Browsing by Author "Galhena, P.B."

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Anti-hepatocarcinogenic Ayurvedic herbal remedy reduces the extent of diethylnitrosamine-induced oxidative stress in rats
(Medknow Publications, 2009) Galhena, P.B.; Thabrew, I.; Mayuri, G.T.; Vasanthi, A.H.R.
Previous studies have shown that a decoction prepared from a mixture of Nigella sativa seeds, Hemidesmus indicus roots and Smilax glabrarhizome, has the potential to protect against diethylnitrosamine (DEN) induced hepatocarcinogenic changes in rats. The present in-vivo investigation with Wistar rats was conducted to determine whether the treatment with above decoction (6 g / kg / day) for a period of 10 weeks can provide protection against DEN (200 mg / kg by a single i.p. injection) mediated changes in (a) lipid peroxidation, (b) glutathione (GSH) concentration, (c) activities of the antioxidant enzymes superoxide dismutase (SOD) and glutathione peroxidase (GPx), and (d) activity of the phase II detoxification enzyme glutathione S-transferase (GST). DEN administration resulted in a significant (p<0.05) enhancement (+ 46.2%) of lipid peroxidation (as assessed by formation of thiobarbituric acid reactive substances, TBARS), accompanied by a decreased GSH concentration (- 21.7% in liver; - 5.9% in blood), and activities of SOD (- 34.3%), GPx (- 49.1%) and GST (- 19.38%). Administration of the decoction to DEN treated animals resulted in a significant (p<0.05) reduction in TBARS production, along with a restoration towards the normal levels, of the other biochemical parameters evaluated. The overall results obtained suggests that, protection against DEN - mediated changes in oxidative stress and enhancement of the activities of enzymes participating in carcinogen detoxification are possible mechanisms utilized by the decoction to mediate its anti-hepatocarcinogenic action.
Anti-Inflammatory activity is a possible mechanism by which the polyherbal formulation comprised of Nigella sativa (Seeds), Hemidesmus indicus (Root), and Smilax glabra (Rhizome) mediates its antihepatocarcinogenic effects
(Hindawi Pub, 2012) Galhena, P.B.; Samarakoon, S.R.; Thabrew, M.I.; Weerasinghe, G.A.; Thammitiyagodage, M.G.; Ratnasooriya, W.; Tennekoon, K.H.
The present study investigated the anti-inflammatory effects of a polyherbal decoction comprised of Nigella sativa, Hemidesmus indicus, and Smilax glabra in order to justify its claimed antihepatocarcinogenic activity. Activation of hepatic nuclear factor-kappa B (NF-κB), IκB kinase (IKK α/β) proteins, and TNFα and IL-6 expression was investigated in diethylnitrosamine- (DEN-) induced C3H mice-bearing early hepatocarcinogenic changes. Acute phase inflammatory response was evaluated by carrageenan-induced rat paw edema formation. Anti-inflammatory mechanisms were also assessed by determining effect on (a) membrane stabilization, (b) nitric oxide (NO) inhibitory activity, and (c) inhibition of leukocyte migration. A significant inhibition of the paw edema formation was observed in healthy rats as well as in rats bearing early hepatocarcinogenic changes with oral administration of the decoction. As with the positive control, indomethacin (10 mg/kg b.w.) the inhibitory effect was pronounced at 3rd and 4th h after carrageenan injection. A notable IKK α/β mediated hepatic NF-κB inactivation was associated with a significant hepatic TNFα down regulation among mice-bearing hepatocarcinogenic changes subjected to decoction treatment. Inhibition of NO production, leukocyte migration, and membrane stabilization are possible mechanisms by which anti-inflammatory effect is mediated by the decoction. Overall findings imply that anti-inflammatory activity could
A Comparison of the cytotoxic potential of standardized aqueous and ethanolic extracts of a polyherbal mixture comprised of Nigella sativa (seeds), Hemidesmus indicus (roots) and Smilax glabra (rhizome)
(Medknow Publications, 2010) Samarakoon, S.R.; Thabrew, I.; Galhena, P.B.; de Silva, D.; Tennekoon, K.H.
