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Inhibition of vascular smooth muscle cell proliferation by red wine polyphenols is associated with downregulation of cyclin a gene expression
Thursday June 29, 2000: Read by lltle Abstracts T.'W33 Proliferation and Differentiation of Smooth Muscle Cells
318
but also of total mortality) increases both with the degree of reduction of the cholesterol levels and with its prolongation in time, we come to conclusion that cardiologists should consider hypercholesterolemia as a chronic disease that must be treated and treated without interruptions. If the attitude towards the cholesterol-lowering therapy changed in the way we are suggesting, the high coronary risk primary prevention and the secondary prevention of CHD could become more efficacious.
T:W32
INTRACELLULAR
LIPID METABOLISM
ThT1 :~/V32 [ Tyrosine phophorylution ofmaerophage by low density lipoprotein from hemodialysis patients and vitamin E M. Sonobe 1'2, Y. Kida 2, T. Ou 2, K. Kimura I , M. Kishino I , M. Mune 1, S. Yukawa I . ~Third Dept. of lnt. Med. Wakayama Medical College;
2Kozagawa hospital, Wakayama, Japan
genesis. The aim of this study is to clarify the somatic distribution and chronological change of D1T. Methods: Aorta, coronary artery and cerebral artery as well as other aortic branches and visceral arteries were systemically examined in autopsy cases of Japanese children and young adults. The age groups were 0 year, 1-10 years, 11-20 years and 21-30 years with 16 cases in each group. Histological sections were stained with Elastica van-Gieson stain, and intima to media (I/M) ratio was measured on a digitalised image. Results: DIT was a organized structure of elastin, proteoglycan and smooth muscle cells and devoid of lipid pool and foamy macrophages. DIT emerged in the coronary artery as early as in 1 month of age and UM ratio increased proportionately with age. The abdominal aorta, carotid artery and iliac artery showed similar trends, though DIT appeared later than the coronary artery. The cerebral artery and other visceral arteries did not develop DIT until third decades in general, and the thickening, if any, was very thin. Conclusion: DIT is different from atherosclerosis, but emerges in atherosclerosis-prone arteries from childhood and thickens with age. These findings suggest an important role of DIT as a ground for atherogenesis.
Objective: We investigated whether interaction of low density lipoprotein (LDL) from hemodialysis patients (HD) and macrophages induces tyrosine phosphorylated proteins in the macrophages. Methods: Human monocyte-derived macrophages were incubated with HD-LDL (100 g/ml) with (E+) or without ( E - ) vitamin E (600 rag/day) dministration. Lysed proteins were divided into Triton-soluble and insoluble fractions. Both fractions (soluble and insoluble) were separated by SDS-PAGE and electroblotted onto a PVDF membrane. Immunoblotting was performed using antibody against phosphotyrosine. Results: Several proteins in the range 50 KDa-100 KDa were found to be phosphorylated constitutively in he macrophages, not affected by the addition of HD-LDL. HD-LDL either from E - or E+ did not induce any tyrosine phosphorylatd proteins. Macrophages pretreated with tyrosine kinase inhibitor, genestein drastically inhibited tyrosine phosphorylation of these proteins. Conclusion: These data suggest that tyrosine autophosphorylated proteins may play a role in the early step of signal transduction in the macrophages.
ThT2:W32 [ Liver stero127 hydroxylase in hamster: Modulation by
steroids and diets C. Lutton, M. Souidi, S. Dubrac, M. Parquet. Physiologie de la Nutrition,
Universit# Paris-Sud, Orsay, France Objective: To investigate, in hamster liver, the effect of various oxysterols, bile acids or a sucrose-rich versus commercial diet on sterol 27-hydroxylase ($27), an ubiquitous enzymatic activity implicated in anti-atherogenic process and bile acid synthesis. Methods: A sensitive assay (Life Sciences 1999, 64, 1585-1593) was used in order to obtain linearity for the accumulation of labelled 27-hydroxycholesterol in hamster mitochondria. Results: In vitro, epicoprostanol and 27 hydroxycholesterol lowered $27 activity (88 and 29%) while hyodeoxycholic acid and 25 hydroxycholesterol were without effect. A sucrose-rich diet induced a marked decrease in $27 activity (-54%) compared with the commercial diet. In vivo, epicoprostanol and hyodeoxycholic acid lowered ( - 4 6 and 70%) as well as the lithogenic diet ( - 5 6 % ) but 25 and 27 hydroxycholesterol were without effect. Conclusions: Hepatic sterol 27 hydroxylase activity can be modulated by steroids and diets. Researching conditions able to activate this activity might be of great interest.
