PMID: 

Cancer Lett. 2018 09 1 ;431:123-141. Epub 2018 May 29. PMID: 29857127

Abstract Title: 

Formononetin-induced oxidative stress abrogates the activation of STAT3/5 signaling axis and suppresses the tumor growth in multiple myeloma preclinical model.

Abstract: 

Aberrant reactions of signal transducer and transcriptional activator (STAT) are frequently detected in multiple myeloma (MM) cancers and can upregulate the expression of multiple genes related to cell proliferation, survival, metastasis, and angiogenesis. Therefore, agents capable of inhibiting STAT activation can form the basis of novel therapies for MM patients. In the present study, we investigated whether the potential anti-cancer effects of Formononetin (FT), a naturally occurring isoflavone derived from Astragalus membranaceus, Trifolium pratense, Glycyrrhiza glabra, and Pueraria lobata, against MM cell lines and human multiple myeloma xenograft tumors in athymic nu/nu mice model are mediated through the negative regulation of STAT3 and STAT5 pathways. Data from the in vitro studies indicated that FT could significantly inhibit cell viability, and induce apoptosis. Interestingly, FT also suppressed constitutive STAT3 (tyrosine residue 705 and serine residue 727) and STAT5 (tyrosine residue 694/699) activation, which correlated with the suppression of the upstream kinases (JAK1, JAK2, and c-Src) in MM cells, and this effect was found to be mediated via an increased production of reactive oxygen species (ROS) due to GSH/GSSG imbalance. Also, FT abrogated STAT3 and STAT5 DNA binding capacity and nuclear translocation. FT induced cell cycle arrest, downregulated the expression of STAT3-regulated anti-apoptotic, angiogenetic, and proliferative gene products; and this correlated with induction of caspase-3 activation and cleavage of PARP. Intraperitoneal administration of FT significantly suppressed the tumor growth in the multiple myeloma xenograft mouse model without exhibiting any significant adverse effects. Overall, our findings indicate that FT exhibits significant anti-cancer effects in MM that may be primarily mediated through the ROS-regulated inhibition of the STAT3 and STAT5 signaling cascade.

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PMID: 

Horm Metab Res. 2014 Oct ;46(11):753-60. Epub 2014 Jun 30. PMID: 24977660

Abstract Title: 

Formononetin inhibits migration and invasion of MDA-MB-231 and 4T1 breast cancer cells by suppressing MMP-2 and MMP-9 through PI3K/AKT signaling pathways.

Abstract: 

Formononetin is a naturally existing isoflavone, which can be found in the roots of Astragalus membranaceus, Trifolium pratense, Glycyrrhiza glabra, and Pueraria lobata. It was found to be associated with inhibition of cell proliferation and cell cycle progression, as well as induction of apoptosis in various cancer cell lines. However, the effect of formononetin on breast cancer cell metastasis remains unclear. In this study, we examined the effect of formononetin on the migration and invasion of breast cancer cells MDA-MB-231 and 4T1 in vitro and in vivo. Our data demonstrated that formononetin did not effectively inhibit the cell viability of MDA-MB-231 and 4T1 in 24 h with the concentration lower than 160 μmol/l. When treated with nontoxic concentration of formononetin, the migration and invasion of MDA-MB-231 and 4T1 cells were markedly suppressed by wound healing assay, chamber invasion assay, and in vivo mouse metastasis model. In vitro, formononetin reduced the expression of matrix metalloproteinase-2 (MMP-2), MMP-9 and increased the expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2. Furthermore, the immunofluorescence and immunoblotting assays indicated that formononetin was very effective in suppressing the phosphorylation of Akt and PI3K. Collectively, these results suggest that formononetin inhibited breast cancer cell migration and invasion by reducing the expression of MMP-2 and MMP-9 through the PI3K/AKT signaling pathway. These findings demonstrate a potentially new therapeutic strategy of formononetin as anti-invasive agent for breast cancer.

