INTRODUCTION:Human milk oligosaccharides (HMOs) arise from tightly regulated glycosylation pathways and exert pleiotropic effects on infant immune, microbial, and neurodevelopmental trajectories. Vitamin D influences gene transcription and cellular metabolism, yet its potential role in mammary glycosylation remains unexplored. We examined whether maternal vitamin D status during lactation is associated with HMO composition and whether effects vary by maternal secretor phenotype. METHODS:We performed a post hoc analysis of two clinical studies: the NICHD vitamin D lactation randomized controlled trial (2005-2012) and the Lactation Immune Trial pilot (n = 88). Serum 25-hydroxyvitamin D [25(OH)D, vitamin D status] and 19 HMOs concentrations were measured at 1 and 4 months postpartum. Secretor status was defined by presence of 2'-fucosyllactose. Statistical analyses were conducted using SAS 9.4 with p < 0.05. RESULTS:The cohort comprised 41% White, 7% Black, and 52% Hispanic women. Black and Hispanic mothers exhibited lower 25(OH)D concentrations than White mothers (p < 0.001), and Hispanic mothers were more likely to be secretors (p = 0.006). In secretors, higher 25(OH)D was significantly associated with increased fucosylated and complex HMOs, including lacto-N-fucopentaose II, lacto-N-hexaose, and fucosyl-disialyl-lacto-N-hexaose I at 1 month, with sustained association for the latter at 4 months. In non-secretors, higher 25(OH)D was associated with lower 6'-sialyllactose at 4 months. Distinct HMO patterns were observed by secretor phenotype. CONCLUSION:Vitamin D status is associated with differential HMO profiles during early lactation, supporting a potential role for vitamin D-mediated regulation of mammary glycosylation that is modified by secretor status.

Human milk contains multiple bioactive components, many of which are influenced by the mother's nutritional status. To identify the impact of maternal vitamin D status on neuroactive compounds, we conducted a post hoc analysis of breast milk and matched infant stool samples from mothers categorized as sufficient or deficient for vitamin D. Neuroactive metabolites were quantified using both targeted and nontargeted liquid chromatography with tandem mass spectrometry (LC-MS/MS). Our findings revealed that breast milk from mothers with sufficient vitamin D levels contained significantly higher concentrations of tryptophan, phenylalanine, and tyrosine compared to milk from mothers with lower vitamin D levels. No significant differences were observed in tryptamine, kynurenine, kynurenic acid, anthranilic acid, quinolinic acid, tyramine, dopamine, epinephrine, or norepinephrine between the two groups. Among SCFAs and other acids, only hexanoic acid was significantly elevated in the breast milk of mothers with sufficient vitamin D. A nontargeted metabolomics analysis of infant stool identified distinct metabolite profiles, where oleamide, vaccenic acid, lacto-N-triaose, and N-acetyl-D-glucosamine varied according to maternal vitamin D levels, indicating that maternal nutrient status may influence the infant gut metabolome. These findings suggest that maternal vitamin D status is associated with neurotransmitter precursor levels in breast milk and a distinct metabolomic profile of infant stool.

