Association between Leptin (G2548A) and Leptin Receptor (Q223R) Polymorphisms with Plasma Leptin, BMI, Stress, Sleep and Eating Patterns among the Multiethnic Young Malaysian Adult Population from a Healthcare University.

Relative leptin resistance in childhood to absolute leptin resistance in maturity suggests sleep, eating behaviour, and the psychological state as probable causes. The current body of research provides inconclusive evidence linking G2548A and Q223R to obesity. Furthermore, we could find very little data that have observed the association between the environment and gene polymorphism, especially in the multiethnic population that exists in Malaysia. This study searched for a possible link between sleeping habits, eating behaviour, and stress indicators with plasma leptin and its genetic variation in young adult Malaysian healthcare students. The study involved 185 first- and second-year medical and dental students from a healthcare university. Polymerase Chain Reaction−Restriction Fragment Length Polymorphism(PCR-RFLP) determined the genotype, Enzyme Linked Immunoabsorbant Assay (ELISA) tested the serum leptin, and a self-administered questionnaire evaluated sleep, eating behaviour, and psychological condition. Gender and ethnicity are linked to fasting plasma leptin levels (p < 0.001). Plasma leptin also affects stress, anxiety, and sadness. Leptin (LEP) and Leptin Receptor (LEPR) polymorphisms were not associated with BMI, plasma leptin, sleep, eating behaviour, or psychological state. Young adult Malaysian Indians were obese and overweight, while Chinese were underweight. These findings imply overweight and obese participants were in stage I of leptin resistance and lifestyle change or leptin therapy could prevent them from becoming cripplingly obese as they age.

The potential protective effect of Commelina nudiflora L. against carbon tetrachloride (CCl4)-induced hepatotoxicity in rats, mediated by suppression of oxidative stress and inflammation.

BACKGROUND: This study aims to assess the hepatoprotective potential of Commelina nudiflora against CCl4-induced hepatic injury in rats. METHOD: Antioxidant activities were determined. Phytochemical analysis was performed by gas chromatography mass spectrometry (GCMS). In the in vivo study, Sprague Dawley rats were pretreated with C. nudiflora (150, 300, and 450 mg kg body weight (b.wt.)) once daily for 14 days followed by two doses of CCl4 (1 ml/kg b.wt.). After 2 weeks, the rats were sacrificed and hepatoprotective analysis was performed. RESULTS: In vitro studies have shown that the extract possessed strong antioxidant activity and has ability to scavenge 2,2-diphenyl-2-picrylhydrazyl-free radicals effectively. GCMS analysis of the C. nudiflora extract revealed the presence of various bioactive compounds. Administration of C. nudiflora significantly reduced the impact of CCl4 toxicity on serum markers of liver damage, serum aspartate transaminase (AST), and alanine transaminase (ALT). C. nudiflora also increased antioxidant levels of hepatic glutathione (GSH) and antioxidant enzymes and ameliorated the elevated hepatic formation of malondialdehyde (MDA) induced by CCl4 in rats. Histopathological examination indicated that C. nudiflora protect the liver from the toxic effect of CCl4 and healed lesions such as necrosis, fatty degeneration, and hepatocyte injury as irregular lamellar organization and dilations in the endoplasmic reticulum. The immunohistochemical studies revealed that pretreatment of C. nudiflora decreased the formation of 4-hydroxy-2-nonenal (HNE)-modified protein adducts and 8-hydroxy-2'-deoxyguanosine (8-OHdG). Furthermore, overexpression of the proinflammatory cytokines TNF-α, IL-6, and prostaglandin E2 is also reduced. CONCLUSION: These findings exhibited the potential prospect of C. nudiflora as functional ingredients to prevent ROS-related liver damage.

Prenatal and postnatal exposure to diazinon and its effect on spermatogram and pituitary gonadal hormones in male offspring of rats at puberty and adulthood.

