coumestrol and Inflammation

Coumestrol is a phytoestrogen found in various plant foods. Increasing evidence ascertained its robust anti-inflammatory, anti-oxidative properties likewise ability to mitigate insulin resistance. Thus, it may be a potential medicine in the treatment of many metabolic disorders, including obesity, type 2 diabetes (T2D) as well as non-alcoholic fatty liver disease (NAFLD). In this study, we aimed to shed some light on its influence on the accumulation of certain lipid fractions and the expression of pro-inflammatory proteins in primary rat hepatocytes during the lipid-overload state. The cells were isolated from the male Wistar rat's liver with the use of collagenase perfusion. It was followed by incubation of the cells with the presence or absence of palmitic acid and/or coumestrol. The accumulation of lipid fractions was assessed by gas-liquid chromatography (GLC) whereas the expression of the proteins was evaluated by the Western blot technique. Treatment with coumestrol in the state of increased fatty acids availability led to the deposition of triacylglycerols rather than diacylglycerols, significantly decreased expression of proinflammatory and profibrotic cytokines, especially interleukin 6 (IL-6) and tumor necrosis factor α (TNF-α), as well as transforming growth factor β (TGF-β), and nuclear factor κβ (NF-κβ). Also, we observed a substantial diminution in proinflammatory enzymes expression. Taking into consideration the direction of the aforementioned changes, we may assume that coumestrol can ameliorate the array of factors leading to the development of steatosis, likewise counteracting progression to steatohepatitis, thus it may be a step forward to the long-awaited breakthrough in the treatment of NAFLD.

Topics: Animals; Coumestrol; Diabetes Mellitus, Type 2; Fatty Acids; Hepatocytes; Inflammation; Liver; Non-alcoholic Fatty Liver Disease; Rats; Rats, Wistar

Diabetes-induced oxidative stress is vital in initiating neuronal damage in the diabetic retina, leading to diabetic retinopathy (DR). This study investigates the possible effects of coumestrol (CMS) on streptozotocin (STZ)-induced DR. First, we established a rat model of DR by STZ injection and a cell model involving high-glucose (HG) exposure of human retinal microvascular endothelial cells (hRMECs). We characterized the expression patterns of oxidative stress indicators, pro-inflammatory cytokines, and pro-apoptotic proteins in hRMECs. Polymerase chain reaction showed sirtuin 1 (SIRT1) to be poorly expressed in the retinal tissues of STZ-treated rats and HG-exposed hRMECs, but its expression was upregulated upon treatment with CMS treatment. Furthermore, CMS treatment attenuated the STZ-induced pathologies such as oxidative stress, inflammation, and cell apoptosis. Consistent with the

Topics: Animals; Apoptosis; Coumestrol; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Disease Models, Animal; Endothelial Cells; Glucose; Humans; Inflammation; Oxidative Stress; Phytoestrogens; Rats; Retina; Retinal Vessels; Sirtuin 1

In the present study, we investigated the anti-catabolic effects of coumestrol, a phytoestrogen derived from herbal plants, against interleukin-1β-induced cartilage degeneration in primary rat chondrocytes and articular cartilage. Coumestrol did not affect the viability of human normal oral keratinocytes and primary rat chondrocytes treated for 24 h and 21 days, respectively. Although coumestrol did not significantly increase the proteoglycan contents in long-term culture, it abolished the interleukin-1β-induced loss of proteoglycans in primary rat chondrocytes and knee articular cartilage. Furthermore, coumestrol suppressed the expression of matrix-degrading enzymes such as matrix metalloproteinase-13, -3, and -1 in primary rat chondrocytes stimulated with interleukin-1β. Moreover, the expression of catabolic factors such as nitric oxide synthase, cyclooxygenase-2, prostaglandin E

Topics: Animals; Cells, Cultured; Chondrocytes; Coumestrol; Humans; Inflammation; Interleukin-1beta; Matrix Metalloproteinases; Metabolism; Osteoarthritis; Phytoestrogens; Rats

Coumestrol, a phytoestrogen found in alfalfa, clover, and beans, has nM affinity for both estrogen receptor-α [ERα] and ERβ. Recently, a novel activity of coumestrol was reported: coumestrol binding to human ERβ represses microglia-mediated inflammation, which is associated with various neurodegenerative diseases, such as multiple sclerosis. In contrast, estradiol binding to ERβ had little or no effect on repression of microglia-mediated inflammation. Coumestrol and estradiol have several structural differences, which suggest that each ligand could induce different conformations in ERβ and, thus, different transcriptional responses in brain microglia. To begin to understand how coumestrol binds to ERβ and ERα, we constructed 3D models of coumestrol with human ERβ and ERα, which were compared to the structures of these ERs with estradiol. Of four possible orientations of coumestrol in ERα and ERβ, one orientation had the most favorable contacts with both ERs. Other phytochemicals may activate ERβ and inhibit inflammation in brain microglia and be useful therapeutics for inflammatory conditions in the brain.

Topics: Binding Sites; Coumestrol; Estrogen Receptor alpha; Estrogen Receptor beta; Humans; Imaging, Three-Dimensional; Inflammation; Models, Molecular; Molecular Conformation; Stereoisomerism; Structure-Activity Relationship

This protocol describes microsphere-based protease assays for use in flow cytometry and high-throughput screening. This platform measures a loss of fluorescence from the surface of a microsphere due to the cleavage of an attached fluorescent protease substrate by a suitable protease enzyme. The assay format can be adapted to any site or protein-specific protease of interest and results can be measured in both real time and as endpoint fluorescence assays on a flow cytometer. Endpoint assays are easily adapted to microplate format for flow cytometry high-throughput analysis and inhibitor screening.

Topics: Animals; Biotinylation; Flow Cytometry; Fluorescence Resonance Energy Transfer; Green Fluorescent Proteins; High-Throughput Screening Assays; Humans; Inflammation; Kinetics; Microspheres; Peptide Hydrolases; Peptides; Reproducibility of Results; Temperature