Objective Evaluate the association between vitamin D (vitD) status and Corona Virus-19 (COVID-19) infection in adults aged 50 years and older. Design Adults ≥50 undergoing COVID-19 testing from July 2020 to December 2021, without prior vaccination, consented to blood analysis. SARS-CoV-2 PCR confirmed current COVID-19 infection. VitD status was assessed via 25(OH)D concentration (LCMS/MS, ZRT Labs, Portland, OR). Sociodemographic data were collected at enrollment. Statistical analyses (SAS 9.4) examined associations between sociodemographics, COVID-19, and vitD status. Multivariate logistic regression analyzed factors linked to COVID-19 or vitD status. Results Of 131 participants, 46.6% were ≥65 years old, 71.0% married, 19.9% Black American, 36.6% male, 38.9% Medicaid/Medicare/self-pay, and 42.8% BMI≥30. VitD status and Black American (P=0.0001) significantly associated with COVID-19 infection (P=0.0001). Black American (P=0.0003), males (P=0.003), and BMI (P=0.007) were inversely associated with 25(OH)D concentration. In a multiple logistic regression model predicting COVID-19 infection, only vitamin D status remained significant after controlling for certain sociodemographic and clinical factors (P<0.0001, OR 0.92, 95% CI 0.89–0.95).Of the 44 COVID-positive participants, 35 (79.6%) were hospitalized and 19 (43.2%) were admitted to the Intensive Care Unit (ICU). Hospitalization due to COVID-19 was associated with age ≥65 years old (P=0.02; OR 12.0, 95% CI 1.34–106.79), male (P=0.02, OR 10.7, 95% CI 1.20–94.73), and 25(OH)D <40 ng/mL (P=0.0006, OR 42.5, 95% CI 3.90–461.01). In multivariate analysis, the association between vitamin D status and the risk of COVID-related hospitalization remained significant and inversely associated (P=0.03, OR 0.88, 95% CI 0.78–0.99).In unadjusted analysis, COVID pneumonia was associated with male sex (P=0.049; OR 4.6, 95% CI 1.06–20.16) and 25(OH)D <40 ng/mL (P=0.006, OR 18.8, 95% CI 1.9–184.10). Participants with COVID infection and 25(OH)D <20 ng/mL were 2.1 times more likely to be admitted to ICU/death (P=0.03). In unadjusted analysis, ICU admission and/or death were linked to age ≥65 years (P=0.0002, OR 16.9, 95% CI 3.63–78.56), Medicaid/Medicare/self-pay insurance status (P=0.004, OR 0.1, 0.04–0.56), and 25(OH)D <20 (P=0.03, OR 3.9, 1.09–13.66) and <40 ng/mL (P=0.03); however, only age ≥65 remained significant in multivariate analysis (P=0.04, OR 6.7, CI 1.05–43.0). Conclusions Lower 25(OH)D concentration was a significant predictor and/or contributor to COVID-19 infection, suggesting the importance of maintaining adequate vitamin D status in reducing infection risk and mitigating severe outcomes.

Preterm infants have an immature intestinal environment featuring microbial dysbiosis. Human milk can improve the composition of the neonatal gut microbiome by supporting commensal species. Milk free fatty acids (FFAs) provide nutritional energy, participate in endogenous signaling, and exert antimicrobial effects. This study examined the growth of individual commensal and pathobiont microbes in response to unesterified unsaturated FFAs found in milk: oleic, linoleic, arachidonic, and docosahexaenoic acid. Select species of commensal and pathobiont genera (Bifidobacterium, Lactobacillus, Streptococcus, Staphylococcus, Enterococcus, Acinetobacter, Pseudomonas, Escherichia, and Klebsiella) were cultured with FFAs. The growth of all commensals, except for L. johnsonii, was significantly inhibited by the highest concentration (1 %) of all FFAs. L. johnsonii was only inhibited by arachidonic acid. In contrast, suppression of pathobionts in response to FFAs was less pronounced. Higher concentrations (0.1 %, 1 %) of docosahexaenoic acid significantly inhibited the growth of five of eight pathobionts. Meanwhile, for oleic, linoleic, and arachidonic acid, only two of eight pathobionts were significantly affected. Intriguingly, the effects for these FFAs were highly complex. For example, S. agalactiae growth was enhanced with 1 % oleic acid but suppressed at 0.01 %; however, the effects were directionally opposite for linoleic acid, i.e., suppressed at 1 % but enhanced at 0.01 %. Our genome analyses suggest that pathobiont survival may be related to the number of gene copies for fatty acid transporters. Overall, the effect of FFAs was dose-dependent and species-specific, where commensal growth was broadly inhibited while pathobionts were either unaffected or exhibited complex, bi-directional responses.

Background: Few studies evaluate the effects of vitamin D status and supplementation on maternal bone mineral density (BMD) during lactation and further lack inclusion of diverse racial/ethnic groups, body mass index (BMI), or physical activity.Objective: Determine the effects of vitamin D treatment/status, feeding type, BMI, race/ethnicity, and physical activity on postpartum women's BMD to 7 months.Methods: Women with singleton pregnancies beginning 4-6 weeks' postpartum were randomized into two treatment groups (400 or 6400 IU vitamin D/day). Participant hip, spine, femoral neck, and whole-body BMD using Dual-energy X-ray absorptiometry (DXA Hologic), serum 25-hydroxyvitamin D [25(OH)D] (RIA; Diasorin), BMI, and physical activity were measured at 1, 4, and 7 months postpartum. A general linear mixed modeling approach was undertaken to assess the effects of vitamin D status [both serum 25(OH)D concentrations and treatment groups], feeding type, race/ethnicity, BMI, and physical activity on BMD in postpartum women.Results: During the 6-month study period, lactating women had 1-3% BMD loss in all regions compared with 1-3% gain in nonlactating women. Higher maternal BMI was associated with less bone loss in femoral neck and hip regions. Black American women had less BMD loss than White/Caucasian or Hispanic lactating women in spine and hip regions. Exclusively breastfeeding women in the 6400 IU vitamin D group had less femoral neck BMD loss than the 400 IU group at 4 months sustained to 7 months. Physical activity was associated with higher hip BMD.Conclusion: While there was BMD loss during lactation to 7 months, the loss rate was less than previously reported, with notable racial/ethnic variation. Breastfeeding was associated with loss in BMD compared with formula-feeding women who gained BMD. Higher BMI and physical activity independently appeared to protect hip BMD, whereas higher vitamin D supplementation appeared protective against femoral neck BMD loss.

