PMID: 

Mol Med Rep. 2018 Jun ;17(6):7721-7729. Epub 2018 Apr 5. PMID: 29620230

Abstract Title: 

Formononetin inhibits colon carcinoma cell growth and invasion by microRNA‑149‑mediated EphB3 downregulation and inhibition of PI3K/AKT and STAT3 signaling pathways.

Abstract: 

Formononetin (Form), a phytoestrogen extracted from the roots of Astragalus membranaceus, is one of the fundamental herbs used in traditional Chinese medicine because of its protective effects against certain malignant tumors. However, its role in colon carcinoma cells and the underlying molecular mechanisms have not been completely elucidated. The present study aimed todemonstrate that Form significantly inhibited the proliferation and invasion of the colon carcinoma cell lines SW1116 and HCT116. Mechanistic studies have suggested that Form suppresses colon carcinoma cell growth by downregulating cell cycle‑associated protein (cyclin D1) expression and arresting the cell cycle at the G0‑G1 checkpoint. Further studies revealed that treatment with Form inhibits matrix metalloproteinase (MMP)2 and MMP9 expression. Aditionally, the results demonstrated that Form significantly increased microRNA (miR)‑149 expression. Following miR‑149 overexpression in SW1116 and HCT116 cells using an miR‑149 mimic, cell viability and Ephrin type‑B receptor 3 (EphB3) levels decreased. Furthermore, the inhibitory effects of Form were associated with phosphatidylinositol 3‑kinase (PI3K)/protein kinase B (AKT) and signal transducer and activator of transcription3 (STAT3) signaling pathways. These results indicated the suppressive effect of Form on colon carcinoma cell proliferation and invasion, possibly via miR‑149‑induced EphB3 downregulation and the inhibition of the PI3K/AKT and STAT3 signaling pathways. Overall, Form may be used as a novel candidate for the clinical treatment of colorectal cancer in the future.

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PMID: 

J Ethnopharmacol. 2018 Jul 15 ;221:91-99. Epub 2018 Apr 13. PMID: 29660466

Abstract Title: 

Formononetin, an isoflavone from Astragalus membranaceus inhibits proliferation and metastasis of ovarian cancer cells.

Abstract: 

ETHNOPHARMACOLOGICAL RELEVANCE: Astragalus membranaceus which was originally described in the Shennong's Classic of Materia Medica, the earliest complete Pharmacopoeia of China written from the Warring States Period to Han Dynasty, has been widely used in Chinese medicine for> 2000 years, especially in the prescription of curing cancer. A. membranaceus has various bioactivities, such as anti-tumor, anti-viral, anti-oxidant, anti-diabetes, anti-inflammation, anti-atherosclerosis, immunomodulation, hepatoprotection, hematopoiesis, neuroprotection and so on. As an important component of A. membranaceus, whether formononetin has a close relationship with its tumor-inhibiting effect on ovarian cancer cell has been investigated.AIM OF STUDY: The present study aimed to demonstrate the anti-proliferation, anti- migration and invasion effects of formononetin on ovarian cancer cells and further explore the underlying molecular mechanisms associated with apoptosis, migration and invasion.MATERIALS AND METHODS: MTT assay was performed to detect the viability of ovarian cancer cells. DAPI staining, Annexin-V assay and assay for mitochondrial membrane potential detected the apoptosis of ovarian cancer cells treated by formononetin. The migration and invasion of ovarian cancer cells which exposed to formononetin were detected by scratch assay and transwell assay. Meanwhile, the protein-level changes of in ovarian cancer cells treated by formononetin were assessed by western blot analysis.RESULTS: MTT assays indicated that cell viability significantly decreased in ovarian cancer cells treated with formononetin. DAPI staining, Annexin-V assay and assay for mitochondrial membrane potential suggested that formononetin suppressed cells proliferation by inducing apoptosis. We detected the expression of apoptosis-related proteins in ovarian cancer cells after treatment with formononetin and found the expression of caspase 3/9 proteins and the ratio of Bax/Bcl-2 were increased in a dose-dependent manner. In addition, wound healing and transwell chamber assays showed that formononetin suppressed the migration and invasion of ovarian cancer cells. And formononetin decreased expression of MMP-2/9 proteins and phosphorylation level of ERK.CONCLUSIONS: The present results demonstrated that formononetin have potential effects on induction of apoptosis and suppression of migration and invasion.

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PMID: 

J Cell Biochem. 2018 09 ;119(9):7377-7387. Epub 2018 May 15. PMID: 29761845

Abstract Title: 

The O-methylated isoflavone, formononetin, inhibits human ovarian cancer cell proliferation by sub G0/G1 cell phase arrest through PI3K/AKT and ERK1/2 inactivation.

