA-seq and immunofluorescence methodologies were used to characterize cell-type-specific distribution of RPS14.
• The similarity in expression patterns between older males and young dysfunctional testes underpins RPS14 as an aging-related biomarker in the context of male infertility.

The ribosome pathway was identified as a potential core mechanism through which RPS14 regulates testicular cell function.

• RPS14 encodes a ribosomal protein (40S ribosomal protein S14), consistent with involvement in ribosome biogenesis and protein synthesis.
• Pathway analyses pointed to the ribosome pathway as the central mechanistic link between RPS14 and testicular cell dysfunction.
• This mechanistic insight suggests that ribosomal dysregulation may underlie senescence-associated spermatogenic failure.

Early-onset transcriptional phenotypes of senescence have been identified in spermatogenic dysfunctional testes of patients at childbearing age.

• The study contextualized its findings within a broader recognized phenomenon of premature or early-onset testicular senescence.
• Limited prior studies had reported specific biomarkers or functions of testicular SAGs in spermatogenic dysfunction of young men.
• The identification of SAG alterations in young patients highlights a gap between chronological age and biological/molecular aging in the testis.