Nanoparticulate Silica Internalization and Its Effect on the Growth and Yield of Groundnut (Arachis hypogaea L.).

In recent years, foliar applications of nanoparticles are increasingly being employed in agricultural fields as fertilizers to enhance crop yields. However, limited studies are available on the foliar uptake of nanoscale nutrients and their interaction with plants. In this study, we reported the effects of foliar spray with varied concentrations of nanoscale silica (N-SiO2) and bulk tetraethyl orthosilicate (TEOS at 2000 ppm) on the growth and yield of groundnut. Nanosilica was prepared by a sol-gel method and characterized by transmission electron microscopy, dynamic light scattering, and X-ray diffraction. The size and zeta potential of N-SiO2 were found to be 28.7 nm and 32 mV, respectively. The plant height, number of branches, total dry weight, SPAD chlorophyll meter reading, photosynthetic rate, water use efficiency, number of nodules, and ascorbic acid content were increased significantly with the N-SiO2 foliar application at 400 ppm over control. The number of filled pods increased significantly by 38.78 and 58.60% with N-SiO2 at 400 ppm application over TEOS and control, respectively. The pod yield per plant in N-SiO2 at 400 ppm increased by 25.52 and 31.7% higher over TEOS and control, respectively. Antioxidant enzyme activities enhanced significantly in N-SiO2 at 200 and 400 ppm over control, indicating a stimulatory effect on the plant growth. In addition, confocal microscopy revealed that fluorescein isothiocyanate (FITC)-N-SiO2 entered through stomata and then transported to vascular bundles via apoplastic movement. Our study for the first time demonstrated that N-SiO2 can significantly modulate multiple complex traits in groundnut through an eco-friendly and sustainable approach.

Antidiabetic Action of Mcy Protein: Studies on Gene Expression and Competitive Binding to Insulin Receptors.

Mcy protein, isolated from the fruits of Momordica cymbalaria, was shown to have antihyperglycemic, antihyperlipidemic activities along with renal as well as hepatoprotective activities in streptozotocin-induced diabetic rats. Mcy protein was shown to have insulin-like structure and/or function and/or insulin secretagogue activity. Hence, the present study was conducted to elucidate the molecular mechanism whereby Mcy protein elicits its therapeutic role and also to know whether the Mcy protein has any structural and functional similarity with insulin. Results of our experiments revealed that the Mcy protein is insulin-like protein. Furthermore, we assessed the effect of treatment with Mcy protein on the glucose transport (levels of glucose transporter, GLUT-2) and on the levels of key regulators of glucose and lipid metabolisms like hepatic glucokinase (GK) and sterol regulatory element-binding protein-1c (SREBP-1c). Our findings demonstrated that Mcy protein elevated the expressions of GK, SREBP-1c, and GLUT-2 that were decreased in diabetic animals. Insulin-receptor binding studies using rat erythrocytes demonstrated that mean specific binding of insulin with insulin receptors was significantly increased in Mcy-treated diabetic rats when compared to diabetic control rats. Scatchard analyses of insulin binding studies yielded curvilinear plots, and the number of receptor sites per cell was found to be 180 ± 21.1 in Mcy-treated diabetic animals and found to be significantly superior to those of diabetic control animals. Kinetic analyses also revealed an increase in the average receptor affinity of erythrocytes of Mcy-treated rats compared to diabetic control rats suggesting acute alteration in the number and affinity of insulin receptors on the membranes of erythrocytes.

Induction of vitellogenesis, methyl farnesoate synthesis and ecdysteroidogenesis in two edible crabs by arachidonic acid and prostaglandins.