Abstract: 

Formononetin is an isoflavone that is extracted from red clovers or soy. It has anti-oxidant, anti-proliferative, and anti-tumor effects against cells in various diseases. Several cohort studies have indicated that phytoestrogen intake, including formononetin, could reduce the risk of various carcinogenesis. In fact, many case-control studies have indicated the potential value of flavonoids as drug supplements in the treatment of many cancer patients. However, the toxic effects and the anti-cancer mechanism of formononetin in ovarian cancer are unknown. We investigated the toxicological mechanism of formononetin in ES2 and OV90 ovarian cancer cells. Formononetin suppressed cell proliferation through sub G0/G1 phase arrest and increased apoptosis in both cell lines. Furthermore, it induced loss of mitochondrial membrane potential and generation of reactive oxygen species in ES2 and OV90 cells. The formononetin-mediated regulation of cell proliferation and apoptosis involved decreased phosphorylation of ERK1/2, P90RSK, AKT, P70S6K, and S6 proteins, and increased phosphorylation of P38 protein in ES2 and OV90 cells. Co-treatment of formononetin with pharmacological inhibitors (LY294002 or U0126) revealed additional anti-proliferative effects on the two human ovarian cancer cell types. Conclusively, the results indicate the potential value of formononetin as an anti-cancer agent in human ovarian cancer.

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PMID: 

Biomed Pharmacother. 2018 Oct ;106:349-354. Epub 2018 Jul 11. PMID: 29966980

Abstract Title: 

Neuroprotective effect of formononetin against TBI in rats via suppressing inflammatory reaction in cortical neurons.

Abstract: 

Traumatic brain injury (TBI) refers to external force-induced brain damage, characterized with necrosis and cell loss in cerebral cortex. Interestingly, a plant-extract named formononetin (FN) is found to possess promising pharmacological activities, including cellular neuroprotection. Thus, we propose that FN may exert biological protection against TBI and discuss the underlying mechanism. In the current study, a rat TBI model was established via Feeney's classical method, followed by different concentrations of FN treatment. Nissl-special and DAPI-labeled stains were utilized to assess the proliferation of cortical neurons nearing lesioned tissue. The contents of interleukin-6 (IL6), tumor necrosis factor (TNF-α), and interleukin-10 (IL10) in serum and the cortical neurons were determined by ELISA. Further, intracephalic IL10 expression levels were detected through immunoassay and RT-PCR. Interestingly, the results exhibited within the FN-treated TBI rat model indicated elevated cortical proliferation. The levels of IL10 in serum and the cortical neurons were increased following FN treatments, while TNF-α and IL6 levels in the blood were decreased. In addition, both mRNA and protein expression levels of IL10 in the FN-treated TBI rat model were up-regulated in a dose-dependent manner. Collectively, our present findings indicate that FN provides effective neuroprotection against TBI, likely by activating IL10 expression in cortical neurons nearing lesioned tissue to inhibit neuroinflammatory reaction.

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PMID: 

Front Pharmacol. 2018 ;9:739. Epub 2018 Jul 18. PMID: 30072892

Abstract Title: 

Formononetin Treatment in Type 2 Diabetic Rats Reduces Insulin Resistance and Hyperglycemia.

Abstract: 

Type 2 diabetic mellitus is a multifactorial metabolic disorder affecting huge population around the world. This indicates that there is an urgent unmet need of cost effective, new treatment strategies for type 2 diabetes mellitus with no or less side effects. Phenolic compounds including isoflavones are known for their beneficial effect in metabolic disorders. The present work was intended to find out efficacy of formononetin, an isoflavone treatment in experimental model of type 2 diabetes. Type 2 diabetes mellitus was induced by feeding high fat diet for 2 weeks prior to streptozotocin administration inrats. Diabetic animals were treated with formononetin for 28 days at three dose level, i.e., 10, 20, and 40 mg/kg body weight orally. The effect of formononetin treatment on various parameters such as plasma glucose, glucose tolerance, insulin, HOMA-IR, lipid profile, hepatic glycogen content, glycohaemoglobin and SIRT1 expression in pancreatic tissue was measured. Histopathological changes in pancreatic tissue were also studied. Results of the study demonstrate that formononetin treatment reduces blood glucose level significantly (

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PMID: 

Biomed Pharmacother. 2018 Oct ;106:1250-1257. Epub 2018 Jul 20. PMID: 30119194

Abstract Title: 

The protective effect of formononetin on cognitive impairment in streptozotocin (STZ)-induced diabetic mice.

