Proteomic profiling of peritoneal rinse fluid sediment separates patients with ovarian cancer from women admitted for cesarean section. A pilot study

Abstract

Objective. There is a great need for additional approaches to discover putative biomarkers for ovarian cancer that accounts for 5% of all cancer deaths among women and is the leading cause of deaths among gynecologic malignancies in the United States. Methods. Peritoneal rinse fluid sediment from seven patients having ovarian cancer and four women admitted for cesarean section was analysed by proteomic techniques using two-dimensional gel electrophoresis and tandem mass spectrometry. Results. Sixteen protein spots were found to be significantly and highly up-regulated in samples from cancer patients as compared with the cesarean section group. Nine of the protein spots were identified as C-terminal fragments of fibrinogen-β (spot Nos. 5404 and 5408), haptoglobin-α₂ (spot Nos. 4101 and 6103), haptoglobin-β (spot Nos. 2502, 2504, 3502 and 4401) and transthyretin (spot No. 4002). Generally, several isoelectric variants were present of each protein. The protein concentration in the sediment of spot No. 506 and spot No. 5404 (identified as fibrinogen-β) was significantly (p < 0.05) and negatively correlated with survival rate of the ovarian cancer patients while for three of the spots a borderline significant (p < 0.07) negative correlation was found. Conclusions. It is suggested that further development of this method may give a basis for identifying suitable diagnostic and prognostic biomarkers for ovarian cancer.     

国人胃癌蛋白质组分析中双向凝胶电泳技术建立及优化

目的　初步建立、优化用于胃癌蛋白质组研究的双向凝胶电泳技术和计算机图像分析技术 ,提高其分辨率及重复性 ,得到一种分析胃癌蛋白质组双向凝胶电泳新方法。方法　提取胃癌组织中蛋白质 ,对以固相pH梯度为第一向双向电泳的关键因素与环节 ,如样品处理、电泳参数和凝胶浓度进行一系列优化。进一步采用双向凝胶电泳分离胃癌组和正常组总蛋白质 ,银染显色、图像分析软件分析电泳图谱。结果　重复性实验结果显示同组样品在 3次不同实验中所得蛋白质斑点数目相对标准差 (变异系数平均值 % ) :胃癌组和正常组分别为 2 3.0 0± 10 .11、2 0 .33± 9.90。变异系数范围 (% ) :胃癌组和正常组分别为 3.80～ 6 0 .10、2 .70～ 5 6 .89。同一蛋白质斑点在 3次实验中等电点、分子量的相对标准差分别为 8.76 %± 5 .14 % ,13.0 0 %± 4 .2 2 %和 10 .84 %± 9.16 %。获得了较好的分辨率和重复性的双向凝胶电泳图谱。结论　蛋白质组双向电泳的建立及优化表明适合于胃癌蛋白质组差异表达研究。     

蛋白质组学技术分析小鼠中脑能量代谢蛋白

目的：采用蛋白质组学技术鉴定小鼠中脑部位与能量代谢相关的蛋白,探讨这些蛋白与神经性疾病的关系。方法提取小鼠中脑蛋白质,采用双向电泳分离,切取蛋白点进行基质辅助激光解吸/电离飞行时间质谱(MALDI-TOF MS)鉴定。结果双向电泳技术结合 MALDI-TOF MS 成功鉴定出24个与能量代谢相关蛋白。结论建立了小鼠中脑部位蛋白质的鉴定方法,为探讨这类蛋白质在神经系统疾病中的作用奠定了基础。     

Cerebrospinal Profiling of Proteins, Lipids, Small Molecules, and Elements: Application to the Study of Migraine Pathophysiology

Background and objectives.—The fundamental reason that the migraine phenotype is experienced by sufferers and not by the more fortunate populace is not known. We believe a common biochemical pathway is involved, and are attempting to reveal this by using an objective and broad survey of molecular composition, rather than the conventional hypothesis-driven approaches that are narrowly focused and subjective.
Methods.—This article discusses the application of CSF bioanalysis in a clinical sample and summarizes preliminary findings, which are described more fully in papers currently in press.
Results.—In migraineurs during a migraine attack, compared with a migraine-free period, changes have been documented in the concentrations of specific proteins, lipids, small molecules, and elements in the CSF. The degree to which these changes are specific to migraine remains to be determined.
Conclusions.—As the CSF profiles for proteins, lipids, small molecules, and elements become better delineated in both healthy individuals and in disease states such as migraine, measurement of changes in CSF composition will provide the potential for improving diagnosis, correlating molecular events with symptoms, developing new therapeutic strategies, and enhancing the ability to monitor the results of therapy.     

