Association of B12 deficiency and anemia synergistically increases the risk of high TNF-α levels among adolescent girls.

We investigated the association between iron status, B12, and inflammatory markers among 101 adolescent girls. We found that B12 showed significant negative association with tumor necrosis factor-alpha (TNF-α) (rs = -0.232, P = 0.020) and positive association with serum ferritin (SF) (rs = 0.209, P = 0.036) among girls. Our results showed that hepcidin discriminates anemic and non-anemic population under normal B12 conditions. The logistic regression analysis revealed that the risk of having higher TNF-α levels was 13.2 times higher in low B12 girls in the presence of anemia compared to the girls having normal hemoglobin and B12 levels.

Effects of oxidation on redox and cytotoxic properties of copper complex of Aß1-16 peptide.

The effect of oxidation on redox and cytotoxic properties of copper complex of amyloid beta (Aß) peptide was studied by gamma radiolysis. The oxidation of Aß1-16 and Aß1-16/Cu(II) complex was carried out using hydroxyl ((•)OH) radicals produced by gamma radiolysis and the products were analyzed using mass spectrometry. The presence of Cu(II) was found to enhance the oxidation of Aß1-16 peptide. The oxidation of residues Asp1, His6, and His13 was enhanced due to their involvement in copper binding. The oxidation of His residues of Aß1-16 peptide, which are chiefly responsible for copper binding, resulted in altered redox properties and subsequently in higher cytotoxicity of the Aß1-16 peptide in SH-SY5Y cells.

Effects of oxidation on copper-binding properties of Aß1-16 peptide: a pulse radiolysis study.

The reaction of hydroxyl radicals ((•)OH) with Aß1-16 peptide was carried out using pulse radiolysis to understand the effect of oxidation of peptide on its copper-binding properties. This reaction produced oxidized, dimeric and trimeric Aß1-16 peptide species. The formation of these products was established with the help of fluorescence spectroscopy and mass spectrometry. The mass spectral data indicate that the major site of oxidation is at His6, while the site for dimerization is at Tyr10. Diethyl pyrocarbonate-treated Aß1-16 peptide did not produce any trimeric species upon oxidation with (•)OH. The quantitative chemical modification studies indicated that one of the three histidine residues is covalently modified during pulse radiolysis. The copper-binding studies of the oxidized peptide revealed that it has similar copper-binding properties as the unoxidized peptide. Further, the cytotoxicity studies point out that both oxidized and unoxidized Aß1-16 peptide are equally efficient in producing free radicals in presence of copper and ascorbate that resulted in comparable cell death.

Heavy cluster knockout reaction (16)O((12)C,2(12)C)(4)He and the nature of the (12)C-(12)C interaction potential.

The heavy cluster knockout reaction (16)O((12)C,2(12)C)(4)He performed for the first time, reveals the true nature of the (12)C-(12)C interaction. The observed cross section is enhanced by almost 2 orders of magnitude over the conventional zero range distorted wave impulse approximation (DWIA) predictions. An attractive (12)C-(12)C optical potential, as obtained in the folding model, does not explain the enhanced cross section in the finite range (FR) DWIA framework. The inclusion of a hard core of fairly long range ∼3.65 fm explains the data. The present investigation of (16)O((12)C,2(12)C)(4)He along with the (12)C-(12)C elastic scattering also proves beyond doubt that the folding model's deep attractive heavy ion potentials are unsuitable to describe the highly overlapping heavy ions. The application of FR-DWIA opens up new avenues to use the heavy core knockout for the detailed investigation of heavy as well as Borromean halo nuclei.

Melatonin-induced changes in ovarian function in the freshwater fish Channa punctatus (Bloch) held in long days and continuous light.

