激光打孔技术用于冷冻精子复苏后体外受精的研究

目的 比较精子冷冻前后体外受精率的变化,以及卵子经过激光打孔后,与新鲜精子及冷冻复苏精子体外受精,比较受精率的变化.方法 冷冻2个近交系小鼠和5个品系的基因工程小鼠的精子,比较新鲜精子和冷冻复苏精子与C57BL/6J小鼠卵子做体外受精的结果;然后对C57BL/6J卵子做激光打孔,再与新鲜及冷冻复苏精子体外受精结果进行比较.结果 (1)新鲜C57BL/6J精子受精率为50.4％,C57BL/6N的受精率为48.6％,远高于冷冻精子复苏后的受精率(17.7％和15.4％);卵子打孔后,再与C57BL/6J及C57BL/6N的新鲜精子或冷冻复苏精子受精,受精率分别为64.6％,59.7％和60.4％,48.3％,冷冻精子复苏后的体外受精率提高显著;(2)5个基因工程小鼠新鲜精子和冷冻精子复苏后与C57BL/6J卵子的受精率分别为:91.5％ vs 28.6％,77.4％vs 52.3％,40.7％ vs 12.7％,56.5％ vs 14.6％,65％vs 6.5％;这5个品系的小鼠新鲜精子和冷冻精子复苏后与C57BL/6J打孔卵子的受精率分别为75％ vs 50.8％,74.3％ vs 35.6％,75.7％ vs32.7％,79.5％ vs 48.9％,59.5％ vs 46.3％,冷冻精子复苏后的受精率较之不打孔卵子,提高显著.结论 激光打孔技术用于冷冻复苏精子的辅助体外受精,可以显著提高冷冻精子复苏后的受精率,使得精子保种更加经济有效.     

两种抗冷冻剂对不同品系小鼠精子冷冻的影响

目的建立一套比较理想的小鼠精子冷冻复苏的方法。方法采用R18S3和FERTIUPTM-CPA两种保护剂,对DBA/2、C57BL/6J、KM和B6.129S7-Ldlrtm1Her/J四个品系的小鼠精子进行冷冻,冷冻精子用三种方法复苏,以体外受精率来评价精子冷冻的效果。结果以R18S3作为冷冻保护剂,DBA/2(73.3%,88.4%,55.6%)和KM(64.9%,60.2%,39.6%)品系小鼠冷冻精子复苏后体外受精率差异显著(P<0.05),C57BL/6J(3.0%,10.3%,3.7%)和B6.129S7-Ldlrtm1Her/J(0%,5.0%,0%)结果差异不显著(P>0.05);以FERTIUPTM-CPA作为冷冻保护剂,DBA/2(33.6%,14.1%,91.6%)、C57BL/6J(8.4%,21.0%,4.9%)和B6.129S7-Ldlrtm1Her/J(8.2%,10.0%,28.9%)品系小鼠冷冻精子复苏后体外受精率差异显著(P<0.05),而KM(48.1%,48.0%,48.1%)小鼠冷冻精子复苏后体外受精率差异不显著(P>0.05)。结论对于DBA/2和KM品系小鼠来说,用R18S3或FERTIUPTM-CPA冷冻精子,选择一种恰当的复苏方法,均可以得到较理想的体外受精率,而C57BL/6J和B6.129S7-Ldlrtm1Her/J品系小鼠无论采用哪种冷冻保护剂,选择何种方法复苏精子,得到的体外受精率都较低。     

不同品系小鼠的体外受精、胚胎冷冻及移植的比较研究

目的　探讨不同品系小鼠的体外受精、胚胎和精子的低温保存效果。方法　本实验分别在中国科学院上海实验动物中心 (SLAC)和日本熊本大学动物资源开发中心 (CARD)对 13个品系小鼠 (C57BL 6J、BALB c、C3H HeJ、ICR、KM、FVB、MRL、NOD、CBA、DBA 2、CD 1、BDF1、B6C3F1)的体外受精 (IVF)率、胚胎培养及移植成绩进行了比较研究。结果　各品系小鼠新鲜精子的IVF率 15 1%～ 87 9% ,冻融精子的IVF率 8%～ 80 % ;冷冻胚胎的复苏率4 2 6 %～ 83 9% ;冻融胚胎移植后的产仔率在 17 8%～ 5 1 8%。结论　遗传背景不同的小鼠体外受精率、冷冻胚胎复苏率和胚胎移植的产仔率差异有显著性。但同一品系两个实验室间的新鲜精子的IVF率、冷冻胚胎的复苏率及移植产仔率差异无显著性 (P >0 0 5 ) ;冻融精子的体外受精率CARD明显高于SLAC(P <0 0 1)。     

