精子DNA损伤与保护

精子DNA损伤是引起不育的重要原因,携有DNA损伤的幸存精子可能逃避体内精子选择机制而成熟并将遗传缺陷传递于后代。因此对精子DNA损伤的研究已成为生殖医学的热点之一。损伤精子DNA的因素主要包括氧化应激、微量元素、精子毒性物质、放射线等,而机体则凭借高度压缩精子DNA、抗氧化系统等机制保护精子DNA的完整性。此外,一些药物如抗氧化剂、黑茶提取物等也可以促使这种保护机制的健全与重建。     

冷冻损伤精子受精后DNA损伤修复机制的研究进展

精子冻存损伤的修复是一个热门话题,在辅助生殖技术(assisted reproductive technology,ART)中起着重要的临床指导意义.     

移动电话电离辐射体外诱导人精子产生活性氧物质损伤精子DNA

近年来有很多关于电离辐射影响人类健康的争论。自从有研究认为使用移动电话产生的电离辐射与精液质量存在相关性后，移动电话电离辐射对人类生殖功能的影响便成了主要的争论点之一。其作用机理至今不清楚，然而人类精子容易被氧化应激产生的氧自由基攻击和自身缺乏抗氧化物酶而受到损伤，氧化应激诱导不仅影响精子受精能力，而且还使精子DNA损伤，导致低生育能力、增加流产率以及后代的疾病发生率包括小儿肿瘤等。     

精子DNA损伤与男性不育的关系

近年来,不育症日益受到社会的广泛关注.发病率约为15％,其中男性因素约占40％[1].在临床实践中,男性不育的评估主要依靠精液分析.然而常规的精液分析(精子密度,精子活动力,精子形态)往往不能全面评估男性的生育力,即部分精液分析正常的人仍无生育能力,而某些精液分析异常的人却能够生育.这无疑给不育症的诊断和治疗带来了困难.有研究表明,男性不育可能与精子DNA的损伤紧密相关[2],不育男性的精子DNA碎片率显著高于正常生育男性的精子DNA碎片率[3].精子DNA损伤形式包括精子DNA碎片、异常的染色质包装和鱼精蛋白缺乏,其中最主要的损伤形式是精子DNA碎片,包括精子DNA双链或单链的断裂.精子DNA的完整性是父系遗传信息传递给子代的前提.精子DNA损伤对受精、胚胎的形态和植入有负影响.如果精子的遗传物质发生严重的损伤,胚胎的发育就会停滞,并导致流产的发生.这里面似乎存在一个阈值,如果精子DNA损伤超过了这个阈值,胚胎发育和妊娠就会受到损害[4].因而,评估精子DNA损伤程度有望成为诊断男性不育的一个指标,并对辅助生殖技术(artificial reproductive technology,ART)有指导意义.目前,已有许多方法用来检测精子DNA损伤,精子DNA损伤的评估在临床上的应用也日益受到重视.本综述的目的就是介绍在这个领域的新进展.     

精子DNA损伤与辅助生殖技术

精子DNA损伤与辅助生殖技术(ART)结局密切相关,精子DNA损伤影响到ART中的受精比率、胚细胞发育、妊娠率和子代的健康。同时,精子低温冷冻保存和诸多体外处理技术均能影响精子DNA的完整性。检测精子DNA断裂指标(DFI),采取适当措施获得DNA无损伤或损伤较小的精子,对改善ART结局十分必要。现就精子DNA损伤与ART相关性的研究做一综述。     

精子DNA损伤与辅助生殖技术

随着辅助生殖技术的广泛开展,精子评估已由传统的精液常规分析向细胞、分子水平深入发展。精子DNA损伤是反映男性生育力的一个新指标。精子DNA损伤的发生机制包括精子染色质包装与分离异常、氧化应激、细胞凋亡异常等。精子染色质结构分析是目前检测精子DNA损伤最常用的方法之一。精子DNA损伤可能与辅助生殖技术治疗结局、复发性自然流产、增加ICSI后代遗传风险相关。采取口服抗氧化药物、取睾丸精子行ICSI、预冻存精子、去除病因以及中医中药等治疗对策可能会降低精子DNA损伤程度,进而提高辅助生殖技术成功率。本文主要就精子DNA损伤的机制与检测方法、DNA损伤与生殖结局以及辅助生殖技术中与DNA损伤相关的治疗对策作一综述。     

