PMID: 

J Cell Physiol. 2019 Aug ;234(8):12701-12713. Epub 2018 Dec 4. PMID: 30515812

Abstract Title: 

Diosmetin inhibits osteoclast formation and differentiation and prevents LPS-induced osteolysis in mice.

Abstract: 

Osteolytic bone diseases are closely linked to the over-activation of osteoclasts and enhancement of bone resorption. It has become a major health issue in orthopedic practice worldwide. Inhibition of osteoclasts is proposed to be the main treatment for osteolytic disorders. Diosmetin (DIO) is a natural flavonoid with properties of antioxidant, anti-infection, and antishock. The effect of DIO on osteoclast differentiation is poorly understood. In this study project, we found that DIO could inhibit osteoclastic formation induced by receptor activator of nuclear factor kappa-B ligand (RANKL) in a dose-dependent manner. The expression of the osteoclast differentiation marker genes, cathepsin K, nuclear factor of activated T-cells 1 (NFATc1), Acp5, Ctr, Atp6v0d2, and Mmp9 were also decreased by the treatment of DIO. In addition, DIO attenuated the formation of actin ring and the ability of bone resorption. Further, the western blotting showed that DIO inhibits the phosphorylation of the mitogen-activated protein kinases signaling pathway induced by RANKL, accompanied by the downregulation of NFATc1 and c-Fos expression. We also found that DIO could reduce the accumulation of reactive oxygen species (ROS) induced by RANKL. In vivo, the study revealed that DIO can significantly reduce LPS-induced osteolysis in mice. Collectively, our study shows that DIO can inhibit osteoclast formation and activation, and could serve as a potential therapeutic drug for osteolytic bone diseases.

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PMID: 

Br J Pharmacol. 2019 Jun ;176(12):2079-2094. Epub 2019 May 11. PMID: 30825187

Abstract Title: 

Diosmetin induces apoptosis and enhances the chemotherapeutic efficacy of paclitaxel in non-small cell lung cancer cells via Nrf2 inhibition.

Abstract: 

BACKGROUND AND PURPOSE: Non-small-cell lung cancer (NSCLC) accounts for up to 80-85% of all lung cancers and has a disappointing prognosis. Flavonoids exert anticancer properties, mostly involving stimulation of ROS production without significant toxicity to normal cells. This study was aimed to delineate the effect of diosmetin, a natural flavonoid, on NSCLC cells and its ability to enhance the antitumour activity of paclitaxel.EXPERIMENTAL APPROACH: NSCLC cells, normal cell lines HLF-1 and BEAS-2B, and immunodeficient mice were chosen as models to study the effects of diosmetin. Changes in cell viability, apoptosis, and ROS were analysed by MTT assay, flow cytometry assay, and fluorescent probe DCFH-DA. Expression of proteins and mRNA was determined by Western blotting and real-time RT-PCR. Growth of xenografted tumours was measured. Spleens and other vital organs were analysed with histological and immunohistochemical techniques.KEY RESULTS: Diosmetin induced selective apoptotic death in NSCLC cells but spared normal cells, via ROS accumulation. Diosmetin induced ROS production in NSCLC cells probably via reducing Nrf2 stability through disruption of the PI3K/Akt/GSK-3β pathway. The in vitro and in vivo xenograft studies showed that combined treatment of diosmetin and paclitaxel synergistically suppressed NSCLC cells. Histological analysis of vital organs showed no obvious toxicity of diosmetin, which matched our in vitro findings.CONCLUSIONS AND IMPLICATIONS: Diosmetin selectively induced apoptosis and enhanced the efficacy of paclitaxel in NSCLC cells via ROS accumulation through disruption of the PI3K/Akt/GSK-3β/Nrf2 pathway. Therefore, diosmetin may be a promising candidate for adjuvant treatment of NSCLC.

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PMID: 

Med Sci Monit. 2019 Mar 27 ;25:2238-2245. Epub 2019 Mar 27. PMID: 30914630

Abstract Title: 

Diosmetin Suppresses Neuronal Apoptosis and Inflammation by Modulating the Phosphoinositide 3-Kinase (PI3K)/AKT/Nuclear Factor-κB (NF-κB) Signaling Pathway in a Rat Model of Pneumococcal Meningitis.

