ABSTRACT
BACKGROUND: Vitrification is a comparatively new technology which applies high concentration cryoprotectant and rapid refrigeration. By the method, the cells were quickly frozen and to avoid damage by ice crystals inside and outside. OBJECTIVE: To compare the effect of four cryoprotectants on morphology and function of ovarian tissue in rats after vitrification. METHODS: The rats were randomly assigned into six groups with 6 rats for each: DMSO + EG, DMSO + EG + sucrose, DMSC +EG + sucrose + acetamide, EG + sucrose + acetamide, ovariectomized, and normal control groups. The ovarian tissues of four freezing groups were treated with the corresponding cryoprotectants, the vitrified ovarian tissues were then resected but not frozen and transplanted; otherwise, tissues were not treated with any treatment in the normal control group. Two weeks after freezing, the tissues were thawed and heterotopic-transplanted into femoribus intemus of hind limb. At 30 days after implantation, vaginal epithelial cells and estrus cycle were observed, while after three months, blood were collected to detect the level of estradiol (E2) and the ovarian tissues were reclaimed to analyze their morphological changes. RESULTS AND CONCLUSION: All ovarian tissues were damaged after cryoprersarvation in four freezing groups. The rates ot healthy primordial follicles were 67.9%, 71.6%, 80.5%, and 59.4%, respectively, while healthy primary follicles were 41.6%, 52.3%, 55.9%, and 36.7%, respectively. In all freezing groups, the rate of the healthy follicles in DMSO + EG + sucrose + acetamide group was higher than DMSO + EG group and EG + sucrose + acetamide group (P < 0.05). No significant difference was found in the proportion of follicles at different development stages among four groups. The typical secondary follicle was not found in four groups. Damaged ovotid showed oocyte pyknosis and vacuolation in cytoplasmic area. There was not typical cell type of all freezing groups. Ovarian autografting gained visible vascularity from surrounding tissue that connected ovarian tissue to form net. There was a lot of blood capillary in transplanted ovarian tissues and clumped primordial follicles in cortical substance. The rates of primary follicles and secondary follicles were lower than primordial follicles. The level of serum estradiol was obviously decreased compared with normal control group (P < 0.01). There was significant difference between DMSO + EG + sucrose + acetamide group and other three freezing groups (P < 0.05). Four kinds of freezing methods have poor effects on different stages of follicles and the structure of ovariarn tissue. DMSO + EG + sucrose + acetamide group is an optimal protocol for cryoprerserving ovarian tissue. Freezing methods still need to explore further because the rats had not appeared disciplinary estrus cycle after ovarian autoqrafting.
ABSTRACT
AIM: To study the effect of the radix sophorae flavescentis on cellular immunity in rats with Pneumocystis Carinii Pneumonia (PCP) induced by long-term use of immunosuppressant, and explore the action of traditional Chinese medicine for the immunological regulation and infectious prevention after organ transplantation. METHODS: The experiment was conducted at Department of Pathobiology, Jilin Medical College from May 2005 to March 2006. Forty adult healthy female SD rats were selected from Harbin Medical University (Certification: 02473146) and randomly divided into experiment group and control group, with 20 rats in each. The model of PCP was set up by glucocorticoid injection subcutaneously to SD rats (25 mg once, 2 times/week). The mixture of sophorae flavescentis was given to stomach with tube in experiment group (3 mL/kg, 2 times/day), and was consisted of radix sophorae flavescentis, ash bark, amur cork-tree, malt, milkvetch root and danshen root. Six weeks later, all the rats were anesthetized and broncholveolar lavage fluids were collected.①Alveolar washing fluid was concentrated 10 times and the levels of the soluble interleukin-2 receptor (sIL-2R) were examined by double antigen sandwich ELISA.②Blood was sampled from rat eyes and the count of lymphocytes in peripheral blood were detected.③The percentages of CD3+ and CD4+ subgroups were assessed with erythrocyte chaplet kit sensitized by antigen. RESULTS: All 40 rats were involved in the result analysis without drop.①The count of lymphocytes in peripheral blood in experiment group was significantly higher than that in control group (5.1?1.3)%, (0.8?0.3)%, P
ABSTRACT
The experiment was performed at the Institute of Treatment and Prevention of Tumor of Jilin Province and Department of Etiology, Jilin Medical College from October 2004 to June 2005. The normal umbilical cord blood (UCB) was provided by a healthy lying-in woman from Changchun Hospital with the permission of the pregnant woman. Full-term normal delivery and UCB of healthy pregnant women was collected by aseptic venepuncture after fetal disengagement, and with Natrium Citricum anticoagulation. After the UCB mononuclear cells were isolated by Ficoll-paque (relative volume mass: 1.077?0.22) density gradient centrifugation, long-term liquid culture system was used for UCB adherent cells, and co-cultured with UCB mononuclear cells. The adherent cells were observed and mononuclear cell cycles were tested with flow cytometer. It showed that adherent layer of UCB increased UCB mononuclear cells expansion as compared with that of non-adherent layer of UCB after co-culture for 7 days. The cell percentage in S+G2+M phase obviously increased, and there was significant difference [(42.7?1.1)%,(35.5?2.8)%,P