Glucagon stimulates hepatic glucose production, in part by promoting the uptake and catabolism of amino acids. Inhibition of the liver glucagon receptor (GCGR) results in elevated plasma amino acids, which triggers the proliferation of pancreatic α-cells, forming a liver-α-cell loop. This study aims to delineate hepatic signaling molecules downstream of GCGR that mediate the liver-α-cell loop. We knocked down liver GCGR, its G-coupled protein GNAS, and two GNAS downstream effectors, PKA and EPAC2 (RAPGEF4). Mice with GCGR, GNAS, and PKA knockdown had similar suppression of hepatic amino acid catabolism genes, hyperaminoacidemia, and α-cell hyperplasia, but those with EPAC2 knockdown did not. We then demonstrated that activating liver PKA was sufficient to reverse hyperaminoacidemia and α-cell hyperplasia caused by GCGR blockade. These results suggest that liver GCGR signals through PKA to control amino acid metabolism and that hepatic PKA plays a critical role in the liver-α-cell loop. ARTICLE HIGHLIGHTS:A liver-α-cell loop exists, where inhibition of the liver glucagon receptor (GCGR) causes hyperaminoacidemia and pancreatic α-cell hyperplasia, but the GCGR downstream factors responsible for these effects are not clear. We silenced GCGR, its G-coupled protein GNAS, and two GNAS downstream effectors, PKA and EPAC2, to assess their effects on the liver-α-cell loop. Inhibition of the GCGR-GNAS-PKA pathway suppresses amino acid catabolism and causes α-cell hyperplasia, whereas PKA activation promotes amino acid catabolism and reduces alpha cell mass even when GCGR is blocked. Our study establishes hepatic PKA as the critical regulator of the liver-α-cell loop.

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Liver steatosis is an increasing health issue with few therapeutic options, partly because of a paucity of exper-imental models. In humanized liver rodent models, abnormal lipid accumulation in transplanted human hepa-tocytes occurs spontaneously. Here, we demonstrate that this abnormality is associated with compromised interleukin-6 (IL-6)-glycoprotein 130 (GP130) signaling in human hepatocytes because of incompatibility between host rodent IL-6 and human IL-6 receptor (IL-6R) on donor hepatocytes. Restoration of hepatic IL-6-GP130 signaling, through ectopic expression of rodent IL-6R, constitutive activation of GP130 in human he-patocytes, or humanization of an Il6 allele in recipient mice, substantially reduced hepatosteatosis. Notably, providing human Kupffer cells via hematopoietic stem cell engraftment in humanized liver mice also corrected the abnormality. Our observations suggest an important role of IL-6-GP130 pathway in regulating lipid accu-mulation in hepatocytes and not only provide a method to improve humanized liver models but also suggest therapeutic potential for manipulating GP130 signaling in human liver steatosis.

Glucagon-like peptide-1 receptor agonists (GLP-1RAs) have become standard-of-care for the pharmacological treatment of obesity by inducing clinically meaningful weight loss through a reduction in appetite. While these agents potently reduce fat mass, the caloric restriction also reduces skeletal muscle mass. Strategies to preserve skeletal muscle in the presence of caloric restriction are needed to improve weight loss quality in obese patents. Trevogrumab (∝-MSTN) and garetosmab (∝-ActA) are fully human monoclonal antibodies that potently inhibit two mediators of skeletal muscle atrophy: myostatin (MSTN) and activin A (ActA). We first tested the effects of ∝-MSTN, ∝-ActA and GLP-1RA (Semaglutide [Sema]) on body composition of diet-induced obese mice either alone or in combination. Sema (-29.8%) and ∝-MSTN/∝-ActA (-23.4%) produced significant reduction in fat mass after 4 weeks of treatment vs baseline. Adding MTSN and ActA blockade to Sema produced an additive effect on fat mass (-54.2%). This combination also increased lean mass by 9.8% vs a 1.9% loss with Sema alone. To test this in primates, we performed a 20-week study with the combined administration of either Sema + ∝-MSTN or Sema + ∝-MSTN and ∝-ActA in obese male cynomolgus monkeys. The combination of ∝-MSTN + Sema was sufficient to cause an additive effect on fat mass -43.9% vs -25.1% for Sema alone. The addition of ∝-ActA to the combination produced a similar reduction in fat (-49.6%). The Sema control group also lost -2.5% lean mass after 20 weeks; despite the significant reduction in fat mass, the addition of ∝-MSTN to Sema slightly increased lean mass over the same period (0.5%); the addition of ∝-ActA increased lean mass even further (6.2%). These data suggest the blockade of MSTN and ActA to GLP-1RA treatment could greatly improve the quality of weight loss with this class of anti-obesity agents. Disclosure J.Mastaitis: Employee; Regeneron Pharmaceuticals Inc., Stock/Shareholder; Regeneron Pharmaceuticals Inc. D.Gomez: Employee; Regeneron Pharmaceuticals Inc., Stock/Shareholder; Regeneron Pharmaceuticals Inc. V.Le rouzic: Employee; Regeneron Pharmaceuticals Inc., Stock/Shareholder; Regeneron Pharmaceuticals Inc. M.Stec: Employee; Regeneron Pharmaceuticals Inc., Stock/Shareholder; Regeneron Pharmaceuticals Inc. N.Khan: Employee; Regeneron Pharmaceuticals Inc., Stock/Shareholder; Regeneron Pharmaceuticals Inc. E.Na: Employee; Regeneron Pharmaceuticals Inc., Stock/Shareholder; Regeneron Pharmaceuticals Inc. T.Mcwilliams: None. S.Min: Employee; Regeneron Pharmaceuticals Inc., Stock/Shareholder; Regeneron Pharmaceuticals Inc. M.Sleeman: Employee; Regeneron Pharmaceuticals Inc. Funding Regeneron Pharmaceuticals Inc.