Abstract: 

Bacterial meningitis has a high mortality rate and can be challenging to diagnose and manage. This study aimed to evaluate the effect of diosmetin in a rat model of Streptococcus pneumoniae meningitis and to investigate the mechanism of action. Forty rats included a treatment group (n=30) that underwent intracisternal injection with S. pneumoniae, and a sham group (n=10) that underwent intracisternal injection with normal saline. In the treatment group, four days before the inoculation of the bacteria, rats were pre-treated with oral diosmetin 100 mg/kg (n=10) and 200 mg/kg (n=10), and the negative control was pre-treated with normal saline (n=10). Bacterial meningitis was confirmed one day after inoculation by cerebrospinal fluid (CSF) bacterial titer and neurological score. In rat brain tissue, levels of inflammatory mediators were determined by enzyme-linked immunosorbent assay (ELISA) and western blot for protein kinase B (Akt), phosphoinositide 3-kinase (PI3K), myeloid differentiation primary response 88 (MyD88), and nuclear factor-kappaB (NF-kappaB), and the TUNEL assay for apoptosis was performed. In the diosmetin-treated group compared with negative control group, the CSF bacterial titer and the level of pro-inflammatory mediators, and the neurological score, were significantly reduced (p

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PMID: 

Med Sci Monit. 2019 Jun 22 ;25:4639-4647. Epub 2019 Jun 22. PMID: 31228347

Abstract Title: 

Anti-Proliferation and Pro-Apoptotic Effects of Diosmetin via Modulating Cell Cycle Arrest and Mitochondria-Mediated Intrinsic Apoptotic Pathway in MDA-MB-231 Cells.

Abstract: 

BACKGROUND Breast cancer is one of the most malignant tumors worldwide. The natural flavonoid diosmetin has been reported to exhibit various pharmacological activities, including anti-cancer effects. This study aimed to investigate the anti-breast cancer effects of diosmetin on MDA-MB-231 cells and to explore the underlying molecular mechanisms of cell apoptosis. MATERIAL AND METHODS The MDA-MB-231 cells were incubated with diosmetin for 24 h. Then, cell viability and lactate dehydrogenase (LDH) leakage were detected using CCK-8 and LDH assay kits, respectively. Inverted fluorescence microscopy and flow cytometry were used to measure the mitochondrial membrane potential (MMP) and intracellular reactive oxygen species (ROS). Cell apoptosis and cell cycle were determined by flow cytometry. The expressions of apoptosis and cell cycle-related genes were determined by Western blotting and qRT-PCR. RESULTS The results revealed that diosmetin exerts significant cytotoxic effects on MDA-MB-231 cells, as indicated by decreased cell viability, increased intracellular ROS accumulation and LDH release, as well as cell cycle arrest in G0/G1 phase, inducing mitochondrial dysfunction and apoptosis. Moreover, diosmetin treatment significantly downregulated the expression levels of Bcl-2 and Cyclin D1, and upregulated that of p53, Bax, caspase 3, cleaved caspase 9, and cleaved caspase 3. CONCLUSIONS These findings demonstrate that diosmetin has anti-proliferative and pro-apoptotic activities against MDA-MB-231 cells via cell cycle arrest and the mitochondria-mediated intrinsic apoptotic pathway. Our results extend the understanding of the anti-tumor mechanism of diosmetin and suggest that it may be of use as an active natural agent for the prevention or treatment of human breast cancer.

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PMID: 

Biomed Pharmacother. 2019 Sep ;117:109091. Epub 2019 Jun 19. PMID: 31228803

Abstract Title: 

Diosmetin inhibits tumor development and block tumor angiogenesis in skin cancer.

Abstract: 

Diosmetin is a natural flavonoid obtained from citrus fruits and some medicinal herbs. Previous studies have reported the anti-cancer activity of diosmetin in some types of tumors. However, it is still unclear whether diosmetin exerts anti-cancer effects, particularly anti-angiogenic effects, in skin cancer. In this study, we used B16F10 melanoma cells and human umbilical vein endothelial cells to investigate the inhibitory effect of diosmetin on cell proliferation, migration and tube formation in vitro. Rat aorta ring assays were performed to determine the effect of diosmetin on ECs sprouting ex vivo. Furthermore, a B16F10 mouse melanoma model was used to observe the effect of diosmetin on tumor growth, angiogenesis, and metastasis in vivo. Our results showed that diosmetin not only suppressed tumor cell proliferation and migration but also induced cell apoptosis via the caspase pathway in B16F10 cells, and potently inhibited tube formation and cell migration in HUVECs. Rat aorta ring assays showed that diosmetin attenuated the ECs sprouting. Moreover, the mouse melanoma model showed that diosmetin significantly delayed tumor growth by inhibiting tumor vessels sprouting and expansion during tumor progression. Notably, diosmetin induced the normalization of tumor vasculature through the downregulation of angiopoietin-2 and the improvement of pericyte coverage, leading to suppression of metastasis formation in lungs and lymph nodes. In conclusion, our results demonstrate that diosmetin suppresses tumor progression and metastasis by inducing tumor cell death and inhibiting tumor angiogenesis as well as normalizing the defective tumor vasculature, suggesting that diosmetin is a potential adjuvant chemotherapy agent.

