Neuregulin-4 protects cardiomyocytes against high-glucose-induced ferroptosis via the AMPK/NRF2 signalling pathway.

BACKGROUND: High glucose levels are key factors and key contributors to several cardiovascular diseases associated with cardiomyocyte injury. Ferroptosis, which was identified in recent years, is a mode of cell death caused by the iron-mediated accumulation of lipid peroxides. Neuregulin-4 (Nrg4) is an adipokine that has protective effects against metabolic disorders and insulin resistance. Our previous study revealed that Nrg4 has a protective effect against diabetic myocardial injury, and the aim of this study was to investigate whether Nrg4 could attenuate the occurrence of high glucose-induced ferroptosis in cardiomyocytes. METHODS: We constructed an in vivo diabetic myocardial injury model in which primary cardiomyocytes were cultured in vitro and treated with Nrg4. Changes in ferroptosis-related protein levels and ferroptosis-related indices in cardiomyocytes were observed. In addition, we performed back-validation and explored signalling pathways that regulate ferroptosis in primary cardiomyocytes. RESULTS: Nrg4 attenuated cardiomyocyte ferroptosis both in vivo and in vitro. Additionally, the AMPK/NRF2 signalling pathway was activated during this process, and when the AMPK/NRF2 pathway was inhibited, the beneficial effects of Nrg4 were attenuated. CONCLUSION: Nrg4 antagonizes high glucose-induced ferroptosis in cardiomyocytes via the AMPK/NRF2 signalling pathway.

Low circulating levels of neuregulin 4 as a potential biomarker associated with the severity and prognosis of obesity-related metabolic diseases: a systematic review.

BACKGROUND: Neuregulin 4 (Nrg4) is a brown adipose tissue-derived adipokine that greatly affects systemic metabolism and improves metabolic derangements. Although abnormal circulating levels of Nrg4 are common in obesity, it remains elusive whether low or elevated levels of this batokine are associated with the onset of metabolic diseases. AIM: To assess Nrg4 levels and its role as a feasible biomarker to predict the severity of obesity, gestational diabetes mellitus (GDM), type 2 diabetes mellitus (T2DM), non-alcoholic fatty liver disease (NAFLD), and cardiovascular diseases (CVD). METHODS: A search for relevant studies was performed systematically using prominent search engines, including PubMed, Google Scholar, and Embase, by following PRISMA guidelines. RESULTS: Ample clinical evidence reported low serum/plasma levels of Nrg4 in obesity and these were inversely proportional to the indices of metabolic syndrome, including body mass index, waist circumference, triglycerides, fasting plasma glucose, and homoeostatic model assessment for insulin resistance as well as high-sensitivity C-reactive protein. Low circulating Nrg4 levels may aid in the prediction of morbid obesity, and subsequent GDM, T2DM, NAFLD, and CVD. CONCLUSION: Current clinical evidence emphasizes that the circulating levels of Nrg4 are decreased in morbid obesity, and it also highlights that Nrg4 May serve as a potential prognostic biomarker for obesity-related metabolic diseases.

Neuregulin 4 Attenuates Podocyte Injury and Proteinuria in Part by Activating AMPK/mTOR-Mediated Autophagy in Mice.

