左卡尼汀在特发性弱精子症患者中的水平测定

Objective To study the relationship between sperm activity and the level of L-carnitine in seminal plasma of patients with idiopathic asthenospermia by HPLC.Methods Experimental group:idiopathic asthenospermia 50 cases,and the control group:male with pregnancy history 10 cases.The semen parameters (volume,sperm density,viability,motility),and L-carnitine level in seminal plasma were detected with high performance liquid chromatography method.Results There were significant difference between A and B group about sperm density and the percentage of progressive motile spermatozoa,and the level of L-carnitine(P＜0.01).Conclusions The decrease of L-carnitine level in patients with idiopathic asthenospermia,is closely related to the low sperm motility,which shows that L-carnitine can be used to treat idiopathic asthenospermia effectively.     

血清睾酮雌二醇比值变化对精子缺陷及生育力的影响

目的 了解血清睾酮(T)/雌二醇(E2)比值变化对精子缺陷程度及男性生育能力的影响.方法对90例已婚育龄男性进行血清T、E2、卵泡刺激素(FSH)、黄体生成素(LH)、催乳素和精液参数、精子形态测定,计算血清T/E2和精子多重缺陷指数.根据血清T/E2分为≤10(1组)、＞10(2组)及＞20(3组)分别比较.结果 1、2、3组精子密度和精子活动力分别为(66.8±39.6)×106、(57.6±33.9)×106、(77.4±26.0)×106/ml和(46.6±16.4)%、(44.9±16.0)%、(53.9±14.4)%,3组间差异无统计学意义(P均＞0.05).1组头部缺陷精子为(81.4±9.4)%,明显高于3组的(74.0±10.7)%(t=2.482,P=0.016);1、2组颈部缺陷精子为(16.9±8.6)%和(12.7±9.8)%,均明显高于3组的(7.1±3.9)%(t=4.113、2.050,P=0.000、0.046);1、2、3组尾部缺陷精子分别为(22.6±14.6)%、(14.6±9.0)%和(6.5±5.7)%,组间差异有统计学意义(t=2.722、3.996、3.110,P=0.008、0.000、0.003).2组精子畸形指数(SDI)(1.4±0.3)明显高于1组(1.3±0.2,t=2.293,P=0.025);2、3组畸形精子指数(TZI)分别为1.6±0.3和1.6±0.2,明显高于1组的1.4±0.2(t=2.285、2.727,P=0.025、0.009).1、2、3组中配偶有生育或妊娠经历者分别为13、16、6例,组间差异无统计学意义(x2=3.285、0.854、0.199,P=0.070、0.355、0.655),但2、3组相对危险度分别为1组的2.4倍和1.8倍.1、2、3组中特发性不育病例分别为25、15、7例,组间差异均无统计学意义(x2=0.735、0.200、0.038,P=0.391、0.655、0.845),1组相对危险度分别为2、3组的1.5倍和1.3倍.相关性分析结果发现,血清T/E2与头、颈和尾部缺陷精子比率呈显著负相关(r=-0.209、-0.316、-0.335,P=0.048、0.002、0.001).结论血清T/E2与精子头、颈及尾部缺陷呈显著负相关,其下降与男性生育概率降低有关,但并不影响精子密度及活动力,表明男性体内维持较高比例的T/E2水平对于精子发生过程中的精子形成阶段及受精能力有重要意义.

