Morphological study of isolated ovarian preantral follicles using fibrin gel plus platelet lysate after subcutaneous transplantation

Ovarian and follicle transplantation may preserve fertility in young cancer survivors. In this study, we have transplanted preantral follicles using fibrin gel as a carrier and fibrin gel supplemented with platelet lysate [PL] as a rich source of angiogenic and growth factors. The purpose of this study was to evaluate the role of fibrin gel and PL in follicle transplantation. In this experimental study, ovaries were taken from 14-day-old Naval Medical Research Institute [NMRI] mice. Preantral follicles were dissected from the ovaries and encapsulated into fibrin gel supplemented with 5, 10, 15 or 20% PL, then transplanted back into the same donor mice. Fibrin gels supplemented with PL that contained preantral follicles were placed in a subcutaneous pocket in the back of the neck of the recipient, donor mouse [the same mouse that follicles were collected]. After 14 days the grafts were processed and embedded in paraffin blocks, then serially sectioned for histological evaluation. We counted the follicles and classified them according to stage [preantral or antral]. Data were presented as mean +/- standard error of mean [SEM] and analysed by analysis of variance [ANOVA] and the Kruskal-Wallistest. The mean percentage of recovered follicles encapsulated and transplanted in each group were 33.30 +/- 2.47 [fibrin gel], 31.96 +/- 1.90 [fibrin gel+5% PL], 34.02 +/- 2.44 [fibrin gel+10% PL], 48.31 +/- 2.06 [fibrin gel+15% PL] and 17.60 +/- 2.79 [fibrin gel+20% PL]. There was a significant increase in the recovery rate of grafted follicles with fibrin gel+15% PL [48.31%; p<0.001]. The percentage of preantral follicles showed no significant difference in all groups [p<0.05]. The percentage of antral follicles showed a significant decrease in follicles grafted with fibrin gel+20% PL when compared to the other groups [11.77%; p<0.005] but no significant difference was observed in the other groups. The use of PL in follicle transplantation can improve ovarian follicular survival rate

In vitro maturation, fertilization and embryo culture of oocytes obtained from vitrified auto-transplanted mouse ovary

The purpose of this study was to investigate the in vitro survival and developmental potential of oocytes obtained from vitrified mouse ovaries transplanted to a heterotopic site. In this experimental study, two-week-old mice were unilaterally ovariectomized after anesthesia. The ovaries were vitrified by cryotop. After two weeks, the ovaries were thawed and autotransplanted to the gluteus muscle tissue. Three weeks later the mice were killed, after which we removed and dissected the transplanted and opposite right ovaries. Cumulus oocyte complexes [COCs] and denuded oocytes were evaluated for in vitro maturation [IVM], in vitro fertilization [IVF] and in vitro development [IVD]. The control group consisted of sevenweek- old age-matched mice ovaries. All vitrified-transplanted [Vit-trans] ovaries contained some oocytes that survived. Following IVM, IVF and IVD, there were 41.7% out of 12 cultured zygotes that reached the 8-cell stage. Our experiment supports the progressive role of long-term graft survival after wholeovarian cryopreservation by vitrification and subsequent heterotopic transplantation. It is possible to recover viable follicles and oocytes that have the ability to develop in vitro

Prevention of di [2-ethylhexyl] phthalate-induced testicular disturbance in mice by Co-administration of L-carnitine

di [2-ethylhexyl] phthalate [DEHP] is widely used in the plastic industry and can induce reproductive toxicity. On the other hand, L-carnitine [LC] plays a crucial role in sperm metabolism and maturation. This study evaluates the effect of LC on body and testis weight, testis tissue, count, motility, viability, morphology, and chromatin quality of epididymal sperm, testicular spermatid number [TSN] per gram testis and daily sperm production [DSP] in LC-treated mice. In this experimental study, adult male NMRI mice [mean age: 4 weeks] were given doses of DEHP and LC by gavaging for 2 weeks. All samples were assessed according to World Health Organization [WHO] criteria. Sperm morphology was assessed using Papanicolaou staining and sperm chromatin quality by aniline-blue staining. The left testes were fixed in Bouin? solution for histological examination and the end slices were stained with hematoxylin and eosin [H and E]. The right testes were homogenized, and then TSN and DSP were calculated with an improved Neubauer haemocytometer and respective frames. Paired t-test, ANOVA, and Kruskal-Wallis tests were utilized for data analysis. Co-administration of DEHP and LC not only prevented significant gains in testicular weight, but also maintained the sperm's normal morphology and chromatin quality [p<0.05]. In addition, LC recovered histological changes, TSN, DSP, and sperm count. These results demonstrated that oral administration of LC partially or generally protects spermatogenesis from DEHP-toxicity in mice

Histochemical study of N-acetygalactosamine containing glycoconjugate in intraductal carcinoma of the breast by HPA lectin

