基于触珠蛋白变化的白血病干细胞移植治疗疗效初探

目的探讨不同的干细胞移植法对白血病患者血浆触珠蛋白(Hp)变化的影响。方法以4例进行间充质干细胞和造血干细胞共移植(MSC/HSCT)的患者和2例采用造血干细胞移植(HSCT)的患者为研究对象,运用双向凝胶电泳(2-DE)对其不同治疗阶段的血浆中的触珠蛋白进行分离,利用MALDI-TOF/TOF MS进行鉴定。结果治疗过程中触珠蛋白β链和α链的血浆含量在移植后呈现明显的递减趋势,并伴有多种翻译后修饰,这种变化趋势在MSC/HSCT治疗的患者中更为显著。结论触珠蛋白有可能成为白血病干细胞移植治疗疗效评价及疗程监测的标志物。     

细胞表面糖链与肿瘤干细胞

近年来,肿瘤治疗手段虽有了很多新的进展,但如何应对复发和转移仍是肿瘤治疗的棘手问题.在白血病及多种实体肿瘤如乳腺癌、结肠癌、肺癌等中都存在数量极少的具有干细胞性质的肿瘤细胞,这类细胞被称为肿瘤干细胞或肿瘤起始细胞.肿瘤干细胞能够抵抗常规的放射和化学治疗,被认为是造成肿瘤复发和转移的主要原因.肿瘤干细胞的表面标志分子用来分离鉴定肿瘤干细胞及作为治疗靶点具有极大潜力,目前被广泛认可的标志物有CD44、CD133等,它们在正常组织干细胞中表达,如人造血干细胞、神经干细胞等,也可以用来分离鉴定一些实体肿瘤中的干细胞[1].选择性地去除肿瘤干细胞而对正常干细胞不产生明显毒性是治愈肿瘤的关键,但目前并没有获得理想的针对肿瘤干细胞的特异性标志物.由于必要的糖转移酶的表达缺陷,肿瘤细胞表面糖链的组成及结构明显改变.新近发现,一些糖链表达于肿瘤干细胞表面,这些糖链对糖生物学基础研究、发现和寻找肿瘤干细胞新的特异性标志物,以及肿瘤的临床诊断和治疗具有积极意义.     

慢性粒细胞白血病病人TCRζ链表达特点

目的:建立实时定量PCR方法检测TCRζ链表达水平的方法,了解慢性粒细胞白血病(CML)外周血TCRζ链表达水平.方法:采用SYBR Green Ⅰ荧光定量PCR,相对定量检测30例CML患者和30例正常人外周血的单个核细胞的TCRζ链表达情况,以β2微球蛋白基因(β2M)作为内参,根据相对定量公式:2-ΔΔCt计算CML病人与正常人TCRζ链表达差异倍数.结果:成功建立SYBR Green荧光实时定量PCR检测TCRζ链表达检测技术.18例CML患者TCRζ链出现表达低于正常人,而12例CML患者TCRζ链出现表达高于正常人.结论:CML病人中TCRζ链表达水平可分为表达下调(60%)和表达上调(40%)2组,提示部分CML病人的细胞免疫缺陷可能与其TCRζ链表达下调有关.     

β-地中海贫血基因治疗的研究进展

β-地中海贫血基因治疗的策略和方法,根据靶向的不同可以分为3类:导入正常的β-珠蛋白基因、导入正常的γ-珠蛋白基因和抑制α-珠蛋白基因的表达.β-地中海贫血动物模型的发展,对疾病基因治疗的进步有重要意义.基因治疗载体系统是关系到基因治疗成败的重要因素之一,其中慢病毒载体的采用是一重要进展.     

