ACE2 pathway regulates thermogenesis and energy metabolism

Identification of key regulators of energy homeostasis holds important therapeutic promise for metabolic disorders, such as obesity and diabetes. ACE2 cleaves angiotensin II (Ang II) to generate Ang-(1-7) which acts mainly through the Mas1 receptor. Here, we identify ACE2 pathway as a critical regulator in the maintenance of thermogenesis and energy expenditure. We found that ACE2 is highly expressed in brown adipose tissue (BAT) and that cold stimulation increases ACE2 and Ang-(1-7) levels in BAT and serum. Ace2 knockout mice (Ace2-/y) and Mas1 knockout mice (Mas1-/-) displayed impaired thermogenesis. Mice transplanted with brown adipose tissue from Mas1-/- display metabolic abnormalities consistent with those seen in the Ace2 and Mas1 knockout mice. In contrast, impaired thermogenesis of Leprdb/db obese diabetic mice and high-fat diet-induced obese mice were ameliorated by overexpression of Ace2 or continuous infusion of Ang-(1-7). Activation of ACE2 pathway was associated with improvement of metabolic parameters, including blood glucose, lipids and energy expenditure in multiple animal models. Consistently, ACE2 pathway remarkably enhanced the browning of white adipose tissue. Mechanistically, we showed that ACE2 pathway activated Akt/FoxO1 and PKA pathway, leading to induction of UCP1 and activation of mitochondrial function. Our data propose that adaptive thermogenesis requires regulation of ACE2 pathway and highlight novel potential therapeutic targets for the treatment of metabolic disorders.

Transglutaminase mediated asprosin oligomerization allows its tissue storage as fibers

Asprosin, the C-terminal furin cleavage product of profibrillin-1, was reported to act as a hormone that circulates at nanomolar levels and is recruited to the liver where it induces G protein-coupled activation of the cAMP-PKA pathway and stimulates rapid glucose release into the circulation. Although derived from profibrillin-1, a multidomain extracellular matrix glycoprotein with a ubiquitous distribution in connective tissues, little is known about the tissue distribution of asprosin. In the current view, asprosin is mainly produced by white adipose tissue from where it is released into the blood in monomeric form. Here, by employing newly generated specific asprosin antibodies we monitored the distribution pattern of asprosin in human and murine connective tissues such as placenta, and muscle. Thereby we detected the presence of asprosin positive extracellular fibers. Further, by screening established cell lines for asprosin synthesis we found that most cells derived from musculoskeletal tissues render asprosin into an oligomerized form. This oligomerization is facilitated by transglutaminase activity and requires an intact fibrillin fiber network for proper linear deposition. Our data suggest a new extracellular storage mechanism of asprosin in oligomerized form which may regulate its cellular bioavailability in tissues.

Evaluation of spatial memory and anti-fatigue function of long-term supplementation of β-alanine and confirmation through cAMP-PKA and apoptosis pathways in mice

With an aim to explore the effects of <i>β</i>-alanine (<i>β</i>-A) on spatial memory and fatigue resistance, Kunming mice were treated with different concentrations of β-A (418, 836, and 2090 mg·kg^-1·day^-1). After gavage feeding with <i>β</i>-A for 10 weeks, results of the maze and MWM tests showed that <i>β</i>-A can enhance spatial learning and memory in mice. After evaluating the fatigue resistance, biochemical parameters (LG, GG, BUN, SOD, and MDA) showed significant differences in the low concentration treatment group compared to control group. Our data demonstrated that the appropriate dose of <i>β</i>-A can alleviate the oxidative stress and muscle fatigue in mice. Subsequently, expression of mRNA of key genes involved in cAMP-PKA pathway (PDE4A, MAPK1, adcy1, cAMP and CREB) was up regulated. Also, expression levels of apoptotic pathway genes were significantly affected as confirmed by qPCR and Western blotting. Our results demonstrated that <i>β</i>-A can enhance spatial learning and memory in mice via regulation of cAMP-PKA and apoptotic pathway.

