Bisphenol A-induced ultrastructural changes in the testes of common marmoset.

BACKGROUND & OBJECTIVES: Bisphenol A (BPA) is an endocrine disruptor that is widely used in the manufacture of polycarbonate plastics, epoxy resins and dental sealants. It is known to have adverse effects on spermatogenesis in rodents. This study was aimed to evaluate the effects of BPA in adult common marmoset owing to its similarities with human spermatogenesis. METHODS: Sixteen marmosets were divided into four groups (n=4 per group) and given oral doses of BPA (2.5, 12.5 and 25 µg/kg BW/day) for 70 days to cover two spermatogenic cycles, and the control group received only vehicle (honey). Testes were processed for histological and transmission electron microscopy studies. RESULTS: Histology of the testis showed sloughing of germ cells into the lumen, increase in interstitial space and vacuolation of Sertoli cell cytoplasm. Ultrastructural analysis of the testis revealed several degenerative effects on the basement membrane, Sertoli cells, Leydig cells and other developing germ cells in the 12.5 and 25 µg/kg BW/day groups as compared to control. INTERPRETATION & CONCLUSIONS: The observed ultrastructural changes caused by BPA in testicular morphology might be indicative of a perturbed sperm production. Considering the genetic and spermatogenic similarities of common marmoset (Callithrix jacchus) and humans, the study findings are of significance. Further studies are, however, needed to elucidate the mechanism of action.

Tamoxifen nanostructured lipid carriers: enhanced in vivo antitumor efficacy with reduced adverse drug effects.

A novel approach of enhancing the Tamoxifen uptake via Intestinal Lymphatic System is executed by developing long chain lipid and oil based nanostructured lipid carrier system (Tmx-NLC). The aim was to achieve improved systemic bioavailability of Tamoxifen, prevent systemic and hepatotoxicity and enhance antitumor efficacy. Following the proof of concept achieved in cell culture experiments and in vivo pharmacokinetic and biodistribution study, the current work focuses on investigation of antitumor efficacy and treatment associated toxicity in murine mammary tumor mice model. The efficacy study demonstrated greater tumor suppression and 100% survival with 1.5 and 3 mg/kg Tmx-NLC compared to 3 mg/kg Tamoxifen suspension and Mamofen(®) (Khandelwal Pharmaceuticals, Mumbai, India). Tmx-NLC treatment for a month demonstrated improved systemic toxicity profile and no evidences of hepatotoxicity. Thus, developed Tmx-NLC could prove to be a promising delivery strategy to confer superior therapeutic efficacy and ability to address the biopharmaceutical and toxicity associated issues of drug.

Buparvaquone loaded solid lipid nanoparticles for targeted delivery in theleriosis.

BACKGROUND: Buparvaquone (BPQ), a hydroxynaphthoquinone derivative, has been investigated for the treatment of many infections and is recommended as the gold standard for the treatment of theileriosis. Theileriosis, an intramacrophage infection is localized mainly in reticuloendotheileial system (RES) organs. The present study investigates development of solid lipid nanoparticles (SLN) of BPQ for targeted delivery to the RES. MATERIALS AND METHODS: BPQ SLN was prepared using melt method by adding a molten mixture into aqueous Lutrol F68 solution (80°C). Larger batches were prepared up to 6 g of BPQ with GMS: BPQ, 2:1. SLN of designed size were obtained using ultraturrax and high pressure homogenizer. A freeze and thaw study was used to optimize type and concentration of cryoprotectant with Sf: Mean particle size, Si: Initial particle size <1.3. Differential scanning calorimetry (DSC), powder X-ray diffraction (XRD) and scanning electron microscope (SEM) study was performed on optimized formulation. Formulation was investigated for in vitro serum stability, hemolysis and cell uptake study. Pharmacokinetic and biodistribution study was performed in Holtzman rat. RESULTS: Based on solubility in lipid; glyceryl monostearate (GMS) was selected for preparation of BPQ SLN. Batches of BPQ SLN were optimized for average particle size and entrapment efficiency at <100 mg solid content. A combination of Solutol HS-15 and Lutrol F68 at 2% w/v and greater enabled the desired Sf/Si < 1.3. Differential scanning calorimetry and powder X-ray diffraction revealed decrease in crystallinity of BPQ in BPQ SLN while, scanning electron microscope revealed spherical morphology. BPQ SLN revealed good stability at 4°C and 25°C. Low hemolytic potential (<8%) and in vitro serum stability up to 5 h was observed. Cytotoxicity of SLN to the U937 cell was low. The macrophage cell line revealed high (52%) uptake of BPQ SLN in 1 h suggesting the potential to RES uptake. SLN revealed longer circulation and biodistrbution study confirmed high RES uptake (75%) in RES organs like liver lung spleen etc. CONCLUSION: The high RES uptake suggests BPQ SLN as a promising approach for targeted and improved delivery in theileriosis.

