Transcription Factor PPARγ Research Articles

Synthetic curcumin analog: inhibiting the invasion, angiogenesis, and metastasis in human laryngeal carcinoma cells via NF-kB pathway.

Laryngeal carcinoma, the most common among head and neck squamous cell carcinoma (HNSCC), induces 1% of all cancer deaths. Curcumin the active constituent of turmeric, is shown to be effective in the treatment of various cancers. In the present study, we explored the mechanistic role of bis-demethoxy curcumin analog (BDMC-A) as a chemotherapeutic agent. We investigated its inhibitory effect on invasion, angiogenesis, and metastasis in human laryngeal carcinoma (Hep-2) cells in comparison with curcumin. The effect of curcumin and BDMC-A on transcription factors (NF-κB, p65, c-Jun, c-Fos, STAT3, 5, PPAR-γ, β-catenin, COX-2, MMP-9, VEGF, TIMP-2) involved in signal transduction cascade, invasion, and angiogenesis in Hep-2 cells were quantified using Western blotting and RT-PCR technique. ELISA was used to measure the pro-inflammatory markers in Hep-2 cells treated with curcumin and BDMC-A. The results showed that BDMC-A inhibits the transcription factors NF-κB, p65, c-Jun, c-Fos, STAT3, STAT5, PPAR-γ and β-catenin, which are responsible for tumor progression and malignancy. Moreover, BDMC-A treatment downregulated MMP-9, VEGF, TGF- β, IL-6 and IL-8 and upregulated TIMP-2 levels. The effects were more significant compared to curcumin. Our overall results revealed that BDMC-A more effectively inhibited the markers of invasion, angiogenesis and metastasis in comparison with curcumin.

Increasing the Power of Polyphenols through Nanoencapsulation for Adjuvant Therapy against Cardiovascular Diseases

Polyphenols play a therapeutic role in vascular diseases, acting in inherent illness-associate conditions such as inflammation, diabetes, dyslipidemia, hypertension, and oxidative stress, as demonstrated by clinical trials and epidemiological surveys. The main polyphenol cardioprotective mechanisms rely on increased nitric oxide, decreased asymmetric dimethylarginine levels, upregulation of genes encoding antioxidant enzymes via the Nrf2-ARE pathway and anti-inflammatory action through the redox-sensitive transcription factor NF-κB and PPAR-γ receptor. However, poor polyphenol bioavailability and extensive metabolization restrict their applicability. Polyphenols carried by nanoparticles circumvent these limitations providing controlled release and better solubility, chemical protection, and target achievement. Nano-encapsulate polyphenols loaded in food grade polymers and lipids appear to be safe, gaining resistance in the enteric route for intestinal absorption, in which the mucoadhesiveness ensures their increased uptake, achieving high systemic levels in non-metabolized forms. Nano-capsules confer a gradual release to these compounds, as well as longer half-lives and cell and whole organism permanence, reinforcing their effectiveness, as demonstrated in pre-clinical trials, enabling their application as an adjuvant therapy against cardiovascular diseases. Polyphenol entrapment in nanoparticles should be encouraged in nutraceutical manufacturing for the fortification of foods and beverages. This study discusses pre-clinical trials evaluating how nano-encapsulate polyphenols following oral administration can aid in cardiovascular performance.

Anti-Obesity and Lipid Metabolism Effects of Ulmus davidiana var. japonica in Mice Fed a High-Fat Diet.

The root bark of Ulmus davidiana var. japonica (Japanese elm) is used in Korea and other East Asian countries as a traditional herbal remedy to treat a variety of inflammatory diseases and ailments such as edema, gastric cancer and mastitis. For this study, we investigated the lipid metabolism and anti-obesity efficacy of ethyl alcohol extract of Ulmus davidiana var. japonica root bark (UDE). First, HPLC was performed to quantify the level of (+)-catechin, the active ingredient of UDE. In the following experiments, cultured 3T3-L1 pre-adipocytes and high-fat diet (HFD)-fed murine model were studied for anti-obesity efficacy by testing the lipid metabolism effects of UDE and (+)-catechin. In the test using 3T3-L1 pre-adipocytes, treatment with UDE inhibited adipocyte differentiation and significantly reduced the production of adipogenic genes and transcription factors PPARγ, C/EBPα and SREBP-1c. HFD-fed, obese mice were administered with UDE (200 mg/kg per day) and (+)-catechin (30 mg/kg per day) by oral gavage for 4 weeks. Weight gain, epididymal and abdominal adipose tissue mass were significantly reduced, and a change in adipocyte size was observed in the UDE and (+)-catechin treatment groups compared to the untreated control group (***p < 0.001). Significantly lower total cholesterol and triglyceride levels were detected in UDE-treated HFD mice compared to the control, revealing the efficacy of UDE. In addition, it was found that lipid accumulation in hepatocytes was also significantly reduced after administration of UDE. These results suggest that UDE has significant anti-obesity and lipid metabolism effects through inhibition of adipocyte differentiation and adipogenesis.

