Ovarian stimulation by exogenous gonadotropin decreases the implantation rate and expression of mouse blastocysts integrins

Integrins are heterodimeric glycoprotein receptors that regulate the interaction of cells with extracellular matrix and may have a critical role in implantation. The aim of this study was to investigate the effect of ovulation induction on the expression of Alpha4, Alphav, Beta1, and Beta3 integrins in mouse blastocyst at the time of implantation. The ovarian stimulated and non-stimulated pregnant mice were sacrificed on the morning of 5[th] day of pregnancy. The blastocysts were collected, and the expression of Alphav, Alpha4, Beta1, and Beta3 integrins was examined using real-time RT-PCR and immunocytochemical techniques, then their ovarian hormones were analyzed at the same time. The implantation sites in uterine horns of other pregnant mice in both groups were determined under a stereomicroscope on the 7[th] day of pregnancy. The results showed that the expression of Alphav, Beta1, and Beta3 integrins in both mRNA and protein levels was significantly lower in the ovarian stimulated group than the control group, and the maximum ratio of expression was belonged to Beta1 molecule [P>0.05]. The implantation rate in superovulated mice was significantly lower than control mice. It was suggested that ovulation induction decreased the expression of Alphav, Beta1, and Beta3 integrins of mouse blastocysts

Ultrastructural changes of corpus luteum after ovarian stimulation at implantation period

To achieve multiple oocytes for in vitro fertilization, ovulation induction is induced by gonadotropins; however, it has several effects on oocytes and embryo quality and endometrium receptivity. The aim of this study was to assess ultrastructural changes of corpus luteum after ovarian induction using human menopausal gonadotropin [HMG] and human chorionic gonadotropin [HCG] during luteal phase at implantation period. Female NMRI mice [6-8 weeks] were divided into control and stimulated groups. In the control group, the mice were rendered pseudopregnant and in the ovarian induction group, the mice were rendered pseudopregnant after the ovarian induction. The samples were obtained from the ovary in each group at the same time during luteal phase at implantation period. Ultrastructural changes were assessed using electron microscopy study. Our results displayed some identifiable changes in ultrastructure of corpus luteum in ovarian induction group. These changes included enhancement of the apoptosis and intercellular space, whereas the angiogenesis was decreased. The findings indicated a decline in organelle density in the cytoplasm of ovarian induction, such as mitochondria, endoplasmic reticulum and polyribosome. Furthermore, chromatin condensation of nuclei was observed in some cells. The ovarian induction using HMG and HCG resulted in some ultrastructural changes on the corpus luteum at implantation period, which could affect on the pregnancy rate

Effect of ovarian stimulation on the endometrial apoptosis at implantation period

Apoptosis is a process that plays an important role during early stage of implantation. The aim of this study was to investigate the incidence of apoptosis in mice endometrium after ovarian stimulation at implantation period. NMRI female mice were divided into two groups: 1] control group, which were rendered pseudopregnant by vaginal stimulation and 2] experimental group, which were stimulated using an intrapritoneal injection of 10 IU hMG followed by another injection of 10 IU hCG after 48 h. In the evening of the second injection, the mice were rendered pseudopregnant the same as control group. Samples were obtained from 1/3 middle part of uterine horns during implantation period. Apoptosis was assessed in two groups at implantation period using light and electron microscopic studies, TUNEL staining and semiquantitative RT-PCR. Our morphological and ultrastructural results showed apoptosis in both groups, while TUNEL analysis showed that the percentage of TUNEL-positive cells was higher in stimulated group than in the control group [P

Transplantation and homing of mouse embryonic stem cells treated with erythropoietin in spleen and liver of irradiated mice

The present study was designed to evaluate the homing potential of mouse embryonic stem cells [ESC] treated with erythropoietin [EPO] in hematopoietic organs such as spleen and liver after transplantation using morphological and immuno-histochemical techniques. Day-four embryoid body [EB]-derived cells were dissociated and re-plated in medium in the presence and absence of EPO for three days. The EPO- and untreated differentiated cells were labeled with 5-bromo-2 deoxyuridine [BrdU] before transplantation and analyzed using flow cytometry and reverse transcription-PCR methods. BrdU-labeled cells were injected via the tail vein into irradiated adult mice in both groups. The spleen colony-forming unit assay [CFU-S] was performed 12 days after transplantation. Immuno-histochemistry was also carried out to trace transplanted cells. The percentage of CD34 positive cells was 5.51 +/- 1.06% in the EPO-treated group and 1.63 +/- 0.225% in untreated group. The RT-PCR analysis showed that the EPO-treated cells expressed epsilon globin, beta H1 globin, RUNX1 and EPO receptor genes, but the beta-major globin gene was not expressed. The number of colonies formed in the spleens of treated group [17.33 +/- 4.726] was significantly different from the control group [6 +/- 1]. The population of BrdU positive cells in spleen of EPO-treated cell-transplanted group was higher than that of the control group. Also, BrdU positive cells were observed in the central vein of the liver sections of EPO-treated and control groups but were not observed in the liver parenchyma. There were not BrdU positive cells in the spleen and liver sections of the sham group. Our results confirm that ESC have the ability to home and form colonies in spleen after transplantation and EPO-treated EB-derived cells caused an increase in the number of colonies in spleen after CFU-S

histological characteristics of cultured oral epithelium in different culture conditions

This study was undertaken to establish the characterization of cultured oral mucosal epithelium and introducing them as an alternative source for reconstruction of ocular surface disease. Human oral epithelial cells were cultured on simple media [DMEM/HF12] as control and co-cultured on mitomycin C-treated 3T3 feeder layer, on the amniotic membrane [AM] without nitrocellulose and the mitotically inactivated 3T3 fibroblast, and on the sandwich layer of AM fastened on the nitrocellulose as insert and 3T3 fibroblast. After 3 weeks, the characteristics of the cells were assessed morphologically and also ultrastructurally using scanning electron microscopy and transmission electron microscopy and immuno-cytochemically. The epithelial cells were cultured on AM spread on nitrocellulose insert and 3T3 feeder layer showed better growth than other groups and all groups of study were shown similar characteristics. The cultured oral epithelial shared the characteristics with corneal epithelium. Thus the oral epithelial could be an alternative source for transplantation