The objective of this research is to study the possible reproductive adverse effects of diazinon on rat offspring exposed in utero and during lactation. Twenty-four Sprague-Dawley female rats (10-12 week old) were randomly assigned to four groups, each consisting of six rats. Group 1 served as the control and these rats were given normal saline orally. Rats in groups 2, 3, and 4 were administered diazinon, dissolved in saline at 10, 15, 30 mg/ kg(-1) body weight, per oral, once daily, during mating, pregnancy and lactation. The male offsprings were examined at puberty and adulthood for body weight, testis weight, epididymis weight, sperm count, motility and morphology, pituitary-gonadal hormone levels. At 30 mg kg(-1) dose, the male offsprings showed a decrease in testicular weight, sperm count, motility, with an increase in abnormal sperm percentage and a decline in pituitary-gonadal hormones, at puberty. Upon attaining adulthood, there was a decrease in testicular weight, sperm count and motility with an increase in abnormal sperm percentage and a decrease in pituitary hormone level. There was evidence of some adverse reproductive effects on the male offspring at the 15 mg/ kg(-1) dose. Most of the adverse effects were irreversible and were evident at both puberty and adulthood in the offsprings, although a few parameters reverted to the normal growth pattern. Diazinon is a reproductive toxicant for male offsprings if exposed during prenatal and postnatal phases.

Lipid peroxidation and decline in antioxidant status as one of the toxicity measures of diazinon in the testis.

The rapid emergence of various pesticides in the market is inevitable due to the demands from agriculture industries and domestic needs to control nuisance pests and to sustain green resources worldwide. However, long-term exposure to pesticide has led to adverse effects on male fertility. Organophosphate diazinon (O,O-diethyl-O-[2-isopropyl-6-methyl-4-pyrimidinyl] phosphorothiote) is an often abusively used pesticide, as it is effective and economical. This study is to determine the adverse effects of low-dose diazinon exposure on the male reproductive system. In this study, 72 Sprague-Dawley rats were segregated into 1, 2, and 8 weeks of exposure groups and further sub-grouped (n = 6) to receive 0, 10, 15, and 30 mg/kg body weight diazinon treatment. Rats were gavaged orally with diazinon and sacrificed under anaesthesia the day after the last exposure. Our results showed that consistent diazinon exposure decreased glutathione and catalase, and increased lipid peroxidation which together lead to diazinon-mediated oxidative stress. Additionally, diazinon increased serum lactate dehydrogenase and decreased serum testosterone, which may have caused sperm and histopathological anomalies. In conclusion, exposure to diazinon caused changes in lipid peroxidation and sperm, and these two effects might be causally linked.

Endosulfan induced early pathological changes in vital organs of rat: a biochemical approach.

AIM: To evaluate the pathogenesis in heart and liver by the early induction of biochemical and antioxidant derangements in rats exposed to endosulfan. MATERIALS AND METHODS: Wistar rats were gavaged with endosulfan (0.5, 1 and 1.5 mg/kg body weight in sunflower oil) for a period of 21 days (single dose at 24 h interval). Control and sunflower oil control groups were also maintained simultaneously. Rats were sacrificed on the 22(nd) day posttreatment. Blood samples, heart and liver were collected and different biochemical parameters such as total protein, cholesterol, triglycerides, amino acids and antioxidant and lipid peroxidation level were measured. Statistical analysis was carried out by one way ANOVA, followed by Bonferroni' post-hoc test. RESULTS: Endosulfan induced a significant increase in the serum levels of total protein, amino acids, triglyceride, total cholesterol, free fatty acid and phospholipid levels in a dose-dependent manner. In the heart and liver, lipid peroxidation was increased significantly in a dose-dependent manner and the antioxidant levels such as superoxide dismutase (SOD), glutathione S-transferase, glutathione peroxidase, and catalase were significantly decreased in a dose-dependent pattern. CONCLUSION: Exposure to endosulfan results in a significant derangement in the biochemical parameters with a decrease in antioxidant levels in the heart and liver. This is an early indication of pathogenesis in the vital organs of rats.