The present study investigated the effect of arachidonic acid (AA) and selected prostaglandins on the regulation of vitellogenesis, ecdysteroidogenesis and methyl farnesoate (MF) synthesis in the freshwater crab Oziotelphusa senex senex and the giant mud crab, Scylla serrata Administration of AA and prostaglandin F2α (PGF2α) and prostaglandin E2 (PGE2) significantly increased ovarian index, oocyte diameter and ovarian vitellogenin levels and ecdysteroid and MF levels in the hemolymph of crabs. Secretions of MF and ecdysteroids from in vitro cultured mandibular organs (MO) and Y-organs (YO) isolated from intermolt crabs injected with AA, PGF2α and PGE2 were greater when compared with controls. In contrast, injection of prostaglandin D2 (PGD2) had no effect on vitellogenesis, ecdysteroid and MF levels in circulation. In vitro secretion of MF from MO explants isolated from avitellogenic crabs incubated with AA, PGF2α and PGE2 increased in a time-dependent manner. Conversely, incubation of YOs isolated from avitellogenic crabs with AA, PGF2α and PGE2 had no effect on secretion of ecdsyteroids. These results implicate prostaglandins in the regulation of reproduction by inducing the synthesis of MF and consequent ecdysteroid synthesis in brachyuran crabs, and provide an alternative molecular intervention mechanism to the traditional eyestalk ablation methodology to induce vitellogenesis and ovarian maturation in crustaceans.

Arsenic aggravated reproductive toxicity in male rats exposed to lead during the perinatal period.

The aim of this study was to assess the reproductive toxic effects of arsenic on adult Wistar rats exposed to lead during the perinatal period. The pregnant rats were allowed ad libitum access to tap water containing 819 mg of lead (Pb) per L or without Pb from conception until weaning. Litter size, survival rate and developmental milestones of the pups delivered by Pb exposed dams were comparable to those of the control rats. Conversely, the pups exposed to Pb during the perinatal period exhibited significant delay in cliff avoidance, negative geotaxis, surface righting reflex, ascending wire mesh and testis descent. The control and perinatal Pb-exposed male rats were maintained on tap water containing 2.3 mg of arsenite (As) per L or without arsenite from the pubertal period (post-natal day 55) to adulthood (post-natal day 115) and assessed for reproductive end points. The results revealed that the (1) relative weights of the testis, epididymis, seminiferous tubules and ventral prostate; (2) daily sperm production; (3) epididymal sperm density and (4) numbers of motile, viable, and HOS tail swelled sperm declined significantly in the rats exposed to either Pb or As. The activity levels of testicular 3ß- and 17ß-hydroxysteroid dehydrogenases were also significantly decreased in the experimental rats. Significant elevation in the levels of reactive oxygen species and lipid peroxidation in association with reduced activities of antioxidant enzymes in the testis and different epididymal regions was recorded in the experimental rats. In the fertility study, although each male in the control and experimental groups produced a copulatory plug and impregnated a female, the mean conception time significantly increased in the experimental groups. The mean number of implantations decreased significantly in the females mated with the experimental males. Moreover, the results of the present study also indicate that reproductive alterations were more deteriorated in the Pb-exposed rats treated with arsenic when compared to individual exposures. In conclusion, the data clearly suggest that reproductive toxicity in male rats exposed to Pb during the perinatal period is exacerbated by As treatment during the pubertal period.

Alleviative effect of resveratrol on polyvinyl chloride-induced reproductive toxicity in male Wistar rats.

The present study was aimed to investigate the effect of PVC on reproductive competence in adult male Wistar rats. Further, the study also encompasses the protective effect of trans-resveratrol on PVC-induced reproductive toxicity in rats. Adult male rats weighing 210-240 g were administered with either PVC at two different doses 100 and 500 mg/kg body weight, orally, daily for 60 days or resveratrol (20 mg/kg body weight/day) through gavage for 60 days on alternate days or both PVC (500 mg/kg body weight) and resveratrol. The results revealed significant reduction in the weights of reproductive organs, epididymal sperm count, viable-, motile-, and HOS-tail coiled sperm and testicular daily sperm production, steroidogenic enzyme activities, serum testosterone levels in PVC treated rats. Conversely the levels of lipid peroxidation increased significantly with a decrease in activity levels of antioxidant enzymes in the testis of PVC exposed rats. Exposure to PVC resulted in reduction in epithelial thickness and seminiferous tubule diameter. No significant changes in the selected reproductive variables were observed in the resveratrol alone treated control rats, whereas, co-administration of resveratrol and PVC resulted in a significant improvement in steroidogenesis and spermatogenesis and mitigated oxidative stress over PVC exposed rats.

Evidence for retinoic acid involvement in the regulation of vitellogenesis in the fresh water edible crab, Oziotelphusa senex senex.

