bluuebunny/crossref_metadata_2025 huggingface dataset:
The J-Shaped Relationship Between Body Mass Index and Mortality in Patients With Covid-19: A Systematic Review and Dose-Response Meta-Analysis
Authors: Bukhari Khulood , Huang Huei-Kai , Hung Duan-Pei , Peng Carol Chiung-Hui , Shih Ming-Chieh , Chang Rachel Huai-En , Tu Yu-Kang , Munir Kashif M
Published: May 2021

Introduction: Several studies have linked obesity to more severe illness and higher mortality in COVID-19 patients. However, the relationship between being underweight and COVID-19 mortality remains inconclusive. Previous dose-response meta-analyses did not enroll or specifically analyze the underweight population. Herein, we conducted a systematic review and dose-response meta-analysis to investigate the relationship between body mass index (BMI) and mortality in both obese and underweight patients with COVID-19. Methods: We searched PubMed, Embase, Cochrane Library, Scopus, and Web of Science databases from inception until August 12, 2020 using the keywords “COVID-19,” “body mass index,” “obesity,” “overweight”, and “underweight.” Three reviewers independently assessed the relevant articles, including the title, abstract, and full text, to identify eligible studies. We performed a two-stage random-effects dose-response meta-analysis, including only studies with at least three quantitative classifications for BMI. The nonlinear trend was evaluated using a restricted cubic splines model with three-knots at the 10th, 50th, and 90th percentiles. A sensitivity analysis was conducted by pooling only those studies which specifically evaluated underweight patients (BMI<18.5 kg/m2). Results: Thirteen studies comprising 25,828 patients were included in the analysis. In the linear model, the mortality of patients with COVID-19 increased by 1.5% for each 1-kg/m2 increase in BMI (pooled relative risk [RR] =1.015, 95% confidence interval [CI] =1.004−1.027). However, a significant non-linear relationship between BMI and mortality was observed (Wald test: Pnon-linearity<0.001). We demonstrated a J-shaped curve, indicating that both underweight and obese patients had a higher mortality than those with normal weight. Interestingly, overweight patients (BMI, 25−30 kg/m2) seemed to have the lowest mortality risk. Using a BMI of 15 kg/m2 as the reference, the RRs of mortality decreased with BMI, and this trend continued until BMI of approximately 28 kg/m2 (RR=0.743, 95% CI=0.576−0.959). The relationship between BMI and mortality was then reversed, and an upward trend was observed when BMI exceeded 30 kg/m2; the RRs (95% CI) at BMIs 30, 35, 40, and 45 kg/m2 were 0.745 (0.570−0.974), 0.841 (0.643−1.100), 1.082 (0.850−1.377), and 1.457 (1.129−1.879), respectively. Conclusion: This study is the first dose-response meta-analysis that showed both underweight and obese COVID-19 patients are at higher risk of increased mortality. A J-curve relationship was demonstrated between BMI and COVID-19 mortality.
The Long-Term Safety and Efficacy of Metreleptin in Various Forms of Partial Lipodystrophy
Authors: Akinci Baris , Neidert Adam , Foss de Freitas Maria Cristina , Hench Rita , Oral Elif A
Published: May 2021

Metreleptin is an approved treatment for patients with generalized lipodystrophy but still under investigation for patients with partial forms of the disease in the US. This open-label single-arm study in partial lipodystrophy (PL) allowed continued treatment of patients who volunteered and completed treatment under our previous 1-year protocol and who derived significant clinical benefit. The primary outcome was percent change in fasting triglyceride levels and the secondary outcome was percent change in hemoglobin A1c levels (time frame: 5 years on metreleptin or last observation carried forward in both instances). The hypothesis was that these metabolic parameters would stay stable over the observation period. The study enrolled 11 participants (11 females; 3 ≤ 18 years; 8 between 18–65 years). Four patients had familial partial lipodystrophy type 2 (FPLD2) caused by LMNA pathogenic variants (R482Q in two subjects, R482W in one subject, and R349W in one subject), two patients had POLD1 associated PL (E1067K), four patients had FPLD1, and one patient had atypical PL. Variants of uncertain significance (VUS) in genes of interest were detected in two patients with FPLD1 and one patient with atypical PL. Four subjects completed 48 months (actual 60 months on metreleptin). Data from 3 subjects were excluded from final data analyses due to early loss to follow-up (n = 1), serious adverse event withdrawal (n = 1; development of persistent neutralizing antibody shortly after 18 months of drug exposure), and death (n = 1). The remaining 4 subjects withdrew from the study either through their own volition or due to the investigator decision (last observation carried forward in these 4 subjects). As expected, the median percent change from baseline triglyceride and hemoglobin A1c levels were not significant (55.76 percent (range: -66.79 to 203.80; p > 0.05), and 2.55 percent (range: -27.52 to 25.00; p > 0.05), respectively). Serious adverse events were reported in 8 of 11 patients and additional adverse events were observed in all patients. Apart from neutralizing antibody development in one subject causing substantial metabolic derangement, adverse events were considered not to be directly related to the study drug. Death was caused by a sudden cardiac event related to the specific molecular etiology (LMNA R349W) in one case. This interventional study provided valuable information on the efficacy and safety of long-term metreleptin in PL. Although nonsignificant fluctuations were observed in triglycerides, hemoglobin A1c levels were generally stable over time and the initial improvement seen in the first year were sustained. Adverse events were common but more likely due to the complex nature of the underlying disease. Although rare, neutralizing antibodies can develop in patients with PL treated with metreleptin.
Morbid Obesity Does Not Impact Mortality, Rate of Endoscopy in Patients With Biliary Acute Pancreatitis: A US Nationwide Analysis
Authors: Shaka Hafeez , Palomera-Tejeda Emmanuel , Achebe Ikechukwu , Asotibe Jennifer Chiagoziem , Pudasaini Garima , Essam Rashad , Warraich Muhammad-Sheharyar
Published: May 2021

Introduction: Morbid obesity (MO) is associated with increased mortality in various conditions including acute pancreatitis. Interventions are challenging in patients with MO due to higher prevalence of comorbidities that may affect airway and cardiopulmonary management. Biliary acute pancreatitis (BAP) is the most common etiology for acute pancreatitis in the US. Population-based studies on the effect of obesity on biliary acute pancreatitis are lacking. This study aimed to assess the impact of MO on outcomes of patients with BAP.