精子DNA损伤在男性不育症中的研究进展

男性不育症是现今困扰我国不孕不育夫妇的一个严重问题。目前临床上对于男性不育的筛查主要取决于精液常规的检查。但由于精液本身的易干扰性,导致其已不能对精子的受精能力及男性的生育能力进行准确的评价。近年来,精子DNA损伤作为一项新的评价精子功能的指标,逐渐成为男性不育症的研究热点。本文就精子DNA损伤的原因、机制及在男性不育症中的研究进展作一综述,为男性不育症的诊断和治疗研究提供依据。     

畸形精子症患者中氧化应激与精子DNA损伤的关系探讨

目的:探讨畸形精子症患者中氧化应激相关指标与精子DNA损伤变化的相关性,以期为男性不育诊治提供理论依据。方法:选取2019年1月至2019年8月在河北医科大学第四医院生殖医学科诊治的101例男性不育症患者作为研究对象,按精子形态将患者精液标本分为两组,A组为精子正常形态≥4,B组为精子正常形态<4,即畸形精子症组,按世...     

不育患者精子DNA链损伤类型的研究及临床意义

目的:观察精子DNA单、双链损伤(SSB、DSB)在男性不育中的特征,探讨DSB与男性不育的关系,为男性不育的诊疗提供新的观察指标与思路。方法:选择男性不育患者60例以及同期因女方因素不孕前来检查的生育力评估正常的健康男性30例作为对照组,分析两组精子DNA损伤及精液主要参数的差异。精子浓度及活力采用计算机辅助精子分析系统,精子存活率分析采用低渗膨胀试验,精子形态采用Diff-Quik染色法,精子DNA损伤采用双尾彗星实验。结果:双尾彗星实验检测精子DNA完整性共有9种彗星模型。不育患者精子DNA损伤指数(DFI)为(33.8±13.1)%,单链损伤指数(SSB-DFI)为(19.2±11.4)%、SSB占所有损伤精子的比率(SSB-DFI/DFI)为(56.8±32.4)%,双链损伤指数(DSB-DFI)为(23.9±13.4)%、DSB占所有损伤精子的比率(DSB-DFI/DFI)为(70.8±19.5)%;与对照组比较[分别为(16.3±7.9)%、(14.9±7.6)%、(91.4±27.8)%、(6.1±2.7)%、(37.4±11.3)%)],SSB-DFI无显著性差异(P0.05),DSB-DFI、DFI和DSB-DFI/DFI显著高于对照组(P均0.01),SSB-DFI/DFI显著降低于对照组(P0.01)。绘制ROC曲线,DSB-DFI/DFI、DSB-DFI及DFI诊断男性不育最佳截断值分别为39.5%、15.85%和18.65%,ROCAUC、敏感度和特异性分别为(0.969,98.3%,90.0%)、(0.912,86.7%,80.0%)、(0.861,90.0%,70.0%)。不育组精子SSB-DFI及SSB-DFI/DFI与精液常规参数均无相关关系(P均0.05),DFI与前向运动精子百分率、精子存活率及正常形态精子百分率呈负相关(P0.05或P0.01),与精子浓度无相关关系(P0.05),精子DSB-DFI及DSB-DFI/DFI与精子浓度、精子存活率、前向运动精子百分率及正常形态精子百分率均呈负相关(P0.05或P0.01)。结论:影响男性不育的DAN损伤因素可能是DSB,而与SSB相关性不大,精子DNA链的损伤类型对男性生殖能力的评估有较大的参考价值。     

单细胞凝胶电泳检测人精子DNA损伤

目的 :利用单细胞凝胶电泳 (SCGE)对精子DNA损伤进行检测 ,了解其在评估男性生殖中的意义。方法 :4 18名男性的精子经计算机辅助分析系统常规检测后进行SCGE ,根据精子细胞核DNA损伤程度分为 5级。结果 :①当精子密度 <2 0× 10 6/ml时 ,SCGE的发生特别是Ⅱ、Ⅲ 级升高 ,但是与其他相比无统计学意义 (P均 >0 .0 5 ) ;②d级精子中 ,Ⅰ、Ⅱ、Ⅲ 级慧星发生升高 ,与a级精子相比差异有显著性 (P <0 .0 5 )。　结论 :SCGE可评价精子的质量和损伤 ,是男性生殖评估的又一较为可靠的检测手段     