Bacterial biofilms of importance to medicine and bioterrorism: proteomic techniques to identify novel vaccine components and drug targets

Biofilms are highly ordered microbial communities enmeshed in a carefully sculpted matrix designed for survival of organisms either in multi- or mono-genus/species in a specific microniche. In human disease, biofilm infections are some of the most recalcitrant to treat. Even with rigorous antibiotic regimens, some biofilms, such as those within the thick airway mucus of cystic fibrosis (CF) patients, persist throughout the course of the disease process. In this editorial, discussion will cover the utility of using advanced proteomic techniques to help identify potential weaknesses in the already impressive defensive armamentarium of biofilm bacteria. Two biofilm systems will be discussed herein, one of which is that of Pseudomonas aeruginosa biofilms within CF airway biofilms. The other is referred to as persistent ‘bioterrorist agent biofilms’ in which Francisella tularensis can grow on surfaces where environmental amoeba can phagocytose them, allowing for growth of F. tularensis within the amoebae.     

Deciphering the complexity of the cancer proteome for diagnostic applications

The proteome is the most functional component encoded in the genome, yet most features of the proteome that are deregulated in cancer cannot be predicted from genomic analysis alone. These include post-translational modifications (PTMs), sub-cellular localization, networks and circuitry, formation of complexes, and functional activity, all of which could play a role or be affected as part of tumorigenesis. Thus, there is a substantial opportunity to elucidate protein alterations in cancer and to translate knowledge into diagnostics and therapeutics. The progress made in mining the cancer proteome for diagnostic applications and the path forward are herein reviewed.     

结肠癌合并糖尿病患者围手术期应用胰岛素方式的观察

目的：探讨结肠癌合并糖尿病患者围手术期胰岛素泵控制血糖的效果。方法：将60例结肠癌合并糖尿病手术治疗患者随机分为两组,观察组35例采用胰岛素泵皮下连续注射,对照组25例采用胰岛素多次皮下注射治疗。观察两组患者空腹血糖、餐后血糖、血糖达标时间及低血糖的发生情况。结果：两组患者均有效控制血糖,比较差异无统计学意义（P〉0.05）,但观察组血糖达标时间短于对照组（P〈0.05）差异有统计学意义,且低血糖的发生少于对照组（P〈0.05）。结论：结肠癌合并糖尿病患者在手术前后通过胰岛素泵控制血糖达标效果满意。     

89株山夫登堡沙门菌脉冲场凝胶电泳分子分型及耐药性分析研究

目的 分析2005-2011年杭州市下城区89株山夫登堡沙门菌的脉冲场凝胶电泳(pulse-field gel electrophoresis,PFGE)分子型别和耐药性,建立杭州市沙门菌指纹图谱数据库。 方法 89株山夫登堡沙门菌用XbaⅠ酶切、PFGE获得电泳图谱,使用BioNumerisc统计软件聚类分析比较同源性。采用K-B法测定菌株对抗生素敏感性。 结果 以相似度85%为判定标准,89株山夫登堡沙门菌可分为10个PFGE带型,其中68株集中在一个PFGE基因型内。不同年份基因型无明显规律性。89株山夫登堡沙门菌对所选11种抗生素敏感性85.19%。 结论 杭州市下城区2005-2011年分离的89株山夫登堡沙门菌有明显的克隆株优势带型存在,对抗生素敏感性较高。     

Desmin在大肠癌与癌旁组织中的表达及意义

目的探讨Desmin在大肠癌与癌旁组织中的的表达及其意义。方法采用液质联用技术对18例新鲜的大肠癌患者的肿瘤组织及癌旁组织检测Desmin的表达情况。结果通过液质联用技术对大肠癌肿瘤组织及癌旁组织进行分离分析,对有差异的蛋白质点进行质谱鉴定,Desmin在癌组织中较癌旁组织中表达降低,并通过免疫印迹进一步验证准确性。结论 Desmin蛋白的低表达可能与大肠癌的癌变有密切的关系。     