Methods: Data was obtained from the Nationwide Inpatient Sample database for 2016 and 2017. Hospital discharges of patients 18 years and over with a principal diagnosis of BAP were included. This cohort was divided based on BMI into nonobese patients (BMI <30) and morbidly obese (MO) patients (BMI >/=40.0). Patients with BMI between 30.0–39.9 were excluded. Primary outcome was inpatient mortality. Secondary outcomes included rate of endoscopic procedures, length of hospital stay (LOS), total hospital charges (THC), discharge diagnoses of hypocalcemia, septic shock, acute renal failure (AKI) and acute respiratory failure (ARF). Multivariate regression analysis was used to adjust for patients’ sociodemographic factors, Charlson comorbidity index as well as hospital characteristics as confounders.

Results: A total of 128995 hospitalizations were principally for BAP, with 75.7% and 12.0% of these patients classified as nonobese and MO respectively. There was a significantly higher proportion of females (66.1 vs 54.5%, p<0.001) and lower mean age (50.1 vs 58.7 years, p<0.001) in patients with MO. There was no significant difference in adjusted odds of mortality (aOR=1.34, 95% CI: 0.88 - 2.03, p=0.174), or rate of endoscopy (aOR 1.00 95% CI: 0.91 - 1.11, p=0.958), in MO compared with patients who were nonobese. However, MO patients had increased mean LOS of 0.8 days (95% CI: 0.5 - 1.0, p<0.001), increased mean THC of $10760 (95% CI: 7077 - 14442, p<0.001), increased odds of hypocalcemia (aOR=1.60, 95% CI: 1.22 - 2.09, p=0.001), septic shock (aOR=2.13, 95% CI: 1.39 - 3.25, p<0.001), and AKI (aOR=1.48, 95% CI: 1.30 - 1.68, p<0.001).

Conclusion: Even though we did not find any significative difference in mortality, patients with MO appear to have and increased LOS and THC, as well as more complications like septic shock, AKI, and hypocalcemia. This calls for a greater recognition of this association for further research studies and to recognize this potential association during clinical practice.
Refractory Hypoglycemia Related to Gastric Bypass Complicated by Hypothyroidism
Authors: Jain Akhil , Alukal Thomas J , Amakye Kingsley , Kagita Navyamani V D , Mustaqeem Ruqqiya , Alam Fahd , Jose Jose , Qiu Jack , Syed Asad , Sharma Sandeep
Published: May 2021

A 29-year-old African-American female was brought to the emergency department for sudden onset nausea, dizziness, and loss of consciousness at work. She was found to have low blood sugar and was fed with juice and crackers. She had history of similar episodes in the past, migraine headache, post-thyroidectomy hypothyroidism, seizures, generalized anxiety disorder, gastroesophageal reflux disease, and Roux-en-Y gastric bypass 9 months ago for morbid obesity related refractory Pseudotumor Cerebri (failed VP Shunting). She lost 41 kg, and had intolerance to foods from dumping syndrome. Her vital signs were within normal limits upon arrival to the ER. Laboratory parameters were remarkable for hypoglycemia of 52 mg/dL. She was immediately given dextrose intravenously. ECG showed sinus bradycardia. Though she regained consciousness, her blood sugar levels were persistently low, the lowest recording being 43 mg/dL. She was placed on a continuous D10 infusion. Further investigations revealed negative toxicology screens for 1st and 2nd generation sulfonylureas, and normal insulin levels at the time of the hypoglycemic event. Her TSH was found to be very high (99 nIU/mL). Her thyroglobulin (2.8 ng/mL), free T4 (0.25ng/dL), thyroglobulin antibody (99.18 uIU/mL), cortisol levels in morning (14.2 ug/mL), and evening (4.6 ug/dL) were found to be within normal range. Abdominal MRI did not reveal Insulinoma or any other pathology. Repeat ECG showed normal sinus rhythm with occasional PVCs. She was treated for nausea with ondansetron, promethazine, scopolamine, and dronabinol. Dextrose infusion was titrated down with rise in blood sugar levels. She confessed non-compliance to thyroid replacement. Therefore, she was initially treated with intravenous thyroxine replacement, which was later switched to oral therapy. Psychiatry evaluation ruled out any eating disorder. She was counselled to eat small frequent meals, increase dietary proteins, and restrict high glycemic index sugars. Educational value: National data indicates an increasing trend for admissions related to morbid obesity and bariatric procedures, especially laparoscopic (vertical) sleeve gastrectomy (see Fig 1). (1) Complex alteration in the signaling pathway to pancreas owing to changing metabolism causes hypoglycemia after bariatric surgery. When compounded by severe untreated hypothyroidism, as in our case, causes severe hypoglycemia needing hospitalization. Patient education for adjustment in dietary habits would have played a key role in preventing episodes of severe hypoglycemia.

