| Project/Area Number |
12671633
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Obstetrics and gynecology
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| Research Institution | TOKYO DENTAL COLLEGE |
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Principal Investigator |
KANEKO Satoru TOKYO DENTAL COLLEGE, DEPARTMENT OF DENTISTRY, LECTURER, 歯学部, 講師 (40214457)
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| Co-Investigator(Kenkyū-buntansha) |
ISHIKAWA Hiromichi TOKYO DENTAL COLLEGE, DEPARTMENT OF DENTISTRY, ASSOCIATE PROFESSOR, 歯学部, 助教授 (60112679)
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| Project Period (FY) |
2000 – 2001
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2001: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2000: ¥2,700,000 (Direct Cost: ¥2,700,000)
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| Keywords | sperm / DNA break / DNA fragmentation / TUNEL method / cell electrophoresis / adenyl purine / 染色体 |
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| Research Abstract |
1. DNA fragmentation in human sperm was observed by means of TUNEL method : To obtain negative control for DNA fragmentation, sperm was purified by the density gradient centrifugation in 99% Percoll and subsequent swim up. DNA nicks caused by DNA fragmentation were labeled with FITC by TdT reaction and counter stain with PI. Fluorescence was observed by the aid of high sensitive CCD camera. When the qualities of human semen (n=129) were classified according to the criteria of WHO, The TUNEL positive rate was higher in oligo-asthenozoospermia (25.9±16.0%, n=24) than normozoospermia(9.4 ±6.2%, n=73).
2. Double strand break in DNA was observed by means of Comet cell electrophoresis. Sperm was mounted in 0.7% agar and treeated with proteinase K and detergent. After electrophoresis at 5.0V for 40 min, the sperm DNA was stained with cyber gold. The sperm with elongated comet image was estimated to be with double strand break and those without the image being under detectable. When the comet positive rate was compared with semen qualities, the rate gave poor correlations with sperm density and motility.
3. The blood plasma and seminal plasma involved adenyl purins. In the former, ATP occupied more than 50% and cAMP was under-detectable. On the other hand, adenosine occupied 57% in the latter and ATP was found to be less than 1.0%. The composition of adeny purines in two body fluids were quite different each other.
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