异种体外受精实验模型的建立及其方法学探讨

建立人和小鼠精子与去透明带金黄地鼠卵的异种体外受精实验模型(SPA)。8例小鼠精子对地鼠卵子的平均受精率为69.9±26.4%;16例次人精子对地鼠卵的平均受精率为55±20%,批内变异系数为14.5%,符合 WHO 标准,技术稳定可靠;PMSG 作用61h 可提高地鼠排卵量。     

棕色卵子形成原因及在体外受精胚胎移植中对妊娠结局的影响

目的:探讨棕色透明带卵子形成的可能原因以及对受精、胚胎质量、妊娠结局的影响。方法:回顾性分析2011-2012年在本中心接受体外受精胚胎移植助孕治疗的703例患者资料,根据透明带的色泽分为棕色透明带组(A组)74例,正常卵子组(B组)629例。比较两组一般临床特征、促性腺激素(Gn)使用天数、Gn用量、人绒毛膜促性腺激素(hCG)日血清激素水平、卵子透明带厚度、受精率、补救胞浆内单精子注射(ICSI)率、优质胚胎率、妊娠率。结果:两组的平均年龄、不孕年限、不孕原因、基础内分泌、hCG日血清雌二醇(E2)、孕激素(P)、黄体生成激素(LH)以及卵子受精率、卵裂率、补救ICSI率、可利用胚胎率差异均无统计学意义(P>0.05);A组的卵子透明带厚度大于B组,差异有统计学意义(P<0.05);A组Gn用量、hCG日血清卵泡刺激素(FSH)水平高于B组,优质胚胎率和妊娠率低于B组,差异均有统计学意义(P<0.05)。结论:棕色透明带的卵子与正常卵子相比受精率、补救ICSI率、卵裂率没有明显变化,透明带厚度有明显差异,但优质胚胎率和妊娠率明显下降,可能与患者Gn用量较多和hCG日血清高FSH水平有关。     

影响小鼠冻融精子体外受精率的主要因素探讨

目的分析影响小鼠精子冷冻成功的因素。方法用含3％脱脂奶和18％棉子糖的冷冻保护剂分别冷冻C57BL／6J、FVB／NJ、B6D2F1、KM和ICR小鼠精子。复苏后，评价小鼠精子的冷冻效果和受精能力。结果各品系小鼠精子冷冻前后的受精率不同；新鲜精子体外受精时，C57BL／6J、FVB／NJ、B6D2F1，KM和ICR五个品系的小鼠的受精率分别为81．4％、87．2％、90．4％、82．7％和79．4％而冻融精子的体外受精率分别为6．7％、46．0％、76．8％、59．2％和31．9％，差异极显著（P〈0．01）。结论遗传背景不同的小鼠品系，精子的受精能力及对冷冻的耐受能力不同。     

获能培养液等因素对遗传工程小鼠冻融精子体外受精率的影响

目的 探讨遗传工程小鼠精子冷冻、复苏及体外受精率的效果,建立简便、经济的遗传工程小鼠保种体系.方法 采用精子冷冻、体外受精和胚胎移植技术,比较了精子获能培养液、雄鼠周龄及精子冻存液等因素对于遗传工程小鼠精子冷冻保存的影响.结果 用CPA精子冻存液冷冻保存精子,精子复苏后用PM精子培养液获能培养,体外受精率在82.49%～91.43%,而HTF精子培养液的体外受精率为14.46%～27.38%,同品系间差异极显著(P<0.01);10～35周龄的雄鼠精子均能成功冷冻、受精,移植后产仔;使用R18S3,CPM,CPA三种不同精子冻存液冷冻遗传工程小鼠精子,复苏后采用PM精子培养液体外受精,受精率分别是75.85%、88.89%和94.27%,移植后得到阳性小鼠;体外受精后的胚胎成功冷冻保存,移植后产仔.结论 采用CPA精子冻存液冷冻保存精子,精子复苏后用PM精子培养液体外受精,能够更有效地保存遗传工程小鼠.     