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PMID: 

PLoS One. 2014 ;9(3):e91019. Epub 2014 Mar 10. PMID: 24614080

Abstract Title: 

Neuroprotective effects of sevoflurane against electromagnetic pulse-induced brain injury through inhibition of neuronal oxidative stress and apoptosis.

Abstract: 

Electromagnetic pulse (EMP) causes central nervous system damage and neurobehavioral disorders, and sevoflurane protects the brain from ischemic injury. We investigated the effects of sevoflurane on EMP-induced brain injury. Rats were exposed to EMP and immediately treated with sevoflurane. The protective effects of sevoflurane were assessed by Nissl staining, Fluoro-Jade C staining and electron microscopy. The neurobehavioral effects were assessed using the open-field test and the Morris water maze. Finally, primary cerebral cortical neurons were exposed to EMP and incubated with different concentration of sevoflurane. The cellular viability, lactate dehydrogenase (LDH) release, superoxide dismutase (SOD) activity and malondialdehyde (MDA) level were assayed. TUNEL staining was performed, and the expression of apoptotic markers was determined. The cerebral cortexes of EMP-exposed rats presented neuronal abnormalities. Sevoflurane alleviated these effects, as well as the learning and memory deficits caused by EMP exposure. In vitro, cell viability was reduced and LDH release was increased after EMP exposure; treatment with sevoflurane ameliorated these effects. Additionally, sevoflurane increased SOD activity, decreased MDA levels and alleviated neuronal apoptosis by regulating the expression of cleaved caspase-3, Bax and Bcl-2. These findings demonstrate that Sevoflurane conferred neuroprotective effects against EMP radiation-induced brain damage by inhibiting neuronal oxidative stress and apoptosis.

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PMID: 

J Sep Sci. 2019 Mar ;42(6):1133-1143. Epub 2019 Feb 25. PMID: 30620132

Abstract Title: 

Screening and isolation of cyclooxygenase-2 inhibitors from Trifolium pratense L. via ultrafiltration, enzyme-immobilized magnetic beads, semi-preparative high-performance liquid chromatography and high-speed counter-current chromatography.

Abstract: 

Nonsteroidal anti-inflammatory drugs reportedly reduce the risk of developing cancer. One mechanism by which they reduce carcinogenesis involves the inhibition of the activity of cyclooxygenase-2, an enzyme that is overexpressed in various cancer tissues. Its overexpression increases cell proliferation and inhibits apoptosis. However, selected cyclooxygenase-2 inhibitors can also act through cyclooxygenase-independent mechanisms. In this study, using ultrafiltration, enzyme-immobilized magnetic beads, high-performance liquid chromatography, and electrospray-ionization mass spectrometry, several isoflavonoids in Trifolium pratense L. extracts were screened and identified. Semi-preparative high-performance liquid chromatography and high-speed counter-current chromatography were then applied to separate the active constituents. Using these methods, seven major compounds were identified in Trifolium pratense L. As cyclooxygenase-2 inhibitors: rothindin, ononin, daidzein, trifoside, pseudobaptigenin, formononetin, and biochanin A, which were then isolated with>92% purity. This is the first report of the presence of potent cyclooxygenase-2 inhibitors in Trifolium pratense L. extracts. The results of this study demonstrate that the systematic isolation of bioactive components from Trifolium pratense L., by using ultrafiltration, enzyme-immobilized magnetic beads, semi-preparative high-performance liquid chromatography, and high-speed counter-current chromatography, represents a feasible and efficient technique that could be extended for the identification and isolation of other enzyme inhibitors.

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PMID: 

Cell Biochem Biophys. 2011 Mar ;59(2):71-7. PMID: 20824388

Abstract Title: 

Oxidative stress and apoptosis in relation to exposure to magnetic field.

