蛋白芯片联合SELDI-TOF-MS在卵巢癌诊断中的应用

目的用蛋白芯片联合表面增强激光解析离子化/飞行时间质谱（SELDI-TOF-MS）分析卵巢癌与对照血清中的差异表达蛋白来筛选与卵巢癌早期诊断和预后相关的肿瘤标志物。方法采用联合SELDI-TOF-MS及WXC2蛋白质芯片,对21例卵巢癌患者（实验组）及24例健康妇女（对照组）血清标本进行检测。结果在相对分子质量0~50 000 Da范围内,检测出69个差异蛋白峰。卵巢癌差异表达明显（P〈0.01）的蛋白峰8个,其中高表达的波峰3个,质荷比分别为5 927.6,8 719.3和11 729.8 Da,低表达的波峰5个,质荷比分别为4 073.8,4 158.4,4 186.7,4 290.2和8 162.5 Da。其中11 729.8 Da蛋白峰已经鉴定为血清淀粉样蛋白A1（SAA1）。结论SELDI-TOF-MS联合蛋白质芯片能够直接检测出卵巢癌患者血清中相对特异的蛋白峰,对于卵巢癌的诊断具有一定的临床意义。     

鳞癌与腺癌患者血清差异蛋白质组分析

目的为鉴定鳞癌与腺癌的血清标志物和病理分型提供依据。方法采用表面增强激光解吸电离飞行时间质谱（SELDI-TOF-MS）技术及其配套的金芯片（Gold Chip）对254份血清（鳞癌120份、腺癌82份,健康人52份）进行蛋白质谱指纹图分析,筛选鳞癌与腺癌的差异蛋白。结果 254份血清共筛选出83个差异表达的蛋白,其中5个差异表达最明显的蛋白质峰的质荷比（m/z）分别为2953、3285、6625、6646、9257。对5个峰强度分别进行F检验,P均〈0.05。结论鳞癌、腺癌患者血清具有明显表达差异的特征蛋白,为癌症的诊断和病理分型研究提供了新思路。     

用飞行质谱技术筛选肝棘球蚴病患者血清蛋白质组学标记物

目的 筛选肝棘球蚴病患者血清蛋白质组学标记物,建立血清蛋白质指纹图诊断模型,评价其应用价值.方法 肝棘球蚴病患者68例,对照组73例(其他肝病患者33例,健康人40例),将全部样品分为训练组(37例)和测试组(67例).用弱阳离子交换蛋白芯片结合表面增强激光解析电离飞行时间质谱(SELDI-TOF-MS)技术检测肝棘球蚴病患者和对照者的血清蛋白质谱.分别比较训练组中37例肝棘球蚴患者和37例对照者、5例肝囊型棘球蚴病(HCE)患者和5例肝泡球蚴病(HAE)患者、8例肝棘球蚴病患者手术前后两组血清蛋白质峰的差异.利用ZJU-PDAS软件进行数据处理,通过支持向量机(SVM)运算建立蛋白质指纹图诊断模型,并用测试组标本采用盲法对模型的灵敏度和特异度进行验证.结果 肝棘球蚴病组和对照组共筛选出9个差异蛋白峰,其中相对分子质量为1044、1047、1073、1075、1338、6453、6649、8714 m/z的8个蛋白质峰在病例组中下调,相对分子质量为5651 m/z的蛋白峰在病例组中上调(P<0.05),盲法验证表明所建模型对肝棘球蚴病诊断的灵敏度为77.4%(24/31),特异度为66.7%(24/36);阳性预测值为66.7%(24/36),阴性预测值为77.4%(24/31).HCE和HAE患者之间筛选出2个差异表达蛋白峰,质荷比分别为8716和2751 m/z(P<0.05);8例肝棘球蚴病患者手术前后两组共筛选出6个差异表达蛋白质峰,相对分子质量分别为1297、1505、1525、1534、5921,5941 m/z(P<0.05).结论 SELDI-TOF-MS技术结合生物信息学方法能筛选出肝棘球蚴病患者血清蛋白质组学标记物,对肝棘球蚴病诊断及预后判断具有潜在的应用价值.     