精子正常形态率、顶体完整率、顶体反应率与卵子体外受精率的相关性研究

目的:通过检测常规体外受精中(in vitro fertilization IVF)精液常规分析正常男性的精子正常形态率、顶体完整率、顶体反应率并分析其与卵子的体外受精率之间的关系,旨在为辅助生殖技术提供几种可靠的判断精子生育力的参数与指标。方法:选择2008年7月1日到2009年12月30日间在广西医科大学第三附属医院生殖医疗中心进行控制性长方案超排卵下行常规IVF-ET的100对夫妻,共100个周期的资料做前瞻性分析,男方精液常规分析属正常范围,分析100份标本精子正常形态率、精子顶体完整率以及精子顶体反应率;观察并记录获卵个数、成熟卵个数、受精卵个数、未受精卵个数,计算其受精率。然后应用计量资料t检验,F检验以及线性相关等统计学方法对精子各项参数与卵子受精率之间的相关性进行分析。结果:实验组1,精液标本按其正常精子形态百分率分为3组,A组:≥5%而<10%、B组:≥10%而<15%、C组:≥15%,卵子体外受精率分别为67.05%、74.51%、82.49%,各组间两两相比差异有统计学意义(P<0.05)。实验组2,按顶体完整率分为两组,A组:≤85%组、B组:>85%组,卵子体外受精率分别为74.25%和82.14%,两组间比较差异有统计学意义(P<0.05)。实验组3,按顶体反应率分为三组,A组:<14%、B组:≥14%而≤25%、C组:>25%,其卵子体外受精率分别为73.45%、77.19%、82.74%,各组间的差异有统计学意义(P<0.05)。实验组4,按精子正常形态百分率分为两组<15%和>15%组,所对应的顶体完整率分别为79.51%、87.27%,两组间比较差异有统计学意义(P<0.05)。实验组5,用多重线性回归分析,精子正常形态率、顶体完整率、顶体反应率的标准化系数和P值分别为0.352、0.497、0.271;0.045、0.005、0.017,各个系数均有统计学意义。其中又以顶体完整率的标准化系数最大。结论:1.对于精液常规分析正常的男性,约有50%存在着不同程度的精子形态异常。卵子的受精率随正常形态精子所占百分率的升高而增高;其正常精子形态率≥5%,仍可得到一个较好的体外受精率,因此可行常规体外受精助孕。2.对于精液常规分析正常的男性,约有65%男性的精子顶体完整率>85%,随着精子顶体完整率的增加,卵子的受精率也明显增高;3.对于精液常规分析正常的男性,其78%男性精子顶体反应率≥14%,在体外受精中卵子的受精率随着精子顶体反应率所占百分比的增加而增高;4.上述三者都是影响卵子受精的重要因素,卵子体外受精率与以上三项指标间存在较强的相关关系,而其中又以顶体完整率对卵子体外受精率的影响程度最大。5.对于精液常规分析正常的男性,若正常精子形态率≥5%,在体外受精助孕前,精子顶体完整率和精子顶体反应率的检测,可不作为常规检查项目。     

FVB小鼠精子冷冻及体外受精的研究

目的：探索FVB小鼠及FVB遗传工程小鼠种系保存的新途径.方法：用孕马血清和人绒毛膜促性腺激素（HCG）超排FVB小鼠,并分别用复苏精子和新鲜精子进行体外受精并将受精卵移植到假孕雌鼠输卵管中,比较两种精子的受精率;对注射HCG后13,15,17h不同时间采集FVB小鼠的卵母细胞进行体外受精,比较其受精率及体外培养至2细胞情况.结果：新鲜精子的受精率（62.4%）略高于冷冻复苏精子的受精率（58.1%）;在注射HCG 13～15 h后取FVB小鼠卵母细胞其受精率效果较好,受精率（60.2%、61.6%）显著高于17h取卵的受精率（51%）,但13,15,17 h采集卵母细胞体外受精后胚胎发育至2细胞及异常卵数无显著性差异.结论：本研究将为FVB小鼠及FVB遗传工程小鼠的种系保存、繁殖提供了基础.     

