卵巢癌早期诊断方法的研究进展

基因芯片与SELDI-TOF蛋白质芯片技术在卵巢癌早期诊断中得到了广泛应用，但SELDI-TOF蛋白质芯片技术存在的内在缺陷（测试结果的不精确性以及数据分析技术的不成熟性）。改进卵巢癌早期诊断的研究策略有：(1)把基因芯片技术和SELDI-TOF蛋白质芯片技术结合起来进行卵巢癌早期诊断研究；(2)采用MALDI-TOF技术来实现血清蛋白质谱的测试；(3)开发更有效的数据挖掘算法。     

核磁共振成像在诊断胎儿先天畸形中的应用——附2例报道及文献复习

目的总结和探讨核磁共振成像在产前诊断胎儿先天畸形的作用。方法对2例产前初步诊断为胎儿先天畸形并行核磁共振检查的临床资料进行分析，同时回顾复习近3年国外有关MRI诊断胎儿畸形的相关文献。结果核磁共振对胎儿先天畸形的诊断有较高的特异性。结论孕晚期MRI可作为B超的重要补充辅助检查项目而提高胎儿先天畸形的确诊率。     

SELDI-TOF-MS技术检测心绞痛患者血浆蛋白指纹图谱的研究

目的： 采用表面增强激光解析/电离飞行时间质谱（SELDI-TOF-MS）技术检测心绞痛患者血浆蛋白质指纹图谱,从中筛选出特异的分子标志物。方法: 应用SELDI-TOF-MS技术及金属离子螯合（IMAC-3）蛋白质芯片对20例心绞痛患者和29例正常对照者血浆样本进行检测,借助生物信息学工具（非线性的支持向量机,SVM）提出心绞痛的诊断模型,并运用留一交叉验证法来评估该模型的判别效能。结果: 用SELDI-TOF-MS技术筛选出由3个有显著差异的蛋白质峰［质荷比（m/z）分别为2 667.3、5 914.0和6 890.5］组合构建的诊断模型,可将20例心绞痛患者和29例正常人全部正确分组,诊断特异性和灵敏度均为100%。结论: SELDI-TOF-MS技术在心绞痛的诊断中具有较高灵敏度和特异性,发现的蛋白质峰可能在心绞痛的发病中起一定作用,血浆中分子标志物的发现有助于心绞痛的早期诊断。     

SELDI-TOF-MS技术在口腔黏膜癌前病变和口腔鳞癌的唾液诊断模型中的应用

目的:分析唾液中蛋白质组对口腔鳞癌患者的动态变化的意义,对口腔鳞癌和癌前病变的相关诊断模型的建立进行初步探索。方法:按标准提取唾液标本,采用SELDI-TOF-MS技术,对17例口腔鳞癌患者、7例区域性转移患者、8例白斑患者和15例健康志愿者的唾液蛋白质指纹图谱进行比较,并采用整合性生物信息学方法建立诊断模型。结果:得到唾液诊断模型Ⅰ,为原发性口腔鳞癌患者与正常人的诊断模型:质荷比分别为5797、2902、3883和4951的4个蛋白质峰。其中5797、2902和3883在原发性口腔鳞癌患者唾液中低表达。4951在原发性口腔鳞癌患者唾液中高表达。敏感性为93.33%,特异性为88.24%,总准确率为90.63%。唾液诊断模型Ⅱ,为OSCC区域性转移患者与正常人的诊断模型:质荷比分别为5446、4934、4968、3296、4800和2901的6个蛋白质峰。其中5446、4934和3296在口腔鳞癌区域性转移患者唾液中高表达。4968、4800和2901在口腔鳞癌区域性转移患者唾液中低表达,敏感性为100.00%,特异性为85.71%,总准确率为95.45%。唾液诊断模型Ⅲ,为白斑患者与正常人的诊断模型:质荷比分别为9515、2230、2238、5694和2646的5个蛋白质峰。其中9515和5694在白斑患者唾液中高表达。2230、2238和2646在白斑患者唾液中低表达,敏感性为93.33%,特异性为85.71%,总准确率为86.96%。结论:结合SELDI-TOF-MS和生物信息学方法,唾液对肿瘤的诊断意义将进一步发挥其价值。本研究根据口腔粘膜上皮癌前病变、癌变、转移的唾液蛋白质组的变化,初步建立了3个早期诊断模型。     

