Author Correction: γδ T cells producing interleukin-17A regulate adipose regulatory T cell homeostasis and thermogenesis.

In the version of this article initially published, three authors (Hui-Fern Kuoy, Adam P. Uldrich and Dale. I. Godfrey) and their affiliations, acknowledgments and contributions were not included. The correct information is as follows:Ayano C. Kohlgruber1,2, Shani T. Gal-Oz3, Nelson M. LaMarche1,2, Moto Shimazaki1, Danielle Duquette4, Hui-Fern Koay5,6, Hung N. Nguyen1, Amir I. Mina4, Tyler Paras1, Ali Tavakkoli7, Ulrich von Andrian2,8, Adam P. Uldrich5,6, Dale I. Godfrey5,6, Alexander S. Banks4, Tal Shay3, Michael B. Brenner1,10* and Lydia Lynch1,4,9,10*1Division of Rheumatology, Immunology and Allergy, Brigham and Women's Hospital, Boston, MA, USA. 2Division of Medical Sciences, Harvard Medical School, Boston, MA, USA. 3Department of Life Sciences, Ben-Gurion University of the Negev, Beersheba, Israel. 4Division of Endocrinology, Department of Medicine, Brigham and Women's Hospital, Boston, MA, USA. 5Department of Microbiology and Immunology, Peter Doherty Institute for Infection and Immunity, University of Melbourne, Parkville, Australia. 6ARC Centre of Excellence in Advanced Molecular Imaging, University of Melbourne, Parkville, Australia. 7Department of General and Gastrointestinal Surgery, Brigham and Women's Hospital, Boston, MA, USA. 8Department of Microbiology and Immunology, Harvard Medical School, Boston, MA, USA. 9School of Biochemistry and Immunology, Trinity College, Dublin, Ireland. 10These authors jointly supervised this work: Michael B. Brenner, Lydia Lynch. *e-mail: mbrenner@research.bwh.harvard.edu; llynch@bwh.harvard.eduAcknowledgementsWe thank A.T. Chicoine, flow cytometry core manager at the Human Immunology Center at BWH, for flow cytometry sorting. We thank D. Sant'Angelo (Rutgers Cancer Institute) for providing Zbtb16-/- mice and R. O'Brien (National Jewish Health) for providing Vg4/6-/- mice. Supported by NIH grant R01 AI11304603 (to M.B.B.), ERC Starting Grant 679173 (to L.L.), the National Health and Medical Research Council of Australia (1013667), an Australian Research Council Future Fellowship (FT140100278 for A.P.U.) and a National Health and Medical Research Council of Australia Senior Principal Research Fellowship (1117766 for D.I.G.).Author contributionsA.C.K., L.L., and M.B.B. conceived and designed the experiments, and wrote the manuscript. A.C.K., N.M.L., L.L., H.N.N., M.S., T.P., and D.D. performed the experiments. S.T.G.-O. and T.S. performed the RNA-seq analysis. A.S.B. and A.I.M. provided advice and performed the CLAMS experiments. A.T. provided human bariatric patient samples. Parabiosis experiments were performed in the laboratory of U.v.A. H.-F.K., A.P.U. and D.I.G provided critical insight into the TCR chain usage of PLZF+ γδ T cells. M.B.B., N.M.L., and L.L. critically reviewed the manuscript.The errors have been corrected in the HTML and PDF version of the article.Correction to: Nature Immunology doi:10.1038/s41590-018-0094-2 (2018), published online 18 April 2018.

γδ T cells producing interleukin-17A regulate adipose regulatory T cell homeostasis and thermogenesis.

γδ T cells are situated at barrier sites and guard the body from infection and damage. However, little is known about their roles outside of host defense in nonbarrier tissues. Here, we characterize a highly enriched tissue-resident population of γδ T cells in adipose tissue that regulate age-dependent regulatory T cell (Treg) expansion and control core body temperature in response to environmental fluctuations. Mechanistically, innate PLZF+ γδ T cells produced tumor necrosis factor and interleukin (IL) 17 A and determined PDGFRα+ and Pdpn+ stromal-cell production of IL-33 in adipose tissue. Mice lacking γδ T cells or IL-17A exhibited decreases in both ST2+ Treg cells and IL-33 abundance in visceral adipose tissue. Remarkably, these mice also lacked the ability to regulate core body temperature at thermoneutrality and after cold challenge. Together, these findings uncover important physiological roles for resident γδ T cells in adipose tissue immune homeostasis and body-temperature control.