CCAAT/增强子结合蛋白β在系统性红斑狼疮患者外周血单个核细胞中的表达及意义

目的探讨系统性红斑狼疮(systemic lupus erythematosus,SLE)患者外周血单个核细胞(peripheral blood mononuclear cell,PBMC)中CCAAT/增强子结合蛋白(CCAAT/enhancer binding proteinsβ,C/EBPβ)的表达及其意义。方法从SLE患者血液中分离血浆和PBMC。通过定量PCR(quantitative PCR,q PCR)和Western blot方法检测SLE患者和健康对照PBMC中C/EBPβm RNA及其功能活性形式LAP(liver-enriched activating protein)蛋白表达水平。诱导THP-1细胞成为单核巨噬细胞,再分别用SLE患者血浆和健康对照血浆刺激24 h后,通过q PCR和Western blot方法检测C/EBPβm RNA和LAP蛋白表达水平。结果 SLE患者外周血PBMC中C/EBPβm RNA及LAP蛋白水平高于正常对照(P0.05)。SLE患者外周血PBMC中C/EBPβm RNA表达水平及LAP蛋白灰度值与SLE疾病严重程度评分(SLEDAI评分)正相关(P0.05,r0)。与正常对照血浆相比,SLE患者血浆刺激THP-1细胞诱导成的单核巨噬细胞24 h后C/EBPβm RNA和LAP蛋白水平显著升高(P0.05)。结论 SLE患者外周血PBMC中C/EBPβ表达显著升高,尤其是在单核巨噬细胞中,提示C/EBPβ表达升高参与SLE炎症发生的过程。     

肺癌与肺良性疾病 特异血清免疫炎性蛋白质N-糖基化修饰差异

目的探索肺癌与肺良性疾病特异血清免疫炎性蛋白质N-糖基化修饰差异。方法应用非变性聚丙烯酰胺凝胶电泳(native-PAGE)从肺腺癌和慢性肺炎患者血清中分离得到一组疾病特异的血清免疫炎性蛋白质复合物(IIRPCs),经胶内酶解、石墨相氮化碳富集分离得到完整糖肽。应用高效液相色谱-串联质谱(HPLC-MS/MS)技术,结合GPSeeker数据库检索软件进行糖肽鉴定。结果共鉴定了来自12种蛋白质的89种糖肽,其中21种仅与肺良性疾病相关,38种仅与肺癌相关。89种糖肽中包括63种糖型,其中10种为肺良性疾病特有糖型,21种为肺癌特有糖型。两种疾病的特有糖型在其结构和连接位点上均存在差异。结论发现了两种疾病血清特异免疫炎性蛋白质N-糖基化修饰的差异,为糖基化修饰在慢性疾病的诊断、预后评估和分子机制等方面的研究提供了新视角。     

N-糖链合成相关β-1,4-半乳糖基转移酶在大鼠坐骨神经损伤后的表达变化

为了分析与 N-糖链合成相关β-1,4-半乳糖基转移酶 - 、 、 (β-1,4-galactosyltransferase , , ,β-1,4-Gal T- , , ) m RNA在正常和损伤坐骨神经组织的表达变化。本研究采用 RT-PCR方法 ,分析 β-1,4-Gal T- 、 、 在小鼠坐骨神经中的表达存在。以扩增的 c DNA片断标记探针 ,Northern blot分析β-1,4-Gal T- 、 、 m RNA在正常和损伤大鼠坐骨神经的表达变化。结果表明 :通过 RT-PCR方法 ,检测到 β-1,4-Gal T- 、 、 在小鼠坐骨神经中的表达存在 ;采用 Northern blot方法 ,发现β-1,4-Gal T- 、 、 在正常大鼠坐骨神经中有表达 ,并在坐骨神经损伤后发生表达变化 ,但三种同功酶的表达变化各不相同。结论 :本实验发现 β-1,4-Gal T- 、 、 在坐骨神经有表达 ,并在坐骨神经损伤后发生表达变化。提示与 N-糖链合成相关的蛋白修饰调节在周围神经损伤后再生和退变过程中可能发挥一定的作用     