Effects of pesticide use on semen quality among farmers in rural areas of Sabah, Malaysia.

OBJECTIVES: To determine the relationship between semen quality and exposure to pesticide residues. METHODS: A cross-sectional study was conducted among male farmers from 3 different communities in Sabah, Malaysia. A total of 152 farmers participated in this study of whom 62 farmers had been exposed to either paraquat or malathion or both to varying extents. Questionnaires were designed to record a history of pesticides exposure and other potential risk factors among farmers. All semen samples were collected, processed and analyzed by qualified personnel based on WHO guidelines. Volume, pH, sperm concentration, motility, morphology and WBC count were examined and recorded. The association between pesticide exposure and semen parameters was highly significant. RESULTS: The mean values of volume, pH, sperm concentration, motility, and WBC count were significantly less in the exposed group than in compared with the non-exposed group, with p<0.005. Those who were exposed to pesticides had greater risk of having abnormal semen parameters than those in with the non exposed group, with p values of less than 0.05. The comparison between semen qualities such as lower sperm count, motility and higher percentage of sperm abnormality of those exposed to different types of pesticides (paraquat and malathion) showed no significant differences. CONCLUSION: The results showed a significant decline in semen quality with a decline in sperm count, motility and higher percent of teratospermia among subjects with pesticide exposure, and those who were exposed to pesticides had significantly 3 to 9 times greater risk of having abnormal semen parameters.

Exogenous leptin administration decreases sperm count and increases the fraction of abnormal sperm in adult rats.

Daily intraperitoneal injection of 5-30 microg/kg body weight of leptin for 42 days to adult rats decreases sperm count and increases the fraction of abnormal sperm.

An organophosphate insecticide methyl parathion (o- o- dimethyl o-4-nitrophenyl phosphorothioate) induces cytotoxic damage and tubular atrophy in the testis despite elevated testosterone level in the rat.

Methyl parathion (MP) is an organophosphate pesticide used in agriculture, although quite often illegally used indoors to contain insects. The present study was planned to investigate the effects of MP on rat testis. Adult male Wistar rats (13-14 weeks) were treated with MP as follows. Experiment 1-0, 1.75, 3.5 or 7 mg/kg i.p. for 5 days and sacrificed on Day 14; experiment 2 and 3- 0, 0.5, or 1 mg/kg i.p. for 12 days, and sacrificed on Days 130 and 77, respectively; experiment 4- 0, 0.75, or 1.5 mg/kg i.p. for 25 days, and sacrificed on Day 17; experiment 5- 0 or 3.5 mg/kg po for 25 days, and sacrificed on Day 17, after the last exposure. MP decreased the body weight and the testis weight in experiments 4 and 5 (p<0.05-0.001) due to decreased food intake and tubular atrophy respectively. MP increased the intra-testicular testosterone level and decreased the LH level in experiments 4 and 5. The seminiferous epithelium showed sloughing of germ cells, vacuoles, focal necrosis, and formation of multinucleated giant cells, cellular degeneration (nuclear pyknosis, halo appearance and shrinkage of nuclei) and tubular atrophy, especially in experiment 4. The degree of testicular damage was higher in experiment 4>5>1>3>2 indicating more effect of prolonged i.p. treatment. Homogenization-resistant spermatid count was decreased in experiments 1, 4 and 5, and MP also decreased the tubular diameter, and epithelial height (p<0.05-0.001). Incidences of stage XIV tubules, number of meiotic figures and elongating spermatids were also decreased, whereas the incidence of tubules showing epithelial sloughing increased (p<0.05-0.001). We conclude that MP is a reproductive toxicant in male rats which causes significant testicular damage in the testis.

Dacarbazine induces genotoxic and cytotoxic germ cell damage with concomitant decrease in testosterone and increase in lactate dehydrogenase concentration in the testis.

