唾液蛋白质组学的研究进展

唾液作为一种成分复杂,在口腔内具有多种生物学功能的液体.它起到润滑、辅助消化、抗菌等诸多作用,而对唾液的功能和其分泌的分子机制的研究离不开对唾液蛋白质的研究.蛋白质组学作为大规模蛋白质分离和识别技术研究蛋白质组的一门学科,已经开始在很多领域得到应用,同时正逐渐成为唾液蛋白组研究中的新技术.本文就蛋白质组学在唾液中的蛋白全表达、生物标记应用等方面内容作一综述.     

牙齿发育早期间充质细胞的差异蛋白分析

目的:应用蛋白质组学方法分析间充质细胞在牙齿发育早期不同阶段的蛋白质差异表达情况,从蛋白水平进一步了解牙齿发育早期的分子机制.方法:制备C57胎鼠起始期(E11.5)、蕾状期(E13.5)冰冻切片,通过激光捕获显微切割技术(LCM)获取两个时期下颌第一磨牙牙胚间充质细胞,采用双向凝胶电泳方法分离蛋白质,比较分析凝胶图像,通过质谱及生物信息学分析鉴定差异蛋白.结果:激光捕获了较纯的目的细胞,得到双向凝胶电泳图谱,比较分析发现E13.5组8个蛋白点明显上调,E11.5组5个蛋白点发生明显上调.质谱分析初步鉴定了蛋白质种类.结论:LCM技术能较好地获取较为均一的目的细胞,并发现牙胚发育起始期与蕾状期间充质细胞蛋白质表达存在差异,为牙胚早期发生的分子机制研究提供一种新的方法.     

蛋白质组学及其在口腔医学中的应用

蛋白质组学是对一个基因组、一种生物、一种细胞/组织所表达的全部蛋白质及其相互作用的研究,蛋白质组的研究为口腔疾病病原学、口颌系统发育、口腔肿瘤转移和早期诊断等研究提供了新的技术平台。本文综述了蛋白质组学在口腔医学领域应用的研究进展。     

蛋白质组学及其在口腔医学中的应用

蛋白质组学是对一个基因组、一种生物、一种细胞／组织所表达的全部蛋白质及其相互作用的研究，蛋白质组的研究为口腔疾病病原学、口颌系统发育、口腔肿瘤转移和早期诊断等研究提供了新的技术平台。本文综述了蛋白质组学在口腔医学领域应用的研究进展。     

用比较蛋白质组学技术筛选口腔黏膜上皮细胞体外癌变的相关蛋白

目的：探讨口腔黏膜上皮细胞在体外癌变不同阶段表达的差异蛋白质。方法：以口腔黏膜上皮细胞体外癌变模型为对象,采用双向凝胶电泳技术和图像分析软件PDQuest分离和分析不同阶段细胞间的差异蛋白质点,采用LC-MS/MS质谱分析系统鉴定差异蛋白质点,采用Gene Ontology Annotation将已知差异蛋白质进行分类。结果：采用双向凝胶电泳技术和图像分析软件PDQuest,得到差异蛋白质点54个,采用LC-MS/MS质谱鉴定后,共得到候选差异蛋白质45个。根据Gene Ontology Annotation分类,按细胞组成分布最多的差异蛋白质位于细胞质和细胞膜,按分子功能分布最多的是磷酸酶活性、催化活性、结构分子功能和钙离子结合功能,按生物过程分布最多的是代谢过程、细胞信号传导和细胞黏附与运动。结论：比较蛋白质组学方法中的双向凝胶电泳和质谱技术,能够很好分离和鉴定口腔黏膜上皮细胞体外癌变模型中不同阶段细胞的差异蛋白质。     

蛋白质组学在口腔医学中的研究进展

蛋白质组学是在人类基因组计划研究发展的基础上形成的新兴学科,它的主要任务是识别、鉴定细胞、组织或机体的全部蛋白质,并分析蛋白质的功能及其模式.随着蛋白分离,鉴定技术和生物信息学的完善,蛋白质组学在各个基础和临床领域得到了长足的发展.在口腔医学领域从鳞癌蛋白质含量对比分析开始,其高通量、高灵敏度的技术特点已逐渐应用于口腔牙体组织、唾液的蛋白质全表达、口腔内环境菌群蛋白功能研究、口腔肿瘤发生发展的蛋白机制表达和生物标记等方面,并取得了相应的成果,我们就蛋白质组学在口腔医学中应用的相关技术及其研究进展作一阐述.     