Abstract：

Objective To study the effect of the serum testosterone/estradiol ratio (T/E2) alteration on sperm defect and fertility. Methods The testosterone, estradiol, FSH, LH, PRL,sperm parameters and sperm morphology of 90 men were analyzed and the T/E2 and multiple anomalies index (MAI) were calculated. The patients were divided into 3 groups; T/E2≤10 (Ⅰ), T/E2＞10 (Ⅱ), and T/E2＞20 (Ⅲ). Results The sperm concentration and motility among the 3 groups were not significantly different (P＞0. 05). The percentages of the sperm whose head, neck and tail were abnormal declined gradually with the increase of the T/E2 in serum. The percentage of sperm head defeet of group Ⅰ was significantly higher than that of group Ⅲ (t=2. 482, P=0. 016) and that of sperm neck defect of groups Ⅰ and Ⅱ were significantly higher than that of group Ⅲ (t=4.113, 2. 050, P=0.000, 0. 046, respectively). The percentage of sperm tail defect among 3 groups was significantly different (t=2. 722, 3. 996, 3. 110, P=0. 008, 0. 000, 0. 003, respectively). The SDI of group Ⅱ was significantly higher than that of group Ⅰ (t= -2. 293, P= 0. 025). But the TZI increased gradually with the increase of the serum T/E2 and the TZI of groups Ⅱ and Ⅲ were significantly higher than that of group Ⅰ (t=2. 285, 2. 727, P=0. 025, 0. 009, respectively). The percentage of the men in group Ⅰ whose partners became pregnant was 29. 5% and those of groups Ⅱ and Ⅲ were 50% and 42.9%, respectively. Although the percentage among three groups was not different statistically (x2 = 3. 285, 0. 854, 0. 199, P= 0. 070, 0. 355, 0. 655, respectively), the relative risks of groups Ⅱ and Ⅲ were 2.4 and 1.8 times of that of group Ⅰ. There were 25, 15, 7 cases of idiopathic infertility among the 3 groups, respectively. The relative risk of I group was 1.5 and 1.3 times of that of Ⅱ and Ⅲ groups. The correlation analysis showed that the T/E2 in serum had significantly negative correlation with the percent of the sperm head or neck or tail defects (r= -0. 209, -0. 316 and -0. 335,respectively and P= 0. 048, 0. 002 and 0. 001, respectively). Conclusions The decrease of T/E2 in serum was correlative with the decrease of fertility probability, but it did not alter the sperm density and the sperm motility. It showed that the level of the T/E2 in serum was important for spermatogenesis and sperm fertilizing capability.     

Low dose gossypol for male contraception

Aim: To ascertain whether the side effects of gossypol, hypokalemia and irreversibility, could be avoided on dose re-duction. Methods: Seventy-seven male volunteers were divided into 3 groups: control (22 cases), 10 mg gossypol(29 cases) and 12.5 mg (26 cases). Serum levels of testosterone, FSH and LH were measured by RIA and potassiumby flame photometry. Spema counts and motility were examined before and regularly after treatment for the evaluationof contraceptive efficacy. Results: The average sperm density and motility started to decrease significantly by theend of month 2 of medication and gradually reached the infertility levels ( < 4 million /mL) in both treated groups. Af-ter that the 10 mg group was asked to take the same dose every other day for up to a total observation period of 16-18months for the maintenance of infertility. Subjects in the 12.5 mg group did not take gossypol any more so as to ob-serve the length of the loading dose required, but in a few, a maintenance dose of 12.5 mg every     

异柠檬酸脱氢酶基因突变在神经胶质瘤发生发展中的作用

Objective To explore the possible role of isocitrate dehydrogenase (IDH) gene mutation in the genesis of glioma. Methods DNA was extracted from 89 glioma FFPE specimens. The primers were designed and synthetized. The exon 4 of IDH1 and IDH2 gene was screened and analyzed through polymerase chain reaction (PCR) and direct DNA sequencing. Results Altogether 36 IDH1 gene mutations were found (40. 4% ) , mainly from WHO Ⅱ and Ⅲ grade astrocytoma (61%-63% ) , oligodondroglioma (33%-66%) and oligoastrocytoma (57% -80%). All mutations were heterozygous, missense and point mutation, which were only found in codon 132 of IDH1. The patients with gene mutation were younger than those without muatation. Conclusion IDH1 gene mutation may play an important role in the tumorigenesis of glioma.     

异柠檬酸脱氢酶基因突变在神经胶质瘤发生发展中的作用

Objective To explore the possible role of isocitrate dehydrogenase (IDH) gene mutation in the genesis of glioma. Methods DNA was extracted from 89 glioma FFPE specimens. The primers were designed and synthetized. The exon 4 of IDH1 and IDH2 gene was screened and analyzed through polymerase chain reaction (PCR) and direct DNA sequencing. Results Altogether 36 IDH1 gene mutations were found (40. 4% ) , mainly from WHO Ⅱ and Ⅲ grade astrocytoma (61%-63% ) , oligodondroglioma (33%-66%) and oligoastrocytoma (57% -80%). All mutations were heterozygous, missense and point mutation, which were only found in codon 132 of IDH1. The patients with gene mutation were younger than those without muatation. Conclusion IDH1 gene mutation may play an important role in the tumorigenesis of glioma.     