BACKGROUND: A decoction (hot-water extract) comprised of Nigella sativa (seeds), Hemidesmus indicus (roots), and Smilax glabra (rhizome) has been reported to prevent chemically-induced hepatocarcinogenic changes in rats and to exert significant cytotoxic effects on human hepatoma (HepG2) cells. However, the decoction used in previous studies to determine cytotoxicity was not standardized. Further, during preparation of pharmaceuticals for clinical use, it is more convenient to use an ethanolic extract. Therefore this study was carried out to (a) develop standardizedaqueous and ethanolic extracts of the plant mixture (N. sativa, H. indicus, and S. glabra) used in the preparation of the original decoction, and (b) compare the cytotoxic effects of these two extracts by evaluating cytotoxicity to the human hepatoma (HepG2) cell line. METHODS: Aqueous and ethanolic extracts have been standardized by evaluating organoleptic characters, physicochemical properties, qualitative and quantitative analysis of chemical constituents, and analysis of High Performance Liquid Chromatography (HPLC) and Thin Layer Chromatography (TLC) profiles. Cytotoxic potentials of the above standardized extracts were compared by evaluating their effects on the survival and overall cell activity of HepG2 cells by use of the 3-(4, 5-dimethylthiazol-2yl) -2, 5 - biphenyl tetrazolium bromide (MTT) and Sulphorhodamine B (SRB) assays. RESULTS: Results from MTT and SRB assays demonstrated that both extracts exerted strong dose-dependent in vitro cytotoxicity to HepG2 cells. The standardized aqueous extract showed a marginally (though significantly, P<0.05) higher cyotoxic potential than the ethanolic extract. Thymoquinone, an already known cytotoxic compound isolated from N. sativa seeds was only observed in the standardized ethanolic extract. Thus, compounds other than thymoquinone appear to mediate the cytotoxicity of the standardized aqueous extract of this poly-herbal preparation. CONCLUSION: It may be concluded that results obtained in the present study could be used as a diagnostic tool for the correct identification of these aqueous or ethanolic extracts and would be useful for the preparation of a standardized pharmaceutical product that may be used in the future for clinical therapy of hepatocellular carcinoma.
Cytotoxic and apoptotic effect of the decoction of the aerial parts of Flueggea leucopyrus on human endometrial carcinoma (AN3CA) cells
(Pharmacotherapy Group, University of Benin, Nigeria, 2014) Samarakoon, S.R.; Kotigala, S.B.; Gammana-Liyanage, I.; Thabrew, I.; Tennekoon, K.H.; Siriwardana, A.; Galhena, P.B.
PURPOSE: To evaluate the anti-cancer potentials of a decoction of Flueggea. leucopyrus (Willd.) on human endometrial carcinoma (AN3CA) cells. METHODS: Decoction was prepared by boiling 60 g of the ground plant material in 1.6 L of distilled water for about 3 h to reduce the volume to 200 mL and then freeze dried. The effect of the decoction on AN3CA cells was determined by evaluating its cytotoxicity by 3-(4, 5-dimethylthiazol-2yl) -2, 5-biphenyl tetrazolium bromide (MTT) and sulphorhodamine B (SRB) assays, as well as its ability to modulate apoptosis (microscopic observation of morphological changes, DNA fragmentation and caspase activity). The antioxidant activity of the decoction was also determined by DPPH assay, and its total polyphenolic and flavonoid content. RESULTS: The decoction exerted a significant dose-dependent cytotoxicity on AN3CA cells as evident from MTT assay IC50 values of 22.09 and 14.60 µg/mL at 24 and 48 h post-incubation, respectively; and SRB assay IC50 values of 28.60 and 15.09 µg/mL at 24 and 48 h post-incubation, respectively. The decoction also enhanced apoptosis as shown by enhanced DNA fragmentation, microscopic observation of nuclear condensation, fragmentation and apoptotic bodies and enhanced caspase 3 and 9 activities, as well as moderately increased radical scavenging activity. CONCLUSION: The cytotoxic and apoptotic effects demonstrated by F. leucopyrus (Willd.) decoction provide supportive evidence for the ethnomedicinal use of this plant for cancer therapy. Copyright of Tropical Journal of Pharmaceutical Research
Effect of standardized decoction of Nigella sativa seed, Hemidesmus indicus root and smilax glabra rhizome on the expression of p53 and p21 genes in human hepatoma cells (HepG2) and mouse liver with chemically-induced hepatocarcinogenesis
(University of Benin, 2012) Samarakoon, S.R.; Thabrew, I.; Galhena, P.B.; Tennekoon, K.H.
