Fractions of Hoslundia opposita Vahl and hoslundin induced apoptosis in human cancer cells via mitochondrial-dependent reactive oxygen species (ROS) generation.

BACKGROUND: Cancer remains one of the leading causalities of several morbidity and mortality with negative impact on global economy due to low workforce and management/treatment cost. A number of conventional therapies have been explored in the management/treatment of cancer including chemotherapeutic intervention, radiotherapy, and surgery. Among these treatment modalities, chemotherapy remains the most popular first line of intervention in management/treatment of cancer, and natural products have been implicated as the main source of antineoplastic agents with phenomenal efficacy. However, current antineoplastic agents suffer from lack of selectivity and specificity necessitating the need for further research in the search for novel anticancer drug molecules. METHODS: In this present study, the anticancer activity of Hoslundia opposita leaves extracts were tested against a number of cell lines including human hepatoma cell line (HepG2), human breast cancer cell lines (MDA-MB-231), intestinal epithelial cell lines (Caco-2), and human keratinocyte HACAT cell lines. A bio-guided fractionation assay and the structural elucidation of the pure isolate (hoslundin) was conducted by 1D and 2D NMR spectroscopy. The cell viability, colony formation, and apoptotic activities were investigated using MTT assay, clonogenic assay, and caspase - 3 and - 7 kits respectively. Flow cytometry was employed in assessing the altered cell cycle expression. The production of the intracellular reactive oxygen species (ROS) levels and the reduction of the mitochondrial membrane potential (MMP) was determined at the cellular level using fluorescent probe dyes dihydro-fluorescin diacetate (DCFH-DA) and tetramethylrhodamin (TMRE), respectively. RESULTS: The H. opposita fractions and its pure isolate (hoslundin) demonstrated a potent cytocidal activity against the tumorigenic cells (HepG2, MDA-MB-231, Caco-2) at concentration ranging from 25 to 100 µg/mL. The inhibition of the colony formation was significantly observed in HepG2 cell lines. More so, the cellular viability of the normal cells (HaCaT) was relatively unchanged in the presence of H. opposita fractions and its isolate proving the selectivity of the compounds towards tumourigenic cells. The H. opposita fractions and hoslundin exerted their anticancer activity via cell cycle arrest with the accumulation of the DNA content at the S-phase, activation of apoptosis in the caspase 3,7 activities and depolarized mitochondrial membrane potential mediated by mitochondrial-dependent ROS generation in the treated tumor cells. CONCLUSION: The anticancer activities of Hoslundia opposita Vahl and hoslundin exhibited significant efficacy against tumor cells and well tolerated in the presence of normal cells making them a potential antineoplastic agent.

Mitoquinol mesylate (MITOQ) attenuates diethyl nitrosamine-induced hepatocellular carcinoma through modulation of mitochondrial antioxidant defense systems.

Diethyl nitrosamine (DEN) induced cirrhosis-hepatocellular carcinoma (HCC) model associates cancer progression with oxidative stress and mitochondrial dysfunction. This study investigated the effects of mitoquinol mesylate (MitoQ), a mitochondrial-targeted antioxidant on DEN-induced oxidative damage in HCC Wistar rats. Fifty male Wistar rats were randomly divided into five groups. Healthy control, DEN, and MitoQ groups were orally administered exactly 10 mg/kg of distilled water, DEN, and MitoQ, respectively for 16 weeks. Animals in the MitoQ + DEN group were pre-treated with MitoQ for a week followed by co-administration of 10 mg/kg each of MitoQ and DEN. DEN + MitoQ group received DEN for 8 weeks, then co-administration of 10 mg/kg each of DEN and MitoQ till the end of 16th week. Survival index, tumour incidence, hematological profile, liver function indices, lipid profile, mitochondrial membrane composition, mitochondrial respiratory enzymes, and antioxidant defense status in both mitochondrial and post-mitochondrial fractions plus expression of antioxidant genes were assessed. In MitoQ + DEN and DEN + MitoQ groups, 80% survival occurred while tumour incidence decreased by 60% and 40% respectively, compared to the DEN-only treated group. Similarly, MitoQ-administered groups showed a significant (p < 0.05) decrease in the activities of liver function enzymes while hemoglobin concentration, red blood cell count, and packed cell volume were significantly elevated compared to the DEN-only treated group. Administration of MitoQ to the DEN-intoxicated groups successfully enhanced the activities of mitochondrial F1F0-ATPase and succinate dehydrogenase; and up-regulated the expression and activities of SOD2, CAT, and GPx1. Macroscopic and microscopic features indicated a reversal of DEN-induced hepatocellular degeneration in the MitoQ + DEN and DEN + MitoQ groups. These data revealed that MitoQ intervention attenuated DEN-induced oxidative stress through modulation of mitochondrial antioxidant defense systems and alleviated the burden of HCC as a chemotherapeutic agent.