尿蛋白标志物模型诊断糖尿病肾病的初步研究

目的探索基于Au蛋白芯片的尿蛋白标志物模型早期快速诊断糖尿病肾病（DN）的应用价值。方法应用表面增强激光解析电离飞行时间质谱（SELDI-TOF-MS）技术及Au芯片检测86例DN患者、40例糖尿病（DM）患者及55名健康人（对照组）尿蛋白质谱。获得的谱图用Biomaker Wizard 3．1软件筛选差异蛋白,通过BPS软件建立决策树辨别分析模型,评价其临床诊断价值。部分筛选的差异蛋白通过比对标准蛋白质谱数据进行鉴定。结果DN患者与对照组尿差异蛋白质峰有37个（P〈0．05）,BPS筛选66916质荷比（m／z）蛋白建立的模型诊断DN敏感度100％,特异度97．78％。对DM和DN患者尿蛋白质谱图分析后得到24个差异蛋白质峰（P〈0．05）,BPS筛选4008、11619、66916m／z蛋白建立的模型区分两者的灵敏度和特异度均为100％。比对标准蛋白质谱数据,DN患者尿中11619、23529、66916、79378m／z蛋白,可能分别为β2-微球蛋白、α1-微球蛋白、自蛋白、转铁蛋白。结论利用SELDI及Au芯片检测尿标志蛋白在鉴别蛋白尿来源、DN快速诊断及肾损害评估中具有重要应用价值。     

儿童双侧肾母细胞瘤差异性表达蛋白的筛选

目的采用表面增强激光解析电离飞行时间质谱(SELDI-TOF-MS)技术筛选双侧肾母细胞瘤(BWT)的差异性表达蛋白。方法选取BWT患儿血清标本10例(BWT组),另选单侧肾母细胞瘤(UWT)患儿血清标本(UWT组)及健康儿童血清标本(对照组)各20例,采用SELDI-TOF-MS技术检测并收集相关数据。结果应用生物信息学方法对所得数据进行分析,得到差异性峰10个(P<0.01)。对比各组差异性表达蛋白峰值数据,筛选出质子数/电荷数(m/z)为5 648 Da的差异性表达蛋白1个,其在BWT组、UWT组及对照组中表达强度分别为3 889.36±1 796.83、2 886.81±1 404.65、432.21±730.42,三组相比,P均<0.01。结论采用SELDI-TOF-MS技术成功筛选出一种BWT差异性表达蛋白,其有望成为BWT早期诊断及预后判断的新的标志物。     

SELDI-TOF-MS联合蛋白质组学实验数据库对结直肠癌血清标志物的鉴定

目的:探讨表面增强激光解析离子化飞行时间质谱仪(SELDI-TOF-MS)联合蛋白质组学实验数据库方法研究结直肠癌患者血清标志物的可行性.方法:应用SELDI-TOF-MS对169例结直肠癌患者与83例正常人血清进行了蛋白质谱检测.通过Biomarker WizardTM与BiomarkerPatternsTM软件分析发现差异蛋白,建立结直肠癌诊断模型;用蛋白质组学实验数据库检索鉴定出有意义的差异蛋白,并用ELISA法对结直肠癌患者及正常人血清样本中差异蛋白表达进行验证.结果:发现34种差异蛋白,以质荷比(m/z)为2873,3163,6121,7778的4种差异蛋白组成的分类树诊断模型.在学习模式下其诊断结直肠癌的灵敏度为91.57%,特异度为90.53%:在测试模式下分别为82.25%.80.72%.其中m/z为7778的蛋白峰强度在结直肠癌患者血清中明显高于正常人(p<0.01),数据库检索结果提示该蛋白为血小板因子4(PF4).ELISA法测定结直肠癌患者血清PF4水平显著高于正常人(p<0.01),其灵敏度及特异度分别为61.1%,76.9%.结论:SELDI-TOF-MS联合蛋白质组学实验数据库的方法简便可行.可为蛋白芯片研究提供新思路;PF4(m/z=7778)在结直肠癌患者血清中明显升高,是结直肠癌患者新的血清学标志物.     