Ablation of PM20D1 reveals N-acyl amino acid control of metabolism and nociception.

N-acyl amino acids (NAAs) are a structurally diverse class of bioactive signaling lipids whose endogenous functions have largely remained uncharacterized. To clarify the physiologic roles of NAAs, we generated mice deficient in the circulating enzyme peptidase M20 domain-containing 1 (PM20D1). Global PM20D1-KO mice have dramatically reduced NAA hydrolase/synthase activities in tissues and blood with concomitant bidirectional dysregulation of endogenous NAAs. Compared with control animals, PM20D1-KO mice exhibit a variety of metabolic and pain phenotypes, including insulin resistance, altered body temperature in cold, and antinociceptive behaviors. Guided by these phenotypes, we identify N-oleoyl-glutamine (C18:1-Gln) as a key PM20D1-regulated NAA. In addition to its mitochondrial uncoupling bioactivity, C18:1-Gln also antagonizes certain members of the transient receptor potential (TRP) calcium channels including TRPV1. Direct administration of C18:1-Gln to mice is sufficient to recapitulate a subset of phenotypes observed in PM20D1-KO animals. These data demonstrate that PM20D1 is a dominant enzymatic regulator of NAA levels in vivo and elucidate physiologic functions for NAA signaling in metabolism and nociception.

Cricotracheal separation: do not get fooled by what you see.

INTRODUCTION: Laryngeal involvement in neck trauma is quite considerable. The presence of a cricotracheal separation type of injury can be easily missed and overlooked, especially if the neck does not show any external signs. Blunt trauma to the neck affects many anatomical structures inside the intact-looking neck that threatens the victim's life. At exploration, the surgeon must be aware of the full impact of the injury on different neck structures. AIM OF WORK: Raise the attention on the proper management of laryngeal trauma victims. PATIENTS AND METHOD: This is a retrospective study carried out on 23 patients who suffered from cricotracheal separation as a result of laryngeal trauma. RESULTS: Cricotracheal separation is a frequent finding in an innocent-looking neck. The mechanism of the trauma itself is an excellent clue to suspect its presence. CONCLUSION: This type of laryngeal injury must be kept in mind and must be suspected. Some recommendations and guidelines are presented on the proper handling of such patients.

Sleeve gastrectomy enhances glucose utilization and remodels adipose tissue independent of weight loss.

Sleeve gastrectomy (SG) induces weight loss-independent improvements in glucose homeostasis by unknown mechanisms. We sought to identify the metabolic adaptations responsible for these improvements. Nonobese C57BL/6J mice on standard chow underwent SG or sham surgery. Functional testing and indirect calorimetry were used to capture metabolic phenotypes. Tissue-specific glucose uptake was assessed by 18-fluorodeoxyglucose (18-FDG) PET/computed tomography, and RNA sequencing was used for gene-expression analysis. In this model, SG induced durable improvements in glucose tolerance in the absence of changes in weight, body composition, or food intake. Indirect calorimetry revealed that SG increased the average respiratory exchange ratio toward 1.0, indicating a weight-independent, systemic shift to carbohydrate utilization. Following SG, orally administered 18-FDG preferentially localized to white adipose depots, showing tissue-specific increases in glucose utilization induced by surgery. Transcriptional analysis with RNA sequencing demonstrated that increased glucose uptake in the visceral adipose tissue was associated with upregulation in transcriptional pathways involved in energy metabolism, adipocyte maturation, and adaptive and innate immune cell chemotaxis and differentiation. SG induces a rapid, weight loss-independent shift toward glucose utilization and transcriptional remodeling of metabolic and immune pathways in visceral adipose tissue. Continued study of this early post-SG physiology may lead to a better understanding of the anti-diabetic mechanisms of bariatric surgery.

Time-Series Aware Metrics for the Evaluation of Intraoperative Electroencephalography-Based Ischemia Detection.

