N-Heterocyclic Carbene Triazolium Salts Containing Brominated Aromatic Motifs: Features and Synthetic Protocol.

In this work, we provide a brief overview of the role of N-aryl substituents on triazolium N-heterocyclic carbene (NHC) catalysis. This synopsis provides context for the disclosed synthetic protocol for new chiral N-heterocyclic carbene (NHC) triazolium salts with brominated aromatic motifs. Incorporating brominated aryl rings into NHC structures is challenging, probably due to the substantial steric and electronic influence these substituents exert throughout the synthetic protocol. However, these exact characteristics make it an interesting N-aryl substituent, because the electronic and steric diversity it offers could find broad use in organometallic- and organo-catalysis. Following the synthetic reaction by NMR guided the extensive modification of a known protocol to enable the preparation of these challenging NHC pre-catalysts.

The influence of vitamin D supplementation on local and systemic inflammatory markers in periodontitis patients: A pilot study.

OBJECTIVES: Vitamin D deficiency/insufficiency is a worldwide public health issue that has been linked to numerous inflammatory disorders, including periodontitis. There is increasing support for a role for adequate vitamin D levels in overall health. Populations with darker skin color have a higher prevalence of vitamin D deficiency/insufficiency and periodontitis. The purpose of this small pilot study was to investigate the influence of 12 weeks of 25(OH)D vitamin D supplementation (VDS) on mediators of systemic inflammation in dark-skinned, periodontitis patients. MATERIALS AND METHODS: A total of 23 patients with moderate to severe periodontitis were randomly assigned to the vitamin D group or placebo group and received intensive single visit scaling and root planning to elicit a systemic inflammatory response. RESULTS: Vitamin D supplementation increased serum 25(OH)D levels approximately 2-fold over baseline levels; moreover, VDS group had reduced peripheral blood CD3 and CD3+CD8+ cytotoxic T lymphocyte (CTLs) counts and reduced pro-inflammatory salivary cytokines. In contrast, VDS group had higher levels of the autophagy-related proteins and other proteins crucial for anti-microbial autophagy in whole blood PBMCs. CONCLUSION: In conclusion, VDS has multiple benefits for reducing systemic inflammation and promoting induction of autophagy-related proteins related to anti-microbial functions.

Bardet-Biedl syndrome proteins 1 and 3 regulate the ciliary trafficking of polycystic kidney disease 1 protein.

Bardet-Biedl syndrome (BBS) and autosomal dominant polycystic kidney disease (ADPKD) are two genetically distinct ciliopathies but share common phenotypes such as renal cysts. Seven BBS proteins form a complex called the BBSome which is localized at the basal body or ciliary axoneme and regulates the ciliary entry or flagellar exit of several signaling molecules. Here, we demonstrate that, unlike the seven-span somatostatin receptor 3 or the leptin receptor that interacts with all subunits of the BBSome, the ADPKD protein polycystin-1 (PC1) interacts with BBS1, BBS4, BBS5 and BBS8, four of the seven components of the BBSome. Only depletion or mutation of BBS1, but not depletion of BBS5 and BBS8, or knockout of BBS4, impairs ciliary trafficking of PC1 in kidney epithelial cells. Depletion of these BBS proteins affects neither the ciliary length nor the plasma membrane targeting of PC1. Expression of a pathogenic BBS3/Arl6 mutant (T31R) that locks Arl6 in the GDP form leads to stunted cilia and inhibition of PC1 on primary cilia. We propose that the 11-span membrane protein PC1 is a BBSome cargo and that the components of the BBSome may possess subunit-specific functions. Moreover, physical interactions between the BBS and ADPKD proteins may underline the overlapping renal phenotypes in these two diseases.

Cathelicidin antimicrobial protein, vitamin D, and risk of death in critically ill patients.

