Antibody blockade of activin type II receptors preserves skeletal muscle mass and enhances fat loss during GLP-1 receptor agonism.

OBJECTIVE: Glucagon-like peptide 1 (GLP-1) receptor agonists reduce food intake, producing remarkable weight loss in overweight and obese individuals. While much of this weight loss is fat mass, there is also a loss of lean mass, similar to other approaches that induce calorie deficit. Targeting signaling pathways that regulate skeletal muscle hypertrophy is a promising avenue to preserve lean mass and modulate body composition. Myostatin and Activin A are TGFß-like ligands that signal via the activin type II receptors (ActRII) to antagonize muscle growth. Pre-clinical and clinical studies demonstrate that ActRII blockade induces skeletal muscle hypertrophy and reduces fat mass. In this manuscript, we test the hypothesis that combined ActRII blockade and GLP-1 receptor agonism will preserve muscle mass, leading to improvements in skeletomuscular and metabolic function and enhanced fat loss. METHODS: In this study, we explore the therapeutic potential of bimagrumab, a monoclonal antibody against ActRII, to modify body composition alone and during weight loss induced by GLP-1 receptor agonist semaglutide in diet-induced obese mice. Mechanistically, we define the specific role of the anabolic kinase Akt in mediating the hypertrophic muscle effects of ActRII inhibition in vivo. RESULTS: Treatment of obese mice with bimagrumab induced a ∼10 % increase in lean mass while simultaneously decreasing fat mass. Daily treatment of obese mice with semaglutide potently decreased body weight; this included a significant decrease in both muscle and fat mass. Combination treatment with bimagrumab and semaglutide led to superior fat mass loss while simultaneously preserving lean mass despite reduced food intake. Treatment with both drugs was associated with improved metabolic outcomes, and increased lean mass was associated with improved exercise performance. Deletion of both Akt isoforms in skeletal muscle modestly reduced, but did not prevent, muscle hypertrophy driven by ActRII inhibition. CONCLUSIONS: Collectively, these data demonstrate that blockade of ActRII signaling improves body composition and metabolic parameters during calorie deficit driven by GLP-1 receptor agonism and demonstrate the existence of Akt-independent pathways supporting muscle hypertrophy in the absence of ActRII signaling.

mTORC1 controls murine postprandial hepatic glycogen synthesis via Ppp1r3b.

In response to a meal, insulin drives hepatic glycogen synthesis to help regulate systemic glucose homeostasis. The mechanistic target of rapamycin complex 1 (mTORC1) is a well-established insulin target and contributes to the postprandial control of liver lipid metabolism, autophagy, and protein synthesis. However, its role in hepatic glucose metabolism is less understood. Here, we used metabolomics, isotope tracing, and mouse genetics to define a role for liver mTORC1 signaling in the control of postprandial glycolytic intermediates and glycogen deposition. We show that mTORC1 is required for glycogen synthase activity and glycogenesis. Mechanistically, hepatic mTORC1 activity promotes the feeding-dependent induction of Ppp1r3b, a gene encoding a phosphatase important for glycogen synthase activity whose polymorphisms are linked to human diabetes. Reexpression of Ppp1r3b in livers lacking mTORC1 signaling enhances glycogen synthase activity and restores postprandial glycogen content. mTORC1-dependent transcriptional control of Ppp1r3b is facilitated by FOXO1, a well characterized transcriptional regulator involved in the hepatic response to nutrient intake. Collectively, we identify a role for mTORC1 signaling in the transcriptional regulation of Ppp1r3b and the subsequent induction of postprandial hepatic glycogen synthesis.

LKB1 acts as a critical brake for the glucagon-mediated fasting response.

As important as the fasting response is for survival, an inability to shut it down once nutrients become available can lead to exacerbated disease and severe wasting. The liver is central to transitions between feeding and fasting states, with glucagon being a key initiator of the hepatic fasting response. However, the precise mechanisms controlling fasting are not well defined. One potential mediator of these transitions is liver kinase B1 (LKB1), given its role in nutrient sensing. Here, we show LKB1 knockout mice have a severe wasting and prolonged fasting phenotype despite increased food intake. By applying RNA sequencing and intravital microscopy, we show that loss of LKB1 leads to a dramatic reprogramming of the hepatic lobule through robust up-regulation of periportal genes and functions. This is likely mediated through the opposing effect that LKB1 has on glucagon pathways and gene expression. Conclusion: Our findings show that LKB1 acts as a brake to the glucagon-mediated fasting response, resulting in "periportalization" of the hepatic lobule and whole-body metabolic inefficiency. These findings reveal a mechanism by which hepatic metabolic compartmentalization is regulated by nutrient-sensing.

A Novel Mouse Model to Analyze Non-Genomic ERα Physiological Actions.

Nongenomic effects of estrogen receptor α (ERα) signaling have been described for decades. Several distinct animal models have been generated previously to analyze the nongenomic ERα signaling (eg, membrane-only ER, and ERαC451A). However, the mechanisms and physiological processes resulting solely from nongenomic signaling are still poorly understood. Herein, we describe a novel mouse model for analyzing nongenomic ERα actions named H2NES knock-in (KI). H2NES ERα possesses a nuclear export signal (NES) in the hinge region of ERα protein resulting in exclusive cytoplasmic localization that involves only the nongenomic action but not nuclear genomic actions. We generated H2NESKI mice by homologous recombination method and have characterized the phenotypes. H2NESKI homozygote mice possess almost identical phenotypes with ERα null mice except for the vascular activity on reendothelialization. We conclude that ERα-mediated nongenomic estrogenic signaling alone is insufficient to control most estrogen-mediated endocrine physiological responses; however, there could be some physiological responses that are nongenomic action dominant. H2NESKI mice have been deposited in the repository at Jax (stock no. 032176). These mice should be useful for analyzing nongenomic estrogenic responses and could expand analysis along with other ERα mutant mice lacking membrane-bound ERα. We expect the H2NESKI mouse model to aid our understanding of ERα-mediated nongenomic physiological responses and serve as an in vivo model for evaluating the nongenomic action of various estrogenic agents.

