Pharmacological properties of durva swaras (Cynodon dactylon L. Pers.) in an ovariectomised rat model mimicking chronic menopausal syndrome.

Hormonal replacement therapy (HRT), as the first-line management of chronic menopausal syndrome (CMS) in women, has limited application due to adverse effects. We aimed to evaluate the therapeutic potential of a herbal alternative (HALT), durva swaras (DS) of Cynodon dactylon L. Pers., in a CMS rat model. Female Sprague-Dawley rats were subjected to Sham and ovariectomy (OVX) surgery. OVX rats received either 0.11 mg/kg oestrogen as a positive treatment control or 1 (DS1), 2 (DS2), and 4 (DS3) g/kg DS for 160 days. Vaginal smear tests indicated the menopausal status. Routine clinical examinations, weekly body weights (BW), serum calcium, proinflammatory cytokines, and reproductive hormones levels were monitored. Clinical chemistry, body composition, bone mineral density (BMD), uterotrophic response, bone morphometry, and histopathology of major organs were evaluated. BW of OVX rats increased by 18-25% compared to Sham. Total fat and fat percentage were significantly elevated in the oestrogen group compared to DS2, DS3, and OVX group. DS treatment groups showed the levels of TNF- α was slightly reduced, while IL-1ß and IL-6 levels were significantly reduced (P < 0.05) compared to the oestrogen treated group. DS treatment restored serum calcium levels, while BMD, bone quality, osteoblast/osteoclast ratio, and collagen levels improved in both DS and oestrogen treatment groups. The uterotrophic assay demonstrated non-oestrogenic activity of DS. Endometrial hyperplastic change was observed in oestrogen-treated rats. The preclinical non-oestrogenic activity of DS has therapeutic potential in CMS through anti-inflammatory and osteo-protective effects. Further clinical research into DS, as a viable HALT to HRT, is required.

Obesity Potentiates the Risk of Drug-Induced Long QT Syndrome - Preliminary Evidence from WNIN/Ob Spontaneously Obese Rat.

Drug-induced long QT syndrome (DI-LQTS) is fatal and known to have a higher incidence in women rather than in men. Multiple risk factors potentiate the incidence of DI-LQTS, but the actual contribution of obesity remains largely unexplored. Correspondingly, the present study is aimed to evaluate the susceptibility of DI-LQTS in WNIN/Ob rat in comparison with its lean counterpart using 3-lead electrocardiography. Four- and eight-month-old female WNIN/Ob and their lean controls were used for the experimentation. Non-invasive blood pressure measurement and total body electric conductivity (TOBEC) analysis were carried out. After the baseline evaluations, animals were anesthetized with Ketamine (50 mg/kg). Haloperidol (12.5 mg/kg single dose) was administered intraperitoneally and ECG was taken at 0, 10, 20, 30, 60 min, and 24 h time points. Myocardial lystes were used to assess the BNP, protein carbonylation, and hydroxyproline content. Adiposity, as assessed by TOBEC, is higher in obese rats with elevated mean arterial blood pressure. Baseline-corrected QT interval (QTc) is significantly higher in the obese rat with a wider QRS complex. The incidence of PVC and VT are more intense in the obese rat. Haloperidol-induced QT prolongation in obese rats was rapidly induced than in lean, which was observed to remain till 24 h in obese groups while normalized in lean controls. Higher levels of BNP, protein carbonylation, hydroxyproline content, and relative heart weights indicated the presence of cardiac hypertrophy. The study provides preliminary evidence that obesity can be a potential risk factor for DI-LQTS with faster onset and longer subsistence.

Amelioration of neuronal cell death in a spontaneous obese rat model by dietary restriction through modulation of ubiquitin proteasome system.

