Deuterium labeling improves the therapeutic index of 3,3'-diselenodipropionic acid as an anticancer agent: insights from redox reactions.

3,3'-Diselenodipropionic acid (DSePA), a selenocystine derivative, has been previously reported as an oral supplement for anticancer/radio-modulation activities. The present study is focused on devising a strategy to synthesize and characterize the deuterated derivative of DSePA and on understanding the effect of deuteration on its therapeutic index by comparing its cytotoxicity in cancerous versus non-cancerous cell types. In this context, the synthesis of 3,3'-diselenodipropionic acid-D8 (D-DSePA) was accomplished in ∼42% yield. Further, the results clearly established that the deuteration of DSePA significantly reduced its cytotoxicity in non-cancerous cell types while retaining its cytotoxicity in cancerous cell lines. Together, D-DSePA displayed a ∼5-fold higher therapeutic index than the non-deuterated derivative for anticancer activity. The biochemical and NMR studies confirmed that the better biocompatibility of D-DSePA than its non-deuterated derivative in non-cancerous cells was due to its ability to undergo slower redox reactions and to cause lesser inhibition of intracellular redox enzymes.

Interaction of a Model Hydrophobic Drug Dimethylcurcumin with Albumin Nanoparticles.

The aim of present study was to investigate the binding interactions of a model hydrophobic molecule, dimethylcurcumin (DMC) with nanoparticle form of bovine serum albumin (BSA) using fluorescence spectroscopy techniques. For this, BSA nanoparticles (size = 62.0 ± 3.5 nm, molecular weight = 11,243 ± 3445 kD) prepared by thermal denaturation method was mixed with DMC in solution and monitored for fluorescence emission of tryptophan (Trp) residue as well as DMC separately. The emission maximum of DMC in nanoparticles form exhibited more blue sift and quenched the excited state of tryptophan (Trp) by six fold higher than in the native form of BSA. By analyzing Trp fluorescence, the mean binding constant (K) estimated for the interaction of DMC with native and nanoparticles forms of BSA was 2.7 ± 0.4 × 104 M-1 and 1.5 ± 0.5 × 105 M-1 respectively. Together these results suggested that DMC experienced a more rigid environment in nanoparticles than in native form of BSA. Additionally the above determined K values were in agreement with those reported previously by absorption techniques. Further direct energy transfer was observed between Trp and DMC, using which the distance (r) calculated between them was 28.25 ± 0.27 Çº in BSA native. Similar analysis involving BSA nanoparticle and DMC revealed a distance of 24.25 ± 1.05 Çº between the hydrophobic core and the ligand. Finally interaction of DMC with BSA was validated through molecular docking studies, which indicated sub-domain IIA as the binding site of DMC. Thus it is concluded that intrinsic fluorescence of protein can be utilized to study the interaction of its different physical forms with any hydrophobic ligand.

Tuning the binding, release and cytotoxicity of hydrophobic drug by Bovine Serum Albumin nanoparticles: Influence of particle size.

To elucidate the effect of particle size of albumin nanoparticles on cellular uptake of a hydrophobic drug, herein we report the release kinetics and cytotoxicity of nanoparticle bound dimethylcurcumin (DMC) in A549 tumor cells. The bovine serum albumin (BSA) nanoparticles were prepared by thermal denaturation and characterized by dynamic light scattering (DLS), zeta (ζ) -potential, circular dichroism (CD) and transmission electron microscope (TEM). The preparation conditions were optimized to obtain nanoparticles with mean hydrodynamic diameters 28.0nm (BSAnp1) and 52.0nm (BSAnp2) and corresponding ζ- potential value of∼-7.0 and -6.0mV, respectively. Interaction of DMC with BSA nanoparticles was investigated by UV-vis, fluorescence and CD spectroscopy. CD studies indicated significant changes in the secondary structure of BSA upon particle formation, as revealed by decrease in the helicity. The cellular uptake of DMC increased with increase in particle size and the toxicity of DMC loaded nanoparticles to A549 cells were found to be consistent with their cellular uptake. Between the two formulations studied, BSAnp2 provided enhanced cellular uptake and can be used as an effective delivery system for hydrophobic drugs like DMC.

Arsenic(V) adsorption mechanism using kaolinite, montmorillonite and illite from aqueous medium.

