Background： L-proline is a natural, nontoxic cryoprotectant that helps cells and tissues to tolerate freezing in a variety of plants and animals. The use of L-proline in mammalian oocyte cryopreservation is rare. In ...Background： L-proline is a natural, nontoxic cryoprotectant that helps cells and tissues to tolerate freezing in a variety of plants and animals. The use of L-proline in mammalian oocyte cryopreservation is rare. In this study, we explored the cryobiological characteristics of L-proline and evaluated its protective effect in mouse oocyte cryopreservation. Methods： The freezing property of L-proline was detected by Raman spectroscopy and osmometer. Mature oocytes obtained from 8-week-old B6D2F 1 mice were vitrified in a solution consisting various concentration of L-proline with a reduced proportion ofdimethyl sulfoxide （DMSO） and ethylene glycol （EG）, comparing with the control group （15% DMSO and 15% EG without L-proline）. The survival rate, 5-methylcytosine （5-mC） expression, fertilization rate, two-cell rate, and blastocyst rate in vitro were assessed by immunofluorescence and in vitro fertilization. Data were analyzed by Chi-square test. Results： L-proline can penetrate the oocyte membrane within 1 min. The osmotic pressure of 2.00 mol/L L-proline mixture is similar to that of the control group. The survival rate of the postthawed oocyte in 2.00 mol/L L-proline combining 7.5% DMSO and 10% EG is significantly higher than that of the control group. There is no difference of 5-mC expression between the L-proline combination groups and control. The fertilization rate, two-cell rate, and blastocyst rate in vitro from oocyte vitrified in 2.00 mol/L L-proline combining 7.5% DMSO and 10% EG solution are similar to that of control. Conclusions： It indicated that an appropriate concentration of L-proline can improve the cryopreservation efficiency of mouse oocytes with low concentrations of DMSO and EG, which may be applicable to human oocyte vitrification.展开更多
目的探讨在冷冻保护剂中添加槲皮素对羊卵巢组织冷冻复苏后卵泡活性的影响。方法将36只性成熟母羊的卵巢皮质组织块随机分配到新鲜对照组(CON组),玻璃化冷冻组(VIT组),添加低、中、高浓度(1、5、10μmol/L)槲皮素的玻璃化冷冻组(VWQ1组...目的探讨在冷冻保护剂中添加槲皮素对羊卵巢组织冷冻复苏后卵泡活性的影响。方法将36只性成熟母羊的卵巢皮质组织块随机分配到新鲜对照组(CON组),玻璃化冷冻组(VIT组),添加低、中、高浓度(1、5、10μmol/L)槲皮素的玻璃化冷冻组(VWQ1组、VWQ2组、VWQ3组),将卵巢组织玻璃化冷冻后复苏、培养,测定各组雌激素水平,分析各组的卵泡计数及形态学改变。采用免疫组织化学法进行组织增殖细胞核抗原分析,采用TUNEL法进行组织凋亡分析,采用免疫组织化学法及可见光比色法进行组织抗氧化能力检测。结果 CON组原始卵泡形态正常比例最高、凋亡细胞计数最少(84.1%,P<0.001;13.92±3.88,P<0.001);VWQ1组的原始卵泡正常形态率高于VIT组(76.5%vs 71.7%,P=0.035),凋亡细胞数少于VIT组(50.96±24.28 vs 73.44±35.47,P=0.017),锰超氧化物歧化酶(SOD-2)蛋白表达量、过氧化氢酶(CAT)含量均高于VIT组[0.54(0.53,0.55)vs 0.32(0.29,0.51),P<0.001;5.60±1.49 vs 3.48±1.21,P=0.008];VWQ3组的原始卵泡正常形态率最低(61.2%,P<0.001),凋亡细胞计数最多(127.12±42.46,P<0.001)。结论低浓度槲皮素可以减轻卵巢组织氧化损伤进而保存卵泡活性,但过高浓度的槲皮素对卵泡活性有害。展开更多
Little mechanical data is available on human arteries because of the difficulty of testing artery samples often obtained from autopsy, while arteries are still considered “fresh”. Various solutions mimicking the phy...Little mechanical data is available on human arteries because of the difficulty of testing artery samples often obtained from autopsy, while arteries are still considered “fresh”. Various solutions mimicking the physiological environment have been used to preserve artery samples from harvesting to testing. Cryopreservation might provide a means to preserve the mechanical properties of arteries for days or weeks after harvesting. The objective of this study is to investigate the effect of several preservation methods, including simplified cryopreservation methods, on the passive mechanical properties of arteries. Eighteen fresh cruciform samples were mechanically tested. Samples were divided in three groups based on preservation medium and freezing method: isotonic saline solution, Krebs-Henseleit buffer solution with dimethyl sulfoxide (DMSO), and dipped in liquid nitrogen. In each group, half of the samples were stored at -20℃ and the other half at -80℃. Two months later, all the tissues were thawed at 4℃ and mechanical tests were repeated. Preservation of arteries for two months in Krebs solution with DMSO (at -20℃ or at -80℃) or in isotonic saline solution at -20℃ were the methods that least changed the mechanical properties of the arteries.展开更多
基金The work was supported by grants from the National Natural Science Foundation of China （No. 31230047, No. 31429004, No. 81571386, and No. 81471508）, the Interdisciplinary Project of Peking University Third Hospital and Chinese Academy of Sciences, Research Fund of National Health and Family Planning Commission of China （No. 201402004）, the Mega-projects of Science Research for the 12th Five-year Plan （No. 2012ba132b05）, and the Key Research Program of the Chinese Academy of Sciences （No. KJZD-EW-TZ-L03-2）.
