Toshiro Ooyama

Comparative effect of lipoxygenase products of arachidonic acid on rat aortic smooth muscle cell migration.

We investigated the effects of mono-hydroxyeicosatetraenoic acids (HETEs) and N-formyl-methionyl-leucyl-phenylalanine (F-Met-Leu-Phe) on rat aortic smooth muscle cell migration in modified Boyden chambers. 12-HETE showed the most potent stimulatory effect on smooth muscle cell migration among the mono-HETEs tested. The optimal concentrations for cell migration were 3 X 10(-15) and 3 X 10(-13) g/ml for 12-HETE and 10(-8) g/ml for 15-HETE, 5-HETE and F-Met-Leu-Phe were inactive with these cells. As 12-HETE is biosynthesized from arachidonic acid by the 12-lipoxygenase pathway in platelets and macrophages, and 15-HETE by the 15-lipoxygenase pathway in granulocytes, the present results indicate an important role for such cells in the early phase of atherosclerosis.

Calcium dependency of aortic smooth muscle cell migration induced by 12-l-hydroxy-5,8,10,14-eicosatetraenoic acid: Effects of A23187, Nicardipine and Trifluoperazine

We have previously reported that 12-L-hydroxy-5,8,10,14-eicosatetraenoic acid (12-HETE), a 12-lipoxygenase product of arachidonic acid in platelets, is a potent chemoattractant for rat aortic smooth muscle cells. In the present study, the mechanism involved in 12-HETE-associated smooth muscle cell migration was investigated in relation to calcium mobilization in the cells. Migration of smooth muscle cells was measured by a filter membrane technique in modified Boyden chambers. Smooth muscle cell migration induced by 12-HETE increased with the increase of extracellular Ca2+ concentration and became maximal at the physiological Ca2+ concentration of 1.25 mM. The calcium ionophore A23187, at concentrations of 0.2 and 2.0 microM, significantly stimulated cell migration. Nicardipine, a potent calcium-entry blocker, significantly inhibited 12-HETE-associated smooth muscle cell migration at concentrations from 10(-9) to 10(-5) M. Concentrations of trifluoperazine from 10(-9) to 10(-5) M and W-7 at 10(-5) M, which are specific inhibitors of calmodulin, also significantly inhibited cell migration induced by 12-HETE. Cytochalasin B at 1.0 and 10 microM, and colchicine at 0.1 and 1.0 microM concentrations drastically inhibited cell migration, indicating that actin-containing microfilaments and microtubules are involved in smooth muscle cell migration. These findings indicated that the stimulation of smooth muscle cell migration by 12-HETE is a highly calcium-dependent process and suggest that 12-HETE might act at the initial stage of smooth muscle cell migration through enhancing calcium influx through plasma membrane and thus stimulating cell migration.

Platelets stimulate aortic smooth muscle cell migration in vitro: Involvement of 12-l-hydroxy-5,8,10,14-eicosatetraenoic acid

The migration of rat aortic smooth muscle cells was measured in modified Boyden chambers. Smooth muscle cells were motile in vitro and their migration was stimulated (time- and dose-dependently) by a platelet-derived factor. Treatment of platelets with indomethacin resulted in a significant increase in smooth muscle cell migration, whereas treatment with 5,8,10,14-eicosatetraenoic acid inhibited it. Purified 12-L-hydroxy-5,8,10,14-eicosatetraenoic acid at a very low concentration (6 x 10(-15)-6 x 10(-13) g/ml) significantly stimulated smooth muscle cell migration. The locomotion induced by 12-L-hydroxy-5,8,10,14-eicosatetraenoic acid was chemokinetic. These findings point to the physiological importance of a platelet 12-lipoxygenase product of arachidonic acid in the early phase of atherosclerosis.

Low serum estradiol levels in subjects with arterial calcification.

The relationship between arterial calcification and serum estradiol levels was studied in 72 postmenopausal female and 59 male subjects. In both sexes, subjects with iliac artery calcification had rather lower serum estradiol levels (8.4 +/- 1.4 pg/ml in the females, and 19.2 +/- 2.5 pg/ml in the males) than controls (16.1 +/- 1.6 pg/ml in the females, and 29.7 +/- 2.4 pg/ml in the males). The bone mineral content of females with iliac artery calcification (0.44 +/- 0.02 g/cm2) was lower than controls (0.52 +/- 0.01 g/cm2); a positive correlation between serum estradiol levels and bone mineral content was found in the females. However, bone mineral content did not significantly differ between males with and without arterial calcification (0.67 +/- 0.03 g/cm2 in the former, and 0.65 +/- 0.02 g/cm2 in the latter). These results indicate that arterial calcification and increased bone resorption are both individual results of estrogen deficiency.