Reference:

1.HCUPnet, Healthcare Cost and Utilization Project. Agency for Healthcare Research and Quality, Rockville, MD. https://hcupnet.ahrq.gov/.

Fig 1: Trends for admissions for Morbid Obesity and Sleeve Gastrectomy
Severe Early-Onset Obesity in Three Adult Patients Harboring Inactivating Mutations of the Leptin Receptor Gene
Authors: Chaves Carolina Marques , Kay Teresa , Anselmo Joao
Published: May 2021

Background: Leptin is secreted by white adipocytes in response to fat storage and binds to leptin receptors (LEPR) expressed all over the body particularly in hypothalamic neurons. This hormone regulates several physiologic functions including energy expenditure and appetite. Clinical Case: We describe the clinical and hormonal findings in 3 adult brothers with body mass index (BMI) of 36.7, 50.7, and 46.1 kg/m2, respectively. There is no history of consanguinity in the family and none of the patients exhibited dysmorphic features. A rapid weight gain was noticed during their first few months of life, associated with permanent hyperphagia. At 2 years old, their BMI was already above 25 kg/m2 (> +3SD), and at 10 years old, it was over 40 kg/m2 (> +3SD). The linear growth was within the expected target heights for their parents. They did not seem to present cognitive impairment. The pubertal development began at 16 to 18 years old, and since then they maintained levels and FSH and LH above the upper limit of normal (15.6±3.7mUI/mL and 12.3±2.2mUI/mL, reference range 1.5–12.4 and 1.7–8.6, respectively), but with normal sexual steroids (estradiol 36.4±16.1pg/mL and total testosterone 445±401ng/dL, reference range 11–44 and 249–836, respectively). The thyroid function was normal and none of the patients suffered from dyslipidemia or diabetes, despite high serum insulin levels 26.4±15.8 mU/L (normal 5–10). Genetic sequencing identified a homozygous mutation of the leptin receptor gene in the 3 brothers: c.2357T> C, p. (Leu786Pro). Their parents were heterozygous for the mutation as well as the patients’ daughters. Homozygous carriers of the mutation presented a significantly higher BMI than their heterozygous family members, 44.5±7.1 Kg/m2 vs 32.2±1.7 Kg/m2 (p=0.023), a significantly increased leptin levels, 80±36.4 ng/ml vs 26.3±9.3 ng/ml (p=0.028), and significantly higher weight, 134.6±16.9 vs 89.2±15,2 kg (p=0.021), respectively. Women had higher BMI than men (42.0 ±12.2 Kg/m2 vs 37.9±7.5 Kg/m2, p=0.496) and also higher percentage of fat (46.4±3.1% vs 34.9±6.8%, p=0.108). Serum levels of leptin in homozygous patients were not significantly higher than those measured in 10 patients with adult-onset morbid obesity, 80± 36.4 ng/ml vs 53.8±24.1 ng/ml, respectively (p=0.149). Therefore, serum leptin is not a useful discriminative maker of LEPR gene mutations. Conclusion: Patients with severe early-onset obesity should have a genetic diagnosis workup. Firstly, because clinical trials with MC4R-agonists have raised expectations regarding the treatment of patients with mutations of the LEPR gene. Secondly, and in contradiction to other reports in the literature, our patients were fertile. Therefore, identification of the mutation allows genetic counseling of these patients and their families, including the possibility of Pre-Natal Diagnosis or Pre-Implantation Genetic Diagnosis.
Endothelin-1 Receptor A Blockade Attenuates Metabolic and Proinflammatory Profile in Mice Fed a High Fat Diet
Authors: Rivera-Gonzalez Osvaldo , Taylor Erin , Speed Joshua S
Published: May 2021

Endothelin-1 (ET-1) is elevated in patients with obesity; however, its contribution to the pathophysiology related to obesity is not fully understood. Obesity is associated with dyslipidemia and insulin resistance, which may in part be mediated by inflammation and alterations to immune cell subsets within the adipose tissue. ET-1 promotes inflammation via the ET-1 type A (ETA) receptor, and blockade of ETA receptors improves dyslipidemia in patients with chronic kidney disease. We hypothesized that ET-1 causes dyslipidemia and inflammation within the adipose tissue of obese mice. To test this hypothesis, C57BL/6J mice were fed either normal diet (NMD) or high fat diet (HFD) for 8 weeks followed by 2 weeks of treatment with either vehicle or atrasentan (ETA receptor antagonist, 10mg/kg/day). HFD mice had significantly higher fat mass than NMD mice, with no significant effect of treatment with atrasentan. HFD mice had significantly higher circulating non-esterified free fatty acids, an effect that was ameliorated in mice treated with atrasentran (1.03±0.07 vs 0.58±0.02 mEq/L, p<0.05). Atrasentan-treated mice had significantly attenuated increase in liver triglycerides compared to non-treated HFD mice (3.8±0.7 vs 7.5±1.3mg/dL respectively, p<0.05). Mice treated with atrasentan had significantly improved glucose tolerance (10150±1031 vs 6563±975 AUC, p<0.05) and insulin tolerance (-2796±386 vs -9825±319 AUC, p<0.05) compared to non-treated insulin-resistant HFD mice. Plasma adiponectin, an insulin sensitizing adipokine that is inversely associated with adiposity and insulin resistance, was significantly increased in atrasentan-treated mice compared to non-treated HFD (4.8±0.1326 vs 6.5±0.3 µg/ml, p<0.05), with no differences in plasma insulin levels. Gene expression analysis of visceral fat showed improved expression of genes negatively associated with insulin resistance that were downregulated in non-treated HFD mice vs. NMD (IRS-1, PPAR-gamma, GLUT4, and adiponectin). Flow cytometric analyses of visceral adipose tissue indicated that HFD mice had a significantly higher number of both CD4+ and CD8+ T cells compared to NMD mice, which was attenuated by treatment with atrasentan. Further, eosinophils, which are important in maintaining adipose tissue health and reducing inflammation, were significantly decreased in HFD mice compared to NMD. Atrasentan treatment abolished the decrease in eosinophils. Taken together, these data indicate that ETA receptor blockade improves peripheral glucose homeostasis, dyslipidemia, and liver triglyceride levels, and also attenuates the proinflammatory immune profile in visceral adipose tissue. These data suggest a potential use for ETA receptor blockers in the treatment of obesity-associated dyslipidemia and insulin resistance.
AM833, a Long Acting Amylin Analogue Induces Hypocalcemia in Young Rats via a Calcitonin Receptor Mediated Mechanism
Authors: Dahl Kirsten , John Linu M , Skyggebjerg Rikke B , Vegge Andreas , Andersen Ninette K , Raun Kirsten
Published: May 2021