In this study, we investigate the effect of neuregulin 4 (NRG4) on podocyte damage in a mouse model of diabetic nephropathy (DN) and we elucidate the underlying molecular mechanisms. In vivo experiments were conducted using a C57BL/6 mouse model of DN to determine the effect of NRG4 on proteinuria and podocyte injury, and in vitro experiments were performed with conditionally immortalized mouse podocytes treated with high glucose and NRG4 to assess the protective effects of NRG4 on podocyte injury. Autophagy-related protein levels and related signaling pathways were evaluated both in vivo and in vitro. The involvement of the adenosine monophosphate-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathway was detected using chloroquine or AMPK inhibitors. The results showed that the AMPK/mTOR pathway was involved in the protective roles of NRG4 against high glucose-mediated podocyte injury. Also, NRG4 significantly decreased albuminuria in DN mice. PAS staining indicated that NRG4 mitigated glomerular volume and mesangium expansion in DN mice. Consistently, western blot and RT-PCR analyses confirmed that NRG4 decreased the expression of pro-fibrotic molecules in the glomeruli of DN mice. The immunofluorescence results showed that NRG4 retained expression of podocin and nephrin, whereas transmission electron microscopy revealed that NRG4 alleviated podocyte injury. In DN mice, NRG4 decreased podocyte apoptosis and increased expression of nephrin and podocin, while decreasing the expression of desmin and HIF1α. Overall, NRG4 improved albuminuria, glomerulosclerosis, glomerulomegaly, and hypoxia in DN mice. The in vitro experiments showed that NRG4 inhibited HG-induced podocyte injury and apoptosis. Furthermore, autophagy of the glomeruli decreased in DN mice, but reactivated following NRG4 intervention. NRG4 intervention was found to partially activate autophagy via the AMPK/mTOR signaling pathway. Consequently, when the AMPK/mTOR pathway was suppressed or autophagy was inhibited, the beneficial effects of NRG4 intervention on podocyte injury were diminished. These results indicate that NRG4 intervention attenuates podocyte injury and apoptosis by promoting autophagy in the kidneys of DN mice, in part, by activating the AMPK/mTOR signaling pathway.

Neuregulin4-ErbB4 signalling pathway is driven by electroacupuncture stimulation to remodel brown adipose tissue innervation.

AIM: To show that electroacupuncture stimulation (ES) remodels sympathetic innervation in brown adipose tissue (BAT) via the bone morphogenic protein 8B (BMP8B)-neuregulin 4 (NRG4)-ErbB4 axis, with somatotopic dependence. MATERIALS AND METHODS: We established a high-fat diet (HFD) model with C57BL/6J mice to measure the thermogenesis and metabolism of BAT. In addition, the sympathetic nerve activity (SNA) was measured with the electrophysiological technique, and the immunostaining of c-Fos was used to detect the central nervous system sources of sympathetic outflows. Finally, the key role of the BMP8B-NRG4-ErbB4 axis was verified by peripheral specific antagonism of ErbB4. RESULTS: ES at the forelimb and abdomen regions significantly up-regulate SNA, whereas ES at the hindlimb region has a limited regulatory effect on SNA but still partially restores HFD-induced BAT dysfunction. Mechanistically, ES at the forelimb and abdomen regions driving catecholaminergic signals in brown adipocytes depends on neural activities projected from the ventromedial nucleus of the hypothalamus (VMH) to the spinal cord intermediolateral column (IML). Notably, the peripheral suppression of ErbB4 in BAT inhibits the thermogenesis and metabolic function of BAT, as well as significantly hindering the SNA activation and metabolic benefits induced by ES. CONCLUSION: These results suggest that ES appears to be an effective approach for remodeling sympathetic innervation in BAT, which is closely related to neuronal activity in the VMH and the NRG4-ErbB4 signaling pathway.

The browning and mobilization of subcutaneous white adipose tissue supports efficient skin repair.

Adipocytes in dermis are considered to be important participants in skin repair and regeneration, but the role of subcutaneous white adipose tissue (sWAT) in skin repair is poorly understood. Here, we revealed the dynamic changes of sWAT during wound healing process. Lineage-tracing mouse studies revealed that sWAT would enter into the large wound bed and participate in the formation of granulation tissue. Moreover, sWAT undergoes beiging after skin injury. Inhibition of sWAT beiging by genetically silencing PRDM16, a key regulator to beiging, hindered wound healing process. The transcriptomics results suggested that beige adipocytes in sWAT abundantly express neuregulin 4 (NRG4), which regulated macrophage polarization and the function of myofibroblasts. In diabetic wounds, the beiging of sWAT was significantly suppressed. Thus, adipocytes from sWAT regulate multiple aspects of repair and may be therapeutic for inflammatory diseases and defective wound healing associated with aging and diabetes.

Adipocyte secreted NRG4 ameliorates age-associated metabolic dysfunction.

