Simultaneous exposure to electromagnetic field from mobile phone and unimpeded fructose drinking during pre-, peri-, and post-pubertal stages perturbs the hypothalamic and hepatic regulation of energy homeostasis by early adulthood: experimental evidence.

The present-day children-adolescents ubiquitously use the mobile phones and unrestrictedly consume fructose-laden diet. Unfortunately, a rise in the incidence of insulin resistance and fatty liver syndrome in young adults has also been recorded. To delineate a possible correlate, the effect of exposure to electromagnetic field (EMF) from the mobile phone and unrestricted fructose intake during pre-, peri-, and post-pubertal stages of development on orexigenic and anorexigenic signals arising from the hypothalamus and liver of rats is investigated here. The study design included four arms, i.e., "Normal", "Exposure Only (ExpO)", "Fructose Only (FruO)", and "Exposure with Fructose (EF)", wherein weaned rats received either "normal chow and drinking water" or "normal chow and fructose (15%) drinking solution" in presence and absence of EMF exposure (2 h/day) for 8 weeks. The results indicate that the total calories consumed by the EF were higher by early adulthood than normal, possibly under the influence of the raised levels of the orexigenic hormone, i.e., ghrelin, and it reflected as raised rate of weight gain. At early adulthood, the EF recorded mitigated response and sensitivity of insulin. Despite EF being a "fed-state", both centrally and peripherally, the glycolysis was restrained, but the gluconeogenesis was raised. Additionally, the altered lipid profile and the glycogen levels indicate that the EF developed fatty liver. The energy homeostasis of the EF was compromised as evidenced by (a) reduced expression of the glucosensors-GLUT2 and glucokinase in the hypothalamus and liver and (b) reduced expression of the cellular energy regulator-AMPK, orexigenic peptide-NPY, and anorexigenic peptide-POMC in the hypothalamus. Taken together, the present study evidences that the exposure to EMFfrom the mobile phone and unrestricted fructose intake during childhood-adolescence impairs the central and peripheral pathways that mediate the glucosensing, glucoregulation, feeding, and satiety behavior by early adulthood.

Molecular and Metabolic Markers of Fructose Induced Hepatic Insulin Resistance in Developing and Adult Rats are Distinct and Aegle marmelos is an Effective Modulator.

The time course of pathogenesis of fructose mediated hepatic insulin resistance (HepIR) is not well-delineated and we chronicle it here from post-weaning to adulthood stages. Weaned rats were provided for either 4 or 8 weeks, i.e., upto adolescence or adulthood, chow + drinking water, chow + fructose, 15% or chow + fructose, 15% + hydroalcoholic extract of leaves of Aegle marmelos (AM-HM, 500 mg/kg/d, po) and assessed for feed intake, fructose intake, body weight, fasting blood sugar, oral glucose tolerance test, HOMA-IR, insulin tolerance test and lipid profile. Activities of enzymes (glucose-6-phosphatase, hexokinase, phosphofructokinase, aldehyde dehydrogenase), hormones (leptin, ghrelin, insulin), insulin signaling molecules (Akt-PI3k, AMPK, JNK) hallmarks of inflammation (TNF-α), angiogenesis (VEGF), hypoxia (HIF-1), lipogenesis (mTOR) and regulatory nuclear transcription factors of de novo lipogenesis and hepatic insulin resistance gene (SREBP-1, FoxO1) that together govern the hepatic fructose metabolism, were also studied. The effect of fructose-rich environment on metabolic milieu of hepatocytes was confirmed using (human hepatocellular carcinoma) HepG2 cells. Using in vitro model, fructose uptake and glucose output from isolated murine hepatocytes were measured to establish the HepIR under fructose environment and delineate the effect of AM-HM. The leaves from the plant Aegle marmelos (L) Correa were extracted, fractionated and validated for rutin content using LC-MS/MS. The rutin content of extract was quantified and correlated with oral pharmacokinetic parameters in rat. The outcomes of the study suggest that the molecular and metabolic markers of fructose induced HepIR in developing and adult rats are distinct. Further, AM-HM exerts a multi-pronged attack by raising insulin secretion, augmenting insulin action, improving downstream signaling of insulin, reducing overall requirement of insulin and modulating hepatic expression of glucose transporter (Glut2). The butanol fraction of AM-HM holds promise for future development.

Aegle marmelos impedes onset of insulin resistance syndrome in rats provided with drinking fructose from weaning to adulthood stages of development - a mechanistic study.