The possible involvement of 13-cis retinoic acid (CRA) in the regulation of ovarian development in Oziotelphusa senex senex was investigated. Injection of CRA, into avitellogenic crabs significantly increased ovarian index, oocyte diameter and ovarian vitellogenin levels. Injection of CRA also resulted in a significant increase in the secretory rates of mandibular organs and Y-organs and circulatory levels of the methyl farnesoate and ecdysteroids. Further, administration of CRA into avitellogenic crabs produced higher amounts of Retinoid X Receptor, Ecdysteroid Receptor, E75 and vitellogenin mRNAs in the hepatopancreas. Mandibular organ and Y-organ explants isolated from avitellogenic crabs secreted more of methyl farnesoate and ecdysteroids respectively when incubated with CRA. Taken together, these observations led us to hypothesize that CRA stimulates ecdysteroidogenesis and methyl farnesoate synthesis, up-regulates EcR, RXR and E75 expression in hepatopancreas, which then induces vitellogenin gene expression. Vitellogenin is subsequently taken up from hemolymph by ovaries ensuing in ovarian maturation.

Forskolin ameliorates mancozeb-induced testicular and epididymal toxicity in Wistar rats by reducing oxidative toxicity and by stimulating steroidogenesis.

In the present study, we have tested the beneficial effects of forskolin in protecting the mancozeb-induced reproductive toxicity in rats. Adult male Wistar rats were exposed to either mancozeb (500 mg/kg body weight/day) or forskolin (5 mg/kg body weight/day) or both for 65 days and analyzed for spermatogenesis and steroidogenesis and testicular and epididymal oxidative toxicity. A significant decrease in daily sperm production, epididymal sperm count, motile, viable, and hypo-osmotic swelling-tail swelled sperm was observed in mancozeb-treated rats. The activity levels of testicular 3ß-hydroxysteroid dehydrogenase and 17ß-hydroxysteroid dehydrogenase and circulatory testosterone levels were significantly decreased in mancozeb-treated rats. Exposure to mancozeb resulted in a significant decrease in glutathione levels and superoxide dismutase and catalase activity levels with an increase in lipid peroxidation levels in the testes and epididymis. Coadministration of forskolin mitigated the mancozeb-induced oxidative toxicity and suppressed steroidogenesis and spermatogenesis.

Serotonin induces ecdysteroidogenesis and methyl farnesoate synthesis in the mud crab, Scylla serrata.

In the current study, we have examined the role of serotonin in regulating the levels of methyl farnesoate and ecdysteroids in the giant mud crab Scylla serrata and validated that serotonin indeed is a reproductive hormone. Administration of serotonin elevated circulatory levels of methyl farnesoate and ecdysteroids in crabs. Since methyl farnesoate and ecdysteroid act through retinoid X receptor (RXR) and ecdysteroid receptor (EcR) respectively and these receptors are involved in the regulation of reproduction in crustaceans, we have determined the mRNA levels of RXR and EcR in hepatopancreas and ovary after serotonin administration. The expression levels of both RXR and EcR increased significantly in the hepatopancreas and ovary of serotonin injected crabs when compared to the controls. In vitro organ culture studies revealed that incubation of Y-orgas and mandibular organ explants in the presence of serotonin resulted in a significant increase in the secretion of ecdysteroids by Y-organs, but without alterations in MF synthesis in mandibular organs. From the above studies it is evident that serotonin stimulates Y organs resulting in increased ecdysteroidogenesis. Though the circulatory levels methyl farnesoate elevated after serotonin administration, organ culture studies revealed serotonin mediated methyl farnesaote synthesis is indirect probably by inhibiting release of mandibular organ inhibiting hormone from eyestalks.

Effects of maternal exposure to aflatoxin B1 during pregnancy on fertility output of dams and developmental, behavioral and reproductive consequences in female offspring using a rat model.