ThT2:W33 II Inhibition of vascular smooth muscle cell proliferation by red wine polyphenols is associated with downregulation of cyclin a gene expression K. Iiiima, M. Yoshizumi, M. Hashimoto, S. Kim, M. Akishita, J. Ako, Y.Q. Liang, N. Sudoh, T. Watanabe, Y. Ohike, K. Toba, K. Hosoda 1, K. Nakahara I , Y. Ouchi. Department of Geriatric Medicine, The University of
Tokyo; l Suntory Research Center, Osaka, Japan Red wine polyphenols have been shown to contribute to the "French paradox" phenomenon consisting of lower mortality from coronary heart disease in the French population. Although vascular smooth muscle cell (VSMC) proliferation plays an important role in the progression of atherosclerotic lesions, the effects of red wine polyphenols (RWP) on VSMC proliferation have not been elucidated. Methods and Results: To examine the effect of RWP on the growth of VSMC, we extracted a total polyphenolic fraction, called RW-PE from red wine using column chromatography. Cultured rat aortic smooth muscle cells (RASMC) were incubated with RW-PF (1-100 microgram/ml) for 72 hr. In cell count and thymidine uptake assay, treatment with RW-PF showed a potent inhibitory effect on cell growth and DNA synthesis in a dose-dependent manner. In contrast, this polyphenolic fraction did not affect the growth of bovine carotid endothelial cells (BCEC). To elucidate the mechanism of this anti-proliferative effect of RW-PF in RASMC, we investigated expression of cyclin A mRNA and cyclin A promoter activity. RW-PF dramatically dowuregulated the cyclin A mRNA and reduced the cyclin A promoter activity in a dose-dependent manner in RASMC but not in BCEC. In addition, RW-PF decreased the binding of nuclear proteins to the activating transcription factor (ATF) site in the cyclin A promoter, and the expression of transcription factors, cyclic AMP-responsive element-binding protein (CREB) and ATF-1, was also downregulated by RW-PF. Conclusion: The dowuregulation of cyclin A gene expression may contribute to the antiproliferative effect of RWP on VSMC through the inhibition of transcription factor expression. 1
ThT3:W33 ] Magnolol induces a regression of aortic hypertrophy in spontaneously hypertensive rats T.-C. Chou. Graduate of Medical Sciences, National Defense Medical Center,
Taipei, Taiwan Objective: To evaluate whether the Chinese herb, magnolol, induces regres-
T:W33
PROLIFERATION AND DIFFERENTIATION SMOOTH MUSCLE CELLS
OF
|
ThT1:W331J Diffuse intimal thickening emerges in atherosderosis-prone arteries in childhood and the intima/media ratio increases with age Y. Nakashima, Y.X. Chen, K. Sueishi. Pathophysiological and Experimental
Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan Objective: Diffuse intimal thickening (DIT) is considered as an adaptive change of arterial intima and suggested to have a close relation to athero-
sion of hypertrophy and possesses antihypertensive effect in spontaneously hypertensive rats (SHR). Methods: After administration of magnolol (50/zg/kg, p.o.) for 6 weeks, the vascular morphological change, blood pressure, endothelium-dependent response, and the 0 2 formation from aorta in SHR were evaluated. Results: Magnolol resulted in a significant reduction of mean blood pressure compared with that of untreated group in SHR (140.2 :t= 3.7 vs 161.0 :t= 7.0 mmHg, P < 0.05) and the endothelium-dependent relaxation in response to acetylcholine was also improved. In addition, the intima/media thickness and area ratio of aorta were significantly lower in magnolol treated group (Table 1). The production of 0 2 from aorta in SHR was also significantly attenuated by magnolol treatment. Conclusion: Our results show that magnolol possesses beneficial effect of regression of vascular hypertrophy and antihypertensive effect in SHR. In
Xllth International Symposium on Atherosclerosis, Stockholm, Sweden, June 25-29, 2000
318
but also of total mortality) increases both with the degree of reduction of the cholesterol levels and with its prolongation in time, we come to conclusion that cardiologists should consider hypercholesterolemia as a chronic disease that must be treated and treated without interruptions. If the attitude towards the cholesterol-lowering therapy changed in the way we are suggesting, the high coronary risk primary prevention and the secondary prevention of CHD could become more efficacious.