It is possible to manipulate fish gonadal function through exogenous melatonin. However gonadal responses to melatonin vary and depend on time, mode and duration of the administration of the hormone. The present study describes the effects of different photoperiods and melatonin treatments on the ovarian kinetics of the fish Channa punctatus. Fish held in long days (LD 14:10) were injected melatonin daily (10 microg im) either at 08.00 or 17.00 h, respectively or immersed in melatonin water (100 microg/l) for 24 or 15 h daily. In another experiment fish held in long days or continuous light (LL; LD 24.00) were immersed in melatonin water (100 microg/l) either for 24 or 15 h daily. Both experiments had appropriate controls. The GSI (Gonadosomatic index=gonadal wt./100g body weight) increased (P<0.01) in fish immersed in melatonin water daily for 24h when compared either with the GSI of control fish or with fish held in melatonin water daily for 15 h (17.00-08.00 h). The GSI decreased (P<0.01) in fish that received melatonin daily by injection mode. Data from follicular kinetics largely corroborate the GSI data. Fish exposed to melatonin water daily for 24h had more vitellogenic follicles (VF) and fewer atretic follicles (AF). There was a general decrease (P<0.01) in previtellogenic follicles (PVF) in all treated groups. Melatonin by injection mode did not affect the number of VF but it significantly increased (P<0.01) the AF. In the other experiment, the GSI increased (P<0.01) in fish held in long days and immersed in melatonin water 24h daily. However, the GSI decreased in fish held in long days and immersed in melatonin water for a restricted period (between 17.00 and 08.00 h). The GSI of fish held in LL and immersed in melatonin water daily for 24h increased (P<0.01), whereas it decreased (P<0.01) in fish that were immersed in melatonin water daily for 15 h. The data from follicular kinetics revealed a decrease in PVF of fish held in LL and in all the melatonin-exposed groups irrespective of the duration of exposure to melatonin. However the VF increased markedly in fish held in LL and immersed in melatonin water daily for 24h. VF of fish held in LL but immersed in melatonin water daily for restricted period decreased. In fish held in long days and immersed in melatonin water daily either for 24 or 15 h the VF number increased (P<0.01). Marked increase in AF was seen fish held in LL and exposed to melatonin-containing water daily for restricted period. The effect of melatonin on reproduction seems to depend on the photoperiod and duration of exposure to melatonin. In fish held either in long days or LL, daily restricted exposure to melatonin tends to inhibit gonads, whereas continuous exposure to melatonin either stimulate or has no effect. The results apparently support the view that duration of exposure to melatonin signals darkness.

Melatonin counteracts the stimulatory effects of blinding or exposure to red light on reproduction in the skipper frog Rana cyanophlyctis.

An earlier study reported that blinding or exposure to red light stimulates reproduction in R. cyanophlyctis. The present study investigates whether melatonin counteracted blinding- or red-light-induced ovarian stimulation. The study consisted of two experiments carried out during the prebreeding (March/April) and breeding phase (August/September) of the reproductive cycle of this species. In experiment 1, exposure to red light for 30 days increased (P<0.01) the gonadosomatic index (GSI) and oviductal weights, whereas exposure to continuous darkness for the same duration decreased (P<0.01) the GSI. Data on follicular kinetics revealed an increase in vitellogenic oocytes and decrease in previtellogenic oocyte numbers following exposure to red lights. Exposure to continuous dark on the other hand increased previtellogenic oocyte and decreased vitellogenic oocyte numbers. In experiment 2, the GSI and the number of vitellogenic oocytes increased in blinded and red-light-exposed frogs. However, treatment with melatonin (15 microg sc) for 30 days to both these groups of frogs reduced the GSI, oviductal weights, and the number of vitellogenic oocytes. The results both confirm earlier finding that blinding and exposure to red light stimulate ovarian growth and also demonstrate that melatonin counteracts blinding or red-light-induced stimulation of ovarian activity. This apart, the findings from experiment 1 demonstrate for the first time that exposure to continuous dark inhibits ovarian growth in this species. Ocular melatonin seems to play an important role in regulation of reproduction in this species.

Pearl millet cysteine protease inhibitor. Evidence for the presence of two distinct sites responsible for anti-fungal and anti-feedent activities.