Abstract: 

The present study was aimed to elucidate the pharmacological effect of Formononetin (FMN) treatment on STZ-induced diabetic cognitive dysfunction. The diabetic model was induced by an intraperitoneally injection of 180 mg/kg STZ. The animals were randomly divided into five groups: control group, streptozocin (STZ, 180 mg/kg) group, STZ + metformin (Met, 200 mg/kg) group, STZ + FMN (25 mg/kg) group, STZ + FMN (50 mg/kg) group. The mice were intragastrically administrated with metformin (Met,200 mg/kg) or FMN (25, 50 mg/kg) once daily for 6 weeks. The blood glucose content and body weight were examined. Morris water maze test and Y maze test were used to evaluate the learning and memory abilities. The cognitive decline was reversed by regulating superoxide dismutase (SOD), malondialdehyde (MDA), tumor necrosis factor-a (TNF-α), interleukin(IL)-1β, IL-6 in serum and hippocampus. The protein expressions of high mobility group box-1 protein (HMGB1), toll like receptor 4 (TLR4), myeloid differentiating factor 88 (MyD88), inhibitor of NF-κB (IκBα), p-IκBα, nuclear factor kappa-B(NF-κB), p-NF-κB, NOD-like receptor 3(NLRP3), apoptosis-associated speck-like protein containing CARD(ASC) and caspase-1 were detected. Furthermore, the SH-SY5Y cells were exposed to high glucose stimulation, FMN (2.5, 5 and 10 μM) treatment, and glycyrrhizin, the selective inhibitor of HMGB1. After an incubation for 22 h, the SH-SY5Y cells were harvested for detection. As a result, FMN treatment effectively attenuated the body weight, learning and memory abilities, as well as the levels of blood glucose, SOD, MDA, TNF-α, IL-1β, IL-6. FMN administration also downregulated the protein expressions of HMGB1, TLR4, MyD88, p-IκB, p-NF-κB, NLRP3, ASC and caspase-1. The inhibition of HMGB1 by glycyrrhizin also confirmed the involvement of HMGB1/TLR4/NF-κB/NLRP3 pathway in high glucose-induced SH-SY5Y cells. In summary, the results suggested that FMN exhibited the protective effect on STZ-induced cognitive impairment possibly via the mediation of HMGB1/TLR4/NF-κB signaling and NLRP3 inflammasome.

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PMID: 

Mol Med Rep. 2018 Dec ;18(6):4821-4830. Epub 2018 Oct 10. PMID: 30320398

Abstract Title: 

Formononetin may protect aged hearts from ischemia/reperfusion damage by enhancing autophagic degradation.

Abstract: 

Myocardial infarction is a leading cause of mortality worldwide, and timely blood/oxygen reperfusion may substantially improve the outcome of infarction. However, ischemia/reperfusion (I/R) may cause severe side effects through excess reactive oxygen species generation. To develop novel methods to relieve I/R induced cell damage, the present study used a component of traditional Chinese medicine. In the present study, isolated heart tissue from aged mice and H9C2 cells with chemically‑induced aging were used as experimental subjects, and it was demonstrated that formononetin was able to alleviate I/R‑induced cell or tissue apoptosis. By applying formononetin to I/R‑damaged tissue or cells, it was demonstrated that formononetin was able to enhance autophagy and thus alleviate I/R‑induced cell damage. Furthermore, it was observed that I/R was able to inhibit lysosomal degradation processes in aged tissues or cells by impairing the lysosome acidification level, and formononetin was able to reverse this process via the re‑acidification of lysosomes. In conclusion, the present study demonstrated that formononetin was able to alleviate I/R‑induced cellular apoptosis in aged cells by facilitating autophagy.

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PMID: 

Drug Des Devel Ther. 2018 ;12:3675-3684. Epub 2018 Nov 1. PMID: 30464399

Abstract Title: 

Formononetin induces vasorelaxation in rat thoracic aorta via regulation of the PI3K/PTEN/Akt signaling pathway.

Abstract: 

