Volume 28, Issue 3

Experimental models used in cardiovascular research range from cellular to whole heart preparations. Isolated whole hearts show higher levels of structural and functional integration than lower level models such as tissues or cellular fragments. Cardiovascular diseases are multi-factorial problems that are dependent on highly organized structures rather than on molecular or cellular components alone. This article first provides a general introduction on the animal models of cardiovascular diseases. It is followed by a detailed overview and a historical perspective of the different isolated heart systems with a particular focus on the Langendorff perfusion method for the study of cardiac arrhythmias. The choice of species, perfusion method, and perfusate composition are discussed in further detail with particular considerations of the theoretical and practical aspects of experimental settings.

Background: Indomethacin is commonly used as a nonsteroidal anti-inflammatory drug (NSAID) to treat inflammation, arthritis and joint pains. Unfortunately, it has a wide range of adverse effects on the physiological system, including gonads. This study aimed to assess possible beneficial effects of black tea extract (BTE) against indomethacin-induced alteration of gonadal hormone levels in male rats. Methods: Adult male rats were divided into Group I (control), Group II (indomethacin, 5 mg/kg body weight [bwt.]; i.p., 21 days), Group III (BTE, 2.5 g tea leaf/dL of water, i.e. 2.5% of aqueous BTE, orally, 21 days) and Group IV (indomethacin+BTE, 21 days). Sperm count and motility, serum luteinising hormone (LH), follicle-stimulating hormone (FSH) and testosterone, along with histopathology of testes were studied. One-way ANOVA, followed by post-hoc t-test were conducted. Results: Indomethacin-treated rats showed significant decrease in testicular weight, sperm count, sperm motility, serum gonadotropins and testosterone concentrations. Histopathology of the testes showed tortuous and distorted seminiferous tubules, marked thickening of the tubular basement membrane, reduced spermatogenesis process (>30%) and marked decrease in the number of interstitial cells of Leydig in indomethacin-treated rats. Interestingly, rats supplemented with BTE showed remarkable improvements in testicular weight gain, sperm count and motility, serum gonadotropins and testosterone concentrations, along with testicular histopathology. Conclusions: The results suggest that BTE might have potential ameliorative effects against sub-chronic indomethacin-induced alteration of gonadal hormone levels in male albino rats.

Background: The mechanisms underlying the onset of labor are not fully understood. Extracellular adenosine 5′-triphosphate (ATP) is known to cause uterine contractions in different species but the exact underlying mechanisms are poorly investigated to date. The aims of this study were to investigate the effect of extracellular ATP on spontaneous uterine contractions from different gestational stages and to elucidate its possible underlying mechanisms. Methods: Longitudinal uterine strips were obtained from rats in different gestational stages (nonpregnant, late-pregnant, and term-pregnant). The effects of 1 mM ATP were examined on uterine contractions generated spontaneously, depolarized by high-KCl (60 mM), induced by oxytocin (5 nM), in the presence of high external Ca 2+ , or in the absence of external Ca 2+ . Results: Application of 1 mM extracellular ATP significantly increased the force of spontaneous contraction in uterine strips obtained from all gestational stages with prominent increase in term-pregnant rats compared to other gestations. ATP significantly increased the force induced by depolarization (122%, p=0.010, n=6), oxytocin (129%, p=0.001, n=7), high-Ca 2+ (145%, p=0.005, n=6) and it was able to cause transient contraction in the absence of external Ca 2+ (33%, p<0.01). Conclusions: Extracellular ATP is able to increase the force and frequency of uterine contractions and its effect increases with the progression of pregnancy and it involves Ca 2+ influx and release. These findings open a new window for clinicians to consider ATP as a therapeutic target to control the uterine activity during difficult labors.