北京地区2010-2015年宋内志贺菌脉冲场凝胶电泳分型及耐药特征分析

目的 分析北京市2010-2015年分离的259株宋内志贺菌脉冲场凝胶电泳（pulsed field gel electrophoresis,PFGE）分型及耐药特征。方法 收集2010-2015年分离到的宋内志贺菌259株,经生化及血清型鉴定后,应用PFGE分型并用最小抑菌浓度法（minimal inhibition concentration,MIC）进行12种药物的耐药检测。结果 259株宋内志贺菌经PFGE图谱聚类分析后,发现宋内志贺菌有A、B两个优势簇,2010-2015年A簇菌株所占构成比逐年增高;耐药结果显示257株为耐药菌株,其中245株为多耐药菌株,占所有耐药菌株的95.33%。对头孢曲松（CRO）的耐药性有明显升高。结论 北京地区近年宋内志贺菌有较高的同源性,A簇菌株已经成为目前北京地区宋内志贺菌的优势簇。对三代头孢类药物CRO等耐药性上升可能与临床上大量应用三代头孢药物以及A簇菌株增多有关,多耐药谱的情况也更加复杂,提示应合理使用抗生素。     

稳定性心绞痛患者血清差异蛋白质的初步分析

摘要：目的：初步分析稳定性心绞痛（SAP）患者血清中差异蛋白质的表达。 方法：收集年龄与性别相匹配的50例SAP初诊患者与50例体检健康者的血清,分别混合血清后去除IgG 和清蛋白,进行PAGE并观察去除效率;进行固相pH梯度SDS-PAGE二维凝胶电泳,对凝胶进行银染后用Image Master 5.00软件分析;对差异点用基质辅助激光解吸电离飞行时间质谱（MALDI-TOF-MS）技术进行测定。 结果：去除IgG和清蛋白前后的SDS-PAGE电泳图效果良好,IgG 和清蛋白去除率均＞80%;分析获得的2个差异点经质谱分析,间-α-胰蛋白酶抑制剂家族重链相关蛋白（inter-α-trypsin inhibitor family heavy-chain related protein,IHRP）、锌指蛋白A20（zinc finger protein A20,A20）在SAP患者血清中表达而在健康人血清中无表达。 结论：IHRP和A20可能在SAP发病机制中起一定作用。     

2014—2018年仙桃市伤寒沙门菌分子分型及耐药性分析

目的了解仙桃市伤寒沙门菌的耐药性及分子分型特征。方法收集2014—2018年分离的29株伤寒沙门菌开展药敏试验、脉冲场凝胶电泳(PFGE)分型及多位点序列分型(MLST)分析。结果药敏结果显示,29株伤寒沙门菌中27株菌对26种临床常见抗菌药物均为敏感/中度敏感,余2株菌有不同程度的耐药性,其中1株菌株耐药谱分布较广,对β-内酰胺/β-内酰胺抑制剂复合物类药物耐药。PFGE结果显示,29株伤寒沙门菌可分为6种PFGE型别,其中流行优势型为HuBXTSal003型别。MLST分型显示,29株菌株共有2种序列型,其中ST2为优势ST型。结论仙桃市伤寒沙门菌株存在优势PFGE带型及优势ST型,有多重耐药菌株出现。     

Colon cancer: Cancer stem cells markers,drug resistance and treatment

Malignant tumours consist of heterogeneous populations of tumour cells. Cancer stem cells (CSC) represent a population of cells within a tumour with highly tumorigenic and chemoresistant properties. These cells may be identified by the expression of CSC markers. There are several key stem cells markers specified for colon cancer: CD133, CD44, ALDH1, ALCAM. These days, a major obstacle to effective cancer management is development of a multidrug resistance (MDR). The principal mechanism responsible for development of MDR phenotype is the over-expression of ABC transporters. Tumours and relapsing tumours after therapy are drived by subpopulations of tumour cells with aggressive phenotype resistant to chemotherapeutics. These cells are called CSC or tumour-initiating cells (TIC). Here we outline recent information about MDR of colon cancer and CSC markers. We have focused on novel therapeutic strategies which have been developed to prevent or overcome MDR. One such strategy is a combination of chemotherapy and modulators of MDR pumps or chemotherapy and monoclonal antibodies against vascular endothelial growth factor VEGF. Colon cancer is characterized by the presence of colon CSC expressing specific stem cell markers. The divergent presence of these markers can help to adjust personalized therapy. The review provides a detailed overview of resistance of colon cancer cells and discusses how the presence of CSC markers can influence therapy and prognosis of patients.     

