- Email: [email protected]
Effects of methionine load on oxidative modification of low density lipoprotein
Thursday June 29, 2000: Poster Abstracts P: W29 Oxidation and Atherogenesis
IThP34:W291
Effects of methionine load low density lipoprotein
I
on
oxidative modification of
A. ~fik, M. Buchffkov~l, A. Pisah'kov~i, S. gomaniv, M. Zeman, S. Stipek. 4 th Dept. Intern. Med., 1st Fac. Med., Charles Univ., Prague, Czech Republic
Objective: Mild hyperhomocyst(e)inemia (hHcy) is an independent risk factor for atherosclerotic vascular diseases. Mechanisms by which hHcy accelerate atherothrombosis are supposed to be multifactorial. Homocyst(e)in (Hcy) and other thiols are able to generate several cytotoxic reactive oxygen species (ROS). Increased generation of ROS initiates lipid peroxidation. Oxidatively modified LDL formed during this process may play important role in the early stages of atherogenesis. Methionine (Met) load leads to a rise of plasma Hcy levels. Methods: Fasting (0 hrs) and postmethionine loading (6 hrs, 0.1 g/kg bw) plasma levels of total Hcy (tHcy) were determined by the HPLC method (Araki and Sako, 1987). Lipoperoxidation of LDL was measured by Cu2+-catalyzed conjugated diene (CD) formation (A234nm) in ultracentrifugally isolated LDL (Esterbauer et al, 1989). Etectrophoretic mobility of LDL before and after in vitro Cu2÷-catalyzed oxidation before and after Met load was measured on agarose (Sebia). Studied group (15 M/8 F) consisted from 8 healthy persons, 9 patients with atherosclerotic vascular disease, and 6 patients with mixed hyperlipidaemia. Results: Post Met-loading changes in levels of tHcy (+152%, P < 0.001), were associated with an increase of concentrations of CD in LDL (+8%, P < 0.10), decrease in duration of lag time ( - 1 2 % , P < 0.01) and increased oxidation rate of LDL (+11% (P < 0.05). Similarly, Met load led to increased electrophoretic mobility of "in vitro" Cu 2+ oxidatively modified LDL (+20%, P < 0.05). Conclusion: Met load, through induced mild hHcy, led to increased amount of CD in LDL, that implicated rise in concentrations of mm-LDL. After Met load decreased duration of lag time, increased oxidation rate of LDL and increased etectrophoretic mobility of LDL were observed as well. The data implicated enhanced oxidation and increased susceptibility of LDL to lipid peroxidation after methionine load. Relation of LDL-cholesteroL, LDL oxidability and antioxidants to endothelium dependent vasodilation in hipercholesterolemia and hypertension
I ThP35:W29
D.S.E Abdaila t , P. Moriet I , EL. Piavnik 2, M.T. Zanella 2, E.C. Pincinato I , M.C. Bertolami 3 . l Faculdade de Ci#ncias Farmac~uticas, Universidade de Sao Paulo; 2 UNIFESP; 31nst. Dante Pazzanese de Cardiologia., S.P, Brazil
Objectives: The purpose of this study was to examine the relationship of LDL-cholesterol, oxidative susceptibility of LDL and antioxidants with the impairment of endothelium dependent vasodilation in hypercholesterolemic (HC), hypertensive (H), hypercholesterolemic/hypertensive (HH) and normolipidemic normotensive subjects (N) (n = 11). Methods: Ascorbate, urate, c~-tocopherol, lycopene and/%carotene were determined by HPLC. The kinetics of LDL oxidation was monitored by incubating LDL with Cu 2+ and following the absorbance in 234 nm for analyzing the lag time, lag rate, log rate and peak time. Mode B ultrasonography was used to measure the blood flow and diameter in response to reactive hyperemia of brachial artery. Results: Plasma ascorbate, lipid soluble antioxidants, the lag time and peak time were lower in H, HC, and HH than in N. The diameter increase percentages in response to hyperemia were 20.4, 12.9, 9.9 and 8.1 for N, H, HC and HH, respectively. LDL cholesterol was negatively correlated to the percentage of diameter increase in response to hyperemia.. Positive correlations were found for the concentration of plasma lipid soluble antioxidants and blood flow increase, as well as, between the lag time of LDL oxidation and the percentage of diameter increase in response to hyperemia. Conclusions: These data suggest that the cholesterol content and oxidability of LDL, as well as, the lipid soluble antioxidants may be important for the impairment of endothelial dependent vasodilation in hypercholesterolemia and hypertension. Supported by FAPESP
,!ThP36:W29
1 The effect of different intensities of exercise o n c¢-tocopherol levels in the plasma and liver of rats I
S. Kinoshita, E. Tsuji. Graduate School of Medical Professions, Kawasaki University of Medical Welfare, Okayama, Japan
Objective: Vitamin E is the major lipid soluble anti-oxidant and may play an important protective role against free radicals produced during exercise.