Objective Our objective was to conduct a secondary, post hoc analysis of the National Institute of Child Health and Human Development (NICHD) vitamin D (vitD) pregnancy study by Hollis et al, which reported on the effect of vitD supplementation in pregnant women and determine the potential interaction between intact parathyroid hormone ( iPTH) concentrations, vitD status, and various comorbidities associated with pregnancy. Women with low 25-hydroxy vitamin D (25(OH)D) concentrations and high iPTH concentrations during pregnancy, known as functional vitamin-D deficiency (FVDD), were more likely to acquire complications also affecting their neonates. Study Design This post hoc analysis of data collected from a diverse group of pregnant women participating in the NICHD vitD pregnancy study was applied to investigate the applicability of the concept of FVDD in pregnancy (Hemmingway, 2018) in identifying potential risks for certain comorbidities of pregnancy. This analysis defines FVDD as maternal serum 25(OH)D concentrations below 20 ng/mL and iPTH concentrations above 65 pg/mL creating a definitive ratio number, 0.308, to classify mothers as having FVDD prior to delivery (PTD). Statistical analyses were performed using SAS 9.4 ( Cary, NC). Results In total, 281 women ( 85 African American, 115 Hispanic, and 81 Caucasian) with 25(OH)D and iPTH concentrations measured at monthly visits were included in this analysis. No statistically significant association was found between mothers classified as having FVDD at baseline or 1-month PTD and hypertensive disorders of pregnancy, infection, or admittance to the neonatal intensive care unit. When combining all comorbidities of pregnancy in this cohort, results showed those with FVDD at baseline, 24 weeks' gestation, and 1- month PTD were more likely to experience a comorbidity (p = 0.001; p = 0.001; p = 0.004, respectively). Those with FVDD 1-month PTD were 7.1 times (confidence interval [CI]: 1.71-29.81) more likely to have preterm birth (<37 weeks) than women without FVDD. Conclusion Participants were more likely to have experienced preterm birth if they met the criteria for FVDD. This study supports the importance of FVDD during pregnancy.

In the neonatal intensive care unit, adequate nutrition requires various enteral products, including human milk and formula. Human milk is typically fortified to meet increased calorie goals, and infants commonly receive vitamin mixes, iron supplements, and less frequently, thickening agents. We examined the growth of 16 commensal microbes and 10 pathobionts found in the premature infant gut and found that formula, freshly pasteurized milk, and donated banked milk generally increased bacterial growth. Fortification of human milk significantly elevated the growth of all microbes. Supplementation with thickeners or NaCl in general did not stimulate additional growth. Vitamin mix promoted the growth of several commensals, while iron promoted growth of pathobionts. These data indicate that pathobionts in the preterm gut have significant growth advantage with preterm formula, fortified donor milk, and supplemented iron and suggest that the choice of milk and supplements may impact the infant gut microbiota.

