| Apostichopus japonicus,which belongs to Echinodermata,Holothuroidea,stichopodidae.A.japonicus is a kind of important marine economy echinoderms.Sea cucumber in the face of predators or adverse environmental conditions(high salinity stress,no oxygen)can be visceral(digestive tract,respiratory tree,gonad)excreted;when the environmental conditions to improve,can timely regeneration of new features to improve the organ.The histology,immunohistochemistry,immunofluorescence and Western blot method of imitation sea cucumber intestine regeneration process of intestinal stem cells were labeled and intestinal regeneration during embryonic development related protein expression and so on.Studied in the process of sea cucumber intestinal regeneration stem cell marker and some important development related protein expression levels in echinoderm regeneration has certain theoretical significance and can provide reference for the study of human organ regeneration and regenerative medicine.The results of double immunofluorescence showed that in intestinal regeneration CD44 proteins in the process of tissue were expressed in the intestinal regenerating;MSI1 protein in the intestinal tissue regeneration expression,you can prove that the two protein can be used as identification of stem cell markers used in sea cucumber Apostichopus japonicus intestinal stem cells.The results of histology showed third days of sea cucumber intestine regeneration,mesenteric marginal thickening.Sixth days of regeneration,further thickening,mesenteric primordia formate.The regeneration of ninth days,the further development of neonatal intestinal bowel,began to form and began to form wrinkles.Regeneration twelfth days,mesenteric further the thickening of intestinal wall into the digestive tract,intestinal tissue regeneration formed completely,divided into intestinal epithelium,connective tissue layer and coelomic epithelial layer.The regeneration of fifteenth days,the digestive tract gradually have the function,organization structure began to rebuild,intestinal epithelial layer composed of multiple layers of cuboidal cells gradually differentiated into columnar cells,muscle layer gradually the formation of the eighteenth day.The regeneration,regeneration stage of digestive tract with digestive function,muscle layer longitudinal muscle layer and muscular layer.The regeneration of digestive tract in twenty-first days,almost reached the normal level.The results of immunohistochemical showed that in intestinal regeneration process and adult sea cucumber,the expression of ?-catenin.Regeneration of the 3-6 day,?-catenin mainly expressed on the edge of the mesentery.Regeneration of the 9th day,?-catenin in the intestinal wall and intestinal expression more.Regeneration of the 12 days,?-catenin expressed the most,mainly concentrated in the regeneration of the bowel.Regeneration 15-18 days,?-catenin with regeneration and expression of less and less,is hardly expressed in muscle layer.Regeneration of the 21 days,?-catenin expression and localization of almost with the same adult sea cucumber.The results of immunofluorescence showed that in the intestinal regeneration process,Wnt6 protein in the intestinal regeneration was expressed in small intestine of the body.In imitation of sea cucumber intestine regeneration process,Wnt6 protein of the expression quantity increased,although with regeneration decreased gradually.In the regeneration of the9-12 days,the expression of capacity and the normal state compared to the highest.The 15-21 days,Wnt6 protein expression gradually decreased,the adult sea cucumber Apostichopus japonicus Wnt6 expressed the lowest level.The results of Western blot showed that Wnt6 protein in intestinal regeneration process expression increasing first and then decreasing and in regenerating the 9 day expression quantity highest.?-catenin protein in imitation of intestinal regeneration process expression is first increase and then decrease,but with Wnt6 protein has certain difference,in intestinal regeneration the 12 day was the highest,regeneration the 21 st day is has been close to the normal level.The experimental results show that CD44 and MSI1 protein further proved as the intestinal stem cell marker for the identification of intestinal stem cells;when the sea cucumber intestine spit,intestinal stem cells may originate from the mesenteric edge;than the mesenteric pass through the pack in a way to form the new gut;Wnt signaling pathway in intestinal regeneration process plays an important role in regulating,the pathway of Wnt6 protein and ?-catenin protein in intestinal regeneration expressed both,have regulating intestinal regeneration occurred. |
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