PPARγ mRNA Expression Research Articles

Danggui Shaoyao powder improves hepatic lipid metabolism in atherosclerosis mice via PPARγ-LXRα-ABCA1 pathway regulation

ObjectiveTo observe the effects of Danggui Shaoyao powder (DSP) on hepatic lipid metabolism and further explore its mechanism of action by peroxisome proliferator-activated receptor (PPARγ)-liver X receptor (LXRα)-adenosine triphosphate (ATP)-binding cassette transporter A1 (ABCA1) pathway regulation. MethodsEight C57BL/6J male mice were selected as the control group, and 24 ApoE−/− male mice were randomly divided into the atherosclerosis model (AS) group, atorvastatin calcium (AC) group, and DSP group (n = 8 each group). To establish an AS model, ApoE−/− mice were fed a high-fat diet for 16 weeks. Pathologic changes in the aortic vasculature and liver were identified using Oil Red O staining. Triglyceride (TG), cholesterol (TC), and low-density lipoprotein cholesterol (LDL-C) levels were determined in the livers using a single-reagent GPO-PAP method. Fluorescence quantitative polymerase chain reaction and western blot were used to observe and evaluate the mRNA and protein expression of the PPARγ-LXRα-ABCA1 intermediates in the liver. ResultsAfter 16 weeks of a high-fat diet, ApoE−/− mice showed more Oil Red O staining in the aorta and liver compared to the CONT group. Compared to the AS group, the DSP and AC treatment reduced aortic plaque and hepatic lipid deposition to varying degrees. Furthermore, DSP significantly reduced the hepatic lipid area in ApoE−/− mice (P < .001) and decreased the levels of TG, TC, and LDL-C in liver (P < .001, P = .027, P < .001, respectively). DSP also significantly increased the levels of PPARγ, LXRα, ABCA1, and ABCG1 mRNA expression, as well as the PPARγ, LXRα, ABCA1, and ABCG1 protein expression in liver. ConclusionDSP improved hepatic lipid metabolism via PPARγ-LXRα-ABCA1 pathway modulation for AS treatment.

Abstract 1426: Lipid-mediated epigenetic activation of PPAR-gamma signaling promotes immune-checkpoint resistance in hepatocellular carcinoma

Abstract Background: Although use of immune-checkpoint inhibitors (ICI) has yielded impressive results in cancer patients, resistance to ICI therapies has increasingly been observed, especially in hepatocellular carcinoma (HCC). Materials and Methods: We generated a novel anti-programmed death-ligand 1 (PD-L1) treatment resistant mouse hepatoma cell Hepa1-6 (PD-L1R) by 7-generation of in vivo selection. To dissect tumor cell-intrinsic resistant mechanism, we performed single-cell RNA-sequencing (scRNA-seq) from anti-PD-L1-treated tumors generated from parental or PD-L1R cells. Expression level of target genes was detected by QPCR, western blot and immunohistochemical analyses. Lipid content and fatty acid uptake was examined by Oil Red O and BIODIPY staining. Peroxisome proliferator-activated receptor gamma (PPARγ) promoter methylation level was analyzed by bisulfite pyrosequencing. To investigate the tumor cell-extrinsic factors, we profiled the myeloid and lymphoid lineages by flow cytometry. Results: ScRNA-seq revealed significant gene enrichments in lipid metabolism and PPARγ signaling pathway in PD-L1R tumors, which could be verified in both mRNA and protein levels. Moreover, we observed evident lipid accumulation in the PD-L1R tumors when compared to the parental tumors by Oil Red O staining. Consistently, PD-L1R tumor cells exhibited higher lipid content and enhanced fatty acid uptake in vitro, as demonstrated by BODIPY493/503 and BODIPY-C16 analyses. Of note, palmitic acid could dose-dependently increase Pparg mRNA expression in parental cells to a similar level with the PD-L1R cells, supporting an important role of lipids in inducing PPARγ expression. Pyrosequencing analysis indicated that PD-L1R cells exhibited lower PPARγ methylation level compared with parental cells. Notably, 5-Azacytidine treatment in parental cells significantly reduced the promoter DNA methylation and increased Pparg mRNA expression to similar extent with the PD-L1R tumor cells, thus indicating DNA hypomethylation-driven PPARγ up-regulation. Flow cytometry analysis showed that ICI resistance was associated with lower cytotoxic CD8+ T cells but higher intermediate Th17 cells, myeloid-derived suppressor cells and T regulatory cells in the tumor microenvironment (TME). Conclusions: Our study suggested that lipid exposure may epigenetically up-regulate PPARγ transcription in the development of anti-PD-L1 resistance. Detailed investigation on the crosstalk between aberrant lipid metabolism and immunosuppressive TME is ongoing. Acknowledgment: This study is supported by RGC General Research Fund (14120621 and 14119023), Li Ka Shing Foundation and CUHK Strategic Seed Funding for Collaborative Research Scheme. Citation Format: Zhewen Xiong, Stephen Lam Chan, Jingying Zhou, Xuezhen Zeng, Weiqin Yang, Haoran Wu, Jianquan Cao, Jie-Ting Low, Michael Wing-Yan Chan, Kevin Yuk-Lap Yip, Joseph Jao-Yiu Sung, Alfred Sze-Lok Cheng. Lipid-mediated epigenetic activation of PPAR-gamma signaling promotes immune-checkpoint resistance in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1426.

