The Experimental Study Of Microencapsulated Newborn Rats Ovaries In Vitro And Allotransplantation

The ovary is the most important endocrine and reproductive organ in the female. Due to premature ovarian failure, ovarian heteroplasia or absence, pathological ovariotomy, and after the actinotheraphy and chemotherapy for those with cancer, the ovary could be lost it's functions. It will be caused some serious problems such as the incretion maladjustment or lost the ability of pregnancy. There are some serious side effects in the traditional hormone replacement therapy (HRT). It will be raise the rate of endometrium pathological changes and the rate of the breast caner. All of these caused by the non- physiological secretion of estrogen. There are many limits in hormone replacement therapy if the patients have the thrombus disease (Crus thrombus phlebitis) or breast cancer, and SLE disease.As the same time, according the improvement of combination of surgery,chemical therapy and radiation therapy, the long term survival rate in young malignancy patients is increased, but these therapy always damage sex organ. It will make serious effect in patients' survival quality and procreate ability. As one of the therapies to recover the ability of female incretion and procreation, ovarian tissue transplantation is paid more attention by the scientists.Transplantation of reproductive organs, including ovarian tissue transplantation , started over 100 years ago using both human and animal models.Because the effect was far from expected, this technique has been abandoned for a long time, however, as the ovarian cryopreservation and the immunity-transplant technology were developed, people find the great potential for ovarian transplantation that has been applied in clinical treatment.Alginate-poly(L)lysine-alginate (APA) membrane has been developed for the microencapsulation of living cells which protects these cells from the immune system. It was preparaed for the first time by Lim and Sun in 1980's. APA microcapsules have semipermeable membranes which permit passage of low molecular weight substance, such as nutrients and oxygen, but not of cells and high molecular weight substance. The structure of the membrane is one layer poly(l)lysine which insert into two layers alignate. The special cells were packaged into the microcapsule membrane. Due to the microcapsule membrane has semi-permeable ability, the small molecules like oxygen and nutrients can pass the membrane to supply the cells that insert into the membrane to keep the cells activity. And the biological activity substance that secreted by the cells can pass the membrane to some special effects. As the same time, the big molecules like cells and immunity substances can not pass the microcapsule membrane to attack the cells that in the microcapsule. It shows that APA microcapsule has good function of immunity isolation.APA microencapsulated biological active molecules, tissue, and cells for allo-or xenograft transplanting have been studied by many researchers. In other incretion deficiency diseases, the scientists found that the microencapsulated cells transplantation can solve the non- physiological secretion of the hormone replacement therapy, and it can avoid the immunity repellency .In the modern society, the scientists already focus on the microencapsulated cells transplantation in medical and biotechnology area. There are already many researches in thyroid gland, pancreas, parathyroidism, pituitary, chromaffin cells.In this study,we use the alginate—polylysine^-alginate(APA) microcapsule as the method of immunity isolation. The APA microencapsulated newborn rats ovaries were prepared with a high-voltage electrostatic system,and then cultured in RPMI 1640 medium containing 20 % FCS at 37t: and 5 % CO2, the growth of APA microencapsulated newborn rats ovaries were monitored. At intervals, the estradiol (E2) and progesterone(P) secreted from APA microencapsulated newborn rats ovaries were measured . The study by electron microscope showed that the growth of microencapsulated newborn rats ovaries can be clearly visualized. Then the APA microcapsules were transplanted to the peritoneal cavitie of castrated adult female rats, and the E2 and P secretion were measured periodically. The recovery and maintaining time of the oestrus cycle were carefully observed, and the endocrine function of transplanted ovaries was determined by measuring serum E2 and P. The transplanted microcapsules were recovered at intervals to explore the feasibility to transplant the APA microencapsulated ovarian tissue.We found that the APA microcapsules prepared in the same condition of high-voltage electrostatic system were round and homogeneous. The newborn rat ovaries were grown in the microcapsules after microencapsulation, and some tissues have the tendency to grow along the microcapsule wall. The E2> P secreted by microencapsulated newborn rats ovaries were shown the permeate of the membrane of APA microcapsules.The concentration of E2 and P was reached the highest levelat 14th and 21st day., the concentration of E2 and P dropped the little bit first and keep stable afterwards. Since the days going ,the develop of granulose cells in ovaries were founded by transmission electron microscope. There were many luteal granulose cells in the APA microencapsulated newborn rat ovaries after 4 weeks in vitro. The conformation of the nucleuses was big and the chromatins were normal. There were many compositions in the cytopalst, like golgi apparatus, endoplasm and mitochondria. It was showed that ovaries have grown well in the microcapsule.Following the results in vitro, the biocompatibility of APA microencapsulated newborn rats ovaries were studied in vivo. After transplanted to the peritoneal cavitie of castrated adult female rats, APA microcapsules were recovered from the peritoneal cavities periodically. The majority of microcapsules were found to be freely floating in the abdominal cavities both experiment and control group. Moreover, gross examination of the abdominal contents did not reveal any tuberculum or tumor growth in all rats of all groups. The level of E2^ P in experiment group were raised gradually ,but the oestrus cycle were not resumed. It was also showed that ovaries have grown well in the microcapsule.In addition, the microencapsulated ovaries in vivo were growing faster than in vitro.In conclusion, the E2 and P were secreted continually and steadily by the APA microencapsulated newborn rats ovaries prepared by high-voltage electrostatic system. And the ovarian function of castrated adult female rats were improved throgh transplantated APA microcapsule. The microcapsules have good biocompatibility in vivo.We found it was feasibility that the APA microencapsulated ovaries was one of technique in ovarian tissue transplantation.