Sperm DNA damage in men from infertile couples

Aim： To investigate the prevalence of high levels of sperm DNA damage among men from infertile couples with both normal and abnormal standard semen parameters. Methods： A total of 350 men from infertile couples were assessed. Standard semen analysis and sperm chromatin structure assay （SCSA） were carried out. Results： Ninety-seven men （28% of the whole study group） had a DNA fragmentation index （DFI） 〉 20%, and 43 men （12%） had a DFI 〉 30%. In the group of men with abnormal semen parameters （n = 224）, 35% had a DFI 〉 20%, and 16% had a DFI 〉 30%, whereas these numbers were 15% and 5%, respectively, in the group of men with normal semen parameters （n = 126）. Men with low sperm motility and abnormal morphology had significantly higher odds ratios （ORs） for having a DFI 〉 20% （4.0 for motility and 1.9 for morphology） and DFI 〉 30% （6.2 for motility and 2.8 for morphology） compared with men with normal sperm motility and morphology. Conclusion： In almost one-third of unselected men from infertile couples, the DFI exceeded the level of 20% above which, according to previous studies, the in vivo fertility is reduced. A significant proportion of men with otherwise normal semen parameters also had high sperm DNA damage levels. Thus, the SCSA test could add to explaining causes of infertility in cases where semen analysis has not shown any deviation from the norm. We also recommend running the SCSA test to choose the appropriate assisted reproductive technique （ART）.     

异柠檬酸脱氢酶基因突变在神经胶质瘤发生发展中的作用

目的 观察异柠檬酸脱氢酶(IDH)基因突变在神经胶质瘤发生、发展中的作用.方法 提取89例胶质瘤蜡块标本DNA,设计合成引物,聚合酶链反应(PCR)扩增覆盖IDH1、IDH2基因4号外显子cDNA片段,正反双向DNA测序,观察基因突变频率和类型,分析基因型与临床表型的关系.结果 89例胶质瘤标本中有36例IDH1突变(40.4%);主要见于Ⅱ、Ⅲ级星形细胞瘤(61%～63%)、少突胶质细胞瘤(33%～66%)以及混合性胶质瘤(57%～80%);全部为4号外显子R132h杂合性、错义、点突变;各型胶质瘤组中突变型患者多较年轻.结论 高频率IDH基因特定催化活性区域碱基突变是胶质瘤发生、发展过程中一种重要的分子事件.

Abstract：

Objective To explore the possible role of isocitrate dehydrogenase (IDH) gene mutation in the genesis of glioma. Methods DNA was extracted from 89 glioma FFPE specimens. The primers were designed and synthetized. The exon 4 of IDH1 and IDH2 gene was screened and analyzed through polymerase chain reaction (PCR) and direct DNA sequencing. Results Altogether 36 IDH1 gene mutations were found (40. 4% ) , mainly from WHO Ⅱ and Ⅲ grade astrocytoma (61%-63% ) , oligodondroglioma (33%-66%) and oligoastrocytoma (57% -80%). All mutations were heterozygous, missense and point mutation, which were only found in codon 132 of IDH1. The patients with gene mutation were younger than those without muatation. Conclusion IDH1 gene mutation may play an important role in the tumorigenesis of glioma.     

邻苯二甲酸二酯与特发性弱精子症的关系

目的 检测人精浆中邻苯二甲酸二酯(DEHP)的浓度来评价DEHP对特发性弱精子患者精液质量的影响.方法 选择特发性弱精子患者50例,非特发性弱精子患者50例,精液正常者50例,分别用非特发性弱精子患者组及精液正常者组作为对照组.运用反相液相高效色谱法测定血浆及精浆中DEHP浓度.并运用SPSS分析软件测定结果进行了分析.结果 特发性弱精子患者组中DEHP均可检出,含量范围在精液中为ND0.10-1.5 mg/L;非特发性弱精子患者组及精液正常组DEHP分别检出15例及13例,精液中为ND0～0.96 mg/L和ND0～0.95mg/L;特发性弱精子组与非特发性弱精子组和正常组精浆中DEHP的浓度有差异(P＜0.05),非特发性弱精子症患者组与正常组精浆中DEHP浓度无差异(p＞0.05),精浆中DEHP浓度与精子活力负相关(p＜0.05).结论 精浆中DEHP可以影响精子质量.

