Oxidative Stress Mediated Cytotoxicity, Cell Cycle Arrest, and Apoptosis Induced by Rosa damascena in Human Cervical Cancer HeLa Cells.

Rosa damascena Mill (Damask rose), belonging to the Rosaceae family, is known for medicinal purposes in traditional medicine system. However, its anticancer activity has not been studied yet in detail. Herein, we aimed to investigate the cytotoxic effects of R. damascena hexane (RA-HE) and methanolic (RA-ME) extracts against human breast (MCF-7), lung epithelial (A-549), and cervical (HeLa) cancer cells. The RA-HE and RA-ME showed more potent cytotoxic effects against HeLa cells with an IC50 of 819.6 and 198.4 µg/ml, respectively. Further, cytotoxic concentrations of most effective extract (RA-ME) were used to evaluate the mechanism of cytotoxicity involved in HeLa cells. A concentration-dependent induction of lipid peroxidation (LPO) and reduction of glutathione (GSH) in HeLa cells treated with 250-1000 µg/ml of RA-ME confirms the association of oxidative stress. We also detected a noteworthy increase in reactive oxygen species (ROS) production and a decline in mitochondrial membrane potential (MMP) level in RA-ME-exposed HeLa cells. Flow cytometric data showed a strong dose-response relationship in cell cycle analysis between subG1 phase in HeLa cells and RA-ME treatment. Similarly, a concentration-dependent increase was recorded with Annexin V assay in HeLa cells going to late apoptosis. In conclusion, our findings suggest that RA-ME-induced cytotoxicity and apoptosis in HeLa cells are mediated by oxidative stress.

Neuroprotective Effects of Withania somnifera on 4-Hydroxynonenal Induced Cell Death in Human Neuroblastoma SH-SY5Y Cells Through ROS Inhibition and Apoptotic Mitochondrial Pathway.

The antioxidant, anti-inflammatory, and anticancer activities of Withania somnifera (WS) are known for a long time. This study was aimed to examine whether WS also diminishes 4-hydroxy-trans-2-nonenal (HNE)-induced neurotoxicity in human neuroblastoma (SH-SY5Y) cell line. The cytotoxic response of HNE (0.1-50 µM) and WS (6.25-200 µg/ml) was measured by MTT assay after exposing SH-SY5Y cells for 24 h. Then neuroprotective potential was assessed by exposing the cells to biologically safe concentrations of WS (12.5, 25, and 50 µg/ml) then HNE (50 µM). Results showed a concentration-dependent protective effect of WS at 12.5, 25, and 50 µg/ml against HNE (50 µM) induced cytotoxicity and cell inhibition. Pre-exposure to WS resulted in a strong inhibition of 24, 55 and 83% in malondialdehyde (MDA) level; 5, 27 and 60% in glutathione (GSH) level; 12, 36 and 68% in catalase activity; 11, 33 and 67% in LDH leakage; and 40, 80 and 120% in cellular LDH activity at 12.5, 25, and 50 µg/ml, respectively, induced by 50 µM HNE in SH-SY5Y cells. The HNE-mediated cellular changes (cell shrinkage, rounded bodies, and inhibition of outgrowth) and increased caspase-3 activity were also prevented by WS. The HNE-induced upregulation of proapoptotic markers (p53, caspase-3, and -9, and Bax) and downregulation of antiapoptotic marker Bcl-2 genes were also blocked by pretreatment with WS. Altogether, our findings indicate that WS possesses a protective potential against HNE-induced neurotoxicity.

Protective effects of Nigella sativa extract against H2 O2 -induced cell death through the inhibition of DNA damage and cell cycle arrest in human umbilical vein endothelial cells (HUVECs).