氧化应激检测在男性不育临床中的应用

活性氧 (ROS)在低浓度发生可以调节正常精子功能 ,而高浓度ROS危害精子的存活力和功能。ROS产生过量与抗氧化系统损伤会发生氧化应激 (OS)。目前认为损伤OS促进了一系列男性生殖功能的病理学改变。ROS介导的精子质膜过氧化损伤 ,可以解释在高比例的不育病人中观察到的精子功能缺陷。ROS产生过多也会破坏精子核内DNA的完整性。DNA碱基对OS敏感 ,这些结构的过氧化能引起碱基修饰、DNA链断裂以及染色质交联。由过量ROS诱发的DNA损伤会加速生殖细胞凋亡过程 ,导致与男性不育相关的精子计数降低 ,以及可以解释近半个世纪以来所观察到的精液质量明显下降现象。近 10年来 ,克里夫兰临床基金会的研究组已经查明了男性不育中OS的关键作用 ;研究的主要目的是将这些重要的知识从实验室转向临床应用。我们设计的研究目标 :①理解精液中OS发生的精确机制 ;②建立准确评估OS状态的实验 ,并进行质控研究 ;③检测OS与精子核DNA损伤之间的相关性 ;④鉴定精液OS评估对男性不育的临床意义     

Time Course of Oxidative Stress During Open-Heart Surgery

Oxidative stress and subsequent lipid peroxidation have been suggested as pathogenetically important for postischaemic reperfusion injury. We studied the time-course of oxidative stress in 14 adults undergoing cardiac surgery, evaluating serum levels of lipid peroxidation products—diene conjugates (DC) and basal and Fe-stimulated thio-barbituric acid reactive substances (TBARS, Fe-TEBARS)—as well as markers of blood antioxidant status—serum antioxidative capacity (AOC) and red blood cell glutathione (RBC-GSH) at 6 perioperative time-points. Arterial TBARS were significantly increased 15 minutes after start of cardiopulmonary bypass, 5 minutes after release of aortic cross-clamp and 15 minutes after cessation of bypass, compared with the preoperative levels (respective means 20.8, 38.5, 34.8 vs 7.5 nmol/g protein, p < 0.05). AOC had decreased at these times (means 21.3, 18.1, 23.2 vs 34.9%, p < 0.05). The TBARS changes correlated with AOC decrease (r = 0.30, p < 0.001). Changes in serum DC and RBC-GSH were not statistically significant. All lipid peroxidation parameters had returned to preoperative levels on the following morning, while antioxidative capacity remained suppressed (28.1%, p < 0.05). These data demonstrate a definite time-course of oxidative stress markers in arterial blood during open-heart surgery.     

精子氧化应激损伤影响因素研究进展

不育症发病率呈全球性增高趋势。据统计,目前不育夫妇占已婚夫妇15%,其中男性不育占50%。这与生活节奏的加快、环境污染和人们不良行为,造成精子数量和质量降低,生殖功能下降等有关。随着自由基理论的建立和发展,人们发现活性氧介导的氧化应激对精子的损伤作用,可能是男性不育的原因之一。本文对精子产生氧化应激损伤的影响因素进行综述。     

表没食子儿茶素没食子酸酯对高糖环境下体外小鼠足细胞活性氧的影响

目的　观察不同浓度表没食子儿茶素没食子酸酯（EGCG）对高糖造成氧化应激状态下体外小鼠足细胞损害的作用并探讨其机制。 方法　以高糖（25 mmol/L）培养的小鼠足细胞为研究对象,维生素E培养为对照。首先以MTT法检测细胞活力,再在激光共聚焦显微镜下以CM-H2DCFDA荧光探针观察不同浓度EGCG（0.2、10、100 μmol/L）刺激足细胞6、12、24 h后活性氧（ROS）生成,并以流式细胞仪定量分析ROS平均荧光强度。RT-PCR法检测足细胞内ROS产生的主要通路NADPH氧化酶各亚基mRNA（ph22phox、p47phox、p67phox）的表达。 结果　高糖刺激下6 h,足细胞ROS生成显著增加（P < 0.01）。正常糖组和甘露醇组培养12 h ROS生成无显著增加（P > 0.05）。EGCG 0.2 μmol/L作用6 h可显著降低高糖环境下体外小鼠足细胞ROS水平（P < 0.01）。与高糖组比较,EGCG（100 μmol/L）显著减少NADPH氧化酶亚基p22phox和p67phox mRNA表达（均P < 0.05）。与维生素Ｅ组比较,EGCG（0.2 μmol/L）和维生素E（0.2 mmol/L）协同作用组显著减少p47phox mRNA表达（P < 0.05）。 结论　EGCG能缓解高糖环境下体外足细胞氧化应激状态,对高糖培养下足细胞有保护作用。     

