Mechanisms of Coreopsis tinctoria Nutt in the treatment of diabetic nephropathy based on network pharmacyology analysis of its active ingredients / Mecanismos de Coreopsis tinctoria Nutt. En el tratamiento de la nefropatía diabética basado en el análisis farmacológico de red de sus principios activos

SUMMARY: Coreopsis tinctoria Nutt. (C. tinctoria Nutt.) can protect diabetic kidneys, but the mechanisms are unclear. This work is to investigate the potential mechanisms of C. tinctoria Nutt. in the treatment of diabetic nephropathy based on network pharmacology analysis of its active ingredients. Twelve small molecular compounds of C. tinctoria Nutt. and targets related to diabetic nephropathy were docked by Discovery Studio 3.0. DAVID database was used for GO enrichment and KEGG pathway analysis. Cytoscape 3.6.1 was used to construct active ingredient-target network. Cell viability was detected with MTT. Glucose consumption was analyzed with glucose oxidase method. Protein expression was measured with Western blot and immunofluorescence. Electron microscopy observed autophagosomes. The core active ingredients of C. tinctoria Nutt. included heriguard, flavanomarein, maritimein, and marein. Twenty-one core targets of the 43 potential targets were PYGM, TLR2, RAF1, PRKAA2, GPR119, INS, CSF2, TNF, IAPP, AKR1B1, GSK3B, SYK, NFKB2, ESR2, CDK2, FGFR1, HTRA1, AMY2A, CAMK4, GCK, and ABL2. These 21 core targets were significantly enriched in 50 signaling pathways. Thirty- four signaling pathways were closely related to diabetic nephropathy, of which the top pathways were PI3K/AKT, insulin, and mTOR, and insulin resistance. The enriched GO terms included biological processes of protein phosphorylation, and the positive regulation of PI3K signaling and cytokine secretion; cellular components of cytosol, extracellular region, and extracellular space; and molecular function of protein kinase activity, ATP binding, and non-membrane spanning protein tyrosine kinase activity. In vitro experiments found that marein increased the expression of phosphorylated AKT/AKT in human renal glomerular endothelial cells of an insulin resistance model induced by high glucose, as well as increased and decreased, respectively, the levels of the microtubule-associated proteins, LC3 and P62. C. tinctoria Nutt. has many active ingredients, with main ingredients of heriguard, flavanomarein, maritimein, and marein, and may exert anti-diabetic nephropathy effect through various signaling pathways and targets.

RESUMEN: Coreopsis tinctoria Nutt. (C. tinctoria Nutt.) puede proteger riñones diabéticos, sin embargo los mecanismos son desconocidos. Este trabajo se realizó para investigar los potenciales mecanismos de C. tinctoria Nutt. en el tratamiento de la nefropatía diabética basado en el análisis de farmacología en red de sus principios activos. Doce compuestos moleculares pequeños de C. tinctoria Nutt. y los objetivos relacionados con la nefropatía diabética fueron acoplados por Discovery Studio 3.0. La base de datos DAVID se utilizó para el enriquecimiento GO y el análisis de la vía KEGG. Se usó Cytoscape 3.6.1 para construir una red de ingrediente-objetivo activa. La viabili- dad celular se detectó mediante MTT. El consumo de glucosa se analizó con el método de glucosa oxidasa. La expresión proteica fue determinada mediante Western blot e inmunofluorescencia. En la microscopía electrónica se observó autofagosomas. Los principales ingredientes activos de C. tinctoria Nutt. incluyeron heriguard, flavanomarein, maritimin y marein. Veintiún de los 43 objetivos potenciales fueron PYGM, TLR2, RAF1, PRKAA2, GPR119, INS, CSF2, TNF, IAPP, AKR1B1, GSK3B, SYK, NFKB2, ESR2, CDK2, FGFR1, HTRA1, AMY2A, CAMK4, GCK y ABL2. Estos 21 objetivos principales se enriquecieron significativamente en 50 vías de señalización. Treinta y cuatro vías de señalización estuvieron estrechamente relacionadas con la nefropatía diabética, de las cuales las principales vías fueron PI3K/ AKT, insulina y mTOR, y resistencia a la insulina. Los términos GO enriquecidos incluyeron procesos biológicos de fosforilación proteica, la regulación positiva de la señalización de PI3K y la secreción de citoquinas; componentes celulares del citosol, región extracelular y espacio extracelular; y la función molecular de la actividad de la proteína quinasa, la unión de ATP y la actividad de la proteína tirosina quinasa que no se extiende por la membrana. Los experimentos in vitro encontraron que la mareína aumentaba la expresión de AKT/AKT fosforilada en células endoteliales glomerulares renales humanas en un modelo de resistencia a la insulina inducida por niveles elevados de glucosa, así como aumentaron y disminuyeron respectivamente, los niveles de las proteínas asociadas a los microtúbulos, LC3 y P62. C. tinctoria Nutt. tiene muchos principios activos, con ingredientes principales de heriguard, flavanomarein, maritimain y marein, y puede ejercer un efecto de nefropatía antidiabética a través de distintass vías de señalización y objetivos.

