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目的:探讨使用非渗透性冷冻保护剂蔗糖快速冻存人类精子的可行性。方法:收集正常精液标本40份,经上游法处理后,收集上游精子悬液,分成6份,1份作为对照,其余5份分别用常规精子冷冻法,0.15、0.20、0.25、0.30 mol/L蔗糖溶液冷冻精子,复苏后比较6组精子活动率、前向运动精子活动率及精子质膜完整性。结果:所有冷冻组复苏后精子活动率、前向运动精子活动率及膜完整率与冷冻前上游组(96.2±1.8)%,(93.8±2.8)%,(99.0±0.8)%比较均显著下降,差异有显著性(P<0.05)。精子活动率结果:蔗糖冷冻组之间比较,0.20 mol/L组解冻后精子活动率高于0.15、0.25、0.30 mol/L组,分别为(55.5±6.3)%、(45.9±6.6)%、(50.4±9.4)%、(45.5±11.2)%,差异有显著性(P<0.05)。0.20、0.25 mol/L蔗糖冷冻组与常规慢速冷冻组(53.6±5.0)%比较,差异均无显著性(P>0.05);0.15、0.30 mol/L蔗糖组均低于常规慢速冷冻组(P<0.05)。前向运动精子活动率结果:0.20 mol/L蔗糖冷冻组与常规冷冻组比较差异无显著性[(44.4±7.4)%vs(42.3±8.1)%,P>0.05];以上两组均高于0.15、0.25、0.30 mol/L蔗糖冷冻组,分别为(37.1±8.3)%、(33.1±9.2)%、(22.0±9.1)%,差异有显著性(P<0.05)。精子膜完整率结果:0.20 mol/L蔗糖冷冻组(70.1±6.9)%高于常规冷冻组、0.15、0.25、0.30 mol/L蔗糖冷冻组,分别为(63.1±6.8)%、(57.7±8.3)%、(63.5±10.7)%、(57.8±12.9)%,差异有显著性(P<0.05);常规冷冻组与0.15、0.25、0.30 mol/L蔗糖冷冻组比较差异无显著性(P>0.05)。结论:终浓度为0.20 mol/L的蔗糖溶液作为人类精子冷冻保护剂是可行的,快速冷冻方法简便、快速、经济、安全有效。
Objective: To explore the feasibility of cryopreservation of human sperm using sucrose, an impermeable cryoprotectant. Methods: 40 samples of normal semen were collected and processed by the upstream method. The sperm suspension was collected and divided into 6 parts and 1 part as control. The other 5 samples were treated by conventional sperm freezing method, 0.15, 0.20, 0.25 and 0.30 mol / L Sucrose solution was used to freeze sperm. After resuscitation, the sperm motility rate, motility sperm motility and sperm plasma membrane integrity were compared. Results: The sperm motility rate, motile sperm motility rate and membrane integrity rate of all frozen groups after resuscitation were significantly decreased compared with those of the pre-freezing group (96.2 ± 1.8%), (93.8 ± 2.8)% and (99.0 ± 0.8)% , The difference was significant (P <0.05). Results: The sperm motility of the 0.20 mol / L group was higher than that of the 0.15,0.25,0.30 mol / L group (55.5 ± 6.3%, 45.9 ± 6.6%, respectively) (50.4 ± 9.4)% and (45.5 ± 11.2)%, respectively, with significant difference (P <0.05). There was no significant difference between 0.20,0.25 mol / L sucrose group and 53.6 ± 5.0% group (P> 0.05), while those of 0.15,0.30 mol / L sucrose group were lower than those of conventional slow freezing group (P <0.05). The results of forward motility sperm motility showed that there was no significant difference between the 0.20 mol / L sucrose group and the conventional group [(44.4 ± 7.4)% vs (42.3 ± 8.1)%, P> 0.05] (37.1 ± 8.3)%, (33.1 ± 9.2)%, (22.0 ± 9.1)%, respectively. The difference was significant (P <0.05). Results showed that the membrane integrity rate of sperm membrane was (63.1 ± 6.8)% and (57.7 ± 8.3)% respectively in the 0.20 mol / L sucrose freezing group (70.1 ± 6.9)% compared with the conventional freezing group, 0.15,0.25,0.30 mol / L sucrose freezing group ), (63.5 ± 10.7)% and (57.8 ± 12.9)%, respectively. There was no significant difference between the conventional freezing group and the 0.15,0.25,0.30 mol / L sucrose freezing group (P> 0.05). CONCLUSION: Sucrose solution with a final concentration of 0.20 mol / L is feasible as cryoprotectant for human sperm. The rapid freezing method is simple, rapid, economical, safe and effective.