INTRODUCTION: Decreased production of cathelicidin antimicrobial protein-18 (hCAP18) has been proposed to be a key mechanism linking decreased 25-hydroxyvitamin D (25D) levels with adverse outcomes among critically ill patients. However, few studies in humans have directly assessed plasma hCAP18 levels, and no study has evaluated the association between hCAP18 levels and adverse outcomes among critically ill patients. METHODS: We performed a single-center, prospective cohort study among 121 critically ill patients admitted to intensive care units (ICUs) between 2008 and 2012. We measured plasma hCAP18, 25D, D-binding protein, and parathyroid hormone levels on ICU day 1. The primary endpoint was 90-day mortality. Secondary endpoints included hospital mortality, sepsis, acute kidney injury, duration of mechanical ventilation, and hospital length of stay. RESULTS: ICU day 1 hCAP18 levels were directly correlated with 25D levels (Spearman's rho (rs) = 0.30, P = 0.001). In multivariate analyses adjusted for age and Acute Physiology and Chronic Health Evaluation II (APACHE II) score, patients with hCAP18 levels in the lowest compared to highest tertile on ICU day 1 had a 4.49 (1.08 to 18.67) greater odds of 90-day mortality, and also had greater odds of sepsis. ICU day 1 levels of other analytes were not associated with 90-day mortality. CONCLUSIONS: Lower 25D levels on ICU day 1 are associated with lower hCAP18 levels, which are in turn associated with a greater risk of 90-day mortality. These findings provide a potential mechanistic basis for the frequently observed association between low 25D levels and poor outcomes in critically ill patients.

Randomized controlled trial of calcitriol in severe sepsis.

RATIONALE: Vitamin D and its metabolites have potent immunomodulatory effects in vitro, including up-regulation of cathelicidin, a critical antimicrobial protein. OBJECTIVES: We investigated whether administration of 1,25-dihydroxyvitamin D (calcitriol) to critically ill patients with sepsis would have beneficial effects on markers of innate immunity, inflammation, and kidney injury. METHODS: We performed a double-blind, randomized, placebo-controlled, physiologic study among 67 critically ill patients with severe sepsis or septic shock. Patients were randomized to receive a single dose of calcitriol (2 µg intravenously) versus placebo. The primary outcome was plasma cathelicidin protein levels assessed 24 hours after study drug administration. Secondary outcomes included leukocyte cathelicidin mRNA expression, plasma cytokine levels (IL-10, IL-6, tumor necrosis factor-α, IL-1ß, and IL-2), and urinary kidney injury markers. MEASUREMENTS AND MAIN RESULTS: Patients randomized to calcitriol (n = 36) versus placebo (n = 31) had similar plasma cathelicidin protein levels at 24 hours (P = 0.16). Calcitriol-treated patients had higher cathelicidin (P = 0.04) and IL-10 (P = 0.03) mRNA expression than placebo-treated patients 24 hours after study drug administration. Plasma cytokine levels (IL-10, IL-6, tumor necrosis factor-α, IL-1ß, and IL-2) and urinary kidney injury markers were similar in calcitriol- versus placebo-treated patients (P > 0.05 for all comparisons). Calcitriol had no effect on clinical outcomes nor were any adverse effects observed. CONCLUSIONS: Calcitriol administration did not increase plasma cathelicidin protein levels in critically ill patients with sepsis and had mixed effects on other immunomodulatory markers. Additional phase II trials investigating the dose and timing of calcitriol as a therapeutic agent in specific sepsis phenotypes may be warranted. Clinical trial registered with www.clinicaltrials.gov (NCT 01689441).

Sixteen-Week Vitamin D3 Supplementation Increases Peripheral T Cells in Overweight Black Individuals: Post hoc Analysis of a Randomized, Double-Blinded, Placebo-Controlled Trial.