A suboptimal in utero environment can have detrimental effects on the pregnancy and long-term adverse "programing" effects on the offspring. Aflatoxin B1 is one of the potent reproductive toxicants and currently detected in both milk and tissues. This article focuses on the effects of prenatal exposure to graded doses of aflatoxin B1 on the pregnancy outcomes of dams and postnatal developments of the female offspring, since these issues have ethological relevance in both animals and humans. Pregnant Wistar rats were injected intramuscularly with vehicle or aflatoxin B1 (10, 20, 50 or 100 µg/kg body weight/day) on days 12-19 of gestation. At parturition, newborns were observed for clinical signs of toxicity and survival. The female offspring were examined through a battery of tests in order to evaluate their developmental, behavioral and reproductive end points. All animals were born alive. The litter size of the aflatoxin B1 treated rats was comparable to the controls. However, the birth weight of the pups in the experimental group was significantly lower when compared to controls. Significant and persistent lags in cliff avoidance, negative geotaxis, surface rightening activity and ascending wire mesh, with a delay in elapsed time for vaginal opening were detected in the female progeny exposed to aflatoxin B1 during embryonic development. The locomotor activity and exploratory behavior in experimental females were significantly decreased than that of controls. Embryonic exposure to aflatoxin B1 also resulted in prolonged stress response, irregular estrus and suppressed fertility output in the progeny at their adulthood. These results indicate that in utero exposure to aflatoxin B1 severely compromised postnatal development of neonatal rats and caused irregular estrus that was accompanied by suppressed fertility output.

Induction of ecdysteroidogenesis, methyl farnesoate synthesis and expression of ecdysteroid receptor and retinoid X receptor in the hepatopancreas and ovary of the giant mud crab, Scylla serrata by melatonin.

Melatonin, a chronobiotic molecule, is known to modulate several physiological functions in crustaceans including reproduction, molting and glucose homeostasis. In our earlier studies (Sainath and Reddy, 2010a), we observed hyperglycemia in crabs after melatonin administration and concluded that melatonin is another crustacean hyperglycemic hormone. In the current study, we have further examined the role of melatonin in regulating the levels of methyl farnesoate and ecdysteroid in the giant mud crab Scylla serrata and determined that melatonin indeed is a reproductive hormone. Further, we have determined partial nucleotide sequences of retinoid X receptor (RXR) and ecdysone receptor (EcR) in S. serrata and also studied the effect of melatonin on expression of these genes. Cloned RXR and EcR possess high sequence similarity with other Brachyuran genes. Administration of melatonin elevated circulatory methyl farnesoate (MF) and ecdysteroid levels in crabs. Since MF and ecdysteroid act through RXR and EcR respectively and these receptors are involved in the regulation of reproduction in crustaceans, we measured the expression levels of RXR and EcR in hepatopancreas and ovary after melatonin administration. The expression levels of both RXR and EcR increased significantly in the hepatopancreas and ovary of melatonin injected crabs when compared to the controls. In vitro culture of mandibular organ (MO) and Y-organ (YO) in the presence of melatonin resulted in a significant increase in the secretion of methyl farnesoate and ecdysteroid respectively. From the above studies it is clear that melatonin stimulates YO and MO, resulting in increased synthesis of ecdysteroids and methyl farnesoate, and thereby inducing reproduction in S. serrata.

Expression of RXR, EcR, E75 and VtG mRNA levels in the hepatopancreas and ovary of the freshwater edible crab, Oziothelphusa senex senex (Fabricius, 1798) during different vitellogenic stages.

The objective of the present study was to investigate the expression profile of retinoid X receptor (RXR), ecdysone receptor (EcR) and ecdysone inducible gene (E75) in the hepatopancreas and ovary of Oziothelphusa senex senex during different vitellogenic stages. RXR, EcR and E75 complementary DNAs (cDNAs) were isolated from the ovaries, while vitellogenin (VtG) cDNA was isolated from the hepatopancreas of vitellogenic female crab. Deduced amino acid sequence of the messenger RNAs (mRNAs) of RXR, EcR and E75 showed more than 80% identity with their respective mRNAs of other brachyurans. VtG mRNA was not detected in the ovary throughout vitellogenic stages. RXR and EcR were significantly increased in the ovaries during vitellogenic stage I. The levels of EcR, E75 and VtG in the hepatopancreas elevated significantly during vitellogenic stages I and II, whereas the levels of RXR elevated only in vitellogenic stage I. During vitellogenic stage III, the levels of RXR, EcR and VtG in the hepatopancreas were significantly decreased. Immunoprecipitation analysis revealed the presence of VtG in the haemolymph, hepatopancreas and ovary extracts from the females but absent in haemolymph and hepatopancreas extract of males. It can be inferred that RXR, EcR and E75 are involved in the regulation of synthesis of VtG in hepatopancreas, whereas in ovary, it is hypothesized that they play an important role in the uptake of VtG from the haemolymph, probably by regulating the levels of vitellogenin receptor. These are the first data showing an association between the expression levels of RXR, EcR and E75 and vitellogenesis and provide an alternative molecular intervention mechanism to the traditional eyestalk ablation to induce vitellogenesis and ovarian maturation in crustaceans.