Background: Stress fracture (SF) is a common injury among military recruits, especially among women, during the army basic training (ABT). The purpose of this study was to evaluate the effects of health habits and physical activity before recruitment on the fitness level and the incidence of SF during the 4-month ABT. Methods: We screened 226 female recruit volunteers (weight: 60.5±10 kg; height: 163±6 cm) from an integrated combat unit and 124 aged-matched female controls (weight: 57.0±8.3 kg, height 162±7 cm) from a non-combat unit. A self-report questionnaire on their habits pertaining to smoking, physical activity, and orthopedic injuries prior to recruitment were analyzed in relation to the incidences of SF during ABT. Results: Aerobic fitness was similar between the two groups. The overall incidence of SFs was 10.2%. Physical training prior to recruitment had no significant effect on the incidence of SF during ABT (11.7% vs. 9.6% in those who trained and did not train before recruitment, respectively) (Odds ratio, OR)=1.24, p=0.236). Nearly 42% of the female recruits smoked regularly, and the incidence of SFs among smokers was 10.5% compared with 9.9% among the non-smokers (OR=1.07, p=0.188). The overall incidence of SFs 12 months after recruitment was 1.78%. The use of contraceptive medication did not affect the incidence of SF: 10.0% among prior-trained vs. 6.4% in non-prior trained (p>0.05) recruits. SFs were not correlated to these variables at the end of the ABT program and 16 months after recruitment. Conclusions: In the present female cohort, physical activity prior to recruitment had no protective effect against SF during or after ABT. The incidence of SFs during the 12-month period after ABT was negligible.

Background: Physical inactivity contributes to many health issues. The WHO-recommended physical activity for adolescents encompasses aerobic, resistance, and bone strengthening exercises aimed at achieving health-related physical fitness. Heart rate variability (HRV) and maximal aerobic capacity (VO2max) are considered as noninvasive measures of cardiovascular health. The objective of this study is to compare the effect of structured and unstructured physical training on maximal aerobic capacity and HRV among adolescents. Methods: We designed a single blinded, parallel, randomized active-controlled trial (Registration No. CTRI/2013/08/003897) to compare the physiological effects of 6 months of globally recommended structured physical activity (SPA), with that of unstructured physical activity (USPA) in healthy school-going adolescents. We recruited 439 healthy student volunteers (boys: 250, girls: 189) in the age group of 12–17 years. Randomization across the groups was done using age and gender stratified randomization method, and the participants were divided into two groups: SPA (n=219, boys: 117, girls: 102) and USPA (n=220, boys: 119, girls: 101). Depending on their training status and gender the participants in both SPA and USPA groups were further subdivided into the following four sub-groups: SPA athlete boys (n=22) and girls (n=17), SPA nonathlete boys (n=95) and girls (n=85), USPA athlete boys (n=23) and girls (n=17), and USPA nonathlete boys (n=96) and girls (n=84). Results: We recorded HRV, body fat%, and VO2 max using Rockport Walk Fitness test before and after the intervention. Maximum aerobic capacity and heart rate variability increased significantly while heart rate, systolic blood pressure, diastolic blood pressure, and body fat percentage decreased significantly after both SPA and USPA intervention. However, the improvement was more in SPA as compared to USPA. Conclusions: SPA is more beneficial for improving cardiorespiratory fitness, HRV, and reducing body fat percentage in terms of magnitude than USPA in adolescent individuals irrespective of their gender and sports activities.

Background: One of the major toxic side effects of methotrexate (MTX) is enterocolitis, for which there is no efficient standard treatment. Nitric oxide overproduction has been reported to play an important role in MTX-induced mucositis. This study was designed to investigate whether pretreatment with aminoguanidine (AG) – a selective iNOS inhibitor – prevents MTX-induced mucositis in rats. Methods: Rats were pretreated with AG (30 and 50 mg/kg body weight) i.p. daily 1 h before MTX (7 mg/kg body weight) administration for 3 consecutive days. After the final dose of MTX, the rats were killed, and the small intestines were used for analysis. Results: The small intestines of MTX-treated rats showed moderate to severe injury. Pretreatment with AG had a dose-dependent protective effect on MTX-induced mucositis. AG pretreatment reduced iNOS protein levels, mucosal nitric oxide levels, and protein tyrosine nitration. AG pretreatment also restored the activities of electron transport chain (ETC) complexes, vital tricarboxylic acid (TCA cycle) enzymes, and mitochondrial antioxidant enzymes. Conclusions: These findings suggest that AG is beneficial in ameliorating MTX-induced enteritis in rats.

