PEGylated AdipoRon derivatives improve glucose and lipid metabolism under insulinopenic and high-fat diet conditions.

The pleiotropic actions of adiponectin in improving cell survival and metabolism have motivated the development of small-molecule therapeutic agents for treating diabetes and lipotoxicity. AdipoRon is a synthetic agonist of the adiponectin receptors, yet is limited by its poor solubility and bioavailability. In this work, we expand on the protective effects of AdipoRon in pancreatic ß-cells and examine how structural modifications could affect the activity, pharmacokinetics, and bioavailability of this small molecule. We describe a series of AdipoRon analogs containing amphiphilic ethylene glycol (PEG) chains. Among these, AdipoRonPEG5 induced pleiotropic effects in mice under insulinopenic and high-fat diet (HFD) conditions. While both AdipoRon and AdipoRonPEG5 substantially attenuate palmitate-induced lipotoxicity in INS-1 cells, only AdipoRonPEG5 treatment is accompanied by a significant reduction in cytotoxic ceramides. In vivo, AdipoRonPEG5 can substantially reduce pancreatic, hepatic, and serum ceramide species, with a concomitant increase in the corresponding sphingoid bases and improves insulin sensitivity of mice under HFD feeding conditions. Furthermore, hyperglycemia in streptozotocin (STZ)-induced insulinopenic adiponectin-null mice is also attenuated upon AdipoRonPEG5 treatment. Our results suggest that AdipoRonPEG5 is more effective in reducing ceramides and dihydroceramides in the liver of HFD-fed mice than AdipoRon, consistent with its potent activity in activating ceramidase in vitro in INS-1 cells. Additionally, these results indicate that the beneficial effects of AdipoRonPEG5 can be partially attributed to improved pharmacokinetics as compared with AdipoRon, thus suggesting that further derivatization may improve affinity and tissue-specific targeting.

ß1 Syntrophin Supports Autophagy Initiation and Protects against Cerulein-Induced Acute Pancreatitis.

Syntrophins are a family of proteins forming membrane-anchored scaffolds and serving as adaptors for various transmembrane and intracellular signaling molecules. To understand the physiological roles of ß1 syntrophin, one of the least characterized members, we generated mouse models to eliminate ß1 syntrophin specifically in the endocrine or exocrine pancreas. ß1 syntrophin is dispensable for the morphology and function of insulin-producing ß cells. However, mice with ß1 syntrophin deletion in exocrine acinar cells exhibit increased severity of cerulein-induced acute pancreatitis. Reduced expression of cystic fibrosis transmembrane conductance regulator and dilation of acinar lumen are potential predisposition factors. During the disease progression, a relative lack of autophagy is associated with deficiencies in both actin assembly and endoplasmic reticulum nucleation. Our findings reveal, for the first time, that ß1 syntrophin is a critical regulator of actin cytoskeleton and autophagy in pancreatic acinar cells and is potently protective against cerulein-induced acute pancreatitis.

Regulation of Dipeptidyl Peptidase-4, its Substrate Chemokines, and Their Receptors in Adipose Tissue of ob/ob Mice.

The physiological function of DPP-4 in proteolytic inactivation of incretins has been well established, however, there is limited information on the expression and the significance of DPP-4 in white adipose tissue with regard to obesity. The objective of the work was to reveal the expression and regulation of DPP-4 in adipocytes and compare the expression and activity of DPP-4 in white adipose tissue and several other organs such as the liver, muscle and kidney. We also investigated the gene expression levels of DPP-4 substrate chemokines, and their receptors in white adipose tissue. DPP-4 was mainly expressed in stromal vascular fraction (SVF), and downregulated in adipose tissue of ob/ob compared with C57BL6/J mice. Mimetic conditions of obese fat in vitro showed that differentiation of mouse primary preadipocytes into adipocytes was associated with marked downregulation of DPP-4 expression. Treatment with TNF-α or ROS even decreased DPP-4 expression in mouse primary adipocytes. Various DPP-4 substrate chemokines were expressed in white adipose tissue and regulated by obesity. The expression of receptors for DPP-4 substrate chemokines was markedly high and tightly regulated by obesity in white adipose tissue. Expression of DPP-4 was reduced in adipose tissues of ob/ob mice. Actions of several substrate chemokines might be potentiated by downregulation of DPP-4, synergistically with upregulation of chemokines and their receptors in adipose tissues of obese mice.

Nur77 gene expression levels were involved in different ACTH-secretion autonomy between Cushing's disease and subclinical Cushing's disease.