Restraint stress exacerbates alcohol-induced reproductive toxicity in male rats.

Cumulative exposure to multiple stresses may lead to aggravating the toxicity of each stress, qualitatively or quantitatively altering biological responses because of toxicological interaction. In this study, we intended to determine the possible effects of restraint stress on reproductive toxicity due to ethanol usage in male rats. Early pubertal male Wistar rats were subjected to either restraint stress (5 h/day) or alcohol intoxication (2 mg/kg body weight) or both for 60 days. Body weights of control and experimental rats were similar during the 60 days of this study. Testes were harvested, weighed, and prepared for enzyme assays, and cauda epididymides were isolated for the determination of density, motility, and viability of stored spermatozoa. Restraint stress or alcohol treatment significantly reduced testis weight and caused significant reductions in steroidogenesis and spermatogenesis. Mean density, motility, and viability of stored spermatozoa were reduced in experimental rats. Plasma testosterone concentrations in rats subjected to restraint stress or alcohol were decreased compared with those of controls, concomitant with increased concentrations of LH and FSH in experimental rats. These data suggest that sub-chronic exposure to restraint stress or alcohol contribute to reduce testicular and epididymal function in exposed rats. The study also suggests that restraint stress exacerbates alcohol-induced reproductive toxicity in rats.

Aflatoxin B1-Induced Reproductive Toxicity in Male Rats: Possible Mechanism of Action.

Aflatoxin B1 (AFB1), one of the most common mycotoxins found in human foods, is principally hepatotoxic; however, it also affects reproduction. The aim of the present study was to elucidate the reproductive toxic effects and possible mechanism of action of AFB1 in rats. Male Wistar rats were injected intramuscularly with doses of 10, 20, or 50 µg AFB1/kg body weight on alternate days from 45 to 100 days of age. Significant reductions in body weights, relative weights of reproductive organs, daily sperm production, epididymal sperm count, viable sperm, motile sperm, and hypoosmotic swelling-tail coiled sperm were observed. Significant decreases in testicular steroidogenic enzymes and serum testosterone levels were also observed indicating decreased steroidogenesis. In silico docking studies illustrated AFB1 binds with steroidogenic acute regulatory (StAR) protein thereby affecting the transport of cholesterol into mitochondria resulting in decreased steroidogenesis.

Hepatopancreas but not ovary is the site of vitellogenin synthesis in female fresh water crab, Oziothelphusa senex senex.

The objective of the present study was to explore the site of synthesis of vitellogenin (Vtg) in fresh water edible crab, Oziothelphusa senex senex. Vtg cDNA fragments were isolated from the hepatopancreas of female crabs using RT-PCR method, and the deduced amino acid sequence of O. senex senex showed more than 60% identity with other brachyuran Vtg sequences. RT-PCR analysis showed that Vtg mRNA can be detected only in hepatopancreas of female Oziothelphusa but not in other tissues including eyestalks, Y-organs, mandibular organs, thoracic ganglion, hypodermis and ovary. Antibodies were raised against vitellin purified from the ovary of O. senex senex. Immunoprecipitation analysis revealed the presence of Vtg in the hepatopancreas of vitellogenic stage I females and in the hemolymph, hepatopancreas and ovary extracts from vitellogenic stage II females but absent in hemolymph and hepatopancreas extract of males. These results suggest that Vtg is synthesized only in hepatopancreas but not in the ovaries of O. senex senex. In addition, Vtg synthesized in hepatopancreas is transported to ovary through hemolymph.