Background: Vitamin D is considered to modulate T-cell function, which has been implicated in the treatment of inflammatory conditions. However, there is limited knowledge on the effects of vitamin D and its influences on circulating T-cell profiles in humans, particularly in overweight Black individuals who are more likely to be vitamin D insufficient (serum 25(OH)D concentrations of ≤20 ng/mL). Thus, this study tested the hypothesis that vitamin D supplementation modulates T-cell composition, which is in a dose-dependent manner. Methods: A 16-week randomized, double-blinded, placebo-controlled trial of vitamin D3 supplementation was undertaken in 70 overweight/obese Black people (mean age = 26 years, 82% female) with 25 hydroxyvitamin D ≤ 20 ng/mL at baseline. Subjects were randomly assigned a supervised monthly oral vitamin D3 equivalent to approximately 600 IU/day (n = 17), 2000 IU/day (n = 18), 4000 IU/day (n = 18), or a placebo (n = 17). Fresh peripheral whole blood was collected and CD3+, CD4+ and CD8+ cell counts and percentages were determined by flow cytometry at baseline and at 16 weeks, among 56 subjects who were included in the analyses. Results: A statistically significant increase in CD3+% in the 2000 IU/day vitamin D3 supplementation group, and increases in CD4+% in the 2000 IU/day and 4000 IU/day vitamin D3 supplementation groups were observed (p-values < 0.05) from the changes in baseline to 16 weeks. Further adjustments for age, sex and BMI showed that 2000 IU/day vitamin D3 supplementation increased in CD3+ count, CD3%, CD4 count, and CD4%, as compared to the placebo group (p-values < 0.05). Moreover, the highest serum 25(OH)D quantile group had the highest CD3% and CD4%. Conclusions: Sixteen-week vitamin D3 supplementation increases peripheral blood T-cell numbers and percentages in overweight/obese Black patients with vitamin D insufficiency. This resulting shift in circulating T-cell composition, particularly the increase in T helper cells (CD4+ cells), suggests that vitamin D supplementation may improve immune function in Black individuals.

Handgrip and sex-specific cardiometabolic risk factors in Hispanic/Latino migrant farmworkers.

Studies have suggested that handgrip strength might be a marker for cardiometabolic risk (CMR), but it has not been studied in Hispanic/Latino farmworker population. This study aimed to characterize absolute and relative handgrip strength in Hispanic/Latino farmworkers, and investigate the sex-specific association between handgrip strength and CMR factors. CMR factors and seated isometric absolute (the sum of both hands) and relative (absolute handgrip strength divided by body mass index) handgrip strengths were collected in 173 Hispanic/Latino farmworkers (mean age 35.1 ± 0.7 years; 49% female). The absolute and the relative handgrip strengths were 89.2 ± 1.8 kg, 3.3 ± 0.1 kg among males, and 56.5 ± 1.9 kg, 1.9 ± 0.1 kg among females, respectively. Age was correlated with absolute (r = - 0.17, p = 0.03) and relative handgrip strengths (r = - 0.28, p < 0.01). In males, absolute handgrip was related to triglycerides (r = - 0.25, p < 0.05), whereas relative handgrip was related to waist circumference (r = - 0.32, p < 0.01), waist/hip circumference ratio (r = - 0.36, p < 0.01), high-density lipoprotein (r = 0.24, p < 0.05), and triglycerides (r = - 0.35, p < 0.01). In females, absolute handgrip was related to fasting plasma glucose (r = - 0.28, p = 0.03), whereas relative handgrip was related to waist circumference (r = - 0.38, p < 0.01) and fasting plasma glucose (r = - 0.22, p < 0.05). Males had lower absolute handgrip strength when their triglycerides levels were at risk (p = 0.021), and lower relative handgrip strength when their plasma glucose (p = 0.034) and triglycerides (p = 0.002) levels were at risk. Females had lower relative handgrip strength when their plasma glucose (p = 0.001) and blood pressure (p = 0.004) were at risk. This study suggests that handgrip strength may be associated with sex-specific CMR factors in a Hispanic/Latino farmworker population.