The regulatory effects of electroacupuncture on type 2 innate lymphoid cell (ILC2) function and browning of white adipose tissue in obese rats: 电针通过调控ILC2功能和白色脂肪棕色化治疗肥胖大鼠的作用机制

ObjectiveTo observe the effects of electroacupuncture (EA) on type 2 innate lymphoid cell (ILC2) function in white adipose tissue (WAT) of high-fat diet (HFD)-induced obese rats to reveal the mechanism underlying weight loss. MethodsMale Winstar rats (n=15) were randomly divided into control, HFD, and HFD+EA groups, with five rats in each group. An obesity model was established by feeding rats with a HFD. EA (2 Hz, 1 mA) was applied to “Zhongwan (CV12)” bilateral “Tianshu (ST25)” “Zusanli (ST36)” and “Sanyinjiao (SP6)” for 15 min, 7 days a week, for a total of 5 weeks. During EA treatment, the HFD group was fed a HFD, whereas rats in the control group were maintained on a normal diet. There was no intervention in either the control or the HFD groups. Body weight was measured weekly. Full-field adipocyte morphology, adipocyte volume, and the number of abdominal WAT were observed under a microscope after sectioning and hematoxylin and eosin staining. The levels of IL-5 (interleukin-5), IL-13 (interleukin-13), IL-33 (interleukin-33), and ST2 (growth stimulating gene 2 protein) in the WAT were assayed using enzyme-linked immunosorbent assay and reverse transcription-polymerase chain reaction (RT-PCR). The expression levels of browning markers including PGC1α（peroxisome proliferator-activated receptor gamma coactivator 1-alpha）, PPARγ (peroxisome proliferator-activated receptor gamma), PRDM16 (histone-lysine N-methyltransferase), and UCP1（uncoupling protein 1 in WAT were detected by western blot analysis and RT-PCR. ResultsAfter a 7-week induction period, rats in both the HFD and HFD+EA groups were 20% heavier than their initial weights before modeling (P<0.01, P<0.001). Following a 5-week EA intervention, the body mass and Lee index of obese rats were significantly reduced (P<0.05). The HE results showed that adipocytes in the HFD group displayed a significant increase in diameter and a decrease in number compared to the control group. However, in the HFD+EA group, white adipocytes exhibited reduced diameter and appeared more regular in shape. In the control group, the proportion of ILC2 was (86.53±2.55)%, whereas in the HFD group, it decreased to (73.62±2.52)%, indicating a significant reduction in ILC2 in the WAT of obese rats. Following EA treatment, the proportion of ILC2 was elevated to (80.54±4.25)%. This analysis revealed a significant decrease in the expression levels of IL-5, IL-13, IL-33, and ST2 in the WAT of obese rats in the HFD group; however, these levels increased after EA intervention. The mRNA levels of PPARγ, PGC1α, PRDM16, and protein levels of UCP1 were decreased in the HFD group (P<0.01, P<0.05, and P<0.001), and the trends were significantly reversed with EA manipulation (P<0.05). RT-PCR and western blotting results revealed that EA remarkably upregulated the mRNA expression of PGC1α, PPARγ, and PRDM16, as well as the protein expression of UCP1 induced by HFD. In the HFD group, PRDM16 expression showed a strong correlation with UCP1 expression, and ILCs were strongly correlated with PGC1α expression. However, these strong correlations did not occur in HFD+EA group. Following EA treatment, the expression of PPARγ displayed a strong correlation with ILC2s, indicating that the altered ILC2 proportion was closely related to browning in WAT. ConclusionEA treatment effectively alleviated obesity by restoring ILC2 function and promoting browning of white adipocytes. ILC2s may play an important role in the effects of EA treatment, but this requires further study.

Vimentin protein is a factor for decreasing breast cancer cell proliferation co-culture with human bone marrow-derived mesenchymal stem cells pre-treated with thiazolidinedione solutions.

Our previous study investigated the levels of soluble growth factors in the conditioned media of bone marrow-derived mesenchymal stem cells (BMSCs) pre-treated with thiazolidinedione solutions. The present study aimed to investigate the complex intracellular proteins extracted from BMSCs pre-treated with pioglitazone and/or rosiglitazone using proteomics. The proliferative effect of the identified protein on MCF-7 cells that interacted non-adhesively with BMSCs pre-treated with pioglitazone and/or rosiglitazone was evaluated using cell culture inserts and conditioned media. The mRNA expression of proliferation and lipid accumulation markers was also evaluated in the interacted MCF-7 cells by reverse transcription-quantitative PCR. Finally, the correlation between the identified protein and fibroblast growth factor 4 (FGF-4) proteinin the conditioned media of the pre-treated BMSCs was evaluated by ELISA. The present study identified vimentin as the specific protein among the complex intracellular proteins that likely plays a role in MCF-7 cell proliferation when the breast cancer cells interacted non-adhesively with BMSCs pre-treated with a combination of pioglitazone and rosiglitazone. The inhibition of this protein promoted the proliferation of MCF-7 cells when the breast cancer cells interacted with pre-treated BMSCs. Gene expression analysis indicated that pre-treatment of BMSCs with a combination ofpioglitazone and rosiglitazone decreased the mRNA expression of Ki67 and proliferating cell nuclear antigen in MCF-7 cells. The pre-treatment did not induce mRNA expression of PPARγ, which is a sign of lipid accumulation. The level of vimentin protein was also associated with the FGF-4 protein expression level in the conditioned media of the pre-treated BMSCs. Bioinformatics analysis revealed that vimentin regulated the expression of FGF-4 through its interaction with SRY-box 2 and POU class 5 homeobox 1. The present study identified a novel intracellular protein that may represent the promising target in pre-treated BMSCs to decrease the proliferation of breast cancer MCF-7 cells for human health and wellness.