唾液蛋白质组学的研究进展

唾液作为一种成分复杂，在口腔内具有多种生物学功能的液体。它起到润滑、辅助消化、抗茵等诸多作用，而对唾液的功能和其分泌的分子机制的研究离不开对唾液蛋白质的研究。蛋白质组学作为大规模蛋白质分离和识别技术研究蛋白质组的一门学科，已经开始在很多领域得到应用，同时正逐渐成为唾液蛋白组研究中的新技术。本文就蛋白质组学在唾液中的蛋白全表达、生物标记应用等方面内容作一综述。     

人乳牙牙髓干细胞和恒牙牙髓干细胞的蛋白质组学比较

目的 采用蛋白质组学方法研究人乳牙牙髓干细胞（SHED）和恒牙牙髓干细胞（DPSC）中的蛋白表达差异.方法 应用双向凝胶电泳技术分离SHED和DPSC的细胞总蛋白.通过比较两种细胞的蛋白组学图谱,确定差异表达的蛋白点,而后对差异点进行基质辅助激光解析电离飞行时间质谱分析和蛋白数据库信息检索,对差异蛋白进行功能分类.结果 建立了SHED和DPSC的蛋白质组图谱,经软件分析出45个差异蛋白点,其中26个表达上调,19个表达下调,再经质谱鉴定出48种蛋白,其生物学功能涉及细胞周期、代谢等.结论 SHED与DPSC中蛋白的差异表达体现了两种细胞在结构和功能上的异同性,为进一步研究SHED和DPSC在增殖、分化中的差异,以及牙齿相关干细胞在组织工程和再生医学研究中的应用提供参考.     

双向电泳分析口腔鳞癌组织蛋白表达差异

目的:利用蛋白质组学方法建立分辨率高和重复性好的人口腔鳞癌组织及正常口腔黏膜组织的双向凝胶电泳图谱.分析其差异表达的蛋白质,为进一步寻找口腔鳞癌标志物奠定基础.方法:采用固相pH梯度双向凝胶电泳分离人口腔鳞癌组织及配对的正常口腔黏膜组织的总蛋白质,凝胶经银染显色后,ImagingMaster 2D图像分析软件进行比较分析,识别差异表达的蛋白质.结果:①癌组织和正常口腔黏膜组织凝胶的平均蛋白质点数分别为2325±390个和2487±281个.②通过比较分析10例口腔鳞癌组织及正常口腔黏膜的双向凝胶电泳图谱,得到差异表达蛋白点数为29个,这29个点在癌组织中均为低表达.结论:本研究建立了分辨率高且重复性较好的人口腔鳞癌组织及其正常口腔黏膜组织的双向凝胶电泳图谱.发现两者间存在一些差异表达的蛋白质,为进一步筛选口腔鳞癌特异性的分子标志物打下了坚实基础.     

变性梯度凝胶电泳分析口腔微生物多样性的研究进展

变性梯度凝胶电泳(DGGE)是近些年微生物分子生态学研究中的热点技术之一.由于DGGE技术具有可靠性强、重现性高和方便快捷等优点,被广泛地应用于微生物群落多样性和动态性分析.下面就DGGE的原理、在口腔微生物研究中的应用、优缺点和应用前景作一.     

The prominent proteins expressed in healthy gingiva: a pilot exploratory tissue proteomics study

Gingiva is a unique tissue which protects the underlying periodontal tissues from consistent mechanical and bacterial aggressions. Molecular analysis of gingiva is likely to improve our understanding of the underlying biological processes at work. The aim of this preliminary exploratory study is to analyze the proteomic profile of healthy gingiva and to detect prominently expressed proteins. Gingival tissue samples were obtained from periodontally healthy individuals who underwent surgical crown lengthening procedure. After protein isolation, two dimensional gel electrophoresis (2DE) gels were prepared for each sample and only protein spots common to all gels were selected to eliminate the bias caused by the effect of individuals on proteomic profile. Following the 2DE; in-gel tryptic digestion and MALDI-TOF/TOF steps were performed for protein identifications. Forty-seven proteins were successfully identified. The identified proteins were classified based on their classes, molecular functions and involvements in biological processes and metabolic pathways. Among them, 14-3-3 protein sigma, Protein DJ-1, Alpha-enolase, Triosephosphate isomerase, Superoxide dismutase, Peroxiredoxin-1, Protein S100-A9, Galectin-7, Annexin A2/A4, Carbonic anhydrase 1 and chaperone proteins are worthy of attention. The proteomic profile of the gingiva reflected its highly dynamic characteristics. Despite complexity of the gingival tissue proteome, 2DE was an effective approach in studying the common protein expression profile of the gingiva. Considering the significance of gingiva in the formation of periodontal diseases, it is important to generate a detailed proteome map of gingival tissue to set up a bridge between molecular events and the disease formation. This study established an initial proteome map of the gingival tissue from healthy individuals.     