超声诊断胎儿羊水过多与畸形的相关性研究

目的探讨羊水过多和胎儿畸形的相关性。方法收集2008年7月～2011年8月产前超声检查孕妇资料,按产科常规超声检查程序扫查胎儿全身各部位,检查胎儿头颅、颜面、脊柱、躯干、四肢、胸腔脏器、腹腔脏器、胎盘及羊水情况。结果 11735例胎儿中发现羊水过多135例,检出率1.2%;伴发畸形31例,发生率23.0%。结论发现羊水过多病例,要加强产前早期进行B超检查可以及时发现先天畸形,及时进行正确处理。     

产前超声对胎儿神经系统畸形的诊断价值

目的探讨超声检查在胎儿神经系统畸形诊断中的价值.方法回顾性分析2003～2008年来我院产科就诊并行常规产前超声检查的孕妇共12000例.结果 12000例孕妇产前超声检查中,检出胎儿畸形153例,发生神经系统畸形64例,其中脑积水29例,脑脊膜膨出9例,无脑儿9例,全前脑3例,Dandy-Walker畸形3例,脊柱裂11例.结论超声检查对产前诊断胎儿神经系统畸形有重要价值,对提高出生人口素质和优生优育有重要意义.     

帕金森病相关血清蛋白质标志物的筛选及鉴定

 目的：探讨并初步鉴定帕金森病患者血清中相关蛋白质作为特异标志物的可能性。方法：应用表面增强激光解吸电离飞行时间质谱(SELDI-TOF-MS)结合纳米磁珠技术检测44例帕金森病和60例健康对照的血清标本,应用生物信息学方法筛选差异蛋白峰,经高效液相色谱（HPLC）分离出差异蛋白,酶解后进行液质联用串联质谱（LC-MS/MS）分析,利用Xcalibur的程序组件BioWorks 3.2完成蛋白质序列数据库鉴定分析。结果：经SELDI-TOF-MS结合纳米磁珠技术筛选出质荷比m/z位于8 937的蛋白质在帕金森病中高表达(帕金森病组表达强度为27.47±16.58,正常组表达强度为5.01±3.47),有显著差异(P<0.01);6 636和8 697的蛋白质在帕金森病中低表达(帕金森病组表达强度为5.43±2.66和20.22±9.57,正常组表达强度为18.85±7.56和51.13±26.22), 有显著差异(P<0.01)。联合上述3种潜在蛋白质标志物,可区分帕金森病组和对照组,其中帕金森病患者检出率为90.0%(27/30),健康者检出率为92.5%（37/40）。对m/z为6 636、8 697和8 937的标志物进行鉴定,结果分别为载脂蛋白C-I、载脂蛋白 C-III 和补体成分3a。结论：鉴定出的载脂蛋白 C-I、载脂蛋白C-III和补体成分3a在帕金森病的诊断中具有一定价值,值得进一步研究和探讨。     

应用SELDI-TOF-MS技术建立胰腺癌筛选血清蛋白指纹图谱模型

目的：建立胰腺癌筛选血清蛋白质指纹图谱模型。方法：应用表面增强激光解析离子化飞行时间质谱（SELDI-TOF-MS）技术,采用弱阳离子交换芯片（CM10）首先对33例胰腺癌患者和31例性别年龄匹配的健康对照血清蛋白表达谱的差异进行了分析,建立决策树分类模型。结果：在质荷比（M/Z）2000～20000范围内,共检测到185个有效蛋白峰,其中12个峰差异有显著性（P〈0.01）。自动选用M/Z为11674.7和7771.2的2个差异蛋白峰,建立胰腺癌决策树分类模型,其灵敏度为96.97%（32/33）,特异度为96.67%（30/31）。结论：应用SELDI-TOF-MS技术可以筛选出胰腺癌相关的血清蛋白标志,建立的决策树模型可能对胰腺癌的诊断具有重要的临床价值。     

蛋白质指纹图谱在乳腺癌诊断与随访中的应用研究

目的:应用表面加强激光解吸电离-飞行时间质谱(SELDI-TOF-MS)和CM10蛋白质芯片检测乳腺癌患者血清蛋白质指纹图谱,筛选候选肿瘤标志物,建立、验证诊断模型并初步探讨其在术后随访监测中的价值。方法:SELDI-TOF-MS技术及CM10芯片检测63例乳腺癌和40例健康人的指纹图谱,ZJU-PDAS软件筛选标志物、建立模型,在另23例乳腺癌和20例健康人中盲法验证模型;并检测16份术后标本以探索其随访监测价值。结果:质荷比(m/z)为3.9kD和5.6kD的2个蛋白质峰(分别命名为BC1和BC2)构建的模型诊断乳腺癌灵敏度为87.30%(55/63)、特异度为95.00%(38/40);盲法验证灵敏度为95.65%(22/23)、特异度为85.00%(17/20);对不同病期患者具有相同诊断效力(P0.05)。BC1手术后表达升高;BC2手术后表达降低,且复发转移组表达高于无瘤生存组(P0.05)。结论:该方法在乳腺癌的诊断、术后随访及肿瘤标志物筛选等方面具有一定价值,值得进一步研究。     

