Kamani Hemamala Tennekoon

Effect of Momordica charantia fruit juice on streptozotocin-induced diabetes in rats.

The oral hypoglycaemic activity of Momordica charantia fruit juice was investigated in rats with streptozotocin-induced diabetes. Oral administration of the juice (10 ml/kg for 30 days) did not show a significant effect, either acute or cumulative, on the ability to tolerate an external glucose load. The glycosylated haemoglobin concentrations were significantly elevated in both juice-treated and untreated diabetic rats and there was no significant difference between the two groups. Viable beta-cells capable of secreting insulin upon stimulation appear to be required for M. charantia to exert its oral hypoglycaemic activity.

Possible hepatotoxicity of Nigella sativa seeds and Dregea volubilis leaves

Aqueous extracts of the seeds of Nigella sativa and mature leaves of Dregea volubilis were administered orally under light ether anaesthesia to male Sprague-Dawley rats for 14 days. Key hepatic enzyme concentrations and histopathological changes in the liver in both treatment groups at the end of 14 days were compared with a control group which received distilled water under identical conditions for 30 days and with a group of normal animals. Serum gamma-glutamyl transferase concentrations were significantly increased in both extract groups while serum alkaline phosphatase concentrations were significantly increased following administration of only D. volubilis when compared with either the control or the normal group. Serum alanine aminotransferase concentrations were significantly increased in both extract groups when compared with the normal group but not with the control group. Degenerative changes in hepatocytes were seen following administration of D. volubilis while consistent significant histopathological changes were not evident following administration of N. sativa.

Effect of Momordica charantia on key hepatic enzymes

The effect of Momordica charantia on certain key hepatic enzymes was investigated using male Sprague-Dawley rats as the animal model. Fruit juice and seed extract of Momordica charantia were administered orally at a daily dose of 1 ml/100 g body weight for 30 days under light ether anaesthesia while the control group received equivalent amounts of distilled water under identical conditions (n = 10 in each case). Serum gamma-glutamyl transferase (P < 0.001) and alkaline phosphatase (P < 0.01-0.001) concentrations were found to be significantly elevated following oral administration of both the fruit juice and the seed extract. Consistent significant histopathological changes in the liver were not observed in either treatment group although the prevalence of dilatation and/or congestion of the central vein sinusoidal system appeared to be twice as high following fruit juice treatment than in the other 2 groups. Thus, Momordica charantia may either contain hepatotoxins capable of causing cellular damage at the molecular level without causing significant histopathological changes or the plant may have an enzyme inducing effect.

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)

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 standardized aqueous 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.

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

BackgroundA 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.MethodsEvaluation 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.ResultsThe 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.ConclusionsOverall findings provide confirmatory evidence to demonstrate that the decoction may mediate its reported anti-hepatocarcinogenic effect, at least in part, through modulation of apoptosis.

Novel sequence variants and a high frequency of recurrent polymorphisms in BRCA1 gene in Sri Lankan breast cancer patients and at risk individuals

BackgroundBreast Cancer is the most commonly diagnosed cancer among Sri Lankan women. Germline mutations in the susceptibility genes BRCA1 and BRCA2 in hereditary breast/ovarian cancer, though low in prevalence, are highly penetrant and show geographical variations. There have been only a few reports from Asia on mutations in BRCA1/2 genes and none from Sri Lanka.MethodsA total of 130 patients with (N = 66) and without (N = 64) a family history of breast cancer, 70 unaffected individuals with a family history of breast cancer and 40 control subjects were analysed for BRCA1 mutations. All but exon 11 were screened by single strand conformation analysis (SSCP) and heteroduplex analysis. PCR products which showed abnormal patterns in SSCP were sequenced. Exon 11 was directly sequenced.ResultsNineteen sequence variants were found in BRCA1 gene. Two novel deleterious frame-shift mutations; c.3086delT/exon11 (in one patient) and c.5404delG/exon21 (in one patient and two of her family members) were identified. A possibly pathogenic novel missense mutation (c.856T>G/exon 11) and three novel intronic variants (IVS7+36C>T, IVS7+41C>T, IVS7+49del15) were characterised. Ten previously reported common polymorphisms and three previously reported intronic variants were also observed.ConclusionAfter screening of 66 patients with family history and 64 sporadic breast cancer patients, 2 deleterious mutations (c.3086delT and c.5404delG) in two families were identified and two more possibly pathogenic mutations (c.856T>G and IVS17-2A>T) in two families were identified.Data baseBRCA1 - Gene Bank: Accession # U14680 Version # 14680.1