The Association of Intraocular Pressure With Obesity and Cardiometabolic Risk in a Young Farmworker Population.

PRCIS: Intraocular pressure (IOP) was found to be significantly correlated with body mass index (BMI), waist circumference, and diastolic blood pressure (DBP) in a farmworker population located in the southeast Georgia, USA. BMI was correlated with IOP, independent of systemic blood pressures. PURPOSE: Elevated IOP is a known risk factor for glaucomatous optic neuropathy and is believed to be associated with obesity and cardiometabolic diseases. The high prevalence of these conditions in the United States necessitates an evaluation of the relationship among obesity, cardiometabolic risks, and IOP among understudied younger populations. MATERIALS AND METHODS: Farmworker data were collected from the annual Costa-Layman Health Fair between 2013 and 2017. Correlations of IOP with demographic factors, obesity, and cardiometabolic risks were analyzed using analysis of covariance, partial Pearson correlations, and linear regressions. RESULTS: In the farmworker population (n=346), the mean IOP was 15.5 mm Hg and the prevalence of ocular hypertension (IOP>21 mm Hg) was 5.5%. BMI, waist circumference, and DBP were significantly correlated (r=0.192, P=0.001; r=0.128, P=0.017; r=0.142, P=0.007, respectively) with IOP when adjusted for age, sex, and ethnicity. Each 10 mm Hg increase in DBP corresponded with a 0.51 mm Hg increase in IOP. With adjustment for age, sex, ethnicity, systolic blood pressure, and DBP, BMI remained significantly correlated with IOP (r=0.166, P=0.002). CONCLUSIONS: Higher IOP is associated with obesity measures including BMI and waist circumference and is correlated with DBP. These findings suggest that BMI is an independent risk factor for elevated IOP.

Vitamin D3 Supplementation Increases Long-Chain Ceramide Levels in Overweight/Obese African Americans: A Post-Hoc Analysis of a Randomized Controlled Trial.

Sphingolipid metabolism plays a critical role in cell growth regulation, lipid regulation, neurodevelopment, type 2 diabetes, and cancer. Animal experiments suggest that vitamin D may be involved in sphingolipid metabolism regulation. In this study, we tested the hypothesis that vitamin D supplementation would alter circulating long-chain ceramides and related metabolites involved in sphingolipid metabolism in humans. We carried out a post-hoc analysis of a previously conducted randomized, placebo-controlled clinical trial in 70 overweight/obese African-Americans, who were randomly assigned into four groups of 600, 2000, 4000 IU/day of vitamin D3 supplements or placebo for 16 weeks. The metabolites were measured in 64 subjects (aged 26.0 ± 9.4 years, 17% male). Serum levels of N-stearoyl-sphingosine (d18:1/18:0) (C18Cer) and stearoyl sphingomyelin (d18:1/18:0) (C18SM) were significantly increased after vitamin D3 supplementation (ps < 0.05) in a dose-response fashion. The effects of 600, 2000, and 4000 IU/day vitamin D3 supplementation on C18Cer were 0.44 (p = 0.049), 0.52 (p = 0.016), and 0.58 (p = 0.008), respectively. The effects of three dosages on C18SM were 0.30 (p = 0.222), 0.61 (p = 0.009), and 0.68 (p = 0.004), respectively. This was accompanied by the significant correlations between serum 25-hydroxyvitamin D3 [25(OH)D] concentration and those two metabolites (ps < 0.05). Vitamin D3 supplementations increase serum levels of C18Cer and C18SM in a dose-response fashion among overweight/obese African Americans.

Effects of Vitamin D3 Supplementation on Epigenetic Aging in Overweight and Obese African Americans With Suboptimal Vitamin D Status: A Randomized Clinical Trial.