In this study, we explored the effect of aqueous extract of leaves of Aegle marmelos (AM) on hepatic carbohydrate metabolism and insulin downstream signalling in rats given fructose (15%) in drinking water from weaning to adulthood. Wistar albino rats (4 weeks old) were randomly divided into normal control (NC), fructose control (FC), and treatment (AMT) groups and were fed for a period of 8 weeks the following diets: chow + water, chow + fructose (15%), and chow + fructose (15%) + AM (500 mg/kg per day, p.o.), respectively. Compared with the NC group, the FC group was found to have significantly (p < 0.05) raised levels of fasting blood glucose, lipid, visceral mass, plasma insulin and leptin, glycogen, and gluconeogenesis enzyme but decreased glycolytic enzyme activity. Raised levels of glucose transporter 2 protein but decreased activity of phosphatidylinositol-3-kinase (PI3K/Akt) and Janus kinase - signal transducer and activator of transcription-3 (JAK-STAT3) in hepatic tissue indicate a state of insulin and leptin resistance in the FC group. A significant (p < 0.05) lowering of physical and glycemic parameters, strengthening of the hepatic glycolytic pathway over the gluconeogenic pathway, and upregulation of the PI3K/Akt and JAK-STAT3 pathways was observed in the AMT group, as compared with the FC group. For the first time, the mechanism underlying the development of insulin resistance syndrome is delineated here, along with the potential of A. marmelos to impede it.

Investigation of anti-asthmatic potential of Kanakasava in ovalbumin-induced bronchial asthma and airway inflammation in rats.

ETHNO-PHARMACOLOGICAL RELEVANCE: Kanakasava is an Indian traditional Ayurvedic formulation containing Datura (Datura metel), Vasaca (Adhatoda vasica), Dhataki (Woodfordia fruticosa) and Grape (Vitis vinifera) extracts as major constituents and used to treat pulmonary diseases including coughing, breathing difficulty and asthma. The present study was designed to assess the safety and therapeutic efficacy of Kanakasava against ovalbumin-induced bronchial asthma and related airway inflammation in rats due to lack of evidence based therapeutic efficacy data. MATERIAL AND METHODS: Male wistar rats were sensitized with allergen (ovalbumin, 40mg/rat+aluminum hydroxide, 2.0mg/rat) and treated orally with standard dexamethasone (2.5mg/kg, b.w.) or Kanakasava (1.23 and 2.46ml/kg, b.w.) from day 1 to day 28. Inflammatory markers, including cell counts and cytokines such as interleukins (IL-4, IL-5, IL-1ß), tumor necrosis factor (TNF-α), leukotriene (LTD-4), immunoglobulin (IgE), nitric oxide and nitrite levels in both blood and broncheo alveolar lavaged fluid (BALF) were analyzed. Abdominal mesentery was studied histologically for mast cell degranulation, whereas lung functions were investigated by spirometer. Method was also developed to quantify gallic acid and ethyl gallate content in Kanakasava by HPTLC for its quality control. RESULTS: None of the rats exhibited mortality and Kanakasava was found to be safe at the tested doses. Treatment with Kanakasava significantly (P<0.01) reversed elevated levels of IgE, cytokines, nitrites and influx of eosinophils and neutrophils in blood and BALF. These findings were further supported by the significant improvement in lung functions (P<0.01) and suppression (P<0.01) of degranulation of mast cells. The content of gallic acid and ethyl gallate in Kanakasava was found to be 1.94% and 0.98%, respectively. CONCLUSION: These findings demonstrated the preventive effect of Kanakasava in allergen induced model of asthma providing scientific basis for its traditional use in Ayurveda, since long time.

Development and validation of High-performance Thin-layer Chromatography Method for Simultaneous Determination of Polyphenolic Compounds in Medicinal Plants.