Bone healing evaluation of nanofibrous composite scaffolds in rat calvarial defects: a comparative study.

We studied the in vivo performance of scaffolds consisting of nanofibrous poly(L-lactic acid) (P) and blend of poly(L-lactic acid/gelatin) (PG) prepared by electrospinning and further composited them with hydroxyapatite (HA) via alternate soaking method, to get poly(L-lactic acid)/hydroxyapatite (PH) and poly(L-lactic acid)/gelatin/hydroxyapatite (PGH) scaffolds respectively. The purpose of this study was to assess and compare bone regeneration potential of electrospun P, PG and electrospun-alternate soaked PH and PGH scaffolds using rat as an animal model by creating two 5 mm circular defects in calvaria. The respective scaffolds were implanted into the defects as one side implantation and both side implantation. Defects left empty served as a negative control for one side implantation and as sham control for both side implantations. The outcomes of the scaffold implantation were determined after 6 and 10 weeks by digital radiography, micro-CT, dual-energy X-ray absorptiometry (DEXA) and histological analysis. PGH scaffold regenerated maximum amount of new bone with high bone mineral density (BMD) into the defects and complete closure occurred in just 6 weeks while other scaffolds failed to close the defects completely. PGH group exhibited highest BMD value after 10 weeks. Histological findings showed abundant osteoblasts and initiation of matrix mineralization in HA containing scaffolds. Masson's trichrome staining showed collagen deposition in all scaffold groups except sham control group. Biochemical and haematological parameters were well with in normal range, indicating no infection due to scaffold implantation. These results prove PGH scaffold as a potential biomaterial for bone regenerative medicine.

In vivo biocompatibility evaluation of electrospun composite scaffolds by subcutaneous implantation in rat.

In vivo biocompatibility of nanofibrous poly-L-lactic acid (P), poly-L-lactic acid/gelatin (PG), poly-L-lactic acid/hydroxyapatite (PH), and poly-L-lactic acid/gelatin/hydroxyapatite (PGH) scaffolds, useful in regenerative medicine and drug delivery, was evaluated by subcutaneous implantation in both male and female rats (n = 5) for up to 90 days. The body weight of each animal in the study was evaluated on a weekly basis, and no significant difference was noticed. Total and differential leukocyte counts displayed no inflammatory reaction due to scaffold implantation. Gross observation and histology of necropsied vital organs exhibited normal morphology of cell types and tissue, denying any systemic adverse reaction on distal organs. Histology of subcutaneous tissue surrounding scaffolds was done to assess any local toxic effect of implants and found all scaffolds to be compatible and nontoxic. Moreover, no remnant of scaffolds was observed in any of the histological sections, suggesting all scaffolds to be biodegradable. All the results in this study confirm that nanofibrous scaffolds P, PG, PH, and PGH are biocompatible and safe for bone tissue engineering application.

Genotoxicity evaluation of asymmetric lipid polymer hybrid nanoparticles of doxycycline hydrochloride following intravenous administration.