The present work investigates the adsorptive behavior of As(V) ions with kaolinite, montmorillonite and illite in aqueous medium as a function of As(V) concentration, pH, contact time and temperature. As(V) adsorption on studied clays were pH dependent and maximum adsorption were achieved in the pH range 2.0-5.0. The adsorption data gave good fits with Langmuir isotherm and yielded Langmuir monolayer capacity of 0.86, 0.64 and 0.52 mg As(V) /g of kaolinite, montmorillonite and illite, respectively. An increase in adsorption temperature resulted in a decrease in the amount of As(V) adsorbed. The results of leaching study showed that, kaolinite was very active clay constituent regarding both As(V) adsorption and mobility. The electrokinetic behavior of both the kaolinite and montmorillonite were modified in the presence of As(V). The shift in isoelectric point indicated that adsorption involves inner sphere surface complexation and strong specific ion adsorption. Kaolinite was successfully tested as an adsorbent for the removal of arsenic from two contaminated groundwater samples containing arsenic in the range 1.36-1.41 mg/L.

Removal of As(V) by Cu(II)-, Ni(II)-, or Co(II)-doped goethite samples.

The present study reports removal of As(V) by adsorption onto laboratory-prepared pure and Cu(II)-, Ni(II)-, and Co(II)-doped goethite samples. The X-ray diffraction patterns showed only goethite as the crystalline phase. Doping of ions in the goethite matrix resulted in shift of d-values. Various parameters chosen for adsorption were nature of adsorbent, percentage of doped cations in goethite matrix, contact time, solution pH, and percentage of adsorbate. It was observed that the pH(pzc) of the goethite surface depended on the nature and concentration of metal ions. The surface area as well as the loading capacity increased with the increase of dopant percentage in goethite matrix. A maximum loading capacity of 19.55 mg/g was observed for 2.7% Cu(II)-doped goethite. The adsorption kinetics for Ni(II), Co(II) and for undoped goethite attained a quasi-equilibrium state after 30 min with almost negligible adsorption beyond this time. In case of Cu(II)-doped goethite samples, the quasi-equilibrium state for As(V) adsorption was observed after 60 min. At each studied pH condition, it was observed that the percentage of adsorption of As(V) decreased in the order Cu(II)-doped goethite > or = Ni(II)-doped goethite > Co(II)-doped goethite > pure goethite. The adsorption followed: Langmuir isotherm, indicating monolayer formation.

Use of oxide minerals to abate fluoride from water.

The removal of fluoride from aqueous solutions has been investigated using various oxide ores such as refractory grade bauxite, feed bauxite, manganese ore, and hydrated oxides of manganese ores (WAD). The refractory grade bauxite showed promising results. The studies were carried out as functions of contact time, pH, concentration of adsorbents, concentration of adsorbate, and temperature. The adsorption was rapid during the initial 5 min but equilibrium was attained within 120 min. The adsorption followed first-order kinetics. The present system followed the Langmuir adsorption isotherm model. Various thermodynamic parameters such as free energy, enthalpy, entropy, and equilibrium constants were calculated. The isosteric heat calculations showed that the adsorption process followed a heterogeneous model.

Roles of FSH and testosterone in the initiation of spermatogenesis in prepubertal rats medically hypophysectomized by a GnRH antagonist.

Experimentally induced medical hypophysectomy in prepubertal rats through treatment of GnRH antagonist for 3 weeks, initiated on the 20th day of age, markedly decreased testicular weight by 85% of that of the controls. Quantitative assessment of spermatogenesis in testicular semithin preparations revealed a significant reduction in the numbers of preleptotene (27.2 +/- 1.6 to 15.6 +/- 0.52) and pachytene (25.8 +/- 0.96 to 5.35 +/- 0.26) spermatocytes and complete absence of any spermatids after the treatment. By contrast, round stage 7 and elongated spermatids were observed in many tubules of the testis in the age-matched control rats. At the end of GnRH antagonist treatment, the blood levels of LH were undetectable, while testosterone and FSH were decreased to 12 and 44% of the controls, respectively. Supplementation of either FSH (ovine FSH 20 micrograms/rat day-1) or testosterone (30 micrograms/rat day-1) enhanced the testicular weight (68%) and the circulatory levels of these hormones, but failed to support quantitatively normal spermatogenesis, which was, however, qualitatively improved. The number of maturing spermatids were comparatively higher in the testosterone-supplemented group that in the FSH-administered group. The latter group had otherwise the highest number of degenerating germ cells per tubule (mean 4.8 +/- 0.1). Testicular weight and stage-specific germ cell counts were restored to normalcy only in rats supplemented with both FSH and testosterone, the critical concentrations of which were important in the initial stages of spermatogenesis. Testosterone alone had a positive effect in terms of germ cell development, while FSH without testosterone was detrimental to the maturing germ cells.

