Synergistic effect of genistein and adiponectin reduces fat deposition in chicken hepatocytes by activating the ERß-mediated SIRT1-AMPK signaling pathway.

Dietary supplementation with bioactive substances that can regulate lipid metabolism is an effective approach for reducing excessive fat deposition in chickens. Genistein (GEN) has the potential to alleviate fat deposition; however, the underlying mechanism of GEN's fat-reduction action in chickens remains unclear. Therefore, the present study aimed to explore the underlying mechanism of GEN on the reduction of fat deposition from a novel perspective: intercellular transmission of adipokine between adipocytes and hepatocytes. The findings showed that GEN enhanced the secretion of adiponectin (APN) in chicken adipocytes, and the enhancement effect of GEN was completely blocked when the cells were pretreated with inhibitors targeting estrogen receptor ß (ERß) or proliferator-activated receptor γ (PPARγ) signals, respectively. Furthermore, the results demonstrated that both co-treatment with GEN and APN or treatment with the medium supernatant (Med SUP) derived from chicken adipocytes treated with GEN significantly decreased the content of triglyceride and increased the protein levels of ERß, Sirtuin 1 (SIRT1) and phosphor-AMP-activated protein kinase (p-AMPK) in chicken hepatocytes compared to the cells treated with GEN or APN alone. Moreover, the increase in the protein levels of SIRT1 and p-AMPK induced by GEN and APN co-treatment or Med SUP treatment were blocked in chicken hepatocytes pretreated with the inhibitor of ERß signals. Importantly, the up-regulatory effect of GEN and APN co-treatment or Med SUP treatment on the protein level of p-AMPK was also blocked in chicken hepatocytes pretreated with a SIRT1 inhibitor; however, the increase in the protein level of SIRT1 induced by GEN and APN co-treatment or Med SUP treatment was not reversed when the hepatocytes were pretreated with an AMPK inhibitor. In conclusion, the present study demonstrated that GEN enhanced APN secretion by activating the ERß-Erk-PPARγ signaling pathway in chicken adipocytes. Subsequently, adipocyte-derived APN synergized with GEN to activate the ERß-mediated SIRT1-AMPK signaling pathway in chicken hepatocytes, ultimately reducing fat deposition. These findings provide substantial evidence from a novel perspective, supporting the potential use of GEN as a dietary supplement to prevent excessive fat deposition in poultry.

Dimethyl fumarate restores Ca2+ dyshomeostasis through activation of the SIRT1 signal to treat nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) is characterized by an excessive lipid accumulation in the liver, with a global prevalence of approximately 25 %. While early-stage steatosis is reversible and can be intervened upon, it has the potential to progress to some serious complications, including cirrhosis and even liver cancer. Dimethyl fumarate (DMF), a derivative of fumaric acid shows promise in intervening in certain diseases. However, the precise effect and underlying mechanism of DMF on hepatic steatosis remain unclear. In this study, we demonstrated that DMF mitigates hepatic steatosis in mice subjected to high-fat/high-cholesterol (HFHC) diets. Meanwhile, our in vivo and in vitro results showed that DMF relieves lipid accumulation, oxidative stress, and endoplasmic reticulum (ER) stress. Mechanically, our findings revealed that the effect of DMF on reducing lipid accumulation is linked to the restoration of Ca2+ homeostasis. Furthermore, we found that activation of the SIRT1 signal by DMF plays an important role in correcting the mishandling of the Ca2+ signal, and knockdown of SIRT1 expression reverses the beneficial role of DMF PA-incubated AML12 cells. In conclusion, our results suggested DMF's amelioration of hepatic steatosis is related to the activation of SIRT1-mediated Ca2+ signaling.

G protein-coupled estrogen receptor 1 ameliorates nonalcoholic steatohepatitis through targeting AMPK-dependent signaling.

Nonalcoholic fatty liver disease (NAFLD), especially nonalcoholic steatohepatitis (NASH), has emerged as a prevalent cause of liver cirrhosis and hepatocellular carcinoma, posing severe public health challenges worldwide. The incidence of NASH is highly correlated with an increased prevalence of obesity, insulin resistance, diabetes, and other metabolic diseases. Currently, no approved drugs specifically targeted for the therapies of NASH partially due to the unclear pathophysiological mechanisms. G protein-coupled estrogen receptor 1 (GPER1) is a membrane estrogen receptor involved in the development of metabolic diseases such as obesity and diabetes. However, the function of GPER1 in NAFLD/NASH progression remains unknown. Here, we show that GPER1 exerts a beneficial role in insulin resistance, hepatic lipid accumulation, oxidative stress, or inflammation in vivo and in vitro. In particular, we observed that the lipid accumulation, inflammatory response, fibrosis, or insulin resistance in mouse NAFLD/NASH models were exacerbated by hepatocyte-specific GPER1 knockout but obviously mitigated by hepatic GPER1 activation in female and male mice. Mechanistically, hepatic GPER1 activates AMP-activated protein kinase signaling by inducing cyclic AMP release, thereby exerting its protective effect. These data suggest that GPER1 may be a promising therapeutic target for NASH.