Treatment of cancers with cytotoxic agents such as alkylating drugs often, but not always results in transient to permanent testicular dysfunction. The present study was planned to investigate the effects of dacarbazine [5-(3,3-dimethyltriazeno) imidazole-4-carboxamide] on testicular function in mice. Swiss albino mice (9-12 weeks old) were treated with 0, 5, 25, 50, or 100mg/kg body weight/day dacarbazine (i.p.) for 5 days at intervals of 24h between treatments. Mice were sacrificed on days 7, 14, 21, 28, 35, 49, and 70 after the last treatment (6 mice/dose/sample time), and the epididymal sperm count, sperm motility, sperm morphology, testicular histopathology (qualitative histopathology, seminiferous tubular diameter and epithelial height), and intra-testicular levels of testosterone and lactate dehydrogenase were assessed. Dacarbazine decreased the body weight only on day 28 at 25mg/kg dose-level, but increased the paired testes weights at 50mg/kg on day 7, at 25-100mg/kg on day 14, and at 25 and 50mg/kg on day 21 (P<0.05-0.01; one-way ANOVA and Bonferroni's post hoc test). The sperm count was decreased on all sampling days except at 5 and 25mg/kg dose-levels on day 70, but with severe oligospermia on days 28 and 35 (P<0.05-0.001). The sperm motility was decreased at 100mg/kg on days 14 and 21, at 5, 25, and 100mg/kg on day 28, and at all dose-levels on day 35 (P<0.05-0.001). Dacarbazine induced both head and tail abnormalities and some sperms with cytoplasmic droplets, but significant increase was seen in all dose groups on days 14 and 21, and at 100mg/kg dose-level on day 35. Drug-induced epithelial sloughing was seen on days 14-35 and other histopathological changes observed were vacuoles and abnormal cells. The STD was increased at 25-100mg/kg on day 7, at all dose-levels on day 14, at 50-100mg/kg on days 21 and 28, but without any effects on days 35-70 (P<0.05-0.001), and the tubular lumen was found dilated. The SE was increased on days 7, 21 and 28 at 100mg/kg and on day 14 at 50-100mg/kg. Dacarbazine reduced the intra-testicular testosterone level at 100mg/kg on day 7, at 5, 50 and 100mg/kg on day 14, at all dose-levels on days 21, 28, and 35, and at 50mg/kg on day 49 (P<0.05-0.001). The intra-testicular lactate dehydrogenase concentration increased at all dose-levels up to day 35, but without any effect on days 49 and 70 (P<0.05-0.001). There was no particular dose-response of dacarbazine on any parameters tested. The sperm count (except on day 7-positive correlation; Pearson product moment correlation) or sperm motility did not have any relation but increase in abnormal sperms showed negative correlation with decrease in testosterone level on days 7, 21 and 28. Decrease in sperm count was in negative correlation on days 14 and 35, and increase in abnormal sperms showed positive correlation on day 35 with increase in LDH level. Finally, the decrease in sperm motility had no correlation with increase in abnormal sperm shapes. We conclude that dacarbazine is genotoxic and cytotoxic to the mouse testis in a transient fashion, and these effects are exerted along with decrease in testosterone and increase in lactate dehydrogenase levels in the testis.

A broad-spectrum organophosphate pesticide O,O-dimethyl O-4-nitrophenyl phosphorothioate (methyl parathion) adversely affects the structure and function of male accessory reproductive organs in the rat.