Hyperoxaluria-induced oxidative stress and antioxidants for renal protection

Renal cellular exposure to oxalate (Ox) and/or CaOx crystals leads to the production of reactive oxygen species (ROS), development of oxidative stress followed by injury and inflammation. Renal injury and inflammation appear to play a significant role in stone formation. ROS are produced from many sources and involve a variety of signaling pathways. Tissue culture and animal model studies show that treatments with anti-oxidants and free radical scavengers reduce Ox/CaOx crystal induced injuries. In addition, CaOx crystal deposition in kidneys is significantly reduced by treatments with antioxidants and free radical scavengers, indicating their efficacy. These results point towards a great potential for the therapeutic application of antioxidants and free radical scavengers to reduce stone recurrence particularly after shock wave lithotripsy, which is itself known to generate ROS and cause renal damage.     

URINARY THYMIDINE GLYCOL AS A BIOMARKER FOR OXIDATIVE STRESS AFTER KIDNEY TRANSPLANTATION

Reactive oxygen species are generated during ischemia-reperfusion tissue injury. Oxidation of thymidine by hydroxyl radicals (HO^˙) causes formation of 5,6-dihydroxy-5,6-dihydrothymidine (thymidine glycol). Thymidine glycol excreted in urine can be used as a biomarker of oxidative DNA damage. The aim of this study was to investigate the oxidative DNA damage in patients showing immediate allograft function after kidney transplantation, and to find out whether this damage correlates with glomerular and tubular lesions.

Time dependent changes in urinary excretion rates of thymidine glycol, but also of total protein, albumin, low molecular weight (α₁-microglobulin, β₂-microglobulin) and high molecular weight proteins (transferrin, IgG, α₂-macroglobulin) were analyzed quantitatively and by polyacrylamide-gel electrophoresis in six patients. Urinary thymidine glycol was determined by a fluorimetric assay in combination with affinity chromatography and HPLC.

After kidney transplantation the urinary excretion rate of thymidine glycol increased gradually reaching a maximum within the first 48 hours (16.56 ± 11.3 nmol/mmol creatinine, ref. 4.3 ± 0.97). Severe proteinuria with an excretion rate of up to 7.2 g/mmol creatinine was observed and declined within the first 24 hours of allograft function (0.35 ± 0.26 g/mmol creatinine). The gel-electrophoretic pattern showed a nonselective glomerular and tubular proteinuria. The initial nonselective glomerular proteinuria disappeared within 48 hours, changing to a mild selective glomerular proteinuria. In this period (12–48 hours) higher levels of thymidine glycol excretion were observed, when compared to the initial posttransplant phase (13.66 ± 9.76 vs. 4.31 ± 3.61 nmol/mmol creatinine; p < 0.05).

An increased excretion of thymidine glycol is seen after kidney transplantation and is explained by the ischemia-reperfusion induced oxidative DNA damage in the kidney. In the second phase higher levels of excretion were observed parallel to the change from a nonselective to a selective glomerular and tubular proteinuria. An explanation may be sought in the repair process of DNA in the glomerular and tubular epithelial cells, appearing simultaneously with functional recovery.     

Oxidative stress and anti-oxidative mobilization in burn injury

A severe burn is associated with release of inflammatory mediators which ultimately cause local and distant pathophysiological effects. Mediators including Reactive Oxygen Species (ROS) and Reactive Nitrogen Species (RNS) are increased in affected tissue, which are implicated in pathophysiological events observed in burn patients. The purpose of this article is to understand the role of oxidative stress in burns, in order to develop therapeutic strategies. All peer-reviewed, original and review articles published in the English language literature relevant to the topic of oxidative stress in burns in animals and human subjects were selected for this review and the possible roles of ROS and RNS in the pathophysiology of burns are discussed. Both increased xanthine oxidase and neutrophil activation appear to be the oxidant sources in burns. Free radicals have been found to have beneficial effects on antimicrobial action and wound healing. However following a burn, there is an enormous production of ROS which is harmful and implicated in inflammation, systemic inflammatory response syndrome, immunosuppression, infection and sepsis, tissue damage and multiple organ failure. Thus clinical response to burn is dependent on the balance between production of free radicals and its detoxification. Supplementation of antioxidants in human and animal models has proven benefit in decreasing distant organ failure suggesting a cause and effect relationship. We conclude that oxidative damage is one of the mechanisms responsible for the local and distant pathophysiological events observed after burn, and therefore anti-oxidant therapy might be beneficial in minimizing injury in burned patients.