PURPOSE: To evaluate in vitro (using human hepatoma HepG2 cells) and in vivo (using mouse liver with diethlynitrosamine (DEN)-induced hepatocarcinogenesis) effect of a standardized decoction on the expression of p53 (tumour suppressor) and p21 (cyclin kinase inhibitor) genes with the long-term goal of developing the formulation into a globally acceptable therapy for hepatocellular carcinoma (HCC). METHODS: The effect of the decoction on (a) mRNA and (b) protein expression of p53 and p21 genes in HepG2 cells and mouse livers with DEN-induced early hepatocarcinogenesis were evaluated by (a) reverse transcription PCR (RT-PCR) and (b) immunohistochemical and Western blot analysis, respectively. RESULTS: The results demonstrated that the decoction significantly (p < 0.001) enhanced the expression of p53 and p21 genes in a time-and dose-dependent manner in HepG2 cells. A dose of 75 μg/ml significantly increased p53 mRNA at 24 and 48 h and p21 mRNA at 12, 24, 48 h of incubation with the decoction (p < 0.01). Induction of hepatocarcinogenesis in mice significantly increased hepatic expression of both p53 and p21 compared to distilled water control (p < 0.001), while treatment with the decoction further enhanced expression of both genes in DEN-induced hepatocarcinogenesis (p < 0.01). CONCLUSION: Overall, the findings demonstrate that the decoction may mediate its reported antihepatocarcinogenic effect, at least in part, through the modulating activities of genes involved in tumour suppression and cell cycle arrest. All rights reserved. © Pharmacotherapy Group. All rights reserved.
Effect of standardized decoction of Nigella sativa seed, Hemidesmusindicus root and Smilax glabra rhizome on the expression of p53 and p21 genes in human hepatoma cells (HepG2) and mouse liver with chemically-induced hepatocarcinogenesis
(Pharmacotherapy Group, University of Benin, Nigeria, 2012) Samarakoon, S.R.; Thabrew, I.; Galhena, P.B.; Tennekoon, K.H.
Purpose: To evaluate in vitro (using human hepatoma HepG2 cells) and in vivo (using mouse liver with diethlynitrosamine (DEN)-induced hepatocarcinogenesis) effect of a standardized decoction on the expression of p53 (tumour suppressor) and p21 (cyclin kinase inhibitor) genes with the long-term goal of developing the formulation into a globally acceptable therapy for hepatocellular carcinoma (HCC). Methods: The effect of the decoction on (a) mRNA and (b) protein expression of p53 and p21 genes in HepG2 cells and mouse livers with DEN-induced early hepatocarcinogenesis were evaluated by (a) reverse transcription PCR (RT-PCR) and (b) immunohistochemical and Western blot analysis, respectively. Results: The results demonstrated that the decoction significantly (p < 0.001) enhanced the expression of p53 and p21 genes in a time- and dose-dependent manner in HepG2 cells. A dose of 75 Âµg/ml significantly increased p53 mRNA at 24 and 48 h and p21 mRNA at 12, 24, 48 h of incubation with the decoction (p < 0.01). Induction of hepatocarcinogenesis in mice significantly increased hepatic expression of both p53 and p21 compared to distilled water control (p < 0.001), while treatment with the decoction further enhanced expression of both genes in DEN-induced hepatocarcinogenesis (p < 0.01). Conclusion: Overall, the findings demonstrate that the decoction may mediate its reported antihepatocarcinogenic effect, at least in part, through the modulating activities of genes involved in tumour suppression and cell cycle arrest.
The gastroprotective effect of ethyl acetate fraction of hot water extract of Trichosanthes cucumerina Linn and its underlying mechanisms
(BioMed Central, 2017) Karunakaran, R.; Ira Thabrew, M.; Thammitiyagodage, G.M.; Galhena, P.B.; Menuka Arawwawala, L.D.A.