Cushing's disease (CD) and subclinical Cushing's disease (subCD) are both diseases caused by adrenocorticotropic hormone (ACTH)-secreting pituitary adenomas. However, ACTH autonomy in subCD is weaker than in CD and there are no Cushingoid features in subCD. The differences of molecular mechanisms in ACTH autonomy between CD and subCD have not yet been reported. Therefore, we aimed to investigate the differences in molecular mechanisms of ACTH-secretion autonomy between CD and subCD. The study included 23 patients [7 CD, 6 subCD, and 10 non-functioning pituitary tumors (NFTs)] who underwent transsphenoidal surgery at the Osaka University Hospital between December 2009 and October 2013. Using quantitative real-time PCR, various ACTH-related gene expressions in tumor tissues from CD, subCD, and NFT were measured such as pro-opiomelanocortin (POMC), POMC transcription factor (Tpit, Pitx1, NeuroD1, and Nur77), POMC peptide processing enzymes (prohormone convertase: PC1/3 and PC2), and ACTH secretion-related factors (corticotropin-releasing hormone receptor 1: CRHR1 and glucocorticoid receptor α: GRα). Only Nur77 mRNA levels were significantly higher in CD than in subCD. Furthermore, we stained 6 CD and 6 subCD with anti-Nur77 antibody. All tumor samples from CD had Nur77 protein positive cells. On the other hand, Nur77 protein was expressed in only one tumor sample from subCD. This sample showed high expression of Nur77 mRNA. Nur77 is an important to regulate POMC transcription and negative-feedback by glucocorticoids. Nur77 gene expression levels might involve different autonomy of ACTH production between CD and subCD.

Clinical significance of fluctuations in thyroid hormones after surgery for Cushing's syndrome.

Patients with Cushing's syndrome (CS) frequently develop hyperthyroidism after surgery due to SITSH (syndrome of inappropriate secretion of TSH) and this SITSH contributed to the symptoms of steroid withdrawal syndrome (SWS). However, the duration of fluctuations in thyroid hormones after surgery for CS remains unknown. The aim of this prospective study was to investigate the clinical course of fluctuation in thyroid hormone level in CS patients after surgery. Thyroid hormone levels [free T3 (FT3), free T4 (FT4) and TSH] and serum cortisol levels were measured before and 1, 3, 6 and 12 months after surgery in 8 patients with active CS (3 pituitary CS and 5 adrenal CS). FT3 levels were above the normal range in 75% of patients up to 6 months after surgery, but returned to the normal range by 12 months. However, TSH levels were not suppressed below the normal range throughout the first 12 months after surgery. Serious symptoms of SWS appeared during the 6-month period after surgery, but disappeared with normalization of thyroid function at 12 months, which was not related to the recovery of function hypothalamus-pituitary-adrenal axis after CS surgery. Therefore, T3 toxicosis could result in deterioration of SWS after surgery for CS. These results indicate that physicians need to take T3 toxicosis into consideration in the pathological evaluation of SWS within 12 months after surgery for CS.

Rapid decline in bone turnover markers but not bone mineral density in acromegalic patients after transsphenoidal surgery.

Growth hormone (GH) and insulin-like growth factor-I (IGF-I) play important roles in maintaining bone metabolism and bone mineral density (BMD) in adulthood, in addition to stimulating longitudinal bone growth in childhood. However, information on the effect of GH excess on bone metabolism and BMD is incomplete and requires further analysis. The aim of this study is to clarify the effect of rapid decline in GH levels after transsphenoidal surgery (TSS) on bone metabolism in acromegalic patients. In this prospective study, 22 patients (11 males and 11 females) with active acromegaly underwent TSS. Bone formation marker (serum bone alkaline phosphatase: BAP), bone resorption marker (urinary type I collagen cross-linked N-telopeptide: urinary NTx) and BMD were measured before and at 3 and 12 months after TSS. BAP was significantly decreased at 12 months after TSS, but not at 3 months. Urinary NTx was significantly decreased at 3 and 12 months after TSS. BMD did not change after TSS. In conclusion, the rapid fall in GH level after TSS had no effect on BMD for up to 12 months after TSS despite the decrease in markers of bone formation and resorption.

Downregulation of the kidney glucagon receptor, essential for renal function and systemic homeostasis, contributes to chronic kidney disease.