原发性胆汁性肝硬化血清蛋白质谱的初步研究

目的筛选与原发性胆汁性肝硬化（PBC）相关的血清蛋白质分子标志物并建立指纹诊断模型。方法收集30例PBC患者血清,30例年龄、性别相匹配正常人,采用弱阳离子磁珠（MB-WCX）为检测介质,基质辅助激光解吸电离质谱（MADIL-TOF-MS）检测得到蛋白质谱,再应用基于磁珠的多维纳升级高效液相色谱及液体蛋白芯片指纹图谱（CLINPROT）系统比较两组血清蛋白质谱。结果在m/z100～10000范围内,检测出的129个蛋白峰中11个蛋白峰差异有统计学意义（P〈0.05）,其中m/z为4963.6、5805.33、1544.45、2292.16、2466.45、2845.85在PBC中表达上调,而m/z为3262.24、3191.2、4090.5、4072.43、3277.46的蛋白峰在PBC中下调;选择m/z为3262.24、3191.2这2个蛋白峰建立的蛋白芯片指纹图谱（CLINPROT）系统对PBC诊断的敏感性为90%,特异性为95%。结论筛选出的11个蛋白分子标志物可能与PBC有关,其中m/z4963在PBC中表达最明显;蛋白芯片指纹图谱（CLINPROT）系统可能对PBC的诊断具有一定临床意义。     

蛋白质指纹图谱技术在肺结核、肺癌鉴别诊断中的应用研究

目的 探索应用表面增强激光解析离子化飞行时间质谱（SELDI-TOF-MS）技术于肺结核、肺癌的鉴别诊断。 方法 肺结核、肺癌患者及正常人各65例,收集其血清标本,采用WCX2芯片技术对血清蛋白进行捕获,用蛋白芯片阅读器PBSⅡ对芯片进行扫描、分析。 结果65例活动性肺结核与65例肺癌患者血清蛋白质谱数据的比较,4个蛋白峰(5 335 m/z、8 048 m/z、11 700 m/z、11 683 m/z)倾向于肺结核,以此鉴别肺癌差异有统计学意义(P<0.01)。该诊断模型判别的总准确率为74.6%（97/130）,灵敏度80.0%(52/65),特异度69.2%(45/65)。 结论 此方法简便、快速,标本用量少,为肺结核、肺癌患者提供了1种新的无创诊断和鉴别诊断方法 。     

用蛋白质芯片技术筛选非小细胞肺癌患者血清中标志蛋白

目的 探讨用蛋白质芯片技术检测血清非小细胞肺癌（NSCLC）标志蛋白筛查肺癌患者的可行性。方法 用蛋白质芯片表面增强激光解吸电离飞行时间质谱仪（SELDI-TOF-MS）技术检测123例肺癌患者和40名正常人血清蛋白质质谱。用数字表法随机抽取94份标本（53例NSCLC，21例小细胞肺癌和20名正常人）作为训练组进行系统训练，将筛选出来的相对分子质量为11493、6429、8245、5336及2536的5个蛋白峰作为一个标志物组合模式，建立分类树模型（即系统训练过程）；用69份未知血清标本（49例NSCLC，20名正常人）作为盲筛组验证该模型。结果 系统显示，在训练组该模式检测NCLC的敏感性和特异性分别为95．9％（71／74）、90．0％（18／20），盲筛组分别为83．7％（41／49）及80．0％（16／20）。结论 蛋白质芯片SELDI-TOF-MS技术能较准确的区分NSCLC患者与健康对照者，该技术为NSCLC的筛查提供了新的有效工具。     

应用SELDI-TOF-MS技术建立肝癌筛选血清蛋白质指纹图谱模型

目的:建立肝癌筛选血清蛋白质指纹图谱模型．方法:用表面加强激光解析电离飞行时间质谱技术(SELDI-TOF-MS)及WCX2蛋白芯片获得新发肝癌、肝硬化患者和正常人血清的蛋白质指纹图谱,用计算机软件进行比较分析,建立肝癌的筛选模型．结果:肝癌患者与健康对照组血清蛋白质指纹图谱之间有5个标志蛋白(4477,8943,5181, 8617,13 761 Da)在肝癌患者血清中高表达,肝癌患者与肝硬化患者血清蛋白质指纹图谱之间2个标志蛋白(4477,13 761 Da)在肝癌患者血清中高表达,1个标志蛋白(4097 Da)在肝癌患者血清中低表达．SELDI-TOF-MS技术的特异性(60／60,100％);敏感度(18／20,90％)．分析系统筛选出4477,8943,13 761,4097 Da标志蛋白建立的肝癌诊断模型．结论:建立的血清蛋白质指纹图谱模型能够区分肝癌与非肝癌患者,SELDI-TOF-MS在肝癌的诊断及肿瘤特异性蛋白质生物标志分子的筛选等方面具有一定价值．     

