| Budget Amount *help |
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1990: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1989: ¥1,200,000 (Direct Cost: ¥1,200,000)
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| Research Abstract |
Several subclones were obtained from the megakaryoblastic cell line, CMK, using the limiting dilution method. Among them, two subclones, CMK6 and CMK11-5, were studied. CMK6 showed more immature megakaryoblast, on the other hand, CMK11-5 showed more mature megakaryoblast both morphologically and in cell surface markers.
CMK cells produced interleukin-6 (IL-6) and were induced both the proliferation and differentiation by IL-6.
The cDNA library contained 1.2x10^7independent clones was prepared from the mRNAs of CMK cells which were induced the differentiation by phorbol ester (TPA) using lambda phage gt11. Two kinds of M13 phage cDNA libraries were also prepared as the probe from the mRNAs of CMK cells which were induced the differentiation by TPA or not. The clones which hybridized with the probe of TPA-treated CMK cells but not with the probe of CMK cells were selected from the cDNA library of TPA-treated CMK cells using differential hybridization method. Thirty six clones, confirmed the specificity using Northern blotting method, were obtained by 5 different experiments and finally 18 clones were obtained with the exception of overlapped clones. Among them, 3 clones were identified as glucose transporter like protein, glutation peroxidase and leukocyte common antigen, but the other clones were unknown. The study for identification is now ongoing.
The hematopoietic stem cells are recognized by the antibody of CD34. The CMK cells also had the characteristics of hematopoietic stem cells such as the positive reaction to the antibody of CD34. The extents of reaction to it were lower in TPA-treated CMK cells and CMK11-5 cells than in CMK cells. These results induce the possibility that new specific substances recognized in stem cells could be identified by screening the mRNAs which disappeared with differentiation.
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