人类卵子改良程序化冷冻和玻璃化冷冻的比较研究

【】目的：研究两种冷冻方法对人类卵子冻融结果的影响。方法：分为程序化冷冻卵子与玻璃化冷冻卵子两组,对于卵子冻融后的复苏率、受精率以及优质胚胎率等指标进行比较。结果：程序化冷冻组卵子的复苏率(71.05%,108/152)显著低于(P<0.05)玻璃化冷冻组(81.75%,103/126),但优质胚胎率(41.07%,23/56)显著高于(P<0.05)玻璃化冷冻组(30.23%,13/53),而受精率、卵裂率及囊胚形成率无统计学差异。结论：玻璃化冷冻卵子可以获得较高的复苏率,但使用改良程序化冷冻试剂盒冻融卵子,可能较玻璃化冷冻获得更好发育潜能。     

Value of D-Dimers in patients with acute aortic dissection

Objective: To evaluatel the value of D-dimers in patients with acute aortic dissection (AAD). Methods: This study consisted of 16 patients with AAD and 27 non-AAD patients. Serum D-dimets were measured by Sta-Liatest D-DI immunoturbidimetric assay. Results: D-dimer level was higher (P ＜ 0.001) in patients with AAD(7.91 ± 5.52 μg/ml) than that in non- AAD group(1.57±1.24 μg/ml). D-dimer was positive (＞0.4 μg/ml) in all patients with AAD and in 10 control group patients (37%). Among patients with acute AAD, D-dimers tended to be higher in Stanford A than in Stanford B (8.67 ± 4.31 μg/ml vs. 3.24±1.27 μg/ml, P ＜0.01). D-dimer values tended to be higher in more extended disease(3.84 ± 1.65 μg/ml, 8.57 ± 3.58 μg/ml and 11.87 ± 5.69 μg/ml in thoracic aorta, thoracic and abdominal aorta, thoracic and abdominal aorta and iliacal arteries, respectively, P ＜ 0.05 for both 8.57 ± 3.58 and 11.87 ± 5.69 vs. 3.84 ± 1.65 ). Including the control group into the analysis, we found a sensitivity of 100%, a negative predictive value of 100%, and a specificity of 66% and a positive predictive value of 64% for D-dimer in diagnosis of AAD in our patients with suspected AAD. Conclusion: D-dimer was elevated in patients with AAD. A negative D-dimer test result could be useful in excluding AAD.     

Research of combined liver-kidney transplantation model in rats

Objective： To set up a simple and reliable rat model of combined liver-kidney transplantation. Methods： SD rats served as both donors and recipients. 4℃ sodium lactate Ringer＇s was infused from portal veins to donated livers,and from abdominal aorta to donated kidneys, respectively. Anastomosis of the portal vein and the inferior vena cava （IVC） inferior to the right kidney between the graft and the recipient was performed by a double cuff method, then the superior hepatic vena cava with suture. A patch of donated renal artery was anastomosed to the recipient abdominal aorta. The urethra and bile duct were reconstructed with a simple inside bracket. Results： Among 65 cases of combined liver-kidney transplantation, the success rate in the late 40 cases was 77.5%. The function of the grafted liver and kidney remained normal. Conclusion： This rat model of combined liver-kidney transplantation can be established in common laboratory conditions with high success rate and meet the needs of renal transplantation experiment.     

BLOOD PRESSURE CHANGE WITH AGE IN SALT-SENSITIVE TEENAGERS

Objective To observe blood pressure change with age in salt-sensitive teenagers whose salt sensitivity were determined by repeated testing.Methods Salt sensitivity was determined through intravenous infusion of normal saline combined with volume-depletion by oral diuretic furosemide in 55 teenagers. After five years, salt sensitivity was re-examined and subject blood pressure was followed up. Blood pressure changes in salt-sensitive teenagers were compared to that of non-salt sensitive teenagers over five years.Results After 5 years, the repetition rate of salt sensitivity determined by intravenous saline loading is 92.7%. In teenagers with salt sensitivity on the baseline, both the systolic blood pressure increments and increment rates were much higher than non-salt sensitive teenagers (12.7±12.1 mmHg vs. 2.8±5.2 mmHg, P＜ 0.01; 12.2%± 12.0% vs. 2.5% ±4.4%, P＜ 0.001,respectively). There was a similar trend for diastolic blood pressure (8.4 ± 6.4 mmHg vs. 3.7 ± 6.4 mmHg, P = 0.052; 13.2% ±10.6 % vs. 6.8%± 10.1%, P = 0.053, respectively).Conclusions Salt sensitivity determined by intravenous saline loading showed good reproducibility. Blood pressure increments with age were much higher in salt-sensitive teenagers than non-salt sensitive teenagers, especially in terms of systolic blood pressure.     

