ObjectiveTo determine if the cryoprotectant solution supplementation with Vitamin C can improve the protective effect of human ovarian tissue cryopreservation by anti-apoptosis. MethodsHuman ovarian cortical tissues were collected from nine patients treated between March 2012 and April 2013. The cortical tissue pieces obtained from each patient were divided into two groups:control (conventional slow freezing) and trial group (slow freezing supplementation with Vitamin C). The preservation effects in the two groups were compared by histology using light microscope and apoptosis assessed by TUNEL assay. ResultsThe proportion of morphologically normal primordial follicles in the trial group was higher than that in the control group (P<0.05). The proportion of apoptotic primordial follicles and stromal cells in the trial group was lower than that in the control group (P<0.05). ConclusionCryoprotectant solutions supplementation with Vitamin C can improve the preservation of primordial follicles and stromal cells. It might be a method worth to try in order to improve the protective effect of human ovarian tissue cryopreservation by inhibiting of apoptosis.
ObjectiveTo compare the effect of cryoprotective vitrification agent with different concentrations on protecting human ovarian tissue. MethodsHuman ovarian biopsy tissues were obtained from nine patients between August 2013 and May 2014. The cortical tissue pieces obtained from each patient were cryopreserved using direct cover vitrification (DCV) with two different concentrations. The vitrification solutions were divided into two groups: high concentration [15% (V/V) dimethyl sulphoxide (DMSO)+15% (V/V) ethylene glycol (EG)+0.5 mol/L sucrose] and low concentration group (12% DMSO+12% EG+0.5 mol/L sucrose). The preservation effects in the two different concentration groups were compared by histologic evaluation using light microscope and apoptosis assessed by terminal dexoynucleotidyl transferase-mediated nick end labeling staining. ResultsThere was no significant difference in the proportion of morphologically normal primordial follicles between high concentration group and low concentration group (P ＞ 0.05) . The proportion of apoptotic primordial follicles in the low concentration group was 29.7% (58/195) , and was 42.1% (69/164) in the high concentration group, with a significant difference between the two groups (P ＜ 0.05) . The proportion of apoptotic stromal cells in the low and high concentration group was 30.2% (162/537) and 41.9% (206/492) respectively with a significant difference (P ＜ 0.05) . ConclusionsVitrification solutions with lower concentration can improve the preservation of the primordial follicles and stromal cells in human ovarian tissue. It is a method worth trying in order to improve the protective effect of vitrification by decreasing the toxicity of vitrification solutions.
ObjectiveTo determine if the anti-apoptosis agent can improve the protective effect of human ovarian tissue with novel vitrification method. MethodsHuman ovarian cortical tissues were collected from ten patients treated between October 2012 and August 2013. The samples obtained from each patient were divided into two groups:control (the novel immersed vitrification) and experimental group (the novel immersed vitrification supplemented with vitamin C). The preservation effects in the two groups were compared by ultrastructure using electronic microscopy, and apoptosis was assessed by terminal deoxynucleotidyl transferase-medicated dUTP nick-end labeling (TUNEL) staining. ResultsThe proportion of normal ultrastructure of oocytes and granulosa cells in the experimental group was higher than that in the control group (P<0.05). The proportion of TUNEL-positive primordial follicles in the experimental group was 21.6% (49/227) and the proportion of TUNEL-positive primordial follicles in the control group was 38.6% (91/236), with a significant difference between the two groups (P<0.001). The proportion of TUNEL-positive stromal cells in the experimental group was (21.33±3.41)% and the proportion of TUNEL-positive stromal cells in the control group was (33.46±3.09)%, also with a significant difference between the two groups (P<0.001). ConclusionAnti-apoptosis agents can improve the preservation of primordial follicles and stromal cells by inhibiting of apoptosis. It may be a method worthy of trying in order to improve the protective effect of human ovarian tissue vitrification.