Genistein alleviates chronic heat stress-induced lipid metabolism disorder and mitochondrial energetic dysfunction by activating the GPR30-AMPK-PGC-1α signaling pathways in the livers of broiler chickens.

The objective of this study was to investigate the preventive effects and mechanisms of genistein (GEN) on production performance and metabolic disorders in broilers under chronic heat stress (HS). A total of 120 male 3-wk-old Ross broilers were randomly assigned to 5 groups: a thermoneutral zone (TN) group maintained at normal temperature (21°C ± 1°C daily), an HS group subjected to cyclic high temperature (32°C ± 1°C for 8 h daily), and 3 groups exposed to HS with varying doses of GEN (50, 100, or 150 mg/kg diet). The experimental period lasted for 3 wk. Here, HS led to a decline in growth performance parameters and hormone secretion disorders (P < 0.05), which were improved by 100 and 150 mg/kg GEN treatment (P < 0.05). Moreover, the HS-induced increases in the liver index (P < 0.01) and abdominal fat rate (P < 0.05) were attenuated by 150 mg/kg GEN (P < 0.05). The HS-induced excessive lipid accumulation in the liver and serum (P < 0.01) was ameliorated after 100 and 150 mg/kg GEN treatment (P < 0.05). Furthermore, the HS-induced decreases in lipolysis-related mRNA levels and increases in lipid synthesis-related mRNA levels in the liver (P < 0.01) were effectively blunted after 100 and 150 mg/kg GEN treatment (P < 0.05). Importantly, the HS-stimulated hepatic mitochondrial energetic dysfunction and decreases in the mRNA or protein levels of peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α), nuclear respiratory factor 1, and mitochondrial transcription factor A in the liver were ameliorated by 150 mg/kg GEN (P < 0.05). Moreover, 50 to 150 mg/kg GEN treatment resulted in a significant increase in the mRNA or protein levels of G protein-coupled estrogen receptor (GPR30), AMP-activated protein kinase (AMPK) α1, phosphorylated AMPKα, and phosphorylated acetyl-CoA carboxylase α. Collectively, GEN alleviated metabolic disorders and hepatic mitochondrial energetic dysfunction under HS, possibly through the activation of GPR30-AMPM-PGC-1α pathways. These data provide a sufficient basis for GEN as an additive to alleviate HS in broilers.

Protective effects of genistein on the production performance and lipid metabolism disorders in laying hens with fatty liver hemorrhagic syndrome by activation of the GPER-AMPK signaling pathways.

The aim of this study was to evaluate the beneficial effects and potential mechanisms of genistein (GEN) on production performance impairments and lipid metabolism disorders in laying hens fed a high-energy and low-protein (HELP) diet. A total of 120 Hy-line Brown laying hens were fed with the standard diet and HELP diet supplemented with 0, 50, 100, and 200 mg/kg GEN for 80 d. The results showed that the declines in laying rate (P < 0.01), average egg weight (P < 0.01), and egg yield (P < 0.01), and the increase of the ratio of feed to egg (P < 0.01) induced by HELP diet were markedly improved by 100 and 200 mg/kg of GEN treatment in laying hens (P < 0.05). Moreover, the hepatic steatosis and increases of lipid contents (P < 0.01) in serum and liver caused by HELP diet were significantly alleviated by treatment with 100 and 200 mg/kg of GEN in laying hens (P < 0.05). The liver index and abdominal fat index of laying hens in the HELP group were higher than subjects in the control group (P < 0.01), which were evidently attenuated by dietary 50 to 200 mg/kg of GEN supplementation (P < 0.05). Dietary 100 and 200 mg/kg of GEN supplementation significantly reduced the upregulations of genes related to fatty acid transport and synthesis (P < 0.01) but enhanced the downregulations of genes associated with fatty acid oxidation (P < 0.01) caused by HELP in the liver of laying hens (P < 0.05). Importantly, 100 and 200 mg/kg of GEN supplementation markedly increased G protein-coupled estrogen receptor (GPER) mRNA and protein expression levels and activated the AMP-activated protein kinase (AMPK) signaling pathway in the liver of laying hens fed a HELP diet (P < 0.05). These data indicated that the protective effects of GEN against the decline of production performance and lipid metabolism disorders caused by HELP diet in laying hens may be related to the activation of the GPER-AMPK signaling pathways. These data not only provide compelling evidence for the protective effect of GEN against fatty liver hemorrhagic syndrome in laying hens but also provide the theoretical basis for GEN as an additive to alleviate metabolic disorders in poultry.

