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Bariatric surgery is emerging as an effective treatment for obesity and the metabolic syndrome. Recently, we demonstrated that Roux-en-Y gastric bypass (RYGB), but not vertical sleeve gastrectomy (VSG), resulted in improvements to white adipose physiology and enhanced brown adipose functioning. Since beneficial alterations to liver health are also expected after bariatric surgery, comparing the post-operative effects of RYGB and VSG on liver physiology is essential to their application in the treatment of non-alcoholic fatty liver disease (NAFLD).The effects of RYGB and VSG on liver physiology were compared using diet induced mouse model of obesity. High-fat diet (HFD) was administered for 12 weeks after surgery and alterations to liver physiology were assessed.Both RYGB and VSG showed decreased liver weight as well as reductions to hepatic cholesterol and triglyceride levels. There were demonstrable improvements to NAFLD activity score (NAS) and fibrosis stage scoring after both surgeries. In RYGB, these beneficial changes to liver function resulted from the downregulation of pro-fibrotic and upregulation anti-fibrotic genes, as well as increased fatty acid oxidation and bile acid flux. For VSG, though similar alterations were observed, they were less potent. However, VSG did significantly downregulate pro-fibrotic genes and showed increased glycogen content paralleled by decreased glycogenolysis which may have contributed to the resolution of NAFLD.RYGB and VSG improve liver physiology and function, but RYGB is more efficacious. Resolutions of NAFLD in RYGB and VSG are achieved through different processes, independent of weight loss.

Understanding the effects of Roux-en-Y gastric bypass (RYGB) and vertical sleeve gastrectomy (VSG) on adipose tissue physiology is important for the treatment of obesity-related metabolic disorders. By using robust mouse models of bariatric surgery that closely resemble those performed in humans, we can compare the effects of RYGB and VSG on adipose physiology in the absence of post-operative confounds such as diet and lifestyle changes. RYGB and VSG were compared using a diet-induced mouse model of obesity. High-fat diet (HFD) was administered post-operatively and changes to white and brown adipose tissue were evaluated, along with alterations to weight, glucose homeostasis, dyslipidemia, and insulin sensitivity. After prolonged exposure to high-fat diet post-operatively, RYGB was effective in achieving sustained weight loss, while VSG unexpectedly accelerated weight gain rates. The resolution of obesity-related comorbidities such as glucose and insulin intolerance, dyslipidemia, and insulin sensitivity was improved after RYGB, but not for VSG. In RYGB, there were improvements to the function and health of white adipose tissue, enhanced brown adipose metabolism, and the browning of subcutaneous white adipose tissue, with no comparable changes seen for these factors after VSG. Some markers of systemic inflammation improved after both RYGB and VSG. There are significantly different effects between RYGB and VSG when HFD is administered post-operatively and robust mouse models of bariatric surgery are used. RYGB resulted in lasting physiological and metabolic changes but VSG showed little difference from that of its sham-operated, DIO counterpart.

The obesity epidemic is associated with an increased prevalence and severity of type 2 diabetes, cardiovascular disease, and nonalcoholic fatty liver disease (NAFLD) . Bariatric surgery has become a widely accepted treatment for individuals with severe obesity and obesity related health problems. Previous studies from our lab have implicated the role of lipocalin-type prostaglandin D2 synthase (L-PGDS) in adipose tissue dysfunction and NAFLD. Our lab has also shown the effect of Roux-en Y gastric bypass (RYGB) on body weight changes, improvement to insulin sensitivity and adipose tissue physiology. In the current study, we examined the effects of L-PGDS using both diet-induced obesity (DIO) and RYGB models. We found reduced expression of L-PGDS in liver of DIO mice after 14 weeks on high fat diet. We then performed RYGB surgery in DIO mice and compared the effects of diets (normal chow and HFD for 6 weeks) on post-operative outcomes of RYGB. Our results show that both surgery and diet contribute to the substantial improvement in liver weight and HOMA-IR. H&E images of liver sections also demonstrate that both diet and surgery play significant roles in reversal of NAFLD. Interestingly, we only found significant increase to liver L-PGDS expression after RYGB when HFD is administered. We conclude that both surgery and diet plays a significant role in modulating the expression of L-PGDS in liver of obese mice. Thus, our study shows that L-PGDS may serve as a novel therapeutic target in treatment of obesity and NAFLD. Disclosure A.Srivastava: None. M.Stevenson: None. T.Palaia: None. C.Hall: None. J.Lee: None. L.Ragolia: None. Funding American Heart Association: (#15GRNT22420001) and The George Link Foundation

Rationale: Increasing prevalence of obesity and its associated risk with cardiovascular diseases demands a better understanding of the contribution of different cell types within this complex disease for developing new treatment options. Previous studies could prove a fundamental role of FTO (fat mass and obesity-associated protein) within obesity; however, its functional role within different cell types is less understood. Objectives: We identify endothelial FTO as a previously unknown central regulator of both obesity-induced metabolic and vascular alterations. Methods and Results: We generated endothelial Fto -deficient mice and analyzed the impact of obesity on those mice. While the loss of endothelial FTO did not influence the development of obesity and dyslipidemia, it protected mice from high-fat diet–induced glucose intolerance and insulin resistance by increasing AKT (protein kinase B) phosphorylation in endothelial cells and skeletal muscle. Furthermore, loss of endothelial FTO prevented the development of obesity-induced hypertension by preserving myogenic tone in resistance arteries. In Fto -deficient arteries, microarray analysis identified upregulation of L-Pgds with significant increases in prostaglandin D 2 levels. Blockade of prostaglandin D 2 synthesis inhibited the myogenic tone protection in resistance arteries of endothelial Fto -deficient mice on high-fat diet; conversely, direct addition of prostaglandin D 2 rescued myogenic tone in high-fat diet–fed control mice. Myogenic tone was increased in obese human arteries with FTO inhibitors or prostaglandin D 2 application. Conclusions: These data identify endothelial FTO as a previously unknown regulator in the development of obesity-induced metabolic and vascular changes, which is independent of its known function in regulation of obesity.

Cerebral cavernous malformations (CCMs) are enlarged leaking vascular lesions in the brain caused by loss-of-function mutations in CCM1/2/3 genes or loss of expression. Although we previously showed that Nogo-B receptor (NgBR) knockout in endothelial cells (ECs) results in CCMs-like cerebrovascular lesions in the mouse embryo, the molecular mechanism by which NgBR regulates CCM1/2 expression has not been elucidated. Here, we show that temporal genetic depletion of NgBR in ECs at both the postnatal and adult stages results in CCM1/2 expression deficiency and consequently CCMs-like lesions such as enlarged vessels, blood-brain barrier (BBB) hyperpermeability, and intracerebral hemorrhage. These cerebrovascular defects in the brain of NgBR endothelial-specific knockout (ecKO) mice can be rescued by adeno-associated virus (AAV)-mediated overexpression of CCM1 and CCM2 genes. To reveal the molecular mechanism, we used RNA-seq analysis to examine changes in the transcriptome. Surprisingly, we found that acetyltransferase HBO1 was downregulated in NgBR deficient ECs. The mechanistic study elucidated that NgBR is required for maintaining the expression of CCM1/2 in ECs via HBO1-mediated histone acetylation. ChIP-qPCR data further demonstrated that loss of NgBR impairs the binding of both the HBO1 and acetylated H4K5/K12 at the promoter of CCM1 and CCM2 genes. Similarly, AAV-mediated overexpression of HBO1 restores the acetylation of H4K5/K12 and rescues the CCMs-like cerebrovascular defects in the brain of NgBR ecKO mice. Our findings on epigenetic regulation of CCM1 and CCM2 provide a perspective that NgBR and HBO1-mediated histone H4 acetylation may be targeted for preventing the onset of CCMs-like cerebrovascular disease.