Is testosterone essential for maintenance of normal morphology in immature rat Leydig cells?

Selective deprivation of gonadotrophins in prepubertal rats by administration of a GnRH antagonist (Ac-D2Nal1, D4ClPhe2, DTrp3, DArg6, DA1a10-GnRH; GnRH code: 103-289-10, National Institutes of Health, USA) for 3 weeks, initiated at 20-22 days of age, induced morphological changes in the Leydig cells, including thickening and indentation of the nuclear margin, pyknosis and elongation of the nuclei. Mean nuclear diameter was reduced to 22% of that in the controls. Under the electron microscope the cells exhibited reduced volume of the nucleus and cytoplasm and the plasma membrane was irregular. This abnormal appearance of the Leydig cells improved marginally in 20-30% of the Leydig cells and their mean nuclear diameter increased to 39% of the control level after FSH supplementation (20 micrograms ovine FSH/day). Normal morphological integrity of the Leydig cells consisting of round or oval nuclei, a smooth nuclear and cellular margin and the original mean nuclear diameter was restored completely when testosterone (30 micrograms/day) was administered to GnRH antagonist-treated rats, with or without simultaneous administration of FSH; in these rats testosterone levels in blood were also restored to normal. These findings indicate that testosterone may be important for the maintenance of normal Leydig cell morphology in the rat.

Effect of fertility regulating agents on motility and zona-free hamster egg penetration by spermatozoa of bonnet monkey.

Administration of STS-557 (17 alpha-cyanomethyl-17 beta-hydroxyestra 4,9(10)-dien-3-one; 12 mg/monkey daily) for 4 weeks either alone or in combination with 20 Aet-1 (testosterone-trans-4-n-butyl cyclohexyl carboxylate; code CDB 1781; 40 mg/monkey single administration) had no significant effect on motility and zona free hamster egg penetration by spermatozoa of bonnet monkey, but continuation of the treatment for 12 weeks reduced (in one monkey treated with STS-557) or abolished (one treated with STS-557 and two with STS-557 + 20 Aet-1) the motility as well as zona-free hamster egg penetration (by spermatozoa of all treated monkeys). Motility and the ability to penetrate zona-free hamster egg returned to normalcy after 10 weeks of withdrawal of treatments. Active immunization of monkeys with ovine FSH (4 weeks after booster) had no adverse effect on motility of spermatozoa but none of the zona-free hamster eggs was fertilized. The correlation between motility and the capacity to penetrate the zona-free hamster eggs by monkey spermatozoa varies with the treatment. Such correlation was apparent in monkeys treated with STS-557 but not in monkeys immunized with ovine FSH.

Adverse effects of chronic GnRH antagonist administration on seminiferous epithelium in adult rats.

Seminiferous epithelium in adult rats was studied by light and electron microscopy after 5 weeks of chronic administration of GnRH antagonist (Ac-D2 Nal 1, D4ClPhe 2, DTrp 3, DArg 6, DAla 10; GnRH code-103-289-10, National Institutes of Health, USA). In these rats, the epithelium showed significant accumulation of vacuoles in more than 80% of the tubules, along with germ cell degeneration and nuclear pyknosis. Disruption in the process of spermatogenesis was also very much evident. In most of the tubules studied (greater than 90%), germ cell development was arrested beyond the pachytene spermatocyte stages. The vacuoles in the seminiferous epithelium were different sizes and when magnified were seen to consist of a thickened outer margin of solid nonfibrous coat within the Sertoli cell cytoplasm. Associated changes in the interstitium showed increased intertubular space but no inflammatory type of response. In actual cell counts, the decrease in the average number of macrophages was 32% and in Leydig cells 23%, while the total number of all types of cells in the interstitium was 30% less than that of the controls. Following the treatment, weights of testis, epididymis, seminal vesicle, and ventral prostate were drastically reduced. Rats treated with testosterone supplementation (60 micrograms/rat day) to GnRH antagonist recovered testicular and epididymal weights to approximately 57% and seminal vesicle and ventral prostate weights by 82.9 and 84%, respectively. Normalcy returned to the tubular epithelium and the interstitial cell counts were restored to original levels.

Differential response of testis and serum gonadotropins to testosterone in rats treated with a gonadotropin releasing hormone antagonist or estradiol 17 beta.