Continuous intraoperative monitoring with electroencephalo2 graphy (EEG) is commonly used to detect cerebral ischemia in high-risk surgical procedures such as carotid endarterectomy. Machine learning (ML) models that detect ischemia in real time can form the basis of automated intraoperative EEG monitoring. In this study, we describe and compare two time-series aware precision and recall metrics to the classical precision and recall metrics for evaluating the performance of ML models that detect ischemia. We trained six ML models to detect ischemia in intraoperative EEG and evaluated them with the area under the precision-recall curve (AUPRC) using time-series aware and classical approaches to compute precision and recall. The Support Vector Classification (SVC) model performed the best on the time-series aware metrics, while the Light Gradient Boosting Machine (LGBM) model performed the best on the classical metrics. Visual inspection of the probability outputs of the models alongside the actual ischemic periods revealed that the time-series aware AUPRC selected a model more likely to predict ischemia onset in a timely fashion than the model selected by classical AUPRC.

Detecting Cerebral Ischemia From Electroencephalography During Carotid Endarterectomy Using Machine Learning.

Monitoring cerebral neuronal activity via electroencephalography (EEG) during surgery can detect ischemia, a precursor to stroke. However, current neurophysiologist-based monitoring is prone to error. In this study, we evaluated machine learning (ML) for efficient and accurate ischemia detection. We trained supervised ML models on a dataset of 802 patients with intraoperative ischemia labels and evaluated them on an independent validation dataset of 30 patients with refined labels from five neurophysiologists. Our results show moderate-to-substantial agreement between neurophysiologists, with Cohen's kappa values between 0.59 and 0.74. Neurophysiologist performance ranged from 58-93% for sensitivity and 83-96% for specificity, while ML models demonstrated comparable ranges of 63-89% and 85-96%. Random Forest (RF), LightGBM (LGBM), and XGBoost RF achieved area under the receiver operating characteristic curve (AUROC) values of 0.92-0.93 and area under the precision-recall curve (AUPRC) values of 0.79-0.83. ML has the potential to improve intraoperative monitoring, enhancing patient safety and reducing costs.

Diagnostic Accuracy of Somatosensory Evoked Potentials and Electroencephalography During Endovascular Treatment of Unruptured Cerebral Aneurysms.

OBJECTIVE: Endovascular treatment (EVT) of unruptured cerebral aneurysms (UCA) offers a safer alternative to clipping. However, it is still associated with an increased risk for Postprocedural Neurological deficit (PPND). Prompt recognition using intraoperative neurophysiologic monitoring (IONM) and intervention can reduce the incidence and impact of new postoperative neurological complications. We aim to evaluate the diagnostic accuracy of IONM in predicting PPND after EVT of UCA. METHODS: We included 414 patients who underwent EVT for UCA from 2014 to 2019. The sensitivities, specificities, and diagnostic odds ratio of somatosensory evoked potentials and electroencephalography monitoring methods were calculated. We also determined their diagnostic accuracy using receiver operating characteristic plots. RESULTS: The highest sensitivity of 67.7% (95% confidence interval {CI}, 34.9%-90.1%) was obtained when either modality had a change. Simultaneous changes in both modalities have the highest specificity of 97.8% (95% CI, 95.8%-99.0%). The area under the receiver operating characteristic curve was 0.795 (95% CI, 0.655-0.935) for changes in either modality. CONCLUSIONS: IONM with somatosensory evoked potentials alone or in combination with electroencephalography has high diagnostic accuracy in detecting periprocedural complications and resultant PPND during EVT of UCA.

Role of Intraoperative Neurophysiological Monitoring in Preventing Stroke After Cardiac Surgery.

BACKGROUND: Perioperative stroke after cardiac surgical procedures carries significant morbidity. Dual intraoperative neurophysiological monitoring with electroencephalography (EEG) and somatosensory-evoked potentials detects cerebral hypoperfusion and predicts postoperative stroke in noncardiac procedures. We further evaluated preoperative risk factors and intraoperative neuromonitoring ability to predict postoperative stroke after cardiac operations. METHODS: All patients who underwent cardiac operations with intraoperative neurophysiological monitoring from 2009 to 2020 at a single academic medical center were retrospectively analyzed. Patients with circulatory arrest were excluded. Risks factors analyzed were sex, age, tobacco use, hypertension, diabetes mellitus, dyslipidemia, atrial fibrillation, prior cerebrovascular accident, cerebrovascular disease, antiplatelet/anticoagulant use, abnormal somatosensory-evoked potentials and EEG baselines, and significant somatosensory-evoked potentials and EEG change as well as their permanence. Patients were divided into 2 groups by 30-day postoperative stroke occurrence. Univariate and multivariate logistical regressions were used for postoperative stroke significant predictors, and Kaplan-Meier curves estimated survival. RESULTS: The study included 620 patients (67.6% men), mean age 65.1 ± 14.1 years, with stroke in 5.32%. In univariate analysis, diabetes (odds ratio [OR], 2.62) and permanence of EEG change (OR, 5.35) were each associated with increased postoperative stroke odds. In multivariate analysis, diabetes (OR, 2.64) and permanent EEG change (OR, 4.22) were independently significantly associated with postoperative stroke. Overall survival was significantly better for patients with no intraoperative neurophysiological monitoring changes (P < .005). CONCLUSIONS: Permanent EEG change and diabetes were significant postoperative stroke predictors in cardiac operations. Furthermore, overall survival out to 10 years postoperatively was significantly higher in the group without intraoperative neurophysiological monitoring changes, emphasizing its important predictive role.