Dietary restriction (DR) has been shown to increase longevity, delay onset of aging, reduce DNA damage and oxidative stress and prevent age-related decline of neuronal activity. We previously reported the role of altered ubiquitin proteasome system (UPS) in the neuronal cell death in a spontaneous obese rat model (WNIN/Ob rat). In this study, we investigated the effect of DR on obesity-induced neuronal cell death in a rat model. Two groups of 40-day-old WNIN/Ob rats were either fed ad libitum (Ob) or pair-fed with lean. The lean phenotype of WNIN/Ob rats served as ad libitum control. These animals were maintained for 6.5months on their respective diet regime. At the end of the study, cerebral cortex was collected and markers of UPS, endoplasmic reticulum (ER) stress and autophagy were analyzed by quantitative real-time polymerase chain reaction, immunoblotting and immunohistochemistry. Chymotrypsin-like activity of proteasome was assayed by the fluorimetric method. Apoptotic cells were analyzed by TUNEL assay. DR improved metabolic abnormalities in obese rats. Alterations in UPS (up-regulation of UCHL1, down-regulation of UCHL5, declined proteasomal activity), increased ER stress, declined autophagy and increased expression of α-synuclein, p53 and BAX were observed in obese rats and DR alleviated these changes in obese rats. Further, DR decreased TUNEL-positive cells. In conclusion, DR in obese rats could not only restore the metabolic abnormalities but also preserved neuronal health in the cerebral cortex by preventing alterations in the UPS.

Effects of Piper nigrum extracts: Restorative perspectives of high-fat diet-induced changes on lipid profile, body composition, and hormones in Sprague-Dawley rats.

CONTEXT: Piper nigrum Linn (Piperaceae) (PnL) is used in traditional medicine to treat gastric ailments, dyslipidemia, diabetes, and hypertension. OBJECTIVE: The present study explores the possible protective effects of P. nigrum extracts on high-fat diet-induced obesity in rats. MATERIALS AND METHODS: High-fat diet-induced obese rats were treated orally with 200 mg/kg bw of different extracts (hexane, ethylacetate, ethanol, and aqueous extracts) of PnL for 42 d. The effects of PnL extracts on body composition, insulin resistance, biochemical parameters, leptin, adiponectin, lipid profile, liver marker enzymes, and antioxidants were studied. RESULTS AND DISCUSSION: The HFD control group rats showed a substantial raise in body weight (472.8 ± 9.3 g), fat% (20.8 ± 0.6%), and fat-free mass (165.9 ± 2.4 g) when compared with normal control rats whose body weight, fat%, and fat-free mass were 314.3 ± 4.4 g, 6.4 ± 1.4%, and 133.8 ± 2.2 g, respectively. Oral administration of ethyl acetate or aqueous extracts of PnL markedly reduced the body weight, fat%, and fat-free mass of HFD-fed rats. In contrast to the normal control group, a profound increase in plasma glucose, insulin resistance, lipid profile, leptin, thiobarbituric acid reactive substance (TBARS), and the activities of lipase and liver marker enzymes, and a decrease in adiponectin and antioxidant enzymes were noted in HFD control rats. Administration of PnL extracts to HFD-induced obese rats significantly (p < 0.05) restored the above profiles. CONCLUSION: PnL extracts significantly reduced the body weight, fat%, and ameliorated HFD-induced hyperlipidemia and its constituents.

Mitochondriogenesis and apoptosis: possible cause of vitamin A-mediated adipose loss in WNIN/Ob-obese rats.