With the progressive aging of society, there is an increasing prevalence of age-related diseases that pose a threat to the elderly's quality of life. Adipose tissue, a vital energy reservoir with endocrine functions, is one of the most vulnerable tissues in aging, which in turn influences systematic aging process, including metabolic dysfunction. However, the underlying mechanism is still poorly understood. In this study, we found that NRG4, a novel adipokine, is obviously decreased in adipocyte tissues and serums during aging. Moreover, delivered recombinant NRG4 protein (rNRG4) into aged mice can ameliorate age-associated insulin resistance, glucose disorders and other metabolic disfunction. In addition, rNRG4 treatment alleviates age-associated hepatic steatosis and sarcopenia, accompanied with altered gene signatures. Together, these results indicate that NRG4 plays a key role in the aging process and is a therapeutic target for the treatment of age-associated metabolic dysfunction.

Neuregulin 2 Is a Candidate Gene for Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder with heterogeneous and complex genetic underpinnings. Our previous microarray gene expression profiling identified significantly different neuregulin-2 gene (NRG2) expression between ASD patients and controls. Thus, we aimed to clarify whether NRG2 is a candidate gene associated with ASD. The study consisted of two stages. First, we used real-time quantitative PCR in 20 ASDs and 20 controls to confirm the microarray gene expression profiling results. The average NRG2 gene expression level in patients with ASD (3.23 ± 2.80) was significantly lower than that in the controls (9.27 ± 4.78, p < 0.001). Next, we conducted resequencing of all the exons of NRG2 in a sample of 349 individuals with ASD, aiming to identify variants of the NRG2 associated with ASD. We identified three variants, including two single nucleotide variants (SNVs), IVS3 + 13A > G (rs889022) and IVS10 + 32T > A (rs182642591), and one small deletion at exon 11 of NRG2 (delGCCCGG, rs933769137). Using data from the Taiwan Biobank as the controls, we found no significant differences in allele frequencies of rs889022 and rs182642591 between two groups. However, there is a significant difference in the genotype and allele frequency distribution of rs933769137 between ASDs and controls (p < 0.0001). The small deletion is located in the EGF-like domain at the C-terminal of the NRG2 precursor protein. Our findings suggest that NRG2 might be a susceptibility gene for ASD.

Biological markers of adipose tissue: Adipokines.

We currently have a large sum of clinical and experimental data documenting the involvement of numerous adipokines in the maintenance of energy homeostasis in healthy individuals and their dysregulation in diseases such as obesity, metabolic syndrome or type 2 diabetes. Despite the impressive discoveries made in this field over many years, much remains to be done before understanding all the physiological and pathological implications, and hoping for the development of other effective and safe therapeutic strategies. Two original adipokines will be taken as examples to illustrate these remarks, chemerin and neuregulin 4.

Lipid metabolic reprogramming mediated by circulating Nrg4 alleviates metabolic dysfunction-associated steatotic liver disease during the early recovery phase after sleeve gastrectomy.

BACKGROUND: The metabolic benefits of bariatric surgery that contribute to the alleviation of metabolic dysfunction-associated steatotic liver disease (MASLD) have been reported. However, the processes and mechanisms underlying the contribution of lipid metabolic reprogramming after bariatric surgery to attenuating MASLD remain elusive. METHODS: A case-control study was designed to evaluate the impact of three of the most common adipokines (Nrg4, leptin, and adiponectin) on hepatic steatosis in the early recovery phase following sleeve gastrectomy (SG). A series of rodent and cell line experiments were subsequently used to determine the role and mechanism of secreted adipokines following SG in the alleviation of MASLD. RESULTS: In morbidly obese patients, an increase in circulating Nrg4 levels is associated with the alleviation of hepatic steatosis in the early recovery phase following SG before remarkable weight loss. The temporal parameters of the mice confirmed that an increase in circulating Nrg4 levels was initially stimulated by SG and contributed to the beneficial effect of SG on hepatic lipid deposition. Moreover, this occurred early following bariatric surgery. Mechanistically, gain- and loss-of-function studies in mice or cell lines revealed that circulating Nrg4 activates ErbB4, which could positively regulate fatty acid oxidation in hepatocytes to reduce intracellular lipid deposition. CONCLUSIONS: This study demonstrated that the rapid effect of SG on hepatic lipid metabolic reprogramming mediated by circulating Nrg4 alleviates MASLD.