Anti-apoptotic effect of Buchholzia coriacea Engl. stem back extracts on AsPC-1 and mechanisms of action.

ETHNOPHARMACOLOGICAL RELEVANCE: Buchholzia coriacea Engl. is popularly called wonderful cola due to its wide ethnomedicinal use for the treatment of various ailments. We investigated the possible cytotoxic effect of its various fractions on human pancreatic cancer cell (AsPC-1) and also determined its mechanisms of action. MATERIALS AND METHODS: The AsPC-1 cells were cultivated and separately treated with 5-fluorouracil (5-FU) or Buchholzia coriacea Engl. bark (BC) (ethanol, aqueous, chloroform or ethyl acetate extract) for 72 h. Cell viability, caspase 3 and mitochondrial membrane potential (ΔΨm) were determined in vitro after the treatment. Nitric oxide (NO) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radicals' scavenging property, ferric reducing power and lipid peroxidation assays were also done to examine the antioxidant effect of BC in vitro. RESULTS: Various extracts of BC, especially at 2500 µg/ml and 5000 µg/ml, increased the AsPC-1 viability while 5-FU decreased it. The activity of caspase 3 was increased by 5-FU but reduced by all concentrations of various extracts of BC. Incubation of AsPC-1 with 5-FU showed the majority of cells having the monomeric form of JC-1 dye (bright green fluorescence), which indicated de-energized mitochondria. However, fluorescence photomicrograph of cells incubated with different concentrations (20, 40 and 100 µg/ml) of BC extracts (aqueous, ethanol, chloroform and ethyl acetate) showed strong JC-1 aggregation (yellow), which indicated mitochondria with intact membrane potentials. BC extracts also scavenged NO and DPPH radicals, inhibited lipid peroxidation and increased ferric reduction, though not as much as ascorbic acid. CONCLUSION: This study suggests that BC elicits anti-apoptotic activity in AsPC-1 by increasing cell viability, decreasing caspase 3 activity, stabilizing the ∆Ψm, and scavenging free radicals. Even though BC is used ethnomedicinally as anti-cancer agent, our findings in the present study suggest that it has pro-cancer potential in-vitro, especially on pancreatic cells. Its anti-apoptotic activity in AsPC-1 could be of clinical significance, especially to counteract the effect of apoptotic agents on pancreatic cells.

Molecular mechanism of mitoquinol mesylate in mitigating the progression of hepatocellular carcinoma-in silico and in vivo studies.

The safety and efficacy of mitoquinol mesylate (MitoQ) in attenuating the progression of hepatocellular carcinoma (HCC) in Wistar rats has been reported. However, the binding modes for MitoQ as well as its molecular mechanisms in cirrhosis and liver cancer have not been fully investigated. This study sought to understand the structural and molecular mechanisms of MitoQ in modulating the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and mitochondrial succinate dehydrogenase (SDH) in cirrhotic-HCC rats. The research indicates that the upregulated Nrf2 expression in cirrhotic-HCC rats was significantly (p < 0.05) reduced by MitoQ while the activity of SDH was significantly (p < 0.05) increased. Analysis of binding modes revealed MitoQ interacts with amino acid residues in the active pocket of tramtrack and bric-a-brac (BTB) and KELCH domains of KEAP1 with average binding affinities of -66.46 and -74.74 kcal/mol, respectively. Also, MitoQ interacted with the key amino acid residues at the active site of mitochondrial complex II with a higher average binding affinity of -75.76 kcal/mol compared to co-crystallized ligand of complex II (-62.31 kcal/mol). Molecular dynamics simulations data showed the binding of MitoQ to be stable with low eigenvalues while the quantum mechanics calculations suggest MitoQ to be very reactive with its mechanism of chemical reactivity to be via electrophilic reactions. Thus, MitoQ modulates expression of Nrf2 and enhances activity of mitochondrial SDH in cirrhotic-HCC rats via its interaction with key amino acid residues in the active pocket of BTB and KELCH domains of KEAP1 as well as amino residues at the active site of SDH. These findings are significant in demonstrating the potential of Nrf2 and SDH as possible biomarkers for the diagnosis and/or prognosis of hepatocellular carcinoma in patients. This study also supports repurposing of mitoQ for the treatment/management of liver cirrhosis and HCC.