Background: The anti-aging property of β-D-mannuronic acid (M2000) as a novel non-steroidal anti-inflammatory and immunosuppressive agent was investigated on several determinants relative to the oxidative stress in an animal model. Methods: Sprague-Dawley rats were used for evaluating the safety and efficacy properties of M2000 on some oxidative stress enzymes, including the following: mitochondrial superoxide dismutase (SOD2), catalase (CAT), glutathione peroxidase (GPX1), glutathione S-transferase (GST), myeloperoxidase (MPO), and inducible nitric oxide synthase (iNOS) gene expression by real-time PCR. Malondialdehyde (MDA), carbonyl protein (PCO) (the lipid and protein oxidation marker, respectively), and total antioxidant capacity (TAC) were tested in serum by biochemical analysis. In addition, cortisol as a steroid hormone was surveyed by chemiluminescence immunoassay after 12 weeks of M2000 consumption. The rats were sacrificed 3 months after daily oral administration of M2000. Results: Our findings revealed the favorable effects of M2000 on several antioxidant enzyme and gene expression, including SOD2, CAT, GPX1, and GST; however, our results were not statistically significant. Moreover, there was no significant difference in MDA and PCO as lipid and protein oxidation markers, TAC, and cortisol compared with the control group following M2000 consumption. A slight weight increase in the M2000-treated group was also observed. Conclusions: Our data showed the anti-aging property of M2000 as a novel designed non-steroidal anti-inflammatory drug (NSAID) with immunosuppressive property on various oxidative stress determinants.

Background: Butter is one of the widely used fats present in the diet. However, there is no satisfactory study available that evaluates the effect of a high-fat diet containing butter as the principal fat on the development of non-alcoholic fatty liver disease (NAFLD). Methods: In the present study, butter was used for the development of steatosis in Chang liver cells in an in vitro study and Swiss albino mice in an in vivo study. In vitro steatosis was established, and butter was compared with oleic acid in Chang liver cells using an oil red O (ORO)-based colorimetric assay. In the in vivo study, a butter-rich special diet was fed for 15 weeks to mice, who showed no significant change in body weight. The expression pattern of phosphatase and tensin homolog ( PTEN ) and miR-21 was compared by reverse transcriptase-PCR. Results and Conclusions: Special diet-fed animals showed downregulated PTEN compared to normal diet-fed animals, while levels of miR-21 remained the same. Elevations in biochemical parameters, viz., triglycerides and liver function tests showed symptoms of onset of NAFLD. Histophathological study of livers of test animals confirmed mild-to-moderate degree of NAFLD.

Background: Like other tissues, adrenocorticotropic hormone (ACTH) can produce its effect on brown adipose tissue (BAT). This study was taken to understand the direct effect of ACTH action on thermogenin gene expression and possible relation with α receptors and caffeine with this hormone. Methods: Brown fat precursor cells were isolated from interscapular BAT of young mice and grown in culture. The cells were exposed to norepinephrine (NE) and other agents. Total RNA was isolated after harvesting the cells, and northern blot analysis was performed. Hybridization was performed with nick translated cDNA probes. Filters were exposed to film, and results were evaluated by scanning. Cyclic adenosine monophosphate (cAMP) was measured by using Amersham assay kit. Results: ACTH stimulates thermogenin gene expression in brown adipocytes. Initiation and maximum stimulations are observed with 0.01 μM and 10 μM (about 45%) of ACTH, respectively, in comparison to 0.1 μM of NE. Maximum response of cAMP is also observed with 10 μM of ACTH (about 64%). Studies with cirazoline and ACTH show that UCP1 mRNA expression is increased significantly with 10 μM of ACTH, whereas cAMP generation is decreased. In the presence of caffeine, ACTH increases cAMP generation and UCP1 gene expression more than twofold. Conclusions: ACTH stimulates thermogenin gene expression in cultured brown adipocytes. The complex interrelationship of ACTH with cirazoline indicates the possibility of relation between the activity of ACTH and α receptors in brown adipocytes. Further stimulation of cAMP generation and thermogenin gene expression is possible with ACTH in conjugation with caffeine and RO 20-1724.