Cerebral cavernous malformations (CCMs) are enlarged leaking vascular lesions in the brain caused by loss-of-function mutations in CCM1/2/3 genes or loss of expression. Although we previously showed that Nogo-B receptor (NgBR) knockout in endothelial cells (ECs) results in CCMs-like cerebrovascular lesions in the mouse embryo, the molecular mechanism by which NgBR regulates CCM1/2 expression has not been elucidated. Here, we show that temporal genetic depletion of NgBR in ECs at both the postnatal and adult stages results in CCM1/2 expression deficiency and consequently CCMs-like lesions such as enlarged vessels, blood-brain barrier (BBB) hyperpermeability, and cerebral hemorrhage. These cerebrovascular defects in the brain of NgBR endothelial-specific knockout (ecKO) mice can be rescued by adeno-associated virus (AAV)-mediated overexpression of CCM1 and CCM2 genes. To reveal the molecular mechanism, we used RNA-seq analysis to examine changes in the transcriptome. Surprisingly, we found that acetyltransferase HBO1 was downregulated in NgBR deficient ECs. The mechanistic study elucidated that NgBR is required for maintaining the expression of CCM1/2 in ECs via HBO1-mediated histone acetylation. ChIP-qPCR data further demonstrated that loss of NgBR impairs the binding of both the HBO1 and acetylated H4K5/K12 at the promoter of CCM1 and CCM2 genes. Similarly, AAV-mediated overexpression of HBO1 restores the acetylation of H4K5/K12 and rescues the CCMs-like cerebrovascular defects in the brain of NgBR ecKO mice. Our findings on epigenetic regulation of CCM1 and CCM2 provide a perspective that NgBR and HBO1-mediated histone H4 acetylation may be targeted for preventing the onset of CCMs-like cerebrovascular disease.

The triglyceride (TG) transfer activity of microsomal triglyceride transfer protein (MTP) is essential for lipoprotein assembly in the liver and intestine; however, its function in adipose tissue, which does not assemble lipoproteins, is unknown. Here we have elucidated the function of MTP in adipocytes.We demonstrated that MTP is present on lipid droplets in human adipocytes. Adipose-specific MTP deficient (A-MttpOur findings demonstrate that adipose-specific MTP deficiency increases ATGL-mediated TG lipolysis and enhances energy expenditure, thereby resisting diet-induced obesity. We speculate that the regulatory function of MTP involving protein-protein interactions might have evolved before the acquisition of TG transfer activity in vertebrates. Adipose-specific inhibition of MTP-ATGL interactions may ameliorate obesity while avoiding the adverse effects associated with inhibition of the lipid transfer activity of MTP.

Reviewed here are multiple mouse models of vertical sleeve gastrectomy (VSG) and Roux-en Y gastric bypass (RYGB) that have emerged over the past decade. These models use diverse approaches to both operative and perioperative procedures. Scrutinizing the benefits and pitfalls of each surgical model and what to expect in terms of post-operative outcomes will enhance our assessment of studies using mouse models, as well as advance our understanding of their translational potential. Two mouse models of bariatric surgery, VSG-lembert and RYGB-small pouch, demonstrate low mortality and most closely recapitulate the human forms of surgery. The use of liquid diets can be minimized, and in mice, RYGB demonstrates more reliable and longer lasting effects on weight loss compared to that of VSG.

Adipose tissue dysfunction is the primary defect in obesity which contributes to the development of dyslipidemia, insulin resistance, cardiovascular diseases, type 2 diabetes, nonalcoholic fatty liver disease and some cancers. Previous study from our lab implicated the role of lipocalin-type prostaglandin D2 synthase (L-PGDS) in glucose intolerance and atherosclerosis. Our lab also showed the development of NAFLD in L-PGDS knockout mice on both low fat and high fat diet. In the present study, we examined the role of L-PGDS in adipose tissue in response to high fat diet. We found decreased expression of L-PGDS in visceral and subcutaneous white adipose tissue (WAT) of C57BL/6 mice after 12 weeks on high fat diet (HFD). Our study showed increased fat content in L-PGDS KO mice as compared to control C57BL/6 mice after 14 weeks on HFD. Histological analysis showed hypertrophied adipocytes in visceral WAT of L-PGDS KO mice as compared to control mice. Furthermore, we found increased expression of the lipogenesis genes LXRα, SREBP-1c and FAS in visceral WAT of L-PGDS KO mice after 14 weeks on HFD. L-PGDS KO mice showed reduced gene expression of adiponectin and PPARγ in visceral WAT on HFD. The plasma adiponectin level was also decreased in L-PGDS KO mice. We conclude that absence of L-PGDS has a deleterious effect on adipose tissue functioning which further reduces insulin sensitivity in adipose tissue. Thus, our study suggests L-PGDS as a novel therapeutic target for the treatment of obesity-associated metabolic syndrome. Disclosure A. Srivastava: None. T. Palaia: None. C. Hall: None. J. Lee: None. M. Stevenson: None. L. Ragolia: None. Funding American Heart Association (15GRNT22420001); George Link, Jr. Foundation Inc. (3500-7294)

The loss function of cerebral cavernous malformation (CCM) genes leads to most CCM lesions characterized by enlarged leaking vascular lesions in the brain. Although we previously showed that NOGOB receptor (NGBR) knockout in endothelial cells (ECs) results in cerebrovascular lesions in the mouse embryo, the molecular mechanism by which NGBR regulates CCM1/2 expression has not been elucidated. Here, we show that genetic depletion of Ngbr in ECs at both postnatal and adult stages results in CCM1/2 expression deficiency and cerebrovascular lesions such as enlarged vessels, blood-brain-barrier hyperpermeability, and cerebral hemorrhage. To reveal the molecular mechanism, we used RNA-sequencing analysis to examine changes in the transcriptome. Surprisingly, we found that the acetyltransferase HBO1 and histone acetylation were downregulated in NGBR-deficient ECs. The mechanistic studies elucidated that NGBR is required for maintaining the expression of CCM1/2 in ECs via HBO1-mediated histone acetylation. ChIP-qPCR data further demonstrated that loss of NGBR impairs the binding of HBO1 and acetylated histone H4K5 and H4K12 on the promotor of the CCM1 and CCM2 genes. Our findings on epigenetic regulation of CCM1 and CCM2 that is modulated by NGBR and HBO1-mediated histone H4 acetylation provide a perspective on the pathogenesis of sporadic CCMs.

Background Problem-based learning (PBL) offers a unique model of remote learning that resembles important facets of students’ future medical careers. The purpose of this study was to determine if online PBL was successful and to identify creative innovations and best practices of online learning in order to better prepare medical students for health care careers in the future. Methods The impact of implementing virtually delivered PBL sessions on student experience and performance in comparison to in-person PBL sessions was evaluated. A 4-point Likert scale question was used to investigate the effect of the transition to online PBL on the students’ PBL process experience. Student PBL process grades were measured on a scale of 5–10, where 5 is “needs substantial improvement” and 10 is “meets expectations”. PBL session grades were compared in one session before and one session after the transition to online PBL using a paired t-test. Data are presented as number of responses per category (percentage), or as a mean +/- standard deviation. Results Based on our experiences, we identified a successful transition to online PBL with no meaningful impact on students’ performance. We also identified several challenges to online PBL and how students demonstrated creative solutions to overcome these challenges. As a result of our experience, we identified several key factors as best practices of implementing online PBL to ensure a smooth transition from in-person sessions for both faculty and students, including extensive preparation, participation of support staff, availability of adequate internet services and equipment, and input from instructional technology and design personnel. Conclusion Successful student performance and experience, along with best practices learned from implementing virtual PBL, provide valuable tools for improving teaching and critical insight into the challenges faced, when preparing students for global health care changes in the future.

The prevalence of obesity has increased steadily over the last few decades and is a major risk factor for type 2 diabetes and cardiovascular disease. Obesity is associated with alterations in adipose tissue functioning leading to metabolic dysfunction. In previous studies, we have reported the role of lipocalin-type prostaglandin D2 synthase (L-PGDS) in glucose intolerance and atherosclerosis. More recently, we demonstrated that L-PGDS KO mice develop NAFLD even on a low fat diet. Our present study explored the role of L-PGDS in adipose tissue. We found decreased expression of L-PGDS in insulin resistant 3T3-L1 adipocytes. Enzymatic inhibition of L-PGDS in 3T3-L1 adipocytes using AT56 impaired insulin signaling by reducing the phosphorylation of AKT protein and the expression and translocation of GLUT4 to plasma membrane. Furthermore, we found that inhibition of L-PGDS reduces the expression of adiponectin and AMPK phosphorylation in 3T3-L1 adipocytes. Rosiglitazone treatment in insulin resistant 3T3-L1 adipocytes significantly increased L-PGDS expression. Treatment with rosiglitazone restored the expression of adiponectin and phosphorylation of AMPK in AT56 treated 3T3-L1 adipocytes. Correspondingly, we found decreased L-PGDS expression in visceral fat of obese humans. Cumulatively, these data confirm that the absence of L-PGDS results in decreased insulin sensitivity via impaired insulin signaling and reduced adiponectin levels in 3T3-L1 adipocytes. Thus, our findings suggest that L-PGDS plays a significant role in maintaining the insulin sensitivity in 3T3-L1 adipocytes and provides a novel opportunity for therapeutic intervention of obesity-associated metabolic syndrome. Disclosure A. Srivastava: None. T. Palaia: None. J. Lee: None. C. Hall: None. M. Stevenson: None. A.G. Fernando: None. L. Ragolia: None. Funding American Heart Association (15GRNT22420001); George Link, Jr. Foundation, Inc.