Amylin is a peptide hormone that is co-secreted with insulin from pancreatic β-cells. It is member of the calcitonin family and activates amylin receptors (AMYRs) in the calcitonin receptor (CTR) family. During recent years, several AMYR agonists have been in clinical development for treatment of diabetes and obesity. In contrast to pramlintide, which is a short acting amylin analogue approved for use in combination with meal-time insulin in patients with diabetes, the more recent development candidates have been dual acting analogues activating both the CTR and the AMYRs. In rodents the CTR plays a significant role in calcium homeostasis, while in humans this has been associated with little long-term effect on the plasma calcium levels (Ganong, 1993; Longo et al., 2011); Rodents are usually chosen as first choice pre-clinical models for in vivo testing and introducing acute and profound hypocalcaemia can lead to general neurological and neuromuscular disturbances, which can be challenging in clinical development programs. AM833 (NNC0174-0833) is a long acting acylated dual acting amylin analogue activating both CTR and AMYRs. When combined with the long acting GLP-1 analogue semaglutide for weight management, both preclinical studies in rat models of obesity and clinical trials in humans with obesity have demonstrated significant weight loss potential. In young rats aged 7–9 weeks, single sc dose of 3–300 nmol/kg AM833 induced a reduction of total and ionized calcium of 40–50% from baseline with lowest level around 12 hours after dosing. A similar response was seen after administration of salmon calcitonin which also activates both CTR and AMYRs. Tachyphylaxis was introduced, with no effect on calcium found after sub chronic daily treatment for 2 weeks. Age played a key role in the sensitivity to calcium lowering with almost no effect found in 11 months old rats. When administering AM1213 (NNC0174-1213), a selective AMYR agonist with little activation of the CTR, no or little calcium lowering of around 10% was seen. These data indicated that the calcium lowering properties are CTR mediated. The induction of hypercalcemia was a rodent specific phenomenon as no calcium lowering outside of normal range was seen in young dogs and rabbits. Clinical data confirmed that CTR induced calcium lowering was not human relevant, but these data showed that understanding physiology and pharmacology in the animal models of investigation as well as human translational relevance is of outmost importance in the progress of an early development program. References: GaNong WF. Calcitonin. In: Review of medical physiology. 16th ed. Appleton & Lange. 1993. Longo DL et al. Calcitonin. In: Harrison’s principles of internal medicine. 18th ed. McGraw-Hill; 2011.
Defining the Contribution of Adipocyte Subpopulations to Dermal White Adipose Tissue
Authors: Leyendecker Mara , Sharma Rita , Luong Quyen , Powell Noah , Harper Brian , Lee Kevin Y
Published: May 2021

Introduction: Our lab has previously identified three distinct subtypes of developmentally and functionally white adipocytes and have shown that they each differentially contribute to adipose depots (Lee, KY et al. EMBO J. 2019). Dermal white adipose tissue (dWAT), a layer of adipocytes embedded in the skin below the dermis, has recently been shown to play a role in crucial physiologic processes including thermogenesis, the regulation of aging, scar formation, and wound healing. The purpose of this proposal is to investigate the contribution of three adipocyte subtypes to dWAT. Objectives: The primary objectives of this project are to determine the number of preadipocytes and adipocytes from each of three subtypes present in dWAT. Methodology: Lineage tracing analysis was performed by crossing transgenic mice harboring cre-recombinase under the control of promoter/enhancer elements of each of the three marker genes, Wilms tumor 1, transgelin, or myoxovirus 1 to dual- fluorescent reporter mice. These three mice lines mark Type 1–3 preadipocytes and adipocytes, respectively. dWAT was collected from X week old mice, and adipocyte identities were determined by confocal microscopy. Preadipocyte contribution of these subpopulations was determined by FACS analysis. Results: We found that Type 2 (~45%) and Type 3 (~25%), but not Type 1 preadipocytes significantly contributed to the dWAT preadipocyte cellular population. We also found a similar pattern for the adipocyte populations. Type 1, 2, and 3 adipocytes were found to comprise ~3%, 17%, and 7% of mature adipocytes, respectively. These studies demonstrate that Type 2 and Type 3 adipocytes contribute to the composition of dWAT. Summary/Conclusion: These studies demonstrate that Type 2 and Type 3 adipocytes and preadipocytes significantly contribute to the composition of dWAT. Since these adipocyte subpopulations have different functional properties, including metabolism and response to inflammatory cytokines, the contribution of these adipocyte subtypes may impact the crucial physiologic processes mediated by dWAT.
Distinct Calcium Signaling for Wildtype, Loss-of-Function and Gain-of-Function Human MC4R Variants
Authors: Kumar Shree S , Mountjoy Kathleen Grace
Published: May 2021