Background: We have previously shown that vitamin D supplementation increases telomerase activity, suggesting an anti-aging effect. In this study, we aim to test the hypothesis that vitamin D supplementation would slow down epigenetic aging, a new marker of biological aging. Methods: A randomized clinical trial was previously conducted among 70 overweight/obese African Americans with serum 25-hydroxyvitamin D [25(OH)D] < 50 nmol/L, who were randomly assigned into four groups of 600 IU/d, 2,000 IU/d, 4,000 IU/d of vitamin D3 supplements or placebo followed by 16-week interventions. Whole genome-wide DNA methylation analysis was conducted in 51 participants. DNA methylation ages were calculated according to the Horvath and the Hannum methods. Methylation-based age acceleration index (∆Age) is defined as the difference between DNA methylation age and chronological age in years. Mixed-effects models were used to evaluate the treatment effects. Results: Fifty-one participants (aged 26.1 ± 9.3 years, 16% are male) were included in the study. After the adjustment of multi-covariates, vitamin D3 supplementation of 4,000 IU/d was associated with 1.85 years decrease in Horvath epigenetic aging compared with placebo (p value = .046), and 2,000 IU/d was associated with 1.90 years decrease in Hannum epigenetic aging (p value = .044). Serum 25(OH)D concentrations were significantly associated with decreased Horvath ∆Age only (p values = .002), regardless of treatments. Conclusions: Our results suggest that vitamin D supplementation may slow down Horvath epigenetic aging. But the effect on Hannum epigenetic aging is not conclusive. Large-scale and longer duration clinical trials are needed to replicate the findings.

Inactive Matrix Gla Protein, Arterial Stiffness, and Endothelial Function in African American Hemodialysis Patients.

BACKGROUND: Matrix Gla protein (MGP) is a vascular calcification inhibitor dependent upon vitamin K for activation. Evidence suggests that elevated plasma inactive MGP levels (desphospho-uncarboxylated MGP, dp-ucMGP; indicating poorer vascular vitamin K status) are associated with greater cardiovascular disease (CVD) risk. Despite African Americans experiencing highest rates of kidney failure and CVD events, relationships between dp-ucMGP and CVD risk markers have not been examined in this population. We investigated vascular vitamin K status (via plasma dp-ucMGP) between African American hemodialysis (HD) patients and healthy controls, and the associations of dp-ucMGP with arterial stiffness and endothelial function in HD patients only. METHODS: In 37 African American HD patients and 37 age- and race-matched controls, plasma dp-ucMGP was measured by enzyme immunoassay as a marker of vascular vitamin K status. Carotid-femoral pulse wave velocity (PWV; arterial stiffness measurement) and brachial artery flow-mediated dilation (FMD; endothelial function measurement) were assessed by applanation tonometry and ultrasound, respectively, in HD patients only. RESULTS: Mean dp-ucMGP levels were 5.6 times higher in HD patients vs. controls (2,139 ± 1,102 vs. 382 ± 181 pmol/l, P < 0.01). Multiple linear regression, adjusting for age, sex, dialysis vintage, diabetes mellitus, CVD history, body mass index, and blood pressure, revealed that dp-ucMGP was independently related to PWV (standardized ß = 0.49) and FMD (standardized ß = -0.53) (both P < 0.01). CONCLUSIONS: Our data suggest that the higher plasma dp-ucMGP concentrations found in African American HD patients may be associated with greater arterial stiffness and endothelial dysfunction.

Associations between Global DNA Methylation and Telomere Length in Healthy Adolescents.