文摘Background： L-proline is a natural, nontoxic cryoprotectant that helps cells and tissues to tolerate freezing in a variety of plants and animals. The use of L-proline in mammalian oocyte cryopreservation is rare. In this study, we explored the cryobiological characteristics of L-proline and evaluated its protective effect in mouse oocyte cryopreservation. Methods： The freezing property of L-proline was detected by Raman spectroscopy and osmometer. Mature oocytes obtained from 8-week-old B6D2F 1 mice were vitrified in a solution consisting various concentration of L-proline with a reduced proportion ofdimethyl sulfoxide （DMSO） and ethylene glycol （EG）, comparing with the control group （15% DMSO and 15% EG without L-proline）. The survival rate, 5-methylcytosine （5-mC） expression, fertilization rate, two-cell rate, and blastocyst rate in vitro were assessed by immunofluorescence and in vitro fertilization. Data were analyzed by Chi-square test. Results： L-proline can penetrate the oocyte membrane within 1 min. The osmotic pressure of 2.00 mol/L L-proline mixture is similar to that of the control group. The survival rate of the postthawed oocyte in 2.00 mol/L L-proline combining 7.5% DMSO and 10% EG is significantly higher than that of the control group. There is no difference of 5-mC expression between the L-proline combination groups and control. The fertilization rate, two-cell rate, and blastocyst rate in vitro from oocyte vitrified in 2.00 mol/L L-proline combining 7.5% DMSO and 10% EG solution are similar to that of control. Conclusions： It indicated that an appropriate concentration of L-proline can improve the cryopreservation efficiency of mouse oocytes with low concentrations of DMSO and EG, which may be applicable to human oocyte vitrification.
文摘为了观察冷冻稀释液中添加不同浓度的抗氧化剂谷胱甘肽(GSH)、绿茶多酚(GTPs)、维生素E(VE)和超氧化物歧化酶(SOD)对金华猪精液冷冻保存效果的影响,分别在冷冻稀释液中添加0.125、0.25、0.5 mmol.L-1的GSH,10、15、20 mmol.L-1的GTPs,1、2、4 mg.mL-1的VE和1 000、2 000、4 000 I U的SOD,采集活率较高的金华猪精液,用上述含抗氧化剂的冷冻稀释液稀释后进行冷冻保存,分别检测4个处理组精子4℃平衡后的活率和解冻后精子活率、质膜完整性、顶体完整率。结果表明,在金华猪精液的冷冻稀释液中分别添加15 mmol.L-1GTPs、2 mg.mL-1VE、4 000 I U SOD可显著提高猪精子4℃平衡后的活率、解冻后精子活率和质膜完整率(P<0.05)。
文摘目的探讨在冷冻保护剂中添加槲皮素对羊卵巢组织冷冻复苏后卵泡活性的影响。方法将36只性成熟母羊的卵巢皮质组织块随机分配到新鲜对照组(CON组),玻璃化冷冻组(VIT组),添加低、中、高浓度(1、5、10μmol/L)槲皮素的玻璃化冷冻组(VWQ1组、VWQ2组、VWQ3组),将卵巢组织玻璃化冷冻后复苏、培养,测定各组雌激素水平,分析各组的卵泡计数及形态学改变。采用免疫组织化学法进行组织增殖细胞核抗原分析,采用TUNEL法进行组织凋亡分析,采用免疫组织化学法及可见光比色法进行组织抗氧化能力检测。结果 CON组原始卵泡形态正常比例最高、凋亡细胞计数最少(84.1%,P<0.001;13.92±3.88,P<0.001);VWQ1组的原始卵泡正常形态率高于VIT组(76.5%vs 71.7%,P=0.035),凋亡细胞数少于VIT组(50.96±24.28 vs 73.44±35.47,P=0.017),锰超氧化物歧化酶(SOD-2)蛋白表达量、过氧化氢酶(CAT)含量均高于VIT组[0.54(0.53,0.55)vs 0.32(0.29,0.51),P<0.001;5.60±1.49 vs 3.48±1.21,P=0.008];VWQ3组的原始卵泡正常形态率最低(61.2%,P<0.001),凋亡细胞计数最多(127.12±42.46,P<0.001)。结论低浓度槲皮素可以减轻卵巢组织氧化损伤进而保存卵泡活性,但过高浓度的槲皮素对卵泡活性有害。
文摘Little mechanical data is available on human arteries because of the difficulty of testing artery samples often obtained from autopsy, while arteries are still considered “fresh”. Various solutions mimicking the physiological environment have been used to preserve artery samples from harvesting to testing. Cryopreservation might provide a means to preserve the mechanical properties of arteries for days or weeks after harvesting. The objective of this study is to investigate the effect of several preservation methods, including simplified cryopreservation methods, on the passive mechanical properties of arteries. Eighteen fresh cruciform samples were mechanically tested. Samples were divided in three groups based on preservation medium and freezing method: isotonic saline solution, Krebs-Henseleit buffer solution with dimethyl sulfoxide (DMSO), and dipped in liquid nitrogen. In each group, half of the samples were stored at -20℃ and the other half at -80℃. Two months later, all the tissues were thawed at 4℃ and mechanical tests were repeated. Preservation of arteries for two months in Krebs solution with DMSO (at -20℃ or at -80℃) or in isotonic saline solution at -20℃ were the methods that least changed the mechanical properties of the arteries.