The immunoneuroendocrine role of melatonin

Abstract: A tight, physiological link between the pineal gland and the immune system is emerging from a series of experimental studies. This link might reflect the evolutionary connection between self-recognition and reproduction. Pinealectomy or other experimental methods which inhibit melatonin synthesis and secretion induce a state of immunodepression which is counteracted by melatonin. In general, melatonin seems to have an immunoenhancing effect that is particularly apparent in immunodepressive states. The negative effect of acute stress or immunosuppressive pharmacological treatments on various immune parameters are counteracted by melatonin. It seems important to note that one of the main targets of melatonin is the thymus, i.e., the central organ of the immune system. The clinical use of melatonin as an immunotherapeutic agent seems promising in primary and secondary immunodeficiencies as well as in cancer immunotherapy. The immunoenhancing action of melatonin seems to be mediated by T-helper cell-derived opioid peptides as well as by lymphokines and, perhaps, by pituitary hormones. Melatonin-induced-immuno-opioids (MHO) and lymphokines imply the presence of specific binding sites or melatonin receptors on cells of the immune system. On the other hand, lymphokines such as -γ-interferon and interleukin-2 as well as thymic hormones can modulate the synthesis of melatonin in the pineal gland. The pineal gland might thus be viewed as the crux of a sophisticated immunoneuroendocrine network which functions as an unconscious, diffuse sensory organ.     

Changes in connexin43, the gap junction protein of astrocytes, during development of the rat pineal gland

Abstract: The abundance of gap junctions between rat pineal astrocytes formed by connexin43 (Cx43) was studied during development. Levels and distribution of Cx43 were measured by immunoblotting and indirect immunofluorescence, respectively. The amount of Cx43 in cells located within the gland was low until about the 7th postnatal day and increased to adult values between the 14th and 21st days postpartum. Although astrocytes, recognized by their vimentin immunoreactivity, were scarce before birth, they were abundant by the 7th postnatal day suggesting that the low levels of Cx43 found at this age corresponded to a low expression of this protein. Localization of the immunoreactivity to Cx43 and vimentin showed a close correlation, indicating that mature or immature pineal astrocytes form gap junctions made of Cx43. Since Cx43 levels attained their adult values at about the time the innervation and the functional state of the gland reached maturity (2–3 weeks after birth), it is proposed that astrocyte gap junctions are involved in the function of the adult rat pineal gland.     

Response of rat pineal melatonin to calcium, magnesium, and lithium is circadian stage dependent

Abstract: Herein we documented the response of pineal melatonin production to electrolytes known to be effective on pineal function in view of a possible circadian stage dependence. We studied the release of melatonin by perifused rat pineal glands at 2 different circadian stages corresponding to the middle of the light and dark periods, i.e., respectively, 7 and 19 HALO (Hours After Light Onset, L:D = 12:12). The initial efflux rates were, as expected, much higher in the perifusates of glands removed from rats sacrificed during the dark phase than of those removed during the light phase. After 3 hr of perifusion, melatonin release reached similar levels which were found constant up to the 8th hr of perifusion, whatever the circadian stage. Perifusion of the glands with physiological concentrations for the rat of calcium (5.2 mmol/1) and magnesium (1.34 mmol/1) resulted in a stimulatory effect on the pineal glands removed from rats sacrificed in the middle of the dark period (19 HALO), whereas no effects were observed on the pineal glands removed from rats sacrificed during the light (7 HALO). Lithium (0.28 and 0.55 mmol/1) was ineffective on melatonin release in pineal glands removed 7 and 19 HALO. Our results show differences in the initial efflux rates of melatonin and in the response of perifused pineal glands to calcium and magnesium according to the circadian stage.     

Conservative management of perforated duodenal diverticulum： A case report and review of the literature

Duodenal diverticula are a relatively common condition. They are asymptomatic, unless they become complicated, with perforation being the rarest but most severe complication. Surgical treatment is the most frequently performed approach. We report the case of a patient with a perforated duodenal diverticulum, which was diagnosed early and treated conservatively with antibiotics and percutaneous drainage of secondary retroperitoneal abscesses. We suggest this method could be an acceptable option for the management of similar cases, provided that the patient is in good general condition and without septic signs.     