Recently, pearl millet cysteine protease inhibitor (CPI) was, for the first time, shown to possess anti-fungal activity in addition to its anti-feedent (protease inhibitory) activity [Joshi, B.N. et al. (1998) Biochem. Biophys. Res. Commun. 246, 382-387]. Characterization of CPI revealed that it has a reversible mode of action for protease inhibition. The CD spectrum exhibited a 35% alpha helix and 65% random coil structure. The intrinsic fluorescence spectrum was typical of a protein devoid of tryptophan residues. Demetallation of Zn2+ resulted in a substantial change in the secondary and tertiary structure of CPI accompanied by the complete loss of anti-fungal and inhibitory activity indicating that Zn2+ plays an important role in maintaining both structural integrity and biological function. The differential response of anti-fungal and inhibitory activities to specific modifiers showed that there are two different reactive sites associated with anti-fungal and anti-feedent activity in CPI located on a single protein as revealed from its N-terminal sequence data (AGVCYGVLGNNLP). Modification of cysteine, glutamic/aspartic acid or argnine resulted in abolition of the anti-fungal activity of CPI, whereas modification of arginine led to an enhancement of the inhibitory activity in solution. Modification of histidine resulted in a twofold increase in the protease inhibitory activity without affecting the anti-fungal activity, whereas modification of serine led to selective inhibition of the protease inhibitory activity. The differential nature of the two activities was further supported by differences in the temperature stabilities of the anti-fungal (60 degrees C) and inhibitory (40 degrees C) activities. Binding of papain to CPI did not abolish the anti-fungal activity of CPI, supporting the presence of two active sites on CPI. The differential behavior of CPI towards anti-fungal and anti-feedent activity cannot be attributed to changes in conformation, as assessed by their CD and fluorescence spectra. The interaction of CPI modified for arginine or histidine with papain resulted in an enhancement of CPI activity accompanied by a slight decrease in fluorescence intensity of 15-20% at 343 nm. In contrast, modification of serine resulted in inhibition of CPI activity with a concomitant increase of 20% in the fluorescence intensity when complexed by the enzyme. This implies the involvement of enzyme-based tryptophan in the formation of a biologically active enzyme-inhibitor complex. The presence of anti-fungal and anti-feedent activity on a single protein, as evidenced in pearl millet CPI, opens up a new possibility of raising a transgenic plant resistant to pathogens, as well as pests, by transfer of a single CPI gene.

Purification and characterization of a cell-surface lectin (Lectin II) from Agrobacterium radiobacter NCIM 2443.

A lectin was isolated from Agrobacterium radiobacter cell surface and purified. It is a monomer of 40 kDa as shown by SDS-PAGE. The lectin has a pI of 9.15 and amino acid composition of the lectin shows that 44% of the amino acids are hydrophobic. The lectin agglutinates rabbit erythrocytes but does not agglutinate human erythrocytes. It does not show specificity for monosaccharides except for D-glucosamine. Fetuin and its N-linked glycopeptide also inhibit the activity of the lectin but greater inhibition is shown by locust bean gum and Nicotiana tobaccum (tobacco) tissue extracts.

Exposure to different spectra of light, continuous light and treatment with melatonin affect reproduction in the Indian desert gerbil Meriones hurrianae (Jerdon).

In two separate experiments, the gonadal response to melatonin treatment and exposure to continuous white light (LL) and different spectra of light (blue, green, yellow and red) was investigated in the Indian desert gerbil Meriones hurrianae. In experiment 1, exposure to green light for 8 weeks increased (p < 0.01) the gonadosomatic index (GSI) and sperm count, whereas exposure to red light decreased (p < 0.01) the GSI and sperm count. The response of accessory organs to the treatment varied. In experiment 2, exposure to LL for 5 weeks decreased the GSI and sperm count, and treatment with melatonin (in the late-light period) not only increased the GSI and sperm count, but also counteracted the inhibitory action of LL and red light. Melatonin treatment stimulated accessory sex organs and the scent gland surface area. The inhibition or stimulation of the reproductive parameters correlated with an increase or decrease in pinealocytes and pinealocyte nuclear diameters. Therefore, the influence of light and its spectra on reproduction may be mediated by the pineal gland, though extrapineal mechanisms cannot be ruled out.

Cysteine protease inhibitor from pearl millet: a new class of antifungal protein.