Intermedin Inhibits the Ox-LDL–Induced Inflammation in RAW264.7 Cells by Affecting Fatty Acid–Binding Protein 4 Through the PKA Pathway

Objectives: Macrophages stimulated by oxidized low-density lipoprotein (ox-LDL) play an important role in the occurrence and progression of atherosclerosis. Fatty acid–binding protein 4 (FABP4), mainly existing in macrophages and adipocytes, can influence lipid metabolism and inflammation regulated by macrophages. Herein, we first established the connection between intermedin (IMD: a new peptide that has versatile biological activities in the cardiovascular system) and FABP4 and then investigated the influence of IMD on ox-LDL-induced changes in RAW264.7 macrophages line.Methods: The bioinformatics analysis, such as gene ontology enrichment and protein–protein interactions, was performed. For ox-LDL–stimulated assays, RAW264.7 was first pretreated with IMD and then exposed to ox-LDL. To explore the cell signaling pathways of IMD on inflammatory inhibition, main signaling molecules were tested and then cells were co-incubated with relevant inhibitors, and then exposed/not exposed to IMD. Finally, cells were treated with ox-LDL. The protein and gene expression of FABP4, IL-6, and TNF-α were quantified by WB/ELISA and RT-qPCR.Results: In the ox-LDL-stimulated assays, exposure of the RAW264.7 macrophages line to ox-LDL reduced cell viability and increased the expression of FABP4, as well as induced the release of IL-6 and TNF-α (all p &lt; 0.05). On the other hand, IMD prevented ox-LDL–induced cell toxicity, FABP4 expression, and the inflammatory level in RAW264.7 (all p &lt; 0.05) in a dose-dependent manner. The inhibition of FABP4 and the anti-inflammatory effect of IMD were partially suppressed by the protein kinase A (PKA) inhibitor H-89.Conclusion: IMD can prevent ox-LDL–induced macrophage inflammation by inhibiting FABP4, whose signaling might partially occur via the PKA pathway.

Protein Kinase SGK2 Is Induced by the β3 Adrenergic Receptor-cAMP-PKA-PGC-1α/NT-PGC-1α Axis but Dispensable for Brown/Beige Adipose Tissue Thermogenesis

Brown and beige adipocytes are specialized to dissipate energy as heat. Sgk2, encoding a serine/threonine kinase, has been identified as a brown and beige adipocyte-specific gene in rodents and humans; however, its function in brown/beige adipocytes remains unraveled. Here, we examined the regulation and role of Sgk2 in brown/beige adipose tissue thermogenesis. We found that transcriptional coactivators PGC-1α and NT-PGC-1α activated by the β3 adrenergic receptor-cAMP-PKA pathway are recruited to the Sgk2 promoter, triggering Sgk2 transcription in response to cold. SGK2 elevation was closely associated with increased serine/threonine phosphorylation of proteins carrying the consensus RxRxxS/T phosphorylation site. However, despite cold-dependent activation of SGK2, mice lacking Sgk2 exhibited normal cold tolerance at 4°C. In addition, Sgk2+/+ and Sgk2−/− mice induced comparable increases in energy expenditure during pharmacological activation of brown and beige adipose tissue with a β3AR agonist. In vitro loss- and gain-of-function studies further demonstrated that Sgk2 ablation or activation does not alter thermogenic gene expression and mitochondrial respiration in brown adipocytes. Collectively, our results reveal a new signaling component SGK2, although dispensable for cold-induced thermogenesis that adds an additional layer of complexity to the β3AR signaling network in brown/beige adipose tissue.