安心颗粒对急诊经皮冠状动脉介入术后生活质量影响的研究

目的：评价使用安心颗粒对急诊经皮冠状动脉介入术（PPCI）术后生活质量的影响.方法：将160例接受PPCI的急性ST段抬高型心肌梗死患者随机分为安心颗粒组（术前顿服安心颗粒8.8g,术后安心颗粒4.4 g/次,每日2次）和对照组（仅接受基础药物治疗）.所有患者均服用阿司匹林、氯吡格雷和阿托伐他汀.分别在入院时、出院前1d、出院后180 d时,应用心肌梗死多维度量表（MIDAS）、中文版SF-36评价量表对患者生活质量评分.并观察术后30 d以内的出血并发症、血小板减少症发生情况.结果：入院时和出院前1d,两组患者的心肌梗死MIDAS、SF-36量表评分比较无差异（P＞0.05）;出院后180 d时,与对照组比较,安心颗粒组MIDAS、SF-36评分明显减低（P＜0.05）;组内与入院时比较,两组出院前1d、出院后180 d时,MIDAS、SF-36评分均降低（P＜0.05）.两组患者在随访期间均无大量出血、少量出血、重度和极重度血小板减少症发生,安心颗粒组有4例、对照组有7例发生不明显出血（P＞0.05）.两组发生轻度血小板减少症的患者数比较无差异（P＞0.05）.结论：PPCI使用安心颗粒,能改善急性ST段抬高型心肌梗死患者的生活质量,且不增加出血风险.     

The comparative studies of the influences of Urapidil and Nicardipine on sino-atrial node function, atrio-ventricular node function and hemodynamics

Objective:To investigate the influences of urapidil and nicardipine on rabbit sinus function,atrio-ventricular node function and hemodynamics.Methods:Thirty-two Angora's rabbits were selected and randomly divided into four groups.U1 group:urapidil 0.25 mg/kg;U2 group:urapidil 0.5 mg/kg;N1 group:nicardipine 10 μg/kg;N2 group:nicardipine 20 μg/kg.All these medicine were administrated within 30 seconds.Measurements were taken before and after the administration of urapidil or nicardipine for the following data:mean blood pressure(MAP),heart rate(HR),sino-atrial conduction time(SACT),maximal sinoatrial recovery time(SNRTmax)corrected sinus node recovery time(CSNRT),index of sinus node recovery time(SNRTI),Wenckebach A-V conduction frequency (WB),and P-R interval.Results:Significant MAP and HR changes were identified in all of the four groups before and after administration of both urapidil and nicardipine.No significant changes could be found in the rest of the parameters.Intergroup analysis showed that SACT and CSNRT of N1 and N2 groups were shorter than those of the U2 group(P＜0.01);the MAP decreased(P＜0.01)and the HR increased drastically(P＜0.01).Conclusions:Neither urapidil(0.25 mg/kg,0.5 mg/kg)nor nicardipine(10μg/kg,20μg/kg)has any significant influence on rabbit sinus function or rabbit atrio-ventricular node function.Nicardipine could be a better choice than urapidil for parafunctional sinus node patients.     

Expression of OPGmRNA and ODFmRNA in the patients of hip fracture due to osteoporosis

Objective：To investigate the gene expression of osteoprotegerin（OPG） and osteoclast differentiation factor（ODF） in the bone tissue of patients with hip fracture due to osteoporosis. Methods：OPGmRNA and ODFmRNA in the bone tissue in 50 cases of osteoporosis sufferers（over 50 years old） with hip fracture（Observer Group） and 30 cases of hip facture sufferers with no osteoporosis（Control group） were analyzed with the Semi-Quantitative RT-PCR method. Results：The mRNA expressed of ODF, OPG were both high in the patients with hip fracture. In the control group, the expression of OPG mRNA was observed, while the expression of ODF mRNA was very slight. Conclusion：Aged patients contained all signals including OPG, ODF that are essential for inducing osteoclastogenesis and promoting bone resorption.     