A cysteine protease inhibitor exhibiting antifungal activity from pearl millet seeds has been purified to homogeneity by ammonium sulphate precipitation and chromatographic procedures involving CM- sephadex and SP-sepharose cation exchange columns. The molecular characterization has revealed its molecular mass as 24 kD and isoelectric point 9.8. The amino acid composition data shows presence of high content of serine and glycine (34 residues/mole) and absence of tryptophan. The inhibitor exhibits potent antifungal activity against Trichoderma reesei, a dead wood fungus with minimum inhibitory dose to inhibit mycelial growth or spore germination is as low as 1 microgram/ml (250 ng/disc). In addition to Trichoderma reesei, the antifungal activity is observed against some important phytopathogenic fungi, namely, Claviceps, Helminthosporium, Curvularia, Alternaria and Fusarium species. To the best of our knowledge, a cysteine protease inhibitor as an antifungal protein is reported for the first time from a plant system.

Changes in ovarian follicular kinetics in intact blinded and parietal shielded frogs exposed to different spectra of light.

Ovarian follicular kinetics and gravimetric changes in the ovary and oviduct were studied in intact, blinded (BL), and parietal shielded (PS) skipper frog Rana cyanophlyctis exposed to different light spectra. The gonadosomatic index (GSI) increased (P < 0.01) in intact and PS frogs, held in green, yellow, and red light. The maximum increase was in red light followed by yellow and green. The GSI of BL frogs also increased (P < 0.001) in white, green, yellow, and red light. However, the GSI of blinded and parietal shielded (BLPS) frogs increased (P < 0.001) in white and all the colored lights compared with controls. The percentage changes of oviductal weights were similar to the ovarian weights. Data on follicular kinetics revealed a decrease in previtellogenic oocytes in intact and PS frogs placed in blue, green, yellow, and red lights. The previtellogenic oocytes of BL and BLPS frogs decreased in white light, while their number did not vary significantly in other spectra. The vitellogenic oocytes of intact and PS frogs increased in all the spectra, with a maximum increase in red. The vitellogenic oocyte count increased in both BL and BLPS frogs held in white light. The green, yellow, and red spectra stimulated vitellogenic oocyte count of BL frogs. The response of BPLS frogs was similar except for a slightly decreased oocyte count in red light. The pattern of vitellogenic oocyte counts in general exhibited a negative correlation with previtellogenic oocyte counts in all the frogs. The atretic follicular numbers did vary significantly in all of the groups. Red light stimulated ovarian activity maximally, followed by yellow and green. As the ovary exhibited varied response to the different spectra of light even in BL frogs, it may be concluded that extraretinal perception of colored light occurs in this species.

Photoperiod and melatonin-induced changes in male reproduction in Indian desert gerbil, Meriones hurrianae (Jerdon).

Exposure to continuous darkness and chronic treatment with melatonin, for six weeks, stimulated reproduction in the male Indian desert gerbil as evidenced by morphometric data. Exposure to continuous light, for same duration, on the other hand inhibited reproduction. The results are opposite to those reported from similar studies on temperate zone species. Surface areas of abdominal scent glands increased following both, exposures to continuous darkness and the treatment with melatonin. Exposure to continuous light decreased the scent gland surface area. Assessment of scent gland activity could be useful in evaluation of reproductive function as they are dependent on sex steroids. Melatonin that mediates photoperiodic influence on reproduction is not always inhibitory to gonads.

Effect of exposure to continuous light and melatonin on ovarian follicular kinetics in the skipper frog, Rana cyanophlyctis.

Ovarian follicular kinetics and gravimetric changes in the ovary and oviducts were studied in the skipper frog, Rana cyanophlyctis, following exposure to continuous light and melatonin treatment during the breeding season. Daily late-afternoon injections of melatonin (15 micrograms subcutaneous) for 30 days decreased the gonadosomatic index (GSI), whereas continuously available melatonin from subcutaneous implants did not influence the GSI compared to those of controls. Exposure to continuous light for 30 days stimulated the GSI, and melatonin given as daily injections prevented the continuous-light induced increase in GSI. Oviductal weights decreased only in the melatonin-injected groups. Data on follicular kinetics revealed a decrease in first-growth-phase (FGP) oocytes and an increase in medium-sized second-growth-phase (MSGP) and large-sized second-growth-phase (LSGP) oocytes following continuous-light exposure. Melatonin administered to continuous-light-exposed frogs did not change the FGP oocyte number; however, it reduced both the MSGP and LSGP oocytes. Melatonin administration to frogs held in a light and dark cycle increased FGP oocytes and decreased MSGP and LSGP oocytes. Atretic follicles increased in all melatonin-treated groups. The results suggest that continuous light stimulates and melatonin inhibits reproductive function in this species.