Fatty liver hemorrhagic syndrome (FLHS) is a nutritional and metabolic disease that seriously threatens the health and performance of laying hens, which is characterized by hepatic steatosis and lipid metabolism disorders. As an isoflavone phytoestrogen, genistein (GEN) exerts many beneficial functions, including alleviating lipid metabolism disorders and anti-inflammatory properties. However, further research is needed on the protective effect and potential mechanism of GEN on the FLHS in laying hens. Here, we found that GEN treatment improved liver injury and decline of production performance in laying hens with FLHS. Moreover, GEN treatment alleviated hepatic steatosis and lipid metabolism disorders through reducing the expression levels of mRNA related to fatty acid transport and synthesis and enhancing the mRNA expression levels of factors associated with fatty acid oxidation in FLHS layers, which may be achieved by activation of the G protein-coupled estrogen receptor­adenosine 5'-monophosphate (AMP)-activated protein kinase signaling pathways. These data not only provide compelling evidence for the protective effects and mechanisms of GEN against FLHS in laying hens but also provide the theoretical basis for GEN to alleviate other metabolic disorders in poultry.

Dietary supplementation of dimethyl itaconate protects against chronic heat stress-induced growth performance impairment and lipid metabolism disorder in broiler chickens.

This study aimed to investigate the protective effects of dietary supplementation of dimethyl itaconate (DI) on chronic heat stress (HS)-induced impairment of the growth performance and lipid metabolism in broiler chickens. 21 days old male Ross 308 broiler chickens (a total of 120, about 700 g body weight) were randomly divided into five treatment groups, including control group, HS group, HS + 50 mg/kg DI group, HS + 150 mg/kg DI group, and HS + 200 mg/kg DI group, and each group contains eight cages of twenty-four broilers. The broiler chickens in the control group were raised in the room (21 ± 1 °C) and fed with a finisher diet for 21 days. The broiler chickens of the HS group and the HS + DI groups were raised in the room (32 ± 1 °C for 8 h/day) and fed with a finisher diet containing DI at 0, 50, 150, and 200 mg/kg diet for 21 days. The results showed that HS-induced decreases in the final body weight (P < 0.01), average daily gain (P < 0.01), and average daily feed intake (P < 0.01) were alleviated by dietary supplementation of DI (P < 0.05). In addition, dietary supplementation of DI attenuated the increases in the liver index (P < 0.01) and abdominal fat rate (P < 0.01) caused by HS in broilers (P < 0.05). Treatment with DI ameliorated HS-induced lipid accumulation in the liver and serum of broiler chickens (P < 0.05). The upregulation of mRNA levels of fat synthesis factors (P < 0.01) and downregulation of mRNA levels of lipolysis-related factors (P < 0.01) caused by HS were markedly blunted after treatment with DI in the liver of broilers (P < 0.05). Broilers exposed to HS exhibited lower phosphorylated protein levels of AMP-activated protein kinase α and acetyl-CoA carboxylase α compared to the control group (P < 0.01), which were improved by treatment with DI (P < 0.01). Collectively, these results demonstrated that dietary supplementation of DI protects against chronic HS-induced growth performance impairment and lipid metabolism disorder in broiler chickens. These results not only provide a theoretical basis for DI to alleviate metabolic disorders but also provide a reference value for DI as a feed additive to improve heat stress in poultry caused by high temperature.

Heat stress (HS) caused by high temperatures can lead to metabolic disorders and decreased growth performance in broilers, which has become a global concern in broiler production. Dimethyl itaconate (DI), as a cell-permeable itaconate derivative, has many benefits in alleviating inflammatory response and antioxidant. However, the beneficial effect of DI on broilers exposed to HS are still unclear. Here, we found that DI treatment improved the decline of growth performance and hormone secretion disorder caused by HS in broiler chickens. Moreover, the treatment with DI alleviated the excessive accumulation of lipids caused by HS through reducing mRNA levels related to liposynthesis and enhancing mRNA levels associated with lipolysis in broiler chickens, which may be achieved by activation of the AMP-activated protein kinase (AMPK) signaling pathway. These data not only provide the potential mechanism for DI to alleviate metabolic disorders but also provide a sufficient theoretical basis for DI as an additive to alleviate HS in broiler chickens.

Dehydroepiandrosterone protects against oleic acid-triggered mitochondrial dysfunction to relieve oxidative stress and inflammation via activation of the AMPK-Nrf2 axis by targeting GPR30 in hepatocytes.