Chamomile as a potential remedy for obesity and metabolic syndrome.

Obesity is an increasing health concern related to many metabolic disorders, including metabolic syndrome, diabetes type 2 and cardiovascular diseases. Many studies suggest that herbal products can be useful dietary supplements for weight management due to the presence of numerous biologically active compounds, including antioxidant polyphenols that can counteract obesity-related oxidative stress. In this review we focus on Matricaria chamomilla, commonly known as chamomile, and one of the most popular medicinal plants in the world. Thanks to a high content of phenolic compounds and essential oils, preparations from chamomile flowers demonstrate a number of pharmacological effects, including antioxidant, anti-inflammatory, antimicrobial and sedative actions as well as improving gastrointestinal function. Several recent studies have shown certain positive effects of chamomile preparations in the prevention of obesity and complications of diabetes. These effects were associated with modulation of signaling pathways involving the AMP-activated protein kinase, NF-κB, Nrf2 and PPARγ transcription factors. However, the potential of chamomile in the management of obesity seems to be underestimated. This review summarizes current data on the use of chamomile and its individual components (apigenin, luteolin, essential oils) to treat obesity and related metabolic disorders in cell and animal models and in human studies. Special attention is paid to molecular mechanisms that can be involved in the anti-obesity effects of chamomile preparations. Limitation of chamomile usage is also analyzed.

Determination of Oxidized Lipids in Commonly Consumed Foods and a Preliminary Analysis of Their Binding Affinity to PPARγ.

Foods rich in poly unsaturated fatty acids (PUFA) are vulnerable to oxidation. While it is well established that endogenously derived oxidized lipids are ligands of the transcription factor PPARγ, the binding ability of diet-derived oxidized lipids is unknown. Our two-fold objective was to determine the oxidized lipid content and PPARγ binding ability of commonly consumed foods. Extracted food lipids were assayed for the peroxide value, conjugated dienes, and aldehydes, and PPARγ binding was assessed by an in vitro PPARγ ligand screening assay. Oxidized lipids were present in all foods tested at the time of purchase, and oxidation did not increase during storage. The peroxide values for walnuts, sunflower seeds, and flax meal were significantly lower at the end of three months as compared to the day of purchase (peroxide value: 1.26 ± 0.13 vs. 2.32 ± 0.4; 1.65 ± 0.23 vs. 2.08 ± 0.09; 3.07 ± 0.22 vs. 9.94 ± 0.75 mEq/kg fat, p ≤ 0.05, respectively). Lipids extracted from French fries had the highest binding affinity (50.87 ± 11.76%) to PPARγ compared to other foods. Our work demonstrates that oxidized lipids are present in commonly consumed foods when purchased, and for the first time demonstrates that some contain ligands of PPARγ.

The B56α subunit of PP2A is necessary for mesenchymal stem cell commitment to adipocyte.

Adipose tissue plays a major role in maintaining organismal metabolic equilibrium. Control over the fate decision from mesenchymal stem cells (MSCs) to adipocyte differentiation involves coordinated command of phosphorylation. Protein phosphatase 2A plays an important role in Wnt pathway and adipocyte development, yet how PP2A complexes actively respond to adipocyte differentiation signals and acquire specificity in the face of the promiscuous activity of its catalytic subunit remains unknown. Here, we report the PP2A phosphatase B subunit B56α is specifically induced during adipocyte differentiation and mediates PP2A to dephosphorylate GSK3β, thereby blocking Wnt activity and driving adipocyte differentiation. Using an inducible B56α knock-out mouse, we further demonstrate that B56α is essential for gonadal adipose tissue development invivo and required for the fate decision of adipocytes over osteoblasts. Moreover, we show B56α expression is driven by the adipocyte transcription factor PPARγ thereby establishing a novel link between PPARγ signaling and Wnt blockade. Overall, our results reveal B56α is a necessary part of the machinery dictating the transition from pre-adipocyte to mature adipocyte and provide fundamental insights into how PP2A complex specifically and actively regulates unique signaling pathway in biology.