Cardiovascular-related mortality after intraoperative neurophysiologic monitoring changes during carotid endarterectomy.

OBJECTIVE: We examined significant intraoperative neurophysiologic monitoring (IONM) changes and perioperative stroke as independent risk factors of long-term cardiovascular-related mortality in patients who have undergone carotid endarterectomy (CEA). METHODS: Records of patients who underwent CEA with IONM at the University of Pittsburgh Medical Center between January 1, 2009 and December 31, 2019 were analyzed retrospectively. Cardiovascular-related mortality was compared between the significant IONM change group and no IONM change group and between the perioperative stroke group and no perioperative stroke group. RESULTS: Our final cohort consisted of 2,090 patients. Patients with significant IONM changes showed nearly twice the rate of cardiovascular-related mortality up to 10 years post-CEA (hazard ratio (HR) = 1.98; 95% confidence interval (CI) [1.20 - 3.26]). Patients with perioperative stroke were four times more likely than patients without perioperative stroke to experience cardiovascular-related mortality (HR = 4.09; 95% CI [2.13 - 7.86]). CONCLUSIONS: Among CEA patients who underwent CEA and who experienced significant IONM changes or perioperative stroke, we observed long-term increased and sustained risk of cardiovascular-related mortality. SIGNIFICANCE: Significant IONM changes are valuable in predicting the risk of long-term outcomes following CEA.

Predicting transient ischemic attack after carotid endarterectomy: The role of intraoperative neurophysiological monitoring.

OBJECTIVE: Transient ischemic attacks (TIA) after carotid endarterectomy (CEA) are not well-studied. We aimed to investigate the characteristics and the predictive role of intraoperative neurophysiological monitoring (IONM) in TIA post-CEA. METHODS: Patients who underwent CEA utilizing IONM from 2009-2020 were included. Analyses included TIA incidence, sensitivity, specificity, and predictive values of IONM, risk factor regression analyses, and mortality Kaplan Meier plots. RESULTS: Out of 2232 patients, 46 experienced TIA, 14 of which were within 24 hours of CEA (p < 0.01). Nine of these patients displayed significant IONM changes during CEA. The odds of TIA increased with somatosensory evoked potential (SSEP) changes (Odds Ratio (OR): 2.48 95% Confidence Interval (CI): 1.14-5.4), electroencephalogram (EEG) changes (OR: 2.65 95% CI: 1.22-5.77), and combined SSEP/EEG changes (OR: 2.98 95% CI: 1.17-7.55). Patients with TIA were less likely to be alive after an average of 4.3 years (OR: 0.5 95% CI: 0.26-0.96). CONCLUSIONS: The odds a patient will have TIA post-CEA are greater in patients with IONM changes. This risk is inversely related to the time post-CEA. SIGNIFICANCE: Changes in IONM during CEA predict postoperative TIA. Post-CEA TIA may increase long-term mortality, thus further research is needed to better elucidate clinical implications of postoperative TIA.

Optimal "Low" Pedicle Screw Stimulation Threshold to Predict New Postoperative Lower-Extremity Neurologic Deficits During Lumbar Spinal Fusions.