Abstract：

Objectives To detect the level of di-2 - ethylhexyl phthalate(DEHP) in serum and semen to evaluate effect of DEHP to idiopathic asthenaspermia. Methods 150 people were assigned into idiopathic asthenaspermia group( n = 50), no idiopathic asthenaspermia group ( n = 50 ) and nomal group ( n = 50 ). The concentrations of DEHP in the human biological samples were measured by using reversed-phase HPLC. Results The DEHP were detected in most of the idiopathic asthenaspermia group 0. 10 ～ 1. 5 mg / L in semen specimens, 15 of no idiopathic asthenaspermia group 0 ～0.96 mg/L in semen specimens, 13 of nomal group 0 ～0.95mg/L in semen specimens. There was difference between semen concentrations of DEHP and sperm molity ( p ＜ 0. 05 ). Conclusions These results suggest that DEHP affect semen quanlity,specialy DEHP in semen specimens.     

Effect of methoxychlor on antioxidant system of goat epididymal sperm in vitro

Aim: To evaluate the effect of methoxychlor on the antioxidant system of goat epididymal sperm. Methods: Epididymis of adult goat was obtained from local slaughter houses and sperm were collected by chopping the epididymis in modified Ringer‘s phosphate solution (RPS). After several washings, the sperm samples were dispersed in RPS and incubated with methoxychlor (1μmol/L, 10 μmol/L and 100 μmol/L) and methoxychlor vitamin C (100 μmol/L each) for 3 h at 32℃. After incubation, the sperm motility and viability were assessed. An aliquot of sperm sample was homogenized, centrifuged and used for the assay of superoxide dismutase, glutathione peroxidase, glutathione reductase and lipid peroxidation. Results: In methoxychlor-mcubated sperm and in sperm co-incubated with methoxychlor and vitamin C, the sperm motility and viability showed no significant changes as compared to the corresponding controls. In methoxychlor-incubated sperm the activity of superoxide dismutase, glutathione reductase and glutathione peroxidase were decreased while lipid peroxidation was increased in a dose-dependent manner. Co-incubation of sperm with methoxychlor and vitamin C showed no changes in the activity of superoxide dismutase, glutathione reductase and glutathione peroxidase and in the level of lipid peroxidation. Conclusion: Methoxychlor induced oxidative stress in epididymal sperm of goats by decreasing the levels of antioxidant enzymes. Co-incubation of sperm with methoxychlor and vitamin C, a natural antioxidant, reversed the effect of methoxychlor.     

Supplemental effect of varying L-cysteine concentrations on the quality of cryopreserved boar semen

Cryopreservation is associated with the production of reactive oxygen species, which leads to lipid peroxidation of the sperm membrane and consequently a reduction in sperm motility and decreased fertility potential. The aim of this study was to determine the optimal concentration of L-cysteine needed for cryopreservation of boar semen. Twelve boars provided semen of proven motility and morphology for this study. The semen was divided into four portions in which the lactose-egg yolk （LEY） extender used to resuspend the centrifuged sperm pellet was supplemented with various concentrations of L-cysteine to reach 0 mmol L^-1 （group Ⅰ, control）, 5 mmol L^-1 （group Ⅱ）, 10 mmol L^-1 （group Ⅲ） and 15 mmol L^-1 （group Ⅳ）. Semen suspensions were loaded in straws （0.5 mL） and placed in a controlled-rate freezer. After cryopreservation, frozen semen samples were thawed and investigated for progressive motility, viability using SYBR-14/EthD-1 staining and acrosome integrity using FITC-PNA/EthD-1 staining. There was a significantly higher （P 〈 0.01） percentage of progressive motility, viability and acrosomal integrity in two L-cysteine-supplemented groups （group Ⅱ and group Ⅲ） compared with the control. There was a biphasic effect of L-cysteine, with the highest percentage of progressive motility, viability and acrosomal integrity in group Ⅲ. In conclusion, 5 or 10 mmol L^-1 was the optimum concentration of L-cysteine to be added to the LEY extender for improving the quality of frozen-thawed boar semen.     