T:W32
INTRACELLULAR
LIPID METABOLISM
ThT1 :~/V32 [ Tyrosine phophorylution ofmaerophage by low density lipoprotein from hemodialysis patients and vitamin E M. Sonobe 1'2, Y. Kida 2, T. Ou 2, K. Kimura I , M. Kishino I , M. Mune 1, S. Yukawa I . ~Third Dept. of lnt. Med. Wakayama Medical College;
2Kozagawa hospital, Wakayama, Japan
genesis. The aim of this study is to clarify the somatic distribution and chronological change of D1T. Methods: Aorta, coronary artery and cerebral artery as well as other aortic branches and visceral arteries were systemically examined in autopsy cases of Japanese children and young adults. The age groups were 0 year, 1-10 years, 11-20 years and 21-30 years with 16 cases in each group. Histological sections were stained with Elastica van-Gieson stain, and intima to media (I/M) ratio was measured on a digitalised image. Results: DIT was a organized structure of elastin, proteoglycan and smooth muscle cells and devoid of lipid pool and foamy macrophages. DIT emerged in the coronary artery as early as in 1 month of age and UM ratio increased proportionately with age. The abdominal aorta, carotid artery and iliac artery showed similar trends, though DIT appeared later than the coronary artery. The cerebral artery and other visceral arteries did not develop DIT until third decades in general, and the thickening, if any, was very thin. Conclusion: DIT is different from atherosclerosis, but emerges in atherosclerosis-prone arteries from childhood and thickens with age. These findings suggest an important role of DIT as a ground for atherogenesis.
Objective: We investigated whether interaction of low density lipoprotein (LDL) from hemodialysis patients (HD) and macrophages induces tyrosine phosphorylated proteins in the macrophages. Methods: Human monocyte-derived macrophages were incubated with HD-LDL (100 g/ml) with (E+) or without ( E - ) vitamin E (600 rag/day) dministration. Lysed proteins were divided into Triton-soluble and insoluble fractions. Both fractions (soluble and insoluble) were separated by SDS-PAGE and electroblotted onto a PVDF membrane. Immunoblotting was performed using antibody against phosphotyrosine. Results: Several proteins in the range 50 KDa-100 KDa were found to be phosphorylated constitutively in he macrophages, not affected by the addition of HD-LDL. HD-LDL either from E - or E+ did not induce any tyrosine phosphorylatd proteins. Macrophages pretreated with tyrosine kinase inhibitor, genestein drastically inhibited tyrosine phosphorylation of these proteins. Conclusion: These data suggest that tyrosine autophosphorylated proteins may play a role in the early step of signal transduction in the macrophages.
ThT2:W32 [ Liver stero127 hydroxylase in hamster: Modulation by
steroids and diets C. Lutton, M. Souidi, S. Dubrac, M. Parquet. Physiologie de la Nutrition,
Universit# Paris-Sud, Orsay, France Objective: To investigate, in hamster liver, the effect of various oxysterols, bile acids or a sucrose-rich versus commercial diet on sterol 27-hydroxylase ($27), an ubiquitous enzymatic activity implicated in anti-atherogenic process and bile acid synthesis. Methods: A sensitive assay (Life Sciences 1999, 64, 1585-1593) was used in order to obtain linearity for the accumulation of labelled 27-hydroxycholesterol in hamster mitochondria. Results: In vitro, epicoprostanol and 27 hydroxycholesterol lowered $27 activity (88 and 29%) while hyodeoxycholic acid and 25 hydroxycholesterol were without effect. A sucrose-rich diet induced a marked decrease in $27 activity (-54%) compared with the commercial diet. In vivo, epicoprostanol and hyodeoxycholic acid lowered ( - 4 6 and 70%) as well as the lithogenic diet ( - 5 6 % ) but 25 and 27 hydroxycholesterol were without effect. Conclusions: Hepatic sterol 27 hydroxylase activity can be modulated by steroids and diets. Researching conditions able to activate this activity might be of great interest.