Abstract: 

We investigated the effect of extremely low-frequency electromagnetic field (ELF-EMF) with pulse trains exposure on lipid peroxidation, and, hence, oxidative stress in the rat liver tissue. The parameters that we measured were the levels of plasma alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase as well as plasma albumin, bilirubin, and total protein levels in 30 adult male Wistar rats exposed to ELF. We also determined the percentage of apoptotic and necrotic cells of the kidney extracts from the animals by flow cytometry method. Apoptotic cell death was further characterized by monitoring DNA degradation using gel electrophoresis. The results showed an increase in the levels of oxidative stress indicators, and the flow cytometric data suggested a possible relationship between the exposure to magnetic field and the cell death. We showed significantly lower necrotic cell percentages in experimental animals compared to either unexposed or sham control groups. However, DNA ladder analyses did not differentiate between the groups. Our results were discussed in relation to the response of biological systems to EMF.

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PMID: 

Ecotoxicol Environ Saf. 2018 Jun 15 ;154:137-144. Epub 2018 Feb 22. PMID: 29459163

Abstract Title: 

Se enhanced phytoremediation of diesel in soil by Trifolium repens.

Abstract: 

A pot-culture experiment was conducted to assess the effects of selenium (Se) (0.5 mg kg) on Trifolium repens exposed to various levels of diesel (0, 15, 20, 25 g kg) for 30 days and 60 days. Exposure to diesel for 60 day led to concentration-dependent decreases in root morphogenesis, chlorophyll content and CAT activity, and to dose-dependent increases in MDA content and SOD activity. The residual diesel concentration in soil increased and the removal efficiency decreased with soil diesel concentration. The chlorophyll content and residual diesel concentration after were slightly higher at 30 days than at 60days. Application of Se to soil increased Trifolium repens tolerance to diesel and significantly increased the phytoremediation effect at 60 days, with a removal rate of 36 ± 8%, compared to 28 ± 7% in the control. These results contribute to the ongoing effort to develop an effective phytoremediation system for soils highly contaminated by diesel.

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PMID: 

Cell J. 2018 Oct ;20(3):412-421. Epub 2018 May 15. PMID: 29845796

Abstract Title: 

Evaluation of Hydro-Alcoholic Extract of Trifolium Pratens L. for Its Anti-Cancer Potential on U87MG Cell Line.

Abstract: 

Objective: Glioblastoma multiforme is the most malignant form of brain tumors. Trifolium pratense L. has been suggested for cancer treatment in traditional medicine. Here we have investigated the effects of T. pratense extract on glioblastoma multiforme cell line (U87MG).Materials and Methods: In this experimental study, the effect of T. pratense extract on cell viability was investigated using trypan blue staining, MTT assay, and lactate dehydrogenase activity measurement. Apoptosis and autophagy cell death were detected by fluorescent staining. Nitric oxide (No) production was measured using Griess reaction. Expression levels of some apoptotic and autophagic-related genes were detected using real-time polymerase chain reaction (PCR). The combination effects of T. pratense extract and temozolomide (TMZ) were evaluated by calculating the combination index and dose reduction index values.Results: After treatment with T. pratense extract, the cell viability was significantly reduced in a time- and dosedependent manner (P

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PMID: 

Food Funct. 2013 Aug ;4(8):1252-62. PMID: 23764910

Abstract Title: 

The preventive effect of lotus seedpod procyanidins on cognitive impairment and oxidative damage induced by extremely low frequency electromagnetic field exposure.