SIRT6 transcriptionally regulates fatty acid transport by suppressing PPARγ.

SUMMARYPathological lipid accumulation is often associated with enhanced uptake of free fatty acids via specific transporters in cardiomyocytes. Here, we identify SIRT6 as a critical transcriptional regulator of fatty acid transporters in cardiomyocytes. We find that SIRT6 deficiency enhances the expression of fatty acid transporters, leading to enhanced fatty acid uptake and lipid accumulation. Interestingly, the haploinsufficiency of SIRT6 is sufficient to induce the expression of fatty acid transporters and cause lipid accumulation in murine hearts. Mechanistically, SIRT6 depletion enhances the occupancy of the transcription factor PPARγ on the promoters of critical fatty acid transporters without modulating the acetylation of histone 3 at Lys 9 and Lys 56. Notably, the binding of SIRT6 to the DNA-binding domain of PPARγ is critical for regulating the expression of fatty acid transporters in cardiomyocytes. Our data suggest exploiting SIRT6 as a potential therapeutic target for protecting the heart from metabolic diseases.

MED1 is a lipogenesis coactivator required for postnatal adipose expansion

MED1 often serves as a surrogate of the general transcription coactivator complex Mediator for identifying active enhancers. MED1 is required for phenotypic conversion of fibroblasts to adipocytes in vitro but its role in adipose development and expansion in vivo has not been reported. Here we show that MED1 is dispensable for adipose development in mice. Instead, MED1 is required for postnatal adipose expansion and the induction of de novo lipogenesis (DNL) genes after pups switch diet from high-fat maternal milk to carbohydrate-based chow. During adipogenesis, MED1 is dispensable for induction of lineage-determining transcription factors (TFs) PPARγ and C/EBPα but is required for lipid accumulation in the late phase of differentiation. Mechanistically, MED1 controls the induction of DNL genes by facilitating lipogenic TF ChREBP-dependent recruitment of Mediator to active enhancers. Together, our findings identify a cell- and gene-specific regulatory role of MED1 as a lipogenesis coactivator required for postnatal adipose expansion.

MED1 is a lipogenesis coactivator required for postnatal adipose expansion.

MED1 often serves as a surrogate of the general transcription coactivator complex Mediator for identifying active enhancers. MED1 is required for phenotypic conversion of fibroblasts to adipocytes in vitro, but its role in adipose development and expansion in vivo has not been reported. Here, we show that MED1 is not generally required for transcription during adipogenesis in culture and that MED1 is dispensable for adipose development in mice. Instead, MED1 is required for postnatal adipose expansion and the induction of fatty acid and triglyceride synthesis genes after pups switch diet from high-fat maternal milk to carbohydrate-based chow. During adipogenesis, MED1 is dispensable for induction of lineage-determining transcription factors (TFs) PPARγ and C/EBPα but is required for lipid accumulation in the late phase of differentiation. Mechanistically, MED1 controls the induction of lipogenesis genes by facilitating lipogenic TF ChREBP- and SREBP1a-dependent recruitment of Mediator to active enhancers. Together, our findings identify a cell- and gene-specific regulatory role of MED1 as a lipogenesis coactivator required for postnatal adipose expansion.

Cyclooxygenase-2 Inhibitor Parecoxib Was Disclosed as a PPAR-γ Agonist by In Silico and In Vitro Assay.