Oxidative stress is known to induce cytotoxicity and apoptosis in endothelial cells and indorse development of atherosclerosis. The aim of this research was to assess the cytoprotective effects of ethanolic extract of Nigella sativa (NSE) against H2 O2 -induced cell death in human umbilical vein endothelial cells (HUVECs) and also study the probable mechanisms through which NSE exhibited cyto-protection. The cytotoxicity was measured by exposing the HUVECs with NSE (10-200 µg/ml) and H2 O2 (25-1000 µM) for 24 h. Then, the HUVECs were pretreated with noncytotoxic doses (10-50 µg/ml) of NSE for 24 h before administration of 200 µM H2 O2 for 24 h. The MTT, NRU, and morphological assays were performed to assess the cytotoxicity and cyto-protection. Potential antioxidant activity of NSE on oxidative stress marker (glutathione [GSH] and lipid peroxidation [LPO]) was also evaluated. The fluorescence probe, DCF-DA, and Rh123 were applied to measure the reactive oxygen species (ROS) level and mitochondrial membrane potential. Moreover, flow cytometric analysis and comet assay were used to study the cell cycle arrest and DNA damage, respectively. The concentrations (10, 30, and 50 µg/ml) of NSE were found to protect HUVECs against H2 O2 (200 µM)-induced cytotoxicity in HUVECs. Pretreatment of HUVECs with NSE significantly reduced the LPO and ROS levels and restored the GSH and loss of MMP induced by H2 O2 . Furthermore, NSE inhibited H2 O2 -induced cell cycle arrest and cellular DNA damage in HUVECs. Altogether, these results suggest that NSE can prevent H2 O2 -induced cell death, and NSE could be a potential candidate that can prevent HUVECs against toxicants.

Petroselinum sativum protects HepG2 cells from cytotoxicity and oxidative stress induced by hydrogen peroxide.

A number of liver diseases are known to be caused by oxidative stress. Petroselinum sativum (P. sativum; parsley) is popular for its anti-inflammatory, antimicrobial, anticancer, antioxidant and antidiabetic activities. However, till date the hepatoprotective potential of chloroform extract of P. sativum (PSA) on hydrogen peroxide (H2O2) induced cytotoxicity and oxidative stress in human liver (HepG2) cells have not been studied. Therefore, this study was framed to evaluate whether the levels of hydrogen peroxide (H2O2) induced cytotoxicity and oxidative stress in HepG2 cells could be diminished by pretreating the cells with PSA. MTT assay, NRU assay, morphological alterations, glutathione (GSH) depletion, lipid peroxidation (LPO), ROS generation and loss of mitochondrial membrane potential (MMP) were assessed by using non-cytotoxic concentrations (5, 10 and 25 µg/mL) of PSA against H2O2 (0.25 mM) induced damage in HepG2 cells. The results demonstrated that pretreatment of HepG2 cells with PSA offered protective properties by lowering the LPO and ROS generation and elevating the cell viability, GSH and MMP levels. Together, these results suggest that PSA has the hepatoprotective effect on H2O2 induced cell death in HepG2 cells.

In-Vitro dual inhibition of protein glycation, and oxidation by some Arabian plants.

BACKGROUND: Diabetes mellitus is a metabolic disorder of epidemic proportion, projected to become the major cause of morbidity and mortality in the world in future. Despite extensive research in understanding this disease at molecular level, and the discovery of new drugs, diabetes and its complications remain largely untreated. Many of the late diabetic complications are associated with the glycation of proteins in the body. Natural flora has long been a rich source for therapeutic agents, especially against diabetes. The present study deals with the anti-glycation properties of some medicinally important plants of Arabian region. METHODS: Twenty-six medicinal plants, commonly found in different regions of Arabian Peninsula, were evaluated for their protein anti-glycation activity by using BSA-MG glycation assay in-vitro. The extracts were incubated with BSA and MG at 37 °C for 9 days, each sample was then examined for the presence of fluorescence (λex 330 nm, and λem 420 nm), which represent the extent of protein glycation. Antioxidant activity was evaluated by using 1,1-diphenyl- 2-picrylhydrazyl (DPPH), iron chelation, and superoxide radical scavenging asaays. RESULTS: The data revealed that out of 26 medicinal plants, five plants viz. Sida cordifolia, Plumbago zeylanica, Tribulus terrestris, Glycyrrhiza glabra, and Rosa indica were active against the in-vitro protein glycation with IC50 values between 0.408- 1.690 mg/mL. Among the active plants, Glycyrrhiza glabra L. was found to be the most potent (IC50 = 0.408 ± 0.027 mg/mL), followed by Rosa indica (IC50 = 0.596 ± 0.0179 mg/mL), and Sida cordifolia L. (IC50 = 0.63 ± 0.009 mg/mL). The antioxidant potential of these plant extracts were also determined by using DPPH (2,2-diphenyl-1-picrylhydrazyl), iron chelation, and superoxide anion radical scavenging assays. Among five plants, Sida cordifolia exhibited a potent anti-oxidant activity in both DPPH and superoxide anion radical scavenging assays (IC50 = 0.005 ± 0.0004, and 0.078 ± 0.002 mg/mL, respectively), followed by Rosa indica (IC50 = 0.023 ± 0.0005 and 0.141 ± 0.003 mg/mL, respectively). CONCLUSIONS: Protein glycation in hyperglycemic conditions involve oxidative changes. Therefore dual inhibition of protein glycation and oxidation are desirable properties in any test substance investigated for therapeutic purposes.