Glycoconjugates are a class of glycoproteins or glycolipids, their terminal sugars are responsible for cell-cell and/or cell-extracellular matrix interactions. Aberrant glycosylation of these compounds are one of the most important aspects of cellular transformation, metastasis and escape of tumoral cells from immune system and resistance to antineoplastic drugs. Recent studies showed that patients with HPA [helix pomatia agglutinin] positive intraductal carcinoma cells have worse prognosis compared to patients with HPA negative cells. The aim of the present study was to define the presence of GalNac terminal sugar in glycoconjugate of different grades of intraductal breast carcinoma and to compare the degree and the pattern of reactivity of tumoral cells to HPA lectin. Material and The paraffin blocks belonging to 20 patients of intraductal carcinoma was chosen from pathology archive of Khatam-Al-Anbia hospital in Zahedan and 5-7 micrometer sections were prepared. Two expert pathologists determined histopathological grading independently. The lectin histochemistry was performed using HPA. The same observers determined histochemical grading. Data were analyzed by NPAR [non-parametric] test of Mann Whitney. Results of this study revealed that the pattern and the degree of histochemical reactivity of neoplastic cells differ in all grades of intraductal carcinoma. Histochemical staining showed significant difference between grades of intraductal carcinoma of the breast [p<0.003]. The lowest reactivity was seen in grade I and the highest in grade III. Furthermore, the reaction of tumoral cells was primarily confined to apical surfaces of cells in grade I, to the Golgi zone in grade II, and to a diffuse cytoplasmic distribution in grade III Our data suggest that the HPA reactivity of tumoral cells were different in all grades of intraductal carcinoma. The tumor cells showed aberrant glycosylation, which occurred in the course of anaplastic changes. It seems that our data suggest a potential and clinically important role of HPA reactivity to predict the invasive nature of malignant tumoral cells of intraductal carcinoma of the breast

Effect of Beta Mercaptaethanol with and without BSO [DL-Buthionine Sulfoximine] on Resumption of Meisosis, in vitro maturation and Embryo development of immature mouse oocytes

The purpose of this study was to evaluated the effect of beta-mercaptoethanol on resumption of meiosis, in vitro maturation of immature mouse oocytes and resulting embryo development with and without BSO [DL-Buthionine sulfoximine] Material and Germinal vesicle [GV] were recovered from 6-8 weeks old NMRI ovaries and cultured in maturation medium in MEMalpha supplemented with 7.5IU/ml hCG, 100mIU/ml rhFSH, 5% FCS [control group] and adding 100 micro m beta-mercaptoethanol [group 1] or with 5mM BSO + 100 micro m beta-mercaptoethanol [group 2] for 24h. The matured oocytes then were fertilized and cultured for 5 days. Fertilization and development were accomplished in T6 medium.The percentage of GV oocytes reaching to metaphase I [or undergo GVBD] were 78.5%, 85%, 86% in control group, group 1 and group 2 respectively, that no significant difference was detected between groups. The proportion of oocytes that progressed to the metaphase II [MII] stage was minimum within 5mM BSO group [group 2] and maximum within beta-mercaptoetanol group [group 1] with significant difference comparing with control and each other [P<=0.05]. The percentage of embryos reaching to morula stage within beta-mercaptoetanol group was significantly higher than the control group [5% and 12.2% respectively]. None of oocytes treated with BSO could pass the 8 cell stage. beta-mercaptoetanol enhances IVM and improves embryo development. While adding BSO into the maturation medium even with beta-mercaptoetanol decreases maturation and declines the embryo development

Cytotoxic and oxidativc stress caused by cadmium and lead on human skin fibroblast cells

Heavy metals are important occupational and environmental pollutants that cause damage to various organs. Although there is no effective therapy for such a poisoning, metallothionein has been shown to play a key role in the detoxification of cadmium [Cd]. Evidence in the literature suggests that superoxide dismutase, glutathione peroxidase, and catalase constitute important defense mechanisms against oxygen toxicity in the cells. The aim of this study was to investigate the effect of cadmium chloride and Pb-acetate on antioxidant enzymes in the human skin fibroblast cells [HF2FF]. The human skin fibroblast [HF2FF] cells were incubated in serum-free medium containing 20 pM CdCI[2] for 18 hr three times a week. The same exposure to an equimolar dose of Pb-acetate was performed, After each exposure and after three times exposure the cells were collected and cell viability, the contents of superoxide dismutase [SOD], catalase, glutathione peroxidase [GSH-Px], GSH and malondialdehyde [MDA] were measured. Cd caused cytotoxicity and inhibition of glutathione peroxidase [GSH-Px] and SOD activity, as well as depletion of the reduced form of glutathione [GSH] in the cell. The level of lipid peroxidation [LP] was increased, but catalase activity was not significantly altered. These defects were increased with repeated exposures. The same exposure to an equimolar dose of Pb-acetate evoked only inhibition of GSH-Px and SOD. The values of GSH, catalase and LP activity remained unchanged. The inhibition of GSH-Px and SOD may be considered as an important biomarker of the toxic effect of metals

Effect of DEHP[Di-2ethyl hexyl-phthalate] on resumption of meiosis and in -virto maturation of mouse oocytes and development of resulting embryos

DEHP [di[2-ethylhexyl] phthalate]] is widely used in plastic industry and some reproductive toxicity has been shown with it. So, this study was designed to evaluate DEHP effects on resumption of meiosis and in vitro maturation of mouse oocytes as well as development of embryos resulted from them. Mice of 4-6 weeks old were administered daily doses of 50, 100, 200 microl of 2.56 micro M DEHP solution for 12 days. Immature mouse oocytes were recovered from all experimental groups and matured in MEM-alpha medium containing 5% FCS with and without 7.5 IU hCG and 100 mIU rFSH. IVF was performed T6 medium. Resumption of meiosis and in vitro maturation were significantly lower in all experimental groups in culture media without hormones compared to controls. Fertilization and embryo development were also significantly decreased in both culture media [with and without hormones]. This study showed the adverse effects of DEHP on in vitro maturation and embryo development in a dose dependent manner