CONTEXT: Quantitative standardization of plant-based products is challenging albeit essential to maintain their quality. AIMS: This study aims to develop and validate high-performance thin-layer chromatography (HPTLC) method for the simultaneous determination of rutin (Ru), quercetin (Qu), and gallic acid (Ga) from Psidium guajava Linn. (PG) and Aegle marmelos (L.) Correa. (AM) and correlate with antioxidant activity. MATERIALS AND METHODS: The stock solution (1 mg/mL) of standard Ru, Qu, and Ga in methanol: Water (1:1) was serially diluted and spotted (5 µL) on slica gel 60 F254 thin-layer chromatography plates. Toluene: Ethyl acetate: Formic acid: Methanol (3:4:0.8:0.7, v/v/v) was selected as mobile phase for analysis at 254 nm. Hydroalcoholic (1:1) extracts of leaves of PG and AM were fractionated and similarly analyzed. Antioxidant activity was also determined using 2, 2-diphenyl-1-picrylhydrazyl assay. RESULTS: The developed method was robust and resolved Ru, Qu, and Ga at Rf 0.08 ± 0.02, 0.76 ± 0.01, and 0.63 ± 0.02, respectively. The intra-day, interday precision, and interanalyst were <2% relative standard deviation. The limit of detection and limit of quantification for Ru, Qu, and Ga were 4.51, 4.2, 5.27, and 13.67, 12.73, 15.98 ng/spot, respectively. Antioxidant activity (Log 50% inhibition) of PG and AM was 4.947 ± 0.322 and 6.498 ± 0.295, respectively. CONCLUSION: The developed HPTLC method was rapid, accurate, precise, reproducible, and specific for the simultaneous estimation of Ru, Qu, and Ga. SUMMARY: HPTLC method for simultaneous determination and quantification of Rutin, Quercetin and Gallic acid, is reported for quality control of herbal drugs.Abbreviations Used: A: Aqueous fraction; AM: Aegle marmelos L. Correa; B: Butanol fraction; C: Chloroform fraction; EA: Ethyl acetate fraction; Ga: Gallic acid; H: Hexane fraction; HA: Hydroalcoholic extract; HPTLC: High-performance thin-layer chromatography; PG: Psidium guajava; Qu: Quercetin; Ru: Rutin.

Interaction of aqueous leaf extract of Aegle marmelos (L.) Corr. with cholinergic, serotonergic and adrenergic receptors: an ex vivo study.

OBJECTIVES: The aim was to study interaction of aqueous leaf extract of Aegle marmelos (AM) with cholinergic, serotonergic, and adrenergic receptor systems using appropriate rat tissues-ileum, fundus and tracheal chain, respectively. MATERIALS AND METHODS: Cumulative concentration-response curves (CRC) were constructed at various doses on each tissue for AM and respective standard agonist. The CRC was again plotted in presence and absence of respective standard antagonist to confirm the interaction of receptor system and AM. RESULTS: AM induced concentration-dependent contractions in isolated rat ileum (0.2-6.4 mg/ml) and fundus (0.2-3.2 mg/ml) that were inhibited significantly (P < 0.05) in the presence of atropine (10(-7) M) and ketanserin (10(-6) M), respectively. The relaxant effect, produced by AM (0.2 mg/ml) on carbachol (10(-5) M) precontracted rat tracheal chain, was also inhibited significantly (P < 0.05) by propranolol (1 ng/ml). CONCLUSION: It may be concluded that AM possesses agonistic activity on cholinergic, serotonergic and adrenergic receptors.

Qualitative and quantitative assessment of four marketed formulations of brahmi.

This study was conducted with the aim to compare two batches each of four popular commercial formulations of Bacopa monnieri (Brahmi), and report, if any, inter-batch variations. The formulations were procured from local market and analyzed for label specifications, uniformity of weight of capsule, identity, purity and strength parameters (total ash content test, acid insoluble ash content, water soluble extractive, alcohol soluble extractive, loss on drying). Bacoside A, one of the pharmacologically active saponin present in B. monnieri, was quantified in all the formulations using UV-spectrophotometer. In addition each formulation was assessed and compared for variation in biological activity using in vitro test for hemolytic activity using human erythrocytes. The results of the study show that there is a wide variation in the quality and content of herbal drugs marketed by different manufacturers. More importantly this study demonstrates that there exists a bigger challenge of batch-to-batch variation in the quality and content of herbal formulations of the same manufacturer. This challenge of providing standardized formulations is being faced by not any one manufacturing house but by all, and may be attributed firstly to, lack of stringent regulations and secondly to high variability in raw material quality.

Calotropis procera root extracts block VEGF-induced angiogenesis: quantitative analysis.

Angiogenesis is controlled by number of growth factors, including vascular endothelial growth factor (VEGF). Plant derived anti-angiogenic molecules acting via VEGF are being investigated for curtailing angiogenesis dependent diseases. In this study, methanolic (CM), n-hexane (CH), ethylacetate (CE) and water (CW) extracts of the roots of Calotropis procera were tested for anti-angiogenic activity. In the chicken egg chorioallantoic membrane (CAM) assay, CM, CH and CE but not CW inhibited VEGF-induced neovascularization in a dose-dependent manner. Of all the tested extracts, CM at the dose of 10, 5 and 2.5 ng most effectively inhibited over 83, 71 and 64%, of neovascularization induced by 10ng of VEGF, respectively. Sponge implantation assay in mice further showed that at the dose of 100ng CM, CH and CE but not CW significantly inhibited neovascularization induced by VEGF (100 ng). Taken together, this study indicates that the root extracts of C. procera may possess anti-angiogenic activity.

