Women and men showed differences in immune-cell composition, gene expression, and genetic regulation in a single-cell analysis of 1,267,758 peripheral blood mononuclear cells from 982 participants in the OneK1K cohort in Australia.
The study, published in The American Journal of Human Genetics, combined single-cell RNA sequencing with genotype data to classify immune cells into 30 transcriptionally distinct cell types and evaluate sex differences in immune-cell proportions, sex-biased gene expression, and genetic regulation at baseline. Researchers downsampled female participants in sex-stratified expression quantitative trait loci analyses to equalize statistical power between sexes.
Men had higher proportions of CD14-positive monocytes, dendritic cells, natural killer cells, proliferating natural killer cells, CD8-positive proliferating cells, effector memory T cells, and central memory T cells. Women had higher proportions of B cells, CD4-positive naive T cells, CD56-positive natural killer cells, and regulatory T cells.
Transcriptomic analyses identified 78 sex-differentially expressed genes across 24 immune-cell populations, including 65 female-biased genes and 16 male-biased genes. Only 16 of the 78 genes were located on sex chromosomes, suggesting that autosomal regulation may contribute substantially to immune sexual dimorphism.
Female-biased genes in several immune-cell populations were enriched for tumor necrosis factor alpha signaling and nuclear factor kappa-light-chain-enhancer of activated B-cell–related pathways. Female-biased expression in CD4-positive cytotoxic T lymphocytes was also enriched for interferon gamma response pathways. In contrast, male-biased genes were enriched for ribosomal and RNA-processing functions.
In genetic analyses, researchers identified 14,432 autosomal expression quantitative trait loci in joint analyses, along with more than 1,000 sex-specific expression quantitative trait loci following sex-stratified filtering and validation. Researchers also identified 51 sex-interacting expression quantitative trait loci, although the analysis was exploratory and used a less stringent false discovery rate threshold.
Several sex-biased regulatory associations involved genes or pathways previously linked to autoimmune disease biology. Researchers identified female-specific regulation of FCGR3A in natural killer cells and sex-interacting regulation of ITGB2 in CD14-positive monocytes. The investigators emphasized that these findings reflected genetic regulatory associations rather than functional validation or demonstrated disease mechanisms.
FCGR3A has previously been associated with systemic lupus erythematosus, and the rs2099684 variant identified in this study has also been linked to Takayasu arteritis in a Han Chinese population, a condition with a strong female predominance. By comparison, the ITGB2 pathway has been implicated in systemic lupus erythematosus and primary Sjögren syndrome, although the specific rs760462 variant identified in this study has no known clinical correlation. Researchers noted that the sex-interacting regulatory signal involving ITGB2 may help explain the female-biased expression observed in monocytes.
Researchers also reported fewer expression quantitative trait loci on sex chromosomes than on autosomes, with generally smaller effect sizes. Many X-chromosome regulatory effects appeared cell-type specific and may not have been detectable in prior bulk RNA-sequencing studies.
The findings should be interpreted as hypothesis-generating. The study analyzed unstimulated immune cells at baseline, and the observed effect sizes may underestimate sex differences that emerge during activated immune states or disease. In addition, the cohort was restricted to participants of European ancestry to maintain genetic homogeneity, which may limit generalizability across populations.
Researchers also acknowledged that hormone levels, menstrual-cycle timing, menopause status, parity, infections, stress, and environmental exposures were not directly measured. The study relied on single time-point sampling, which may have obscured dynamic immune variation related to circadian rhythms or hormonal fluctuations over time.
“These results highlight that genes that are sexually dimorphic at baseline could potentially vary in their response in immune disease between the sexes in a cell-type-specific way,” wrote lead study researcher Seyhan Yazar, of the Garvan Institute of Medical Research in Sydney, Australia, and colleagues.
Disclosures: The researchers reported no competing interests. The study was supported by Australian research fellowships and grants, including funding from the National Health and Medical Research Council, MS Australia, the Australian Research Council, and the Royal Hobart Hospital Research Foundation. Funders had no role in study design, data collection and analysis, publication decision, or manuscript preparation.