卡格列净联合度拉糖肽对老年糖尿病肾病患者血糖水平及肾功能的影响

目的:研究卡格列净联合度拉糖肽治疗对老年糖尿病肾病患者血糖及肾功能的影响.方法:将我院 2021 年1 月至 2022 年8 月收治的82例老年糖尿病肾病患者,随机分为对照组、观察组(n=41),分别采用度拉糖肽(皮下注射,1.5 mg·次-1,1 次·w-1)以及卡格列净(口服,100 mg·次-1,Qd)联合度拉糖肽治疗.治疗 3 m后比较两组临床疗效,采用葡萄糖氧化酶法检测空腹血糖(Fasting blood glucose,FBG)、餐后 2 h血糖(Postprandial blood glucose 2 hour,2hPBG)水平,采用比浊法检测 24 h尿蛋白,采用全自动生化分析仪检测血肌酐(Serum creatinine,Scr)、尿素氮(Urea nitrogen,BUN)水平,采用双抗体夹心酶联免疫法检测血清转化生长因子-β1(Recombinant transforming growth factor beta 1,TGF-β1)、长正五聚蛋白 3(Pentraxin 3,Ptx3)、α-klotho蛋白水平,采用放射免疫分析法测定肿瘤坏死因子-α(Tumor necrosis factor-α,TNF-α)水平、不良反应发生率.结果:治疗 3 m后,观察组临床总有效率高于对照组,血糖水平、肾功能指标、血清TGF-β1、TNF-α、Ptx3 水平低于对照组,α-klotho蛋白水平高于对照组(P＜0.05);两组不良反应发生率对比,无显著差异(P＞0.05).结论:卡格列净联合度拉糖肽治疗老年糖尿病肾病患者疗效显著,可有效控制血糖,改善肾功能,促进病情恢复,且安全可行.     

食管癌患者手术治疗前后血清TGF-α、TGF-β_1和血浆VEGF检测的临床意义

目的：探讨了食管癌患者手术治疗前后血清TGF-α、TGF-β1和血浆VEGF水平的变化及意义。方法：应用放射免疫分析和酶免法对33例食管癌患者进行了手术治疗前后血清TGF-α、TGF-β1和血浆VEGF测定,并与35名正常健康人作比较。结果：手术前食管癌患者血清TGF-α、TGF-β1和血浆VEGF水平非常显著地高于正常人组（P〈0.01）,经手术治疗6个月则与正常人组比较无显著性差异（P〉0.05）。结论：检测食管癌患者血清中TGF-α、TGF-β和血浆VEGF水平的变化对观察病情和预后判定具有重要的临床价值。     

急性白血病患者治疗前后血清TNF-α和血浆VEGF检测的临床意义

目的：探讨急性白血病患者治疗前后血清TNF-α和血浆VEGF水平的变化及临床意义。方法：分别应用放射免疫分析和酶联免疫法对31例急性白血病患者进行了治疗前后血清TNF-α和血浆VEGF检测,并以30名正常健康人做比较。结果：急性白血病患者治疗前血清TNF-α和血浆VEGF水平显著高于正常人组（P〈0.01）,经化疗和中西医结合治疗后6个月与正常人组比较仍有显著性差异（P〈0.05）。结论：急性白血病的发生发展与血清TNF-α和血浆VEGF水平密切相关。     

Pattern of glycosylation of Sindbis virus envelope proteins synthesized in hamster and chicken cells

The tryptic glycopeptides of the Sindbis virus envelope glycoproteins E1 and E2 grown in BHK and chick cells were purified by gel filtration followed by high-pressure liquid chromatography. Each of the purified glycopeptides was analyzed by N-terminal sequencing to identify from which of the potential glycosylation sites it was derived. The type of oligosaccharide chain attached to each glycopeptide was determined from gel filtration analysis of the pronase-digested glycopeptides, and the relative incorporation of radiolabeled galactose, mannose, and glucosamine into each glycopeptide was used to confirm these determinations. The glycosylation patterns for the two proteins were essentially identical in the two host cells. The E2 glycosylation sites at Asn196 and Asn318 contained exclusively complex-type and simple-type oligosaccharide chains, respectively. In E1, the glycosylation site at Asn139 contained only complex-type chains, but the site at Asn245 contained a mixture of simple (75-85%) and complex (15-25%) type chains. These results are discussed in relation to previously reported results and a prediction as to the relative importance of the different glycosylation sites to the function of the proteins is made.     