BACKGROUND: Previously, we reported that vitamin A-enriched diet (129 mg/kg diet) intake reduces the adiposity development in obese rats of WNIN/Ob strain. Here, we hypothesize that dose lesser than 129 mg of vitamin A/kg diet would also be effective in ameliorating the development of obesity in these rats. METHODS: Five-month-old male lean and obese rats designated as A & B were divided into four subgroups (I, II, III and IV) consisting of 8 rats from each phenotype and received diets containing 2.6 mg (control group), 26 mg, 52 mg and 129 mg vitamin A/kg diet as retinyl palmitate for 20 weeks. Body composition and morphological analysis of brown adipose tissue (BAT) was analyzed. Expression of uncoupling protein 1 (UCP1), retinoic acid receptor α (RARα) and retinoid X receptor α (RXRα) in BAT and levels of Bcl2 and Bax in epididymal white adipose tissue (eWAT) were determined by immunoblotting. RESULTS: Vitamin A supplementation to obese rats at doses of 52 and 129 mg/kg diet showed reduced body weight gain and adiposity compared to control diet-fed obese rats receiving 2.6 mg of vitamin A/kg diet. In BAT of obese rats, vitamin A supplementation at doses of 26 and 52 mg of vitamin A/kg diet resulted in increased UCP1 expression with concomitant decrease in RARα and RXRα levels compared to control diet-fed obese rats. Further, transmission electron microscopy study revealed an increase in number of BAT mitochondria of obese rats supplemented with 26 and 52 mg of vitamin A/kg diet. Also, obese rats fed on 52 mg/kg diet resulted in increased apoptosis by altering the ratio of Bcl2 to Bax protein levels in eWAT. Notably, most of these changes were not observed in lean rats fed vitamin A-enriched diets. CONCLUSION: In conclusion, chronic consumption of 52 mg of vitamin A/kg diet seems to be an effective dose in ameliorating obesity possibly through mitochondriogenesis, UCP1-mediated thermogenesis in BAT and apoptosis in eWAT of obese rats. Therefore, the role of dietary vitamin A in correcting human obesity would be of unquestionable relevance and can only be addressed by future studies.

Collagen type-I leads to in vivo matrix mineralization and secondary stabilization of Mg-Zr-Ca alloy implants.

Biodegradable magnesium-zirconia-calcium (Mg-Zr-Ca) alloy implants were coated with Collagen type-I (Coll-I) and assessed for their rate and efficacy of bone mineralization and implant stabilization. The phases, microstructure and mechanical properties of these alloys were analyzed using X-ray diffraction (XRD), optical microscopy and compression test, respectively, and the corrosion behavior was established by their hydrogen production rate in simulated body fluid (SBF). Coll-I extracted from rat tail, and characterized using fourier transform infrared (FT-IR) spectroscopy, was used for dip-coating the Mg-based alloys. The coated alloys were implanted into the femur bones of male New Zealand white rabbits. In vivo bone formation around the implants was quantified by measuring the bone mineral content/density (BMC/BMD) using dual-energy X-ray absorptiometry (DXA). Osseointegration of the implant and new bone mineralization was visualized by histological and immunohistochemical analysis. Upon surface coating with Coll-I, these alloys demonstrated high surface energy showing enhanced performance as an implant material that is suitable for rapid and efficient new bone tissue induction with optimal mineral content and cellular properties. The results demonstrate that Coll-I coated Mg-Zr-Ca alloys have a tendency to form superior trabecular bone structure with better osteoinduction around the implants and higher implant secondary stabilization, through the phenomenon of contact osteogenesis, compared to the control and uncoated ones in shorter periods of implantation. Hence, Coll-I surface coating of Mg-Zr-Ca alloys is a promising method for expediting new bone formation in vivo and enhancing osseointegration in load bearing implant applications.

Zirconium, calcium, and strontium contents in magnesium based biodegradable alloys modulate the efficiency of implant-induced osseointegration.

Development of new biodegradable implants and devices is necessary to meet the increasing needs of regenerative orthopedic procedures. An important consideration while formulating new implant materials is that they should physicochemically and biologically mimic bone-like properties. In earlier studies, we have developed and characterized magnesium based biodegradable alloys, in particular magnesium-zirconium (Mg-Zr) alloys. Here we have reported the biological properties of four Mg-Zr alloys containing different quantities of strontium or calcium. The alloys were implanted in small cavities made in femur bones of New Zealand White rabbits, and the quantitative and qualitative assessments of newly induced bone tissue were carried out. A total of 30 experimental animals, three for each implant type, were studied, and bone induction was assessed by histological, immunohistochemical and radiological methods; cavities in the femurs with no implants and observed for the same period of time were kept as controls. Our results showed that Mg-Zr alloys containing appropriate quantities of strontium were more efficient in inducing good quality mineralized bone than other alloys. Our results have been discussed in the context of physicochemical and biological properties of the alloys, and they could be very useful in determining the nature of future generations of biodegradable orthopedic implants.

Mg-Zr-Sr alloys as biodegradable implant materials.