Fatty liver hemorrhage syndrome (FLHS) seriously threatens the health and performance of laying hens, and the occurrence and development of FLHS are closely related to oxidative damage and inflammation; thus, diets supplemental with activated substances to relive the oxidative stress and inflammation maybe effectively control the occurrences of FLHS. Dehydroepiandrosterone (DHEA) has beneficial effects in fat-reduction, anti-oxidation and anti-inflammation, and it was widely applied to alleviate multiple metabolic-related diseases; however, there are few reports on whether DHEA can prevent against metabolic-related diseases by modulating oxidative stress and inflammation, especially FLHS in laying hens. Herein, present study aimed to investigate the regulatory actions and potential molecular mechanism of DHEA on inflammation and oxidative stress triggered by oleic acid (OA)-stimulation in primary chicken hepatocytes and chicken hepatocellular carcinoma cell line (LMH). The results showed that DHEA significantly alleviated oxidative stress challenged by OA-stimulation via activation of AMP-activated protein kinase (AMPK)-nuclear factor-erythroid 2-related factor 2 (Nrf2) signaling pathway in hepatocytes, which led to relieving effect of DHEA on inflammatory by inhibiting mitogen-activated protein kinases (MAPKs) and nuclear factor κB (NF-κB) signaling pathways. Mechanistically, we found that the activation of AMPK-Nrf2 signaling pathway by DHEA treatment was mediated by G-protein coupled estrogen receptor (GPR30/GPER) in OA-stimulated hepatocytes. Further investigation found that DHEA activated the GPR30-mediated AMPK-Nrf2 signaling pathways to increase antioxidant capacity and inhibit mitochondrial reactive oxygen species (ROS) overproduction, which thereby inhibiting the activation of ROS-induced MAPK and NF-κB signaling pathways in OA-stimulated hepatocytes. Overall, these data demonstrated that DHEA attenuates the oxidative stress and inflammation triggered by OA-stimulation, and these beneficial effects of DHEA are achieved by activating the GPR30-mediated AMPK-Nrf2 signaling to prevent the impairment of mitochondrial function, and thereby inhibiting the activation of ROS-induced MAPK and NF-κB signaling pathways in hepatocytes. These results revealed the effects and mechanisms of DHEA on oxidative stress and inflammation, and also provide substantial information to support it as a potential nutritional supplement in preventing the occurrences of FLHS in laying hens and other metabolic-related diseases in animals and humans.

Dehydroepiandrosterone alleviates oleic acid-induced lipid metabolism disorders through activation of AMPK-mTOR signal pathway in primary chicken hepatocytes.

The incident of lipid metabolism disorders has obviously increased under the undue pursuit of efficiency, which had seriously threatened to the health development of poultry industry. As an important cholesterol-derived intermediate, though dehydroepiandrosterone (DHEA) has the fat-reduction effect in animals and humans, but the underlying mechanism still poorly understood. Herein, the present study aimed to investigate the regulatory effects and its molecular mechanism of DHEA on disturbance of lipid metabolism induced by oleic acid (OA) in primary chicken hepatocytes. The hepatocytes were treated with 0, 0.1, 1, 10 µM DHEA for 4 h, and then supplemented with 0 or 0.5 mM OA stimulation for another 24 h. Our findings demonstrated that DHEA treatment effectively reduced TG content and alleviated lipid droplet deposition in OA-induced hepatocytes. DHEA inhibited the lipogenesis related factors (ACC, FAS, SREBP-1c, and ACLY) mRNA level and increased the lipolysis key factors (CPT-1 and PPARα) mRNA levels. In addition, DHEA obviously elevated the protein levels of CPT-1A, p-ACC, and ECHS1; whereas decreased the protein levels of FAS and SREBP-1 in hepatocytes stimulated by OA. Furthermore, DHEA promoted the phosphorylation of AMP-activated protein kinase (AMPK) and inhibited the phosphorylation of mammalian target of rapamycin (mTOR). Mechanistically, the hepatocytes were pre-treated with AMPK inhibitor compound C or AMPK activator AICAR before addition of DHEA treatment, and the results certified that DHEA activated cAMP/AMPK pathway and which subsequently led the inhibition of mTOR signal, which finally reduced the fat excessive accumulation in OA-stimulated hepatocytes. Collectively, our study unveiled that DHEA protects against the lipid metabolism disorders triggered by OA stimulation through activation of AMPK-mTOR signaling pathway, which prompts the value of DHEA as a potential nutritional supplement in regulating the lipid metabolism and its related disease in poultry.