In a search for effective PPAR-γ agonists, 110 clinical drugs were screened via molecular docking, and 9 drugs, including parecoxib, were selected for subsequent biological evaluation. Molecular docking of parecoxib to the ligand-binding domain of PPAR-γ showed high binding affinity and relevant binding conformation compared with the PPAR-γ ligand/antidiabetic drug rosiglitazone. Per the docking result, parecoxib showed the best PPAR-γ transactivation in Ac2F rat liver cells. Further docking simulation and a luciferase assay suggested parecoxib would be a selective (and partial) PPAR-γ agonist. PPAR-γ activation by parecoxib induced adipocyte differentiation in 3T3-L1 murine preadipocytes. Parecoxib promoted adipogenesis in a dose-dependent manner and enhanced the expression of adipogenesis transcription factors PPAR-γ, C/EBPα, and C/EBPβ. These data indicated that parecoxib might be utilized as a partial PPAR-γ agonist for drug repositioning study.

PPARγ and Diabetes: Beyond the Genome and Towards Personalized Medicine.

Full and partial synthetic agonists targeting the transcription factor PPARγ are contained in FDA-approved insulin-sensitizing drugs and used for the treatment of metabolic syndrome-related dysfunctions. Here, we discuss the association between PPARG genetic variants and drug efficacy, as well as the role of alternative splicing and post-translational modifications as contributors to the complexity of PPARγ signaling and to the effects of synthetic PPARγ ligands. PPARγ regulates the transcription of several target genes governing adipocyte differentiation and glucose and lipid metabolism, as well as insulin sensitivity and inflammatory pathways. These pleiotropic functions confer great relevance to PPARγ in physiological regulation of whole-body metabolism, as well as in the etiology of metabolic disorders. Accordingly, PPARG gene mutations, nucleotide variations, and post-translational modifications have been associated with adipose tissue disorders and the related risk of insulin resistance and type 2 diabetes (T2D). Moreover, PPARγ alternative splicing isoforms-generating dominant-negative isoforms mainly expressed in human adipose tissue-have been related to impaired PPARγ activity and adipose tissue dysfunctions. Thus, multiple regulatory levels that contribute to PPARγ signaling complexity may account for the beneficial as well as adverse effects of PPARγ agonists. Further targeted analyses, taking into account all these aspects, are needed for better deciphering the role of PPARγ in human pathophysiology, especially in insulin resistance and T2D. The therapeutic potential of full and partial PPARγ synthetic agonists underlines the clinical significance of this nuclear receptor. PPARG mutations, polymorphisms, alternative splicing isoforms, and post-translational modifications may contribute to the pathogenesis of metabolic disorders, also influencing the responsiveness of pharmacological therapy. Therefore, in the context of the current evidence-based trend to personalized diabetes management, we highlight the need to decipher the intricate regulation of PPARγ signaling to pave the way to tailored therapies in patients with insulin resistance and T2D.

Thymoquinone, a Dietary Bioactive Compound, Exerts Anti-Inflammatory Effects in Colitis by Stimulating Expression of the Colonic Epithelial PPAR-γ Transcription Factor.

Inflammatory bowel diseases (IBD) are chronic inflammatory disorders with increasing incidence and prevalence worldwide. Here, we investigated thymoquinone (TQ), a naturally occurring phytochemical present in Nigella sativa, for anti-inflammatory effects in colonic inflammation. To address this, we used in vivo (mice) and in vitro (HT-29 cells) models in this investigation. Our results showed that TQ treatment significantly reduced the disease activity index (DAI), myeloperoxidase (MPO) activity, and protected colon microscopic architecture. In addition, TQ also reduced the expression of proinflammatory cytokines and mediators at both the mRNA and protein levels. Further, TQ decreased phosphorylation of the activated mitogen-activated protein kinase (MAPK) signaling pathway and nuclear factor kappa B (NF-κB) proteins and enhanced colon epithelial PPAR-γ transcription factor expression. TQ significantly decreased proinflammatory chemokines (CXCL-1 and IL-8), and mediator (COX-2) mRNA expression in HT-29 cells treated with TNF-α. TQ also increased HT-29 PPAR-γ mRNA, PPAR-γ protein expression, and PPAR-γ promoter activity. These results indicate that TQ inhibits MAPK and NF-κB signaling pathways and transcriptionally regulates PPAR-γ expression to induce potent anti-inflammatory activity in vivo and in vitro models of colon inflammation.