弱精子症大鼠模型的建立及左旋肉碱对其改善作用的相关研究

目的 研究弱精子症大鼠模型的建立及左旋肉碱(L-肉碱)与精子质量的关系.方法 24只雄性SD大鼠随机均分成3组,分别连续灌胃20d,A组(对照组):0.5%羟甲基纤维素钠(溶剂);B组:400mg/kg奥硝唑悬液:C组:400mg/kg奥硝唑悬液+100mg/kg左旋肉碱.末次给药24h后,麻醉处死所有大鼠,分别检测各组精子密度、活力、形态正常率以及附睾总L-肉碱和游离L-肉碱浓度.结果 A组、B组及C组的精子密度差异无统计学意义(P＞0.05);A组、C组精子活力、精子形态正常率及附睾总L-肉碱、游离L-肉碱浓度均明显高于B组(P＜0.05),A组与C组精子活力、精子形态正常率及附睾总L-肉碱、游离L-肉碱浓度比较,差异均无统计学意义.精子活力与附睾总L-肉碱、游离L-广肉碱浓度呈正相关(r=0.645,P＜0.05:r=0.676,P＜0.05),精子形态与附睾总L-肉碱、游离L-肉碱浓度呈正相关(r=0.557,P＜0.05;r=0.583,P＜0.05),均相关性其具有统计学意义.结论 奥硝唑可以降低大鼠精子活力、精子形态正常率,以及附睾L-肉碱水平,附睾L-肉碱浓度与精子活力、精子形态正常率呈正相关,L-肉碱对精子质量有改善作用.     

细胞外信号调节激酶和p38丝裂原活化蛋白激酶的表达水平对人精子活动力的影响

目的:比较细胞外信号调节激酶(ERK)、P38丝裂原活化蛋白激酶(P38MAPK)的磷酸化和蛋白表达水平在正常精子和弱精子症患者精子中的差异,旨在探讨ERK和P38MAPK表达水平与精子活动力的相关性。方法:收集正常精液(精子浓度≥20×106/ml,a级精子≥25%或a+b级精子≥50%)和弱精子症患者精液(精子浓度≥20×106/ml,a级精子<25%或a+b级精子≤40%)各20份,洗涤后提取精子总蛋白,采用Western印迹方法分别检测ERK、P38MAPK的磷酸化和蛋白表达水平。结果:ERK和P38MAPK在正常精子和弱精子症患者精子中均有表达,ERK、P38MAPK蛋白表达水平和P38MAPK磷酸化水平在弱精子症患者组显著升高(P<0.05),而两组间ERK磷酸化水平差异无统计学意义(P>0.05)。结论:人精子ERK、P38MAPK蛋白表达水平和P38MAPK磷酸化水平升高可能是精子活动力低下的原因之一。     

男性年龄与精子顶体酶活性及精子DNA碎片指数的相关性分析

目的探讨男性年龄与精子顶体酶活性、精子DNA碎片指数(DFI)的相关性。方法选取2016年1~8月在我院生殖医学中心就诊的436例不育症男性患者为研究对象,所有患者均行精液常规检查、精子顶体酶活性检查和(或)精子DFI分析。将患者按年龄分为<30岁、30~39岁、≥40岁3组,分析各组的精液常规、顶体酶活性及精子核DFI的差异及其相关性。结果不同年龄段患者的体重指数(BMI)、禁欲天数、精液量无显著性差异(P>0.05);年龄≥40岁组患者的前向运动精子百分率、活动精子百分率及精子顶体酶活性显著低于<30岁和30~39岁组(P<0.05);≥40岁组患者的精子DFI显著高于<30岁和30~39岁组(P<0.05)。年龄与前向运动精子百分率及活动精子百分率之间呈负相关(P<0.05),但是相关性较弱。精子顶体酶活性与精子正常形态率、前向运动精子百分率、非前向运动精子百分率、活动精子百分率呈正相关(P<0.05);精子DFI与年龄、禁欲天数、前向运动精子百分率呈正相关(P<0.01),与精液量、精子浓度、活动精子百分率呈负相关(P<0.05);精子顶体酶活性和DFI之间无相关性(P>0.05)。结论年龄增长会导致精液前向运动精子百分率、活动精子百分率、精子顶体酶活性、DFI等参数改变,直接或间接影响男性生育力。说明年龄对男性不育的影响是多方面的,建议有生育需求的大龄(≥40岁)男性尽早进行生育咨询与评估。     