Genistein activated SIRT1-AMPK signaling pathway mediated by ERß-FOXO1-Nampt to reduce fat accumulation in chicken hepatocytes.

Excessive fat accumulation in broiler chickens would seriously threaten the poultry industry. It leads to lower feed conversion rate and worse meat quality. Even worse, it harms the consumers' health due to the intake of high-fat chicken products. Dietary supplements with bioactive ingredients have been considered an effective way to solve this problem. Genistein is the primary phytoestrogen in soybean. Its fat-reduction effect has been reported, but the molecular mechanism is unclear. The present study found that genistein reduced lipid droplets accumulation by regulating lipid metabolism-related factors expression in chicken hepatocytes. The research showed that genistein significantly increased phosphor (p)-AMP-activated protein kinase (p-AMPK) and Sirtuin 1 (SIRT1) protein expressions. The effect of genistein on reducing lipid droplets accumulation and upregulating p-AMPK protein level was blocked entirely when pretreated with SIRT1 inhibitor. These results implied that SIRT1 is required to activate AMPK. Furthermore, genistein treatment significantly upregulated the SIRT1 protein level when pretreated with AMPK inhibitor. We demonstrated that the activation of estrogen receptor ß-Forkhead box O1-Nicotinamide phosphoribosyl transferase (ERß-FOXO1-Nampt) signaling pathway upregulated the NAD+ concentration in hepatocytes, and activated SIRT1 ultimately. In summary, we demonstrated that genistein suppressed lipid droplets accumulation in chicken hepatocytes by activating SIRT1-AMPK. The SIRT1-AMPK signaling pathway was mediated by ERß-FOXO1-Nampt. These findings increase our understanding of the mechanisms of genistein on fat reduction, and provide compelling evidence for it as a nutritional supplement to prevent excessive fat deposition and lipid metabolism-related diseases in animals and even humans.

Dehydroepiandrosterone activates the GPER-mediated AMPK signaling pathway to alleviate the oxidative stress and inflammatory response in laying hens fed with high-energy and low-protein diets.

Fatty liver hemorrhagic syndrome (FLHS) seriously threatens the layer industry due to it can cause a sudden decline in egg production and acute death, and dietary supplement with bioactive substance is considered an effective way to prevent the FLHS occurrence. Dehydroepiandrosterone (DHEA) is a popular dietary supplement and it possesses anti-oxidative and anti-inflammatory functions; however, the effect and underlying mechanism about DHEA in protecting against the occurrence and development of FLHS remain elucidated. The current results showed that DHEA relieved HELP-induced decrease of egg productivity and liver injury in laying hens. Meanwhile, DHEA markedly enhanced the antioxidant capacity and then alleviated oxidative stress via activation of nuclear factor (erythroid-derived 2)-like 2 (NRF-2) signal in laying hens fed with HELP diets. In addition, DHEA significantly alleviated HELP-stimulated systemic inflammatory response by suppressing the overproduction of hepatic pro-inflammatory factors in laying hens, and further found this beneficial effect was achieved by blocking the activation of NF-κB pathway. Furthermore, we found that DHEA promoted the AMP-activated protein kinase α (AMPKα) activation and increased the G-protein-coupled estrogen receptor (GPER) expression level in laying hens fed with HELP diets. In summary, our data demonstrated that DHEA attenuates oxidative stress and inflammation through the activation of GPER-AMPK signal axis in laying hens fed with HELP diets. These results might facilitate an understanding of the benefits and mechanism of DHEA on the development of FLHS, and provide sufficient data to support it as a dietary supplement to control the FLHS-related metabolic diseases in chickens.

Activated AMP-activated protein kinase prevents hepatic steatosis, oxidative stress and inflammation in primary chicken hepatocytes.

Fatty liver hemorrhagic syndrome (FLHS) in laying hens, a nutritional metabolic disorder disease, can lead to the decline of laying rate, shortening of laying peak period and increase of mortality, which seriously constrain the sustainable development of layer industry. Until now, there is no effective strategies can prevent and control the occurrence of fatty liver hemorrhagic syndrome in laying hens. The AMP-activated protein kinase (AMPK), a major sensor of cellular energy status, acts a crucial role in regulating lipid metabolism, oxidative stress and inflammatory responses in body. However, the potential molecular mechanisms about AMP-activated protein kinase signal in controlling the occurrence of fatty liver hemorrhagic syndrome are remain unclear. In present study, we found that the phosphorylated AMP-activated protein kinase (Thr172) protein level was markedly reduced in palmitic acid plus oleic acid (PO)-induced primary chicken hepatocytes. Moreover, blocked AMP-activated protein kinase signal by AMP-activated protein kinase inhibitor compound C obviously exacerbated lipid metabolism disorders, oxidative stress and inflammatory response triggered by palmitic acid plus oleic acid in primary chicken hepatocytes. Nevertheless, the lipid metabolism disorders, oxidative stress and inflammatory response challenged by palmitic acid plus oleic acid were obviously alleviated through activation of AMP-activated protein kinase signal with AMP-activated protein kinase activator AICAR in hepatocytes. In addition, we found that the beneficial effects of AMP-activated protein kinase signal in relieving lipid metabolism disorders, oxidative stress and inflammatory response are achieved by activating the nuclear factor erythroid 2-related factor 2 (NRF-2)/kelch-like ECH-associated protein 1 (KEAP1) pathway and inhibiting the NF-κB pathway in PO-stimulated primary chicken hepatocytes. Collectively, our data demonstrated that AMP-activated protein kinase acts as a potential target for the prevention of fatty liver hemorrhagic syndrome occurrence in laying hens.