The glucagon receptor (GCGR) in the kidney is expressed in nephron tubules. In humans and animal models with chronic kidney disease, renal GCGR expression is reduced. However, the role of kidney GCGR in normal renal function and in disease development has not been addressed. Here, we examined its role by analyzing mice with constitutive or conditional kidney-specific loss of the Gcgr. Adult renal Gcgr knockout mice exhibit metabolic dysregulation and a functional impairment of the kidneys. These mice exhibit hyperaminoacidemia associated with reduced kidney glucose output, oxidative stress, enhanced inflammasome activity, and excess lipid accumulation in the kidney. Upon a lipid challenge, they display maladaptive responses with acute hypertriglyceridemia and chronic proinflammatory and profibrotic activation. In aged mice, kidney Gcgr ablation elicits widespread renal deposition of collagen and fibronectin, indicative of fibrosis. Taken together, our findings demonstrate an essential role of the renal GCGR in normal kidney metabolic and homeostatic functions. Importantly, mice deficient for kidney Gcgr recapitulate some of the key pathophysiological features of chronic kidney disease.

Adipogenin Dictates Adipose Tissue Expansion by Facilitating the Assembly of a Dodecameric Seipin Complex.

Adipogenin (Adig) is an evolutionarily conserved microprotein and is highly expressed in adipose tissues and testis. Here, we identify Adig as a critical regulator for lipid droplet formation in adipocytes. We determine that Adig interacts directly with seipin, leading to the formation of a rigid complex. We solve the structure of the seipin/Adig complex by Cryo-EM at 2.98Å overall resolution. Surprisingly, seipin can form two unique oligomers, undecamers and dodecamers. Adig selectively binds to the dodecameric seipin complex. We further find that Adig promotes seipin assembly by stabilizing and bridging adjacent seipin subunits. Functionally, Adig plays a key role in generating lipid droplets in adipocytes. In mice, inducible overexpression of Adig in adipocytes substantially increases fat mass, with enlarged lipid droplets. It also elevates thermogenesis during cold exposure. In contrast, inducible adipocyte-specific Adig knockout mice manifest aberrant lipid droplet formation in brown adipose tissues and impaired cold tolerance.

Obstructive sleep apnea syndrome causes a pseudo-Cushing's state in Japanese obese patients with type 2 diabetes mellitus.

Activation of the hypothalamic-pituitary-adrenal axis has been reported in some patients with the obstructive sleep apnea syndrome (OSAS). In current study, we investigated whether OSAS affect the screening test for subclinical Cushing's disease using 0.5 mg overnight dexamethasone suppression test (DST) in Japanese obese diabetic patients with OSAS. Among Japanese obese patients with type 2 diabetes mellitus who had been hospitalized in our department, we selected 20 patients with moderate to severe untreated OSAS (apnea-hypoxia index, AHI, of ≥15 events/hour). All patients underwent 0.5 mg DST. The same test was repeated in patients with positive response of it within a few days after continuous positive airway pressure (CPAP) therapy. We found that five patients showed positive response of DST (25%). Three of these patients continued to use CPAP, and they showed normal response of DST after CPAP therapy. Serum cortisol after 0.5 mg DST measured before CPAP therapy correlated significantly with fasting serum cortisol level (r=0.764, p<0.0001), but not with various clinical parameters, including AHI (p=0.784), body mass index (p=0.984), waist circumference (p=0.957), HbA1c (p=0.261), fasting plasma glucose (p=0.420) and HOMA-IR (p=0.500). Our study show that OSAS causes a pseudo-Cushing's syndrome in obese patients with type 2 diabetes mellitus, which phenomena can be reversed by CPAP therapy.

HSP47 levels determine the degree of body adiposity.

Adiposity varies among individuals with the influence of diverse physiological, pathological, environmental, hormonal, and genetic factors, but a unified molecular basis remains elusive. Here, we identify HSP47, a collagen-specific chaperone, as a key determinant of body adiposity. HSP47 expression is abundant in adipose tissue; increased with feeding, overeating, and obesity; decreased with fasting, exercise, calorie restriction, bariatric surgery, and cachexia; and correlated with fat mass, BMI, waist, and hip circumferences. Insulin and glucocorticoids, respectively, up- and down-regulate HSP47 expression. In humans, the increase of HSP47 gene expression by its intron or synonymous variants is associated with higher body adiposity traits. In mice, the adipose-specific knockout or pharmacological inhibition of HSP47 leads to lower body adiposity compared to the control. Mechanistically, HSP47 promotes collagen dynamics in the folding, secretion, and interaction with integrin, which activates FAK signaling and preserves PPARγ protein from proteasomal degradation, partly related to MDM2. The study highlights the significance of HSP47 in determining the amount of body fat individually and under various circumstances.