Nanoparticles, being small (<1,000 nm) in size, provide high surface area-to-volume ratio as compared with the bulk materials which increase the concern about their potential toxicities. The present investigation was undertaken to evaluate the genotoxic potential of asymmetric lipid polymer hybrid nanoparticles of doxycycline hydrochloride (DH lipomer) following intravenous route. DH lipomer was prepared by modified nano-precipitation method as reported earlier. Doxycyline loading was found to be 20 ± 2.5 %. Average particle size of DH lipomer and blank lipomer was 512 ± 8 and 520 ± 6 nm, respectively. Micronucleus (MN) assay was performed in adult healthy Swiss mice whereas chromosomal aberration (CA) test and comet assay were performed in healthy Holtzman rats following intravenous administration. Animals were divided into two sets, male and female, each set comprising of six groups (n = 5/group), viz., three test groups, blank lipomer (BL), vehicle control (VC), and positive control. Groups treated with 1.5 mg/kg BW DH lipomer did not show micronuclei formation in bone marrow cell, DNA damage, and CA, respectively, as compared with VC, suggesting no genotoxicity. On the other hand 3 and 6 mg/kg BW revealed significant (P > 0.001) increase in micronuclei formation, DNA damage, and chromosomal aberrations. Furthermore, BL (6 mg/kg BW) did not reveal genotoxic response in any of the tests, suggesting lipomer components as non-genotoxic. No sex-dependent variation in genotoxicity was observed. This study therefore suggests the potential safety of the proposed dose of DH lipomer at 1 mg/kg BW. An interesting highlight of the study is safety of lipomer matrix which could be exploited for other biomedical application.

Polyethylene sebacate: genotoxicity, mutagenicity evaluation and application in periodontal drug delivery system.

The objective of the present study is to evaluate Polyethylene sebacate (PES) for its toxicity profile including oral toxicity, genotoxicity and mutagenicity. PES was synthesised, and characterised by gel permeation chromatography, FTIR, (1)H-NMR, differential scanning calorimetry and X-ray diffraction. Oral toxicity studies revealed PES to be nontoxic up to 3000 mg/kg body weight with no significant changes in serum biochemistry. The standard battery of genotoxicity tests including micronucleus test, chromosomal aberration and comet assay revealed PES as nongenotoxic. Mutagenicity of PES was evaluated using the Ames microplate format mutagenicity assay sample kit using TA98 and TA100 strains of Salmonella typhimurium, both in presence and absence of Aroclor 1254 induced rat liver S9. Ames assay confirmed PES to be nonmutagenic. Periodontal implants of PES of varying roxithromycin/PES ratios and different diameter were prepared. A decrease in in vitro drug release was seen with increase in diameter of the implants. Release rates, however, increased with increase in PES concentration, and were attributed to decreased crystallinity of roxithromycin, confirmed by the DSC thermographs and XRD spectra. Roxithromycin release from the implants followed Higuchi kinetics and exhibited controlled release. The results suggest PES as a safe polymer for biomedical and pharmaceutical applications.

Effect of active immunization with the C-terminal 67-94 (R-28) region of human seminal plasma inhibin on the fecundity of adult male rabbits.

PROBLEM: To characterize the specificity of antibodies to the synthetic C-terminal 67-94 (R-28) peptide of human seminal plasma inhibin (hSPI), and to examine the effect of active immunization on the fecundity of adult male animals. METHOD OF STUDY: The specificity of R-28-DT antibodies was tested using enzyme-linked immunosorbent assay, immunohistochemical and immunofluorescence staining, and Western blotting. For fertility studies, adult male rats and rabbits were immunized and mated with females of the same strain. RESULTS: Rabbit antibodies to R-28-DT recognized native hSPI, as demonstrated by sperm agglutination in vivo and in vitro, on the sperm head by immunofluorescence staining, and in the columnar epithelial cells of the prostate by immunohistochemical staining. Immunization with the R-28-DT conjugate elicited a poor antibody response in male rats and their fecundity remained unaffected, while in male rabbits it elicited a good immune response with reduction in their fertility. CONCLUSION: R-28-DT antibodies recognized the native hSPI in the prostatic epithelium and agglutinated washed rat, rabbit, monkey, and human spermatozoa in vitro. Immunization of rabbits caused agglutination of spermatozoa resulting in a decrease in their fecundity. The conjugated R-28 peptide of hSPI offers promise as a male contraceptive.