Pioglitazone, a peroxisome proliferator‑activated receptor γ agonist, induces cell death and inhibits the proliferation of hypoxic HepG2 cells by promoting excessive production of reactive oxygen species.

Hypoxia is a hallmark of solid tumors. Hypoxic cancer cells adjust their metabolic characteristics to regulate the production of cellular reactive oxygen species (ROS) and facilitate ROS-mediated metastasis. Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that regulates the transcription of fatty acid metabolism-related genes that have a key role in the survival and proliferation function of hypoxic cancer cells. In the present study, mRNA expression in HepG2 cells under chemically induced hypoxia was assessed. The protein expression levels of hypoxia-inducible factor 1α (HIF-1α) were measured using western blotting. Following treatment with the PPARγ agonist pioglitazone, cell viability was assessed using a Cell Counting Kit-8 assay, whilst cell proliferation and death were determined using 5-ethynyl-2'-deoxyuridine incorporation staining, and calcein-acetoxymethyl ester and propidium iodide staining, respectively. Cellular ROS production was assessed using dihydroethidium staining. Cobalt chloride was used to induce hypoxia in HepG2 cells, which was evaluated using HIF-1α expression. The results revealed that the mRNA expression of PPARγ, CD36, acetyl-co-enzyme A dehydrogenase (ACAD) medium chain (ACADM) and ACAD short-chain (ACADS) was downregulated in hypoxic HepG2 cells. The PPARγ agonist pioglitazone decreased the cell viability of hypoxic HepG2 cells by inhibiting cell proliferation and inducing cell death. Following treatment with the PPARγ agonist pioglitazone, hypoxic HepG2 cells produced excessive ROS. ROS-mediated cell death induced by the PPARγ agonist pioglitazone was rescued with the antioxidant N-acetyl-L-cysteine. The downregulated mRNA expression of PPARγ, CD36, ACADM and ACADS was not reverted by a PPARγ agonist in hypoxic HepG2 cells. By contrast, the PPARγ agonist suppressed the mRNA expression of BCL2, which was upregulated in hypoxic HepG2 cells. In summary, the PPARγ agonist stimulated excessive ROS production to inhibit cell proliferation and increase the death of hypoxic HepG2 cells by decreasing BCL2 mRNA expression, suggesting a negative association between PPARγ and BCL2 in the regulation of ROS production in hypoxic HepG2 cells.

Effect of YC-1102 on the Improvement of Obesity in High-Fat Diet-Induced Obese Mice.

Obesity is one of the major risk factors for metabolic diseases worldwide. This study examined the effects of YC-1102, an extract derived from the roots of Rosa multiflora, on 3T3-L1 preadipocytes and high-fat diet (HFD)-induced obese mice. In vivo experiments involved the oral administration of YC-1102 (100, 150, and 200 mg/kg body weight) daily to mice for eight weeks. YC-1102 was found to downregulate the expressions of PPARγ and C/EBPα during adipogenesis, inhibiting adipocyte differentiation and upregulating the expression of PGC-1α for energy metabolism to enhance mitochondrial biogenesis and fatty acid oxidation. It has been shown that daily administration of YC-1102 to mice receiving a HFD prevented an increase in body weight and the accumulation of body fat. YC-1102 administration also reduced TG, TC, and LDL cholesterol levels, as well as glucose and leptin levels, and increased adiponectin levels, thus effectively inhibiting the metabolism of lipids. YC-1102-treated mice showed significant reductions in the mRNA expression of PPARγ and C/EBPα. The levels of PGC-1α involved in energy metabolism increased significantly in the YC-1102-treated mice when compared to the HFD-treated mice. According to the findings of this study, YC-1102 has a dual mechanism that reduces transcription factors that promote the differentiation of adipocytes and increases transcription factors that promote energy consumption.

Molecular Approach to Identify Anti-inflammatory Potential of Stevioside in HFD-induced Type 2 Diabetic Rats: Evidence From in Vivo Study

Stevioside is a natural sweetener derived from the leaves of the Stevia rebaudiana plant. It has gained popularity as a sugar substitute due to its intense sweetness without adding calories or affecting blood sugar levels, making it a suitable option for people with diabetes or those looking to reduce their sugar intake. Studies have shown that stevioside has glucose lowering effects. Previous studies have shown that it has significant role in skeletal muscle but its role on expression of inflammatory signaling molecules in adipose tissue against high diet and sucrose-induced type-2 diabetes in experimental rats is yet to be done. The current research was undertaken to investigate if stevioside could also exert its antidiabetic effects by circumventing adipocyte induced inflammation, a key driving factor for insulin resistance in obese individuals. Effective dose of stevioside (20 mg/kg b.wt) was administered orally for 45 days to high fat diet and sucrose induced type-2 diabetic rats. Interestingly, stevioside treatment restores the elevated serum levels of proinflammatory cytokines including tumor necrosis factor-α (TNF-α) and sterol regulatory element binding protein-1c (SREBP-1c) and enhances Peroxisome Proliferator–activated receptor-γ (PPAR-γ) in adipocytes of diabetic rats. The gene expression of IR, GLUT4 and PPAR-γ mRNA were also significantly activated in stevioside treated groups but reduced IL-1 beta, IlL-6, IKKB, TNF-alpha and NFkB mRNA expression in diabetic adipose tissue. More importantly, stevioside acts very effectively as metformin to circumvent inflammation and insulin resistance in diabetic rats. Our results clearly show that stevioside inhibits obesity induced insulin resistance by ameliorating the inflammatory events and upregulating insulin signalling molecules. Keywords: Stevioside, HFD-T2DM, pro inflammation, insulin signalling; adipose tissue, obesity; signaling pathways; Therapautics.

