[Expression profiles of the extracellular matrix-associated genes during rat liver regeneration].
ABSTRACT It has been well known that the extracellular matrix (ECM) plays an important role in cell polarization, cell adhesion, cell proliferation, morphogenesis and differentiation. For the sake of the further in-depth investigation of the changes and actions of ECM in liver regeneration (LR) at gene transcriptional level, the ECM-associated genes were obtained by databases searching and literature retrieving, subsequently their expression profiles in rat regenerating liver were detected using Rat Genome 230 2.0 array, then LR-associated genes were identified based on comparison of the gene expression difference between sham operation (SO) group and partial hepatectomy (PH) group. A total of 97 genes were verified to be LR-associated. The initially and totally expressed number of these genes occurring in initial phase of LR, G0/G1 transition, cell proliferation, cell differentiation and structure-functional reconstruction were 49, 19, 43, 5 and 84, 51, 369, 144, respectively, illustrating that expression of the ECM-associated genes were initiated mainly in the early phase, working in differ-ent phases. Their expression similarity was classified into 5 groups including only up-, predominantly up-, only down-, predominantly down-, and equal in up-regulated and down-regulated, involving 38, 21, 21, 10 and 7 genes, respectively; the number of up-regulated expressed genes and down-regulated expressed ones was 411 and 186; their expression patterns were categorized into 24 types, showing that the physiological and biochemical activities in LR were characterized by phase, diversity and intricacy. According to expression profiles and expression patterns of the ECM-associated genes in LR, it was confirmed that the levels of the below-listed genes in expression increased at the corresponding phases of LR, including fibronectin-associated genes at early phase in LR, and collagen-associated genes at middle phase.
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ABSTRACT: Recruitment and proliferation of Thy-1+ oval cells is a hallmark of liver regeneration after 2-acetylaminofluorene (2-AAF)/partial hepatectomy (PHx) in rats. To understand the molecular mechanism underlying this process, we investigated the role of connective tissue growth factor (CTGF), one of the candidate genes differentially expressed in Thy-1+ oval cells, in this liver injury model. Northern and Western analyses were performed to examine the induction of CTGF in total liver homogenate. Quantitative real-time polymerase chain reaction (PCR), immunofluorescent staining, and in situ hybridization were performed to confirm the expression and localization of CTGF in Thy-1+ oval cells. Finally, a known inhibitor of CTGF synthesis, Iloprost, was administered to 2-AAF/PHx treated rats to investigate the effect of Iloprost on oval cell response. CTGF was found to be up-regulated at both the RNA and protein levels and occurred concurrently with an up-regulation of transforming growth factor beta1 (TGF-beta1). Sorted Thy-1+ oval cells expressed a high level of CTGF gene in a quantitative PCR assay. Colocalization of Thy-1 antigen and ctgf signals by in situ hybridization further confirmed that Thy-1+ oval cells were a source of CTGF. Iloprost administration blocked CTGF induction in treated animals but did not affect TGF-beta1 expression. The inhibition of CTGF induction by Iloprost was associated with a significant decrease in oval cell proliferation and a lower level of alpha-fetoprotein expression as compared with control animals. These results show that CTGF induction is important for robust oval cell response after 2-AAF/PHx treatment in rats.Gastroenterology 07/2005; 128(7):2077-88. · 12.82 Impact Factor
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ABSTRACT: By virtue of their multifunctional nature, proteoglycans (PGs) are thought to govern the process of cell movement in numerous physiological and pathological contexts, spanning from early embryonic development to tumour invasion and metastasis. The precise mode by which they influence this process is still fragmentary, but evidence is accruing that they may affect it in a multifaceted manner. PGs bound to the plasma membrane mediate the polyvalent interaction of the cell with matrix constituents and with molecules of the neighbouring cells' surfaces; they modulate the activity of receptors implicated in the recognition of these components; and they participate in the perception and convergence of growth- and motility-promoting cues contributed by soluble factors. Through some of these interactions several PGs transduce to pro-motile cells crucial intracellular signals that are likely to be essential for their mobility. A regulated shedding of certain membrane-intercalated PGs seems to provide an additional level of control of cell movement. Coincidentally, matrix-associated PGs may govern cell migration by structuring permissive and non-permissive migratory paths and, when directly secreted by the moving cells, may alternatively create favourable or hostile microenvironments. To exert this latter, indirect effect on cell movement, matrix PGs strongly rely upon their primary molecular partners, such as hyaluronan, link proteins, tenascins, collagens and low-affinity cell surface receptors, whereas a further finer control is provided by a highly regulated proteolytic processing of the PGs accounted by both the migrating cells themselves and cells of their surrounding tissues. Overall, PGs seem to play an important role in determining the migratory phenotype of a cell by initiating, directing and terminating cell movement in a spatio-temporally controlled fashion. This implies that the "anti-adhesive and/or "anti-migratory" properties that have previously been assigned to certain PGs may be re-interpreted as being a means by which these macromolecules elaborate haptotaxis-like mechanisms imposing directionality upon the moving cells. Since these conditions would allow cells to be led to given tissue locations and become immobilized at these sites, a primary function may be ascribed to PGs in the dictation of a "stop or go" choice of the migrating cells.Matrix Biology 10/2005; 24(6):400-17. · 3.19 Impact Factor
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ABSTRACT: Several molecular techniques require high-quality RNA, free from DNA. Various methods have been described to obtain RNA to be used in expression studies or as starting material in differential display-reverse transcriptase (dd-RT-PCR), for which high-quality RNA free from DNA is an essential requirement. In this report, we compare three different methods to isolate RNA from Gram-positive bacteria: (1) An acid-phenol extraction protocol. (2) The "RNeasy mini kit" from QIAGEN (Valencia, CA, USA). (3) The "SV Total RNA Isolation System" from Promega (Madison, WI, USA).The QIAGEN-kit delivers the highest amount of RNA with the highest purity. Slot blot analysis and dd-RT-PCR confirm the absence of DNA contamination and Northern blot analysis and dd-RT-PCR show high quality of the extracted RNA. This RNA extraction method thus addresses current problems by permitting rapid and safe isolation with high yields of intact RNA for subsequent analysis.Journal of Microbiological Methods 05/2001; 44(3):235-8. · 2.16 Impact Factor