Correlation between neuregulin receptor degradation protein-1 and Crohn's disease.

Neuregulin receptor degradation protein-1 (Nrdp1) is a newly discovered E3 ligase that plays a role in the apoptosis process of multiple diseases. Previous studies has shown that Nrdp1 exerted a proapoptotic effect in cardiac diseases. The purpose of this study is to investigate the potential involvement of Nrdp1 in the pathological processes of inflammatory bowel disease (IBD). To create a mouse model of experimental colitis, trinitrobenzenesulfonic acid (TNBS) was administered and the severity of colitis was assessed based on changes in weight and histological scores. Using Western blot and immunohistochemistry, significant increase in Nrdp1 expression was observed in intestinal epithelial cells (IECs). This was accompanied with the up-regulation of cleaved PARP and active caspase-3 in IECs, indicating a potential function in IECs. To study this further, we built an in vitro model of tumor necrosis factor-alpha (TNF-α)-induced apoptosis using human IEC line HT-29 cells. When Nrdp1 was knocked down, a decrease in apoptosis was observed, suggesting that Nrdp1 may play a proapoptotic role in IEC apoptosis. The mechanism behind this phenomenon is associated with the suppression of downstream targets of Nrdp1, such as protein kinase B (AKT). Furthermore, immunohistochemistry analysis in patients with Crohn's disease (CD) and normal controls supported the same results as observed in experimental colitis. We conclude that Nrdp1 may be a promising new therapeutic target for ameliorating IBD in humans.

Neuregulin 4 (Nrg4) cooperates with melatonin to regulate the PRL expression via ErbB4/Erk signaling pathway as a potential prolactin (PRL) regulator.

Neuregulin-4 (Nrg4) and melatonin play vital roles in endocrine diseases. However, there is little discussion about the function and potential mechanism of Nrg4 and melatonin in prolactin (PRL) regulation. The human normal pituitary data from Gene Expression Profiling Interactive Analysis (GEPIA) database was used to explore the correlation between NRG4 and PRL. The expression and correlation of NRG4 and PRL were determined by Immunofluorescence staining (IF) and human normal pituitary tissue microarray. Western Blot (WB) was used to detect the expression of PRL, p-ErbB2/3/4, ErbB2/3/4, p-Erk1/2, Erk1/2, p-Akt and Akt in PRL-secreting pituitary GH3 and RC-4B/C cells treated by Nrg4, Nrg4-small interfering RNA, Erk1/2 inhibitor FR180204 and melatonin. The expression of NRG4 was significantly positively correlated with that of PRL in the GEPIA database and normal human pituitary tissues. Nrg4 significantly increased the expression and secretion of PRL and p-Erk1/2 expression in GH3 cells and RC-4B/C cells. Inhibition of Nrg4 significantly inhibited PRL expression. The increased levels of p-Erk1/2 and PRL induced by Nrg4 were abolished significantly in response to FR180204 in GH3 and RC-4B/C cells. Additionally, Melatonin promotes the expression of Nrg4, p-ErbB4, p-Erk1/2, and PRL and can further promote the expression of p-Erk1/2 and PRL in combination with Nrg4. Further investigation into the function of Nrg4 and melatonin on PRL expression and secretion may provide new clues to advance the clinical control of prolactinomas and hyperprolactinemia.

Nicotine use disorder and Neuregulin 3: Opportunities for precision medicine.

Nicotine use disorder remains a major public health emergency despite years of trumpeting the consequences of smoking. This is likely due to the complex interplay of genetics and nicotine exposure across the lifespan of these individuals. Genetics influence all aspects of life, including complex disorders such as nicotine use disorder. This review first highlights the critical neurocircuitry underlying nicotine dependence and withdrawal, and then describes the cellular signaling mechanisms involved. Finally, current genetic, genomic, and transcriptomic evidence for new drug development of smoking cessation aids is discussed, with a focus on the Neuregulin 3 Signaling Pathway.