弱精子症患者精子线粒体MTCYB、MTATP6基因的检测

目的:探讨精子线粒体MTCYB、MTATP6基因突变与弱精子症的关系。方法:提取80例成年男性弱精子症和20例活力正常者的精子mtDNA,设计PCR引物并扩增MTCYB、MTATP6基因,PCR产物纯化后进行序列测定和BLAST序列比对。结果:在80例弱精子症样本中有60例同时扩增出MTCYB、MTATP6片段,16例仅扩增出MTATP6片段,4例仅扩增出MTCYB片段。在20例精子活力正常的样本中均同时扩增出MTCYB、MTATP6片段。弱精子症样本中MTCYB和MTATP6基因的缺失率分别为20%和5%。扩增片断的序列分析发现,在弱精子症样本中,MTATP6基因出现G8887A的点突变,突变率为20%,而MTCYB基因未见有规律的突变。精子活力正常的样本中MTCYB、MTATP6基因未检测到明显的点突变。结论:精子线粒体MTCYB和MTATP6的基因缺失以及MTATP6基因的G8887A突变可能影响成年男性的精子活力。     

刺五加水提物体外对弱精子症患者精子运动参数的影响

目的:研究不同浓度的刺五加水提物体外对弱精子症患者精子运动的影响,探讨其可能的作用机制。方法:将35例弱精子症患者经手淫法获得并通过上游优化处理的精子,与不同浓度刺五加水提物共同孵育30、60、120、180 m in后,应用计算机辅助精子分析系统(CASA)观察不同浓度(2.5、5、10、20 g/L)刺五加水提物对人精子各运动参数的作用。结果:不同浓度刺五加水提物能明显提高弱精子症患者精子的运动能力,在浓度为5 g/L和10 g/L时,精子活率(MOT)、前向运动精子百分率、曲线运动速度(VCL)、直线运动速度(VSL)和平均路径速度(VAP)与空白对照组相比,差异均有显著性(P<0.05)。结论:刺五加水提物在体外能明显改善弱精子症患者精子运动能力,其最佳浓度为10 g/L。     

正常男子和弱精子症者精子中3β-羟基甾体脱氢酶的研究

目的：测定精子中3β-羟基甾体脱氢酶（HSD）。方法：采用酶组织化学方法对27例精子活率正常和20例精子活率低下的男子精子进行了研究。结果：3β-HSD主要分布在精子颈部，其着色面积不等，精子活率正常组3β-HSD着色面积为616．48±108．48μm2／100个精子，精子活率低下组为354．37±50．58μm2／100个精子，两组差异极显著（P＜0．001）。结论：3β-HSD可能通过某种途径间接参与精子的运动。     

重组人睾丸精子结合蛋白对人精子运动参数的影响

目的:探讨重组人睾丸精子结合蛋白(TSBP)对体外培养人精子运动参数的影响。方法:将22例生育男性的精液经Percoll密度梯度离心后,分别与0.01mg/ml及0.1mg/ml的重组His6-TSBP在体外共同孵育1h或3h,同时设立对照组,Western印迹检测重组His6-TSBP与精子膜的结合情况,计算机辅助精液分析(CASA)系统测定重组His6-TSBP对精子运动参数的影响。将12例弱精子症患者的精液按同样方法处理,检测重组His6-TSBP对弱精子症患者精子运动参数的影响。结果:0.1mg/ml重组His6-TSBP与生育男性精子作用1h可以提高体外培养精子的前向运动百分率(a+b级精子百分率),培养3h后前向运动百分率和活率均有所提高,差异具有显著性(P<0.05);0.01mg/ml重组His6-TSBP对检测各指标均无显著性影响。0.1mg/ml重组His6-TSBP与弱精子症患者精子作用3h可以提高精子前向运动百分率(P<0.05),但对活率无显著影响。结论:0.1mg/ml的重组His6-TSBP在体外可以提高生育男性精子的前向运动百分率和活率及弱精子症患者精子的前向运动百分率。     