Genistein accelerates glucose catabolism via activation the GPER-mediated cAMP/PKA-AMPK signaling pathway in broiler chickens.

Genistein, the most abundance of phytoestrogens in soybeans, has beneficial effects in regulating metabolism-related disease; however, there is few available literatures about whether genistein regulates glucose metabolism that in turn affects the lipid accumulation in animals or humans. The current study showed that genistein promoted glucose uptake by enhancing glucose transporter-2 (GLUT2) protein level; and it also increased the activity of phosphofructokinase-1 (PFK) and pyruvate dehydrogenase (PDH), and the mRNA level of succinate dehydrogenase (SDH) both in broiler chickens or hepatocytes. Moreover, genistein obviously increased the p-LKB1 and p-AMPKα protein levels both in vivo and in vitro. Furthermore, the enhancement of genistein on glucose uptake and catabolism were reversed in hepatocytes pre-treated with AMPK inhibitor Compound C, and the increasing of genistein on the p-LKB1 and p-AMPKα protein levels were also reversed in hepatocytes pre-treated with PKA inhibitor H89. Importantly, the results showed that genistein simultaneously increased the estrogen receptor ß (ERß) and G protein-coupled estrogen receptor (GPER) protein levels, but the elevation effect of genistein on cAMP content was completely reversed in hepatocytes pre-treated with GPER antagonist G15, rather than ERß inhibitor PHTPP. Meanwhile, the increasing of p-LKB1 and p-AMPKα protein levels induced by genistein were also reversed in hepatocytes pre-treated with G15. Collectively, our data demonstrated that genistein improves glucose metabolism via activating the GPER-mediated cAMP/PKA-AMPK signaling pathway. These findings provide theoretical basis for genistein as a promising nutritional supplemental to alleviate metabolism disorders and related diseases in animals or even humans.

Dehydroepiandrosterone exacerbates nigericin-induced abnormal autophagy and pyroptosis via GPER activation in LPS-primed macrophages.

As a widely acknowledged FDA-approved dietary supplement or over-the-counter medicines, dehydroepiandrosterone (DHEA) exerts anti-inflammatory and immunomodulatory function. Pyroptosis is an important form of programmed cell death (PCD), and which acts a key role in the body's anti-infection and inflammatory responses. But the effects and mechanisms of DHEA on pyroptosis remain unclear. Here, we found that DHEA inhibited the NLRP3 inflammasome components expression by blocking inflammatory signals in lipopolysaccharide (LPS)-primed macrophages, and prevented the bacterial toxin nigericin (Nig)-induced NLRP3 inflammasome assembly. However, DHEA exacerbated NLRP3-independent cell death in Nig-treated inflammatory macrophages. During this process, DHEA induced the abnormal autophagy, which reflected as the blocking of autophagic flux and the accumulation of autophagy receptor p62 (SQSTM1) protein. In addition, DHEA caused a burst of reactive oxygen species (ROS) and activated extracellular signal-regulated kinase (ERK) phosphorylation in LPS plus Nig-stimulated macrophages but not in LPS-treated macrophages. Mechanistically, the present study certified that the activation of G protein-coupled estrogen receptor (GPER) signal mediated the cell death induced by DHEA in Nig-stimulated inflammatory macrophages, as GPER specific inhibitor G15 alleviated the abnormal autophagy and ultimately prevented the gasdermin D (GSDMD)-mediated pyroptosis induced by DHEA. Collectively, DHEA can exacerbate Nig-induced abnormal autophagy and pyroptosis via activation of GPER in LPS-primed macrophages, which prompts us the potential application value of DHEA in anti-infection or anti-tumor immunity.

4-Octyl Itaconate Prevents Free Fatty Acid-Induced Lipid Metabolism Disorder through Activating Nrf2-AMPK Signaling Pathway in Hepatocytes.