Emerging evidence suggests that epigenetics regulates telomere dynamics in adults. However, the relationship between these pathways in children and youth remains unknown. Thus, we examined this association in 542 healthy adolescents aged 14 to 18 years old (44.8% African Americans; 55.2% females). Global DNA methylation level (%5-mC) was quantified using ELISA method. Leukocyte telomere length (LTL) was defined as relative telomere to single copy gene (T/S) ratio. Multiple linear regression models, adjusted for age, gender, ethnicity, Tanner stage, BMI, PA, and batch effect, revealed that %5 mC was associated with LTL (adjusted ß = 0.17, p < 0.01). %5 mC accounted for 5.0% of the variation for LTL. A significant gender interaction was identified (p < 0.01). There was an association between %5 mC and LTL in females (all ps < 0.01), but not in males. Further sensitivity analyses by race revealed similar associations in African Americans and whites (all ps < 0.03). The present study, for the first time, shows that lower levels of global DNA methylation are associated with shorter telomere lengths in youth, which may decrease genome stability and augment the susceptibility to diseases. Longitudinal studies are warranted to establish the effects of global DNA methylation on LTL maintenance over time.

Dose responses of vitamin D3 supplementation on arterial stiffness in overweight African Americans with vitamin D deficiency: A placebo controlled randomized trial.

BACKGROUND: Clinical trials are scant and equivocal on whether vitamin D can ameliorate arterial stiffness, particularly in populations at high risk for vitamin D deficiency and cardiovascular disease (CVD). This study determined the dose-response effects of vitamin D3 supplementation on arterial stiffness in overweight African Americans with vitamin D deficiency. METHODS: Seventy overweight African Americans (aged 13-45 years) with serum 25-hydroxyvitamin D [25(OH)D] levels ≤ 20 ng/mL were randomized to monthly oral supplementation of 18,000 IU (~600 IU/day, n = 17), 60,000 IU (~2000 IU/day, n = 18), or 120,000 IU (~4000 IU/day, n = 18) of vitamin D3 or placebo (n = 17) for 16-weeks. The arterial stiffness measurements, carotid-femoral pulse wave velocity (PWV) and carotid-radial PWV, were assessed by applanation tonometry at baseline and 16 weeks. RESULTS: Vitamin D3 supplementation demonstrated a dose-response increase in serum 25(OH)D concentrations between groups (P<0.01). A significant downward linear trend was observed for carotid-femoral PWV (P<0.01), as the mean changes in carotid-femoral PWV across the four treatment groups were 0.13 m/s (95% CI: -0.24, 0.51 m/s) for placebo, 0.02 m/s (95% CI: -0.34, 0.38 m/s) for 600 IU/day group, -0.11 m/s (95% CI: -0.50, 0.27 m/s) for the 2,000 IU/day group, and -0.70 m/s (95% CI: -1.07, -0.32 m/s) for the 4,000 IU/day group. Findings were similar for carotid-radial PWV (P = 0.03), as the mean changes in carotid-radial PWV across the four treatment groups were 0.24 m/s (95% CI: -0.45, 0.92 m/s) for placebo, 0.09 m/s (95% CI: -0.54, 0.73 m/s) for 600 IU/day group, -0.57 m/s (95% CI: -1.20, 0.07 m/s) for the 2,000 IU/day group, and -0.61 m/s (95% CI: -1.25, 0.02 m/s) for the 4,000 IU/day group. CONCLUSION: Arterial stiffness was improved by vitamin D3 supplementation in a dose-response manner in overweight African Americans with vitamin D deficiency.

Race/Ethnicity-Specific Association of Vitamin D and Global DNA Methylation: Cross-Sectional and Interventional Findings.