OBJECTIVE: Previous studies have shown that pedicle screw stimulation thresholds ≤6-8 mA yield a high diagnostic accuracy of detecting misplaced screws. Our objective was to determine the optimal "low" stimulation threshold to predict new postoperative neurologic deficits and identify additional risk factors associated with deficits. METHODS: We included patients with complete pedicle screw stimulation testing who underwent posterior lumbar spinal fusion surgeries from 2010-2012. We calculated the diagnostic accuracy of pedicle screw responses of ≤4 mA, ≤6 mA, ≤8 mA, ≤10 mA, ≤12 mA, and ≤20 mA to predict new postoperative lower-extremity (LE) neurologic deficits. We used multivariate modeling to determine the best logistic regression model to predict LE deficits and identify additional risk factors. Statistics software packages used were Python3.8.5, NumPy 1.19.1, Pandas 1.1.1, and SPSS26. RESULTS: We studied 1179 patients who underwent 8584 pedicle screw stimulations with somatosensory evoked potential and free-run electromyographic monitoring for posterior lumbar spinal fusion. Twenty-five (2.1%) patients had new LE neurologic deficits. A stimulation threshold of ≤8 mA had a sensitivity/specificity of 32%/90% and a diagnostic odds ratio/area under the curve of 4.34 [95% confidence interval: 1.83, 10.27]/0.61 [0.49, 0.74] in predicting postoperative deficit. Multivariate analysis showed that patients who had pedicle screws with stimulation thresholds ≤8 mA are 3.15 [1.26, 7.83]× more likely to have postoperative LE deficits while patients who have undergone a revision lumbar spinal fusion surgery are 3.64 [1.38, 9.61]× more likely. CONCLUSIONS: Our results show that low thresholds are indicative of not only screw proximity to the nerve but also an increased likelihood of postoperative neurologic deficit. Thresholds ≤8 mA prove to be the optimal "low" threshold to help guide a correctly positioned pedicle screw placement and detect postoperative deficits.

Progenitor Cells from the Adult Human Inner Ear.

Loss of inner ear hair cells leads to incurable balance and hearing disorders because these sensory cells do not effectively regenerate in humans. A potential starting point for therapy would be the stimulation of quiescent progenitor cells within the damaged inner ear. Inner ear progenitor/stem cells, which have been described in rodent inner ears, would be principal candidates for such an approach. Despite the identification of progenitor cell populations in the human fetal cochlea and in the adult human spiral ganglion, no proliferative cell populations with the capacity to generate hair cells have been reported in vestibular and cochlear tissues of adult humans. The present study aimed at filling this gap by isolating colony-forming progenitor cells from surgery- and autopsy-derived adult human temporal bones in order to generate inner ear cell types in vitro. Sphere-forming and mitogen-responding progenitor cells were isolated from vestibular and cochlear tissues. Clonal spheres grown from adult human utricle and cochlear duct were propagated for a limited number of generations. When differentiated in absence of mitogens, the utricle-derived spheres robustly gave rise to hair cell-like cells, as well as to cells expressing supporting cell-, neuron-, and glial markers, indicating that the adult human utricle harbors multipotent progenitor cells. Spheres derived from the adult human cochlear duct did not give rise to hair cell-like or neuronal cell types, which is an indication that human cochlear cells have limited proliferative potential but lack the ability to differentiate into major inner ear cell types. Anat Rec, 303:461-470, 2020. © 2019 The Authors. The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists.

Obesity-Linked PPARγ S273 Phosphorylation Promotes Insulin Resistance through Growth Differentiation Factor 3.

The thiazolidinediones (TZDs) are ligands of PPARγ that improve insulin sensitivity, but their use is limited by significant side effects. Recently, we demonstrated a mechanism wherein TZDs improve insulin sensitivity distinct from receptor agonism and adipogenesis: reversal of obesity-linked phosphorylation of PPARγ at serine 273. However, the role of this modification hasn't been tested genetically. Here we demonstrate that mice encoding an allele of PPARγ that cannot be phosphorylated at S273 are protected from insulin resistance, without exhibiting differences in body weight or TZD-associated side effects. Indeed, hyperinsulinemic-euglycemic clamp experiments confirm insulin sensitivity. RNA-seq in these mice reveals reduced expression of Gdf3, a BMP family member. Ectopic expression of Gdf3 is sufficient to induce insulin resistance in lean, healthy mice. We find Gdf3 inhibits BMP signaling and insulin signaling in vitro. Together, these results highlight the diabetogenic role of PPARγ S273 phosphorylation and focus attention on a putative target, Gdf3.

Timing of Lymphedema After Treatment for Breast Cancer: When Are Patients Most At Risk?