OBJECTIVE:To evaluate the levels of inflammatory cytokines and symptom survey scores in patients diagnosed with Menière's disease or vestibular migraine from a single center by a single neurotologist compared to control subjects with no history of dizziness or migraine. STUDY DESIGN:Cross-sectional pilot study. SETTING:Single-center tertiary referral center in Charleston, SC. PATIENTS:Patients were recruited from the neurotology clinic at a tertiary referral center. Patients with definite Menière's disease or definite vestibular migraine as defined by the Barany consensus criteria were eligible. Control subjects presented to clinic without dizziness, vertigo, fluctuating hearing loss, or a history of migraine. MAIN OUTCOME MEASURES:Questionnaire scores include DHI, SF-20, CFQ, PHQ-9, PSWQ, GAD-7, NVI, DCS, VM-PATHI, and MD-POSI. Circulating and in vitro levels of cytokines include ENA-78, GROα, IFN-α2a, IFN-γ, IL-10, IL-1α, IL-1β, IL-1RA, IL-2, IL-4, IL-5, IL-6, IL-8, MCP-1, MCP-2, MDC, MIP-1α, MIP-1β, and TNF-α. Cytokine levels were compared with effect size analysis. RESULTS:There were 20 Menière's disease, 20 vestibular migraine, and 10 control patients enrolled in this study. Episode frequencies ranged from three per week to two to five per year in the MD group and daily to 1 every 2 to 3 months in the VM group. When patient-derived PBMC samples were compared to vestibular migraine, TNF-α ( d = -0.427 [-0.879, 0.025]) and IFN-γ ( d = -0.818 [-1.313, -0.323]) were found to be higher in Menière's disease, whereas ENA-78 ( d = -0.652 [-1.361, 0.056]) was found to be lower. No differences were found when cytokines were measured following stimulation with LPS. CONCLUSIONS:This pilot study suggests Menière's disease patients may have higher levels of TNF-α and IFN-γ and lower levels of ENA-78 than vestibular migraine patients when measured following in vitro release from patient-derived PBMC. Increased sample size, optimized blood draw timing, and more specific PBMC stimulation may help us further elucidate inflammatory pathways implicated in these disorders.

Introduction: Vitamin D (VitD) has been shown to impact neurodevelopment. Studies have shown that higher 25-hydroxy-vitamin D (25(OH)D) concentrations (the indicator of vitD status) may be associated with better neurodevelopmental outcomes, although current data are conflicting. This study examined the relationship between total circulating 25(OH)D concentrations and neurodevelopmental outcomes in 3–5-year-old (3–5 yo) children. Methods: In this study, pregnant women were randomized to receive 400 (standard dose), 2000, or 4000 IU vitD3/day. Offspring then underwent the Brigance Screen at 3–5 yo. The 25(OH)D concentration was measured at birth and 3–5 yo. Relationships between Brigance scores and 25(OH)D and Brigance scores and vitamin D binding protein (VDBP) genotype were examined. Results: Higher 25(OH)D at the time of testing was associated with better overall performance on neurodevelopmental testing as measured by the Brigance quotient (B = 0.208, p = 0.049). Scores were then broken down into sub-scores. Children born to mothers in the 2000 IU/day group scored higher on the Brigance language component of the assessment versus the standard dose group (B = 4.667, p = 0.044). The group of children who had the Gc1f-1s or Gc1f-2 genotypes scored higher on the Brigance academic component (B = 9.993, p < 0.001) and lower on the Brigance language component versus the 1f1f genotype (B = −9.313, p < 0.001). Children with the Gc1s-1s, Gc1s-2, or Gc2-2 genotypes also scored lower than the Gc1f-1f genotype (B = −6.757, p = 0.003). Conclusion: These results suggest that higher 25(OH)D concentrations early in life and higher doses of maternal vitamin D supplementation during pregnancy may have a positive association with neurodevelopmental outcomes. This study also suggests that the VDBP genotype is associated with neurodevelopment and differentially affects various fields of neurodevelopment.