Verbesina encelioides: cytotoxicity, cell cycle arrest, and oxidative DNA damage in human liver cancer (HepG2) cell line.

BACKGROUND: Cancer is a major health problem and exploiting natural products have been one of the most successful methods to combat this disease. Verbesina encelioides is a notorious weed with various pharmacological properties. The aim of the present investigation was to screen the anticancer potential of V. encelioides extract against human lung cancer (A-549), breast cancer (MCF-7), and liver cancer (HepG2) cell lines. METHODS: A-549, MCF-7, and HepG2 cells were exposed to various concentrations of (10-1000 µg/ml) of V. encelioides for 24 h. Further, cytotoxic concentrations (250, 500, and 1000 µg/ml) of V. encelioides induced oxidative stress (GSH and LPO), reactive oxygen species (ROS) generation, mitochondrial membrane potential (MMP), cell cycle arrest, and DNA damage in HepG2 cells were studied. RESULTS: The exposure of cells to 10-1000 µg/ml of extract for 24 h, revealed the concentrations 250-1000 µg/ml was cytotoxic against MCF-7 and HepG2 cells, but not against A-549 cells. Moreover, the extract showed higher decrease in the cell viability against HepG2 cells than MCF-7 cells. Therefore, HepG2 cells were selected for further studies viz. oxidative stress (GSH and LPO), reactive oxygen species (ROS) generation, mitochondrial membrane potential (MMP), cell cycle arrest, and DNA damage. The results revealed differential anticancer activity of V. encelioides against A-549, MCF-7 and HepG2 cells. A significant induction of oxidative stress, ROS generation, and MMP levels was observed in HepG2 cells. The cell cycle analysis and comet assay showed that V. encelioides significantly induced G2/M arrests and DNA damage. CONCLUSION: These results indicate that V. encelioides possess substantial cytotoxic potential and may warrant further investigation to develop potential anticancer agent.

Protective effect of Lepidium sativum seed extract against hydrogen peroxide-induced cytotoxicity and oxidative stress in human liver cells (HepG2).

CONTEXT: Garden cress [Lepidium sativum (Brassicaceae)] has been widely used to treat a number of ailments in traditional medicine. The pharmacological and preventive potential of Lepidium sativum, such as anti-inflammatory, antipyretic, antihypertensive, anti-ashthamatic, anticancer, and anti-oxidant, are well known. OBJECTIVE: The present investigation was designed to study the protective effects of chloroform extract of Lepidium sativum seed (LSE) against oxidative stress and cytotoxicity induced by hydrogen peroxide (H2O2) in human liver cells (HepG2). MATERIALS AND METHODS: Cytotoxicity of LSE and H2O2 was identified by (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), neutral red uptake (NRU) assays, and morphological changes in HepG2. The cells were pre-exposed to biologically safe concentrations (5-25 µg/ml) of LSE for 24 h, and then cytotoxic (0.25 mM) concentration of H2O2 was added. After 24 h of the exposures, cell viability by MTT, NRU assays, and morphological changes in HepG2 were evaluated. Further, protective effects of LSE on reactive oxygen species (ROS) generation, mitochondrial membrane potential (MMP), lipid peroxidation (LPO), and reduced glutathione (GSH) levels induced by H2O2 were studied. RESULTS: Pre-exposure of LSE significantly attenuated the loss of cell viability up to 48% at 25 µg/ml concentration against H2O2 (LD50 value = 2.5 mM). Results also showed that LSE at 25 µg/ml concentration significantly inhibited the induction of ROS generation (45%) and LPO (56%), and increases the MMP (55%) and GSH levels (46%). DISCUSSION AND CONCLUSION: The study suggests the cytoprotective effects of LSE against H2O2-induced toxicity in HepG2. The results also demonstrate the anti-oxidative nature of LSE.