Intermedin1-53 Inhibits NLRP3 Inflammasome Activation by Targeting IRE1α in Cardiac Fibrosis

Intermedin (IMD), a paracrine/autocrine peptide, protects against cardiac fibrosis. However, the underlying mechanism remains poorly understood. Previous study reports that activation of Nucleotide-binding oligomerization domain (NOD)-like receptor family pyrin domain containing 3 (NLRP3) inflammasome contributed to cardiac fibrosis. In this study, we aimed to investigate whether IMD mitigates cardiac fibrosis by inhibiting NLRP3. Cardiac fibrosis was induced by angiotensin II (Ang II) infusion for 2 weeks in rats. Western blot, real-time PCR, histological staining, immunofluorescence assay, RNA sequencing, echocardiography and hemodynamics were used to detect the role and the mechanism of IMD in cardiac fibrosis. Ang II infusion resulted in rat cardiac fibrosis, shown as over-deposition of myocardial interstitial collagen and cardiac dysfunction. Importantly, NLRP3 activation and endoplasmic reticulum stress (ERS) was found in Ang II treated rat myocardium. Ang II infusion decreased the expression of IMD and increased the expression of the receptor system of IMD in the fibrotic rat myocardium. IMD treatment attenuated the cardiac fibrosis and improved cardiac function. In addition, IMD inhibited the upregulation of NLRP3 markers and ERS markers induced by Ang II. In vitro, IMD knockdown by small interfering RNA significantly promoted the Ang II-induced cardiac fibroblast and NLRP3 activation. Moreover, silencing of inositol requiring enzyme 1 α (IRE1α) blocked the effects of IMD inhibiting fibroblast and NLRP3 activation. Pre-incubation with PKA pathway inhibitor H89 blocked the effects of IMD on the anti-ERS, anti-NLRP3 and anti-fibrotic response. In conclusion, IMD alleviates cardiac fibrosis by inhibiting NLRP3 inflammasome activation via suppressing IRE1α and cAMP/PKA pathway.

Mitochondrial active Ras2 protein promotes apoptosis and regulated cell death in a cAMP/PKA pathway-dependent manner in budding yeast.

In previous papers, using the eGFP-RBD3 probe, which binds Ras-GTP with high affinity, we showed that activated Ras proteins are localized to the plasma membrane and in the nucleus in wild-type Saccharomyces cerevisiae cells growing exponentially on glucose, while an aberrant accumulation of activated Ras in mitochondria correlates to mitochondrial dysfunction, accumulation of ROS and an increase of apoptosis. In this paper, we show that lack of TPS1, which is known to trigger apoptosis in S. cerevisiae, induces localization of active Ras proteins in mitochondria, confirming the above-mentioned correlation. Next, by characterizing the ras1Δ and ras2Δ mutants concerning localization of active Ras proteins and propensity to undergo cell death, we show that active Ras2 proteins, which accumulate in the mitochondria following addition of acetic acid, a well-known pro-apoptotic stimulus, might be the GTPases involved in regulated cell death, while active Ras1 proteins, constitutively localized in mitochondria, might be involved in a pro-survival molecular machinery. Finally, by characterizing the gpa2Δ and cyr1Δ mutants concerning the propensity to undergo cell death, we show that active mitochondrial Ras proteins promote apoptosis through the cAMP/PKA pathway.

Bilateral Adrenal Hyperplasia: Pathogenesis and Treatment

Bilateral adrenal hyperplasia is a rare cause of Cushing’s syndrome. Micronodular adrenal hyperplasia, including the primary pigmented micronodular adrenal dysplasia (PPNAD) and the isolated micronodular adrenal hyperplasia (iMAD), can be distinguished from the primary bilateral macronodular adrenal hyperplasia (PBMAH) according to the size of the nodules. They both lead to overt or subclinical CS. In the latter case, PPNAD is usually diagnosed after a systematic screening in patients presenting with Carney complex, while for PBMAH, the diagnosis is often incidental on imaging. Identification of causal genes and genetic counseling also help in the diagnoses. This review discusses the last decades’ findings on genetic and molecular causes of bilateral adrenal hyperplasia, including the several mechanisms altering the PKA pathway, the recent discovery of ARMC5, and the role of the adrenal paracrine regulation. Finally, the treatment of bilateral adrenal hyperplasia will be discussed, focusing on current data on unilateral adrenalectomy.