A study of the potential anticancer activity of Mangifera zeylanica bark: Evaluation of cytotoxic and apoptotic effects of the hexane extract and bioassay-guided fractionation to identify phytochemical constituents

The present study investigated the potential anticancer activity of the bark of Mangifera zeylanica, an endemic plant in Sri Lanka that has been traditionally used for cancer therapy. Cytotoxic and apoptotic effects were investigated in vitro using sulphorodamine assay, acridine orange and ethidium bromide staining, caspase-3 and −7 activity, DNA fragmentation and reverse transcription-quantitative polymerase chain reaction in estrogen receptor positive MCF-7 and triple-negative MDA-MB-231 breast cancer cell lines, SKOV-3 ovarian cancer cell line and MCF-10A normal mammary epithelial cells. Hexane extract demonstrated increased levels of cytotoxicity in cancer cells (IC50, 86.6–116.5 µg/ml) compared with normal cells (IC50, 217.2 µg/ml). Chloroform extract demonstrated increased cytotoxicity to normal cells (IC50, 92.9 µg/ml) compared with cancer cells (IC50, 280.1–506.5 µg/ml). Exposure to the hexane extract led to morphological changes characteristic of apoptosis and DNA fragmentation in the three cancer cell lines. Caspase-3 and −7 were significantly activated in MDA-MB-231 and SKOV-3 cells, indicating the occurrence of caspase-dependent apoptosis in these cells, and caspase-independent apoptosis in MCF-7 cells. Furthermore, upregulation of proapoptotic Bcl-2-associated X protein occurred in the three cancer cell lines, and antiapoptotic survivin was downregulated in MCF-7 and SKOV-3 cells; by contrast, tumor protein p53 was upregulated only in MCF-7 cells, suggesting p53-mediated apoptosis in MCF-7 cells and p53-independent apoptosis in the remaining cancerous cell lines. In addition, fraction M1 obtained from bioactivity-guided fractionation of the hexane extract demonstrated increased cytotoxicity in cancer cells (IC50, 15.4–38.7 µg/ml) compared with normal cells (IC50, 114.6 µg/ml), with the highest cytotoxicity observed in MDA-MB-231 triple-negative breast cancer cells. The hexane extract of M. zeylanica bark contained polyphenols and flavonoids, and caused free radical scavenging activity. Its gas chromatography-mass spectrometry profile revealed the presence of long-chain hydrocarbons, including β-sitosterol and β-amyrin. Fraction M1 contained seven unknown compounds and a small number of known non-cytotoxic compounds. Collectively, results obtained in the present study indicate that the hexane extract of M. zeylanica bark mediates cytotoxic activities through induction of apoptosis in three cancer cell lines; thus, the hexane extract may be used to isolate novel anti-cancer compounds.

Association of -2548 G/A Polymorphism in the Leptin Gene with Preeclampsia/ Pregnancy-Induced Hypertension

Objectives. To investigate −2548 G/A polymorphism of LEP gene, plasma leptin and soluble leptin receptor in preeclampsia/pregnancy induced hypertension. Methods. Sixty-two patients and 63 controls (normal pregnancies) studied in the third trimester. Leptin and soluble leptin receptor measured by enzyme immunosorbent assay and −2548 G/A polymorphism by polymerase chain reaction followed by restriction enzyme digestion. Results. Frequency of the AA genotype was significantly (p < 0.001) higher in patients. The A allele conferred a relative risk of 1.67 for the disease (p < 0.0001). Preeclampsia/pregnancy induced hypertension significantly (p < 0.001 to 0.0001) affected leptin and soluble leptin receptor concentrations, free leptin index and leptin normalized to body mass index. Genotype significantly (p < 0.05) influenced only the soluble leptin receptor concentrations. Conclusions. Preeclampsia/pregnancy induced hypertension appears to be associated with higher circulating leptin and lower SLR levels, and with the AA genotype of −2548 G/A polymorphism of the leptin gene.