OBJECTIVES: Understanding of the influence of vitamin D deficiency on epigenome will provide novel insights into the chronic disease risk. We tested our hypotheses that 1) vitamin D deficiency is associated with global hypomethylation and this association may be race/ethnicity dependent; and 2) vitamin D supplementation will increase global DNA methylation level. METHODS: A two-stage design, cross-sectional observation followed by a 16 week randomized, double- blinded, placebo-controlled trial (RCT) of vitamin D3 supplementation, was undertaken. Global DNA methylation level (percentage of 5-methylcytosine, %5-mC) was quantified using leukocyte DNA with the MethylFlashTM Methylated DNA Quantification kit (Epigentek). Global methylation data was obtained from 454 Caucasians and African Americans (42%) in the observation cohort and 58 African Americans with vitamin D deficiency in the dose responsive RCT. RESULTS: In the cross-sectional study, African Americans had lower %5-mC than Caucasians (P = 0.04). A significant interaction was detected between plasma 25(OH)D and race on %5-mC (P = 0.05), as a positive association was observed between plasma 25(OH)D and %5-mC in African Americans (ß = 0.20, p<0.01), but not in Caucasians (ß = 0.03, p = 0.62). In the 16-week RCT, a dose-response benefit of vitamin D3 supplementation was observed for %5-mC, as indicated by a significant linear upward trend (-0.01 ± 0.01%, placebo; 0.11 ± 0.01%, ~600 IU/day; 0.30 ± 0.01%, ~2,000 IU/day; and 0.65 ± 0.01%, ~4,000 IU/day group; P-trend = 0.04). CONCLUSIONS: Vitamin D deficiency is associated with global hypomethylation in African Americans. Vitamin D3 supplementation increases global DNA methylation in a dose-response manner in African Americans with vitamin D deficiency.

Correction: Race/Ethnicity-Specific Association of Vitamin D and Global DNA Methylation: Cross-Sectional and Interventional Findings.

[This corrects the article DOI: 10.1371/journal.pone.0152849.].

Dose and time responses of vitamin D biomarkers to monthly vitamin D3 supplementation in overweight/obese African Americans with suboptimal vitamin d status: a placebo controlled randomized clinical trial.

BACKGROUND: A critical need exists to better understand the physiological sequel of vitamin D supplementation in obese individuals and African Americans. The aim was to comprehensively evaluate dose- and time-responses of a panel of vitamin D biomarkers to vitamin D supplements in this population. METHODS: We conducted a 16-week randomized, double-blinded, and placebo-controlled clinical trial. Seventy overweight/obese African Americans (age 13-45 years, 84 % females) with 25-hydroxyvitamin D [25(OH)D] concentrations ≤20 ng/mL were randomly assigned to receive a supervised monthly oral vitamin D3 of 18,000 IU (~600 IU/day, n = 17), 60,000 IU (~2000 IU/day, n = 18), 120,000 IU (~4000 IU/day, n = 18), or placebo (n = 17). RESULTS: There were significant dose- and time-responses of circulating 25(OH)D, 1,25-dihydroxyvitamin D [1,25(OH)2D], and intact parathyroid hormone (iPTH), but not fibroblast growth factor-23 (FGF-23), phosphorus and urine calcium to the vitamin D supplements. The mean 25(OH)D concentrations in the 2000 IU and 4000 IU groups reached ≥30 ng/mL as early as 8-weeks and remained at similar level at 16-weeks. The increase of 25(OH)D was significantly higher in the 4000 IU group than all the other groups at 8-weeks. The increase of 1,25(OH)2D was significantly higher in the 2000 IU and 4000 IU groups than the placebo at 8-weeks. Only the 4000 IU compared to the placebo significantly reduced iPTH at 8- and 16-weeks. CONCLUSIONS: Our RCT, for the first time, comprehensively evaluated time- and dose- responses of vitamin D supplementation in overweight/obese African Americans with suboptimal vitamin D status. Circulating 25(OH)D, 1,25(OH)2D, and iPTH, but not FGF-23, phosphorus and urine calcium, respond to vitamin D supplementation in a time- and dose-response manner. By monthly dosing, 2000 IU appears to be sufficient in achieving a 25(OH)D level of 30 ng/mL in this population. However, importantly, 4000 IU, rather than 2000 IU, seems to suppress iPTH. If replicated, these data might be informative in optimizing vitamin D status and providing individualized dosing recommendation in overweight/obese African Americans. TRIAL REGISTRATION: ClinicalTrials.gov number: NCT01583621, Registered on April 3, 2012.