Placental metabolic profiling in gestational diabetes mellitus: An important role of fatty acids

AimGestational diabetes mellitus (GDM) is the most common metabolic disorder during pregnancy. Accumulating studies have reported metabolites that are significantly associated with the development of GDM. However, studies on the metabolism of placenta, the most important organ of maternal‐fetal energy and material transport, are extremely rare. This study aimed to identify and discuss the relationship between differentially expressed metabolites (DEM) and clinical parameters of the mothers and newborns.MethodsIn this study, metabolites from 63 placenta tissues (32 GDM and 31 normal controls) were assayed by ultra‐performance liquid chromatography‐high resolution mass spectrometry (UPLC‐HRMS).ResultsA total of 1297 annotated metabolites were detected, of which 87 significantly different in GDM placenta. Lipids and lipid‐like molecules accounted for 62.1% of DEM as they were significantly enriched via the “biosynthesis of unsaturated fatty acids” and “fatty acid biosynthesis” pathways. Linoleic acid and α‐linolenic acid appeared to be good biomarkers for the prediction and diagnosis of GDM. In addition, the level of PC(14:0/18:0) was negatively correlated with neonatal weight. 14 metabolites significantly different in male and female offspring, with the most increase in female newborns.ConclusionEven if maternal blood glucose level is well controlled, there are still metabolic abnormalities in GDM. Lipids and lipid‐like molecules were the main differential metabolites, especially unsaturated fatty acids.     

Separation, identification of uremic middle molecules, and preliminary study on their toxicity

BACKGROUND: Compounds accumulating in uremic serum with molecular mass from 300 to 5000 Da are called uremic middle molecules (UMMs). In our previous work, two UMM fractions A and B were obtained from uremic sera, urine, and normal urine by gel permeation chromatography (GPC), and six UMMs from subfraction A3 of uremic plasma and normal urine were purified and characterized. METHODS: Urine and serum samples from uremic patients and healthy subjects were isolated by GPC, ion exchange chromatography (IEC), and reversed-phase high-performance liquid chromatography (RP-HPLC). Moreover, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) were used to characterize the compounds. The effects of subfraction A3 on renal function were studied in rabbit models with chronic renal failure (CRF). RESULTS: A compound with molecular weight 1007.94 in subfraction A3 was determined to be an octapeptide by mass spectrometry, with an amino acid sequence of Val-Val-Arg-Gly-Cys-Thr-Trp-Trp. Two CRF rabbits injected with A3 died in 5 days, while the other two CRF rabbits (no injection) survived a few days. By multistep chromatography and MALDI-TOF MS, another 11 endogenous compounds were found not only in the subfraction B9 of uremic sera but also in that of normal urine. CONCLUSION: Seventeen endogenous middle molecular compounds were found in fractions A and B of uremic plasma and normal urine, among them an octapeptide with M(W) 1007.94 in subfraction A3. Preliminary experimental results on rabbits indicate that subfraction A3 could accelerate the death of rabbits with CRF.     

Metabolic abnormalities of apolipoprotein B-containing lipoproteins in non-insulin-dependent diabetes: a stable isotope kinetic study

BACKGROUND: Kinetic abnormalities of apolipoprotein B (apoB)-containing lipoproteins in noninsulin-dependent diabetes mellitus (NIDDM) remain poorly understood. To get further insight into these abnormalities we performed a stable isotope kinetic experiment comparing the metabolism of apoB-containing lipoproteins in moderately severe NIDDM patients and healthy control subjects. METHODS: The study was performed in the fed state. Subjects underwent a primed infusion of 0.7 mg kg(-1) of L-[1-(13)C]leucine followed by a 16-h constant infusion of 0.7 mg kg(-1) h(-1). [13C]Leucine enrichment in apoB was measured by gas chromatography/combustion/isotope ratio mass spectrometry. RESULTS: In NIDDM patients, we observed a 3.49- and 4.52-fold increase of very-low-density lipoprotein (VLDL) and intermediate-density lipoprotein (IDL) apoB plasma concentrations, respectively (P<0.01). VLDL apoB production was increased by 41% (P<0.05) and fractional catabolic rate towards IDL and low-density lipoprotein (LDL) was decreased by 61% (P<0.05). The increased IDL apoB plasma concentration was also related to a major catabolic defect (-78%; P<0.01). For most patients, plasma LDL apoB concentration was comparable to that of controls. Nevertheless, LDL apoB metabolism was impaired in NIDDM subjects, with both a decreased LDL catabolic rate (-28%; P<0.05) and a trend towards a diminished synthesis. CONCLUSION: NIDDM is associated with multiple apoB metabolism abnormalities that are potentially atherogenic. In addition to the increased number of circulating VLDL and IDL particles, the increased residence time observed on all apoB-containing lipoproteins may promote the development of atherosclerotic lesions, by potentiating their oxidizability.     