Metabolome analysis reveals that cyclic adenosine diphosphate ribose contributes to the regulation of differentiation in mice adipocyte.

Adipocytes play a key role in energy storage and homeostasis. Although the role of transcription factors in adipocyte differentiation is known, the effect of endogenous metabolites of low molecular weight remains unclear. Here, we analyzed time-dependent changes in the levels of these metabolites throughout adipocyte differentiation, using metabolome analysis, and demonstrated that there is a positive correlation between cyclic adenosine diphosphate ribose (cADPR) and Pparγ mRNA expression used as a marker of differentiation. We also found that the treatment of C3H10T1/2 adipocytes with cADPR increased the mRNA expression of those marker genes and the accumulation of triglycerides. Furthermore, inhibition of ryanodine receptors (RyR), which are activated by cADPR, caused a significant reduction in mRNA expression levels of the marker genes and triglyceride accumulation in adipocytes. Our findings show that cADPR accelerates adipocytic differentiation via RyR pathway.

Effects of nanoselenium supplementation on lactation performance, nutrient digestion and mammary gland development in dairy cows

The objective of this experiment was to evaluate the influence of nanoselenium (NANO-Se) addition on milk production, milk fatty acid synthesis, the development and metabolism regulation of mammary gland in dairy cows. Forty-eight Holstein dairy cows averaging 720 ± 16.8 kg of body weight, 66.9 ± 3.84 d in milk (dry matter intake [DIM]) and 35.2 ± 1.66 kg/d of milk production were divided into four treatments blocked by DIM and milk yields. Treatments were control group, low-Se (LSe), medium-Se (MSe) and high-Se (HSe) with 0, 0.1, 0.2 and 0.3 mg Se, respectively, from NANO-Se per kg dietary dry matter (DM). Production of energy- and fat-corrected milk (FCM) and milk fat quadratically increased (p < 0.05), while milk lactose yields linearly increased (p < 0.05) with increasing NANO-Se addition. The proportion of saturated fatty acids (SFAs) linearly decreased (p < 0.05), while proportions of monounsaturated fatty acids (MUFAs) linearly increased and polyunsaturated fatty acids (PUFAs) quadratically increased. The digestibility of dietary DM, organic matter (OM), crude protein (CP), neutral detergent fiber (NDF) and acid detergent fiber (ADF) quadratically increased (p < 0.05). Ruminal pH quadratically decreased (p < 0.01), while total VFA linearly increased (p < 0.05) with increasing NANO-Se addition. The acetic to propionic ratio decreased (p < 0.05) linearly due to the unaltered acetic molar percentage and a quadratical increase in propionic molar percentage. The activity of CMCase, xylanase, cellobiase and pectinase increased linearly (p < 0.05) following NANO-Se addition. The activity of α-amylase increased linearly (p < 0.01) with an increase in NANO-Se dosage. Blood glucose, total protein, estradiol, prolactin, IGF-1 and Se linearly increased (p < 0.05), while urea nitrogen concentration quadratically decreased (p = 0.04). Moreover, the addition of Se at 0.3 mg/kg from NANO-Se promoted (p < 0.05) mRNA and protein expression of PPARγ, SREBP1, ACACA, FASN, SCD, CCNA2, CCND1, PCNA, Bcl-2 and the ratios of p-ACACA/ACACA and BCL2/BAX4, but decreased (p < 0.05) mRNA and protein expressions of Bax, Caspase-3 and Caspase-9. The results suggest that milk production and milk fat synthesis increased by NANO-Se addition by stimulating rumen fermentation, nutrients digestion, gene and protein expressions concerned with milk fat synthesis and mammary gland development.

Angong Niuhuang Wan inhibit ferroptosis on ischemic and hemorrhagic stroke by activating PPARγ/AKT/GPX4 pathway