Detection of an outbreak of methicillin-resistant Staphylococcus aureus with reduced susceptibility to glycopeptides in a French hospital

Staphylococcus aureus isolates were screened for reduced susceptibility to glycopeptides with an initial glycopeptide agar screening test, followed by confirmation of the strains thus identified by two Etest strip techniques and population analysis. This procedure detected 48 methicillin-resistant S. aureus (MRSA) isolates with reduced susceptibility to glycopeptides from 24 patients among 883 MRSA isolates tested. The dissemination of a single clone was confirmed by pulsed-field gel electrophoresis.     

Affinity chromatography of branched oligosaccharides in rat liver beta-glucuronidase

Rat liver microsomal and lysosomal beta-glucuronidase-derived glycopeptides were obtained by extensive Pronase digestion followed by N-[14C]acetylation and desialylation by neuraminidase treatment. These glycopeptides were studied by sequential chromatography on lectin-affinity columns such as concanavalin A, lentil lectin, Phaseolus vulgaris erythroagglutinin, Ricinus communis agglutinin I, Triticum vulgaris agglutinin, Glycine max agglutinin and Ulex europaeus agglutinin. Using serial lectin affinity chromatography approach combined with neuraminidase treatment allowed us to show the unexpected presence of complex tri- and/or tetraantennary type glycans (40.8 and 17.0% for microsomal and lysosomal enzyme, respectively). Moreover, the application of neuraminidase treatment revealed that complex biantennary type glycans, present on lysosomal beta-glucuronidase, are almost fully sialylated while the same type of glycans present on microsomal enzyme do not contain sialic acid. Furthermore, the results obtained confirmed that microsomal and lysosomal beta-glucuronidases possess high mannose and/or hybrid type glycans (19.6 and 36.6%, respectively), and complex biantennary type glycans (38.9 and 46.4%, respectively).     

Ostra białaczka o mieszanym fenotypie – jak rozpoznać, jak leczyć?

In 2009, the World Health Organization has published an updated classification of proliferative diseases, whereby biphenotypic acute leukemia and acute bilineal leukemia, which previously were two separate diseases, were replaced by a single name, mixed phenotype acute leukemia (MPAL). Incidence of MPAL is not exactly known and varies in different publications from 0.5 to 5% of all leukemias in children. In these patients genetic disorders are particularly important for both diagnosis and prognosis. However, due to the still unsatisfactory outcome, the main problem remains optimal therapeutic strategy of patients with MPAL. The majority of research points to better outcomes of MPAL treatment with usage of therapeutic programs for the treatment of acute lymphoblastic leukemia (ALL), although the results are worse than in patients with ALL. An attempt to standardize the therapeutic approach and further improve is the project iBFM AMBI2012.     

Detection of new mutations conferring resistance to linezolid in glycopeptide-intermediate susceptibility Staphylococcus hominis subspecies hominis circulating in an intensive care unit

Glycopeptides and linezolid are the most widely used antibiotics to treat infections by methicillin-resistant Staphylococcus spp. We report the presence of various isolates of methicillin-resistant S. hominis subsp. hominis with resistance to linezolid and reduced susceptibility to glycopeptides. We studied ten blood culture isolates of S. hominis subsp. hominis from nine patients admitted to our hospital. Etest was used to study susceptibility to antibiotics commonly prescribed against staphylococci. Domain V region of the 23S rRNA gene was amplified and sequenced to detect possible mutations that confer resistance to linezolid. Pulsed-field gel electrophoresis (PFGE) was used for the clonality study of isolates. All isolates were resistant to oxacillin, gentamicin, levofloxacin, cotrimoxazole, and linezolid, and susceptible to tigecycline and daptomycin. Nine of the isolates were resistant to erythromycin and clindamycin, and showed heterogeneous resistance to glycopeptides. C2190T, G2603T, and G2474T mutations were detected in domain V of the 23S rRNA gene. PFGE showed the presence of two different clones. This report alerts to the possible appearance of clinical strains of methicillin-resistant staphylococci with intermediate resistance to glycopeptides, resistance to linezolid, and multiple resistance to other second-line antibiotics.     