Abstract: 

The present study investigated the effects of lotus seedpod procyanidins (LSPCs) administered by oral gavage on the cognitive deficits and oxidative damage of mice at extremely low frequency electromagnetic field (ELF-EMF) exposure (50 Hz, 8 mT, 28 days). The results showed that 90 mg kg⁻¹ LSPCs treatment significantly increased body weight compared with the ELF-EMF group at ELF-EMF exposure and effectively maintained liver index, thymus index, kidney index and spleen index close to normal. A water maze test indicated that learning and memory abilities of the ELF-EMF group deteriorated significantly with ELF-EMF exposure when compared with the control group, but the ELF-EMF + LSPCs90 group had remarkably improved learning and memory abilities compared with the ELF-EMF group. Malondialdehyde (MDA), reactive oxygen species (ROS), nitric oxide (NO) and nitric oxide synthase (NOS) mostly exhibited significant increases, while the activities of glutathione peroxidase (GPx), catalase (CAT) and superoxide dismutase (SOD) decreased significantly under ELF-EMF exposure in the ELF-EMF group. LSPCs (especially 60, 90 mg kg⁻¹) administration decreased MDA, ROS, NO content andlowered NOS activity in LSPCs treatment groups. Furthermore, LSPCs (60, 90 mg kg⁻¹) treatment significantly augmented GPx, CAT, SOD activity in the hippocampus and serum. Pathological observation showed that number of pyramidal cells of the CA1 and CA3 regions of the hippocampus of the LSPCs treatment groups was significantly greater than the ELF-EMF group. All the data suggested that the LSPCs can effectively prevent learning and memory damage and oxidative damage caused by the ELF-EMF, most likely through the ability of LSPCs to scavenge oxygen free radicals and to stimulate antioxidantenzyme activity.

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PMID: 

Res Pharm Sci. 2018 Aug ;13(4):324-331. PMID: 30065765

Abstract Title: 

Protective effect of hydroalcoholic extracts of. on pancreaticβ cell line (RIN-5F) against cytotoxicty of streptozotocin.

Abstract: 

Natural plants have traditionally been used throughout the world for their anti-diabetic effects. The aim of the present study was to investigate the protective effect of hydroalcoholic extract of. () on streptozotocin (STZ) cytotoxicity and insulin concentration from RIN-5F pancreaticβ cell line. In this study, possible cytoprotective action ofextract (using pre-treatment, simultaneous, and post-treatment schedules) against STZ (30 mM) was evaluated using MTT assay. Apoptosis was quantified by fluorescent dye staining. Also, the effect of extract on insulin secretion in low and high glucose media was examined. Data were analyzed by one-way ANOVA test and

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PMID: 

Environ Sci Pollut Res Int. 2017 Feb ;24(5):5078-5088. Epub 2016 Dec 21. PMID: 28004364

Abstract Title: 

Cytotoxic effects of moderate static magnetic field exposure on human periphery blood mononuclear cells are influenced by Val16Ala-MnSOD gene polymorphism.

Abstract: 

Technological advancement has increasingly exposed humans to magnetic fields (MFs). However, more insights are necessary into the potential toxicity of MF exposure as a result of genetic variations related to oxidative metabolism. Therefore, the following study has assessed an in vitro cytotoxic effect of static magnetic field (SMF) (5 mT) on cells with Val16Ala polymorphism (AA, VA, and VV) in the manganese superoxide dismutase gene. Homozygous Val16Ala-superoxide dismutase 2 (SOD2) genotypes present oxidative imbalance that is associated with risk to several chronic degenerative diseases (VV produces less efficient and AA moreefficient SOD2 enzyme). Blood samples from healthy adult subject carriers with different Val16Ala-SOD2 genotypes were obtained and exposed to MF at different times (0, 1, 3, 6 h). The cytotoxic effect as well as oxidative stress was evaluated after incubation of 24 h at 37 °C. In addition, apoptosis induction has been analyzed by flow cytometry as well as Bcl-2-associated X protein (BAX), B-cell lymphoma 2 (BCL-2), and caspases 8 and 3 gene expression. SMF cytotoxic effect has been observed in AA cells at all times of exposure, whereas AV cells presented higher mortality only after 6 h of exposure at SMF. Higher apoptosis induction has been observed in AA cells when compared to VV and AV cells. These results suggest a toxicogenetic SMF effect related to an imbalance in SOD2 activity.

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