Ethnopharmacological relevanceAngong Niuhuang Wan (AGNHW) is a prescription from traditional Chinese medicine (TCM) that has been used for centuries to treat ischemic stroke (IS) and hemorrhagic stroke (HS). According to a recent study, targeting ferroptosis might be effective in the management of IS and HS. However, the ferroptosis-related effects and mechanisms of AGNHW have not yet been reported. Aim of the studyThis research examines the anti-ferroptosis mechanisms of AGNHW in the treatment of IS and HS. Materials and methodsA system pharmacological approach including in vivo experiment, UHPLC-Q-Orbitrap HRMS, network pharmacology, molecular docking, microscale thermophoresis, and in vitro experiment was utilized to study the anti-ferroptosis mechanisms of AGNHW against IS and HS. ResultsIn vivo experiments indicated that AGNHW enhanced nerve function, decreased cerebral infarct volume, ameliorated histological brain injuries, improved the structural integrity of the blood-brain barrier, ameliorated the mitochondrial dysfunction and morphology disruption, and inhibits ROS, LPO and Fe2+ accumulations in IS and HS rats. Using UHPLC-Q-Orbitrap HRMS, the key ingredients of AGNHW-containing serum were identified as bilirubin, berberine, baicalin, and wogonoside. According to the network pharmacology analyses, AGNHW could inhibit ferroptosis by modulating the PPAR and PI3K/AKT signaling pathways. The core targets are PPARγ, AKT, and GPX4. Molecular docking and microscale thermophoresis experiments further revealed that the key ingredients have strong interactions with ferroptosis-regulating core proteins. Moreover, in vitro experiment results showed that AGNHW alleviated ferroptosis injury induced by erastin in PC12 cells, increased cell viability, reduced the LPO and Fe2+ levels, and up-regulated mRNA expressions of PPARγ, AKT, and GPX4. AGNHW also up-regulated protein expressions of PPARγ, p-AKT/AKT, and GPX4 in IS and HS rats. ConclusionsAGNHW attenuated ferroptosis in treating IS and HS by targeting the PPARγ/AKT/GPX4 pathway. This work reveals AGNHW's anti-ferroptosis mechanism against IS and HS, but it also develops an integrated approach to demonstrate the common characteristics of drugs in treating different diseases.

Dipeptidyl Peptidase 4 Stimulation Induces Adipogenesis-Related Gene Expression of Adipose Stromal Cells.

Adipogenesis has emerged as a new therapeutic target for regulating metabolism and achieving anti-inflammatory and anti-atherosclerotic effects via the release of adiponectin. However, at present, the effects and mechanism of action of dipeptidyl peptidase 4 (DPP4) stimulation on adiponectin production and adipogenesis have not been clarified. Here, we investigated the effects of DPP4 stimulation with monocyte chemoattractant protein-1 (MCP-1) on platelet-derived growth factor receptor alpha (PDGFRα) expression in adipose tissue and blood adiponectin levels. Stromal vascular fractions (SVFs) purified from human subcutaneous adipose tissue and inguinal adipose tissue of obese and diabetic (Leprdb/db) mice were treated with 50 ng of MCP-1 and plasma from control (Lepr+/+) mice supplemented with 10 ng or 50 ng of MCP-1. Treatment of SVFs from human subcutaneous adipose tissues with 50 ng of MCP-1 significantly increased AdipoQ, DPP4, peroxisome proliferator-activated receptor gamma (PPARγ), fatty-acid-binding protein (FABP4), and SERBF1 mRNA expression. MCP-1-supplemented plasma increased adiponectin, CCAAT-Enhancer-binding protein alpha (C/EBPα), DPP4, IL-33, and PDGFRα mRNA expression and adiponectin and DPP4 protein expression, while decreasing the expression of IL-10 mRNA in SVFs compared with the levels in the plasma treatment group. MCP-1-supplemented plasma was shown to increase PPARγ, PPARγ2, adiponectin, DPP4, and FABP4 and decrease IL-10 mRNA expression in PDGFRα cells from adipose tissue. Meanwhile, MCP-1-supplemented plasma increased MCP-1, PDGFRα, TNFα, adiponectin, and IL-1β and decreased IL-10 and FOXP3 mRNA expression in DPP4 cells. Moreover, the injection of MCP-1-supplemented plasma into adipose tissue increased the proportion of DPP4+ cells among PDGFRα+ cells from adipose tissue and plasma adiponectin levels of Leprdb/db mice compared with the levels in the plasma injection group. Our results demonstrate that DPP4+ cells are important adipose progenitor cells. Stimulation of DPP4 with MCP-1 increases adipogenesis-related gene expression and the population of DPP4+ cells among PDGFRα+ cells in SVFs and blood adiponectin levels. DPP4 stimulation could be a novel therapy to increase local adipogenesis and systemic adiponectin levels.

Abstract 11649: Serum Adiponectin Levels Correlate With Atrial PPARγ and TGFβ1 mRNA Expression in Patients Receiving Cardiac Surgery: Involvement of PPARγ on Atrial Fibrotic Remodeling

Introduction: The peroxisome proliferator-activated receptor gamma (PPAR γ) plays important roles in regulating processes related to fibrogenesis. On the other hand, adiponectin, a marker for PPARγ activity, is a hormone involved in maintaining energy homeostasis, and exerts anti-profibrotic effects in various organs. However, the roles of PPARγ and adiponectin on atrial fibrotic remodeling are still unknown. Hypothesis: We investigated the association of serum adiponectin levels and atrial gene expression of adiponectin, PPARγ and fibrosis-related genes in cardiovascular surgery patients including heart valve disease. Methods: Atrial mRNA expression levels including adiponectin and PPARγ were analyzed by real-time reverse transcription polymerase chain reaction (RT-PCR) in 59 cases (sinus rhythm (SR) 24 patients, atrial fibrillation (AF) 35 patients) in which left atrium (LA) tissues were removed from the appendage during surgery. The immunohistochemical studies were used to identify the protein expression. The serum levels of adiponectin were measured by ELISA. Results: The immunohistochemical staining showed the expression of adiponectin, PPARγ and PPARα. The significant correlations were observed among the mRNA expression levels of extracellular matrix (ECM) (COL1A1, COL3A1, fibronectin) and the fibrosis-related genes such as TGFβ1, TIMP1, MMP2, NOX2, CTGF and PDGFD attributed to the fibrosis of myocardium. The mRNA expression level of PPARγ, but not PPARα, positively correlated with that of the ECM and the fibrosis-related gene expression level. The mRNA expression level of adiponectin also positively correlated with that of ECM. Patients with AF had higher atrial expression level of adiponectin, PPARγ, COL1A1 and TGFβ than those with SR. Serum adiponectin levels significantly correlated with atrial mRNA expression level of PPARγ and TGFβ1. Patients with AF had serum higher levels of adiponectin than those with SR. Conclusions: Atrial PPARγ and adiponectin associate with atrial fibrotic remodeling in cardiovascular surgery patients. Serum adiponectin levels correlate with atrial PPARγ and TGFβ1 mRNA expression, and might have potential utility as a biomarker in atrial fibrotic remodeling.