应用羊水细胞间期荧光原位杂交技术诊断胎儿Down综合征

目的 建立以未培养的羊水间期细胞直接制片进行荧光原位杂交(FISH)的简便方法并应用于Down综合征产前诊断。方法 Down综合征患者外周血染色体标本,以及10例产前诊断的孕妇,在妊娠16～20周行羊膜腔穿刺术,羊水细胞制片并以胃蛋白酶进行简单处理后,用位于染色体21q22.3区域的人类基因组BAC克隆探针进行荧光原位杂交。结果 全部杂交成功,信号强,背景低。讨论 该技术简便、稳定、易掌握,适用于检测胎儿Down综合征的快速产前诊断。     

Anti-cardiolipin antibodies in fetal blood and amniotic fluid derived from patients with the anti-phospholipid syndrome

The aim of this study was to investigate whether, in patients with antiphospholipid syndrome, anticardiolipin antibodies pass from mother to offspring sera and amniotic fluid. Eleven patients with antiphospholipid syndrome (study group) and 11 healthy controls, matched by maternal and gestational age (control group) were prospectively examined for the presence of anticardiolipin antibodies in the cord blood during labour, and amniotic fluid during vaginal or Caesarean delivery. Three neonates (27.3%) in the study group had anticardiolipin antibodies in the cord blood, while none had them in the control group. Anticardiolipin antibodies were detected in the amniotic fluid in six (54.5%) of the study group pregnancies, compared with none in the control group. No adverse neonatal outcome was noted except for significantly lower (P < 0.0006) mean birth weight in the study group. Anticardiolipin antibodies can pass the placenta and be detected in fetal cord blood and amniotic fluid. This finding might be used in the future for the assessment of pregnancies with antiphospholipid syndrome.     

SELDI-TOF-MS技术筛选NSCLC血清肿瘤标志物

目的 分析非小细胞肺癌(NSCLC)血清蛋白表达谱的改变,筛选并建立NSCLC血清蛋白的差异表达谱。方法 应用表面增强激光解析电离化飞行时间质谱(SELDI-TOF-MS,简称SELDI-TOF或SELDI)技术,分析100例NSCLC患者和100例正常对照血清,获得蛋白表达图谱。用Biomarker Pattern(BPS)软件分析NSCLC差异蛋白并初步建立诊断模型。通过盲筛进一步验证诊断模型。结 果 发现NSCLC患者血清与正常人有16个显著差异的蛋白波峰,显著高表达8个（P<0.001）。显著低表达8个（P<0.001）。经BPS软件分析,建立分类树模型,其诊断的灵敏度为100％,特异度为100％。100例样本盲筛验证结果显示,其灵敏度为96.15％,特异度为95.83％。无吸烟史患者较有吸烟史患者血清中显著高表达蛋白质波峰有3个（P<0.05）。鳞癌患者较腺癌患者血清中显著高表达蛋白质波峰有2个（P<0.01）。不同病理分期患者之间未发现差异蛋白峰。结 论 SELDI-TOF-MS 技术可筛选出NSCLC差异性蛋白并建立NSCLC诊断的分类树模型,有望成为NSCLC早期诊断的辅助指标。     

Hepatocyte growth factor concentration in the early second-trimester amniotic fluid does not predict fetal growth at birth.

The purpose of this study was to evaluate whether hepatocyte growth factor (HGF) concentrations in the early second-trimester amniotic fluid predict fetal growth at birth. HGF and insulin-like growth factor-I (IGF-I) concentrations in the early second-trimester amniotic fluid were measured in 12 pregnancies with small for gestational age (SGA) infants, 84 pregnancies with appropriate for gestational age (AGA) infants, and eight pregnancies with large for gestational age (LGA) infants. HGF concentrations were measured from the early second-trimester amniotic fluid samples using an enzyme-linked immunosorbent assay. IGF-I concentrations were measured from the early second-trimester amniotic fluid samples using an immunoradiometric assay. Maternal age in AGA group (34.2 +/- 5.5 years) was significantly lower than in SGA (37.9 +/- 3.0 years) and LGA (37.6 +/- 3.3 years) groups (P < 0.05). There were no significant differences for parity or gestational age at amniocentesis among the groups. There were significant differences for birth age, birth weight, neonatal height, and placental weight among the groups (P < 0.05). HGF concentrations in SGA, AGA and LGA groups were 16.9 +/- 6.6, 16.7 +/- 9.0 and 20.2 +/- 14.8 ng/ml respectively (not significant). There was no correlation between amniotic fluid HGF concentrations and birth weight, height or placental weight. There were also no significant differences for amniotic fluid IGF-I concentrations among the three groups. These results suggest that differences in HGF concentrations in the early second-trimester amniotic fluid do not predict fetal growth at birth. Further study is needed to clarify the role of high HGF concentrations in early second-trimester amniotic fluid during pregnancy.     