Novel Mg-Zr-Sr alloys have recently been developed for use as biodegradable implant materials. The Mg-Zr-Sr alloys were prepared by diluting Mg-Zr and Mg-Sr master alloys with pure Mg. The impact of Zr and Sr on the mechanical and biological properties has been thoroughly examined. The microstructures and mechanical properties of the alloys were characterized using optical microscopy, X-ray diffraction and compressive tests. The corrosion resistance was evaluated by electrochemical analysis and hydrogen evolution measurement. The in vitro biocompatibility was assessed using osteoblast-like SaOS2 cells and MTS and haemolysis tests. In vivo bone formation and biodegradability were studied in a rabbit model. The results indicated that both Zr and Sr are excellent candidates for Mg alloying elements in manufacturing biodegradable Mg alloy implants. Zr addition refined the grain size, improved the ductility, smoothed the grain boundaries and enhanced the corrosion resistance of Mg alloys. Sr addition led to an increase in compressive strength, better in vitro biocompatibility, and significantly higher bone formation in vivo. This study demonstrated that Mg-xZr-ySr alloys with x and y ≤5 wt.% would make excellent biodegradable implant materials for load-bearing applications.

Obesity associated pathophysiological & histological changes in WNIN obese mutant rats.

BACKGROUND & OBJECTIVES: WNIN/Ob (obese and euglycaemic) and WNIN/GR-Ob (obesity with impaired glucose tolerance), were isolated and established from Wistar rat stock (WNIN). Both strains showed physical, physiological and biochemical indices related to obesity. We present here haematology, histology and pathophysiological changes between the phenotypes of these strains, lean (+/+), carrier (+/-) and obese (-/-). METHODS: A total of 72 animals of equal gender consisting of three phenotypes were used for the study. Haematology, organ weights were measured and histopathology of the tissues studied using standard procedures. In 12 lean and obese rats (equal gender) of WNIN/GR-Ob group morphometry of pancreatic islets was done immunohistochemically (IHC). RESULTS: Obese rats of both the strains showed normal haematology (except low platelet count), but exhibited changes in the organ weights and in histopathology in organs like liver, kidney, brain and testis/ovary. Hyperplasia of adipocytes was seen in obese rats as compared to lean and carrier. IHC of obese rat pancreas showed that both islet density and volume were significantly (P<0.05) increased compared to lean littermates. INTERPRETATION & CONCLUSIONS: The histological and pathophysiological changes seen in these mutants were in tune with obese phenotype exhibited by these animals.

Infertility in WNIN Obese Mutant Rats-Causes?

We are maintaining two obese mutant rat strains (WNIN/Ob and WNIN/GR-Ob) in our animal facility since 1997. These rat colonies are perpetuated by crossing heterozygous littermates, since the obese phenotypes of both genders turned out be infertile. The present study revealed the reasons for this infertility. The male obese rats, though appeared normal in terms of sperm count, sperm motility and testis histology, however found wanting in terms of libido. This appeared to be due to low circulating testosterone levels seen in these animals, which should also account for low testis and accessory gland weights seen in them. The females exhibited delayed puberty, in terms of days taken for opening of vagina, irregular oestrus cycles and had small ovaries and short and stumpy uterine horns. The FSH peak observed in control lean animals during oestrus stage of the sexual cycle and also the E2 peak of normal oestrus cycle was conspicuously absent in these animals. They also showed elevated levels of progesterone throughout the sexual cycle. Thus the infertility seen in these mutants could be attributed to their abnormal gonadosteroid levels and the resulting anatomical and physiological defects.

Chronic maternal dietary chromium restriction modulates visceral adiposity: probable underlying mechanisms.