Radioimmunoassay of pig pancreatic elastase

Elastase can be considered a proteolytic enzyme capable of specifically cleaving elastin in connective tissue. Since the early concept of Balo and Banga [ 1 ] , it has been suggested that elastase of pancreatic origin might be responsible, via the systemic circulation route, for the development of atherosclerosis [2]. However, there has been no firm evidence to support such a hypothesis due to the lack of a reliable method for measurement of the serum elastase content. We have been studying a radioimmunoassay for the measurement of pig pancreatic elastase and succeeded in measuring the serum elastase content of pigs. This is a report on this radioimmunoassay and the results of cross immunity between pig, rat and human pancreatic elastase.

Effective isolation method of human pancreatic elastase-1 by use of immunoadsorbent affinity chromatography

Abstract The isolation of elastase from human pancreatic juice was achieved in a one-step procedure utilizing affinity chromatography of elastase—antibody Sepharose gel. Isolated elastase was an anionic enzyme, elastase-1. Another elastase, elastase-2, and proelastase were not obtained by this method. The procedure presented here is simpler and faster than the conventional elastase-1 isolation procedure.

The Clinical Study of Hyponatremia in the Elderly

老年者原因不明の低Na血症例について腎機能を検討し次の結論を得た.(1) 対象群は尿Na排泄量の少い群 (第一群) と, 多い群 (第2群) に分類可能であった.(2) 第1群では血清BUN高値, CCr低値, PSP色素排泄能低下, 尿濃縮能低下, 尿稀釈能正常, 及び水排泄能正常を示した.(3) 第2群では血清BUN正常, CCr低値, PSP色素排泄能低下, 尿濃縮能低下の傾向, 尿稀釈能低下及び水排泄能低下を示した.(4) 第1群は, Na再吸収過程は正常に保たれていると考えられ, むしろ, 水再吸収過程に問題があるように思われた. 従って, 本群における低Na血症発症過程に腎機能障害が深く関与していた可能性は少いと思われる.(5) 第2群においてはNa再吸収過程は障害されており, 本群の低Na血症の発症には, 腎のNa調節機構の障害が深く関与している可能性がある.

The Interrelationship between Arterial Calcification and Serum Estradiol Levels and Bone Mineral Content in the Aged

動脈壁石灰化と血清 estradiol 値および骨塩含量の関係を明らかにする目的で, 老年者女性77名, 男性59名 (平均年齢はそれぞれ71歳と70歳) 計136名を対象に臨床的検索を行なった. 動脈壁石灰化の有無はレ線写真上肉眼的に判定し, 血清 estradiol は radioimmunoassay にて測定し, 骨塩含量は Bone Mineral Analyzer にて橈骨含塩量 (以下RMCと略す) を測定した. その結果 (1) 動脈壁石灰化と血清 estradiol 値との関係は, 女性では腹部大動脈に石灰化を有する群の血清 estradiol 値 (11.5±1.3pg/ml) は, 石灰化を有さない群のそれ (15.7±2.5pg/ml) に比し有意に低く, (p<0.01), また腸骨動脈に石灰化を有する群の血清 estradiol 値 (8.4±1.4pg/ml) は, 石灰化を有さない群のそれ (16.1±1.6pg/ml) に比し有意に低かった.(p<0.001). 男性では, 腸骨動脈に石灰化を有する群の血清 estradiol 値 (19.2±2.5pg/ml) は, 石灰化を有さない群のそれ (29.7±2.4pg/ml) に比し有意に低かった. (p<0.01). (2) 動脈壁石灰化と骨塩含量の関係は, 女性では腹部大動脈に石灰化を有する群のRMC (0.46±0.02g/cm2) は, 石灰化を有さない群のそれ(0.52±0.02g/cm2) に比し有意に低く (p<0.02), また腸骨動脈に石灰化を有する群のRMC (0.44±0.02g/cm2) は, 石灰化を有さない群のそれ (0.52±0.01g/cm2) に比し有意に低かった. (p<0.001). 一方男性においては, 動脈壁石灰化とRMCの間には関係が認められなかった. (3) 血清 estradiol 値とRMCとの関係は, 血清 estradiol 値をX, RMCをYとすると, 女性ではY=0.0025X+0.4551という関係式が成立し, 両者の間に正の相関関係の存在することが認められた. (p<0.02), 一方, 男性では両者の間に関係が認められなかった. 以上の結果は, 内因性 estrogen 不足が動脈壁石灰化を促進させる可能性があるという点については, 本質的には性差がなく, また estrogen は骨吸収を介する以外の系で-おそらくは動脈壁に対する直接の作用により, 動脈壁をCa沈着から保護するという可能性を示唆する. 女性においては血清 estrogen 低値による動脈への直接の影響に加えて, さらに骨からのCa遊出の増大が加わっている可能性が考えられる.