BACKGROUND: Antacids, anticholinergic drugs, histamine H2- receptor antagonists and irreversible proton pump inhibitors have been used for the treatment of gastric ulcers. However, prolonged use of these drugs may lead to series of adverse effects such as diarrhea, headache, rash, hypertension, muscular and joint pain. Therefore, there is an urgent need of more effective and safer treatments with fewer side effects. The aim of the present study was to scientifically evaluate the gastroprotective activity of fractions of the hot water extract of Trichosanthes cucumerina Linn (Family: Cucurbitaceae) aerial parts with a view to identifying the fraction with the best gastroprotective activity and the possible mechanism/s by which this fraction exert gastroprotection. METHODS: Gastroprotective activity of hexane fraction (HF), ethyl acetate fraction (EF), butanol fraction (BF) and aqueous fraction (AF) were evaluated by the assessment of ability to reduce the ulcer index in ethanol-induced rat model and the mode of action by which the most active fraction mediating gastroprotection. RESULTS: EF showed the maximum gastroprotection effect followed by BF and AF. EF (75 mg/kg) exhibited significantly higher gastroprotection compared to the reference drugs. Further investigations with two lower doses of EF confirmed that EF can mediated a significant and dose dependent gastroprotection. The rats treated with the EF showed significant reduction in free acidity (45%), total acidity (by 48%) in the gastric juice, increased the amount of mucus produced by the rat gastro mucosa and potent antihistamine activity (by 25.6%). EF was also rich in phenolic compounds and flavonoids. CONCLUSION: Gastroprotective mechanism of EF is possibly involves inhibition of acidity, elevation in mucus content, inhibition of histamine and antioxidant mechanisms.
In silico identification and in vitro validation of alpha-hederin as a potent inhibitor of Wnt/β-catenin signaling pathway in breast cancer stem cells.
(Springer-Verlag, GmbH, 2024) Saliu, T.P.; Seneviratne, N.N.; Faizan, M.; Rajagopalan, U.; Perera, D.C.; Adhikari, A.; Senathilake, K.S.; Galhena, P.B.; Tennekoon, K.H.; Samarakoon, S.R.
Cancer stem cells (CSCs) play a vital role in metastasis, recurrence and chemoresistance in breast cancer. β-catenin, which is a frequently over activated protein in CSCs, binds to T-cell factor/lymphoid enhancer factor (Tcf/Lef) family transcription factors leading to ectopic expression of Wnt pathway responsive genes necessary for the maintenance and action of CSCs. With the aim of identifying a small molecules that can effectively eliminate CSCs, molecular docking studies were performed against the Tcf/Lef binding hotspot on β-catenin using a library of 100 natural or synthetic small molecules. Small molecule ligands giving docking energy better than - 7 kcal/mol were further investigated by binding interactions analysis and molecular dynamics (MD) simulations. These compounds were then investigated in vitro, for cytotoxicity against CSCs isolated from MDA-MB-231 triple negative breast cancer cells. Alpha-hederin (AH) was identified as the only compound in the selected library that has cytotoxicity against breast CSCs. AH was further investigated for it's ability to regulate Wnt pathway target genes (Cyclin D1 and CD44)and the tumor suppressor p53by real-time quantitative PCR. Absorption, distribution, metabolism, excretion and toxicity properties of the AH was predicted in silico. AH significantly down regulated the transcription of Cyclin D1 and CD44 while up-regulating the transcription of p53. AH was predicted to have acceptable drug likeness. Although AH is currently known to inhibit the growth of various cancer cells in vitro, present study demonstrated for the first time that it is a potent inhibitor of Wnt/β-catenin signaling pathway and induce apoptosis in breast CSCs.
In vitro anticancer effect of gedunin on human teratocarcinomal (NTERA-2) cancer stem-like cells
(Hindawi, 2017) Tharmarajah, L.; Samarakoon, S.R.; Ediriweera, M.K.; Piyathilaka, P.; Thennakoon, K.H.; Senathilake, K.S.; Rajagopalan, U.; Galhena, P.B.; Thabrew, I.
Gedunin is one of the major compounds found in the neem tree (Azadirachta indica). In the present study, antiproliferative potential of gedunin was evaluated in human embryonal carcinoma cells (NTERA-2, a cancer stem cell model) and peripheral blood mononuclear cells (PBMCs), using Sulforhodamine (SRB) and WST-1 assays, respectively. The effects of gedunin on expression of heat shock protein 90 (HSP90), its cochaperone Cdc37, and HSP client proteins (AKT, ErbB2, and HSF1) were evaluated by real-time PCR. Effects of gedunin on apoptosis were evaluated by (a) apoptosis associated morphological changes, (b) caspase 3/7 expression, (c) DNA fragmentation, (d) TUNEL assay, and (e) real-time PCR of apoptosis related genes (Bax, p53, and survivin). Gedunin showed a promising antiproliferative effect in NTERA-2 cells with IC50 values of 14.59, 8.49, and 6.55 μg/mL at 24, 48, and 72 h after incubations, respectively, while exerting a minimal effect on PBMCs. Expression of HSP90, its client proteins, and survivin was inhibited and Bax and p53 were upregulated by gedunin. Apoptosis related morphological changes, DNA fragmentation, and increased caspase 3/7 activities confirmed the proapoptotic effects of gedunin. Collectively, results indicate that gedunin may be a good drug lead for treatment of chemo and radiotherapy resistant cancer stem cells.