Methyl parathion (MP) is an organophosphate pesticide used in agriculture, but also illegally used to spray homes and businesses to control insects. The present study was designed to investigate adverse effects of MP on accessory reproductive organs. Male Wistar rats aged 13-14 weeks were treated and sacrificed as follows. Experiment 1: 0.0 (water vehicle), 1.75, 3.5 or 7mg/kg (i.p.) for 5 days and sacrificed on day 14; experiment 2: 0.0, 0.5 or 1mg/kg (i.p.) for 12 days and sacrificed on day 130; experiment 3: 0.0, 0.5 or 1mg/kg (i.p.) for 12 days and sacrificed on day 77; experiment 4: 0.0, 0.75 or 1.5mg/kg (i.p.) for 25 days and sacrificed on day 17 and experiment 5: 0.0 or 3.5mg/kg (p.o.) for 25 days and sacrificed on day 17, after the last exposure. The accessory reproductive organs were removed, weighed and processed for histopathological analysis. Structural qualitative changes such as epithelial cell morphology and luminal observations were carried out for each organ in all experiments. Epididymis of one side was homogenized and biochemical estimations of acid phosphatase (ACP), cholesterol, total protein, uric acid, and Vitamin C were conducted by calorimetric methods in experiments 4 and 5. In experiment 1 the organ weights did not change; in experiment 2, the epididymal weight increased (P<0.001); in experiment 3, the weights of ductus deferens decreased at 1mg/kg and that of seminal vesicle decreased at both dose-levels (P<0.001). In experiments 4 and 5, weights of epididymis and prostate decreased, whereas in experiment 5, weights of ductus deferens and seminal vesicle increased (P<0.05-0.001). The sperm density was normal in control, moderately decreased in experiment 1 at 3.5 and 7mg/kg; in experiment 2 at 1mg/kg, and in experiment 5 at 3.5mg/kg, and severely decreased in experiment 3 at 1mg/kg and in experiment 4 at both dose-levels. The epithelial necrosis and nuclear pyknosis were seen in experiments 1, 3, 4 and 5, whereas nuclear degeneration was seen in experiment 1 and 4 and germ cells in the lumina of epididymis were seen in experiment 4. The nuclear pyknosis in the ductus deferens was seen in all experiments, except at 1.75mg/kg in experiment 1 and at 0.5mg/kg in experiment 3. Brush border disruption in the ductus deferens was seen in experiments 1 and 4; sperms were seen in the lumen in experiment 1 at 7mg/kg, and in experiments 4 and 5. The vacuoles in the epithelium were seen in experiments 1 and 4 and immature germ cells were seen in the lumen in experiment 4. The ACP and Vitamin C levels decreased in experiment 4 at both dose-levels, and in experiment 5 all biochemical parameters tested found decreased (P<0.01-0.001). The present results indicate that MP affects the structure and function of accessory reproductive organs in the rat.

The antiviral drug ribavirin reversibly affects the reproductive parameters in the male Wistar rat.

The present study was planned to evaluate the toxic effects of ribavirin on the reproductive parameters in the male Wistar rat. Rats (11--13 weeks old) were treated with 5 injections (i.p.) of 20, 100 or 200 mg/kg/day ribavirin at intervals of 24 h. The testes were processed for histopathological analysis on days 14, 35, 70 and 105 after the last exposure. The parameters studied were body weight, the weights of the testis, epididymis, seminal vesicle and prostate, seminiferous tubular diameter (STD), epithelial height (SE), epithelial sloughing, incidence of stage XIV tubules, sperm abnormality and total serum level of testosterone. Data were analysed by ANOVA and the Bonferroni post hoc test for significances between different groups. There was a decrease in body weight and organ weights, excluding those of the testis and epididymis, against control at higher dose-levels. Ribavirin induced the formation of vacuoles, gaps and sloughing of the seminiferous epithelium. The STD, SE and the incidences of stage XIV tubules decreased on days 14 and 35. Ribavirin also induced the formation of sperm with microcephaly and cephalocaudal junction defects, with or without fibrils jetting out. All these morphological defects recovered to control limit by day 105. The serum level of testosterone was decreased at all dose-levels and time points, although recovery had started by day 105. In conclusion, ribavirin is gonadotoxic in male rats but the effects are reversible after a period of 105 days. However, the endocrine-disrupting properties of ribavirin persist beyond this period.