Effect of nitro-conjugated linoleic acid on the inflammatory response of murine macrophages activated with lipopolysaccharide derived from Prevotella intermedia.

Nitro-conjugated linoleic acid (NO2-CLA) has been observed to manifest salutary signaling responses, including anti-inflammatory and antioxidant properties. Here, the authors have explored the influence and underlying mechanisms of NO2-CLA on the proinflammatory reaction of murine macrophages that were challenged with lipopolysaccharide (LPS) derived from Prevotella intermedia, a putative periodontopathic bacterium. Treatment of LPS-activated RAW264.7 cells with NO2-CLA notably dampened the secretion of iNOS-derived NO, IL-1β and IL-6 as well as their gene expressions and significantly enhanced the markers for M2 macrophage polarization. NO2-CLA promoted the HO-1 expression in cells challenged with LPS, and tin protoporphyrin IX, an HO-1 inhibitor, significantly reversed the NO2-CLA-mediated attenuation of NO secretion, but not IL-1β or IL-6. We found that cells treated with NO2-CLA significantly increased mRNA expression of PPAR-γ compared to control cells, and NO2-CLA significantly reverted the decrease in PPAR-γ mRNA caused by LPS. Nonetheless, antagonists to PPAR-γ were unable to reverse the NO2-CLA-mediated suppression of inflammatory mediators. In addition, NO2-CLA did not alter the p38 and JNK activation elicited by LPS. Both NF-κB reporter activity and IκB-α degradation caused by LPS were notably diminished by NO2-CLA. NO2-CLA was observed to interrupt the nuclear translocation and DNA binding of p50 subunits caused by LPS with no obvious alterations in p65 subunits. Further, NO2-CLA attenuated the phosphorylation of STAT1/3 elicited in response to LPS. We propose that NO2-CLA could be considered as a possible strategy for the therapy of periodontal disease, although additional researches are certainly required to confirm this.

Dual mechanism of silver nanoparticle-mediated upregulation of adipogenesis in mouse fibroblasts (3T3-L1) in vitro

Silver nanoparticles (AgNPs) are widespread in the environment due to the increase in their application e.g. in medicine as part of hard-to-heal wound dressings. Many studies have revealed easy diffusion of AgNPs into deep skin layers through damaged epidermis and contact with e.g. fibroblasts. Therefore, the aim of this study was to evaluate the impact of small-size AgNPs (10 nm) in ppm concentrations on the adipogenesis process in mouse embryo fibroblasts (3T3-L1). The results showed a decrease in the metabolic activity, followed by an increase in the reactive oxygen species (ROS) level in a dose- and time-dependent manner (0–20 ppm). The increased caspase-3 activity was observed only at the highest concentration (20 ppm) of AgNPs. Further analysis showed the ability of the tested NPs to increase the lipid accumulation in adipocytes, similar to ROSI [peroxisome proliferator-activated receptor gamma (PPARγ) agonist], measured by Oil-Red-O staining. Moreover, the analyses evidenced the ability of AgNPs to increase the lipoxygenase activity and malondialdehyde levels, which is probably based on ROS-dependent enhancement of lipid hydroperoxidation. Lastly, a significant increase in the PPARγ, Adiponectin, Resistin, Vegf, and Serpine mRNA expression was shown 6 h after the induction of the differentiation process. Based on the obtained results, it can be concluded that small-size AgNPs increase adipogenesis via ROS- and PPARγ-based mechanisms with potential engagement of crosstalk with the aryl hydrocarbon receptor, which is important due to the widespread application of AgNPs in medicine. However, more studies are needed to elucidate the full mechanism of these NPs in the tested cell model in depth.

Bo's abdominal acupuncture improves disordered metabolism in obese type 2 diabetic rats through regulating fibroblast growth factor 21 and its related adipokines.