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PMID: 

Phytother Res. 2019 Dec 13. Epub 2019 Dec 13. PMID: 31833613

Abstract Title: 

Diosmetin exhibits anti-proliferative and anti-inflammatory effects on TNF-α-stimulated human rheumatoid arthritis fibroblast-like synoviocytes through regulating the Akt and NF-κB signaling pathways.

Abstract: 

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by inflammation and proliferation of synovial tissues. Diosmetin is a bioflavonoid possessing an anti-inflammatory property. Herein, we aimed to study the effects of diosmetin on the inflammation and proliferation of RA fibroblast-like synoviocytes MH7A cells. MH7A cell proliferation was measured using cell counting kit-8 assay. Cell apoptosis was examined using flow cytometry. The production of inflammatory cytokines including interleukin (IL)-1β, IL-6, IL-8, and matrix metalloproteinase-1 (MMP-1) was measured using enzyme-linked immunosorbent assay (ELISA). Results showed that diosmetin inhibited tumor necrosis factor-α (TNF-α)-induced proliferation increase in MH7A cells in a dose-dependent manner. Diosmetin treatment resulted in an increase in apoptotic rates and a reduction in TNF-α-induced production of IL-1β, IL-6, IL-8, and MMP-1 in MH7A cells. Furthermore, diosmetin inhibited TNF-α-induced activation of protein kinase B (Akt) and nuclear factor-κB (NF-κB) pathways in MH7A cells. Suppression of Akt or NF-κB promoted apoptosis and inhibited TNF-α-induced proliferation increase and production of IL-1β, IL-6, IL-8, and MMP-1 in MH7A cells, and diosmetin treatment enhanced these effects. Taken together, these findings suggested that diosmetin exhibited anti-proliferative and anti-inflammatory effects via inhibiting the Akt and NF-κB pathways in MH7A cells.

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PMID: 

Environ Toxicol. 2020 Feb 15. Epub 2020 Feb 15. PMID: 32061149

Abstract Title: 

Protective effect of Diosmin against benzo(a)pyrene-induced lung injury in Swiss Albino Mice.

Abstract: 

Diosmin, a naturally occurring flavonoid commonly present in citrus fruit, is known to exhibit anti-inflammatory, antimutagenic, antioxidant, and free radical scavenging as well as blood lipid lowering activities among others. Diosmin has also been used for the treatment of various diseases including diabetes mellitus and Alzheimer's disease. Our study explores the role of Diosmin in pulmonary toxicity (lung injury) induced by environmental contaminant benzo(a)pyrene [B(a)P]. Swiss Albino Mice (SAM) were administered with either Diosmin 100 or 200 mg/kg body weight daily for 14 days and then challenged with a single dose of B(a)P. On the 15th day, animals were sacrificed; lung tissues and blood were collected for molecular analysis. B(a)P administration in mice induced the thickening of lung epithelium, damaged alveolar architecture, andpromoted inflammatory cell infiltration in the lung tissues. Also, B[a]P significantly increased the expression of NF-kB, COX-2, IL-6, Bax, cleaved caspase 3, and cleaved PARP proteins and decreased antioxidant enzyme levels. Diosmin-100 and Diosmin-200 significantly attenuated the damage to lung epithelium, alveolar architecture, and reduced inflammatory cell infiltration in the lung tissues of mice. Diosmin significantly (P 

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PMID: 

J Nat Prod. 2020 Feb 21. Epub 2020 Feb 21. PMID: 32083866

Abstract Title: 

Diosmin Treats Lipopolysaccharide-Induced Inflammatory Pain and Peritonitis by Blocking NF-κB Activation in Mice.