Regulation of gene expression associated with LC‐PUFA metabolism in juvenile tambaqui ( Colossoma macropomum ) fed different dietary oil sources

Given the ecological unsustainability of using fish oil (FO) as the main source of long-chain polyunsaturated fatty acids (LC-PUFA), the investigation of alternative dietary LC-PUFA sources is crucial for aquaculture. Tambaqui (Colossoma macropomum) is a valuable economic aquaculture resource in Brazil, capable of endogenously elongating and desaturating linoleic (LA; 18:2n-6) and α-linolenic (ALA; 18:3n-3) acids to longer and physiologically vital LC-PUFA. Yet, it is unclear how this pathway is regulated by different oil sources in tambaqui. Thus, we designed an experiment with 2 different oil diets (fish oil—FO and vegetable oil—VO) at 2 different concentrations (5% or 10%) to investigate the molecular regulation of fatty acid biosynthesis in this species. We observed no differences in survival, body weight gain (BWG) and feed conversion ratio between the treatments. Gene expression analysis shows that fads2 and elovl5 are up-regulated in liver, while fads2 and elovl2 are up-regulated in brain of tambaqui fed with VO. The transcription factors pparβb and pparγ are also up-regulated in the brain by VO diet, when compared to FO diet. The VO diet also contributed to the biosynthesis of LC-PUFA in liver and specifically DHA in the brain. Overall, our approach shows that lipid metabolism-relevant genes are regulated by different dietary lipid sources.

Analysis of ANGPTL8 promoter activity and screening of related transcription factors in bovine.

Analysing the molecular regulation mechanism of fat deposition in yellow cattle can provide a theoretical basis for the breeding of excellent beef cattle. ANGPTL8 (angiopoietin-like protein 8) promotes the formation of lipid droplets during adipocyte differentiation. To explore the promoter active region of ANGPTL8 and predict potential transcription factors, we further provide a theoretical basis for the functional analysis and regulatory mechanism of ANGPTL8 in adipogenesis. The promoter region of bovine ANGPTL8 was cloned by overlap extension PCR. Online software was used to predict potential transcription factor binding sites, and it identified PPARγ, SREBP1, C/EBPα, and Znf423 transcription factor binding sites in ANGPTL8 promoter region. A luciferase reporter gene vector which contained different deletion fragments of the ANGPTL8 promoter was constructed. Then, the vectors were cotransfected into 293T cells with the internal control plasmid pRL-TK by cationic liposomes, and the relative fluorescence intensity was detected by a microplate reader. The results of the luciferase activity analysis showed that the core promoter area of ANGPTL8 was in the -885/-227bp region of the 5' flanking sequence, while just two SREBP1 binding sites occurred in this area. When SREBP1 was knocked down by siRNA, the expression level of ANGPTL8 was reduced, and we speculated that SREBP1 may be an important transcription factor regulating ANGPTL8 transcription.

PPARγ marks splenic precursors of multiple nonlymphoid-tissue Treg compartments

Foxp3+CD4+ regulatory T cells (Tregs) regulate most types of immune response as well as several processes important for tissue homeostasis, for example, metabolism and repair. Dedicated Treg compartments-with distinct transcriptomes, T cell receptor repertoires, and growth/survival factor dependencies-have been identified in several nonlymphoid tissues. These Tregs are specifically adapted to function and operate in their home tissue-When, where, and how do they take on their specialized characteristics? We recently reported that a splenic Treg population expressing low levels of the transcription factor PPARγ (peroxisome proliferator-activated receptor gamma) contains precursors of Tregs residing in visceral adipose tissue. This finding made sense given that PPARγ, the "master regulator" of adipocyte differentiation, is required for the accumulation and function of Tregs in visceral adipose tissue but not in lymphoid tissues. Here we use single-cell RNA sequencing, single-cell Tcra and Tcrb sequencing, and adoptive-transfer experiments to show that, unexpectedly, the splenic PPARγlo Treg population is transcriptionally heterogeneous and engenders Tregs in multiple nonlymphoid tissues beyond visceral adipose tissue, such as skin and liver. The existence of a general pool of splenic precursors for nonlymphoid-tissue Tregs opens possibilities for regulating their emergence experimentally or therapeutically.

Glitazones, PPAR-γ and Neuroprotection.