Dynamic effects of ventral hippocampal NRG3/ERBB4 signaling on nicotine withdrawal-induced responses.

Tobacco smoking remains a leading cause of preventable death in the United States, with approximately a 5% success rate for smokers attempting to quit. High relapse rates have been linked to several genetic factors, indicating that the mechanistic relationship between genes and drugs of abuse is a valuable avenue for the development of novel smoking cessation therapies. For example, various single nucleotide polymorphisms (SNPs) in the gene for neuregulin 3 (NRG3) and its cognate receptor, the receptor tyrosine-protein kinase erbB-4 (ERBB4), have been linked to nicotine addiction. Our lab has previously shown that ERBB4 plays a role in anxiety-like behavior during nicotine withdrawal (WD); however, the neuronal mechanisms and circuit-specific effects of NRG3-ERBB4 signaling during nicotine and WD are unknown. The present study utilizes genetic, biochemical, and functional approaches to examine the anxiety-related behavioral and functional role of NRG3-ERBB4 signaling, specifically in the ventral hippocampus (VH) of male and female mice. We report that 24hWD from nicotine is associated with altered synaptic expression of VH NRG3 and ERBB4, and genetic disruption of VH ErbB4 leads to an elimination of anxiety-like behaviors induced during 24hWD. Moreover, we observed attenuation of GABAergic transmission as well as alterations in Ca2+-dependent network activity in the ventral CA1 area of VH ErbB4 knock-down mice during 24hWD. Our findings further highlight contributions of the NRG3-ERBB4 signaling pathway to anxiety-related behaviors seen during nicotine WD.

Neuregulin 4 mediates the metabolic benefits of mild cold exposure by promoting beige fat thermogenesis.

Interorgan crosstalk via secreted hormones and metabolites is a fundamental aspect of mammalian metabolic physiology. Beyond the highly specialized endocrine cells, peripheral tissues are emerging as an important source of metabolic hormones that influence energy and nutrient metabolism and contribute to disease pathogenesis. Neuregulin 4 (Nrg4) is a fat-derived hormone that protects mice from nonalcoholic steatohepatitis (NASH) and NASH-associated liver cancer by shaping hepatic lipid metabolism and the liver immune microenvironment. Despite its enriched expression in brown fat, whether NRG4 plays a role in thermogenic response and mediates the metabolic benefits of cold exposure are areas that remain unexplored. Here we show that Nrg4 expression in inguinal white adipose tissue (iWAT) is highly responsive to chronic cold exposure. Nrg4 deficiency impairs beige fat induction and renders mice more susceptible to diet-induced metabolic disorders under mild cold conditions. Using mice with adipocyte and hepatocyte-specific Nrg4 deletion, we reveal that adipose tissue-derived NRG4, but not hepatic NRG4, is essential for beige fat induction following cold acclimation. Furthermore, treatment with recombinant NRG4-Fc fusion protein promotes beige fat induction in iWAT and improves metabolic health in mice with diet-induced obesity. These findings highlight a critical role of NRG4 in mediating beige fat induction and preserving metabolic health under mild cold conditions.

Effects of oleoylethanolamide supplementation on the expression of lipid metabolism-related genes and serum NRG4 levels in patients with non-alcoholic fatty liver disease: A randomized controlled trial.