Pregnancy is a unique time when amplified sex steroid concentrations promote an escalation in vitamin D binding protein (DBP) synthesis, associated with increased total vitamin D and metabolites, including 25-hydroxyvitamin D (25(OH)D). Free 25(OH)D concentration increases disproportionately to total 25(OH)D during pregnancy, likely an adaptation to supply the woman and fetus with readily available 25(OH)D. Highlighting the importance of the calcium metabolic stress during pregnancy, the interactional relationship between serum 25(OH)D and PTH has been evaluated. Maternal total 25(OH)D and total 25(OH)D/iPTH are measures of vitamin D status and biomarkers for potential pregnancy complications. It has been proposed that free 25(OH)D and free 25(OH)D/iPTH could be better indicators of vitamin D status and predictors of pregnancy complications such as gestational diabetes (GDM), hypertensive disorders of pregnancy, and preterm delivery. This study aims to determine if free 25(OH)D and its association with PTH are more accurate predictors of comorbidities of pregnancy than total 25(OH)D and its association with PTH. In this post hoc analysis of the Kellogg Pregnancy Study, a double-blind randomized placebo-controlled trial, participants included 297 women with singleton pregnancies: 191 participants were randomized into a group receiving a daily prenatal (400 IU vitamin D3) while 196 received a prenatal plus extra supplementation (4400 IU vitamin D3). Blood and urine samples were collected monthly. 297 participants' serum total 25(OH)D concentrations were measured using radioimmunoassay at baseline (visit 1) and 5-7 months' gestation (visit 6-7). 93 participants' serum free 25(OH)D and PTH concentrations were measured using ELISA and immunoradiometric assay, respectively, at visit 1 and 6-7; 66 participants had paired samples and were included in this analysis. Data were analyzed using SAS 9.4, Cary, N.C. or SPSS v28, IBM Corporation, Armonk, N.Y. Results were considered significant with a p < 0.05. A significant relationship exists between the ratio of total 25(OH)D/iPTH and free 25(OH)D/iPTH grouped by total 25(OH)D ≥ 30 ng/mL and < 30 ng/mL as an indicator of maternal vitamin D status. There was a statistically significant relationship between lower mean free 25(OH)D/iPTH and the development of GDM at visit 1 (p = 0.0003) and at visit 6-7 (p = 0.001) while total 25(OH)D/iPTH and GDM were significantly related only at visit 1 (p = 0.029). In this exploratory cohort, neither free 25(OH)D/iPTH nor total 25(OH)D/iPTH were significantly associated with increased incidence of preterm delivery, hypertensive disorders, or combined comorbidities of pregnancy. An univariate logistic regression evaluating the outcome of gestational diabetes while independently controlling for independent factors showed the ratio of free 25(OH)D/iPTH was more closely associated with gestational diabetes than the ratio of total 25(OH)D/iPTH, although neither were significant. This proof-of-concept analysis suggests that the ratio of free 25(OH)D/iPTH is associated with the development of gestational diabetes throughout pregnancy while total 25(OH)D/iPTH is only associated with the outcome early in pregnancy. Further investigation is warranted to explore this relationship between calcium metabolic stress during pregnancy with a larger cohort to improve validity,reproducibility, and relevance to other pregnancy comorbidities.

endogenous HA localization and staining intensity, and stool microbial compositions are pending.

The oleic acid/alpha-lactalbumin complex HAMLET (human alpha-lactalbumin made lethal to tumors) is cytotoxic to various cancerous cell lines and is assembled from alpha-lactalbumin (ALA) and free oleic acid (OA). HAMLET is also cytotoxic to normal immature intestinal cells. It remains unclear if HAMLET, experimentally assembled with OA and heat, can spontaneously assemble in frozen human milk over time. To approach this issue, we used a set of timed proteolytic experiments to evaluate the digestibility of HAMLET and native ALA. The purity of HAMLET in human milk was confirmed by ultra high performance liquid chromatography coupled to tandem mass spectrometry and western blot to resolve the ALA and OA components. Timed proteolytic experiments were used to identify HAMLET in whole milk samples. Structural characterization of HAMLET was performed by Fournier transformed infrared spectroscopy and indicated a transformation of secondary structure with increased alpha-helical character of ALA upon binding to OA.

Atrial natriuretic peptide (ANP) and its receptors natriuretic peptide receptor (NPR)-A and NPR-C are all highly expressed in alveolar epithelial type II cells (AEC2s) in the late-gestation ovine fetal lung and are dramatically decreased postnatally. However, of all the components, NPR-C stimulation inhibits ANP-mediated surfactant secretion. Since alveolar oxygen increases dramatically after birth, and steroids are administered to mothers antenatally to enhance surfactant lung maturity, we investigated the effects of O2 concentration and steroids on NPR-C-mediated surfactant secretion in AEC2s. NPR-C expression was highest at 5% O2 while being suppressed by 21% O2, in cultured mouse lung epithelial cells (MLE-15s) and/or human primary AEC2s. Surfactant protein-B (SP-B) was significantly elevated in media from both in vitro and ex vivo culture at 13% O2 versus 21% O2 in the presence of ANP or terbutaline (TER). Both ANP and C-ANP (an NPR-C agonist) attenuated TER-induced SP-B secretion; this effect was reversed by dexamethasone (DEX) pretreatment in AEC2s and by transfection with NPR-C siRNA in MLE-15 cells. DEX markedly reduced AEC2 NPR-C expression, and pregnant ewes treated with betamethasone showed reduced ANP in fetal sheep lung fluid. These data suggest that elevated O2 downregulates AEC2 NPR-C and that steroid-mediated NPR-C downregulation in neonatal lungs may provide a novel mechanism for their effect on perinatal surfactant production.