糖尿病患者泌尿道感染病原菌及耐药性分析

目的 分析我院住院糖尿病患者泌尿道感染病原菌分类及耐药性情况,为临床经验性使用抗菌药物提供参考.方法 回顾性分析2011年1月至2012年12月我院收治的217例糖尿病合并泌尿道感染患者,对其尿培养结果阳性的病原菌的分布及耐药性进行了统计分析.结果 217例住院糖尿病伴泌尿道感染患者中段尿标本中,检出病原菌237株,其中革兰阴性菌占73％,革兰阳性菌占21.1％,真菌占5.91％.其中单一菌株占90.78％,两种菌株占9.22％.革兰阴性菌对氨苄西林、复方磺胺、四环素等抗菌药物耐药率＞60.0％,革兰阳性菌对青霉素、红霉素、四环素的耐药率＞70.1％.结论 住院糖尿病伴泌尿道感染患者尿标本中检出主要病原菌为革兰阴性杆菌,其中感染菌中大肠埃希菌居首位.     

Global and comparative protein profiles of the pronotum of the southern pine beetle, Dendroctonus frontalis

The southern pine beetle (Dendroctonus frontalis Zimmermann) kills all pines within its range and is among the most important forest pest species in the US. Using a specialized mycangium surrounded by gland cells in the pronotum, adult females culture, transport, and inoculate two fungi into beetle galleries during oviposition. These fungal symbionts, to varying degrees, exclude antagonistic fungi and provide nutrients to larvae. However, the mechanisms (e.g. secreted antibiotic chemicals or nutrients, proteins or pathways) by which this relationship is maintained are not known. Here we present the first global and differential proteome profile of the southern pine beetle pronotum. Two-dimensional polyacrylamide electrophoresis, tandem mass spectrometry, and database searches revealed that the majority of pronotal proteins were related to energy-yielding metabolism, contractile apparati, cell structure, and defence. The identified proteins provide important insights into the molecular and biochemical processes of, and candidates for functional genomics to understand mycangia and pronotum functions in, the southern pine beetle.     

沙门菌监测点汤卜逊沙门菌分离株的分子分型和耐药性分析

目的应用分子分型技术对2007年上海和重庆市沙门菌监测点的50株汤卜逊沙门菌进行分子流行病学分析和抗生素敏感性测定,了解上海和重庆两地菌株的分子分型特征和药物敏感性特征。方法抗生素敏感性测定采用微量肉汤稀释法,分子分型方法包括脉冲场凝胶电泳（PFGE）和多位点串联重复序列分析（MLVA）。结果药敏试验显示78%的菌株存在多重耐药,其中磺胺甲恶唑和四环素的耐药率最高,其次是甲氧苄胺嘧啶;对头孢类抗生素（头孢噻肟、头孢三嗪、头孢他啶、头孢噻呋）均未产生耐药性。PFGE将50株菌分为15个带型,30株重庆分离株间有较高的相似性;MLVA分析显示除Sal16位点外,其余检测位点在所有待检菌株中没有差别。结论分子分型支持汤卜逊沙门菌引起的暴发以及散发。目前MLVA分型应用于汤卜逊沙门菌分子分型时,分型能力低于PFGE,需要进一步优化。     

外泌体与卵巢癌耐药关系的研究进展

目的： 探讨新型冠状病毒肺炎（COVID-19）合并糖尿病酮症酸中毒（DKA）的临床特征,提升诊疗水平。方法： 分析2020年2月10日到17日在华中科技大学同济医学院附属同济医院抢救治疗的3例COVID-19合并DKA昏迷患者的临床资料。结果： 3例患者中1例为男性,2例为女性;2例为2型糖尿病,1例可能为1型糖尿病;病程中患者随机静脉血糖均大于30 mmol/L,血气分析示pH值均小于7.0;1例心肌酶谱显著升高,考虑心肌炎可能;1例血胰腺淀粉酶和脂肪酶显著升高,胰腺损伤严重;2例患者炎性因子（白介素、肿瘤坏死因子）等显著升高;3例患者均出现了凝血功能障碍。3例患者出现DKA后3~4 d病情迅速恶化,其中2例出现心跳骤停,1例经ICU抢救后病情平稳转入普通病房继续治疗。结论： COVID-19诊治期间要积极预防诱因,避免DKA昏迷发生;DKA一旦发生常病情极危重,可合并胰腺、心脏、肝脏等多脏器的功能损害,出现不良结局。