Leveraging Menstrual Effluent to Investigate Vitamin D and Human Decidualization: A Potential Link with Fertility

CONTEXT: Low vitamin D levels have been associated with female infertility, but the underlying mechanisms are not understood. OBJECTIVES: To examine vitamin D treatment, in vitro and in vivo, on human endometrial stromal cell (eSC) decidualization, including mediating pathways and to assess the feasibility of vitamin D measurement in menstrual effluent (ME). DESIGN: ME, ME-derived endometrial stromal cells, and peripheral plasma were collected from participants in the Research Outsmarts Endometriosis (ROSE) study (2017-2024) and the Investigation of Vitamin D and Menstrual Cycles trial (inVitD) (2022-2024; NCT05050916). SETTING: ROSE study: North America; inVitD study: Detroit, Michigan, and Durham, North Carolina. PARTICIPANTS: Healthy females aged 19 to 40 years. The inVitD trial participants provided ME pre- and post-cholecalciferol supplementation. INTERVENTION: Oral cholecalciferol supplementation (50 000 IU/week or 4200 IU/week × 3 months) was provided to vitamin D deficient inVitD trial participants. MAIN OUTCOME MEASURE: First, we examined the effect of active vitamin D (calcitriol) on ME-eSC decidualization in vitro. Second, we compared ME-eSC decidualization pre- and post-oral cholecalciferol supplementation. Mechanistic studies investigated target signaling proteins and mRNA expression. Third, we compared levels of vitamin D biomarkers in peripheral plasma vs ME. RESULTS: Calcitriol increased eSC decidualization measured by insulin-like growth factor-binding protein 1 (P < .0001) and prolactin levels (P < .0001). Calcitriol and decidualization alone induced VDR mRNA expression. Calcitriol reduced protein kinase B (AKT) and proline-rich AKT substrate of 40 kDa phosphorylation during decidualization. Decidualization capacity increased following in vivo cholecalciferol supplementation (P = .07). Peripheral plasma 1,25-dihydroxyvitamin D levels were correlated with ME levels (r = 0.66, P = .01). CONCLUSION: Vitamin D enhances eSC decidualization, likely through reduced AKT signaling and downstream events, highlighting its potential role in fertility. Vitamin D biomarkers were measurable in ME.