Genotoxic and cytotoxic effects in the bone marrow of rats exposed to a low dose of paraquat via the dermal route.

The genotoxic effect of the herbicide paraquat was studied in rat bone-marrow by means of the micronucleus assay. Paraquat at dose levels of 6, 15 and 30 mg/kg body weight was given to rats in a single application via the dermal route. Marrow was collected at 24, 48 and 72 h after the application. The micronucleus assay was done as recommended by standard procedures. Paraquat gave rise to an increase in the number of micronuclei in a dose-dependent manner. The number of micronucleated polychromatic erythrocytes showed a maximum at 48 h and the toxicity was further prolonged, as there was no complete recovery at 72 h. These findings suggest a genotoxic effect of paraquat even after exposure via dermal application.

Effect of tamoxifen on spermatogenesis and tubular morphology in rats.

AIM: To observe the effect of tamoxifen citrate on spermatogenesis and tubular morphology in rats. METHODS: The effect of tamoxifen citrate i.g. at doses of 400 and 800 mg.kg(-1).day(-1) in 0.1 mL olive oil for 30 days on seminiferous tubular morphology, seminiferous epithelial diameter (STD), epithelial height (SEH), epididymal sperm count and percent abnormal sperm were evaluated at day 1, 12 and 36 after treatment. Controls were given the vehicle. RESULTS: The higher dose resulted in tubular atrophy on day 31. The STD, SEH and sperm count were decreased and the abnormal spermatozoa increased in a dose-dependent manner with the maximal effect on day 36. CONCLUSION: Tamoxifen citrate induces tubular shrinkage and atrophy and sperm abnormality at a dose-dependent manner.

Tamoxifen induced multinucleated cells (symplasts) and distortion of seminiferous tubules in rat testis.

AIM: To evaluate the effect of tamoxifen citrate on male reproductive system of rat. METHODS: Groups of male rats were gavaged with tamoxifen at doses of 200 mg x kg(-1) x d(-1), 400 mg x kg(-1) x d(-1) or 800 mg x kg(-1) x d(-1) in 0.1 mL olive oil for 10 consecutive days. Controls were treated with 0.1 mL olive oil. Rats were anesthetized and killed on d 3, d 15 or d 35 after the last dose. Testes were collected, processed for paraffin embedding, sectioned at 5 microm thickness, stained with HE and analyzed microscopically. RESULTS: There was a dose-dependent increase in the occurrence of seminiferous tubular distortion with germinal cell sloughing. The highest dose increased the number of multinucleated giant cells on d 3 and d 15. CONCLUSION: Tamoxifen citrate induces multinucleated giant cells and germinal epithelial sloughing in a dose-dependent manner and these changes are detrimental to male fertility.

Toxic effects of 5-Fluorouracil on sperm count in wistar rats.

The antimetabolite, 5-fluorouracil is widely used in the treatment of cancers. Although its toxic effects on testis causing germinal epithelial sloughing, tubular atrophy and generation of multinucleated cells were reported, its effect on spermatogenesis has not been studied. Hence the present study was conducted to evaluate the effects of 5-fluorouracil on epididymal sperm count. Male Wistar rats were employed in the study (n=5 per group). The animals were injected (i.p) with five consecutive doses of 5-fluorouracil (10, 20, 30mg/kg b.w) at an interval of 24h and the control with 0.1ml-distilled water. Samples were obtained at 14, 35, 42 and 70 days after injection. Rats were sacrificed, a laparatomy was performed and epididymes were collected in 1ml phosphate buffered saline (pH 7.2), minced, filtered and stained with 1% aqueous eosin Y. An aliquot was taken in leucocyte pipette, diluted with phosphate buffered saline and sperm count was done as per the standard procedure. Data were analyzed by Mann Whitney U test. The results of this study revealed that 5 - fluorouracil significantly decreased the sperm count in a dose- and time-dependent manner.