Nonalcoholic fatty liver disease (NAFLD), characterized with oxidative stress and hepatic steatosis, is a serious threat to human health. As a specific activator of nuclear factor E2-related factor 2 (Nrf2), the 4-octyl itaconate (4-OI) has the beneficial effects in antioxidant and anti-inflammation; however, whether 4-OI can alleviate hepatic steatosis and its mechanism is still unknown. The present study was aimed at investigating the protective effects of 4-OI on free fat acid- (FFA-) induced lipid metabolism disorder and its potential molecular mechanism in hepatocytes. The results showed that 4-OI treatment markedly alleviated FFA-induced oxidative stress and excessive lipid accumulation in hepatocytes. Mechanistically, 4-OI significantly suppressed the overproduction of reactive oxygen species (ROS) through activation of Nrf2; the downregulation of ROS level induced a downregulation of AMP-dependent protein kinase (AMPK) phosphorylation level which finally ameliorated excessive lipid accumulation in FFA-stimulated hepatocytes. In general, our data demonstrated that 4-OI relieves the oxidative stress and lipid metabolism disorder in FFA-stimulated hepatocytes; and these beneficial effects were achieved by activating the Nrf2-AMPK signaling pathway. These data not only expand the new biological function of 4-OI but also provide a theoretical basis for 4-OI to protect against lipid metabolism disorders and related diseases, such as NAFLD.

The sex steroid precursor dehydroepiandrosterone prevents nonalcoholic steatohepatitis by activating the AMPK pathway mediated by GPR30.

The prevalence of nonalcoholic steatohepatitis (NASH) caused by estrogen deficiency increased sharply in recent decades and has become a major threat to liver health in postmenopausal women. There is no effective strategy to control the incidence and development of NASH. Dehydroepiandrosterone (DHEA) is the most abundant circulating steroid with immune and metabolic regulatory properties, and its level markedly declines with increasing age in humans. Importantly, DHEA can convert into active sex hormones depending on the local needs of target tissues with little diffusion, which serves to avoid systemic side-effects from other tissues' exposure to estrogen. Here, we found that DHEA prevented the incidence and development of NASH, which is characterized by the reduction of hepatic steatosis, fibrosis, and inflammation in female mice fed with high-fat/high-cholesterol diets and effectively attenuated lipid accumulation, inflammatory response, and oxidative stress in palmitic acid-challenged hepatocytes. Mechanistically, in vitro and in vivo studies showed that the anti-NASH function of DHEA depended on its biotransformation into estrogen rather than androgen, and which up-regulates the expression of G protein-coupled estrogen receptor (GPR30), a non-classical estrogen receptor. The activation of GPR30-mediated AMP-activated protein kinase signaling is a necessary prerequisite for the alleviative effects of DHEA on NASH. Collectively, our data show the mechanisms of DHEA treatment and its effects on NASH that were previously overlooked; the data also show that GPR30 can be used as a target for treating lipid metabolism disorders and related diseases, such as NASH. Furthermore, these findings have the potential to help researchers develop new strategies for preventing NASH in postmenopausal women.

Dehydroepiandrosterone alleviates intestinal inflammatory damage via GPR30-mediated Nrf2 activation and NLRP3 inflammasome inhibition in colitis mice.

Dehydroepiandrosterone (DHEA) is a popular dietary supplement that has anti-inflammatory, anti-oxidant and immune-regulating role; meanwhile, it also can effective in the protection of inflammation diseases such as inflammatory bowel disease (IBD), but the underlying mechanisms remain elusive. Here, we demonstrated that DHEA inhibits excessive inflammation response and enhances gut barrier function via activating the G protein-coupled receptor 30 (GPR30). GPR30-induced the ERK phosphorylation and p62 accumulation led to the activation of nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway, which subsequently inhibited the reactive oxygen species (ROS) overproduction and finally alleviated the intestinal barrier dysfunction. Furthermore, DHEA blocked the p38-induced NLRP3 inflammasome activation in both LPS-stimulated colon epithelial cells and macrophages. In addition, in vivo results showed that DHEA and GPR30 agonist G1 attenuated inflammatory responses and gut barrier dysfunction in colitis mice, while the GPR30 specific inhibitor G15 abrogated these beneficial effects of DHEA. Cumulatively, our study unveiled that DHEA is an effective anti-inflammatory agent and suggested that GPR30 could as a potential target for the treatment of IBD.

Comprehensive genomic analysis and characterization of a new ST 174 type Klebsiella variicola strain isolated from chicken embryos.