BACKGROUND: This study investigated the effects of oleoylethanolamide (OEA) supplementation on the expression levels of SIRT1, AMPK, PGC-1α, PPAR-γ, CEBP-α and CEBP-ß genes and serum neuregulin 4 (NRG4) levels in patients with non-alcoholic fatty liver diseases (NAFLD). METHODS: Sixty obese patients with NAFLD were equally allocated into either OEA or placebo group for 12 weeks. The mRNA expression levels of genes were determined using the reverse transcription polymerase chain reaction (RT-PCR) technique. Serum NRG4 level was also assessed using an enzyme-linked immunosorbent assay (ELISA) kit. RESULTS: At the endpoint, mRNA expression levels of SIRT1(p = 0.001), PGC-1α (p = 0.011) and AMPK (p = 0.019) were significantly higher in the OEA group compared to placebo group. However, no significant differences were observed in the expression levels of PPAR-γ, CEBP-α and CEBP-ß between the two groups. Serum NRG4 levels significantly increased in the OEA group compared with the placebo group after controlling for confounders (p = 0.027). In the OEA group, significant relationships were found between percent of changes in the expression levels of the SIRT1, AMPK and PGC-1α as well as serum NRG4 level with percent of changes in some anthropometric measures. Moreover, in the intervention group, percent of changes in high-density lipoprotein cholesterol was positively correlated with percent of changes in the expression levels of the SIRT1 and AMPK. While, percent of changes in triglyceride was inversely correlated with percent of changes in the expression levels of SIRT1. CONCLUSION: OEA could beneficially affect expression levels of some lipid metabolism-related genes and serum NRG4 level. "REGISTERED UNDER IRANIAN REGISTRY OF CLINICAL TRIALS IDENTIFIER NO: IRCT20090609002017N32".

ER-PM Junctions on GABAergic Interneurons Are Organized by Neuregulin 2/VAP Interactions and Regulated by NMDA Receptors.

Neuregulins (NRGs) signal via ErbB receptors to regulate neural development, excitability, synaptic and network activity, and behaviors relevant to psychiatric disorders. Bidirectional signaling between NRG2/ErbB4 and NMDA receptors is thought to homeostatically regulate GABAergic interneurons in response to increased excitatory neurotransmission or elevated extracellular glutamate levels. Unprocessed proNRG2 forms discrete clusters on cell bodies and proximal dendrites that colocalize with the potassium channel Kv2.1 at specialized endoplasmic reticulum-plasma membrane (ER-PM) junctions, and NMDA receptor activation triggers rapid dissociation from ER-PM junctions and ectodomain shedding by ADAM10. Here, we elucidate the mechanistic basis of proNRG2 clustering at ER-PM junctions and its regulation by NMDA receptors. Importantly, we demonstrate that proNRG2 promotes the formation of ER-PM junctions by directly binding the ER-resident membrane tether VAP, like Kv2.1. The proNRG2 intracellular domain harbors two non-canonical, low-affinity sites that cooperatively mediate VAP binding. One of these is a cryptic and phosphorylation-dependent VAP binding motif that is dephosphorylated following NMDA receptor activation, thus revealing how excitatory neurotransmission promotes the dissociation of proNRG2 from ER-PM junctions. Therefore, proNRG2 and Kv2.1 can independently function as VAP-dependent organizers of neuronal ER-PM junctions. Based on these and prior studies, we propose that proNRG2 and Kv2.1 serve as co-regulated downstream effectors of NMDA receptors to homeostatically regulate GABAergic interneurons.

Neuregulin modulates hormone receptor levels in breast cancer through concerted action on multiple signaling pathways.

The Neuregulins (NRGs) are growth factors that bind and activate ErbB/HER receptor tyrosine kinases. Some reports have described an interplay between this ligand-receptor system and hormonal receptors in breast cancer. However, the mechanisms by which NRGs regulate hormonal receptor signaling have not been sufficiently described. Here, we show that in breast cancer cells the activation of NRG receptors down-regulated ERα through a double mechanism that included post-transcriptional and transcriptional effects. This regulation required the concerted participation of three signaling routes: the PI3K/AKT/mTOR, ERK1/2, and ERK5 pathways. Moreover, these three routes were also involved in the phosphorylation of ERα at serines 118 and 167, two residues implicated in resistance to endocrine therapies. On the other hand, NRGs conferred resistance to fulvestrant in breast cancer cells and this resistance could be reversed when the three pathways activated by NRGs were simultaneously inhibited. Our results indicate that estrogen receptor-positive (ER+) breast tumors that can have access to NRGs may be resistant to fulvestrant. This resistance could be overcome if strategies to target the three main pathways involved in the interplay between NRG receptors and ERα could be developed.

The relevance of EGFR, ErbB receptors and neuregulins in human adipocytes and adipose tissue in obesity.