Microarray comparative genomic hybridization (CGH)-based prenatal diagnosis for chromosome abnormalities using cell-free fetal DNA in amniotic fluid

Cell-free fetal DNA (cffDNA) in the supernatant of amniotic fluid, which is usually discarded, can be used as a sample for prenatal diagnosis. For rapid prenatal diagnosis of frequent chromosome abnormalities, for example trisomies 13, 18, and 21, and monosomy X, using cffDNA, we have developed a targeted microarray-based comparative genomic hybridization (CGH) panel on which BAC clones from chromosomes 13, 18, 21, X, and Y were spotted. Microarray-CGH analysis was performed for a total of 13 fetuses with congenital anomalies using cffDNA from their uncultured amniotic fluid. Microarray CGH with cffDNA led to successful molecular karyotyping for 12 of 13 fetuses within 5 days. Karyotypes of the 12 fetuses (one case of trisomy 13, two of trisomy 18, two of trisomy 21, one of monosomy X, and six of normal karyotype) were later confirmed by conventional chromosome analysis using cultured amniocytes. The one fetus whose molecular-karyotype was indicated as normal by microarray CGH actually had a balanced translocation, 45,XY,der(14;21)(q10;q10). The results indicated that microarray CGH with cffDNA is a useful rapid prenatal diagnostic method at late gestation for chromosome abnormalities with copy-number changes, especially when combined with conventional karyotyping of cultured amniocytes.     

Chronic chorioamnionitis displays distinct alterations of the amniotic fluid proteome

Acute chorioamnionitis of infectious origin and chronic chorioamnionitis of immunological origin are two major placental lesions of spontaneous preterm birth with elevated amniotic fluid interleukin-6 and CXCL10 concentrations, respectively. The changes in the amniotic fluid proteome associated with intra-amniotic infection and acute chorioamnionitis are well defined, yet alterations unique to chronic chorioamnionitis remain to be elucidated. This study was conducted to determine those amniotic fluid proteins changing specifically in the presence of chronic chorioamnionitis. Amniotic fluid obtained from acute chorioamnionitis, chronic chorioamnionitis and gestational age-matched controls were analysed by two-dimensional (2D) difference in gel electrophoresis and MALDI-TOF analyses. The type of histological inflammation was used to define each condition in preterm labour cases (n = 125) and term not in labour cases (n = 22), and the amniotic fluid concentrations of interleukin-6, CXCL8, CXCL10 and prostaglandin F(2α) were also measured by specific immunoassays. Among preterm labour cases, 31 differentially expressed proteins were identified in chronic chorioamnionitis cases as compared to both acute chorioamnionitis and control cases. Importantly, glycodelin-A, which maintains maternal tolerance against an allogeneic fetus, was decreased in chronic chorioamnionitis, while haptoglobin was increased. We report the amniotic fluid proteome of chronic chorioamnionitis for the first time, and the findings herein strongly suggest that there is a pathophysiological association between the changes of immunomodulatory proteins in the amniotic fluid and chronic chorioamnionitis, a histological manifestation of maternal anti-fetal allograft rejection.     

S100B protein expression in the amnion and amniotic fluid in pregnancies complicated by pre-eclampsia

Our aim was to investigate the expression of S100B protein in the amnion and to assess the amniotic fluid concentration in pregnancies complicated by pre-eclampsia. Samples were obtained from women who developed pre-eclampsia (n = 7), pre-eclampsia with intrauterine growth retardation (IUGR) (n = 4), normotensive IUGR (n = 7) and gestational hypertension (n = 4) during pregnancy and healthy controls who delivered at term (n = 35). To determine the difference in the expression of S100B in the amnion, we performed immunohistochemistry, western blot analysis and RT-PCR. Using enzyme-linked immunosorbent assay (ELISA), we assessed the S100B concentration in amniotic fluid. The S100B mRNA expression in the amnion of pre-eclamptic patients and patients with pre-eclampsia with IUGR was significantly higher than that in the control. The amniotic fluid S100B protein concentration of the pre-eclampsia and normotensive IUGR cases was significantly higher than that of the control. This study shows that amnion could be a source responsible for the increased concentration of S100B in amniotic fluid. In pre-eclampsia, reactive oxygen species (ROS) are generated by oxidative stress. Some pathological conditions that develop during pregnancy and are related to hypoxic stress can affect the elevation of S100B concentration in the amnion.     