To investigate the effect of Bo's abdominal acupuncture (BOAA) on fibroblast growth factor 21 (FGF21) and its related adipokines in type 2 diabetes mellitus (T2DM) rats. This study established obese T2DM rat model by high-fat diet (HFD) with a dose of streptozotocin (STZ, 30 mg/kg). Obese T2DM rats were randomly subdivided into four groups (n = 10): negative, BOAA, conventional acupuncture (COA group) and metformin group (Met group) groups. The biochemical parameters, mRNAs, and proteins were analyzed using enzyme-lined immunoassays kits, quantitative polymerase chain reaction and Western blot. Treatment with BOAA attenuated the histopathological changes in visceral fat and restored the alterations in the levels of body weight, fasting blood glucose (FBG), homeostasis model assessment for insulin resistance (HOMA-IR). BOAA treatment significantly decreased the levels of triglyceride, total cholesterol, low density lipoprotein cholesterol, leptin, and increased the serum levels of high-density lipoprotein cholesterol, fibroblast growth factor 21 (FGF21), adiponectin (ADP), peroxisome proliferator-activated receptor γ (PPAR-γ), C-peptide (C-P) in obese T2DM rats. Furthermore, BOAA treatment significantly increased the mRNA expressions of FGF21, ADP, leptin, PPAR-γ, PPAR-α and adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK). Besides, BOAA treatment upregulated the protein expressions of fibroblast growth factor receptors3 (FGFR3), PPAR-α, extracellular signal-regulated kinase (ERK), phosphorylated ERK (p-ERK), AMPK, p-AMPK, Liver kinase B1 (LKB1), phosphorylated LKB1 (p-LKB1), acetyl-CoA carboxylase (ACC) and phosphorylated ACC (p-ACC), while downregulated the protein expressions of FGF21 and PPAR-γ in visceral fat. BOAA treatment reduced FBG and body weight, and improved insulin sensitivity through regulating FGF21 signaling pathway and its related adipokine in obese T2DM rats.

Gastrodia elata extract exerts anti-obesity effects on obese mice caused by a high-starch diet by suppressing SREBP-1 and ChREBP

ObjectiveGastrodia elata (GE), a natural treatment, is extensively used in traditional Korean medicine. The study's goal was to look into how GE affected obese mice that had been fed a high-starch diet (HSD). MethodsThe C57BL/6 J mice used in this investigation were given HSD to make them obese. Gastrodia elata extract (GEE) was administered orally to the mice throughout the course of the next five weeks in doses of 100, 200, and 500 mg/kg. When the experiment was over, we measured the animals body weight, liver and fat weight, blood biochemical parameters, and gene expression that is related to obesity. ResultsThe overall body weight of animals was dramatically lowered via GEE oral treatment. In comparison to the HSD group, the serum and liver TG levels were considerably lower in the GEE-treated group. Additionally, regional fatty tissues and liver weight markedly dropped in the GEE-treated batch in comparison to the HSD batch. We observed considerably lower levels of PPAR-γ, C/EBP-α, and ChREBP mRNA expression level in the GEE-treated group in addition to lower levels of SREBP-1, ACC, and FAS protein expression. ConclusionThe studies show that GEE has anti-obesity advantages in the mouse model of obesity caused by HSD. Despite the fact that we utilized mice as a model for the experiment rather than people, who have different biochemical pathways, the finding has significant implications for how GEE generally functions.

Dietary supplementation of sodium butyrate enhances lactation performance by promoting nutrient digestion and mammary gland development in dairy cows

This experiment was to evaluate the influence of sodium butyrate (SB) addition on milk production, ruminal fermentation, nutrient digestion, and the development and metabolism regulation of the mammary gland in dairy cows. Forty Holstein dairy cows averaging 710 ± 18.5 kg body weight, 72.8 ± 3.66 d in milk (DIM), and 41.4 ± 1.42 kg/d milk production were divided into four treatments blocked by DIM and milk production. Treatments were control group, low SB, medium SB, and high SB with 0, 100, 200 and 300 g/d of SB addition per cow, respectively. The study lasted for 105 d. Production of milk, milk protein and lactose quadratically increased (P < 0.05), while fat-corrected milk, energy-corrected milk and milk fat yields linearly increased (P < 0.05) with increasing SB addition. The digestibility of dietary dry matter, organic matter, and crude protein linearly increased (P < 0.05), whereas the digestibility of ether extract, neutral detergent fibre, and acid detergent fibre quadratically increased (P < 0.05). Ruminal pH quadratically decreased (P = 0.04), while total volatile fatty acids (VFA) quadratically increased (P = 0.03) with increasing SB addition. The acetic acid to propionic acid ratio increased (P = 0.03) linearly due to the unaltered acetic acid molar percentage and a linear decrease in propionic acid molar percentage. Ruminal enzymatic activity of carboxymethyl-cellulase and α-amylase, populations of total bacteria, total anaerobic fungi, total protozoa, Ruminococcus albus, R. flavefaciens, Butyrivibrio fibrisolvens, Fibrobacter succinogenes, and Ruminobacter amylophilus linearly increased (P < 0.05). Blood glucose, urea nitrogen, and non-esterified fatty acids linearly decreased (P < 0.05), while total protein concentration linearly increased (P = 0.04). Moreover, the addition of SB at 200 g/d promoted (P < 0.05) mRNA and protein expression of PPARγ, SREBF1, ACACA, FASN, SCD, CCNA2, CCND1, PCNA, Bcl-2, GPR41, and the ratios of p-Akt/Akt and p-mTOR/mTOR, but decreased (P < 0.05) mRNA and protein expressions of Bax, caspase-3, and caspase-9. The results suggest that milk production and milk fat synthesis increased with SB addition by stimulating rumen fermentation, nutrient digestion, gene and protein expressions concerned with milk fat synthesis and mammary gland development.

Serelaxin inhibits LPS-induced inflammatory response in cardiac fibroblasts by activating PPAR-γ and suppressing NF-κB signaling pathway.