Melatonin and exposure to constant light/darkness affects ovarian follicular kinetics and estrous cycle in Indian desert gerbil Meriones hurrianae.

Melatonin mediates photoperiodic influence on reproduction and constant light and darkness affect pineal biosynthesis of melatonin. The present study was undertaken to assess the effects of melatonin and drastic photoperiodic changes on reproduction in a tropical desert species with a fossorial lifestyle. Ovarian follicular kinetics and estrous cycle were studied in the Indian desert gerbil Meriones hurrianae, after treatment with melatonin and exposure to constant light (LL) and darkness (DD) regimes. Melatonin treatment increased (P < 0.001) ovarian weights without changing the uterine weights. While exposure to LL decreased (P < 0.001) both ovarian and uterine weights, exposure to DD had no effect on these weights. Follicular kinetics of growing and regressing follicles revealed that ovaries of melatonin-treated and DD-exposed animals had significantly more growing follicles. Melatonin treatment increased all types of growing follicles, especially antral and Graafian follicles. Exposure to DD increased all types of growing follicles, with the medium sized antral and Graafian follicles being significant (P < 0.01). In contrast to stimulation of follicular growth by melatonin and DD, LL caused regression of all stages of follicular growth and also reduced the number of small preantral follicles. Melatonin treatment increased (P < 0.001) the length of estrous cycle (5.08 to 7.29 days). Gerbils treated with melatonin, exposed to LL and DD, had a longer (P < 0.001) metestrus. Animals held in LL, had the least number (P < 0.001) of estrous smears (1 in 30 days). The results suggest that melatonin is involved in growth of ovarian follicles in the Indian desert gerbil.

Diurnal sensitivity in melatonin-induced hematological changes in the male albino rat.

The influence of melatonin on diurnal changes in the hematological profile was investigated in male albino rats. Following treatment with melatonin, under two different experimental protocols, blood samples were collected at 06.00, 12.00, 18.00 and 24.00 h from separate groups of animals for detailed hematological analysis. In experiment 1, melatonin (25 micrograms, s.c.) treatment, once daily at 17.00 h for 4 weeks, led to an alteration in the rhythm of RBC production with a decrease in its counts at 12.00 h. The total WBC counts were increased at 06.00 and 12.00 h and decreased at 18.00 and 24.00 h. In experiment 2, twice-daily administration of melatonin (once at 09.00 h and again at 17.00 h), for 2 weeks, resulted in a decrease in total RBC counts at 06.00 and 24.00 h, whereas at 18.00 h there was an increase, compared to the respective control values. The total WBC counts increased at 06.00, 12.00 and 18.00 h and decreased at 24.00 h. Erythrocyte indices like mean cell volume (MCV), mean cell hemoglobin (MCH) and mean cell hemoglobin concentration (MCHC), in both experiments, correlated with RBC, hemoglobin and hematocrit values. The WBC differential counts in experiment 1 revealed a decrease in total neutrophils at 24.00 h, whereas in experiment 2 there was a general decrease in their number. While in experiment 1 the total lymphocyte number was increased at 06.00 h and decreased at 18.00 and 24.00 h, in experiment 2 it was increased except at 24.00 h. It may be concluded that melatonin has a modulatory role in hemopoiesis and its rhythms. The stimulatory effect of melatonin on WBC supports its purported immunopotentiating action.

Ovarian follicular kinetics in response to enucleation and parietal shielding in Rana cyanophlyctis.