Ethanolic extract of Camellia sinensise licited hypoglycemic but lacked antimalarial properties in Plasmodium berghei-infected diabetic mice.

The in vivo antimalarial and antidiabetic activity of extract of Camellia sinensis (ECS) in alloxan-induced diabetic and Plasmodium berghei-infected mice were investigated. Eighty-four BALB/c mice divided into sets 1 & 2 infected with P. berghei and 2 & 3 injected with alloxan received either distilled water, ECS (300mg/kg), Chloroquine (CQ-10mg/kg) or Metformin (250mg/kg). Results showed significant increases (p<0.05) in percentage parasitaemia of P. berghei-infected mice treated with ECS and P. berghei-diabetic mice. Furthermore, ECS significantly decreased (p<0.05) blood glucose and PCV in diabetic and P. berghei-diabetic mice.  ECS regenerated pancreatic islet cells in P. berghei-infected-diabetes but lacked appreciable antimalarial activity.

Alterations of Antioxidant Status and Mitochondrial Succinate Dehydrogenase Activity in the Liver of Wistar Strain Albino Rats Treated with by Ethanol Extracts of Annona senegalensis Pers (Annonaceae) Stem Bark.

Numerous ethnomedicinal uses have been attributed to different parts of Annona senegalensis (ASE), including its uses as food and food additives. The present study investigated toxicological and antioxidant effects of 28 days administration of ethanol extracts of ASE stem bark to Wistar strain albino rats. Acute toxicity test was done to determine lethal dose in Wistar rats while sub-acute toxicity test was conducted on rats divided into four groups (A - control, B - 50 mg/kg, C - 100 mg/kg, D - 150 mg/kg, respectively and treated for 28 days. Oxidative stress markers in liver and kidney as well as hepatic succinate dehydrogenase activity in the mitochondrial and post mitochondrial fractions (PMF) were evaluated. The LD50 value of ASE was > 2,000 mg/kg. White blood cell counts gradually increased, but red blood cell counts and haematocrits level decreased significantly (p < 0.05) by about 50%. Liver enzymes in the serum and mitochondrial succinate dehydrogenase activity increased significantly (p < 0.05). Superoxide dismutase and catalase activities also increased in liver mitochondria and PMF while malondialdehyde (MDA) and reduced glutathione levels increased only in the PMF. Furthermore, only MDA levels increased significantly in the kidney after 28 days extract administration. Histopathological examination showed hepatic necrosis and no obvious signs of nephrotoxicity. Anona senegalensis is relatively safe, but prolonged ingestion could induce oxidative stress and impair ATP synthesis through the modulation of the activity of mitochondrial succinate dehydrogenase.

Assessment of liver antioxidant status and mitochondrial membrane composition of Plasmodium berghei-infected mice treated with selected antimalarials.

The present study was aimed at determining and comparing the effects of Artecxin (ART), P - Alaxin (P-ALA), Lonart (LON) and Chloroquine (CQ) on oxidative stress parameters and mitochondrial membrane composition in the course of malaria infection. Six groups of five mice each categorized as healthy control (non-parasitized non-treated group), parasitized-non-treated (PnT), parasitized-chloroquine-treated (positive control), parasitized-Artecxin, -Lonart and -P-Alaxin-treated groups were used for the study. Hepatic antioxidant status was assessed with levels of malondialdehyde (MDA) and reduced glutathione (GSH) as well as activity of superoxide dismutase (SOD) and catalase (CAT) in the post mitochondrial and mitochondrial fractions. Mitochondrial membrane integrity was also evaluated with activity of succinate dehydrogenase and levels of phospholipids, cholesterol and proteins in the liver mitochondria. Results revealed that treatment of parasitized mice with the antimalarial drugs significantly (p<0.05) decreased hepatic malondialdehyde (MDA) and mitochondrial membrane phospholipids compared to parasitized untreated group. On the other hand, significantly (p<0.05) elevated succinate dehydrogenase (SDH) activity, mitochondrial membrane cholesterol level, GSH concentration, catalase (CAT) and superoxide dismutase (SOD) activity in the post mitochondrial fraction were obtained. Thus, antimalarial drugs distort mitochondrial membrane integrity and electron transfer but reduce the malaria-induced oxidative stress on the host.