Anticancer Activity of Chloroform Extract and Sub-fractions of Nepeta deflersiana on Human Breast and Lung Cancer Cells: An In vitro Cytotoxicity Assessment.

BACKGROUND: Cancer is one of the major causes of death worldwide. The plant-derived natural products have received considerable attention in recent years due to their diverse pharmacological properties including anticancer effects. Nepeta deflersiana (ND) is used in the folk medicine as antiseptic, carminative, antimicrobial, antioxidant, and for treating rheumatic disorders. However, the anticancer activity of ND chloroform extract has not been explored so far. OBJECTIVES: The present study was aimed to investigate the anticancer activities of chloroform Nepeta deflersiana extract and various sub-fractions (ND-1-ND-15) of ND against human breast cancer cells (MCF-7) and human lung cancer cells (A-549). MATERIALS AND METHODS: The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide and neutral red uptake assays, and cellular morphological alterations using phase contrast light microscope were studied. Cells were exposed with 10-1000 µg/ml of sub-fractions of ND for 24 h. RESULTS: Results showed that selected sub-fractions of the chloroform extract significantly reduced the cell viability of MCF-7 and A-549 cells, and altered the cellular morphology in a concentration-dependent manner. Among the sub-fractions, ND-10 fraction showed relatively higher cytotoxicity compared to other fractions whereas, ND-1 did not cause any cytotoxicity even at higher concentrations. The A-549 cells were found to be more sensitive to growth inhibition by all the extracts as compared to the MCF-7 cells. CONCLUSION: The present study provides preliminary screening of anticancer activities of chloroform extract and sub-fractions of ND, which can be further used for the development of a potential therapeutic anticancer agent. SUMMARY: Nepeta deflersiana extract exhibit cytotoxicity and altered the cellular morphology. Sub-fractions of the chloroform extract of Nepeta deflersiana reduced the cell viability of MCF-7 and A-549 cells. Among the sub-fractions, ND-10 fraction showed relatively higher cytotoxicity. The A-549 cells were found to be more sensitive as compared to the MCF-7 cells. Abbreviations used: MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide; NRU: Neutral red uptake; DMEM: Dulbecco's modified eagle medium; FBS: Fetal bovine serum; PBS: Phosphate buffer saline; DMSO: Dimethyl sulfoxide.

Hepatoprotective potential of Lavandula coronopifolia extracts against ethanol induced oxidative stress-mediated cytotoxicity in HepG2 cells.

The present investigations were carried out to study the protective potential of four extracts (namely petroleum ether extract (LCR), chloroform extract (LCM), ethyl acetate extract (LCE), and alcoholic extract (LCL)) of Lavandula coronopifolia on oxidative stress-mediated cell death induced by ethanol, a known hepatotoxin in human hapatocellular carcinoma (HepG2) cells. Cells were pretreated with LCR, LCM, LCE, and LCL extracts (10-50 µg/ml) of L. coronopifolia for 24 h and then ethanol was added and incubated further for 24 h. After the exposure, cell viability using (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and neutral red uptake assays and morphological changes in HepG2 cells were studied. Pretreatment with various extracts of L. coronpifolia was found to be significantly effective in countering the cytotoxic responses of ethanol. Antioxidant properties of these L. coronopifolia extracts against reactive oxygen species (ROS) generation, lipid peroxidation (LPO), and glutathione (GSH) levels induced by ethanol were investigated. Results show that pretreatment with these extracts for 24 h significantly inhibited ROS generation and LPO induced and increased the GSH levels reduced by ethanol. The data from the study suggests that LCR, LCM, LCE, and LCL extracts of L. coronopifolia showed hepatoprotective activity against ethanol-induced damage in HepG2 cells. However, a comparative study revealed that the LCE extract was found to be the most effective and LCL the least effective. The hepatoprotective effects observed in the study could be associated with the antioxidant properties of these extracts of L. coronopifolia.