The clinicopathological and immuohistochemical analysis of solid-pseudopapillary tumor of the pancreas： report of 9 cases

Objective：To investigate the clinical features, pathological characteristics and immunophenotype of solid-pseudopapillary tumor of the pancreas（SPTP）. Methods：Nine surgically treated cases of SPTP were retrospectively reviewed. Hematoxylin and Eosin（HE） staining and immunohistochemical staining were used to analyze all cases, and the general clinical data was collected. Results：Six patients were asymptomatic except for a palpable mass. Two patients complained of vague-epigastric pain. One patient appeared jaundice. The tumor was encapsulated and solid tissues alternately with cystic tissues. Histologically, the histological structure of solid portion was pseudopapillary with a fibrovascular core. Tumor cells were uniform and medium-sized which were arranged in sheets ets or nests or pseudopapillary patterns. Immunohistochemical studies demonstrated that SPTP proved positive in vimentin（9/9 cases）, AAT（9/9 cases）, NSE（9/9 cases）, ACT（7/9 cases）, CK20（2/9 cases）, CgA（1/9 cases）, S-100（3/gcases）, PR（4/gcases）, Syn（3/9 cases） and CD56（5/9cases）, negative in CEA and ER. Conclusion：SPTP is a tumor predominantly occurring in young women frequently without special symptoms. This tumor has various characteristical histological patterns with different immunophenotype.     

Dynamic Changes in Serum Estradiol and Progesterone levels in Patients of Premenstrual Syndrome with Adverse Flow of Liver-qi

Objective:To probe into the influence of changes of ovarian hormones on the pathogenesis of the specific sub-type premenstrual syndrome(PMS)and reveal partial microcosmic mechanisms of adverse flow of liver-qi.Methods:Estradiol(E2)and progesterone(P)levels in serum were determined at different phases of menstrual cycle by radioimmunoassay.Results:In the group of PMS with adverse flow of liver-qi.the secretive peak value Of E2 and P at the follicular phase significantly decreased,and the secretive peak value at the luteal phase did not come into being.Conclusions:Low E2 and P secretive peak at the follicular phase and absence of secretive peak at the luteal phase is one of the microcosmic mechanisms of PMS with adverse flow of liver-qi.One of the pathophysiologic mechanisms of specific sub-type PMS is probably the continuous low level of E2and P.     

Real-time Three-dimensional Echocardiography in Assessment of Left Ventricular and Right Ventricular Volumes

Real-time three-dimensional echocardiography (RT3DE)is a new ultrasound technique that enables dynamic threedimensional visualization and quantification of the heart in real time. Investigation of feasibility and methodology of RT3DE in determining left ventricular (LV) and right ventricular (RV) volumes, RT3DE was performed in 35 normal adults using Philips SONOS 7500 system with a 2-4 MHz matrix array transducer. The 60°×60° "pyramid" volume database was obtained and analyzed on a TomTec echo workstation. Both LV and RV volumes were calculated with four 3DE methods (i.e. apical 2, 4, 8, and 16-plane) through manually tracing ventricular endocardial borders in end diastole and end systole. Stroke volumes were then calculated. LV volume was also measured by 2DE Simpson's rule using GE VIVID 7 ultrasound machine.     

SCREENING FOR A 21-CHROMOSOME ABNORMALITY IN PREIMPLANTED EMBRYOS OF ELDERLY WOMEN

Increasing maternal age is the only etiological factor unequivocally linked to Down's syndrome in humans. The occurrence rate of newborns with Down's syndrome is about 1/220 in women over 35 years old. However, the occurrence rate in embryos fertilized in vitro, of the elder woman is unclear. Using FISH we screened the number of chromosome 21 in preimplanted embryos of 5 elderly women (average age, 38.4 years) to study the feasibility and necessity of screening trisomy 21 in embryos in patients over 35 years old at the in vitro fertilization (IVF) center.