OBJECTIVE: We demonstrated previously that chronic maternal micronutrient restriction altered the body composition in rat offspring and may predispose offspring to adult-onset diseases. Chromium (Cr) regulates glucose and fat metabolism. The objective of this study is to determine the long-term effects of maternal Cr restriction on adipose tissue development and function in a rat model. RESEARCH DESIGN AND METHODS: Female weanling WNIN rats received, ad libitum, a control diet or the same with 65% restriction of Cr (CrR) for 3 months and mated with control males. Some pregnant CrR mothers were rehabilitated from conception or parturition and their pups weaned to control diet. Whereas some CrR offspring were weaned to control diet, others continued on CrR diet. Various parameters were monitored in the offspring at three monthly intervals up to 15-18 months of age. RESULTS: Maternal Cr restriction significantly increased body weight and fat percentage, especially the central adiposity in both male and female offspring. Further, the expression of leptin and 11 beta-hydroxysteroid dehydrogenase 1 genes were significantly increased in CrR offspring of both the sexes. Adipocytokine levels were altered in plasma and adipose tissue; circulating triglyceride and FFA levels were increased, albeit in female offspring only. Rehabilitation regimes did not correct body adiposity but restored the circulating levels of lipids and adipocytokines. CONCLUSIONS: Chronic maternal Cr restriction increased body adiposity probably due to increased stress and altered lipid metabolism in WNIN rat offspring, which may predispose them to obesity and associated diseases in later life.

Prenatal and perinatal zinc restriction: effects on body composition, glucose tolerance and insulin response in rat offspring.

Maternal undernutrition increases the risk of adult chronic diseases, such as obesity and type 2 diabetes. This study evaluated the effect of maternal zinc restriction in predisposing the offspring to adiposity and altered insulin response in later life. Seventy-day-old female Wistar/NIN rats received a control (ZnC) or zinc-restricted (ZnR) diet for 2 weeks. Following mating with control males, a subgroup of the ZnR dams were rehabilitated with ZnC diet from parturition. Half the offspring born to the remaining ZnR dams were weaned onto the ZnC diet and the other half continued on the ZnR diet throughout their life. Body composition, glucose tolerance, insulin response and plasma lipid profile were assessed in male and female offspring at 3 and 6 months of age. The ZnR offspring weighed less than control offspring at birth and weaning and continued so until 6 months of age. Rehabilitation regimens corrected the body weights of male but not female offspring. Maternal zinc restriction increased the percentage of body fat and decreased lean mass, fat-free mass and fasting plasma insulin levels in both male and female offspring at 6 months of age. Also, glucose-induced insulin secretion was decreased in female but not male offspring. Despite the differences in fasting insulin and the area under the curve for insulin, the fasting glucose and the area under the curve for glucose were in general comparable among offspring of different groups. Rehabilitation from parturition or weaning partly corrected the changes in the percentage of body fat but had no such effect on other parameters. Changes in plasma lipid profile were inconsistent among the offspring of different groups. Thus chronic maternal zinc restriction altered the body composition and impaired the glucose-induced insulin secretion in the offspring.

Does maternal dietary mineral restriction per se predispose the offspring to insulin resistance?

BACKGROUND: Maternal undernutrition is hypothesized to predispose the offspring to disease in adult life. The relevance of maternal macronutrient deficiency has been well studied but not that of micronutrients. OBJECTIVE: To assess the effect of maternal dietary mineral restriction per se on oral glucose tolerance (OGT), insulin resistance (IR) and fat metabolism in offspring. DESIGN: Female weanling Wistar/NIN rats received a control or a 50% mineral-restricted (MR) diet for 12 weeks, by which time MR rats had lower plasma Fe, Zn, Mg and Ca concentrations. Following mating with control males, a third of the MR dams were shifted to the control diet from parturition. Half of the pups born to the remaining MR dams were weaned onto the control diet while the other half continued on the MR diet. RESULTS: Pregnant MR dams had a higher abortion rate, body weights of their pups at birth and weaning were lower and rehabilitation had no beneficial effect. No offspring had impaired OGT, and IR status was comparable among different groups on postnatal days 40, 70, 100 or 180. Compared with controls, total body electrical conductivity measurements indicated significantly higher body fat %, lower lean body mass and fat-free mass in MR offspring besides elevated plasma triacylglycerols. Mineral rehabilitation from parturition or weaning had little effect on these changes, which did not appear to be due to increased oxidative stress. CONCLUSIONS: Maternal MR per se resulted in an increase in body fat and in plasma triacylglycerol concentrations in the offspring. These changes had, however, no discernable effect on insulin sensitivity over the first 180 days of life.