There is controversy over whether African Americans have higher vitamin D requirements than recommended by the Institute of Medicine. We previously reported that maintaining serum 25(OH)D above 30 ng/mL does not prevent age-related bone loss. Herein, we report that bone strength is also unaffected by maintaining this level in this population. The role of vitamin D in bone strength has not been investigated in the African American (AA) population. A 3-year randomized controlled trial was designed to examine the effect of vitamin D supplementation on physical performance, bone loss, and bone strength in healthy older AA women. A total of 260 postmenopausal AA women, ages ≥ 60 years were randomized to a vitamin D3 or placebo arm. Vitamin D3 dose was adjusted to maintain serum 25OHD > 30 ng/mL. Bone mineral density, femoral axis length, and femoral neck (FN) width were measured by dual-energy X-ray absorptiometry. Composite indices of FN strength [compression strength index (CSI), bending strength index (BSI), and impact strength index (ISI)] were computed. The mean age of participants was 68.2 ± 4.9 years. Baseline characteristics between groups were similar. The average dose of vitamin D3 was 3490 ± 1465 IU/day in the active group. The mean serum 25OHD was 46.8 ± 1.2 ng/mL versus 20.7 ± 1.1 ng/mL in the active versus placebo group. Serum 25OHD did not correlate with any composite indices. The longitudinal differences observed in FN width, CSI, BSI, and ISI in both groups were not statistically significant (all p values > 0.05). Further, there was no group × time interaction effect for any of the composite indices (all p values > 0.05). Maintaining serum 25OHD > 30 ng/mL (75 nmol/L) does not affect bone strength in older AA women. There is no evidence to support vitamin D intake greater than the recommended RDA by the Institute of Medicine in this population for bone strength.

Objectives To examine the effect of 25-hydroxyvitamin D (25(OH)D) levels recommended by Endocrine Society guidelines (>30 ng/mL) on cognition in healthy older African-American women over 3 years. Design Randomized, double-blind, placebo-controlled clinical trial. Setting Bone Mineral Research Center at New York University Winthrop Hospital. Participants Healthy postmenopausal African American women aged 65 and older (N=260; mean age 68.2 ± 4.9; 46% college education or higher). Intervention Half of the women were randomized to receive vitamin D (adjusted to achieve a serum level > 30 ng/mL) with calcium (diet and supplement total of 1,200 mg), and half were randomized to receive placebo with calcium (1,200 mg). Measurements Cognitive assessments every 6 months using the Mini-Mental State Examination (MMSE) to detect cognitive decline. Mean MMSE scores were calculated over time for both groups. Those with MMSE scores less than 21 at baseline were excluded. Results The average dose of vitamin D3 was 3,490 ± 1,465 IU per day, and average serum 25(OH)D at 3 years was 46.8 ± 1.2 ng/mL in the active group and 20.7 ± 1.1 ng/mL in the placebo group. Serum 25(OH)D concentration was maintained at greater than 30 ng/mL in 90% of the active group. Over the 3-year period, MMSE scores increased in both groups (p Conclusion There was no difference in cognition over time between older African-American women with serum concentrations of 25(OH)D of 30 ng/mL and greater than those taking placebo. There is no evidence to support vitamin D intake greater than the recommended daily allowance in this population for preventing cognitive decline. J Am Geriatr Soc 67:81-86, 2019.

Rationale Vitamin D deficiency is associated with bone loss, poor muscle strength, falls and fracture. This information in older African Americans (AAs) is sparse. Objective The study of the relationship of P hysical performance, O steoporosis prevention with vitamin D in older A frican Americans (PODA) is a randomized, double-blind, placebo-controlled 3-year trial examining the effect of vitamin D on bone loss and physical performance in older AA women. Methods 260 healthy AA women aged > 60 years were assigned to receive placebo or vitamin D 3 . Initial vitamin D 3 dose was determined by the baseline serum 25OHD level, and adjusted further to maintain serum 25OHD between 30 and 69 ng/ml. Subjects with baseline 25OHD levels ≤ 8 ng/ml or ≥ 26 ng/ml were excluded. Objective measures of neuromuscular strength [Short Physical Performance Battery (SPPB), grip strength and 6-minute walking distance (6MWD)] and bone mineral density (BMD) were obtained. Results SPPB gait speed, grip strength and 6MWD showed a significant positive correlation with free 25OHD. 1 pg/ml increase in free 25OHD predicted a 32% increase in the odds of having higher gait speed and a 1.42 lb. increase in grip strength. No significant differences in BMI, BMD, muscle mass, grip strength, serum total 25OHD and free 25OHD were observed between groups. None of the measures of physical performance showed an association with baseline serum 25OHD. Conclusions This is the first study to show an association between free 25OHD and physical performance. These findings indicate a positive relationship of free 25OHD with gait speed and grip strength in older AA women. Further studies are needed to understand the role of free 25OHD.

Non-alcoholic fatty liver disease (NAFLD) is an emerging risk factor for type 2 diabetes mellitus, cardiovascular disease, and all-cause mortality. Previously, we demonstrated that lipocalin-type prostaglandin D

The transcription factor aryl hydrocarbon receptor nuclear translocator-like protein-1 (BMAL1) is an essential regulator of the circadian clock, which controls the 24-h cycle of physiological processes such as nutrient absorption. To examine the role of BMAL1 in small intestinal glucose absorption, we used differentiated human colon adenocarcinoma cells (Caco-2 cells). Here, we show that BMAL1 regulates glucose uptake in differentiated Caco-2 cells and that this process is dependent on the glucose transporter sodium-glucose cotransporter 1 (SGLT1). Mechanistic studies show that BMAL1 regulates glucose uptake by controlling the transcription of SGLT1 involving paired-homeodomain transcription factor 4 (PAX4), a transcriptional repressor. This is supported by the observation that clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated endonuclease Cas9 (Cas9) knockdown of PAX4 increases SGLT1 and glucose uptake. Chromatin immunoprecipitation (ChIP) and ChIP-quantitative PCR assays show that the knockdown or overexpression of BMAL1 decreases or increases the binding of PAX4 to the hepatocyte nuclear factor 1-α binding site of the SGLT1 promoter, respectively. These findings identify BMAL1 as a critical mediator of small intestine carbohydrate absorption and SGLT1.

Nonalcoholic fatty liver disease (NAFLD) is an emerging risk factor for type 2 diabetes mellitus (T2DM), cardiovascular disease, and all-cause mortality. Obesity is a major determinant of the high prevalence of NAFLD and T2DM worldwide. Previously, we demonstrated that lipocalin-type prostaglandin D2 synthase (L-PGDS) knockout mice show increased glucose intolerance and accelerated atherosclerosis. In the present study, we investigated the role of L-PGDS in causing NAFLD in L-PGDS knockout (KO) and control C57BL/6 mice (n=6/group) on both low fat and high fat diets for 14 weeks. We observed that L-PGDS KO mice remain slightly lighter in weight compared to control mice, yet develop NAFLD faster, even on the low fat diet. In addition, we found elevated fasting glucose and insulin levels in L-PGDS KO mice with increased lipid accumulation in the liver with time on both diets as compared to controls. We also found total cholesterol and LDL levels were significantly elevated in L-PGDS KO mice. Finally, lipogenesis marker proteins such as SREBP-1c and LXRα, and CD36, a fatty acid transporter protein, were also found to be increased in L-PGDS KO mice on both diets as compared to control mice. These data confirm that absence of L-PGDS results in hyperinsulinemia and dyslipidemia. Therefore, we conclude that L-PGDS plays a significant role in developing NAFLD independent of obesity and may serve as a novel therapeutic target for the treatment of NAFLD. Disclosure A. Srivastava: None. T. Palaia: None. C. Hall: None. J. Lee: None. M. Stevenson: None. L. Ragolia: None. Funding George Link, Jr. Foundation, Inc.; American Heart Association

Purpose: Bone Mineral density (BMD) is a predictor of fracture risk. Beyond BMD, the role of vitamin D for bone strength is not known in African American (AA) population. Objective: To examine the effect of vitamin D supplementation on femoral neck (FN) strength. Design and Setting: A 3-year randomized, double-blind, placebo-controlled clinical trial. Participants: 260 healthy postmenopausal AA women aged 65 years and older. Main Outcome Measures: Femoral axis length and width were measured by dual-energy x-ray absorptiometry. We combined these variables with BMD, body weight and height and computed composite indices of FN strength: compression strength index (CSI), bending strength index (BSI), and impact strength index (ISI). Results: Mean age of participants was 68.2 ± 4.9 years. Baseline characteristics between groups were similar. The average dose of vitamin D3 was 3490 ± 1465 IU/day. Serum 25(OH)D concentration was maintained at above 30 ng/mL in the active group. The average serum 25(OH)D at the 3-year period was 46.8 ± 1.2 ng/mL in the active group compared to 20.7 ± 1.1 ng/mL in the placebo group. Linear mixed effects models demonstrated a significant linear increase in FN width, BSI and ISI over time in both active and placebo group (all p values 0.05). Conclusion: These findings indicate no treatment effect of vitamin D on bone strength in African Americans. Maintaining serum 25(OH)D above 30 ng/ml did not affect bone strength. There is no evidence to support vitamin D intake greater than the recommended RDA by the Institute of Medicine in this population for bone health.