PURPOSE: The purpose of the study was to determine when the risk of lymphedema is highest after treatment of breast cancer and which factors influence the time course of lymphedema development. METHODS AND MATERIALS: Between 2005 and 2017, 2171 women (with 2266 at-risk arms) who received surgery for unilateral or bilateral breast cancer at our institution were enrolled. Perometry was used to objectively assess limb volume preoperatively, and lymphedema was defined as a ≥10% relative arm-volume increase arising >3 months postoperatively. Multivariable regression was used to uncover risk factors associated with lymphedema, the Cox proportional hazards model was used to calculate lymphedema incidence, and the semiannual hazard rate of lymphedema was calculated. RESULTS: With a median follow-up of 4 years, the overall estimated 5-year cumulative incidence of lymphedema was 13.7%. Significant factors associated with lymphedema on multivariable analysis were high preoperative body mass index, axillary lymph node dissection (ALND), and regional lymph node radiation (RLNR). Patients receiving ALND with RLNR experienced the highest 5-year rate of lymphedema (31.2%), followed by those receiving ALND without RLNR (24.6%) and sentinel lymph node biopsy with RLNR (12.2%). Overall, the risk of lymphedema peaked between 12 and 30 months postoperatively; however, the time course varied as a function of therapy received. Early-onset lymphedema (<12 months postoperatively) was associated with ALND (HR [hazard ratio], 4.75; P < .0001) but not with RLNR (HR, 1.21; P = .55). In contrast, late-onset lymphedema (>12 months postoperatively) was associated with RLNR (HR, 3.86; P = .0001) and, to a lesser extent, ALND (HR, 1.86; P = .029). The lymphedema risk peaked between 6 and 12 months in the ALND-without-RLNR group, between 18 and 24 months in the ALND-with-RLNR group, and between 36 and 48 months in the group receiving sentinel lymph node biopsy with RLNR. CONCLUSIONS: The time course for lymphedema development depends on the breast cancer treatment received. ALND is associated with early-onset lymphedema, and RLNR is associated with late-onset lymphedema. These results can influence clinical practice to guide lymphedema surveillance strategies and patient education.

CalR: A Web-Based Analysis Tool for Indirect Calorimetry Experiments.

We report a web-based tool for analysis of experiments using indirect calorimetry to measure physiological energy balance. CalR simplifies the process to import raw data files, generate plots, and determine the most appropriate statistical tests for interpretation. Analysis using the generalized linear model (which includes ANOVA and ANCOVA) allows for flexibility in interpreting diverse experimental designs, including those of obesity and thermogenesis. Users also may produce standardized output files for an experiment that can be shared and subsequently re-evaluated using CalR. This framework will provide the transparency necessary to enhance consistency, rigor, and reproducibility. The CalR analysis software will greatly increase the speed and efficiency with which metabolic experiments can be organized, analyzed per accepted norms, and reproduced and will likely become a standard tool for the field. CalR is accessible at https://CalRapp.org/.

A selective gut bacterial bile salt hydrolase alters host metabolism.

The human gut microbiota impacts host metabolism and has been implicated in the pathophysiology of obesity and metabolic syndromes. However, defining the roles of specific microbial activities and metabolites on host phenotypes has proven challenging due to the complexity of the microbiome-host ecosystem. Here, we identify strains from the abundant gut bacterial phylum Bacteroidetes that display selective bile salt hydrolase (BSH) activity. Using isogenic strains of wild-type and BSH-deleted Bacteroides thetaiotaomicron, we selectively modulated the levels of the bile acid tauro-ß-muricholic acid in monocolonized gnotobiotic mice. B. thetaiotaomicron BSH mutant-colonized mice displayed altered metabolism, including reduced weight gain and respiratory exchange ratios, as well as transcriptional changes in metabolic, circadian rhythm, and immune pathways in the gut and liver. Our results demonstrate that metabolites generated by a single microbial gene and enzymatic activity can profoundly alter host metabolism and gene expression at local and organism-level scales.

Phosphorylation of Beta-3 adrenergic receptor at serine 247 by ERK MAP kinase drives lipolysis in obese adipocytes.