The intestinal microbiota of the preterm neonate has become a major research focus, with evidence emerging that the microbiota influences both short and long-term health outcomes, in the neonatal intensive care unit and beyond. Similar to the term microbiome, the preterm gut microbiome is highly influenced by diet, specifically formula and human milk use. This study aims to analyze next-generation products including preterm formula, human milk-oligosaccharide term formula, and preterm breastmilk. We used a culture-based model to differentially compare the growth patterns of individual bacterial strains found in the human intestine. This model probed 24 strains of commensal bacteria and 8 pathobiont species which have previously been found to cause sepsis in preterm neonates. Remarkable differences between strain growth and culture pH were noted after comparing models of formulas and between human milk and formula. Both formula and human milk supported the growth of commensal bacteria; however, the formula products, but not human milk, supported the growth of several specific pathogenic strains. Computational analysis revealed potential connections between long-chain fatty acid and iron uptake from formula in pathobiont organisms. These findings indicate that there is a unique profile of growth in response to human milk and formula and shed light into how the infant gut microbiota could be influenced.

Background: Vitamin D (vitD) plays a major role in maintenance of bone mineral homeostasis. It is unknown if bone mineral content (BMC) and bone mineral density (BMD) differ between infants who receive direct vitD supplementation and those who receive vitD indirectly via their mother's breast milk, while she received a high dose of vitD. It is hypothesized that there would be no differences in BMC or BMD by treatment group.Design/Methods: Randomized, double-blind trial to compare BMD and BMC of infants who received direct vitD (400 IU vitD(3)/day) in addition to their mother receiving standard dosage (400 IU vitD(3)/day) versus infants whose mothers were their only source of vitD and were given high-dose supplementation (6,400 IU vitD(3)/day). Participants were exclusively breastfeeding mothers and their infant consuming only human milk. Infant BMC and BMD were measured by dual-energy X-ray absorptiometry (DXA) scans of the infant's total body using Hologic Discovery A Densitometer and analyzed using Hologic Infant software at 1, 4, and 7 months of age.Results: Infant BMC and BMD did not differ significantly at 1, 4, or 7 months of age between direct and indirect supplementation arms. The mean difference in BMC from 1 to 7 months was 1.624 and 1.464 g for the 400 and 6,400 IU groups, respectively, (p = 0.5); the mean difference in BMD over this same period was 0.042 and 0.032 g/cm(2) for the 400 and 6,400 IU groups, respectively (p = 0.2). Although some differences among races were observed, this did not reflect changes in bone growth between the treatment arms.Conclusion: High-dose vitD supplementation of mothers during lactation provided an efficacious alternative to direct supplementation of infants, as evidenced by noninferior infant BMD and BMC.Clinical Trial Registration number: NCT00412074.

These findings indicate a unique profile of growth in response to human milk and formula. Since alterations in the infant gut microbiota have linked to childhood health and development, understanding how bacteria differentially utilize whole milk or formula may provide insights into which microbes drive diet-related microbiota shifts.

It is unknown if vitamin D (vitD) sufficiency in breastfeeding mothers can lead to physiological outcomes for their children that are discernible from infant vitD sufficiency per se. In a 3-month, randomized vitD supplementation study of mothers and their exclusively breastfeeding infants, the effects of maternal vitD sufficiency were determined on infant plasma concentrations of 25-hydroxyvitamin D (i.e., vitD status) and 11 cytokines. An inverse correlation was seen between maternal vitD status and infant plasma TNF concentration (r = −0.27; p < 0.05). Infant whole blood was also subjected to in vitro antigenic stimulation. TNF, IFNγ, IL-4, IL-13, and TGFβ1 responses by infant leukocytes were significantly higher if mothers were vitD sufficient but were not as closely correlated to infants’ own vitD status. Conversely, IL-10 and IL-12 responses after antigenic challenge were more correlated to infant vitD status. These data are consistent with vitD-mediated changes in breast milk composition providing immunological signaling to breastfeeding infants and indicate differential physiological effects of direct-infant versus maternal vitD supplementation. Thus, consistent with many previous studies that focused on the importance of vitD sufficiency during pregnancy, maintenance of maternal sufficiency likely continues to affect the health of breastfed infants.