Background There is a paucity of data describing the effects of coronavirus disease 2019 on placental pathology, especially in asymptomatic patients. Although the pathophysiology of coronavirus disease 2019 is not completely understood, there is emerging evidence that it causes a severe systemic inflammatory response and results in a hypercoagulable state with widespread microthrombi. We hypothesized that it is plausible that a similar disease process may occur in the fetal-maternal unit. Objective This study aimed to determine whether coronavirus disease 2019 in term patients admitted to labor and delivery, including women without coronavirus disease 2019 symptomatology, is associated with increased placental injury compared with a cohort of coronavirus disease 2019–negative controls. Study Design This was a retrospective cohort study performed at NYU Winthrop Hospital between March 31, 2020, and June 17, 2020. During the study period, all women admitted to labor and delivery were routinely tested for severe acute respiratory syndrome coronavirus 2 regardless of symptomatology. The placental histopathologic findings of patients with coronavirus disease 2019 (n=77) who delivered a singleton gestation at term were compared with a control group of term patients without coronavirus disease 2019 (n=56). Controls were excluded if they had obstetrical or medical complications including fetal growth restriction, oligohydramnios, hypertension, diabetes, coagulopathy, or thrombophilia. Multivariable logistic regression models were performed for variables that were significant (P Results In univariable analyses, coronavirus disease 2019 cases were more likely to have evidence of fetal vascular malperfusion, that is, presence of avascular villi and mural fibrin deposition (32.5% [25/77] vs 3.6% [2/56], P Conclusion Despite the fact that all neonates born to mothers with coronavirus disease 2019 were negative for severe acute respiratory syndrome coronavirus 2 by polymerase chain reaction, we found that coronavirus disease 2019 in term patients admitted to labor and delivery is associated with increased rates of placental histopathologic abnormalities, particularly fetal vascular malperfusion and villitis of unknown etiology. These findings seem to occur even among asymptomatic term patients.

Background Glucagon-like peptide-1 (GLP-1) level was significantly increased post Vertical Sleeve Gastrectomy (VSG), an effect believed to contribute to its beneficial cardiometabolic effects. Objective To validate the beneficial GLP-1 mediated cardiometabolic effects post VSG using GLP-1 antagonist (exendin 9-39) in Zucker diabetic fatty rats. Methods Animals were divided into three (n = 5) groups: (i) sham, (ii) VSG, and (iii) VSG received exendin 9–39 (GLP-1 receptor antagonist). The study was performed over 12 weeks and parameters were measured 12 weeks post-surgery. Results and discussion As expected, fasting blood glucose and insulin levels were improved post VSG due to enhanced GLP-1 secretion. However, both fasting glucose and insulin levels were impaired in the presence of GLP-1 antagonist. Baseline total cholesterol level pre-surgery was 100±1 mg/dl which remained unchanged in the VSG group but significantly increased to 140±8 mg/dl in the presence of antagonist. Interestingly, post-surgery there was a nearly 70% reduction in triglyceride level in the VSG group compared to sham which was overcome in the presence of antagonist. Myographic studies using aortic rings showed no significant change between groups. Additionally, blood pressure and heart rate also remained unchanged in all groups. Serum bile acid and L-PGDS levels increased post VSG but significantly decreased in the presence of antagonist, suggesting a strong association with GLP-1 and a novel mechanism of action. Conclusion Enhanced GLP-1 secretion post VSG imparted beneficial cardiometabolic effects on blood glucose, insulin, total cholesterol, triglyceride, bile acids and L-PGDS levels which were abated in the presence of GLP-1 antagonist., Highlights • GLP-1 increases post-VSG 30 min after glucose load. • Post-VSG GLP-1 secretion is associatged with lower cholesterol and triglycerides. • Bile acids and L-PGDS increase post-VSG and are inhibited in the presence of GLP-1 antagonist. • Heart rate, blood pressure and myograph profile remain unchanged.

Background Vertical sleeve gastrectomy (VSG) ameliorates metabolic complications in obese and diabetic patients through unknown mechanisms. Objective The objective of this study was to investigate the role of lipocalin-type prostaglandin D 2 synthase (L-PGDS) in glucose regulation in response to VSG using L-PGDS knock-out (KO), knock-in (KI), and C57BL/6 (wild type) mice. Setting Winthrop University Hospital Research Institute. Methods Animals were divided into 6 groups: L-PGDS KO sham/VSG (n = 5), L-PGDS KI sham/VSG (n = 5), and C57BL/6 (wild type) sham/VSG (n = 5). Related parameters were measured in fasting animals after 10 weeks. Results Our intraperitoneal glucose tolerance tests and homeostatic model assessment insulin resistance results showed significant glycemic improvement 10 weeks post-VSG in both C57BL/6 and KI groups compared with the sham group. In contrast, the KO group developed glucose intolerance and insulin resistance similar to or greater than the sham group 10 weeks post-VSG. Interestingly, weight gain was insignificant 10 weeks post-VSG in all the groups and even trended higher in the KO group compared with sham. Peptide YY levels in the KO group post-VSG were slightly increased but significantly less than other groups. Similarly, the KO group showed significantly less leptin sensitivity in response to VSG compared with the KI group. Total cholesterol level remained unchanged in all groups irrespective of sham or surgery but interestingly, the KO group had significantly higher cholesterol levels. In parallel, adipocyte size was also found to be significantly increased in the KO group post-VSG compared with the sham group. Conclusion Our findings propose that L-PGDS plays an important role in the beneficial metabolic effects observed after VSG.

Non-alcoholic fatty liver disease (NAFLD) is an emerging risk factor for type 2 diabetes mellitus, cardiovascular disease, and all-cause mortality. Previously, we demonstrated that lipocalin-type prostaglandin D2 synthase (L-PGDS) knockout mice show increased glucose intolerance and accelerated atherosclerosis. In the present study, we investigated the role of L-PGDS in mediating NAFLD utilizing L-PGDS knockout (KO) and control C57BL/6 mice fed either low fat (LFD) or high fat diet (HFD) for 14 weeks. Our present study demonstrates that L-PGDS KO mice remain slightly lighter in weight compared to control mice, yet develop NAFLD faster and eventually progress to the more severe non-alcoholic steatohepatitis (NASH). We found increased lipid accumulation in the liver of KO mice over time on both diets, as compared to control mice. The L-PGDS KO mice showed elevated fasting glucose and insulin levels and developed insulin resistance on both LFD and HFD. Lipogenesis marker proteins such as SREBP-1c and LXRα were increased in L-PGDS KO mice after 14 weeks on both diets, when compared to control mice. We replicated our in vivo findings in vitro using HepG2 cells treated with a combination of free fatty acids (oleic and palmitic acid) and exposure to a L-PGDS inhibitor and prostaglandin D2 receptor (DP1) antagonists. We conclude that the absence or inhibition of L-PGDS results in dyslipidemia, altered expression of lipogenesis genes and the acceleration of NAFLD to NASH, independent of diet and obesity. We propose L-PGDS KO mice as a useful model to explore the pathogenesis of NAFLD and NASH, and L-PGDS as a potential therapeutic target for treatment.