Klebsiella variicola is a widespread opportunistic pathogen that causes infections in humans and animals. Herein a novel Klebsiella strain, AHKv-S01, was isolated and identified from dead chicken embryos in Anhui, China. Its genome contained a circular chromosome of 5,505,304 bp, with 5244 protein-coding genes, and an integrative conjugative element region containing 79 ORF sequences. AHKv-S01 was given a new sequence type number-174. Phylogenetic analyses showed that rpoB partial nucleotide sequences were highly reliable for identifying Klebsiella spp. Most of the 340 unique genes of AHKv-S01 were involved in cell envelop biogenesis, transcription, transport, and metabolic processes. Moreover, AHKv-S01 was sensitive to several antibiotics, but it showed strong resistance to penicillins, macrolides, and lincosamide. The genome contained three drug efflux pump superfamilies, ß-lactamase genes, and fosfomycin resistance-related genes. Most drug resistance genes showed amino acid mutations. Multiple virulence and pathogenic factors were also identified, and they were mainly related to adhesion, secretion, iron acquisition, and immune evasion. Chicken embryo lethality assay results revealed that the 7-day chicken embryo lethality rate was 80%, 40%, and 50% for AHKv-S01, K. pneumoniae ATCC10031, and K. pneumoniae CICC24714, respectively. The median lethal dose of AHKv-S01 was 39.9 CFU/embryo. Even low infection levels of AHKv-S01 caused a significant reduction in chicken embryo hatchability. Severe pathological changes to the liver, heart, and brain tissues of embryos infected with AHKv-S01 were observed, and these changes appeared earlier in the heart and brain than in the liver. To conclude, our results provide a foundation for further studies aiming to assess the potential risk of K. variicola to poultry populations and production yields.

Dehydroepiandrosterone attenuates LPS-induced inflammatory responses via activation of Nrf2 in RAW264.7 macrophages.

Dehydroepiandrosterone (DHEA) is the major steroid hormone in humans and animals, which can regulate the body's inflammatory responses. However, the detail mechanism of this beneficial function is still poorly understood. The present study aimed to explore the anti-inflammation effect of DHEA and its underlying molecular mechanism in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. The findings showed that DHEA significantly inhibited the inflammation-related mediators production and pro-inflammatory cytokines expression level. Further research found that DHEA obviously blocked the LPS-stimulated PI3K/AKT, MAPK and NF-κB activation in RAW 264.7 cells. Meanwhile, DHEA enhanced the autophagy-dependent Keap1 protein degradation, subsequently activated the Nrf2 pathway to alleviate the redox imbalance and inflammatory responses. In conclusion, our data demonstrated that DHEA suppresses inflammatory responses through the activation of Nrf2 and inhibition of NF-κB in LPS-stimulated macrophages.

Genistein activated adenosine 5'-monophosphate-activated protein kinase-sirtuin1/peroxisome proliferator-activated receptor γ coactivator-1α pathway potentially through adiponectin and estrogen receptor ß signaling to suppress fat deposition in broiler chickens.

Genistein can be used as a dietary additive to control fat deposition in animals, while its mechanism is poorly understood. In this study, a total of 144 male broilers were randomly divided into 4 groups. Birds were fed standard diets supplemented with 0, 50, 100 or 150 mg of genistein/kg from 21 to 42 d of age. Results showed that genistein treatment decreased the relative weight of abdominal fat and triglyceride contents in broiler chickens. Genistein downregulated hepatic lipid droplets accumulation and upregulated the activity of lipoprotein lipase and hepatic lipase and the concentration of adiponectin. Furthermore, the liver X receptor α, sterol regulatory element-binding protein 1c (SREBP-1c), acetyl-CoA carboxylase (ACC), and fatty acid synthase (FAS) mRNA expressions were decreased, whereas adiponectin receptor 2, peroxisome proliferator-activated receptor α, adipose triglyceride lipase, and carnitine palmitoyl transferase-I (CPT-I) mRNA abundances were increased in the liver of broilers treated with genistein. In addition, genistein increased the NAD+ concentration and NAD+/NADH ratio in the liver. Genistein increased estrogen receptor ß (ERß), forkhead box O1, nicotinamide phosphoribosyl transferase, sirtuin1 (SIRT1), phospho (p)-adenosine 5'-monophosphate-activated protein kinase (AMPK), peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), p-ACC, and CPT-I protein levels, whereas the SREBP-1c and FAS levels were decreased. These data indicated that genistein might reduce fat accumulation in broiler chickens via activating the AMPK-SIRT1/PGC-1α signaling pathway. The activation of this signaling pathway might be achieved by its direct effect on improving the adiponectin secretion or its indirect effect on upregulation of ERß expression level through paracrine acting of adiponectin.