弱精子症患者精子中CRISP2基因及蛋白的初步研究

目的:探讨弱精子症患者精子中富含半胱氨酸的分泌蛋白2(CRISP2)mRNA及蛋白的表达水平,探讨其与精子活力的关系及相关的分子机制。方法:收集成年男性弱精子症患者精液78例,活力正常精液70例作为对照组,50% Percoll离心分离纯化后,提取精子总RNA及总蛋白,采用RT-PCR,SYBR Green实时定量PCR和Western印迹技术检测CRISP2 mRNA及蛋白相对表达量。结果:CRISP2基因在弱精子症患者精子中的表达量明显低于其在正常对照组精子中的表达量,下降达4.3倍;Western印迹发现CRISP2蛋白在弱精子症组较正常对照组下降达1.71倍,两组之间的表达量有明显的统计学差异(P<0.05)。结论:CRISP2基因及蛋白在弱精子症患者精子中的表达下调可能与精子活力下降有密切联系,提示CRISP2基因及蛋白可能成为弱精子症发病机制研究的一个分子靶标。     

精子Tektin-2基因单核苷酸多态性与特发性弱精子症的相关性研究

目的探讨Tektin-2基因的单核苷酸多态性(SNP)位点rs12043423与特发性弱精子症的相关性。方法采用病例对照法,随机选取特发性弱精子症患者192例作为弱精子症组,另募集同期208例精子活力正常的不育男性作为不育症组和213例精液正常的已生育男性作为正常对照组,所有研究对象均进行精液分析,对三组患者Tektin-2基因的SNP位点rs12043423进行基因分型,比较三组间的基因型和等位基因频率,并且进行与特发性弱精子症的关联分析。结果(1)弱精子症组Tektin-2基因的SNP位点rs12043423的CC基因型频率显著低于正常对照组及不育症组,TT基因型频率则显著增加(P<0.05),而CT基因型频率在三组间的分布频率无显著性差异(P>0.05)。弱精子症组C等位基因的分布频率显著低于正常对照组和不育症组,而T等位基因的频率显著高于正常对照组和不育症组(P<0.05)。不育症组和正常对照组比较,不同基因型的分布频率及等位基因的频率在两组间均无显著性差异(P>0.05)。(2)弱精子症组与正常对照组比较,Tektin-2基因突变(杂合子[CT]和纯合子[TT])的发生率为61.5%vs.50.2%,TT基因型与弱精子症的风险因素分析结果为[OR=1.968,95%CI(1.041,3.723),P=0.035];弱精子症与不育症组比较,Tektin-2基因突变(CT+TT)的发生率为61.5%vs.51.5%,TT基因型与弱精子症的风险因素分析结果为[OR=1.918,95%CI(1.014,3.630),P=0.043]。结论Tektin-2基因rs12043423的多态性位点TT基因型和T等位基因增加特发性弱精子症的易感性,在特发性弱精子症的发展中可能是危险因素。     

精子发生相关新基因KLHL-10在无精子症及少弱精子症患者中突变筛查分析

目的:探讨精子发生相关新基因KLHL-10突变与无精子症及少、弱精子症之间的关系。方法:收集临床上不明原因的、非阻塞性无精子症及少、弱精子症患者(分别为11例、196例和118例)共325份外周血标本以及100份正常生育男性的外周血标本,抽提其DNA,采用PCR技术、变性高效液相色谱技术以及测序等手段对全部DNA样本进行KLHL-10基因的突变筛查。结果:在少精子症患者组及正常生育男性组中各发现1例及3例在1号外显子有C88→A的新的杂合突变,系同义突变;在少精子症患者组、弱精子症患者组及正常生育男性组中各检出3例、1例及4例在2号外显子有C424→A的新的杂合突变,也系同义突变;尚未发现有该基因的错义突变或微缺失。结论:KLHL-10基因错义突变或微缺失不是引起本组无精子症及少、弱精子症患者的主要致病原因,该基因在男性不育症的诊断价值有待进一步确定。