| Co-Investigator(Kenkyū-buntansha) |
IKEHARA Yuzuru Aichi Cancer Center Research Institute, Laboratory of Pathology, Researcher, 腫瘍病理学部, 研究員 (10311440)
NAKANISHI Hayao Aichi Cancer Center Research Institute, Laboratory of Pathology, Section head, 腫瘍病理学部, 室長 (20207830)
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| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2000: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1999: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
1) Qie of the most important results of this project is that we couldexploit a very effective method to isolate the smallest target of mtestinalized gastric glands under conditions in which the tissue RNA was well preserved After glands in pyloric mucosa were isolated by a crypt isolation technique, they were fixed in 70 % ethanol in 20-30minutes and then stained by Alcian-blue (AB) staining to identify goblet cells. In addition to definition of morphological characteristics of the isolated glands under phase-contrast microscopy, AB staining allowed us to discriminate between gastric and intestinal type glands. Synthesis of CDNA was successfully from accomplished isolated glands using the RT-PCR method.
2) However, differential display could not be achieved because it was too difficult to synthesize sufficient CDNA from only one isolated gland.
3) Isolated glands of each subtype of intestinal metaplasia (IM), ten were collected and processed for genomic analyses. With the three subtypes
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of isolated glands (IM (-), moderate IM, and severe IM), the expression of mRNAs of homeobox genes, Cdx-1 and Cdx-2, revealedto be important transcriptional factors introducing and maintaining the differentiation of large or small intestinal mucosal epithelial cells in the immature cells of the crypts, were investigated by the semi-quantitative RT-PCR method The expression of β-actin mRNA was also examined as a positive control. The intestinalized gastric glands consistently expressed both Cdx-1 and Cdx-2 mRNAs, levels generally increasing with severity of IM. No obvious differences in mRNA levels between Cdx-1 and Cdx-2 were apparent. Moreover, when mRNA levels of MUC5AQ Pgll, MUC2, villin, defensinS were examined, isolated glands without IM, associated with gastric and intestinal mixed-type IM, and intestinal-type IM showed different expression patterns. Immunohistochemical analyses with antibodies to their gene products demonstrated close correlation between mRNA and protein expression.
4) In conclusion, it is suggested that some genes, which have important roles in differentiation and maintenance of the intestinal mucosal epithelial cells, might control the appearance of IM in gastric mucosa.
5) The interrelationships among these genes are now under investigation. Less
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