Garima Shakya

Influence of stabilizers on the production of disulfiram-loaded poly(lactic-co-glycolic acid) nanoparticles and their anticancer potential

AIM Our hypothesis was to prove that surface modifiers themselves can be used as stabilizers and that their entrapment efficiency is directly influenced by the type of stabilizers used. MATERIALS & METHODS Particle size and the polydispersity index of the nanoparticles (NPs) were measured by dynamic light scattering, whereas the morphology of the NPs was studied by scanning electron microscopy. Percentage nanoparticle yield, entrapment efficiency and drug loading capacity were measured by ultraviolet absorbance. The physical rigidity, robustness and drug releasing capability of these NPs were also assessed. CONCLUSION Physiochemical characterization and the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay suggest that polysorbate 80 has the dual capability of being a stabilizer and a surface modifier in addition to having better drug entrapment properties than Pluronic® 188. Disulfiram, the drug that was loaded on these NPs, is also observed for the first time to show significant anticancer potential against hepatocellular carcinoma (Hep3B) cell lines.

Anti-proliferative and apoptosis-triggering potential of disulfiram and disulfiram-loaded polysorbate 80-stabilized PLGA nanoparticles on hepatocellular carcinoma Hep3B cell line

There is an emerging trend to restudy known drugs for their anti-cancer potential. One such anti-alcoholic drug, disulfiram, with significant anti-cancer potential was studied for its efficacy against Hep3B cell lines, an in vitro model of hepatocellular carcinoma. Simultaneously, we intended to study the effect of polysorbate 80-stabilized PLGA nanoparticles and its DSF-loaded counterpart. Cell and nuclear staining, comet assay, flow cytometry and Western blots were performed. Results suggest that cell proliferation was inhibited by DSF and its PLGA nanoparticles through cell cycle arrest, triggering activation of apoptotic pathways that culminates with cell death. DSF loaded nanoparticles when compared with free DSF, showed significantly lesser effect due to its sustained drug-releasing property, while empty nanoparticles showed negligible influence on Hep3B cells. Our results suggest that DSF alone contributes to cell death, while polysorbate 80-stabilized PLGA nanoparticles show sustained drug release patterns that would potentially lower dosage regimens.

Hepatoprotective role of kaempferol during alcohol- and ΔPUFA-induced oxidative stress.

Abstract Background: The present study aimed to analyze the effect of kaempferol on oxidative stress induced by alcohol and thermally oxidized polyunsaturated fatty acid (ΔPUFA) in male albino Wistar rats. Methods: The rats were divided into four groups. The animals in group 1 served as the normal group (standard diet), group 2 served as the hepatotoxic group (alcohol+ΔPUFA), group 3 served as the treated group (alcohol+ΔPUFA+kaempferol), and group 4 served as kaempferol control. The levels of marker enzyme γ-glutamyl transferase (GGT), lipid peroxidation markers [thiobarbituric acid reactive substances (TBARS) and lipid hydroperoxides (LH)], enzymatic antioxidants (catalase, superoxide dismutase, and glutathione peroxidase), and nonenzymatic antioxidants (reduced glutathione, vitamin E, and vitamin C) were analyzed in liver to evaluate the effects of kaempferol. Results: The levels of GGT, TBARS, and LH were significantly increased in liver of the alcohol+ΔPUFA group and were found to be reduced on treatment with kaempferol. The levels of both enzymatic and nonenzymatic antioxidants were decreased in liver of the alcohol+ΔPUFA group and were found to be restored on treatment with kaempferol. Conclusions: From the results obtained, we conclude that kaempferol protects the liver against alcohol- and ΔPUFA- induced oxidative stress.

Phytochemical screening and analysis of antioxidant properties of aqueous extract of wheatgrass.

