[Effects of Vitrification with Self-made Carriers and Slow Programmed Freezing on Ovarian Tissue of Sheep].

Objective To investigate the effects of self-made carriers on the cryopreservation of ovarian tissue of sheep. Methods Thirty-two ovaries were randomly assigned to fresh group,programmed freezing group,self-made carrier I vitrification group,and self-made carrier Ⅱ vitrification group.The morphology,proliferation,apoptosis,and estrogen level of the ovarian tissue in each group were observed. Results After cryopreservation,the morphology normal rate of the primordial follicles in programmed freezing group,self-made carrier I vitrification group,and self-made carrier Ⅱ vitrification group were 74.2%,72.8%,and 72.3%,respectively,lower than that(83.7%)in the fresh group(χ2=13.079,P=0.004).The percentage of normal primary follicles in programmed freezing group was lower than that in the fresh group(χ2=12.486,P=0.000).The percentage of normal primary follicles showed no significant difference between vitrification groups and fresh group(P=1.000,P=0.972).There was no significant difference in estrogen level or the positive expression rate of PCNA among the 4 groups(F=0.363,P=0.780;χ2=0.359,P=0.949).The number of apoptotic cells in cryopreservation groups was significantly higher than that in the fresh group(F=37.584,P=0.000),and it was significantly higher in the programmed freezing group than in the two vitrification groups(F=18.992,P=0.000). Conclusion Compared with slow programmed freezing,the vitrification with self-made carriers could well preserve the activity of cells in large sheep ovarian tissue blocks.

Preparation of transparent zein films for cell culture applications.

In our previous studies, zein has shown good cell compatibility in both films and porous scaffolds. To use the zein film in microfluidic devices or as a cell culture substrate, swelling and excessive degradation should be avoided. Moreover, the film should be transparent. In this study, we found that the zein film could maintain good transparency even after swelling when it was treated for 20min at 121°C, 100% relative humidity and 103.4kPa, especially for the zein film without the addition of a plasticizer. FT-IR and XRD analysis showed that the ratio of α-helix of zein structure decreased and the ratio of its ß-sheet increased. Proliferation of NIH 3T3 cells on the transparent zein film was as cytocompatible as the untreated one and the cell culture plate. The improved transparency of zein films after swelling will expand the application field of zein.

Cell adhesion on zein films under shear stress field.

Vascular implants after implantation need to improve the ability of cells to withstand flow-shear stress. As such, we want to test whether zein films could improve the flow-shear stress resistance of cells by control of their surface morphology. We chose Collagen, poly L-lactic acid (PLLA) and three types of zein as the coating films and evaluated the flow-shear stress resistance of NIH3T3, and EA.hy926 on these respective films. The results showed that the retention of two cell lines on Collagen film was better than PLLA and zein films. The cell retention of EA.hy926 on Zein 3 film with higher roughness was better than Zein 1 film with a flat surface in the first 2h. The cell retention of NIH3T3 on a rougher surface was always better than the smoother one under flow-shear stress condition for 6h. Observation of cell morphologies showed that the aspect ratio changed significantly for NIH3T3 cells upon flow-shear stress condition, as shown by reduced numbers of pseudopodia, increased cell rounding and shrinkage. Zein 3 film with higher roughness improved the flow-shear stress resistance of cells and might be used in vascular implant coatings.

[Study on positive rate of blocking antibody in women with recurrent spontaneous abortion administered by route and frequency of paternal lymphocyte immunotherapy].

OBJECTIVE: To investigates factors affecting the positive rate of blocking antibody treated by paternal lymphocyte immunotherapy in patients with recurrent spontaneous abortion (RSA). METHODS: From January 2008 to August 2012, 326 RSA cases undergoing treatment in Infertility Center of Qilu Hospital were studied retrospectively. Those patients were divided into 2 groups randomly: 260 cases in intradermal injection group were administered via bilateral forearm intradermal injections for immunotherapy once 21 days, then the blocking antibody was determined after 2 (23 cases) , 3(73 cases), 4 (74 cases) , 5(90 cases) times respectively, while in subcutaneous injection group, the 66 cases were administered via subcutaneous injection once 21 days, the blocking antibody measured after 3 times; In both cases, the blocking antibody was all determined 2 weeks later. The positive rate of blocking antibodies and the rate of successful pregnancy was recorded, and then followed up after the blocking antibody turning positive. RESULTS: (1) Positive rate of blocking antibodies:the positive rate of blocking antibodies were 17% (4/23) , 58% (42/73), 72% (53/74) and 84% (76/90) in the 2, 3, 4, and 5 times of intradermal injection group, respectively (P < 0.05). In subcutaneous injection group, the positive rate of blocking antibodies was 38 % (25/66), which was significantly lower than that in group intradermal injection receiving 3 times immunotherapy (P < 0.05). (2) The rate of pregnancy:the 176 patients out of 200 patients were pregnant when antibody was positive after immunotherapy, with 71.6% (126/176) of patients gained successful pregnancy(the length of pregnancy more than 5 months). CONCLUSIONS: The route and frequency of administration of immunotherapy could influence the positive rate of blocking antibody. The rate of successful pregnancy will be increased after blocking antibody turning positive.