Adult rats treated with a GnRH antagonist (Ac D2Nal1, D4Cl Phe2, DTrp3, DArg6, DAla10 GnRH; code: 103-289-10, National Institutes of Health, USA) for 5 weeks (250 micrograms/kg b.w) showed multiple degrees of impairment and atrophy of the genital organs concomitant with decreased serum levels of testosterone, LH and FSH. Inhibition of spermatogenesis was characterized by germ cell degeneration and overall decline in different cell numbers and in particular, spermatids of any kind were completely absent. Testosterone supplementation (60 micrograms/rat/day, sc) to GnRH antagonist treated rats, for the same period, significantly elevated the weights of the sex organs, and the serum levels of hormones. Spermatogenesis was improved both qualitatively and quantitatively; albeit failed to be restored back to control levels. Treatment with estradiol 17 beta (1 microgram/rat/day) for 5 weeks had insignificant effect on spermatogenesis but the weights of the genital organs (seminal vesicles by 19% and ventral prostate by 40%) and the levels of serum hormones (LH by 24%, FSH 22% and testosterone by 25%) were otherwise reduced. Administration of testosterone either alone or in combination with estradiol 17 beta had only a marginal effect on spermatogenesis or on other reproductive parameters. The results indicate a positive shift in the response of the testis and serum levels of gonadotropins to testosterone supplementation in rats treated with either GnRH antagonist or estradiol 17 beta.

Sperm production and fertility of bonnet monkeys (Macaca radiata) following immunization with ovine follicle stimulating hormone.

Adult male bonnet monkeys were rendered oligospermic but not azoospermic following active immunization with ovine follicle stimulating hormone. The percentage of sperms in the semen having good motility was reduced with a concomitant increase in the sperm ATPase activity. Eight out of 10 immunized monkeys failed to impregnate females of proven fertility after mating for consecutive three cycles while the remaining two impregnated the cohabitated females during the third cycle at a time when the antibody titer was reduced. Active immunization with ovine follicle stimulating hormone may not produce complete azoospermia but renders adult male monkeys infertile provided sufficient antibody titer is maintained.

Effects of STS-557 and 20 Aet-1 on sperm functions and serum level of testosterone in bonnet monkey (Macaca radiata).

Treatment of adult male bonnet monkeys with STS-557 (17 alpha-cyanomethyl 17 beta-hydroxy estra-4, 9(10)-dien-3-one; 12 mg/monkey daily for 15 weeks; i.m.) reduced the sperm count from the 9th week, leading close to azoospermia on the 2nd post-treatment week which persisted until the 10th post-treatment week and normalcy was restored on the 13th post-treatment week. The sperm motility was reduced from the 9th week of treatment to 8th post-treatment week. The fertilizing ability of spermatozoa (formation of swollen sperm head or pronuclei in zona-free hamster eggs) was abolished from the 8th week of treatment to the 7th post-treatment week. The serum level of testosterone was reduced from the 2nd to 12th week of treatment (data on subsequent weeks were not collected). When 20 Aet-1 (testosterone-trans-4-n butyl cyclohexyl carboxylate; 40 mg/monkey, single; i.m.) was administered on the first day of STS-557 treatment, the sperm count was reduced from the 10th week, with near azoospermia ensuing on the 13th week which continued until the 8th post-treatment week; recovery was observed on the 12th post-treatment week. The motility was low from the 8th week of treatment to 6th week after withdrawal of treatment. The fertilizing ability of spermatozoa was abolished from the 8th week to the 12th week of treatment (data on last week of treatment were not collected). The serum level of testosterone was maintained within normal range except on the 2nd, 6th and 12th week of treatment. 20 Aet-1 alone had no significant effect on these parameters.(ABSTRACT TRUNCATED AT 250 WORDS)

Abdominal suture proctopexy for complete prolapse of rectum.

Eighteen patients, 13 males and 5 females, with complete rectal prolapse have been treated by suture proctopexy during the period 1987-1989 with no operative mortality and very little morbidity. There was only one case of recurrence during a 3-year follow-up for which a repeat identical procedure was successful. There was some disturbance in faecal continence pre-operatively in 14 cases, of whom, 13 cases were improved by suture proctopexy. There was no significant change in bowel habit postoperatively, with the exception of 2 patients who had constipation postoperatively but one reverted to normal on re-education of bowel habit. All the patients had normal urinary and sexual function. This relatively simple procedure was well tolerated by the patients.

GnRH antagonist treatment affects nuclear size and membrane associated indentations in rat Leydig cells.