ThT2:W33 II Inhibition of vascular smooth muscle cell proliferation by red wine polyphenols is associated with downregulation of cyclin a gene expression K. Iiiima, M. Yoshizumi, M. Hashimoto, S. Kim, M. Akishita, J. Ako, Y.Q. Liang, N. Sudoh, T. Watanabe, Y. Ohike, K. Toba, K. Hosoda 1, K. Nakahara I , Y. Ouchi. Department of Geriatric Medicine, The University of
Tokyo; l Suntory Research Center, Osaka, Japan Red wine polyphenols have been shown to contribute to the "French paradox" phenomenon consisting of lower mortality from coronary heart disease in the French population. Although vascular smooth muscle cell (VSMC) proliferation plays an important role in the progression of atherosclerotic lesions, the effects of red wine polyphenols (RWP) on VSMC proliferation have not been elucidated. Methods and Results: To examine the effect of RWP on the growth of VSMC, we extracted a total polyphenolic fraction, called RW-PE from red wine using column chromatography. Cultured rat aortic smooth muscle cells (RASMC) were incubated with RW-PF (1-100 microgram/ml) for 72 hr. In cell count and thymidine uptake assay, treatment with RW-PF showed a potent inhibitory effect on cell growth and DNA synthesis in a dose-dependent manner. In contrast, this polyphenolic fraction did not affect the growth of bovine carotid endothelial cells (BCEC). To elucidate the mechanism of this anti-proliferative effect of RW-PF in RASMC, we investigated expression of cyclin A mRNA and cyclin A promoter activity. RW-PF dramatically dowuregulated the cyclin A mRNA and reduced the cyclin A promoter activity in a dose-dependent manner in RASMC but not in BCEC. In addition, RW-PF decreased the binding of nuclear proteins to the activating transcription factor (ATF) site in the cyclin A promoter, and the expression of transcription factors, cyclic AMP-responsive element-binding protein (CREB) and ATF-1, was also downregulated by RW-PF. Conclusion: The dowuregulation of cyclin A gene expression may contribute to the antiproliferative effect of RWP on VSMC through the inhibition of transcription factor expression. 1
ThT3:W33 ] Magnolol induces a regression of aortic hypertrophy in spontaneously hypertensive rats T.-C. Chou. Graduate of Medical Sciences, National Defense Medical Center,
Taipei, Taiwan Objective: To evaluate whether the Chinese herb, magnolol, induces regres-
T:W33
PROLIFERATION AND DIFFERENTIATION SMOOTH MUSCLE CELLS
OF
|
ThT1:W331J Diffuse intimal thickening emerges in atherosderosis-prone arteries in childhood and the intima/media ratio increases with age Y. Nakashima, Y.X. Chen, K. Sueishi. Pathophysiological and Experimental
Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan Objective: Diffuse intimal thickening (DIT) is considered as an adaptive change of arterial intima and suggested to have a close relation to athero-
sion of hypertrophy and possesses antihypertensive effect in spontaneously hypertensive rats (SHR). Methods: After administration of magnolol (50/zg/kg, p.o.) for 6 weeks, the vascular morphological change, blood pressure, endothelium-dependent response, and the 0 2 formation from aorta in SHR were evaluated. Results: Magnolol resulted in a significant reduction of mean blood pressure compared with that of untreated group in SHR (140.2 :t= 3.7 vs 161.0 :t= 7.0 mmHg, P < 0.05) and the endothelium-dependent relaxation in response to acetylcholine was also improved. In addition, the intima/media thickness and area ratio of aorta were significantly lower in magnolol treated group (Table 1). The production of 0 2 from aorta in SHR was also significantly attenuated by magnolol treatment. Conclusion: Our results show that magnolol possesses beneficial effect of regression of vascular hypertrophy and antihypertensive effect in SHR. In
Xllth International Symposium on Atherosclerosis, Stockholm, Sweden, June 25-29, 2000