OBJECTIVE To screen the phytochemical constituents and study antioxidant properties of the aqueous extract of the wheatgrass. METHODS The current study was focused on broad parameters namely, phytochemical analysis, gas chromatography-mass spectrometry analysis and antioxidant properties in order to characterize the aqueous extract of wheatgrass as a potential free radical quencher. RESULTS The phytochemical screening of the aqueous extract of wheatgrass showed the presence of various secondary metabolites but the absence of sterols and quinone in general. Wheatgrass was proved to be an effective radical scavenger in all antioxidant assays. The gas chromatography-mass spectrometry analysis confirmed the presence of diverse category of bioactive compounds such as squalene, caryophyllene and amyrins in varying percentage. CONCLUSIONS From the results obtained, we conclude that wheatgrass aqueous extract contains various effective compounds. It is a potential source of natural antioxidants. Further analysis of this herb will help in finding new effective compounds which can be of potent use in pharmacological field.

Methanol extract of wheatgrass induces G1 cell cycle arrest in a p53-dependent manner and down regulates the expression of cyclin D1 in human laryngeal cancer cells-an in vitro and in silico approach

Background: Deregs been implicated in the malignancy of cancer. Since many years investigation on the traditional herbs has been the focus to develop novel and effective drug for cancer remedies. Wheatgrass is a medicinal plant, used in folk medicine to cure various diseases. The present study was undertaken to gain insights into antiproliferative effect of methanol extract of wheatgrass. Materials Methods: Cell viability was assessed via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and Lactate Dehydrogenase assays. Cell cycle was analyzed by flow cytometry. Western blot was performed to determine the p53 and cyclin D1 levels. In silico docking interaction of the 14 active components (identified by high-performance liquid chromatography/gas chromatography-mass spectroscopy) of the methanol extract was tested with cyclin D1 (Protein Data Bank ID: 2W96) and compared with the reference cyclin D1/Cdk4 inhibitor. Results: Methanol extract of wheatgrass effectively reduced the cell viability. The cell cycle analysis showed that the extract treatment caused G1 arrest. The level of cyclin D1 was decreased, whereas p53 level was increased. Molecular docking studies revealed interaction of seven active compounds of the extract with the vital residues (Lys112/Glu141) of cyclin D1. Conclusion: These findings indicate that the methanol extract of wheatgrass inhibits human laryngeal cancer cell proliferation via cell cycle G1 arrest and p53 induction. The seven active compounds of the extract were also found to be directly involved in the inhibition of cyclin D1/Cdk4 binding, thus inhibiting the cell proliferation.

Inhibition of metastasis and angiogenesis in Hep-2 cells by wheatgrass extract – an in vitro and in silico approach

Abstract Metastasis is the major hindrance in the treatment of all cancers, including laryngeal squamous cell carcinoma. Intensive researches are under way to identify the effective natural polyphenols with anti-metastatic ability for cancer treatment. Wheatgrass, an herbal plant has been reported to show anticancer effects. Hence, in this study, we aimed to analyze the anti-metastatic effect of methanol extract of wheatgrass (MEWG). The levels of metastatic marker proteins were determined by western blot. PI3K and AKT levels were determined by real time (RT)-PCR analysis. In silico molecular docking was done to check the interaction of the 14 components (identified by HPLC/GCMS) of MEWG with PI3K and AKT. MEWG effectively decreased the metastatic protein expressions, namely VEGF, MMP-9 and COX-2 and increased TIMP-2. RT-PCR results showed reduced m-RNA levels of both PI3K and AKT when compared to control. Molecular docking studies revealed interaction of most of the identified compounds of the extract with the important residues of PI3K and AKT. These findings indicate that MEWG inhibits metastasis and angiogenesis in Hep-2 cells possibly via PI3K/AKT due to the cumulative effect of polyphenols and other constituent present in extract. The compounds of the extract were also found to be directly involved in inhibition of AKT/PI3K, thus could help to restrain metastasis.

Hypoglycaemic role of wheatgrass and its effect on carbohydrate metabolic enzymes in type II diabetic rats.

