| Project/Area Number |
05454183
|
|---|
| Research Category |
Grant-in-Aid for General Scientific Research (B)
|
| Allocation Type | Single-year Grants |
|---|
| Research Field |
Experimental pathology
|
|---|
| Research Institution | Oita Medical University |
|---|
Principal Investigator |
YAMAMOTO Shunsuke Oita Medical Univ.Pathology, Professor, 医学部, 教授 (90040188)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
AKIZUKI Shinichiro Oita Medical Univ.Pathology, assistan, 医学部, 助手 (80159334)
|
|---|
| Project Period (FY) |
1993 – 1994
|
| Project Status |
Completed (Fiscal Year 1994)
|
| Budget Amount *help |
¥6,900,000 (Direct Cost: ¥6,900,000)
Fiscal Year 1994: ¥1,800,000 (Direct Cost: ¥1,800,000)
Fiscal Year 1993: ¥5,100,000 (Direct Cost: ¥5,100,000)
|
|---|
| Keywords | MS2 / metalloproteinase / disintegrin / CD14 / TNFalpha / transgene / recombinant / dismtegrin / collagenase / CDNA / Kupffer / monoclonal antibody |
|---|
| Research Abstract |
MS2 : 1.Recombinant proteins for metalloproteinase (MTP) and disintegrin (DI) domains of mouse MS2 (mMS2) and human MS2 (hMS2) were produced. DI activity of hMS2 DI protein was observed and inhibited by anti-DI antibody. Monoclonal antibodies (mAbs) for DI proteins of mMS2 and hMS2 were produced. Flowcytometry and immunocytomemical analysis confirmed expression of MS2 on mouse and human monocytes and granulocytes using these mAbs. 2.cDNAs for hMS2 and truncated froms of hMS2 were cloned and sequenced. 3.Cis regulatory regions for the constitutive and LPS-inducible expression of the mMS2 gene were determined by chloramphenycol acetyltransferase (CAT) assay.
CD14 : 1.cDNA for rabbit CD14 was cloned and sequenced. 2.Recombinant mouse CD14 (rCD14) and polyclonal antibodies and mAbs for rCD14 were produced. Flowcytometry and immunocytochemistry analysis showed expression of CD14 in granulocytes and monocytic cells. Monoclonal antibodies suppressed TNFalpha production by LPS from macrophage c
… More
ell lines in vitro. 3.Three nuclear proteins capable of binding CD14 responsive cis element were separated on DNA affinity column. 4.Kupffer cells were suggested to serve as a scavenger receptor by analysing mRNA expressions of CD14, CD18 and cytokines including TNFalpha , IL-1beta and IL-6 in the liver and macrophages after stimulation with LPS.5.We found that CD14 enhanced sensitivity to LPS by facilitating interactions between LPS and a CD14-associated LPS-signalling molecule (s) using two lipid A analogs, lipid IV_A Rhodobacter spheroides lipid A.6.Recombinant aminoterminal 109 amino acids of mouse CD14 was produced. 7.we found that TNFalpha production from mouse peritoneal macrophages by LPS was not inhibited by anti-rCD14.8.Transgenic mice (TM) carrying metallothionein promoter-driven native and truncated forms of the mouse CD14 gene were produced. The transgenic CD14 genes were expressed in the liver in mice fed in normal diet, whereas they were expressed in the small intestine as well as in the liver after drinkig zinc containing water. Concentration of soluble CD14 was significantly higher in TM with the truncated CD14 gene (TMT) than in TM with the native CD14 gene (TMN) or nontransgenic mice. TNFalpha level was significantly higher in TMT than in TMN and normal mice. Less
|
