(OR02-07-23) Exploring the Impact of Maternal Vitamin D Status on the Proteomic Composition of Human Milk

Objectives: Investigate the potential association between maternal vitD status during lactation and variations in the proteomic profile of human milk.

Methods: In an exploratory clinical trial, 10 women were randomized to receive a daily multivitamin containing 400 IU vitD₃ and either placebo (0 IU vitD₃) or 6000 IU D₃ gummies (Church & Dwight, Ewing, NJ) starting at 1-month post-partum until 4 months. The study team and participants were blinded to treatment. Monthly blood samples were analyzed for total circulating 25(OH)D, used as an indicator of maternal vitD status during lactation, with a target threshold of 100 nmol/L. Serum calcium and urinary calcium/creatinine ratios were monitored as safety measures. Milk samples were collected from the left breast after cleansing and until fully emptied, and whole milk samples were frozen at -20^oC until processed. Protein identification and quantification were performed using LC-MS/MS (Orbitrap Lumos MS) and proteins were identified, quantified, and normalized using MaxQuant software.

Results: 1,250 proteins were identified and the data filtered to retain proteins quantified in >70% of milk samples. To investigate the association between maternal vitD status and human milk proteome, proteins were further filtered based on their intensities with high (≥ 100 nmoL/L) or low ( < 50 nmoL/L) plasma 25(OH)D concentrations using Student's t-test with permutation-based correction for multiple hypothesis testing. With a threshold FDR < 0.05, 32 proteins were increased with higher 25(OH)D concentrations, including 3 proteins with Vitamin D Binding Protein (VDBP) transcription factor binding sites: SerpinAl, GANAS, HAPLNl identified by Toppgene (ToppFun). In milk samples from mothers with high 25(OH)D, there were 77 proteins that were decreased and corresponded to innate and metabolic immune pathways. Further statistical evaluation across the continuum of 25(OH)D concentration is ongoing.

Conclusions: Our findings suggest that maternal vitD status has a differential impact on the human milk proteome profile, with distinct patterns observed in the milk of women with low vs higher total circulating 25(OH)D concentrations.

Funding Sources: Funded in part by The Stone Foundation, MUSC; Digestive Diseases Center, MUSC; SCTR) Institute, NIH/NCAT Grant number UL1 TR000062.