2-Buten-4-Olide (2-B4O) suppresses feeding in neonate and developing rat pups.

The factors that control the satiety behavior and consequently the milk intake in neonates are complex. Mechanical stimulus of gastric filling and emptying are understood to be critical players, though not exclusive. Recently, sugar acid namely, 2-Buten-4-Olide (2-B4O) has been suggested as an endogenous satiety substance in studies conducted on adult rats and monkeys. However, the role of 2-B4O in neonate and suckling rat pups is not well elucidated and this study investigates the same. 2-B4O (50, 75mg/kg, ip) was injected in rat pups on post-natal days (0-2, 4-6 and 12-14) and the mean difference in body weight at 0.5, 1.2 and 24h following treatment was recorded. A dose dependent effect on body weight that correlated with the age of the pups was recorded. In another set of experiment, 2-B4O was micro-infused bilaterally into lateral hypothalamus of 12-14days old pups (1microl of 10microg/microl). Within 30min of micro-infusion, 2-B4O initiated a significant reduction of body weight that lasted up to 24h following treatment. The results of the present study implicate a critical role of 2-B4O as an endogenous feeding suppressant even in neonate and suckling rat pups.

Anti-tumor studies with extracts of Calotropis procera (Ait.) R.Br. root employing Hep2 cells and their possible mechanism of action.

Anti-tumor potential of root extracts of Calotropis procera: methanolic extract (CM), hexane extract (CH), aqueous extract (CW) and ethylacetate extract (CE) and its possible mechanism against Hep2 cancer cells has been investigated. Cellular proliferation activities were assayed by tetrazolium bromide (MTT) colorimetry. Morphological changes of cancer cells were observed under inverted microscope and cell cycle parameters were determined by flow cytometry following propidium iodide staining. Treatment with the extracts at various doses of 1, 5, 10 and 25 microg/ml revealed that CM, CH and CE possessed cytotoxicity, whereas CW did not have cytotoxic effect. CE (10 microg/ml) showed strongest cytotoxic effect (96.3%) on Hep2 at 48 hr following treatment, whereas CM and CH showed cytotoxicity of 72.7 and 60.5%, respectively. Extract-treated cells exhibited typical morphological changes of apoptosis. Results of flow cytometric analysis clearly demonstrated that root extracts initiated apoptosis of Hep2 cells through cell cycle arrest at S phase, thus preventing cells from entering G2/M phase. Results of the study indicate that the root extracts of C. procera inhibit the proliferation of Hep2 cells via apoptotic and cell cycle disruption based mechanisms.

Therapeutic potential of green tea: a new horizon in drug discovery.

Worldwide, tea is one of the most widely consumed beverages. Green tea consumption is especially popular in China, Japan and other Asian countries. It has been found to be rich in polyphenolic compounds, of which catechins are the major constituents. A large number of clinical and preclinical studies have explored its pharmacologic activities. It holds promise as an antioxidant, anti-inflammatory, antibacterial, antiarteriosclerotic, cardioprotective, neuroprotective and anticarcinogenic agent, to name a few. This review summarizes the pharmacodynamics and pharmacokinetics of green tea polyphenols and explores their future as novel drugs for both health and disease conditions.

Evaluation of the effect of Withania somnifera root extracts on cell cycle and angiogenesis.

In the Indian System of Medicine, the medicinal plant, Withania somnifera Dunal (Solanaceae) finds application for numerous ailments including cancer. This study explores the mechanism(s) underlying this property. The hydroalcoholic extract of the roots (WS) was partitioned between chloroform (WS-chloroform) and water (WS-water). Further, WS-chloroform was fractionated (A1-A12) by reverse-phase column chromatography and their withanolide content was quantified by high-performance liquid chromatography (HPLC). Preliminarily, the anti-proliferative activity of all the extracts and fractions was screened against human laryngeal carcinoma (Hep2) cells by microculture tetrazolium assay (MTT). Two extracts (WS and WS-chloroform) and three fractions (A4, A5 and A6) negatively affected Hep2 viability at the concentration of 25 microg/ml and these were further investigated pharmacologically. Flow cytometry revealed cell cycle block and accumulation of hypoploid (sub G1) cells as the mode of anti-proliferative activity of all but A4. Their anti-angiogenic potential was investigated by a chickchorio-allantoic membrane (CAM) wherein a significant inhibition (p<0.0001) of vascular endothelium growth factor (VEGF), induced neovascularization was recorded. The effect was confirmed in vivo by mouse sponge implantation method. These findings suggest that the roots of Withania somnifera possess cell cycle disruption and anti-angiogenic activity, which may be a critical mediator for its anti-cancer action.