The large sialoglycoprotein of human lymphocytes. I. Distribution on T and B lineage cells as revealed by a monospecific chicken antibody

Chickens were immunized with highly purified large sialoglycoprotein of human lymphocytes (L-LSGP; gp 150) which was isolated from neuraminidase-treated normal peripheral blood lymphocytes by affinity chromatography to HP-Sepharose and further purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Antibodies isolated from plasma and egg yolk were highly specific for desialylated L-LSGP (apparent molecular mass approximately 150 kDa). The antigenic sites recognized by the antibodies are probably located in the peptide moiety of the molecule since antibody binding to lymphocytes was not inhibited by a variety of different sugars or abrogated by absorption on various erythrocytes. In immunofluorescence experiments, greater than or equal to 75% of neuraminidase-treated thymocytes and peripheral blood lymphocytes, virtually all E+ cells and T4+ or T8+ T chronic lymphocytic leukemia (CLL) cells were stained by anti-gp 150. A small fraction (approximately 10%) of thymocytes and a larger fraction (greater than or equal to 30%) of T CLL cells in some patients were stained before neuraminidase treatment. Thymocytes appear to contain considerably lower amounts of a less sialylated form of L-LSGP than peripheral blood lymphocytes. In contrast to blast cells of 5-day concanavalin A or leucoagglutinin cultures (greater than or equal to 90% anti-gp150+) only about 50% of the blast cells generated in 5-day mixed leukocyte cultures were anti-gp150+. The large majority (greater than or equal to 75%) of both the anti-gp150+ and anti-gp150- cells were T3+ and T11+.(ABSTRACT TRUNCATED AT 250 WORDS)     

Demonstration and characterization of polyoma-tumor-associated antigenic components using preparative and analytical polyacrylamide gel electrophoresis

Glycoproteins from surface of polyoma tumor cells extracted with 3M KCl were separated in preparative electrophoresis and elution profile showed eight fractions, which were further separated by polyacrylamide gel electrophoresis and SDS-PAGE. Twenty polypeptides were detected, their molecular weights were in range 10000-155000 but most of them had Mr 17000-68000 and isoelectric points ranged between 3.9 and 5.5. Results of immunodiffusion after absorption of immune sera suggested that tumor associated antigens were present in fraction II in which four glycopeptides of molecular weights 14,000, 67,000, 36,000, 18,000 were found. These polypeptides had isoelectric points 5.3, 5.9, 6.4 and 6.6 respectively. Antigens of H-2 system were detected in fraction IV, which contained three glycopeptides of molecular weights 18000, 37000 and 48000 and isoelectric points 6.0, 5.8, and 6.7 respectively.     

The carbohydrates of influenza virus

Summary Two carbohydrate fractions can be obtained from egg-grown influenza virus after Pronase digestion followed by gel chromatography. One fraction contains glycopeptides (MW ca. 2000–2600) which represent the carbohydrate side chains of the viral glycoproteins. The constituent sugars of this material are fucose, galactose, glucosamine, and mannose, and their position within the side chain has been partially elucidated by methylation studies. The other fraction (MW>6000) appears to be mucopolysaccharide and is composed of fucose, galactose, glucose, galactosamine, and glucosamine.With 1 Figure     

Separation of A- and B-type Glycopeptides of Semliki forest virus by concanavalin A affinity chromatography and preliminary characterization of the B-type glycopeptides

A- and B-type glycopeptides of Semliki forest virus were separated by affinity chromatography on concanavalin A (Con A)-Sepharose. The A-type glycopeptides were identified by their high galactose content and the B-type glycopeptides, by their reactivity with α-mannosidase and endo-β-N-acetylglucosaminidase H. The purity and the nature of the separated glycopeptides can also be established by gel filtration because the viral A- and B-type glycopeptides are of different size. The degradative experiments suggested that the glycans of the B-type glycopeptides contained, on the average, four to six distally located α-mannose residues, probably one centrally located β-mannose residue, and two vicinal N-acetylglucosamine residues close to the peptide part of the molecule.