Context There is limited information on the influence of vitamin D on physical performance in black Americans. Objective To determine if maintenance of serum 25(OH)D >75 nmol/L prevents a decline in physical performance. Design The Physical Performance, Osteoporosis and Vitamin D in African American Women (PODA) trial had a prospective, randomized, placebo controlled, double-dummy design with two arms: one of which is placebo vitamin D3 adjusted to maintain serum 25(OH)D >75 nmol/L. Patients The target population was healthy elderly black women with serum 25(OH)D between 20 and 65 nmol/L. The trial was 3 years in duration with measurement of physical performance every 6 months: grip strength, Short Physical Performance Battery (SPPB), 10 chair rises, and 6-minute walk distance. A total of 260 women entered the study and 184 completed 3 years. Mean age was 68.2 years. Baseline 25(OH)D was 53 nmol/L; total SPPB was 11 (10 to 12). Setting Research center in an academic health center. Main outcomes measure Prevention of decline in physical performance measures. Intervention Participants were randomly assigned to placebo or active vitamin D. Vitamin D3 dose was adjusted to maintain serum 25(OH)D >75 nmol/L. Results There was a decline with time in grip strength and the 6-minute walk test. The SPBB increased with time. There were no substantial differences between the placebo and active vitamin D3 groups with respect to the temporal patterns observed for any of the performance measures. Conclusion There is no benefit of maintaining serum 25(OH)D >75 nmol/L in preventing the decline in physical performance in healthy black American women.

Background Limited information is available on the influence of vitamin D on falls in older high-functioning black American women. Endocrine Society guidelines propose serum 25(OH)D levels over 30 ng/mL. Objective To determine if maintenance of serum 25(OH)D above 30 ng/mL protects against falls. Design The Physical Performance, Osteoporosis and Vitamin D in African American Women (PODA) trial had a prospective, randomized, placebo-controlled, double-dummy design with two arms: one with placebo and one with vitamin D3 adjusted to maintain serum 25(OH)D above 30 ng/mL. The primary outcomes were the prevention of bone loss and the decline in physical performance. Patients The target population was healthy black women older than 60 years with serum 25(OH)D between 8 and 26 ng/mL. The trial was 3 years in duration with a falls questionnaire administered every 3 months. A total of 260 women entered the study, and 184 completed the 3 years. Mean age was 68.2 years. Setting Research center in an academic health center. Main outcomes measure Prevention of falls. Intervention Participants were randomly assigned to placebo or active vitamin D. Vitamin D3 dose was adjusted to maintain serum 25(OH)D above 30 ng/mL in the active group using a double-dummy design. Results Baseline 25(OH)D was 22 ng/mL. Mean serum 25(OH)D reached 47 ng/mL in the active group compared with 21 ng/mL in the placebo group. There were 14.2% falls in the previous year recalled at baseline. During the study, 46% reported falling in the treatment group compared with 47% in the placebo group. There was no association of serum 25(OH)D or vitamin D dose with the risk of falling. Conclusions There is no benefit of maintaining serum 25(OH)D above 30 ng/mL compared with the Institute of Medicine recommendation (20 ng/mL) in preventing falls in healthy older black American women. J Am Geriatr Soc 67:1043-1049, 2019.

Hepatosteatosis is strongly associated with hyperinsulinaemia and obesity. Previously, we demonstrated accelerated atherosclerosis and impaired glucose tolerance in L-PGDS knockout (KO) mice. Interestingly, L-PGDS KO mice also exhibit hyperinsulinemia on high fat diet in a time-dependent manner. In this study, we investigated the role of L-PGDS on liver pathophysiology using L-PGDS KO mice compared to control C57BL/6 mice) (n=6/group). Mice were kept on high fat diet (60% fat) for 14 weeks and all parameters were measured initially and at 4-, 8- and 14-weeks post high fat diet. Data were analyzed by unpaired t-test or one-way ANOVA where appropriate with p Disclosure S. Kumar: None. T. Palaia: None. C. Hall: None. J. Lee: None. M. Stevenson: None. L. Ragolia: None.

Background: Lipocalin prostaglandin D2 synthase (LPGDS) is a bile acid binding protein that has been found to have a significant role in glucose metabolism and appetite. In performing sleeve gastrectomy (SG) and gastric bypass in rodent models, we have also demonstrated LPGDS appears to have an important role in the beneficial metabolic effects seen after surgery. We have sought to correlate if LPGDS has a significant clinical effect in humans for surgical weight loss. Methods: Serum LPGDS levels were obtained in patients undergoing SG before, and then one month following surgery. Serum LPGDS was also measured in subjects undergoing a very low calorie diet (VLCD) program before, and then one month following the start of the program. LPGDS levels were measured by ELISA and analyzed. Results: Subjects that underwent surgical loss went from 304.45 +/-74 pounds before surgery to 271.4 +/- 70 pounds one month following surgery. Subjects that underwent VLCD went from 273.1 +/- 65 pounds before the program to 255 +/-61 pounds one month after starting the program. LPGDS levels for surgical weight loss increased from 579.9 +/-161 ng/mL before the program to 694.6 +/- 448 ng/mL one month after surgery. LPGDS for VLCD was 583.1 +/- 187 ng/mL before the program and 598.4 +/- 165 ng/mL one month after starting the program. Conclusion: LPGDS levels appear to increase after sleeve gastrectomy but not VLCD, although this trend is not statistically significant. Further study is needed to confirm this trend. Disclosure R.G. Lau: None. S. Kumar: None. J. Lee: None. T. Palaia: None. S. Islam: None. C. Brathwaite: None. L. Ragolia: None.

Black Americans have lower levels of serum 25(OH)D but superior bone health compared to white Americans. There is controversy over whether they should be screened for vitamin D deficiency and have higher vitamin D requirements than recommended by the Institute of Medicine (IOM). The purpose of this trial was to determine whether Vitamin D supplementation in elderly black women prevents bone loss. A total of 260 healthy black American women, 60 years of age and older were recruited to take part in a two-arm, double-dummy 3-year randomized controlled trial (RCT) of vitamin D3 versus placebo. The study was conducted in an ambulatory clinical research center. Vitamin D3 dose was adjusted to maintain serum 25(OH)D above 75 nmol/L. Bone mineral density (BMD) and serum were measured for parathyroid hormone (PTH), C-terminal crosslink telopeptide (CTX), and bone-specific alkaline phosphatase (BSAP) every 6 months. Baseline serum 25(OH)D3 was 54.8 ± 16.8 nmol/L. There was no group × time interaction effect for any BMD measurement. For all BMD measurements, except for total body and spine, there was a statistically significant negative effect of time (p < 0.001). An equivalency analysis showed that the treatment group was equivalent to the control group. Serum PTH and BSAP declined, with a greater decline of PTH in the treatment group. The rate of bone loss with serum 25(OH)D above 75 nmol/L is comparable to the rate of loss with serum 25(OH)D at the Recommended Dietary Allowance (RDA) of 50 nmol/L. Black Americans should have the same exposure to vitamin D as white Americans. © 2018 American Society for Bone and Mineral Research.

It has been proposed that serum free 25-hydroxyvitamin D [25(OH)D] may better reflect vitamin D action than total 25(OH)D. An ELISA for serum free 25(OH)D has recently become available, permitting direct assay.To determine whether serum free 25(OH)D provides additional information in relation to calcium absorption and other biomarkers of vitamin D action compared to total serum 25(OH)D.Ambulatory research setting in a teaching hospital.Serum free 25(OH)D measured in a previously performed study of varied doses of vitamin D3 (placebo and 800, 2000, and 4000 IU) on calcium absorption, PTH, procollagen type 1 N-terminal propeptide, and C-terminal telopeptides of type I collagen. Free 25(OH)D was measured by ELISA. Calcium absorption was measured at baseline and at 10 weeks using stable dual calcium isotopes.Seventy-one subjects completed this randomized, placebo-controlled trial. Baseline group mean free and total 25(OH)D varied from 4.7 ± 1.8 to 5.4 ± 1.5 pg/mL, and from 23.7 ± 5.9 to 25.9 ± 6.1 ng/mL, respectively. Participants assigned to the 4000-IU dose arm achieved free 25(OH)D levels of 10.4 pg/mL and total 25(OH)D levels of 40.4 ng/mL. Total and free 25(OH)D were highly correlated at baseline and after increasing vitamin D dosing (r = 0.80 and 0.85, respectively). Free 25(OH)D closely reflected changes in total 25(OH)D. PTH was similarly correlated at baseline and follow-up with total and free 25(OH)D. Serum C-terminal telopeptides of type I collagen had a moderate positive correlation with total and free 25(OH)D at follow-up. The serum 1,25-dihydroxyvitamin D change increased significantly with the change in 25(OH)D but not with the change in free 25(OH)D.There was no advantage from measuring free over total 25(OH)D in assessing the response of calcium absorption, PTH, and markers of bone turnover to vitamin D. Free 25(OH)D responded to increasing doses of vitamin D in a similar fashion to total 25(OH)D.