Presence of immunoreactive growth hormone and prolactin in the ovine pineal gland

Abstract: The use of antisera raised against bovine growth hormone (GH) and ovine prolactin (PRL) enabled the detection of related immunoreactive (ir) sequences of proteins in ovine pineal tissue. The isolation of PRL-like ir-material was accomplished using a 0.25 M ammonium sulphate (pH 5.5) extraction followed by ethanol precipitation, whereas the resulting 2.0 M ammonium sulphate (pH 7.0) precipitate contained a GH-like immunoreactivity. Gel chromatography of the GH-like immunoreactivity (Sephadex G-100) indicated the presence of several GH-like fragments ranging in the M_r range of 7,000 to 55,000. Analyses of the PRL-like ir-material found in pineal tissue on HPLC using a TSK 545-DEAE column led to the resolution into a single peak of immunoreactivity. A single peak of activity was also observed following chromatofocusing and hydrophobic interaction chromatography of the ir-peak from the TSK 545-DEAE column. The PRL-like ir-material inhibited the binding of [¹²⁵I]ovine PRL-S14 to anti-ovine PRL antibodies without showing an affinity for binding to anti-rat PRL or anti-bovine GH antibodies. Scatchard analysis of the binding of pineal PRL-like ir-material and pituitary ovine PRL-S14 to liver membranes from day-20 pregnant rats revealed similar affinity constants (K_a of 4.7 ± 0.2 × 10⁹ M^-1). In addition, the replication of Nb 2 Node rat lymphoma cells was stimulated by pineal PRL-like ir-material, an effect known to be specific for lactogenic hormones. The pineal PRL-like immunoreactivity appeared on sodium dodecyl sulfate polyacrylamide gels as a single major band of M_r 24,000. The functional status of PRL-and GH-like ir-material in the ovine pineal remains to be determined, but evidence is presented that the overall protein synthesis rate of the rat pineal responded to circulating concentrations of PRL.     

Microbiology of human immunodeficiency virus anorectal disease

PURPOSE: Individuals who are seropositive for the human immunodeficiency virus are at high risk for opportunistic infection and anorectal disorders. Little prospective information is available regarding anorectal pathogens in these patients. METHODS: One hundred sixty-three HIV-seropositive patients presented to the colorectal clinic between 1989 and 1992. Forty-seven (29 percent) patients were thought to have an infectious process and were prospectively studied using a standardized multiculture protocol. RESULTS: Mean age was 33 (range, 19–59) years. All were male; high-risk behavior accounted for 87 percent of HIV transmissions. Presenting complaints included anorectal pain (79 percent), pus per anum (28 percent), and blood per anum (26 percent). Examination revealed perianal tenderness (60 percent), condyloma (38 percent), perianal ulcers (38 percent), and anal fissures (34 percent). Sixty-six sets of cultures were performed; 28 patients had one set, 15 had two sets, and 4 had three sets. Thirty-two of these 47 patients (68 percent) had positive cultures including herpes (50 percent), cytomegalovirus (25 percent),Neisseria gonorrhoeae (16 percent), chlamydia (16 percent), acidfast bacilli (2 percent), and others (9 percent). Six of 32 patients with positive cultures had more than one organism cultured. Sixteen (50 percent) patients with positive cultures were treated medically, 8 (25 percent) were treated surgically and 8 (25 percent) were treated with both modalities. Sixty-one procedures were performed on 17 patients for condylomata. Eighteen patients had 20 procedures for abscesses, 50 percent of whom had positive cultures for other than common bowel flora; all improved. Fourteen patients underwent 33 procedures for perianal fistulas.Mycobacterium fortuitum was cultured from one patient who required 13 procedures for abscesses and fistulas. Forty-five (96 percent) patients were followed for an average of 12.5 months ±2.9 SEM (range, 1–94 months). Symptoms were improved or resolved in 22 of 32 (69 percent) patients with positive cultures and in 11 of 13 (84 percent) with negative cultures. CONCLUSIONS: Specific pathogens may often be identified in human immunodeficiency virus-seropositive patients with anorectal disorders if aggressively sought. Although patients without specific pathogens identified may be expected to improve with planned empiric treatment, positive identification allows more directed therapy.