There is compelling evidence for human melanocortin-4-receptor (hMC4R) playing a critical function regulating energy balance; yet signal transduction pathways contributing to this are unclear. The hMC4R activates multiple signaling pathways, including induced increases in cAMP and mobilization of intracellular calcium ([Ca2+]i). Recent evidence showed cAMP signaling was not a good predictor for hMC4R variant-associated obesity. We hypothesize that hMC4R mobilization of [Ca2+]i plays an important role in regulating energy balance. To test this, we developed a robust high-throughput Fura-2 ratiometric fluorescent assay to quantitatively measure [Ca2+]i in vitro. We compared basal and α-melanocyte stimulating hormone (α-MSH) activation of [Ca2+]i for hMC4R-wildtype (WT) and hMC4R-variants stably expressed in HEK293 cells. The loss-of-function variants studied were two obesity-associated variants (R7H and R18L) known to exhibit cAMP signaling similar to WT, two obesity-associated variants (H76R and L250Q) known to exhibit cAMP-constitutive activity (CA) compared to WT, and one overweight-associated variant (H158R) known to exhibit cAMP-CA compared to WT. The gain-of-function variants (V103I and I251L) studied are known to exhibit cAMP signaling similar to WT. The data for basal [Ca2+]i were pooled from three independent experiments performed with WT and all variants in each assay. Data (mean ± SEM) were analyzed using one-way ANOVA with Dunnett’s multiple comparisons. The data (mean ± SEM) for α-MSH activation of hMC4R were pooled from three independent experiments and analyzed using non-parametric sum of squares F-test for maximum best-fit values and EC50. The α-MSH activated assays were performed with each hMC4R variant alongside WT. WT hMC4R and non-CA loss-of-function variants exhibited similar basal and α-MSH activated [Ca2+]i (WT: EC50 = 1.44 nM; R7H: EC50 = 1.40 nM; R18L: EC50 = 1.12 nM). The CA loss-of-function variants exhibited significantly (p < 0.0001) increased basal [Ca2+]i compared with WT (WT = 97.6 ± 0.9 nM; H76R = 114.2 ± 1.7 nM; L250Q = 112.1 ± 2.6 nM; H158R = 110.7 ± 1.8 nM) and significantly lower EC50’s compared with WT (H76R: EC50 = 0.07 nM; p = 0.0019; L250Q: EC50 = 0.09 nM; p = 0.0066; H158R: EC50 = 0.14 nM; p = 0.0009). The gain-of-function hMC4R variants exhibited significantly (p < 0.0001) decreased basal [Ca2+]i compared with WT (WT = 97.6 ± 0.9 nM; V103I: = 86.4 ± 0.9 nM; I251L = 87.5 ± 1.0 nM) and significantly (p = 0.0001) increased α-MSH stimulated maximum [Ca2+]i compared with WT (WT = 224.5 ± 13.6 nM; V103I = 288.2 ± 31.5 nM; I251L = 295.6 ± 20.0 nM). To summarize, we show three distinct patterns of hMC4R-associated calcium signaling; (1) WT and non-CA loss-of-function, (2) CA loss-of-function and (3) non-CA gain-of-function. Future studies are required to understand how hMC4R mobilization of [Ca2+]i might contribute to the regulation of energy balance.
Elevated Serum Uric Acid Is a Facilitating Mechanism for Insulin Resistance Mediated Accumulation of Visceral Adipose Tissue
Authors: Fernandez-Chirino Luisa , Antonio-Villa Neftali Eduardo , Bello-Chavolla Omar Yaxmehen , Almeda-Valdes Paloma
Published: May 2021

Background: Serum uric acid (SUA) is related to cardiometabolic conditions such as insulin resistance (IR) and visceral adipose tissue (VAT) accumulation, which have a thoroughly explored bidirectional relationship. Here, we aimed to clarify the nature of the role uric acid plays inside this relationship, alongside the underlying causality mechanism.

Methods: We conducted a population-based cross-sectional study comprising 8,504 subjects from a joint cohort composed from both NHANES 2003–2004 and 2011–2012 cycles and ENSANUT Medio Camino 2016. We performed mixed effects linear regression models using HOMA2-IR, adipoIR, and METS-VF as indicators of both peripheral and adipose tissue IR and VAT accumulation, indicating the subject’s cohort of origin as a random effect. Furthermore, we performed multiple mediation analyses to assess a potential causal mechanism and ROC curves to establish cut-off points for identification of IR and visceral obesity using SUA. Finally, with an additional dataset comprised of 226 subjects with both euglycemic hyperinsulinemic clamp (EHC) and dual X-ray absorptiometry (DXA) measurements for IR and VAT accumulation, we performed a network of confirmatory mediation analyses including adiponectin measurements. Results: We found that SUA has a mediating role inside the bidirectional relationship between IR and visceral obesity, and it is part of an underlying causality mechanism which includes adiponectin. The proportion of the mechanism mediated by SUA is greater when stated that IR (in either peripheral or adipose tissue) leads to VAT accumulation (14.90%[13.20%-17.00%] and 15.54%[13.61%-18.00%]) instead of the opposite direction (4.88%[3.06%-7.00%] and 8.13%[5.91%-10.00%]). This result was strengthened by a mediation analysis network using the gold-standard measurements where we observed that the joint effect of SUA and adiponectin mediated 16.32% [8.84%-26.00%] for the effect of IR and VAT accumulation and 12.52% [3.23%-23.00%] in the opposite direction. Cut-off points for SUA to predict peripheral IR were 6.1 mg/dL and 4.8 mg/dL, for males and females respectively. For visceral obesity, cut-offs were 6.4 mg/dL and 4.8 mg/dL for males and females. SUA had a high negative predictive value for all assessments.

