The Experimental Study On Culture In Vitro And Transplantation Of Frozen-thawed Human Ovarian Tissue

Objective1. To compare the influence on the development of follicles inthawing human ovarian tissue between different culture methods,in order to seek bettermethods of cultivaion.2. To investigate the survival of tissue and follicles aftertransplanting human thrawing ovarian tissue,in order to reduce graft ischemic injury.Methods Ovarian tissues of15ovarian teratoma patients in Obstetric and Gynecologyof the First Affiliated Hospital of Anhui Medical University from January to April,2012.Using vitrification technology preservation,and vitro culture using different tissueculture methods after recovery. According to different methods,culture were dividedinto three groups: The thawed ovarian tissues were cultured alone(group A),Coculturing the thawed ovarian tissues with the decidua monolayer cells(group B),Culturing the thawed ovarian tissues with Matrige-coated flask(group C). After14days,fixing some tissue and detecting the total follicle survival rate(TFSR)and the growfollicle rate(GFR) of every groups, some tissue from groupB and groupC weretransplanted subcutaneously into nude mice. The transplant group were divided intothree groups depending on the different transplanted ovarian tissue(four nude mice ineach group,eight pieces transplant ovarian tissue): transplanting thrawing ovarian tissuedirectly as control(O group), transplanting after thrawing human ovarian tissueco-cultured with human decidua monolayer cells(B1group), transplanting afterthrawing human ovarian tissue cultured by Matrige medium(C1group). A month aftertransplantation,taking out the graft for histological examination.Results1.TFSR and GFR of A group were significant lower than group B and C(P<0.05); There were no difference between group B and C.2.Compared with group O,the recovery and survival of transplanted ovarian tissue of group B1and C1and GFR of follicles from recovered ovarian tissue had no significant difference(P>0.05). Therewere no significant difference in the recovery、survival and GFR of follicles between Band C groups(P>0.05).Conclusions The primordial follicles developed in different speed in different culturesystems. Results of culturing the thawing ovarian tissues with the decidua monolayercells or Matrige are better than culturing alone, but there were no obvious differencebetween these two culture methods. We consider replacing Matrige medium by humandecidua monolayer cells to avoid the contamination of animal origin,but we also need alarge sample research for widely use. Reasons of bad transplant outcome:1.Drawn frompoor tissue,the follicle density in ovarian cortex of teratoma patients is lower thannormal people.2.The assessment method of ovarian tissue viability is limited,thedetected sections of ovarian tissue can not be completely representative of the wholetransplanted tissue.3.The transplant position is not good. Although nude mice canprovide sufficient grow space for graft, but the vascular subcutaneously into nude miceis not as rich muscle,which may lead to slow revascularization after transplantation.Based on factors above, the situation of the survival and growth of transplanted ovariantissue can be improved by changing unfavorable factors, and to provide more data tofuther elucidate the influence of in vitro culture on transplant outcomes.