Investigation of the antioxidant activity of a herbal decoction with anti-carcinogenic properties
(Sri Lanka Association for the Advancement of Science, 2007) Galhena, P.B.; Thabrew, M.I.; Thammitiyagodage, M.G.
Modulation of apoptosis in human hepatocellular carcinoma (HepG2 cells) by a standardized herbal decoction of Nigella sativa seeds, Hemidesmus indicus roots and Smilax glabra rhizomes with anti- hepatocarcinogenic effects
(BioMed Central, 2012) Samarakoon, S.R.; Thabrew, I.; Galhena, P.B.; Tennekoon, K.H.
BACKGROUND A standardized poly-herbal decoction of Nigella sativa seeds, Hemidesmus indicus roots and Smilax glabra rhizomes used traditionally in Sri Lanka for cancer therapy has been demonstrated previously, to have anti-hepatocarcinogenic potential. Cytotoxicity, antioxidant activity, anti-inflammatory activity, and up regulation of p53 and p21 activities are considered to be some of the possible mechanisms through which the above decoction may mediate its anti-hepatocarcinogenic action. The main aim of the present study was to determine whether apoptosis is also a major mechanism by which the decoction mediates its anti-hepatocarcinogenic action. METHODS Evaluation of apoptosis in HepG2 cells was carried out by (a) microscopic observations of cell morphology, (b) DNA fragmentation analysis, (c) activities of caspase 3 and 9, as well as by (d) analysis of the expression of pro-apoptotic (Bax) and anti-apoptotic (Bcl-2) proteins associated with cell death. RESULTS The results demonstrated that in HepG2 cells, the decoction can induce (a) DNA fragmentation and (b) characteristic morphological changes associated with apoptosis (nuclear condensation, membrane blebbing, nuclear fragmentation and apoptotic bodies). The decoction could also, in a time and dose dependent manner, up regulate the expression of the pro-apoptotic gene Bax and down regulate expression of anti-apoptotic Bcl-2 gene (as evident from RT-PCR analysis, immunohistochemistry and western blotting). Further, the decoction significantly (p < .001) enhanced the activities of caspase-3 and caspase-9 in a time and dose dependent manner. CONCLUSIONS Overall findings provide confirmatory evidence to demonstrate that the decoction may mediate its reported anti-hepatocarcinogenic effect, at least in part, through modulation of apoptosis.
Protective effect of a polyherbal aqueous extract comprised of nigella sativa (Seeds), hemidesmus indicus (Roots), and smilax glabra (Rhizome) on bleomycin induced cytogenetic damage in human lymphocytes
(Hindawi Pub. Co., 2017) Galhena, P.B.; Samarakoon, S.S.R.; Thabrew, M.I.; Paul, S.F.D.; Mani, C.
This study was carried out to determine the chemoprotective potential of a polyherbal aqueous decoction comprised of Nigella sativa (seeds), Hemidesmus indicus (roots), and Smilax glabra (rhizome) against bleomycin induced cytogenetic damage in human lymphocytes. Isolated peripheral blood lymphocytes (PBLs) were exposed to bleomycin at a dose of 40 µg/mL for 2 hrs in the presence or absence of different doses of the decoction (100, 300, and 600 µg/mL). Modulatory effect of the decoction on bleomycin induced cytogenetic damage was evaluated by (a) degree of chromosomal aberrations (CA), (b) formation of micronuclei (MN), and (c) induction of γH2AX foci in lymphocytes exposed to bleomycin. Lymphocytes pretreated with the decoction showed that a significant reduction (p < 0.05) in bleomycin induced (a) stable and unstable chromosome aberrations (CA), (b) MN formation, and (c) formation of γH2AX foci, when compared to lymphocytes treated only with bleomycin. The decoction by itself did not induce any significant cytogenetic damage in PBLs. Overall results of the present study confirm that the decoction can attenuate the cytogenetic damage mediated by bleomycin in human PBLs.