Potential biomarkers found by protein profiling may provide insight for the macrovascular pathogenesis of diabetes mellitus

Diabetes mellitus (DM) is an alarming threat to health of mankind, yet its pathogenesis is unclear. The purpose of this study was to find potential biomarkers to serve as indicators for the pathogenesis of DM in a time course manner. Based on our previous findings that oxidative stress occurred at week 8, aorta lysate and sera of 102 streptozotocin (STZ)-induced diabetic and 85 control male Sprague-Dawley rats were obtained at the 4th, 8th and 12th week after STZ injection. The protein profiles were studied employing surface-enhanced laser desorption/ionization time-of-flight mass spectrometry technology in attomole sensitivity range. In the aorta, a multiple biomarker panel was discovered at the 4th week. At the 8th week, 4 biomarkers were found, while at the 12th week, 3 biomarkers were identified. In the sera, a triplet of 3 peaks and 2 biomarkers were all discovered to have 100% classification accuracy rate to differentiate the DM and control groups at all time intervals. Besides, 2 biomarkers were also found to have high classification value at week 12. Comparing the aorta and sera from DM and non-DM rats, a bundle of potential biomarkers with significant changes in peak intensities and high classification values were found. Two of the serum biomarkers matched with islet amyloid polypeptide and resistin in the SWISS-PROT knowledgebase. Validation has been conducted using immunoassay kits. These potential biomarkers may provide valuable insight on the pathogenesis of DM and macrovascular complications.     

Macrophage inhibitory cytokine 1 in fetal membranes and amniotic fluid from pregnancies with and without preterm labour and premature rupture of membranes

The placenta and fetal membranes are the site of expression of macrophage inhibitory cytokine (MIC-1), a member of the transforming growth factor (TGF)-beta superfamily. We hypothesized that MIC-1 may act as an immune regulator in pregnancy complications associated with intrauterine inflammation. Decidual cells, chorionic trophoblasts and amnion epithelial cells were identified by immunohistochemistry as the predominant MIC-1-containing cell type in term membranes. Amnion and choriodecidual explants all produced MIC-1 in culture, the latter having the greatest production rate (206 +/- 74.5 pg/mg tissue/24 h, n=6; mean +/- SEM). Production was not responsive to stimulation by pro-inflammatory cytokines. MIC-1 was detectable in 217 transabdominal amniotic fluid (AF) samples taken from 15 to 41 weeks gestation, concentrations ranging from 0.9-51.1 ng/ml. AF MIC-1 concentrations in pregnancies with premature rupture of membranes (PROM) or preterm labour, either with or without microbial invasion of the amniotic cavity, were not significantly different from those delivered at term either with or without labour. Treatment with MIC-1 (0.25-25 ng/ml) did not alter production of interleukin-6 or -8 by amnion or choriodecidual cells in vitro. We conclude that AF MIC-1 is derived from the fetal membranes and decidua, but that MIC-1 is unlikely to be involved in the pathophysiology of preterm birth or PROM.     

未培养羊水荧光原位杂交快速检测60例胎儿常见染色体数目异常

目的 应用细菌人工染色体(bacterial artificial chromosome,BAC)克隆自行制备荧光探针,对胎儿常见染色体数目异常(13,18,21,X,Y)行快速产前诊断.方法 利用中国医学遗传学国家重点实验室BAC库中相应染色体特异位点克隆,自行制备荧光探针.经外周血淋巴细胞染色体杂交验证后,用于胎儿未培养羊水的快速荧光原位杂交fluorescence in situ hybridization,FISH)检测,已检测60例羊水标本.结果 所有探针特异性均为100%,杂交成功率为97.86%;FISH结果与常规核型分析一致,检出21三体2例,18三体1例.有两例涉及其它染色体的结构异常未能检出.结论 自制探针用于未培养羊水快速FISH,使用标本量少、快速、简便,可有效检出上述5种染色体的数目异常,但本法不能检出其他染色体的数目异常和结构异常,其应用仍有一定局限性.