Sarcopenia, originally defined as age-related low muscle mass and function, has also been used to describe the loss of muscle mass and quality among certain populations, such as sarcopenic obesity (SO) . SO refers to loss of muscle and function in obese subjects; however, given age is not a factor in SO, we classified SO as obesity with low lean muscle mass (OLLMM) . Given the paucity of data in OLLMM, the purpose was to describe the prevalence of OLLMM in the US. Data from NHANES 2017-2018 were used to estimate the prevalence of OLLMM in adults ≥20 yrs of age. OLLMM was defined as DEXA-assessed low appendicular lean mass, adjusted for BMI (men <0.789, women <0.512) and obesity (body fat%: men >25%, women >35%) . Since DEXA was only measured in those aged 20-59 in NHANES 2017-2018, we utilized logistic regression models to predict OLLMM from NHANES 1999-20for those ≥60 yrs. The prevalence of OLLMM was estimated overall, by sex, age group, and diabetes status (prediabetes and T2DM) . We extrapolated results to the US using NHANES sampling weights. Of the 4174 adults (representing 181M US adults) , we identified 827 (28.8M, 15.9% of the US pop.) with OLLMM. Older adults had higher prevalence of OLLMM as did those with prediabetes and T2DM (Figure 1) . The prevalence of OLLMM in the US is high. While the prevalence of OLLMM was higher with age, its prevalence was also high in diabetics, regardless of age. Clinicians should monitor for OLLMM among obese T2DM patients. Disclosure D.J. Murdock: Employee; Regeneron Pharmaceuticals Inc. Stock/Shareholder; Regeneron Pharmaceuticals Inc. N. Wu: Employee; Regeneron Pharmaceuticals Inc. Stock/Shareholder; Regeneron Pharmaceuticals Inc. D.J. Glass: Employee; Regeneron Pharmaceuticals Inc. Stock/Shareholder; Regeneron Pharmaceuticals Inc. J.S. Grimsby: Employee; Regeneron Pharmaceuticals Inc. Stock/Shareholder; Regeneron Pharmaceuticals Inc. R.A. Calle: Employee; Regeneron Pharmaceuticals Inc. Stock/Shareholder; Regeneron Pharmaceuticals Inc. S. Donahue: Employee; Regeneron Pharmaceuticals Inc. Stock/Shareholder; Regeneron Pharmaceuticals Inc. M. Sleeman: Employee; Regeneron Pharmaceuticals Inc. Stock/Shareholder; Regeneron Pharmaceuticals Inc. R.J. Sanchez: Employee; Regeneron Pharmaceuticals Inc. Stock/Shareholder; Regeneron Pharmaceuticals Inc. Funding Supported by Regeneron Pharmaceuticals, Inc.

Leptin directly suppresses the activity of orexigenic neurons in the hypothalamic arcuate nucleus (ARC). We examined c-Fos-like immunoreactivity (CFLIR) as a marker of ARC neuronal activity in db/db mice devoid of the signaling form of the leptin receptor (LRb) and s/s mice that express LRbS1138 [which is defective for STAT3 (signal transducer and activator of transcription) signaling]. Both db/db and s/s animals are hyperphagic and obese. This analysis revealed that CFLIR in agouti related peptide-expressing orexigenic ARC neurons is basally elevated in db/db but not s/s mice. Consistent with these observations, electrophysiologic evaluation of a small number of neurons in s/s animals suggested that leptin appropriately suppresses the frequency of IPSCs on ARC proopiomelanocortin (POMC) neurons that are mediated by the release of GABA from orexigenic ARC neurons. CFLIR in POMC neurons of s/s mice was also increased compared with db/db animals. Thus, these data suggest that, although LRb→STAT3 signaling is crucial for the regulation of feeding, it is not required for the acute or chronic regulation of orexigenic ARC neurons, and the activation of STAT3-mediated transcription by leptin is not required for the appropriate development of leptin responsiveness in these neurons.

Tuberoinfundibular peptide of 39 residues (TIP39) was identified as a potent parathyroid hormone 2 receptor (PTH2R) agonist. Existing anatomical data also support the suggestion that TIP39 is the PTH2R's endogenous ligand, but a comprehensive comparison of TIP39 and PTH2R distributions has not been performed. In the present study, we compared the distributions of TIP39 and PTH2R on adjacent mouse brain sections. In addition, we determined the locations of PTH2R‐expressing cell bodies by in situ hybridization histochemistry and by labeling β‐galactosidase driven by the PTH2R promoter in knockin mice. An excellent correlation was found between the distributions of TIP39‐containing fibers and PTH2R‐containing cell bodies and fibers throughout the brain. TIP39 and the PTH2R are abundant in medial prefrontal, insular, and ectorhinal cortices, the lateral septal nucleus, the bed nucleus of the stria terminalis, the fundus striati, the amygdala, the ventral subiculum, the hypothalamus, midline and intralaminar thalamic nuclei, the medial geniculate body, the periaqueductal gray, the ventral tegmental area, the superior and inferior colliculi, the parabrachial nuclei, the locus coeruleus, subcoeruleus and periolivary areas, and the nucleus of the solitary tract. Furthermore, even the subregional distribution of TIP39‐ and PTH2R‐immunoreactive fibers in these regions showed remarkable similarities, providing anatomical evidence that TIP39 may act on the PTH2R. Based on these observations and on previous pharmacological data, we propose that TIP39 is an endogenous ligand of the PTH2R and that they form a neuromodulator system, which is optimally positioned to regulate limbic, endocrine, and auditory brain functions. J. Comp. Neurol. 502:563–583, 2007. Published 2007 Wiley‐Liss, Inc.