| Project/Area Number |
12672135
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (C)
|
| Allocation Type | Single-year Grants |
|---|
| Section | 一般 |
|---|
| Research Field |
Biological pharmacy
|
|---|
| Research Institution | Kyoto Pharmaceutical University |
|---|
Principal Investigator |
SATO Takashi Kyoto Pharmaceutical University, School of Pharmaceutical Sciences, Professor, 薬学部, 教授 (40065917)
|
|---|
| Project Period (FY) |
2000 – 2002
|
| Project Status |
Completed (Fiscal Year 2002)
|
| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 2002: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2001: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 2000: ¥1,400,000 (Direct Cost: ¥1,400,000)
|
|---|
| Keywords | ceramide / phospholipase A_2 / extracellular signal-regulated kinase 1 / 2 / protein tyrosine phosphatase / phosphatidic acid / cholesterol ester / cyclooxygenase-2 / nuclear factor κB / スフィンゴミエリナーゼ / 酸化LDL / 活性酸素種 / nuclear factor-κB / extracellular signal-regulated kinase / アラキドン酸 / プロスタグランジンD_2 / 血小板 |
|---|
| Research Abstract |
In the study, the regulatory mechanism of cytosolic phospholipase A_2 (cPLA_2) by ceramide, and the implication of ceramide in the pathogenesis of atherosclerosis and nephropathy via acceleration of cPLA_2 activation were investigated. The results obtained are as follows.
1) When rabbit platelets were stimulated with Ca^<2+> ionophore or Ca^<2+> was added into platelet lysate, Ca^<2+>-dependent association of cPLA_2 with platelet membranes was assessed by increase in cPLA_2 protein using immunoblot analysis and in the activity in membranes. Ceramide apparently increased the association.
2) In rat basophilic leukemia cells stimulated with antigen, ceramide accelerated orthovanadate-sensitive protein tyrosine phosphatase activation, which induces dephosphorylation of extracellular signal-regulated kinase 1/2 to attenuate the potency of cPLA_2 activation. Furthermore, ceramide suppressed antigen-induced phospholipase D activation and subsequent formation of phosphatidic acid, which is chara
… More
cterized to be substrate for cPLA_2. These events lead to decrease in cPLA_2 activity, hence, providing evidence that ceramide acts as a negative regulator in agonist-stimulated cPLA_2 activation.
3) Cholesterol ester formation stimulated by the uptake of oxidized low-density lipoproteins in macrophages was fully dependent on cPLA_2 activation. Under the condition de novo synthesis of ceramide was elicited. Furthermore, pretreatment of the cells with sphingomyelinase enhanced the ester formation. These results indicate that de novo-synthesized ceramide accelerates cholesterol ester formation via cPLA_2 activation leading to foam cell formation, and suggest that ceramide is implicated in the initiation and development of atherosclerosis.
4) Stimulation of mesangial cells with tumor necrosis factor α (TNFα) increased expression of cyclooxygenase-2 and secretory PLA_2 following activation of transcription factor, nuclear factor-κB. Pretreatment of the cells with sphingomyelinase enhanced these responses and the enhancement was fully dependent on the production of reactive oxygen species. TNFα also activated sphingomyelinase to produce ceramide from sphingomyelin. These results suggest that ceramide facilitates production of prostaglandins and is implicated in the pathogenesis of renal diseases. Less
|