263
Exercise induces free radical formation in muscles and liver, resulting in oxidative damage, such as lipid peroxidation. The amount of damage depends on exercise intensity, training state and the tissue examined. The present study investigated the effects of exercise intensities on a-tocopherol levels in the plasma and liver of rats. Methods: Male Wistar rats were divided into a sedentary control group and exercise groups. The exercise groups were forced to exercise by treadmill running at 30%, 60% and 80% of maximum oxygen uptake (VO2max) for 120 minutes. One group was sacrificed immediately and others at 6 hours after the exercise period. 0t-Tocopherol levels in the plasma and liver were analyzed by HPLC. Results: In the group sacrificed immediately after the exercise period, plasma ~-tocopherol levels significantly decreased in the 60% and 80%VO2max groups compared to the control. In all exercise groups, plasma ct-tocopherol levels were significantly higher 6 hours after the exercise period than in those sacrificed immediately after the exercise period. On the other hand, liver c~-tocopherol levels decreased in all exercise groups 6 hours after the exercise period compared with those sacrificed immediately after the exercise period. The decrease was especially significant in the 80 VO2max exercise group. Conclusions: Although regular physical exercise is widely accepted to be a protective factor against the development of atherosclerosis, these results indicate that strenuous exercise raises the consumption of a-tocopherol and induces oxidative stress in the liver of rats. 1
T h P 3 7 : W 2 9 J Micronised fenofibrate effect on serum paraoxonase
activity in patients with coronary heart disease Gy. Paragh, I. Seres, Z. Balogh, M. Harangi, L. Illy4s, L Boda, Zs. Varga, P. Kovfics. First Department of Medicine University of Debreeen, Hungary
Objective: To evaluate the effect of micronised fenofibrate on the serum paraoxonase and lipoprotein levels in CHD patients with type II b hyperlipidemia. Methods: 52 patients were involved in the study (36 males, 16 females). The mean BMI was 27.13 5 : 5 . 3 7 kg/m2. The effects of daily 200 mg micronised fenofibrate (Lipidil ®) on serum cholesterol, lipoproteins, triglyceride, apolipoproteins and fibrinogen levels as well as on liver and kidney functions were determined. The serum paraoxonase activity was measured spectropbotometricalty using paraoxon as substrate. Results: During the three month study it was observed that following the effect of micronised fenofibrate, the serum triglyceride level (from 3.25 + 0.66 mmol/l to 1.86 -t- 0.89 mmol/l, p < 0.001) cholesterol level (from 6.7 41.88 mmol/l to 5.83 + 1.4 mmol/1, p < 0.001) were significantly decreased, while the protective high-density lipoprotein (from 1.05 4- 0.23 mmol/l to 1.29 + 0.51 mmol/1; p = 0.005) was significantly increased. The low-density lipoprotein (from 4.22 + 1.83 mmol/l to 3.7 + 1.31 mmol/l; p = 0.028) and apolipoprotein B-100 (from 1.44 + 0.44 G/I to 1.26 ~ 0.40 G/l; p < 0.008) were also significantly decreased while HDL associated apotipoprotein AI (from 1.49 4-4-0.23 G/1 to 1.60 4- 0.26 G/l; p = 0.) was significantly increased. Both the serum paraoxonase activity (189 4- 104 U/I, vs. 260 4- 64 U/l, p < 0.03) and the standardized values for HDL (PON/HDL) were significantly increased (from 162 4- 75 to 191 + 78, p < 0.05) while the PON/apoA ratio (from 131 4- 73 to 161 4- 64 p = 0.39) was not changed significantly. Contusion: The serum paraoxonase activity was lower in CHD patents with type lib hyperlipoproteinaemia compared to age matched healthy control subjects. The three month treatment with daily 200 mg micronised fenofibrate is thought to normalize the lipid profile and improve the antioxidant status by increasing serum paraoxonase activity in these patients.
I ThP38:W29 I Potential role of oxysterols and lysolecithin in the oxidised LDL-induced decrease in albumin synthesis in HEPG2 cells I
E. Bourdon 1, N. Loreau I , J. Davignon 2 , L. Bernier2, D. Blache I . INSERM U498; I Biochimie des lipoproteines, Universitd de Bourgogne, 7, bd Jeanne d'Arc Dijon, France; 2Clinical Research Institute of Montreal, Montreal, Canada
Objectives: Oxidant stress is increasingly thought to be a key element in atherosclerosis especially due to the occurrence of oxidised LDL (OxLDL). Plasma albumin concentration is inversely correlated with the mortality risk and albumin can be therefore considered as an antiatherogenic protein, due in part to its antioxidant properties. In the present work, we aimed to study the influences of OxLDL on albumin synthesis in HepG2 cells.