Dpp4+ interstitial progenitor cells contribute to basal and high fat diet-induced adipogenesis.

OBJECTIVE: The capacity to generate new adipocytes from precursor cells is critical for maintaining metabolic health. Adipocyte precursor cells (APCs) constitute a heterogenous collection of cell types; however, the contribution of these various cell types to adipose tissue expansion in vivo remains unknown. The aim of the current study is to investigate the contribution of Dpp4+ progenitors to de novo adipogenesis. METHODS: Single cell analysis has identified several transcriptionally distinct subpopulations of APCs, including Dpp4+ progenitor cells concentrated in the connective tissue surrounding many organs, including white adipose tissue (WAT). Here, we generated a Dpp4CreER mouse model for in vivo lineage tracing of these cells and their downstream progeny in the setting of basal or high fat diet (HFD)-stimulated adipogenesis. RESULTS: Dpp4CreER mice enabled specific temporal labeling of Dpp4+ progenitor cells within their native connective tissue niche. Following a dietary chase period consisting of chow or HFD feeding for 18 weeks, Dpp4+ progenitors differentiated into mature adipocytes within the gonadal and subcutaneous WAT. HFD stimulated adipogenic contribution from Dpp4+ cells in the gonadal but not the subcutaneous depot. Flow cytometry analysis revealed that Dpp4+ progenitors give rise to DPP4(-)/ICAM1+ preadipocytes in vivo. HFD feeding did not perturb the flux of Dpp4+ cell conversion into ICAM1+ preadipocytes in gonadal WAT. Conversely, in subcutaneous WAT, HFD feeding/obesity led to an accumulation of ICAM1+ preadipocytes without a corresponding increase in mature adipocyte differentiation. Examination of non-classical murine visceral depots with relevance to humans, including omentum and retroperitoneal WAT, revealed robust contribution of Dpp4+ progenitors to de novo adipogenesis, which was further stimulated by HFD. CONCLUSION: Our data demonstrate that Dpp4+ interstitial progenitor cells contribute to basal adipogenesis in all fat depots and are recruited to support de novo adipogenic expansion of visceral WAT in the setting of HFD-induced obesity.

Intravital Microscopy for the Study of Hepatic Glucose Uptake.

The liver is central in maintaining glucose homeostasis. Indeed, impaired hepatic glucose uptake has been implicated in the development of hyperglycemia in type II diabetes (T2D) and non-alcoholic fatty liver disease (NAFLD). However, current approaches to evaluate glucose mobilization rely on indirect measurements that do not provide spatial and temporal information. Here, we describe confocal-based intravital microscopy (IVM) of the liver that allows the identification of hepatocyte spatial organization and glucose transport. Specifically, we describe a method to fluorescently label hepatic landmarks to identify different compartments within the liver. In addition, we outline an in vivo fluorescent glucose uptake assay to quantitatively measure glucose mobilization in space and time. These protocols allow direct investigation of hepatic glycemic control and can be further applied to murine models of liver disease. © Published 2021. This article is a U.S. Government work and is in the public domain in the USA. Basic Protocol 1: Mouse surgical procedure and positioning for liver intravital imaging Basic Protocol 2: Fluorescent labeling and intravital imaging of mouse hepatic compartments Basic Protocol 3: Mouse hepatic glucose uptake assay and intravital imaging analysis.

Physical models can provide superior learning opportunities beyond the benefits of active engagements.

The essence of molecular biology education lies in understanding of gene expression, with subtopics including the central dogma processes, such as transcription and translation. While these concepts are core to the discipline, they are also notoriously difficult for students to learn, probably because they cannot be directly observed. While nearly all active learning strategies have been shown to improve learning compared with passive lectures, little has been done to compare different types of active learning. We hypothesized that physical models of central dogma processes would be especially helpful for learning, because they provide a resource that students can see, touch, and manipulate while trying to build their knowledge. For students enrolled in an entirely active-learning-based Cell & Molecular Biology course, we examined whether model-based activities were more effective than non-model based activities. To test their understanding at the beginning and end of the semester, we employed the multiple-select Central Dogma Concept Inventory (CDCI). Each student acted as their own control, as all students engaged in all lessons yet some questions related to model-based activities and some related to clicker questions, group problem-solving, and other non-model-based activities. While all students demonstrated learning gains on both types of question, they showed much higher learning gains on model-based questions. Examining their selected answers in detail showed that while higher performing students were prompted to refine their already-good mental models to be even better, lower performing students were able to construct new knowledge that was much more consistent with an expert's understanding. © 2018 The Authors. Biochemistry and Molecular Biology Education published by Wiley Periodicals, Inc. on behalf of International Union of Biochemistry and Molecular Biology., 46(5):435-444, 2018.

Experimental models for evaluating non-genomic estrogen signaling.

Non-genomic effects of estrogen receptor α (ERα) signaling have been described for decades. However, the mechanisms and physiological processes resulting solely from non-genomic signaling are poorly understood. Challenges in studying these effects arise from the strongly nucleophilic tendencies of estrogen receptor, and many approaches to excluding ERα from the nucleus have been explored over the years. In this review, we discuss past strategies for studying ERα's non-genomic action and current models, specifically H2NES ERα, first described by Burns et al. (2011). In vitro and preliminary in vivo data from H2NES ERα and H2NES mice suggest a promising avenue for pinpointing specific non-genomic ERα action.