The transcriptional factor PPAR-γ belongs to the nuclear receptor family, which has become a potential therapeutic target for several neurodegenerative diseases and metabolic disorders. Interestingly, PPAR-γ has been reported to have beneficial effects in various chronic neurological conditions via upregulation of its transcriptional co-activator PGC-1α and followed by regulation of multiple molecular events. Although several factors contribute to the progression of neurodegeneration, the dysfunction of PGC-1α expression is primarily interlinked with the pathogenesis of major neurodegenerative diseases. This review gives an insight that ligand-dependent activation of PPAR-γ by glitazones could initiate the structural conformational changes of the secondary proteins, thus recruiting the PGC-1α to form a stable regulatory complex that hampers the various molecular pathways contributing to neurodegeneration. The promising outcomes of the preliminary in silico studies included in this review support that PPAR-γ dependent activation of central PGC-1α signaling by novel glitazones is an encouraging strategy to enhance the oxy-radicals detoxifying system, antiinflammatory responses, and mitochondrial biogenesis required for neuroprotection in various neurodegenerative conditions.

Coleus forskohlii and Garcinia indica extracts attenuated lipid accumulation by regulating energy metabolism and modulating gut microbiota in obese mice

Obesity is related to energy imbalance and energy metabolism. In this study, we investigated the anti-obesity effects of Garcinia indica extract (GIE), Coleus forskohlii extract (CFE), and the combinations of these two extracts in a 3T3-L1 cells and high-fat diet (HFD)-induced obese mice. In vitro, GIE showed better effect on TG content than CFE, CFE showed better effect on glycerol released than GIE, and the combinations of GIE and CFE showed both effects compared with GIE and CFE alone. In vivo, GIE, LMIX (0.005% GIE + 0.025% CFE), and HMIX (0.01% GIE + 0.025% CFE) down-regulated adipogenesis-related transcription factors PPARγ and C/EBPα protein expression, CFE promoted lipolysis by up-regulated p-HSL and p-PKA protein expression, and four supplementations promoted fatty acid β-oxidation by up-regulating CPT-1A and PPARα protein expression to decrease lipid accumulation in adipose tissue. Moreover, we found that CFE, LMIX and HMIX, except GIE exert increasing the abundance of Bacteroides caccae compared with HFD group. Overall, GIE, CFE, and the combinations of GIE and CFE were able to decrease body weight and adipocyte size by promoting fatty acid β-oxidation and modulating gut microbiota in HFD-induced obese mice.

TRPA1 Agonist Cinnamaldehyde Decreases Adipogenesis in 3T3-L1 Cells More Potently than the Non-agonist Structural Analog Cinnamyl Isobutyrate.

The cinnamon-derived bioactive aroma compound cinnamaldehyde (CAL) has been identified as a promising antiobesity agent, inhibiting adipogenesis and decreasing lipid accumulation in vitro as well as in animal models. Here, we investigated the antiadipogenic effect of cinnamyl isobutyrate (CIB), another cinnamon-derived aroma compound, in comparison to CAL in 3T3-L1 adipocyte cells. In a concentration of 30 μM, CIB reduced triglyceride (TG) and phospholipid (PL) accumulation in 3T3-L1 pre-adipocytes by 21.4 ± 2.56 and 20.7 ± 2.05%, respectively. CAL (30 μM), in comparison, decreased TG accumulation by 37.5 ± 1.81% and PL accumulation by 28.7 ± 1.83%, revealing the aldehyde to be the more potent antiadipogenic compound. The CIB- and CAL-mediated inhibition of lipid accumulation was accompanied by downregulation of essential adipogenic transcription factors PPARγ, C/EBPα, and C/EBPβ on gene and protein levels, pointing to a compound-modulated effect on adipogenic signaling cascades. Coincubation experiments applying the TRPA-1 inhibitor AP-18 demonstrated TRPA1 dependency of the CAL, but not the CIB-induced antiadipogenic effect.

Lipid-Lowering Effects of Lotus Leaf Alcoholic Extract on Serum, Hepatopancreas, and Muscle of Juvenile Grass Carp via Gene Expression.