Marein regulates insulin resistance in diabetic nephropathy mice by inducing autophagy / Marein regula la resistencia a la insulina en ratones con nefropatía diabética induciendo la autofagia

SUMMARY: Marein is the main active substance of Coreopsis tinctoria nutt. It not only has anti-oxidation and anti-tumor effects, but also can lower blood lipid, prevent high blood glucose, improve insulin resistance, inhibit gluconeogenesis and promote glycogen synthesis. However, the exact mechanism of its action is still unclear. Here, we explored the effect and mechanism of Marein on insulin resistance. The mice were divided into db/m, db/db, metformin+db/db, and marein+db/db groups. The body weight and kidney weight were recorded. Serum biochemical and renal function tests were measured after 8 weeks of continuous administration. Kidney tissues were subjected to HE staining, PAS staining, and Masson staining. The effect of marein on PI3K/Akt signal and autophagy pathway was detected by Western blot. After 8 weeks of Marein intervention, the body weight and kidney weight of mice did not change significantly, but the fasting blood glucose and blood lipid levels were significantly reduced than db/db group. Marein significantly improved the insulin resistance index, increased serum adiponectin and improved glucose and lipid metabolism disorders of db/db mice. Moreover, marein improved the basement membrane thickness of glomeruli and tubules, improved glomerular sclerosis and tubular fibrosis, as well as renal insufficiency, thereby protecting kidney function and delaying the pathological damage. Furthermore, marein increased the expression of PI3K and the phosphorylation of Akt/Akt (Ser473), and promoted the expression of LC3II/I, Beclin1 and ATG5. Additionally, it promoted the expression of FGFR1 in the kidney of db/db mice, and promoted the increase of serum FGF21 and FGF23. Marein has a protective effect on the kidneys of diabetic mice. It protects diabetic nephropathy by regulating the IRS1/PI3K/Akt signaling pathway to improve insulin resistance. Therefore, marein may be an insulin sensitizer.

RESUMEN: Marein es la principal sustancia activa de Coreopsis tinctoria nutt. No solo tiene efectos antioxidantes y antitumorales, sino que también puede reducir los lípidos en sangre, prevenir la glucemia alta, mejorar la resistencia a la insulina, inhibir la gluconeogénesis y promover la síntesis de glucógeno. Sin embargo, el mecanismo exacto de su acción aún no está claro. Se analizó el efecto y el mecanismo de Marein sobre la resistencia a la insulina. Los ratones se dividieron en grupos db / m, db / db, metformina + db / db y mareína + db / db. Se registró el peso corporal y el peso de los riñones. Se midieron las pruebas de función renal y bioquímica sérica después de 8 semanas de administración continua. Los tejidos renales se sometieron a tinción HE, tinción PAS y tinción Masson. El efecto de la mareína sobre la señal de PI3K / Akt y la vía de autofagia se detectó mediante Western blot. Al término de 8 semanas de tratamiento con mareína, el peso corporal y el peso de los riñones de los ratones no cambiaron significativamente, pero los niveles de glucosa en sangre y lípidos en sangre en ayunas se redujeron significativamente en relación a los del grupo db / db. Marein mejoró significativamente el índice de resistencia a la insulina, aumentó la adiponectina sérica y mejoró los trastornos del metabolismo de la glucosa y los lípidos de los ratones db / db. Además, la mareína mejoró el grosor de la membrana basal de los glomérulos y túbulos, mejoró la esclerosis glomerular y la fibrosis tubular, así como la insuficiencia renal, protegiendo la función renal y retrasando el daño patológico. Además, la mareína aumentó la expresión de PI3K y la fosforilación de Akt / Akt (Ser473), y promovió la expresión de LC3II / I, Beclin1 y ATG5. Además, promovió la expresión de FGFR1 en el riñón de ratones db / db y el aumento de FGF21 y FGF23 en suero. Marein tiene un efecto protector sobre los riñones de ratones diabéticos. Protege la nefropatía diabética regulando la vía de señalización IRS1 / PI3K / Akt para mejorar la resistencia a la insulina. Por tanto, la mareína puede ser un sensibilizador a la insulina.

