维生素D对精子活力的影响及其机制研究

发布时间：2018-04-08 20:46

本文选题：维生素D　切入点：精子活力　出处：《南方医科大学》2017年硕士论文

【摘要】：目的在世界范围内,男性不育占不孕不育总人数的40-50%,它影响大约7%的所有男性。在男性不育的众多原因中,维生素D被认为与男性的生育能力有直接关系。本研究旨在探索维生素D与男性精液质量的关系,为研究维生素D对男性生殖的具体作用机制提供更全面的研究方向。并在体外以维生素D的活性物质--1,25(OH)2D3处理精子,研究其对精子的作用机制。方法第一部分血清和精浆维生素D与精子参数的临床相关性分析根据纳入标准和排除标准收集我院生殖医学中心男科门诊就诊的男性患者以及正常体检者精液标本,按WHO《人类精液检查与处理实验室手册》(第五版)标准,应用WLJY-9000型伟力彩色精子质量检测系统检测精子浓度、精子总数、精子总活率、前向运动精子百分率以及相应运动学参数[曲线速度(VCL)、直线速度(VSL)、平均路径速度(VAP)];吖啶橙染色法检测精子DNA碎片指数(DFI)和高DNA可染性(HDS);应用改良巴氏染色法以WHO标准严格判读精子形态,评估正常形态精子百分率;应用电化学发光法检测血清和精浆维生素D水平。分析血清和精浆维生素D与精液各参数之间的相关性。第二部分1,25(OH)2D3对精子运动能力的影响及其作用机制根据纳入标准和排除标准收集我院生殖医学中心男科门诊就诊的男性患者以及正常体检者的精液标本,用percoll液密度梯度离心法提取纳入研究人群的精子,以不同浓度(0、0.01nM、0.1nM、1nM、10nM)的 1,25(OH)2D3处理精子,在不同时间点(0、15min、30min、60min、90min)用WLJY-9000型伟力彩色精子质量检测系统检测精子各运动参数;根据前期结果,选择最适浓度的1,25(OH)2D3处理精子30min后,检测精子的爬高能力、cAMP的含量以及精子PKA的活性、NADH和ATP的含量,分析各指标的变化。结果1.临床试验中,血清维生素D的水平与精子各参数之间无相关性(p0.05);精浆维生素D与精子总活率(r = 0.308,p = 0.026)、前向运动精子百分率(r= 0.299,p = 0.031)、曲线速度(r = 0.386,p = 0.005)、直线速度(r = 0.348,p = 0.011)、路径速度(r = 0.383,p = 0.005)之间呈显著正相关。2.1,25(OH)2D3处理精子后,在作用30分钟且浓度为0.1nM时,精子的各项运动参数增加显著(p0.001),精子的爬高高度显著增加(p0.01),精子NADH、ATP和cAMP含量显著增加(P0.05),精子PKA活性显著增强(p0.05)。结论1.精浆维生素D可能参与精子运动能力的调节。2.活性形式的维生素D可能通过cAMP/PKA和NADH途径促进精子线粒体电子呼吸链的作用,增加ATP的产量,从而增强精子的活力。
[Abstract]:Objective in the world, male infertility accounts for 40-50% of the total number of infertile women, which affects about 7% of all men.Among the many causes of male infertility, vitamin D is thought to be directly related to men's fertility.The purpose of this study was to explore the relationship between vitamin D and male semen quality, and to provide a more comprehensive research direction for the study of the specific mechanism of vitamin D on male reproduction.The spermatozoa were treated with the active substance of vitamin D in vitro, and the mechanism of its action on spermatozoa was studied.Methods the clinical correlation between serum and seminal plasma vitamin D and sperm parameters was analyzed. According to the inclusion and exclusion criteria, semen samples were collected from male patients in andrology clinic and normal persons in our center of reproductive medicine.According to the WHO Manual for the Laboratory of Human semen examination and processing (fifth edition), the sperm concentration, total sperm count and total sperm motility were detected by using the WLJY-9000 type Weili color sperm quality detection system.Forward motility sperm percentage and its corresponding kinematic parameters [curve velocities, linear velocities, VSLs, average path velocities]; acridine orange staining for the detection of sperm DNA fragment index (DNA) and high DNA dyeability (WHO); using modified pasteurian staining method to detect WHOThe standard strictly interprets sperm morphology,The percentage of normal spermatozoa was evaluated and the levels of vitamin D in serum and seminal plasma were measured by electrochemiluminescence (ECL).The correlation between serum and seminal plasma vitamin D and semen parameters was analyzed.The second part: the effect of OHH 2D3 on sperm motility and its mechanism of action. According to the inclusion and exclusion criteria, semen samples were collected from male patients from andrology outpatient clinics of our Reproductive Medicine Center, as well as normal physical examination subjects.The content of camp in sperm and the content of nadh and ATP in sperm PKA were determined after the treatment of 30min with the optimum concentration of 1o 25 OHH 2D3, and the changes of each index were analyzed.Result 1.In clinical trials,There was a significant positive correlation between path velocities (r = 0.383, p = 0.005).At the concentration of 0.1nM for 30 minutes, the motility parameters of spermatozoa increased significantly (p0.001), the climbing height of spermatozoa increased significantly (p0.01), the contents of NADH-ATP and cAMP increased significantly, and the activity of PKA increased significantly (p0.05).Conclusion 1.Seminal plasma vitamin D may be involved in the regulation of sperm motility.The active form of vitamin D may promote the action of mitochondrial electron respiratory chain through cAMP/PKA and NADH pathway, and increase the production of ATP, thus enhancing sperm motility.
【学位授予单位】：南方医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R698.2

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