Compared with wild grass carp (Ctenopharyngodon idellus), intensively cultured fish displayed disordered lipid metabolism, showing excess lipid deposition in the hepatopancreas and muscle. Lotus leaf prevents fat accumulation in humans and may have similar effects on fish. This study explored the regulatory mechanisms by which the dietary addition of an alcoholic extract of lotus leaf (AELL) reduced lipid deposition in the hepatopancreas and muscle of juvenile grass carp. The fish (average initial weight: 34.00 ± 0.40 g) were fed four experimental diets containing different AELL levels (0, 0.07, 0.14, and 0.21%) for 8 weeks. Serum components, lipid droplet size, triacylglycerol (TAG) content, enzymatic activities, and mRNA levels of genes related to lipid metabolism in the hepatopancreas and muscle were analyzed. The results show that dietary AELL supplementation significantly reduced the TAG content and lipid droplet area in the histological sections as well as the fatty acid synthase (FAS) activity in both the hepatopancreas and muscle but enhanced the activities of lipoprotein lipase (LPL) and carnitine palmitoyltransferase I (CPT1) in both tissues. In addition, dietary AELL supplementation decreased the mRNA expression of genes involved in fatty acid uptake (cd36, fatp1/fatp4/fatp6, fabp10/fabp11, acsl1/acsl4) and de novo lipid synthesis (pgd, g6pd, and fasn) as well as the transcription factors pparg and srebf1 in the hepatopancreas and muscle but increased the mRNA levels of genes relating to lipid catabolism (cpt1a, lipe, pnpla2, lpl), lipid transportation (apob), and the transcription factor ppara in both tissues. In conclusion, dietary AELL supplementation reduced lipid accumulation in the hepatopancreas and muscle by affecting the gene expression of proteins with known effects on lipid metabolism in juvenile grass carp.

LMWF5A suppresses cytokine release by modulating select inflammatory transcription factor activity in stimulated PBMC

BackgroundDysregulation of transcription and cytokine expression has been implicated in the pathogenesis of a variety inflammatory diseases. The resulting imbalance between inflammatory and resolving transcriptional programs can cause an overabundance of pro-inflammatory, classically activated macrophage type 1 (M1) and/or helper T cell type 1 (Th1) products, such as IFNγ, TNFα, IL1-β, and IL12, that prevent immune switching to resolution and healing. The low molecular weight fraction of human serum albumin (LMWF5A) is a novel biologic drug that is currently under clinical investigation for the treatment of osteoarthritis and the hyper-inflammatory response associated with COVID-19. This study aims to elucidate transcriptional mechanisms of action involved with the ability of LMWF5A to reduce pro-inflammatory cytokine release.MethodsELISA arrays were used to identify cytokines and chemokines influenced by LMWF5A treatment of LPS-stimulated peripheral blood mononuclear cells (PBMC). The resulting profiles were analyzed by gene enrichment to gain mechanistic insight into the biologic processes and transcription factors (TFs) underlying the identified differentially expressed cytokines. DNA-binding ELISAs, luciferase reporter assays, and TNFα or IL-1β relative potency were then employed to confirm the involvement of enriched pathways and TFs.ResultsLMWF5A was found to significantly inhibit a distinct set of pro-inflammatory cytokines (TNFα, IL-1β, IL-12, CXCL9, CXCL10, and CXCL11) associated with pro-inflammatory M1/Th1 immune profiles. Gene enrichment analysis also suggests these cytokines are, in part, regulated by NF-κB and STAT transcription factors. Data from DNA-binding and reporter assays support this with LMWF5A inhibition of STAT1α DNA-binding activity as well as a reduction in overall NF-κB-driven luciferase expression. Experiments using antagonists specific for the immunomodulatory and NF-κB/STAT-repressing transcription factors, peroxisome proliferator-activated receptor (PPAR)γ and aryl hydrocarbon receptor (AhR), indicate these pathways are involved in the LMWF5A mechanisms of action by reducing LMWF5A drug potency as measured by TNFα and IL-1β release.ConclusionIn this report, we provide evidence that LMWF5A reduces pro-inflammatory cytokine release by activating the immunoregulatory transcription factors PPARγ and AhR. In addition, our data indicate that LMWF5A suppresses NF-κB and STAT1α pro-inflammatory pathways. This suggests that LMWF5A acts through these mechanisms to decrease pro-inflammatory transcription factor activity and subsequent inflammatory cytokine production.