OBJECTIVE: To investigate the potential role of EGFR, ErbBs receptors and neuregulins in human adipose tissue physiology in obesity. METHODS: Gene expression analysis in human subcutaneous (SAT) and visceral (VAT) adipose tissue in three independent cohorts [two cross-sectional (N = 150, N = 87) and one longitudinal (n = 25)], and in vitro gene knockdown and overexpression experiments were performed. RESULTS: While both SAT and VAT ERBB2 and ERBB4 mRNA increased in obesity, SAT EGFR mRNA was negatively correlated with insulin resistance, but did not change in obesity. Of note, both SAT and VAT EGFR mRNA were significantly associated with adipogenesis and increased during human adipocyte differentiation. In vitro experiments revealed that EGFR, but not ERBB2 and ERBB4, gene knockdown in preadipocytes and in fully differentiated human adipocytes resulted in decreased expression of adipogenic-related genes. ERBB2 gene knockdown also reduced gene expression of fatty acid synthase in fully differentiated adipocytes. In addition, neuregulin 2 (NRG2) mRNA was associated with expression of adipogenic genes in human adipose tissue and adipocytes, and its overexpression increased expression of EGFR and relevant adipogenic genes. CONCLUSIONS: This study demonstrates the association between adipose tissue ERBB2 and obesity, confirms the relevance of EGFR on human adipogenesis, and suggests a possible adipogenic role of NRG2.

Brown adipose tissue-derived Nrg4 alleviates endothelial inflammation and atherosclerosis in male mice.

Brown adipose tissue (BAT) activity contributes to cardiovascular health by its energy-dissipating capacity but how BAT modulates vascular function and atherosclerosis through endocrine mechanisms remains poorly understood. Here we show that BAT-derived neuregulin-4 (Nrg4) ameliorates atherosclerosis in mice. BAT-specific Nrg4 deficiency accelerates vascular inflammation and adhesion responses, endothelial dysfunction and apoptosis and atherosclerosis in male mice. BAT-specific Nrg4 restoration alleviates vascular inflammation and adhesion responses, attenuates leukocyte homing and reduces endothelial injury and atherosclerosis in male mice. In endothelial cells, Nrg4 decreases apoptosis, inflammation and adhesion responses induced by oxidized low-density lipoprotein. Mechanistically, protein kinase B (Akt)-nuclear factor-κB signaling is involved in the beneficial effects of Nrg4 on the endothelium. Taken together, the results reveal Nrg4 as a potential cross-talk factor between BAT and arteries that may serve as a target for atherosclerosis.

Functional EGF domain of the human neuregulin 1α produced in Escherichia coli with accurate disulfide bonds.

BACKGROUND: Neuregulins comprise a large family of growth factors containing an epidermal growth factor (EGF) domain. NRG1 acts in signaling pathways involved in proliferation, apoptosis, migration, differentiation, and adhesion of many normal cell types and in human diseases. The EGF domain of NRG1 mediates signaling by interaction with members of the ErbB family of receptors. Easy access to correctly folded hNRG1α EGF domain can be a valuable tool to investigate its function in different cell types. MATERIALS AND METHODS: The EGF domain of hNRG1α was produced in Escherichia coli in fusion with TrxA and purified after cleavage of TrxA. Conformation and stability analyses were performed by using biophysical methods and the disulfide bonds were mapped by mass spectrometry. The activity of the hNRG1α EGF domain was demonstrated in cell proliferation and migration assays. RESULTS: Approximately 3.3 mg of hNRG1α EGF domain were obtained starting from a 0.5 L of E. coli culture. Correct formation of the three disulfide bonds was demonstrated by mass spectrometry with high accuracy. Heat denaturation assays monitored by circular dichroism and dynamic light scattering revealed that it is a highly stable protein. The recombinant EGF domain of hNRG1α purified in this work is highly active, inducing cell proliferation at concentration as low as 0.05 ng/mL. It induces also cell migration as demonstrated by a gap closure assay. CONCLUSION: The EGF domain of hNRG1α was produced in E. coli with the correct disulfide bonds and presented high stimulation of HeLa cell proliferation and NDFH cell migration.