New halogenated constituents from Mangifera zeylanica Hook.f. and their potential anti-cancer effects in breast and ovarian cancer cells.

ETHNOPHARMACOLOGICAL RELEVENCE Mangifera zeylanica Hook.f. (Anacardiaceae) is a plant endemic to Sri Lanka. Its bark has been used in traditional and Ayurvedic medicine for the treatment of various diseases including some cancers. AIM OF THE STUDY This study was planned to isolate and identify potentially cytotoxic compounds from the bark of M. zeylanica, which may have contributed to its ethno pharmacological use in the treatment of cancer. MATERIALS AND METHODS The chloroform extract of M. zeylanica bark which is cytotoxic to breast and ovarian cancer cells was fractionated using column chromatography and preparative reversed phase high performance liquid chromatography to isolate four compounds. Structures of the isolated compounds were elucidated by means of (1)H- and (13)C NMR spectroscopy, and mass spectrometric techniques. Cytotoxic potential of the isolated compounds was tested in MDA-MB-231 (triple negative breast cancer), MCF-7 (estrogen receptor positive breast cancer), SKOV-3 (ovarian epithelial cancer) and MCF-10A (normal mammary epithelial) cells by SRB assay. Human cancer drug target real-time PCR array was carried out to analyze regulation of possible cancer drug target genes in compound 2 treated triple negative breast cancer cells. DPPH radical scavenging and caspase 3 and 7 induction in response to isolated compounds were also studied. RESULTS Two new halogenated compounds, bromomangiferic acid (1), and chloromangiferamide (2) along with two known compounds quercetin (3), and catechin (4), were isolated from the bark of Mangifera zeylanica for the first time. Interestingly, chloromangiferamide showed cytotoxicity only to triple negative breast cancer cells [IC50:73.19±0.87µM (24h), 56.29±0.86µM (48h)] with no cytotoxicity to other two cancer cell lines or to normal mammary epithelial cells. Quercetin and catechin were cytotoxic to all three cancer cell lines while bromomangiferic acid had no effect. Chloromangiferamide significantly regulated expression of genes associated with apoptosis, drug metabolism, cell cycle, receptor tyrosine kinase signaling, protein kinases, histone deacetylases, growth factors and receptors, topoisomerases, PI-3 kinases and phosphatases in triple negative breast cancer cells. CONCLUSION Selective cytotoxic activity in triple negative breast cancer cells and regulation of some cancer drug target genes by chloromangiferamide indicate that it can be used to develop a potential chemotherapeutic agent for triple negative breast cancer cells.

Lactational amenorrhea/anovulation and some of their determinants: a comparison of well-nourished and undernourished women.

OBJECTIVE To study the effect of maternal nutritional status and some other possible determinants on lactational amenorrhea/anovulation. DESIGN Prospective matched-pairs study. SETTING Postpartum wards and community and academic settings. PATIENT(S) Thirty matched pairs of otherwise healthy, well-nourished (body mass index > or = 26.00 kg/m2) and undernourished (body mass index < or = 19.00 kg/m2) postpartum women were selected. INTERVENTION(S) Infant feeding pattern was recorded weekly, and infant weights, maternal body mass index, and maternal PRL levels were estimated every 4 weeks until resumption of menstruation. Ovulatory activity was determined using urinary estrone and pregnanediol glucuronide concentrations. MAIN OUTCOME MEASURE(S) Time of resumption of menstruation, regular and ovulatory. RESULT(S) Well-nourished women resumed regular menstruation significantly earlier than undernourished women but resumed ovulatory menstruation at almost the same time. Undernourished women had fewer anovulatory cycles preceding first postpartum ovulation and a higher prevalence of formula feeding. Effect of body mass index on lactational amenorrhea became nonsignificant when nonintroduction of formula feeds, maternal age, and socioeconomic status were controlled for. CONCLUSION(S) Improved maternal nutritional status has no significant effect on fertility: ovulation is not advanced despite early resumption of regular menstruation.