Conclusions: Elevated SUA acts as mediator inside the bidirectional relationship between IR and VAT accumulation. Its role appears to be larger when considering adipose tissue IR as the promoter for VAT accumulation.
HepatocyteGHR/STAT5b Signaling Protects Against Liver Injury in NAFLD/NASH Mice Models Independent of Steatosis
Authors: del Río-Moreno Mercedes , Vazquez-Borrego Mari C , Mahmood Mariyah , Sarmento-Cabral Andre , Guzman Grace , Subbaiah Papasani V , Cordoba-Chacon Jose , Kineman Rhonda Denise
Published: May 2021

Non-alcoholic fatty liver disease (NAFLD) represents a spectrum of pathologies ranging from simple steatosis to non-alcoholic steatohepatitis (NASH) that can lead to cirrhosis and hepatocellular carcinoma. Clinical and mouse studies indicate GH-signaling is reduced in NAFLD. We reported that chow-fed mice, with adult-onset, hepatocyte-specific GH receptor knockdown (aHepGHRkd) develop steatosis, and with age, a mild NASH-like phenotype. In the present study, we sought to determine if aHepGHRkd accelerates the development of steatosis and fibrosis in the context of diets shown in wild-type male mice, after 6 months of feeding, to produce mild NASH (60% fat [lard] + sucrose in the drinking water [HFS] or a severe NASH-like phenotype (40% fat, with partially hydrogenated corn-oil; 2% cholesterol; 20% fructose [HFCF]). Since aHepGHRkd is associated with a reduction in active STAT5b, aHepGHRkd mice were treated with either a hepatocyte-specific adenoviral-associated vector that expresses constitutively active STAT5b (AAV-TBGp-STAT5bCA = STAT5bCA) or a AAV-Null vector. After only 3 months of feeding either the HFS or HFCF diet, aHepGHRkd, but not GHR-intact controls, mice exhibited clear fibrosis, associated with higher levels of plasma alanine aminotransferase (ALT). STAT5bCA treatment of aHepGHRkd mice reduced fibrosis, as well as plasma ALT. Of note, hepatic TG content did not differ between the treatment groups, within diet. Preliminary studies used GC-MS to reveal aHepGHRkd, in the context of HFS diet, increased hepatic fatty acid ratios indicative of enhanced de novo lipogenesis, while STAT5bCA reversed this effect. These results suggest GHR/STAT5b may protect against liver injury not by controlling absolute fat accumulation, but by modifying the fatty acid composition of hepatic lipids. Finally, in order to determine if STAT5bCA could also reverse established diet-induced NASH, wild-type mice were fed the HFCF diet for 6 months and then treated with AAV-STAT5bCA or AAV-Null vectors, and followed for an additional 3 months. Preliminary findings show STAT5bCA modestly reduced liver weight with no changes in TG content. However, STAT5bCA prevented the rise in plasma ALT observed in Null-treated controls. Of note, some mice developed hepatic tumors, where the number and size of visible tumors was reduced by STAT5bCA. Importantly, in all models examined thus far, changes in the liver phenotype could not be clearly attributed to changes in systemic metabolism, supporting a direct action of GHR/STAT5b signaling on liver health. Taken together, these results suggest that enhancing hepatocyte STAT5b activity could prevent/treat diet-induced NASH. How STAT5b mediates these effects, and if there are other players involved, remains to be elucidated.
Prenatal Exposure to the Endocrine Disrupting Chemical DEHP Impacts Reproduction-Related Gene Expression in the Pituitary
Authors: Ge Xiyu , Weis Karen , Raetzman Lori
Published: May 2021

Phthalates are chemicals used in various common products including plastics and medical devices, leading to widespread contact. Phthalate exposure during embryonic development can cause changes in puberty timing, reduced fertility and genital abnormalities. Previous studies on prenatal exposure to Di-(2-ethylhexyl) phthalate (DEHP) in mice indicated that it disrupts pituitary-gonadal feedback and alters reproductive performance in the offspring, however, the mechanism behind this is unknown. We hypothesize that prenatal exposure to DEHP during a critical period of embryonic development (e15.5 to e18.5) will cause sex-specific disruptions in reproduction-related functions in the pituitary in offspring due to interference with androgen and aryl hydrocarbon receptor (AhR) signaling. In order to discover the direct effects of DEHP on the reproduction-related functions in the pituitary, we performed both in vivo dosing and in vitro pituitary culture experiments. First, we dosed pregnant CD-1 mice with corn oil, the antiandrogen flutamide or DEHP from gestational day 15.5 to 18.5, then collected the pituitaries of the offspring on postnatal day 0. We found that prenatal DEHP exposure caused a significant increase in Fshb specifically in males, and flutamide caused significant increases in both Lhb and Fshb in males. Besides, DEHP exposure significantly increased AhR pathway related gene Cyp1b1 in both males and females. In the in vitro experiment, we took whole pituitaries from e16.5 embryos and cultured them in media containing DEHP, MEHP and/or AhR antagonist for 72hrs. We found that the DEHP metabolite MEHP was actually the chemical that exerted the effects directly at the level of the pituitary. Similar to in vivo experiments, Cyp1a1 and Cyp1b1 mRNA level were increased in pituitaries treated with MEHP in both sexes and the induction could be reduced by co-treatment with AhR antagonist. The mRNA level of Lhb, Fshb and Gnrhr were significantly decreased in both sexes by MEHP and co-treatment with AhR antagonist did not restore mRNA levels. The induction of Cyp1a1/Cyp1b1 gene in both in vivo and in vitro experiments indicates the possible activation of AhR by DEHP/MEHP. The in vitro experiment with AhR antagonist further proved that the induction of Cyp1a1/Cyp1b1 was indeed due to AhR activation directly at the level of the pituitary. The difference between in vivo and in vitro experiments in terms of gonadotropin gene expression indicates multiple mechanisms should be involved in the regulation of gonadotropin gene expression in vivo including androgen-related pathways and possibly AhR-related pathways. In summary, our data suggest that phthalates can directly affect the function of the pituitary in terms of regulation of reproductive- related genes. This indicates that pituitary impacts of phthalates could contribute to reproductive dysfunction observed in exposed mice and humans.
Thyroid Gland and Male Reproductive Anomalies Among Fuel Handlers in Gampaha District, Sri Lanka
Authors: Kottahachchi Dulani Chathurika , Warnakularuriya Tania , De silva Deepthi , Madagoda Kushan , Wadasinghe Dilesha , Dayanath Bktpi , Siyambalapitiya Sajith , Devanarayana Niranga
Published: May 2021