Background: Ruta montana (L.) is known as a medicinal plant with many beneficial effects, including those that can be used in the treatment of diabetes. The objective of the study was to investigate the antidiabetic effect of this plant in diabetic rat. Methods: This study investigated the effects of an aerial part aqueous extract (APAE) of Ruta montana (L.) (RM) at a dose of 5 mg/kg on blood glucose levels in normal and streptozotocin (STZ)-induced diabetic rats. Histopathological changes were also evaluated in liver and pancreas both in normal and STZ-induced rats. The effect of this aqueous extract on glucose tolerance was demonstrated in normal rats. Furthermore, the relative organ weight (ROW) of liver, kidney, pancreas, and brown adipose tissue were evaluated after 15 days of daily oral administration of the aqueous extract. Results: Both single and repeated oral doses of APAE (5 mg/kg) produced significant reductions in the blood glucose levels in normal and STZ-induced rats. Oral glucose tolerance test results showed that, after the administration of 3 g/kg of glucose, RM APAE (5 mg/kg) improved the increase in blood glucose levels in normal rats at the 30th min (p<0.01) and 90th min (p<0.001). Conclusions: RM APAE exhibits a potent hypoglycemic effect in normal rats and an antidiabetic effect in STZ-induced rats. This finding supports the use of this plant in traditional Moroccan medicine for diabetes management.

Background: In this ethnopharmacological study, methanolic extract of the aerial plant parts of Phragmites karka (Family: Poaceae) and its petroleum ether and carbon tetrachloride fractions were investigated for bioactivities in Swiss-albino mice, namely, analgesic, central nervous system (CNS) depressant, hypoglycemic, and antidiarrheal activity. Methods: The cold methanolic extract of the aerial plant parts of Phragmites karka (MEPK) was first prepared, and it was then further fractionated as petroleum ether (PEFMEPK) and carbon tetrachloride (CTFMEPK) fractions. Analgesic activity was performed employing acidic acid-induced writhing test, central analgesic effect by radiant heat tail-flick method. CNS depressant activity was evaluated by phenobarbitone-induced sleeping time test. Hypoglycemic activity was tested by glucose tolerance test (GTT). Antidiarrheal activity was evaluated by castor oil-induced diarrhea method. For all in vivo tests, doses of 200 and 400 mg/kg body weight were used. Results: In the mice model, the MEPK, PEFMEPK, and CTFMEPK fractions showed significant peripheral analgesic activity at a dose of 400 mg/kg body weight with percentage of inhibition of acetic acid-induced writhing 77.67 (p<0.001), 33.50 (p<0.001), and 40.29 (p<0.001), respectively, compared to the standard dichlofenac (60.68%, p<0.001) group. The hypoglycemic properties of MEPK, PEFMEPK, and CTFMEPK extracts were evaluated in normoglycemic mice where the reduction of blood glucose level after 30 min of glucose load were 69.85%, 78.91%, and 72.73%, respectively, and for standard glibenclamide, the reduction was 72.85%. All results were significant (p<0.05). In the case of the CNS depressant activity by phenobarbitone-induced sleeping time test, the crude ME significantly reduced sleep latency by 57.14% and increased the duration of sleep by 63.29% compared to the control, which were comparable to that of standard diazepam (65.71% and 77.62%, respectively). Among all the extract and fractions, methanolic extract showed the maximum antidiarrheal effect. The methanolic extract at 200 mg/kg dose induced a significant decrease in the total number of defecation in 4 h (69.05% of inhibition, p<0.001) and at 400 mg/kg dose showed 76.19% of inhibition (p<0.001). Conclusions: In light of the available literature, these findings represent the first experimental investigation of biological activities of P. karka in the perspective of their traditional use.