OBJECTIVE: The inappropriate release of free fatty acids from obese adipose tissue stores has detrimental effects on metabolism, but key molecular mechanisms controlling FFA release from adipocytes remain undefined. Although obesity promotes systemic inflammation, we find activation of the inflammation-associated Mitogen Activated Protein kinase ERK occurs specifically in adipose tissues of obese mice, and provide evidence that adipocyte ERK activation may explain exaggerated adipose tissue lipolysis observed in obesity. METHODS AND RESULTS: We provide genetic and pharmacological evidence that inhibition of the MEK/ERK pathway in human adipose tissue, mice, and flies all effectively limit adipocyte lipolysis. In complementary findings, we show that genetic and obesity-mediated activation of ERK enhances lipolysis, whereas adipose tissue specific knock-out of ERK2, the exclusive ERK1/2 protein in adipocytes, dramatically impairs lipolysis in explanted mouse adipose tissue. In addition, acute inhibition of MEK/ERK signaling also decreases lipolysis in adipose tissue and improves insulin sensitivity in obese mice. Mice with decreased rates of adipose tissue lipolysis in vivo caused by either MEK or ATGL pharmacological inhibition were unable to liberate sufficient White Adipose Tissue (WAT) energy stores to fuel thermogenesis from brown fat during a cold temperature challenge. To identify a molecular mechanism controlling these actions, we performed unbiased phosphoproteomic analysis of obese adipose tissue at different time points following acute pharmacological MEK/ERK inhibition. MEK/ERK inhibition decreased levels of adrenergic signaling and caused de-phosphorylation of the ß3-adrenergic receptor (ß3AR) on serine 247. To define the functional implications of this phosphorylation, we showed that CRISPR/Cas9 engineered cells expressing wild type ß3AR exhibited ß3AR phosphorylation by ERK2 and enhanced lipolysis, but this was not seen when serine 247 of ß3AR was mutated to alanine. CONCLUSION: Taken together, these data suggest that ERK activation in adipocytes and subsequent phosphorylation of the ß3AR on S247 are critical regulatory steps in the enhanced adipocyte lipolysis of obesity.

Association Between Precautionary Behaviors and Breast Cancer-Related Lymphedema in Patients Undergoing Bilateral Surgery.

Purpose This study examined the lifestyle and clinical risk factors for lymphedema in a cohort of patients who underwent bilateral breast cancer surgery. Patients and Methods Between 2013 and 2016, 327 patients who underwent bilateral breast cancer surgery were prospectively screened for arm lymphedema as quantified by the weight-adjusted volume change (WAC) formula. Arm perometry and subjective data were collected preoperatively and at regular intervals postoperatively. At the time of each measurement, patients completed a risk assessment survey that reported the number of blood draws, injections, blood pressure readings, trauma to the at-risk arm, and number of flights since the previous measurement. Generalized estimating equations were applied to ascertain the association among arm volume changes, clinical factors, and risk exposures. Results The cohort comprised 327 patients and 654 at-risk arms, with a median postoperative follow-up that ranged from 6.1 to 68.2 months. Of the 654 arms, 83 developed lymphedema, defined as a WAC ≥ 10% relative to baseline. On multivariable analysis, none of the lifestyle risk factors examined through the risk assessment survey were significantly associated with increased WAC. Multivariable analysis demonstrated that having a body mass index ≥ 25 kg/m2 at the time of breast cancer diagnosis ( P = .0404), having undergone axillary lymph node dissection ( P = .0464), and receipt of adjuvant chemotherapy ( P = .0161) were significantly associated with increased arm volume. Conclusion Blood pressure readings, blood draws, injections, and number or duration of flights were not significantly associated with increases in arm volume in this cohort. These findings may help to guide patient education about lymphedema risk reduction strategies for those who undergo bilateral breast cancer surgery.

Genetic Depletion of Adipocyte Creatine Metabolism Inhibits Diet-Induced Thermogenesis and Drives Obesity.

Diet-induced thermogenesis is an important homeostatic mechanism that limits weight gain in response to caloric excess and contributes to the relative stability of body weight in most individuals. We previously demonstrated that creatine enhances energy expenditure through stimulation of mitochondrial ATP turnover, but the physiological role and importance of creatine energetics in adipose tissue have not been explored. Here, we have inactivated the first and rate-limiting enzyme of creatine biosynthesis, glycine amidinotransferase (GATM), selectively in fat (Adipo-Gatm KO). Adipo-Gatm KO mice are prone to diet-induced obesity due to the suppression of elevated energy expenditure that occurs in response to high-calorie feeding. This is paralleled by a blunted capacity for ß3-adrenergic activation of metabolic rate, which is rescued by dietary creatine supplementation. These results provide strong in vivo genetic support for a role of GATM and creatine metabolism in energy expenditure, diet-induced thermogenesis, and defense against diet-induced obesity.