Abstract: 

Gram-negative bacterial infections induce inflammation and pain. Lipopolysaccharide (LPS) is a pathogen-associated molecular pattern and the major constituent of Gram-negative bacterial cell walls. Diosmin is a citrus flavonoid with antioxidant and anti-inflammatory activities. Here we investigated the efficacy of diosmin in a nonsterile model of inflammatory pain and peritonitis induced by LPS. Diosmin reduced in a dose-dependent manner LPS-induced inflammatory mechanical hyperalgesia, thermal hyperalgesia, and neutrophil recruitment to the paw (myeloperoxidase activity). Diosmin also normalized changes in paw weight distribution assessed by static weight bearing as a nonreflexive method of pain measurement. Moreover, treatment with diosmin inhibited LPS-induced peritonitis as observed by a reduction of leukocyte recruitment and oxidative stress. Diosmin reduced LPS-induced total ROS production (DCFDA assay) and superoxide anion production (NBT assay and NBT-positive cells). We also observed a reduction of LPS-induced oxidative stress and cytokine production (IL-1β, TNF-α, and IL-6) in the paw. Furthermore, we demonstrated that diosmin inhibited LPS-induced NF-κB activation in peritoneal exudate. Thus, we demonstrated, using a model of nonsterile inflammation induced by LPS, that diosmin is a promising molecule for the treatment of inflammation and pain.

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PMID: 

Mol Biol Rep. 2020 Feb 22. Epub 2020 Feb 22. PMID: 32088816

Abstract Title: 

The synergistic anti-proliferative effect of the combination of diosmin and BEZ-235 (dactolisib) on the HCT-116 colorectal cancer cell line occurs through inhibition of the PI3K/Akt/mTOR/NF-κB axis.

Abstract: 

One of the most lethal malignancies worldwide is colorectal cancer (CRC). Alterations in various signalling pathways, including PI3K-mTOR and NF-κB, have been reported in CRC with subsequent dysregulation of proliferation, apoptosis, angiogenesis and, questionably, autophagy processes. BEZ-235 (dactolisib) is a dual PI3K-mTOR inhibitor with potent anti-tumour activity. However, the observed toxicity of BEZ-235 necessitated the termination of its clinical trials. Hence, we aimed to evaluate the potential long-lasting anti-carcinogenic effects of adding diosmin (DIO, a natural NF-κB inhibitor) to BEZ-235 in HCT-116 CRC cells. The median inhibitory concentrations (IC50s) of BEZ-235 and/or DIO were evaluated in the HCT-116 CRC cell line. Caspase-3 activity was assessed colorimetrically, and p-Akt, NF-κB, CD1, VEGF and LC3B levels were assessed by ELISA. Additionally, LC3-II and P62 gene expression were assessed using qRT-PCR. The observed CIs (combination indices) and DRIs (dose reduction indices) confirmed the synergistic effectof DIO and BEZ-235. Co-administration of both drugs either in combination-1 (1 μM for BEZ-235, 250 μM for DIO) or in combination-2 (0.51 μM for BEZ-235 + 101.99 μM for DIO) inhibited the PI3K/Akt/mTOR/NF-κB axis, leading to the induction of apoptosis (via active caspase-3), and the inhibition of proliferation marker (CD1), angiogenesis marker (VEGF), autophagy protein (LC3B) and altered effects on LC3-IIandP62 gene expression. Our results reveal the synergistic chemotherapeutic effects of DIO combined with BEZ-235 in the HCT-116 CRC cell line and encourage future preclinical andclinical studies of this combination with reduced BEZ-235 concentrations to avoid its reported toxicity.

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PMID: 

Regul Toxicol Pharmacol. 2020 Feb 19:104622. Epub 2020 Feb 19. PMID: 32087353

Abstract Title: 

Role of Diosmin in protection against the oxidative stress induced damage by gamma-radiation in Wistar albino rats.

Abstract: 

The benchmark of this study is to evaluate the radio protective efficiency of diosmin, a natural citrus flavone of hesperidin derivative on radiation-induced damage in wistar albino rats. Rats orally administered two diosmin doses (100 and 200 mg/kg body wt.) for one month (day after day) prior to exposure to high gamma radiation single dose (8Gy) or cumulative dose (10Gy). To evaluate the radio protective efficiency of diosmin various biochemical estimations, histopathological alterations as well as comet assay and caspase-3 activityfor assessment of apoptosis were performed. Results indicated that radiation-induced decline in the levels of antioxidant parameters (SOD and GSH), increased lipid peroxidation, DNA damage and apoptosis were improved by pre-administration of diosmin. Diosmin dose (200 mg/kg body wt.) restored theantioxidant status to near normal and reduced lipid peroxidation, DNA and tissue damage. These results were confirmed by histopathological examinations, which showed that pre-administration of diosmin protected the liver and kidney of albino rats against gamma-irradiation induced damage. Hence, it has been illustrated that diosmin might be an effective radio protector against radiation-induced damage in rats. Moreover, diosmin alone pretreated group did not show any biochemical alterations or DNA damage indicating the protective nature of the drug.

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