Proteomic analysis of protein components in periodontal ligament fibroblasts

BACKGROUND: Characterization of periodontal ligament (PDL) fibroblast proteome is an important tool for understanding PDL physiology and regulation and for identifying disease-related protein markers. PDL fibroblast protein expression has been studied using immunological methods, although limited to previously identified proteins for which specific antibodies are available. METHODS: We applied proteomic analysis coupled with mass spectrometry and database knowledge to human PDL fibroblasts. RESULTS: We detected 900 spots and identified 117 protein spots originating in 74 different genes. In addition to scaffold cytoskeletal proteins, e.g., actin, tubulin, and vimentin, we identified proteins implicated with cellular motility and membrane trafficking, chaparonine, stress and folding proteins, metabolic enzymes, proteins associated with detoxification and membrane activity, biodegradative metabolism, translation and transduction, extracellular proteins, and cell cycle regulation proteins. CONCLUSIONS: Most of these identified proteins are closely related to the extensive PDL fibroblasts' functions and homeostasis. Our PDL fibroblast proteome map can serve as a reference map for future clinical studies as well as basic research.     

人牙髓细胞向成牙本质细胞分化的蛋白质组学研究

目的 采用蛋白质组学方法分析人牙髓细胞向成牙本质细胞分化过程中细胞蛋白表达谱的改变,揭示特征蛋白在分化过程中所起的作用.方法对人牙髓细胞进行矿化诱导,提取诱导前后细胞总蛋白,双向电泳分离蛋白质,DeCyder V6.0软件确定差异蛋白点,质谱鉴定差异蛋白质.结果双向电泳确认46个差异蛋白质斑点,质谱鉴定20个蛋白斑点,差异蛋白质涉及细胞周期调节、能量代谢、信号传导等细胞生物学过程.结论蛋白质组学技术可高通量筛选与人牙髓细胞向成牙本质细胞分化相关的功能蛋白.     

Analisi tissutale proteomica della tasca parodontale. Uno studio pilota

ObjectivesThe aim of this study was to analyse in the same subject, in sites where no periodontopathogenic bacteria were detectable, pocket-associated and neighboring healthy interproximal tissues, with a view to qualifying proteins associated with the periodontal damage.Materials and methodsFifteen healthy patients, suffering from moderate to advanced chronic periodontitis and presenting at least one intrabony defect and a neighboring not-damaged interproximal site, were enrolled. Patients underwent osseous resective surgery. During surgery pocket-associated and clinically healthy tissues were harvested for proteomic analyses.ResultsIn both pocket-associated and clinically healthy tissues, 19 differently expressed proteins were successfully identified. In particular, 8 proteins – annexin A2; actin cytoplasmic 1 (2 spots); carbonic anhydrase 1; carbonic anhydrase 2; Ig kappa chain C region (2 spots) and flavin reductase – were overexpressed, while 11 proteins – tropomyosin alpha-4 chain; 14-3-3 protein sigma; 14-3-3 protein zeta/delta; alpha-enolase; heat shock protein beta-1 (2 spots); triosophosphate isomerase; peroxiredoxin-1; fatty acid-binding protein, epidermal; protein S100-A9 and galectin-7 – were underexpressed in the pathological tissue of all patients.ConclusionsThe preliminary data indicate a differential expression of proteins that may play important roles in the prevention of cellular damage by stress, in mediating the immune response as well as in tissue regeneration. The proteomic profile study of pocket tissue may be crucial to appreciate both the pathogenesis and the therapy of periodontitis.     