Diabetes is associated with disturbances in the normal levels of both insulin and glucagon, both of which play critical roles in the regulation of glycemia. Recent studies have found lipocalin-type prostaglandin D2 synthase (l-PGDS) to be an emerging target involved in the pathogenesis of type-2 diabetes. This study focused on the effect of l-PGDS on glucagon secretion from cultured pancreatic Alpha TC-1 Clone 6 cells. When cells were treated with various concentrations of l-PGDS (0, 10, 50, and 100 ug/ml) for 2 h in 1 mM glucose; glucagon secretion decreased to 670±45, 838±38, 479±11, and 437±45 pg/ml, respectively. In addition, pancreatic islets were isolated from C57BL/6 mice and stained for prostaglandin D2 receptors, DP1 and DP2, using immunohistochemistry. Our results showed that these islets express only the DP1 receptor. Pancreatic islets were then stained for alpha and beta cells, as well as DP1, to find the primary location of the receptor within the islets using immunofluorescence. Interestingly, DP1 receptor density was found primarily in alpha cells rather than in beta cells. Our study is the first to report a correlation between l-PGDS and glucagon secretion in alpha cells. Based on our obtained results, it can be concluded that higher concentrations of l-PGDS significantly reduced the secretion of glucagon in alpha cells, which may contribute to the pathogenesis of diabetes as well as offer a novel therapeutic site for the treatment of diabetes., Highlights • Alpha TC1 Clone 6) derived from C57BL/6 mice express DP1 receptor. • High [l-PGDS] significantly reduce the secretion of glucagon in alpha cells. • DP1 receptor density is found primarily in alpha cells rather than beta cells.

African Americans have a lower total serum 25-hydroxyvitamin D [25(OH)D] but superior bone health. This has been referred to as a paradox. A recent publication found that free serum 25(OH)D is the same in black and white individuals. However, the study was criticized because an indirect method was used to measure free 25(OH)D. A direct method has recently been developed.We hypothesized that although total serum 25(OH)D is lower in African Americans, free serum 25(OH)D measured directly would not differ between races.White and black healthy postmenopausal women were matched for age and body mass index. Serum total 25(OH)D, PTH, 1,25-dihydroxyvitamin D, vitamin D binding protein (VDBP), and bone density were measured. Measurement of free 25(OH)D was carried out using an ELISA.The study was conducted at an ambulatory research unit in a teaching hospital.A cross-racial comparison of serum free 25(OH)D was performed.A propensity match resulted in the selection of a total of 164 women. Total 25(OH)D was lower in black women (19.5 ± 4.7 vs 26.9 ± 6.4 ng/mL), but a direct measurement of free 25(OH)D revealed almost identical values (5.25 ± 1.2 vs 5.25 ± 1.3 ng/mL) between races. VDBP was significantly lower in blacks when using a monoclonal-based ELISA but higher with a polyclonal-based ELISA. Serum PTH, 1,25-dihydroxyvitamin D, and bone density were higher in African Americans.Free serum 25(OH)D is the same across races despite the lower total serum 25(OH)D in black women. Results comparing VDBP between races using a monoclonal vs a polyclonal assay were discordant.

Inflammation plays a crucial role in neurodegenerative diseases, but the irritants responsible for this response remain largely unknown. This report addressed the hypothesis that hypochlorous acid reacts with dopamine to produce melanic precipitates that promote cerebral inflammation. Spectrophotometric studies demonstrated that nM amounts of HOCl and dopamine react within seconds. A second-order rate constant for the reaction of HOCl and dopamine of 2.5 × 10(4)M(-1)s(-1) was obtained by measuring loss of dopaminergic fluorescence due to HOCl. Gravimetric measurements, electron microscopy, elemental analysis, and a novel use of flow cytometry confirmed that the major product of this reaction is a precipitate with an average diameter of 1.5 μm. Flow cytometry was also used to demonstrate the preferential reaction of HOCl with dopamine rather than albumin. Engulfment of the chlorodopamine particulates by phagocytes in vitro caused these cells to release TNFα and die. Intrastriatal administration of 10(6) particles also increased the content of TNFα in the brain and led to a 50% loss of the dopaminergic neurons in the nigra. These studies indicate that HOCl and dopamine react quickly and preferentially with each other to produce particles that promote inflammation and neuronal death in the brain.

The objective of the study was to investigate the role of prostaglandin D2 during pregnancy and its mediator Lipocalin-type prostaglandin D2 synthase (L-PGDS) as a predictor of preterm birth (PTB). Transgenic L-PGDS (+/+), L-PGDS (-/-) and C57BL/6 control pregnant mice models were used to determine the effect of DP1 and DP2 receptor antagonists in lipopolysaccharide (LPS)-induced PTB mice. In addition, L-PGDS levels were measured in the cervicovaginal secretions (CVS) of 370 pregnant women using ELISA and further processed for isoform detection using 2-D gel electrophoresis. Our results found that C57BL/6 control mice (n = 26), transgenic L-PGDS (+/+) (n = 26), demonstrated an 89% and 100% preterm birth in LPS (intraperitoneal injection, 20mg/kg) induced mice model respectively. Interestingly, the incidence of PTB was significantly reduced to 40% in L-PGDS (-/-) knockout mice (n = 26). DP1 and DP2 receptor antagonists (0.264 μg/day, dose of 0.1 μg/μl with the flow of 0.11 μl/h for 28 day using Alzet pumps) were used to investigate the effect in LPS-induced PTB in C57BL/6 mice and found 3.3-fold increase in viable pups after LPS-induction. In addition, L-PGDS levels were measured in CVS samples and found that PTB women (n = 296) had two-fold higher levels compared to full term births (n = 74) and established a significant inverse correlation between levels of L-PGDS and days to expected delivery by using 370 preterm birth CVS samples. Elevated L-PGDS levels in the CVS of women may be considered as a potential biomarker for PTB in future. Secondly, the use of DP1 and DP2 receptor antagonists may represent novel tocolytic agents for the treatment of PTB.

Summary Background/objective Lipocalin Prostaglandin D2 synthase (LPGDS) contributes to the production of PGD2, which has been associated with adipogenesis. In this study, we aimed to investigate the role of PGD2 on obesity through its DP1 and DP2 receptor signaling using intraperitoneal injection of their respective agonists and antagonists. Methods ApoE−/− mice were divided into five groups: vehicle control (n = 5), DP1 receptor agonist (n = 5), DP1 receptor antagonist (n = 5), DP2 receptor agonist (n = 5), and DP2 receptor antagonist (n = 5), and the study was carried out for 10 weeks. Results Despite being on high fat diet, mice receiving DP1 receptor agonist sustained a significant inhibition of weight gain throughout the study gaining only 11.4% body weight compared to the controls gaining 61% body weight. Interestingly, parallel to the body weight, the DP1 receptor agonist group showed a significant reduction in food intake throughout the study. Consistently, fasting leptin, insulin and bile acids levels were elevated in the DP1 receptor agonist group compared to controls. As expected, there was a significant reduction in fasting glucose level in DP1 receptor agonist group. At last, as a result of weight gain inhibition, DP1 receptor agonist also imparted cardiovascular benefits showing significant reduction in aortic wall thickness, intima, adventia and lumen size. Conclusion Based on the obtained results, we believe DP1 receptor agonism inhibited diet induced weight gain possibly through controlling appetite which consequently imparted beneficial cardiometabolic effects. DP1 receptor agonism may represent a novel therapeutic target for the management of obesity.