In semi-thin preparations of rat testis treated with GnRH antagonist (Ac-D2Nal 1, D4ClPhe 2, DTrp 3, DArg 6, DAla 10; GnRH code- 103-289-10, National Institutes of Health, USA) for a period of 5 weeks, Leydig cells showed a reduction of 18% in the mean nuclear area as compared to the controls. The number of cells with nuclear membrane indentations increased by more than 40% as compared to the controls. Nuclear condensation was evident in the form of dark, electron-dense patches of heterochromatin present all along the rough, uneven boundary of the nuclear wall. In some cells, indentations in the nucleus appeared in the form of deep grooves extending into the core of the nucleus. Shrinkage in the cytoplasm of the cells was very much apparent. Circulating levels of LH and testosterone were very low. LH decreased from 29.6 +/- 2.9 ng/ml to 0.55 +/- 0.018 ng/ml and testosterone from 7.8 +/- .29 ng/ml to 0.5 +/- .02 ng/ml. The mean nuclear area of Leydig cells from other groups of rats treated with estradiol 17-beta (1 microgram/rat per day), testosterone (60 micrograms/rat per day) or combinations of testosterone and estradiol or GnRH antagonist and testosterone in the same regime and for the same period remained unaltered. On the other hand, cells appeared normal when testosterone was supplemented (60 micrograms/rat/day) in rats treated with GnRH antagonist. The mean nuclear area of the Leydig cells recovered significantly (p less than 0.001), but was not completely restored.

Effect of STS-557 (17 alpha-cyanomethyl 17 beta-hydroxy-estra-4, 9(10)-dien-3-one) on blood hormone levels, the testis, accessory sex organs and fertility of rats.

Treatment with STS-557 (17 alpha-cyanomethyl 17 beta-hydroxy-estra-4,9(10)-dien-3-one; 10 mg kg-1 daily s.c.) for 4 weeks induced atrophy of the seminiferous tubules in adult rats with a reduction in tubule diameter and in the number of round spermatids at stage VII. Elongated spermatids were not detected. Leydig cells were atrophied from the second week of treatment with a concomitant decrease in blood levels of testosterone. The blood levels of FSH and LH were reduced from the third week of treatment. The weight of the reproductive organs was reduced after STS-557 treatment. The treatment induced sterility in 50% of rats after 2 weeks of treatment but after 4 weeks none of the treated males mated. Normal fertility and normal levels of testosterone and FSH were restored after 6 weeks and LH after 4 weeks of withdrawal of treatment. All other parameters studied recovered to pretreatment levels 6 weeks after withdrawal of treatment. STS-557 could act on the pituitary-gonadal axis (reducing gonadotrophin secretion) as well as directly affecting the Leydig cells. The consequent reduction in the blood levels of testosterone in combination with reduced gonadotrophins was presumably responsible for the suppression of spermatogenesis.

Contraceptive potentiality of STS-557--a feasibility study in male bonnet monkey (Macaca radiata).

STS-557 (17 alpha-cyanomethyl-17 beta-hydroxy-estra-4, 9(10)-diene-3-one) was administered (i.m.) to two groups (4 in each group) of adult male bonnet monkeys at a daily dose of 10 mg/monkey for 12 weeks (first group) and 5 mg/monkey for 14 weeks (second group). Treatment with the 10 mg dose resulted in a significant decline in the count, motility, acrosin and hyaluronidase activities and the fertilizing ability (zona-free hamster egg penetration assay) of spermatozoa by the 6th week of initiation of treatment. The circulating level of STS-557 was low after one week and increased from the 2nd week of treatment when the serum level of testosterone was significantly reduced. Complete recovery was observed by the 11th week after withdrawal of treatment. The treatment with the 5 mg dose had minor and inconsistent effect on the motility, hyaluronidase and acrosin activity, and the fertilizing ability of spermatozoa in addition to the blood level of testosterone. STS-557 may have the potentiality to be used as a chemical contraceptive in the male but compensation for the reduced level of blood testosterone may be necessary.

Effects of STS 557 on semen and sperm functions in bonnet monkey (Macaca radiata)--a preliminary report.

Intramuscular administration of STS 557 (17 alpha C-cyanomethyl-17 beta-hydroxy-estra-4,9(10)-diene-3-one) at a daily dose of 12 mg/monkey for 8 weeks induced severe oligospermia to complete azoospermia by the 9th week of initiation of treatment with concomitant decrease in the serum level of testosterone. The sperm motility was reduced from the 3rd week of treatment. Complete recovery was observed by the 14th week after withdrawal of treatment. STS 557 may have the potentiality to be used as a chemical contraceptive in the male.

Effect of intra-uterine administration of dimethylsulfoxide on fertility of rats.

A single intra-uterine injection of 60 microliters of dimethylsulfoxide prevented implantation when administered before mating and induced resorption of the conceptus in rats when given during early pregnancy.