1995 Fiscal Year Final Research Report Summary
Early diagnosis of borderline disease of mycosis fungoides using in situ PCR method
| Project/Area Number |
06670202
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Human pathology
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| Research Institution | Saitama Medical School |
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Principal Investigator |
ARAI Eiichi Saitama Medical School, Instructor, 医学部, 講師 (60167228)
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| Project Period (FY) |
1994 – 1995
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| Keywords | PCR / In situ hybridization / T-cell receptor / gamma-chain / Paraffin section / Cutaneous lymphoma / Mycosis fungoides / Pseudolymphoma |
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| Research Abstract |
As a preliminary study in the first year, we modified the method of Algata et al. (Diagn Mol Pathol 3 : 275,1994) by setting nine and three primers into 12 and three segments of V_<gamma> and J_<gamma>-region, respectively. In addition, we developed a semi-nested PCR method setting new inner primers in the side of J_<gammas> for application to paraffin sections. As the result we had 15 positive out of 33 cases of cutaneous T-cell ylmphoma (CTCL). In the second year we identified in these postitive cases effective primer sets with which we performed PCR amplification of TCR-gamma chain on paraffin sections. Moreover, on the same section we made in situ demonstration using labelled oligonucleotide placed between 5'-primer and 3'-primer.Positive dots were seen in nuclei by DAB staining or alkaliphosphatase staining.
By electron microscopy, immnohistochimstry, and DNA analysis, we studied four CTCL cases which had been classified as pseudolymphomas because they regressed spontaneously. As the result we clarified that the spontaneous regression was induced by cataclysmic apoptosis (Majno : Am J Pathol 146 : 3,1995), although all these cases qualified histologically as malignant lymphomas and with Southerm blotting demonstration of clonal lymphocytic proliferation. Moreover we demonstrated the existence of imcomplete apoptosis in a case of infantile digital fibromatosis which did not regeress until 14 years of age.
In the next project we will try to increase sensitivity in detective the rearrangedTCR-gamma chain from minute areas in paraffin sections by PCR method using temperature gradient gel electropheresis (TGGE) and denaturing GGE.
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