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Screening Differentially Gene Expression Profiling Of Labial Glands In Primary Sj(?)gren Syndrome Patients Using CDNA Chip And Study Of The Functions Of Some Genes

Posted on:2008-04-27Degree:DoctorType:Dissertation
Country:ChinaCandidate:Y H WangFull Text:PDF
GTID:1114360218956050Subject:Rheumatology
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Sj(O|¨)gren syndrome(SS) is a systemic chronic autoimmune disease that mainly affects the exocrine glands and usually presents as persistent dryness of the mouth and eyes due to the functional impairment of the salivary and lacrimal glands. The common histopathological feature of all organs affected is a potentially progressive lymphocytic infiltration. The exact etiopathogenisis and pathogenesis are still not well known. But modern study has indicated that occurrence and evolution of SS, which is a complicated biological process with multiple phases and multiple steps, are the same as those of many other diseases. On the molecular level, progress of SS which is regulated synergistic action of multiple genes is involved in this intricate process. Gene chip is a newly emerging biological technique. It can win the aim to screen out differentially expressed genes effectively utilizing slight specimens in a short time, with the advantage of large scale, high-throughput and high efficiency. Through Gene chip, we can get the message of genes expression of SS labial glands in a overall situation. This noval technique provides forceful means to screen out the genes related to SS and to study gene therapy. Our research is the first time to study the differentially expressed genes in labial glands obtained from patients with primary SS and from healthy subjects by gene analysis SS by gene chip in our own country. Our aim was to achieve a more comprehensive and complete picture of ongoing disease mechanisms and biologic pathways activated in SS. and select some genes to study their functions which may be contribute to development of primary SS.Gene Expression Profiling of Labial Glands Clearly Distinguishes Primary Sj(O|¨)gren Syndrome Patients from Healthy Control Subjects using cDNA ChipObjective To identify the differentially expressed genes by comparing the gene expression profiles of labial glands between primary SjOgren syndrome(pSS) and healthy controls. Some screened genes that may be associated with SS were selected for further pathogenesis studying at mRNA and protein level. Materials and methods In this study, 3 patients with primary SS were selected and diagnosed according to the American-european Consensus Group criteria.The control group was composed of 3 subjects. Total RNA was isolated from every biopsy specimen of labial glands respectively. Then partes aequales of purified total RNA was mixed in pSS and healthy controls respectively. The cRNA probes prepared from total RNA were hybridized with two identical genechips containing 4 7000 human transcripts-Affymetrix HG_U133A plus 2.0, genes. After signal detection, Image procession compared and data analysis were performed with Affymetrix GCOS1.2. The differentially expressed genes were screened. The functions of the differentially expressed genes were analyzed by bioinformations according to Gene Ontology. Results 181 significantly differentially expressed genes were detected by genechips. 96 of them were ESTs(Expressed Sequence Tag) and the others were genes with complete sequences. Among the 181 genes, 128 showed higher expression and 53 showed lower expression. Their functions were involved in Cytokine and receptor, immune, apoptosis, cell diffe rentiation, transcription, metabolism, signal transduction, and cell cycle,et al. Conclusion Gene chip, as a new biological technique, can win the aim to screen out differentially expressed genes effectively, with advantages of large scale, high-throughput and high efficiency. Through Gene chip, we can get the overall genes expression message of SS labial glands.. This noval technique provides forceful means to screen out the genes related to SS and study gene therapy.mRNA transcription and protein expression of MMP-3,MMP-9,OPN,DMP1 in labial glands of pSS patients and healthy controlObjective Selected osteopontin(OPN) and Dentin matrix protein 1(DMP1) from differentially expression genes screened by gene chip. To investigate the mRNA transcription and protein expression of OPN,DMP1 and their respective known parter MMP-3 and MMP-9. Possible role of OPN,DMP1,MMP-3 and MMP-9 in primary SS were analyzed. Materials and Methods The biopsy specimens from Sj(O|¨)gren syndrome patients and control were collected in the stomatological clinic of Peking Union Medical College Hospital. For this study, 24 patients with primary SS were selected and diagnosed according to the American-European Consensus Group criteria. The control group was composed of 14 subjects.①Total RNA from labial glands was extracted. The target genes andβ-actin acted as templates to perform reverse transcript polymerase chain reaction(RT-PCR). Then mRNA expression of target genes were semi-quantitated by the OD ratio of the target genes toβ-actin bands on gel under densitometry.②Immunohistocheical analysis was used to determine the expression of proteins of targets genes in labial glands from patients and healthy control.③Comparition between two groups and correlation study were done by using the software of statistics Pakage for social science(SPSS11.0). Results In contrast to healthy subjects, the mRNA transcription and protein level of target genes in labial glands in primary Sj(O|¨)gren syndrome patients were significantly increased statistically(P<0.05). The expression of OPN was positively correlated with the lymphocyte foci-score(LFS).Conclusion Our observation that the proteolytic action of MMPs toward ECM macromolecules is increased in SS patients provides a rationale for understanding organization observed in the basal lamina and apical surface of acini in these patient. Our results provide new evidence that acinar and ductal cell from the LSGs Of SS patients has a molecular potential, with increased capacity to markedly disorganize their ECM-environment and,thus,damage their architecture and functionality. MMP can be suggested that they may play a role in the pathogenesis of Sj(O|¨)gren syndrome. However, OPN and DMP1 as the genes studied for the frist time in SS, their role is to be elucidated.
Keywords/Search Tags:primary Sj(O|¨)gren syndrome, labial gland, gene chip, differentially expressed genes, reverse transcript polymerase chain reaction, immunohistochemistry
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