| Project/Area Number |
61570419
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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 |
Circulatory organs internal medicine
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| Research Institution | Ehime University |
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Principal Investigator |
MURAKAMI Eiki (1987) Ehime University , Lecturer, 医学部附属病院, 講師 (90110832)
高田 泰治 (1986) 愛媛大, 医学部, 講師 (20127898)
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| Co-Investigator(Kenkyū-buntansha) |
FUJIWARA Yasushi Ehime University , Associate, 医学部, 助手 (80156934)
SEKIYA Michihito Ehime University , Associate, 医学部附属病院, 助手 (50171343)
NISHIYAMA Seiichi Ehime University , ex-Associate, 医学部, 元助手 (60145084)
TAKADA Yasuharu Ehime Unversity , ex-Lecturer, 医学部附属病院, 元講師 (20127898)
村上 英紀 愛媛大学, 医学部, 助手 (90110832)
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| Project Period (FY) |
1986 – 1987
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| Project Status |
Completed (Fiscal Year 1987)
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| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1987: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1986: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Keywords | Juxtaglomerular cell / Renin / Density gradient sedimentation / Electron microscopy / Angiotensin / 電子顕微鏡 / カルシウムイオン / カルモジュリン / カルデスモン / ミオシン軽鎖キナーゼ |
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| Research Abstract |
The renin-angiotensin system plays a major role in the homeostasis of blood pressure and sodium-water balance. The pathophysiological alterations of this system are mediated by change in the rate of renin secretion. Renal renin is synthesiszed and secreted by juxtaglomelular (JG) cells. In-vitro model for studying renin secretion is necessary because multiple factors influence renin secretion in vivo. The most suitable in-vitor model appears to be the culture of highly purified JG cells. We established the method to get highly purified JG cells by enzymic dispersion and density gradient sedimentation discribed below.
Rat kidneys were perfused in situ with citrate buffer. After removal of the kidneys, renal cortex was minced and incubated with collagenase solution. The suspension was poured over a 25-um nylon mesh and single cells passing the sieve were washed with culture medium. To improve the enrichment of JG cells, we centrifuged the cells in a discontinuous Percoll-density gradient solution (total 25 ml, density; 1.02, 1.04, 1.06, 1.08 and 1.10). About 35 10^6 cells were placed on the top of percoll-density solution and centifuged at 1,000 g for 30 min. Ten different cell-fractions (2.5 ml in each fraction) were obtained and numbered 1 to 10 from top to bottom portion of the centrifused tube. The renin activity was specifically observed in the cells of fraction 5. The cell viability of this fraction was greater than 80% by examining the ability to excrude trypan blue. The results of electron microscopy showed that JG cells were observed in 16% of the celels in this fraction.
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