Xltth International Symposium on Atherosclerosis, Stockholm, Sweden. June 25-29, 2000
IThP34:W291
Effects of methionine load low density lipoprotein
I
on
oxidative modification of
A. ~fik, M. Buchffkov~l, A. Pisah'kov~i, S. gomaniv, M. Zeman, S. Stipek. 4 th Dept. Intern. Med., 1st Fac. Med., Charles Univ., Prague, Czech Republic
Objective: Mild hyperhomocyst(e)inemia (hHcy) is an independent risk factor for atherosclerotic vascular diseases. Mechanisms by which hHcy accelerate atherothrombosis are supposed to be multifactorial. Homocyst(e)in (Hcy) and other thiols are able to generate several cytotoxic reactive oxygen species (ROS). Increased generation of ROS initiates lipid peroxidation. Oxidatively modified LDL formed during this process may play important role in the early stages of atherogenesis. Methionine (Met) load leads to a rise of plasma Hcy levels. Methods: Fasting (0 hrs) and postmethionine loading (6 hrs, 0.1 g/kg bw) plasma levels of total Hcy (tHcy) were determined by the HPLC method (Araki and Sako, 1987). Lipoperoxidation of LDL was measured by Cu2+-catalyzed conjugated diene (CD) formation (A234nm) in ultracentrifugally isolated LDL (Esterbauer et al, 1989). Etectrophoretic mobility of LDL before and after in vitro Cu2÷-catalyzed oxidation before and after Met load was measured on agarose (Sebia). Studied group (15 M/8 F) consisted from 8 healthy persons, 9 patients with atherosclerotic vascular disease, and 6 patients with mixed hyperlipidaemia. Results: Post Met-loading changes in levels of tHcy (+152%, P < 0.001), were associated with an increase of concentrations of CD in LDL (+8%, P < 0.10), decrease in duration of lag time ( - 1 2 % , P < 0.01) and increased oxidation rate of LDL (+11% (P < 0.05). Similarly, Met load led to increased electrophoretic mobility of "in vitro" Cu 2+ oxidatively modified LDL (+20%, P < 0.05). Conclusion: Met load, through induced mild hHcy, led to increased amount of CD in LDL, that implicated rise in concentrations of mm-LDL. After Met load decreased duration of lag time, increased oxidation rate of LDL and increased etectrophoretic mobility of LDL were observed as well. The data implicated enhanced oxidation and increased susceptibility of LDL to lipid peroxidation after methionine load. Relation of LDL-cholesteroL, LDL oxidability and antioxidants to endothelium dependent vasodilation in hipercholesterolemia and hypertension
I ThP35:W29
D.S.E Abdaila t , P. Moriet I , EL. Piavnik 2, M.T. Zanella 2, E.C. Pincinato I , M.C. Bertolami 3 . l Faculdade de Ci#ncias Farmac~uticas, Universidade de Sao Paulo; 2 UNIFESP; 31nst. Dante Pazzanese de Cardiologia., S.P, Brazil
Objectives: The purpose of this study was to examine the relationship of LDL-cholesterol, oxidative susceptibility of LDL and antioxidants with the impairment of endothelium dependent vasodilation in hypercholesterolemic (HC), hypertensive (H), hypercholesterolemic/hypertensive (HH) and normolipidemic normotensive subjects (N) (n = 11). Methods: Ascorbate, urate, c~-tocopherol, lycopene and/%carotene were determined by HPLC. The kinetics of LDL oxidation was monitored by incubating LDL with Cu 2+ and following the absorbance in 234 nm for analyzing the lag time, lag rate, log rate and peak time. Mode B ultrasonography was used to measure the blood flow and diameter in response to reactive hyperemia of brachial artery. Results: Plasma ascorbate, lipid soluble antioxidants, the lag time and peak time were lower in H, HC, and HH than in N. The diameter increase percentages in response to hyperemia were 20.4, 12.9, 9.9 and 8.1 for N, H, HC and HH, respectively. LDL cholesterol was negatively correlated to the percentage of diameter increase in response to hyperemia.. Positive correlations were found for the concentration of plasma lipid soluble antioxidants and blood flow increase, as well as, between the lag time of LDL oxidation and the percentage of diameter increase in response to hyperemia. Conclusions: These data suggest that the cholesterol content and oxidability of LDL, as well as, the lipid soluble antioxidants may be important for the impairment of endothelial dependent vasodilation in hypercholesterolemia and hypertension. Supported by FAPESP
,!ThP36:W29
1 The effect of different intensities of exercise o n c¢-tocopherol levels in the plasma and liver of rats I
S. Kinoshita, E. Tsuji. Graduate School of Medical Professions, Kawasaki University of Medical Welfare, Okayama, Japan
Objective: Vitamin E is the major lipid soluble anti-oxidant and may play an important protective role against free radicals produced during exercise.