Comparative proteomic analysis of human whole saliva

Human saliva performs a wide variety of biological functions that are critical for the maintenance of the oral health. Various functions include lubrication, buffering, antimicrobial protection, and the maintenance of mucosal integrity. In addition, whole saliva may be analysed for the diagnosis of human systemic diseases, since it can be readily collected and contains identifiable serum constituents. By using proteomic approach, we have established a reference proteome map of human whole saliva allowing for the resolution of greater than 200 protein spots in a single two-dimensional polyacrylamide gel. Fifty-four protein spots, comprised of 26 different proteins, were identifies using N-terminal sequencing, mass spectrometry, and/or computer matching with protein database. Ten proteins, whose levels were significantly different when bleeding had occurred in the oral cavity, were discussed in this study. These 10 proteins include -1-antrypsin, apolipoprotein A-I, cystatin A, SA, SA-III, and SN, enolase I, hemoglobin β-chain, thioredoxin peroxiredoxin B, as well as a prolactin-inducible protein. The proteomic approach identifies candidates from human whole saliva that may prove to be of diagnostic and therapeutic significance.     

Comparative proteomic profiling of human dental pulp stem cells and periodontal ligament stem cells under in vitro osteogenic induction

ObjectiveThis study aimed to compare the proteomic profiling of human dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs) under in vitro osteogenic induction, which imitates the microenvironment during osteo-/odontogenesis of DPSCs and PDLSCs.DesignThe proteomic profiles of osteoinduced DPSCs and PDLSCs from a single donor were compared using the isobaric tag for relative and absolute quantitation (iTRAQ) technique and subsequent bioinformatics analysis.ResultsA total of 159 differentially expressed proteins in PDLSCs and DPSCs were identified, 82 of which had a higher expression level in PDLSCs, while 77 were more highly expressed in DPSCs. Among these enriched proteins, certain members from the collagen, heat shock protein and protein S100 families may distinguish osteoinduced PDLSCs and DPSCs. Gene ontology (GO) classification revealed that a large number of the enriched terms distinguishing PDLSCs and DPSCs are involved in catalytic activity, protein binding, regulation of protein metabolic processes and response to stimulus. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated several involved pathways, including the fatty acid biosynthesis pathway, pantothenate and CoA biosynthesis pathway, arachidonic acid metabolism pathway and PPAR signaling pathway. Further verification showed that the mineralization and migration capacities of PDLSCs were greater than those of DPSCs, in which heat shock protein beta-1, Protein S100-A10 and S100-A11 may play a part.ConclusionsLess than 5% of the differentially expressed proteins make up the comparative proteomic profile between osteoinduced PDLSCs and DPSCs. This study helps to characterize the differences between osteoinduced PDLSCs and DPSCs in vitro.     

Gingival crevicular fluid and its immune mediators in the proteomic era

The discovery of biomarkers for periodontal disease requires an in‐depth understanding of the molecular basis of the initiation and progression of the disease. The gingival crevicular fluid is a biological medium suitable for identifying and measuring such biomarkers because it can be easily and noninvasively sampled from the immediate vicinity of the affected tissues. An ever‐expanding pool of gingival crevicular fluid proteins associated with periodontal health or disease has been catalogued over the years, particularly with the recent implementation of proteomic technologies. ‘Proteomics’ refers to the large‐scale study of entire arrays of proteins expressed by a genome and present in a cell, tissue, biological fluid or organism. Hence, such technologies provide a broad qualitative and quantitative insight of the proteins present in gingival crevicular fluid. Pertinent studies have amassed on the information gathered to date on protein signatures in periodontal health and disease, and have confirmed the nature of the immunological host response. This review discusses the application of proteomic technologies in characterizing the molecular networks present in gingival crevicular fluid, their potential for discovery of biomarkers that are meaningful for clinical practice, and the associated technical challenges.     

Towards second-generation proteome analysis of murine enamel-forming cells

Proteome analysis of rat enamel-forming cells, initiated over a decade ago, has provided valuable insights to enamel biology. In preparation for a more comprehensive, second-generation proteomic exploration, we evaluated an updated microsample-profiling strategy that comprises sequential extraction of enamel epithelium, parallel one- and two-dimensional gel electrophoresis, and mass spectrometric sequence analysis. The results indicated that several hundred proteins, representing various cellular compartments (including membranes), are amenable to identification with a starting tissue volume of < 10  µ l. With its increased proteomic depth and breadth, this straightforward approach constitutes a major advance from the first-generation work (10-fold increased proteome coverage), although care was needed to ensure a comparably high stringency of protein identification. Expression proteomics has an exciting potential to elucidate the inner workings of murine enamel epithelial cells, leading to an improved understanding of enamel in health and disease.