A Preliminary Study On The Correlation Between β-1, 4-galactosyltransferase-I And Keloids Collagen Synthesis

PartⅠExpression ofβ-1,4-galactosyltransferase-I in KeloidsObjective: To observe the expression level ofβ-1,4-GalT-I in keloids, to analyze the correlation betweenβ-1,4-GalT-I and the synthesis of collagen, and to preliminarily investigate the role ofβ-1,4-GalT-I during the fibrotic process of keloid.Methods: The samples obtained, 10 keloids, 6 normal skin and 7 hypertrophic scars, were prepared for frozen sections and subjected to hematoxylin eosin stain. The type, content and distribution of collagen fibers were detected by Sirius red stain and polarization microscopy method. HP content was measured by the kit, and then the total collagen content of the tissue was assessed according to that. The expression level ofβ-1,4-GalT-I in keloids was evaluated by semi-quantitative RT-PCR analysis. The relative expression level ofβ-1,4-GalT-I mRNA was indicated by the ratio of the optical density value ofβ-1,4-GalT-I to that of GAPDH. The Stata7.0 software was used in one-factor analysis of variance and comparison between every two groups. And correlation analysis was used to judge the dependablity between the expression level ofβ-1,4-GalT-I and the collagen content in tissues.Results: (1) The collagen fibers in keloids appeared thicker and more irregular than in normal dermis. Keloids contained increased fibroblasts, capillary vessels and infiltration of inflammatory cells, large and thick collagen fibers composed of numerous fibrils closely packed together. (2) As we found, type I collagen fibers were closely packed thick fibrils giving an intense birefringence with yellow-red color, whereas these green collagen fibers forming in thin fibril giving a weak greenish birefringence were mainly type III collagen fibers. In keloids indicated by image analysis, the proportion of type I collagen was (71.53±4.03) %, and type III collagen was (28.47±4.03) %. The collagen I-to-collagen III ratio was (2.56±0.53). Compared with negative controls, there was statistical significance (P<0.01), while compared with positive controls there was no differences (P>0.05). (3) As showed the HP content in each group respectively was (10.98±1.55), (5.38±0.47) and (10.21±1.25)μg/mg. The statistical result was as the same as the above. It hinted that the fibrosis degree of keloids and hypertrophic scars was very visible. (4) The relative level ofβ-1,4-GalT-I mRNA in keloids was (0.2995±0.0825), which was extraordinary higher than that of normal tissues, (0.0451±0.0287). Also there was no difference between keloids and hypertrophic scars; the latter was (0.2769±0.0975). According to the above results, we found there was similar variation tendency between the elevated level ofβ-1,4-GalT-I mRNA and the increased collagen content. Furthermore, they had the positive correlation with each other (r=0.9507, P<0.01).Conclusion: There is close relation between the level ofβ-1,4-GalT-I and collagen composition in keloids. It indicates thatβ-1,4-GalT-I may participate the pathogenesis of keloids and play an important role in the synthesis of collagen. PartⅡExpression of Galβ-1,4-GlcNAc in KeloidsObjective: To investigate the expression level and location ofβ-1,4-glycosidic bond in keloids, and to approach the role of glycoprotein galactosylation during the formation of keloids.Methods: The glycoprotein glycosylation level was measured by RCA-I lectin blotting. Then the immunofluorescence histochemistry was applied to detect the expression of Galβ-1→4GlcNAc in keloids, in which the galactose-specific lectin, RCA-I lectin can specially bind Galβ-1→4G1cNAc. And double immunofluorescent staining was chosen to observe colocalization of galactose-containing glycans and procollagenα1 type I.Results: (1) There were some impalpable changes detected in keloids at about 30KD and 40KD of glycoproteins by this assay. The same result was observed in hypertrophic scars. (2) Through the fluorescence microscope we observed that signals of positive increased in keloids compared with normal skin, and the receptor of RCA-I uniformly located on the fibroblasts membrane and in the cytoplasm. (3) It also revealed that galactose-containing glycans in keloids had colocalization with procollagenα1 type I; the distribution was as the same as the single maker. And the amount significantly augmented. The same result was observed in frozen sections of hypertrophic scar.Conclusion: These results suggests that the level of Galβ-1,4GlcNAc is changed in keloids andβ-1,4-galactosylated carbohydrate chains may participate the reparative process by mediating modify of fibrosis.