263
Exercise induces free radical formation in muscles and liver, resulting in oxidative damage, such as lipid peroxidation. The amount of damage depends on exercise intensity, training state and the tissue examined. The present study investigated the effects of exercise intensities on a-tocopherol levels in the plasma and liver of rats. Methods: Male Wistar rats were divided into a sedentary control group and exercise groups. The exercise groups were forced to exercise by treadmill running at 30%, 60% and 80% of maximum oxygen uptake (VO2max) for 120 minutes. One group was sacrificed immediately and others at 6 hours after the exercise period. 0t-Tocopherol levels in the plasma and liver were analyzed by HPLC. Results: In the group sacrificed immediately after the exercise period, plasma ~-tocopherol levels significantly decreased in the 60% and 80%VO2max groups compared to the control. In all exercise groups, plasma ct-tocopherol levels were significantly higher 6 hours after the exercise period than in those sacrificed immediately after the exercise period. On the other hand, liver c~-tocopherol levels decreased in all exercise groups 6 hours after the exercise period compared with those sacrificed immediately after the exercise period. The decrease was especially significant in the 80 VO2max exercise group. Conclusions: Although regular physical exercise is widely accepted to be a protective factor against the development of atherosclerosis, these results indicate that strenuous exercise raises the consumption of a-tocopherol and induces oxidative stress in the liver of rats. 1
T h P 3 7 : W 2 9 J Micronised fenofibrate effect on serum paraoxonase
activity in patients with coronary heart disease Gy. Paragh, I. Seres, Z. Balogh, M. Harangi, L. Illy4s, L Boda, Zs. Varga, P. Kovfics. First Department of Medicine University of Debreeen, Hungary
Objective: To evaluate the effect of micronised fenofibrate on the serum paraoxonase and lipoprotein levels in CHD patients with type II b hyperlipidemia. Methods: 52 patients were involved in the study (36 males, 16 females). The mean BMI was 27.13 5 : 5 . 3 7 kg/m2. The effects of daily 200 mg micronised fenofibrate (Lipidil ®) on serum cholesterol, lipoproteins, triglyceride, apolipoproteins and fibrinogen levels as well as on liver and kidney functions were determined. The serum paraoxonase activity was measured spectropbotometricalty using paraoxon as substrate. Results: During the three month study it was observed that following the effect of micronised fenofibrate, the serum triglyceride level (from 3.25 + 0.66 mmol/l to 1.86 -t- 0.89 mmol/l, p < 0.001) cholesterol level (from 6.7 41.88 mmol/l to 5.83 + 1.4 mmol/1, p < 0.001) were significantly decreased, while the protective high-density lipoprotein (from 1.05 4- 0.23 mmol/l to 1.29 + 0.51 mmol/1; p = 0.005) was significantly increased. The low-density lipoprotein (from 4.22 + 1.83 mmol/l to 3.7 + 1.31 mmol/l; p = 0.028) and apolipoprotein B-100 (from 1.44 + 0.44 G/I to 1.26 ~ 0.40 G/l; p < 0.008) were also significantly decreased while HDL associated apotipoprotein AI (from 1.49 4-4-0.23 G/1 to 1.60 4- 0.26 G/l; p = 0.) was significantly increased. Both the serum paraoxonase activity (189 4- 104 U/I, vs. 260 4- 64 U/l, p < 0.03) and the standardized values for HDL (PON/HDL) were significantly increased (from 162 4- 75 to 191 + 78, p < 0.05) while the PON/apoA ratio (from 131 4- 73 to 161 4- 64 p = 0.39) was not changed significantly. Contusion: The serum paraoxonase activity was lower in CHD patents with type lib hyperlipoproteinaemia compared to age matched healthy control subjects. The three month treatment with daily 200 mg micronised fenofibrate is thought to normalize the lipid profile and improve the antioxidant status by increasing serum paraoxonase activity in these patients.
I ThP38:W29 I Potential role of oxysterols and lysolecithin in the oxidised LDL-induced decrease in albumin synthesis in HEPG2 cells I
E. Bourdon 1, N. Loreau I , J. Davignon 2 , L. Bernier2, D. Blache I . INSERM U498; I Biochimie des lipoproteines, Universitd de Bourgogne, 7, bd Jeanne d'Arc Dijon, France; 2Clinical Research Institute of Montreal, Montreal, Canada
Objectives: Oxidant stress is increasingly thought to be a key element in atherosclerosis especially due to the occurrence of oxidised LDL (OxLDL). Plasma albumin concentration is inversely correlated with the mortality risk and albumin can be therefore considered as an antiatherogenic protein, due in part to its antioxidant properties. In the present work, we aimed to study the influences of OxLDL on albumin synthesis in HepG2 cells.
Xltth International Symposium on Atherosclerosis, Stockholm, Sweden. June 25-29, 2000