Background: The maximal calcium absorption in response to vitamin D has been proposed as a biomarker for vitamin D sufficiency. Objective: The objective was to determine whether there is a threshold beyond which increasing doses of vitamin D, or concentrations of serum 25-hydroxyvitamin D [25(OH)D], no longer increase calcium absorption. Design: This was a placebo-controlled, dose-response, randomized, double-blind study of the effect of vitamin D on calcium absorption in healthy postmenopausal women. Seventy-six healthy postmenopausal women were randomly assigned to placebo or 800 IU (20 mg), 2000 IU (50 mg), or 4000 IU (100 mg) vitamin D3 for 8 wk. The technique of dual isotopes of stable calcium was used with a calcium carrier to measure calcium absorption at baseline and after 8 wk. Results: Seventy-one women with a mean 6 SD age of 58.8 6 4.9 y completed the study. The mean calcium intake was 1142 6 509 mg/d and serum 25(OH)D was 63 6 14 nmol/L at baseline. A statistically significant linear trend of an increase in calcium absorption adjusted for age and body mass index with increasing vitamin D3 dose or serum 25(OH)D concentration was observed. A 6.7% absolute increase in calcium absorption was found in the highest vitamin D3 group (100 mg). No evidence of nonlinearity was observed in the dose-response curve. Conclusions: No evidence of a threshold of calcium absorption was found with a serum 25(OH)D range from 40 to 130 nmol/L. Calcium absorption in this range is not a useful biomarker to determine nutritional recommendations for vitamin D. This trial was registered at clinicaltrials.gov as NCT01119378. Am J Clin Nutr 2014;99:624‐31.

Vertical sleeve gastrectomy (VSG) ameliorates metabolic complications in obese and diabetic patients through unknown mechanisms.The objective of this study was to investigate the role of lipocalin-type prostaglandin DWinthrop University Hospital Research Institute.Animals were divided into 6 groups: L-PGDS KO sham/VSG (n = 5), L-PGDS KI sham/VSG (n = 5), and C57BL/6 (wild type) sham/VSG (n = 5). Related parameters were measured in fasting animals after 10 weeks.Our intraperitoneal glucose tolerance tests and homeostatic model assessment insulin resistance results showed significant glycemic improvement 10 weeks post-VSG in both C57BL/6 and KI groups compared with the sham group. In contrast, the KO group developed glucose intolerance and insulin resistance similar to or greater than the sham group 10 weeks post-VSG. Interestingly, weight gain was insignificant 10 weeks post-VSG in all the groups and even trended higher in the KO group compared with sham. Peptide YY levels in the KO group post-VSG were slightly increased but significantly less than other groups. Similarly, the KO group showed significantly less leptin sensitivity in response to VSG compared with the KI group. Total cholesterol level remained unchanged in all groups irrespective of sham or surgery but interestingly, the KO group had significantly higher cholesterol levels. In parallel, adipocyte size was also found to be significantly increased in the KO group post-VSG compared with the sham group.Our findings propose that L-PGDS plays an important role in the beneficial metabolic effects observed after VSG.

Obesity and diabetes are closely associated with hyperactivation of the hypothalamic–pituitary–adrenal (HPA) axis. In this study, the diet-induced obese C57BL/6 mouse was used to test the hypothesis that chronically elevated metabolic parameters associated with the development of obesity such as cholesterol and glucose can aggravate basal HPA axis activity. Because the lipocalin-type prostaglandin D2synthase (L-PGDS) knockout (KO) mouse is a model of accelerated insulin resistance, glucose intolerance, and obesity, it was further hypothesized that HPA activity would be greater in this model. Starting at 8 weeks of age, the L-PGDS KO and C57BL/6 mice were maintained on a low-fat or high-fat diet. After 20 or 37 weeks, fasting metabolic parameters and basal HPA axis hormones were measured and compared between genotypes. Correlation analyses were performed to identify associations between obesity-related chronic metabolic changes and changes in the basal activity of the HPA axis. Our results have identified strong positive correlations between total cholesterol, LDL-cholesterol, glucose, and HPA axis hormones that increase with age in the C57BL/6 mice. These data confirm that obesity-related elevations in cholesterol and glucose can heighten basal HPA activity. Additionally, the L-PGDS KO mice show early elevations in HPA activity with no age-related changes relative to the C57BL/6 mice.

A local melanocortin system is active during tissue injury and inflammation. Thus far this system has been described as autocrine in nature where local production of pro-opiomelanocortin (POMC) peptides by leukocytes feeds back on melanocortin receptor (MC-R) expressing immune cells to quell inflammatory cytokine production. Here we present evidence that POMC peptides may generate extracellular matrix (ECM) changes by inducing matrix production by cells of the mesenchymal lineage through activation of the MC2-R. Using immunoblot, we determined that mouse aorta-derived mesenchymal progenitor cells express both MC2-R and MC3-R. These progenitors respond to treatment with ACTH by increasing collagen matrix synthesis as assessed by picrosirius red stain and (3)H-proline incorporation. ACTH also induces transient increases in intracellular calcium ([Ca(2+)](i)) as assessed using the fluorescent Ca(2+) indicator, fura-2. The ACTH-induced changes in [Ca(2+)](i) are consistent with MC2-R signaling and consist of both an intracellular release and an extracellular influx of Ca(2+). Both mouse aortic mesenchymal progenitors and mouse macrophage cells express POMC and the prohormone convertase 1/3 (PC1/3) indicating they have the potential to contribute to the local production of POMC peptides. These data demonstrate functional MC2-R expression in mouse aorta-derived mesenchymal progenitors and implicate both macrophage and mesenchymal cells as relevant sources of local POMC peptides.

Background. Endothelial Microparticles (EMPs) are small vesicles shed from activated or apoptotic endothelial cells and involved in cellular cross-talk. Whether EMP immunophenotypes vary according to stimulus in Diabetes Mellitus (DM) is not known. We studied the cellular adhesion molecule (CAM) profile of circulating EMPs in patients with and without Diabetes Mellitus type 2, who were undergoing elective cardiac catheterization.Methods and Results. EMPs were analyzed by flow cytometry. The absolute median number of EMPs (EMPs/L) specific for CD31, CD105, and CD106 was significantly increased in the DM population. The ratio of CD62E/CD31 EMP populations reflected an apoptotic process.Conclusion. Circulating CD31+, CD105+, and CD106+ EMPs were significantly elevated in patients with DM. EMPs were the only independent predictors of DM in our study cohort. In addition, the EMP immunophenotype reflected an apoptotic process. Circulating EMPs may provide new options for risk assessment.

Lipocalin-type prostaglandin D(2) synthase (L-PGDS) is a highly glycosylated protein found in several body fluids. Elevated L-PGDS levels have been observed in the serum of patients with renal impairment, diabetes mellitus, and hypertension. Recently, we demonstrated the ability of L-PGDS to induce apoptosis in a variety of cell types including epithelial cells, neuronal cells, and vascular smooth muscle cells (VSMCs). The aim of this study was to investigate the effect several site-directed mutations had on L-PGDS-induced apoptosis in order to identify potential sites of regulation. Point mutations created in a glycosylation site (Asn51), a protein kinase C phosphorylation site (Ser106), and the enzymatic active site (Cys65) all inhibited L-PGDS-induced apoptosis as determined by both terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labeling (TUNEL) and caspase3 activity. We also compared the L-PGDS isoforms present in GK rat serum to WKY control serum using two-dimensional gel electrophoresis and observed distinct differences which vanished after PNGase F glycolytic digestion. We conclude that post-translational modification of L-PGDS, by either glycosylation or phosphorylation, enhances its apoptotic activity and inhibits VSMC hyperproliferation and postulate that this process is altered in type 2 diabetes.

Background Roux-en-Y gastric bypass (RYGB) may improve cardiometabolic risk through alteration of bile acids and L-PGDS levels. Objective The objective of this study was to investigate the effect of RYGB on aortic wall thickness, in relation to bile acid and L-PGDS metabolism. Methods Zucker diabetic fatty (ZDF) rats were divided into two groups, ad lib (n = 4), and RYGB (n = 6). Bile acid and L-PGDS were measured presurgery and fourteen weeks post-surgery. Results Elevation of bile acid levels following RYGB in Zucker Diabetic Fatty (ZDF) rodents was observed, as compared to ad lib . RYGB in ZDF rodents led to a significantly decreased aortic wall thickness (25%) as compared to ad lib control. Although bile acid metabolism is implicated in these alterations, other mediators are likely involved. Our laboratory has demonstrated lipocalin prostaglandin D2 synthase (L-PGDS) is a kno n cardiometabolic modulator that also functions as a bile acid binding protein. Therefore, L-PGDS levels were measured and a significant elevation was observed with RYGB compared to ad lib control. Conclusion Based on these findings, RYGB showed beneficial effect on aortic wall thickness, possibly through bile acids and L-PGDS elevation in a severely obese and diabetic rodent model.