Diabetes mellitus (DM) is a leading cause of morbidity and mortality in the world. Insulin resistance and insulin insufficiency is the major factor for the prognosis of type II diabetes. Consistent high glucose level leads to multiple secondary complications in diabetic patients. Hence, hypoglycaemic drugs are of significance for reducing the risk of secondary complications in type II diabetes. Various hypoglycaemic drugs are already available in the market, but they are associated with several side effects. Therefore, traditional herbs have emerged as safer alternative for effective hypoglycaemic treatment. The juvenile grass of common wheat is known as wheatgrass (WG). It is commonly used as a health drink and has potent antioxidant efficacy. It has been used to cure DM in folk medicine. The current study was planned to test the hypoglycaemic effect and pathways regulated by WG on DM. We analysed the glucose and insulin levels in plasma, the activity of glucose oxidative enzymes, hexokinase and glucose 6 phosphate dehydrogenase, in serum and glycogen levels in liver of the male albino Wistar rats. Activity of glucose oxidative enzymes and the levels of insulin and liver glycogen were decreased in rats with diabetes, but they were reversed on treatment with WG. Hence, we conclude that WG can act as a potent anti-hyperglycaemic agent.

Disulfiram and disulfiram-loaded poly-[lactide-co-glycolic acid] nanoparticles modulate metastatic markers and proteasomal activity in hepatocarcinoma Hep3b cell line

Abstract Hepatocellular carcinoma (HCC) results in significantly high mortality rates due to its subtle metastatic expressions. Exorbitant costs of anticancer drugs have lead to the concept of repositioning standard drugs for their anticancer potential. One such antialcoholic drug, disulfiram (DSF), has been reported to show significant cytotoxicity (IC50 6 μM) against hepatocarcinoma cells. Hence, we studied its antimetastatic and proteasome-inhibiting potential to ascertain its efficacy against metastatic hepatocarcinoma. In addition, we also studied the influence of the components of polysorbate 80-stabilised poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) on metastastatic markers and proteasome complexes. A significant reduction in antimigratory assays (0.1 μM) and gelatin zymography (0.5 μM) was observed. A Western blot analysis furthermore confirmed the modulation of metastatic markers. Chymotrypsin-like activity was significantly inhibited at 2 μM of DSF, and in silico docking studies show comparable gliding scores of DSF against standard antimetastatic drugs. Our experiments suggest a significant antimetastatic and proteasome-inhibiting potential of DSF and its loaded NPs. We also infer that polysorbate 80 and PLGA neither show interference with drug behaviour nor inhibit metastasis and proteasome activity, thus behaving as components of a neutral vector. However, they may potentially manipulate the pharmacokinetics of DSF for achieving maximum therapeutic efficacy, in addition to targeted drug delivery.

Hepatoprotective Role of Wheatgrass on Alcohol and ΔPUFA-Induced Oxidative Stress in Rats

ABSTRACT Alcohol abuse is recognized as the most common cause for the development of various abnormalities including liver disease. Excessive free radicals are generated during the metabolism of ethanol. The ingestion of alcohol along with polyunsaturated fatty acid (PUFA) aggravates the production of free radicals and enhances the oxidative stress. Medicinal plants contain active phytocomponents, which are the principal healthcare resources. We aimed to analyze the effect of wheatgrass extract on alcohol and thermally oxidized PUFA (ΔPUFA) induced oxidative stress in male albino Wistar rats. The levels of marker enzymes gamma-glutamyl transferase (GGT), alkaline phosphatase (ALP), lipid peroxidative markers; thiobarbutric acid reactive substances (TBARS) and lipid hydroperoxides (LH), the levels of enzymatic (catalase [CAT], superoxide dismutase [SOD], glutathione peroxidase [GPx]) and nonenzymatic (reduced glutathione [GSH], vitamin E, vitamin C) antioxidants were analyzed in liver to evaluate the effects of wheatgrass. The levels of TBARS and LH were significantly (p ≤ .05) increased in alcohol + ΔPUFA group, which were found to be reduced on treatment with wheatgrass. The levels of both enzymatic and nonenzymatic antioxidants were significantly (p ≤ .05) decreased in alcohol + ΔPUFA group, which were found to be restored on treatment with wheatgrass. From the results obtained, we conclude that wheatgrass protects the liver against alcohol and ΔPUFA induced oxidative stress.