Ovarian follicular kinetics, gravimetric changes in the ovary and the oviduct, were studied in the skipper frog, Rana cyanophlyctis, following enucleation (ENX) and parietal shielding (PS), during the breeding reason. PS was to prevent extraoptic photoreception by the pineal via the frontal organ. The gonadosomatic index (GSI) was high in ENX and PS frogs (P < 0.01). The highest GSI occurred in frogs that were both ENX and PS (P < 0.01). Percentage of oviductal weight decreased (P < 0.01) in PS frogs and increased (P < 0.01) in those subjected to both ENX and PS. Oviductal weight of ENX frogs was unchanged. Data on follicular kinetics revealed a decrease (P < 0. 01) in the ENX group in the number of first growth phase (FGP) oocytes and atretic follicles (AF), whereas the number of medium-sized secondary growth phase (MSGP) oocytes and large-sized secondary growth phase (LSGP) oocytes increased (P < 0.01). There was a similar pattern of follicular kinetics in ENX+PS frogs. However, following PS, the number of FGP oocytes and AF increased (P < 0.01) and decreased (P < 0.01) in the MSGP and LSGP oocytes. The lower oviductal weight in PS frogs correlated with reduced SGP oocytes, the major source of estrogen. The importance of lateral eyes and the frontal organs in the transduction of environmental signals to the neuroendocrine-reproductive axis is indicated in this species. That recruitment of FGP to SGP is inhibited only in the PS group suggests that the frontal organ plays an important role in ovarian follicular kinetics.

Influence of melatonin on blood morphology in male albino mice.

Influence of melatonin on diurnal changes in hematological profiles was examined in male albino mice, at six hourly interval. Melatonin treatment either once daily (25 micrograms at 1700 hrs) or twice daily (25 micrograms at 0900 and 1700 hrs) for two weeks resulted in an alternation in the RBC rhythm. Generally the effect was suppressive though at some times the counts were marginally increased. Total WBC counts were increased and there was apparently a change in their rhythm too. Interestingly the differential WBC counts exhibited different patterns in, once and twice daily melatonin-treated mice. Melatonin given once daily predominantly stimulated the absolute lymphocyte count whereas the twice daily regimen predominantly stimulated the neutrophil count. This perhaps is related to the varied exposures of the animals to melatonin in the two experimental setups. Erythrocyte indices like mean cell volume, mean cell hemolglobin and mean cell hemoglobin concentration in both the experiments correlated with hemoglobin and hematocrit values. It may be concluded that melatonin has a modulatory role in hemopoiesis and its rhythms. The stimulatory effect of melatonin on WBC supports its purported immunopotentiating action.

Effect of long-term administration of melatonin on adrenal histomorphology of the Indian desert gerbil Meriones hurrianae Jerdon.

Melatonin treatment for 8 weeks resulted in a significant increase (P < 0.01) in weights of the adrenal glands, in both male and female, M. hurrianae. The weight increase was not related to the time of administration of melatonin. Histology of adrenal glands in melatonin treated gerbils revealed a stimulatory response. In treated gerbils, adrenal cortex exhibited significant hypertrophy with enlarged nuclei and degranulated cytoplasm. Signs of hypertrophy were also evident in the adrenal medulla. The physiological significance of this response is not yet clear. It remains to be clarified if the hypertrophy indicates enhanced synthesis of corticosteroids.

Effect of long-term administration of melatonin on eyes and harderian glands in albino rat.

Pineal hormone melatonin is also synthesized by the ocular structures, retina and Harderian gland. The physiological significance of such production is not fully understood. Daily treatment with melatonin (25 micrograms/rat) for 8 weeks led to an increase in weight of eye balls and fluid within it (P < 0.05). Weight of the Harderian gland in the control and treated rats was not significantly different, though the histological observation revealed an increase in cell height and reduction in lumen diameter in the treated animals. Inter luminal deposits of porphyrin content appeared to be more in the Harderian glands of melatonin treated rats. The retinal tissue indicated degenerative changes in the melatonin treated rats. The results suggest melatonin's influence on level of ocular fluid, structure/function of retina and harderian gland.

Melatonin in the control of the estrous cycle of the Indian desert gerbil (Meriones hurrianae: Jerdon).

Daily administration of melatonin for 8 weeks, either in the morning (9 a.m.) or late in the afternoon (5 p.m.), resulted in significant prolongation of the estrous cycle in the Indian desert gerbil Meriones hurrianae. The gonadosomatic index increased significantly (p < 0.01) in the melatonin-treated gerbils compared to the control animals. Histological study of the ovaries revealed enlargement of follicles and corpora lutea with hypertrophied granulosa cells in the melatonin-treated gerbils in comparison to their controls. The data suggest that the mechanism involved in the gonadal response to melatonin may not be the same in the temperate and tropical species.