Lipocalin-type prostaglandin D2 synthase (L-PGDS) has recently been linked to a variety of pathophysiological cardiovascular conditions including hypertension and diabetes. In this study, we report on the 50% increase in L-PGDS protein expression observed in vascular smooth muscle cells (VSMC) isolated from spontaneously hypertensive rats (SHR). L-PGDS expression also increased 50% upon the differentiation of normotensive control cells (WKY, from Wistar-Kyoto rats). In addition, we demonstrate differential effects of L-PGDS treatment on cell proliferation and apoptosis in VSMCs isolated from SHR versus WKY controls. L-PGDS (50 microg/ml) was able to significantly inhibit VSMC proliferation and DNA synthesis and induce the apoptotic genes bax, bcl-x, and ei24 in SHR but had no effect on WKY cells. Hyperglycemic conditions also had opposite effects, in which increased glucose concentrations (20 mm) resulted in decreased L-PGDS expression in control cells but actually stimulated L-PGDS expression in SHR. Furthermore, we examined the effect of L-PGDS incubation on insulin-stimulated Akt, glycogen synthase kinase-3beta (GSK-3beta), and ERK phosphorylation. Unexpectedly, we found that when WKY cells were pretreated with L-PGDS, insulin could actually induce apoptosis and failed to stimulate Akt/GSK-3beta phosphorylation. Insulin-stimulated ERK phosphorylation was unaffected by L-PGDS pretreatment in both cell lines. We propose that L-PGDS is involved in the balance of VSMC proliferation and apoptosis and in the increased expression observed in the hypertensive state is an attempt to maintain a proper equilibrium between the two processes via the induction of apoptosis and inhibition of cell proliferation.

Recent evidence suggests that glycogen-associated protein phosphatase 1 (PP-1(G)) is essential for basal and exercise-induced glycogen synthesis, which is mediated in part by dephosphorylation and activation of glycogen synthase (GS). In the present study, we examined the potential role of site-specific phosphorylation of PP-1(G) in heat-shock-induced glycogen synthesis. L6 rat skeletal-muscle cells were stably transfected with wild-type PP-1(G) or with PP-1(G) mutants in which site-1 (S1) Ser(48) and site-2 (S2) Ser(67) residues were substituted with Ala. Cells expressing wild-type and PP-1(G) mutants, S1, S2 and S1/S2, were examined for potential alterations in glycogen synthesis after a 60 min heat shock at 45 degrees C, followed by analysis of [(14)C]glucose incorporation into glycogen at 37 degrees C. PP-1(G) S1 mutation caused a 90% increase in glycogen synthesis on heat-shock treatment, whereas the PP-1(G) S2 mutant was not sensitive to heat stress. The S1/S2 double mutant was comparable with wild-type, which showed a 30% increase over basal. Heat-shock-induced glycogen synthesis was accompanied by increased PP-1 and GS activities. The highest activation was observed in S1 mutant. Heat shock also resulted in a rapid and sustained Akt/ glycogen synthase kinase 3 beta (GSK-3 beta) phosphorylation. Wortmannin blocked heat-shock-induced Akt/GSK-3 beta phosphorylation, prevented 2-deoxyglucose uptake and abolished the heat-shock-induced glycogen synthesis. Muscle glycogen levels regulate GS activity and glycogen synthesis and were found to be markedly depleted in S1 mutant on heat-shock treatment, suggesting that PP-1(G) S1 Ser phosphorylation may inhibit glycogen degradation during thermal stimulation, as S1 mutation resulted in excessive glycogen synthesis on heat-shock treatment. In contrast, PP-1(G) S2 Ser phosphorylation may promote glycogen breakdown under stressful conditions. Heat-shock-induced glycogenesis appears to be mediated via phosphoinositide 3-kinase/Akt-dependent GSK-3 beta inactivation as well as phosphoinositide 3-kinase-independent PP-1 activation.

Background Roux-en-Y gastric bypass (RYGB) ameliorates type 2 diabetes (T2DM) and obesity through alteration in gastrointestinal (GI) hormones. Objective The objective of this study was to investigate the effect of RYGB on GI hormones and cardiometabolic parameters in Zucker diabetic fatty (ZDF) rodents. Setting Winthrop University Hospital, Research and Academic Center Methods Animals were divided into 3 groups, pair-fed (n = 4), ad lib (n = 4), and RYGB (n = 5). This study was carried out for 4 weeks and all related parameters were measured pre- and postsurgery in fasted obese diabetic Zucker rodents. Results Postoperatively, RYGB significantly decreased fasting blood glucose by 32% compared with ad lib. Plasma insulin and leptin levels were also found to be significantly decreased, by 66% and 38%, respectively, after surgery. Moreover, both glucose-dependent insulinotropic polypeptide (GIP) and peptide tyrosine-tyrosine (PYY) were significantly increased after RYGB—by 300% and 51%, respectively. Glucagon-like peptide-1 (GLP-1) levels were also increased, but the increase was not statistically significant. Total cholesterol levels of the RYGB group remained unchanged for 4 weeks. However, total cholesterol in the ad lib and pair-fed groups increased by 25% and 34%, respectively, compared with initial levels. The cholesterol/high-density lipoprotein (HDL) ratio was decreased in the RYGB group by 14% and 30% compared with the ad lib and pair-fed group, respectively. The RYGB group had a significant decrease in aortic wall thickness of 25% compared with the ad lib and pair-fed groups. Similarly, the RYGB group had a 20-unit (mm Hg) decrease in systolic blood pressure compared with the presurgical value. Conclusion RYGB has beneficial cardiometabolic effects through alterations in GI hormones in a severely obese and diabetic rodent model.

The steroid receptor coactivator 1 (SRC1) regulates key metabolic pathways, including glucose homeostasis. SRC1−/− mice have decreased hepatic expression of gluconeogenic enzymes and a reduction in the rate of endogenous glucose production (EGP). We sought to determine whether decreasing hepatic and adipose SRC1 expression in normal adult rats would alter glucose homeostasis and insulin action. Regular chow-fed and high-fat-fed male Sprage-Dawley rats were treated with an antisense oligonucleotide (ASO) against SRC1 or a control ASO for 4 wk, followed by metabolic assessments. SRC1 ASO did not alter basal EGP or expression of gluconeogenic enzymes. Instead, SRC1 ASO increased insulin-stimulated whole body glucose disposal by ∼30%, which was attributable largely to an increase in insulin-stimulated muscle glucose uptake. This was associated with an approximately sevenfold increase in adipose expression of lipocalin-type prostaglandin D2 synthase, a previously reported regulator of insulin sensitivity, and an approximately 70% increase in plasma PGD2 concentration. Muscle insulin signaling, AMPK activation, and tissue perfusion were unchanged. Although GLUT4 content was unchanged, SRC1 ASO increased the cleavage of tether-containing UBX domain for GLUT4, a regulator of GLUT4 translocation. These studies point to a novel role of adipose SRC1 as a regulator of insulin-stimulated muscle glucose uptake.

Objective: Atherosclerosis contributes to 30% of deaths in systemic lupus erythematosus (SLE) and cardiovascular mortality is 50% higher in rheumatoid arthritis (RA) than in the general population....

Prostaglandin D 2 synthase induces apoptosis in pig kidney LLC-PK1 cells. Background Prostaglandin D 2 synthase (PGD 2 S), a unique member of the lipocalin family, is found at elevated levels in the serum of patients with renal impairment and has recently been implicated as a new biochemical marker of renal insufficiency. The aim of this study was to investigate the apoptotic effects of PGD 2 S on a pig kidney epithelial cell line (LLC-PK1) and to investigate the effects of prostaglandins and growth factors on this process. Methods Apoptosis was detected by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labeling (TUNEL), annexin V staining, and electron microscopy. Results A four- to fivefold increase in apoptosis was observed in PGD 2 S-treated cells as compared with controls and the apoptosis appeared to act via caspase-3. A cyclooxygenase-2 inhibitor, anti-PGD 2 S antibody, and selenium all significantly inhibited the apoptosis induced by PGD 2 S; however, none had any effect on the apoptosis induced by the known apoptotic inducer camptothecin. Furthermore, prostaglandins E 1 and E 2 , known to induce mitogen-activated protein (MAP) kinase phosphorylation and exhibit cytoprotective effects, both inhibited PGD 2 S-induced apoptosis, while prostaglandin H 2 had no significant effect. Growth factors such as insulin, insulin-like growth factor-1, and platelet-derived growth factor also decreased PGD 2 S-induced apoptosis. In addition, PGD 2 S isolated from human serum seemed slightly more effective at inducing apoptosis than recombinantly expressed protein. Conclusions We report on the induction of apoptosis by PGD 2 S in LLC-PK1 pig kidney epithelial cells, and speculate that the accumulation of PGD 2 S in the serum of kidney failure patients may further exacerbate renal problems and is most likely regulated by other prostaglandins and growth factors.