Introduction:Fuel handlers at petrol stations are continuously exposed to organic solvents from fuel and vehicle emissions. Endocrine disrupting chemicals (EDC) are present in fuel, which are harmful to endocrine organs. Thyroiditis and hypogonadism are reported among fuel handlers. Thyroid gland and male reproductive function anomalies were investigated among fuel handlers in the Gampaha district of Sri Lanka. Method: 43 were recruited from 6 fuel stations in the Gampaha district for the study and 28 age matched male workers who were not exposed to fuel in an occupational setting were recruited as controls. Thyroid gland was examined clinically and TSH, free T4, FSH, LH and Testosterone were done on all the participants. TPO antibody and a thyroid scan was done on the fuel handlers. Results: Median (IQR) age was 38 years (27-46 years). The mean TSH value was 1.62 IU/mL (1.15-2.35) vs 1.33 IU/mL (0.83-1.79) respectively in study and control populations with significantly higher levels in the study population (p=0.023). The median (IQR) TSH value above the reference range was identified in 7% of fuel handlers and all controls were within the normal range, while 16.9% of fuel handlers had a derangement in the TPO levels. On examination, only one control had a small goiter but his T4 and TSH levels were normal. On ultrasound thyroid scans, benign nodules were seen in 2 fuel handlers. TPO levels did not correlate with the TSH levels among the fuel handlers (r=-0.078, p=0.652). Inability to sustain an erection was reported by 35.5% fuel handlers which was significantly higher than controls who reported 5.6% (p=0.019). Premature ejaculation was reported by 27.9% of fuel handlers which was significantly higher than controls (p=0.023). The testosterone levels were significantly higher among fuel handlers compared to controls (p=0.048). The FSH and LH levels positively correlated with each other as expected in each subgroup and the total population (p<0.005). The TSH levels significantly negatively correlated with the testosterone levels among the fuel handlers. (r=- 0.338, p=.0.023). When the fuel handlers with premature ejaculation was considered the FSH, LH, Testosterone levels were not significantly different between the two groups, however the duration of employment was significantly longer among those reporting premature ejaculation. (p=0.024). Conclusion: There are thyroid and reproductive abnormalities among those exposed to fuel in an occupational setting. Disturbances to sexual functions may also be related to alteration of autonomic functions. Limiting exposure to fuel vapor will eliminate these detrimental effects and we propose self-service fuel pumps to be the best alternative to avoid occupational health hazards among fuel handlers.
Tildacerfont for the Treatment of Patients With Classic Congenital Adrenal Hyperplasia: Results From a 12-Week Phase 2 Clinical Trial in Adults With Classic CAH
Authors: Auchus Richard Joseph , Merke Deborah P , Madu Ivy-Joan , Nakhle Samer , Sarafoglou Kyriakie , Huang Michael , Moriarty David , Barnes Chris , Newfield Ron S
Published: May 2021

Background: Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD) is an autosomal recessive disorder characterized by insufficient cortisol production resulting in excess adrenocorticotropic hormone (ACTH) and adrenal androgen production. Standard-of-care therapy with glucocorticoids (GC) is suboptimal due to the difficulty of balancing control of the ACTH-driven androgen excess against the serious long-term side effects associated with chronic supraphysiologic GC exposure. Tildacerfont, a second-generation corticotropin-releasing factor type-1 (CRF1) receptor antagonist, lowers excess ACTH, and thus has the potential to reduce adrenal androgen production and to allow for GC dosing closer to physiologic doses. A prior study demonstrated that tildacerfont was effective in reducing ACTH, 17-hydroxyprogesterone (17-OHP) and androstenedione (A4) after 2 weeks of therapy. Here we report results from an open-label 12-week extension study. Methods: Subjects met either of the following criteria: 1) completion of prior study or 2) treatment naïve to tildacerfont with 17-OHP >800 ng/dL while on a stable GC regimen (excluding dexamethasone). Subjects were treated with oral tildacerfont at 400 mg once daily for 12 weeks. Efficacy and safety parameters were assessed at baseline through Week 12. Results: Subject characteristics (n=8) are as follows: median (range) age was 44.5 years (26-67 years; 5 females), median (range) body mass index 30.8 kg/m2 (22-41 kg/m2). In month 3, in the participants with poor control of disease at baseline (elevations in all key biomarkers: ACTH, 17-OHP, and A4) (n=5), maximum mean percentage reductions for ACTH, 17-OHP and A4 were 84%, 82%, and 79%, respectively. In this subgroup, 60% of subjects achieved ACTH normalization and 40% achieved A4 normalization during treatment. Tildacerfont treatment maintained, and did not suppress, biomarkers in participants with good control of disease at baseline (A4 below upper limit of normal) (n=3). Overall, tildacerfont was well tolerated with no serious adverse events.