Background: Formononetin (FMN) is an isoflavone that produces arterial vasodilation. However, the underlying molecular mechanisms are unclear.Purpose: The purpose of this study was to explore the vasorelaxant effect and the potential mechanism of FMN in vascular endothelium in isolated rat aorta.Methods: The thoracic aortas of Sprague Dawley rats were isolated to test the arterial reactivity in the presence of FMN with or without inhibitors. Bioinformatics analyses, including a Bioinformatics Analysis Tool for Molecular Mechanism of Traditional Chinese Medicine and molecular docking methods, were performed to predict therapeutic targets responsible for the vascular protection produced by FMN. We used rat aortic endothelial cells (RAOECs) as an in vitro model to verify the potential mechanism through molecular biological analyses. The production of nitric oxide (NO) metabolites were evaluated via an NO assay kit according to the manufacturer's instruction. The mRNA expression of eNOS was analyzed by polymerase chain reaction, and the protein levels of PTEN, phosphorylated Akt, and eNOS were measured by Western blot.Results: We found that FMN dilated rat aortic rings in a concentration-dependent manner, which was reduced by endothelium denudation and eNOS inhibition. The bioinformatics analyses indicated that FMN activity was associated with the PI3K/PTEN/Akt signaling pathway. Molecular biological studies demonstrated that FMN significantly elevated the levels of NO and eNOS mRNA and markedly increased the protein expression of phosphorylated Akt and eNOS in RAOECs, and decreased PTEN compared with a dimethyl sulfoxide group.Conclusion: FMN performs vasorelaxation of the thoracic aorta through activating the PI3K/PTEN/Akt signaling pathway.

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PMID: 

Biomed Pharmacother. 2019 Jan ;109:2084-2089. Epub 2018 Nov 26. PMID: 30551465

Abstract Title: 

Formononetin and metformin act synergistically to inhibit growth of MCF-7 breast cancer cells in vitro.

Abstract: 

Many breast cancer patients suffer from obvious side effects induced by chemotherapy. Formononetin (FM), one kind ingredient of Chinese herbal medicine, has been suggested to inhibit MCF-7 breast cancer cells. And recently metformin (MET) has gained more attention as a potential anti-cancer drug. The aim of this study was to investigate the synergistic effects of FM and MET on the proliferation of MCF-7 cells and to clarify the possible molecular mechanism involved. MCF-7 cells were treated with various concentrations of FM (40 and 80 μM) or FM (40 and 80 μM) combined with MET (150 μM) for 48 h. Cell proliferation was tested by an methyl tetrazolium (MTT) (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide) assay. The percentage of apoptotic cells was measured by flow cytometry. The expression level of b-cell lymphoma/leukemia-2 (bcl-2) mRNA was examined by RT-PCR, while the expression levels of phosphorylated extracellular signal-regulated kinases (p-ERK1/2) and bcl-2 protein were detected by Western blotting. Compared with untreated cells, 40 μM and 80 μM FM efficiently inhibited proliferation and increased apoptosis in MCF-7 cells. Additionally, 40 μM and 80 μM FM greatly downregulated bcl-2 mRNA expression when compared with untreated cells. Furthermore, the protein expression of bcl-2 and p-ERK1/2 was significantly reduced by 40 μM and 80 μM FM. The cytotoxic effect of FM was more remarkable when 150 μM MET was added. Taken together, the combinational use of FM and MET enhanced cell growth inhibition, and the induction of apoptosis in MCF-7 cells mediated by the ERK1/2 signaling pathway.

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PMID: 

Life Sci. 2019 Feb 15 ;219:109-121. Epub 2019 Jan 11. PMID: 30641085

Abstract Title: 

Formononetin attenuates kidney damage in type 2 diabetic rats.

Abstract: 

AIM: Diabetic nephropathy is the commonly developed complication of vasculature in type 2 diabetic patients. Chronic hyperglycemia leads to nephropathy in diabetics because of the formation of excessive reactive oxygen species and advanced glycation end products which is reflected in the form of glomerulosclerosis, tubular atrophy and interstitial fibrosis. As per the various reports reduction in SIRT1 expression in kidney tissue is key factor in the development of nephropathy in diabetes because its reduction in tissue is linked with excessive formation of ROS. Formononetin is a polyphenolic compound reported for its effect on SIRT1 and ROS.MAIN METHODS: Type 2 diabetes was induced in rats by diet modification using high fat diet for fifteen days prior to streptozotocin regimen (35 mg/kg, i.p.). Treatment of formononetin was started after confirmation of diabetes and continued for 16 weeks. Formononetin was administered orally to the diabetic animals at the dose of 10. 20 and 40 mg/kg.KEY FINDINGS: Formononetin treatment for 16 week was able to control hyperglycemia and insulin resistance in diabetic animals. It has also been reduced triglyceride and cholesterol in blood. Formononetin treatment reduced blood concentration of creatinine, blood urea nitrogen and increased albumin concentration. Formononetin treatment alsoenhanced creatinine clearance in diabetic animals. Oxidative stress burden was also reduced significantly after formononetin treatment along with increased SIRT1 expression in kidney tissues of diabetic animals.SIGNIFICANCE: Formononetin is a potential molecule which increases the expression of SIRT1 in kidney tissue of diabetic. Thus formononetin is an effective molecule to control nephropathy in type 2 diabetes mellitus.

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