Serelaxin has an inhibitory effect on fibrosis. However, whether the anti-fibrotic effects of serelaxin are achieved by inhibiting the inflammatory response has not been clarified. This study aimed to investigate the role of serelaxin in lipopolysaccharide (LPS)-induced inflammation in cardiac fibroblasts (CFs) and elucidate the underlying mechanisms.CFs were isolated from adult rat hearts. The effect of serelaxin on the inhibition of the inflammatory response after LPS induction was examined. Cell viability was measured by MMT assay. Cell proliferation was determined using the cell-counting kit-8. The levels of inflammatory cytokines IL-1β, IL-6, TNF-α, and IL-10 were measured using an enzyme-linked immunosorbent assay (ELISA). The mRNA levels of α-SMA, collagen I/III, MMP-2, MMP-9, IL-1β, IL-6,TNF-α, IL-10, IκBα, p-IκBα, p65 subunit of nuclear factor-kappa B (NF-κB) and peroxisome proliferator-activated receptor-γ (PPAR-γ) were assessed by real-time quantitative PCR (RT-qPCR). The protein levels of α-SMA, collagen I/III, MMP-2, MMP-9, IκBα, p-IκBα, p65, p-p65 and PPAR-γ were examined by western blotting.Serelaxin inhibited LPS-induced IL-1β, IL-6, TNF-α, α-SMA, and collagen I/III, and elevated the expression of IL-10, MMP-2 and MMP-9. Moreover, LPS-induced activation of NF-κB pathway was suppressed by serelaxin treatment. Further studies showed that serelaxin did not significantly increase the expression of PPAR-γ mRNA and protein, but activated PPAR-γ activity, and the PPAR-γ inhibitor GW9662 reversed the inhibitory effect of serelaxin on IL-1β, IL-6, and TNF-α production. These results suggest that serelaxin alleviates cardiac fibrosis by stimulating PPAR-γ via a ligand-independent mechanism which subsequently abolish the expression of NF-κB signalling pathway.

Duhuo Jisheng decoction alleviates neuroinflammation and neuropathic pain by suppressing microglial M1 polarization: a network pharmacology research

BackgroundNeuropathic pain (NP) is the most prevalent form of chronic pain resulting from nerve damage or injury. Despite the widespread use of Duhuo Jisheng decoction (DHJSD) in traditional Chinese medicine (TCM) to treat chronic pain, the mechanism underlying its analgesic action remains unclear.MethodsUsing network pharmacology, we obtained DHJSD and NP-related target information from public databases to construct protein–protein interactions (PPI) and compound-target networks based on common target genes. These networks were further analyzed using gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG). The interaction between molecules was verified through molecular docking using AutoDock Tools software. Additionally, we treated a chronic constriction injury (CCI) rat model with DHJSD and determined the mechanical withdrawal threshold (MWT). We used an enzyme-linked immunosorbent assay kit to determine the levels of inflammatory cytokines. Furthermore, qRT-PCR was employed to analyze ACHE, NOS2, MAPK3, PTGS2, AKT1, and PPARG mRNA expression, and immunofluorescence was used to evaluate changes in microglia.ResultsOur screening of compounds and targets identified 252 potential targets of DHJSD associated with NP. PPI analysis, along with GO and KEGG analyses, revealed that the potential mechanism of DHJSD in NP treatment may be related to inflammatory reactions, the IL-17 signaling pathway, MAP kinase activity, and endocrine activity. Based on molecular docking, the core target showed significant affinity for DHJSD's active components. Moreover, DHJSD treatment repaired the CCI-induced inflammatory reaction in the spinal cord while regulating the expression of ACHE, NOS2, MAPK3, PTGS2, AKT1, and PPARG mRNA. Immunofluorescence results indicated that the active components of DHJSD may regulate microglial M1 polarization to improve neuroinflammation, PPARG may have been involved in the process.ConclusionThe multi-component, multi-target, and multi-pathway actions of DHJSD provide new insights into its therapeutic mechanism in NP.

Autologous Platelet Lysate Is an Alternative to Fetal Bovine Serum for Canine Adipose-Derived Mesenchymal Stem Cell Culture and Differentiation.

The use of fetal bovine serum (FBS) in regenerative medicine raises serious ethical and scientific concerns. We have cultured and differentiated the canine mesenchymal stem cells (cMSCs) in five different media combinations of autologous platelet lysate (A-PL) and FBS; consisting of 0% A-PL and 10% FBS (M-1), 2.5% A-PL and 7.5% FBS (M-2), 5% A-PL and 5% FBS (M-3), 7.5% A-PL and 2.5% FBS (M-4), and 10% A-PL and 0% FBS (M-5). The cMSCs were evaluated for their doubling time, differentiation efficiency, and expression of CD73, CD90, CD105, and PDGFRα. The mRNA expression of NT5E, THY1, ENG, PPARγ, FABP4, FAS, SP7, BGLAP, and SPP1 was also assessed. The results indicated non-significant differences in cellular proliferation/viability; positive expression of surface markers, and PDGFRα with substantial adipo/osteogenic differentiation. The expression of adipogenic (PPARγ, FABP4, FAS), and osteogenic (SP7, BGLAP, SPP1) genes were higher (p < 0.05) in the M5 group. In conclusion, A-PL in cMSCs culture did not negatively affect cellular proliferation and viability but also enhanced their genetic potential for multilineage differentiation. Our results indicate that A-PL can be used as an alternative for FBS to develop potent cMSCs under good manufacturing practice protocol for regenerative medicine.