Conclusions: This is the first study of 12 weeks’ duration for a novel, non-steroidal mechanism-of-action agent for the treatment of 21-OHD. Results of this study show that tildacerfont was generally well-tolerated and effective in achieving meaningful reductions in ACTH and A4 in poorly controlled patients over 12 weeks. In addition, this is the first, non-steroidal therapeutic to show evidence of ACTH and A4 normalization over 12 weeks of therapy. Longer term future studies will evaluate whether treatment with tildacerfont can achieve further clinical benefits and allow reduction of GC doses while controlling relevant disease biomarkers.
Bile Acids Induce IQ Containing GTPase Activating Protein 1 to Alleviate Liver Injury
Authors: Sen Anushna , Erickson Hanna , Anakk Sayeepriyadarshini
Published: May 2021

Bile acids (BA) are cholesterol metabolites synthesized in the liver. In addition to their role as detergents, bile acids can function as signaling molecules via the activation of nuclear and G-protein coupled receptors, FXR and TGR5. BA signaling is associated with inflammation, cell proliferation, as well as apoptosis in the liver. Excessive accumulation of bile acids in the liver (cholestasis) is observed in many diseased states. We found that the scaffold protein IQGAP1 was induced in mouse models of cholestasis and in patients. In vitro BA treatment of liver cell-line, HepG2 also induced IQGAP1 in a dose-dependent manner. To investigate the role of this IQGAP1 induction, we fed wild-type male mice 0.1% 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet for 3days, 1week, and 2weeks. Serum bile acid and hepatic IQGAP1 induction increased concomitantly with time. We next treated Iqgap1-/- male mice similarly with DDC and found a 3-fold increase in serum bile acid levels compared to treated wild-type mice. Liver injury markers ALT, AST, and bilirubin were also increased. Although Iqgap1-/- mice had similar gene expression of key BA regulators, Fxr and Shp as wild-type mice, the alteration in expression of BA transporters and reduced cell-cell junction protein levels may contribute to the elevated BAs levels. We quantified gene expression of pro-inflammatory and proliferative mediators using qRT-PCR and examined the liver histology with immuno-staining. We found Iqgap1-/- livers expressed increased pro-inflammatory mediators and proliferative genes like Il-6 and cyclin D1 along with MAPK signaling targets, Fos and Egr-1. To further examine how IQGAP1 functions as a mediator of bile acid signaling, we have generated IQGAP1 domain deleted constructs and are examining their protein-protein interactions with Mass Spec. Based on our results, IQGAP1 induction subsequent to accumulation of BAs is protective against injury in the liver.
Differential Activity and Utilization of Glucocorticoid Receptor Binding Sites Yields Transcriptional Heterogeneity
Authors: Hoffman Jackson Andrew , Trotter Kevin W , Day Christopher R , Ward James M , Rodriguez Joseph , Archer Trevor K
Published: May 2021

Synthetic glucocorticoids such as Dexamethasone (Dex) are widely prescribed drugs used to treat a variety of human diseases including auto-immune disorders, asthma, cancer, and COVID-19. The transcriptional response to glucocorticoids is elicited by the Glucocorticoid Receptor (GR), which enters the nucleus upon Dex treatment and interacts with thousands of enhancer elements throughout the genome. We recently demonstrated that the Dex response in human breast cancer cells is highly heterogeneous and that individual cells have unique transcriptional responses to Dex. To examine whether this heterogeneity arises from differential utilization of distinct GR-bound enhancers, we focused on the Dex response at the DNA Damage Inducible Transcript 4 (DDIT4) gene. Using a variety of genomic techniques, we identified four GR binding sites (GBSs) 18-30kb upstream of the DDIT4 TSS with differential patterns of chromatin accessibility, histone acetylation, SWI/SNF recruitment, and enhancer RNA (eRNA) transcription. To determine whether these GBSs had unique requirements for DDIT4 transcription, we used CRISPR-CAS9 to generate homozygous deletions of each site. Using ChIP-seq, 4C-seq, single molecule fluorescent in situ hybridization (smFISH), and RT-PCR, we demonstrated GR binding to these GBSs was independent and each GBS deletion had unique effects on DDIT4 and eRNA transcription, local histone acetylation, and chromatin looping. Deletion of any of the first three GBSs resulted in delayed and/or decreased induction of DDIT4 transcription whereas deletion of the fourth GBS resulted in significant upregulation of both DDIT4 and eRNA transcription. Thus, three of the GBSs acted as enhancers of DDIT4 expression while the fourth functioned as a suppressor. Strikingly, smFISH also revealed that these enhancers contributed to cellular heterogeneity, as deleting the GBSs altered the frequency and amplitude of DDIT4 transcription across cell populations. Taken together, these results demonstrate that individual GBSs uniquely contribute to cell-to-cell heterogeneity within the transcriptional response of DDIT4 to Dex. Furthermore, they underscore the possibility that targeted modification of individual GBSs could be utilized to tailor custom, patient-specific strategies for the treatment of human diseases.