Marein inhibited oxidative stress and inflammation of high glucose induced HBZY-1 cells via AMPK signaling pathway / 中国药理学通报

Aim To study the role of marein mediated AMPK signaling pathway in delaying oxidative stress, inflammation and fibrotic protein expression in diabetic nephropathy ( DN ). Methods In vitro diabetic nephropathy model was established by HG + PA induced rat mesangial cells ( HBZY-1 ) , and the cultured HBZY-1 cells were divided into normal control group (NG), HG + PA( GlucoselOO mmol • L'1 + Palmitic acid 250 [imo\ • L'1, HG +PA) model group, HG + PA + marein with different doses of 25 p,mol • L"1, 50 p,mol • L"1, 100 jimol • L"1, and 200 junol • L"1 groups. MTS was used to detect the effect of marein on HBZY-1 cell proliferation, and the optimal concentration was selected. Western blot was used to test the protein expression of NOX4, TGF-fU, MCP-1, a-SMA, FN, Collagen VI. Adenosine monophosphate activated( AMPK) protein kinase family of AMPK7I, p-AMPK a expression were measured. Results Marein inhibited high glucose palmitate-induced proliferation of HBZY-1 cells, down-regulated NOX4, TGF-(31, MCP-1, cx-SMA, FN and Collagen VI expression in model cells. Meanwhile, marein up-regulated both AMPK 7I and p-AMPKa expression. Conclusions Marein may inhibit the HBZY-1 cell proliferation, oxidative stress, inflammation and fibrosis factors expression in HG + PA induced HBZY-1 cell by activating of both AMPK 7I and AMPK signaling pathway, thus delaying renal injury in diabetic nephropathy.

Inhibition of metabolic disorders in vivo and in vitro by main constituent of Coreopsis tinctoria / 中草药·英文版

Objective: To investigate the effects of the ethyl acetate extract of Coreopsis tinctoria (EAEC) on insulin resistance (IR) in rats fed a high-fat diet. Methods: Male Sprague-Dawley (SD) rats were fed a HFD (60% fat) supplemented with EAEC for 8 weeks. The administration of EAEC to the rats with HFD-induced insulin resistance reduced hyperglycemia, plasma levels of insulin, and steatosis in the liver. Metabolomic study was used to analyze the metabolic levels of the high glucose-treated cells, control cells and marein-treated cells. Results: High glucose and high fat conditions caused a significant increase in blood glucose, insulin, serum TC, TG and LDL-C levels, leading to abnormal IR in rats. However, treatment with EAEC protects against HFD-induced IR by improving the fasting serum glucose homeostasis and lipid homeostasis. The high glucose conditions significantly decreased glycogen synthesis and increased PEPCK, G6Pase and Krebs cycle-related enzyme protein levels, leading to an abnormal metabolic state in HepG2 cells. However, treatment with marein improved IR by increasing glucose uptake and glycogen synthesis and by downregulating PEPCK and G6Pase protein levels. The statistical analysis of the HPLC/MS data demonstrated that marein could restore the normal metabolic state. Conclusion: The results revealed that EAEC ameliorates IR in rats, and marein has the potential to improve IR by ameliorating glucose metabolism disorders.