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Evidence for the involvement of specific regions of AI apolipoprotein in mobilisation of intracellular cholesterol
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Oral session abstracts / Atherosclerosis 115 (Suppl.) (1995) $3-$42 017 Treatment of Hyperlipidaemia
101
102
EFFECTS OF PIOGLITAZONE ON SERUM LIPIDS AND ATHEROSCLEROS1S IN RABBITS WITH HYPERCHOLESTEROLEMIA J~ Sato, T. Inamura, H. Nishimura, S. Shirabe, M. Kanazawa, Y. Notoya, T. Hayashi, H. Ito Dept. of Internal. Med., Tokyo Med. Coll.
CLINICAL UTILITY OF THE H.E.L.P.-TREATMENT IN HEART TRANSPLANT PATIENTS U. BrandP, C. Engelschalk2, B. Meiser ~, K. Wenke ~, J. Thiery2, D. SeideF, B. Reichart ~ Clinic for Heart Surgery' and Institute of Clinical Chemistry~, University Hospital GroBhadern, Munich, FRG
A new oral agent, Pioglitazone has been developed for the treatment of NIDDM. In the present study, we administered Pioglitazone to rabbits with hypercholesterolemia, in order to determine its effects on serum lipids and atherosclerosis. Methods: Male Japan White rabbits (2.5 Kg) were used. The animals were fed with diet containing 0.5% cholesterol (group HC) or 0.5% cholesterol and 300 p.p.m. Pioglitazone (group HP) at 200 g/day. Serum lipids were measured at the start and after 10 weeks of treatment. After 10 weeks, the descending aorta was removed under anesthesia, and the lipid deposit lesion area in the intima was measured. Results: As for serum lipids, T-CHO significantly increased from the pretreatment level in the HC group at 10 weeks, whereas HDL-C and T-G significantly decreased. T-CHO significantly increased from the pretreatment level in the HP group at 10 weeks; HDL-C decreased slightly, but not significantly; T-G significantly decreased. There were no significant differences in T-CHO, T-G or LDL-C between the two groups at 10 weeks, but HDL-C was significantly higher, and the lipid deposit lesion area in the intima was significantly lower in the HP group. Sununary: These results suggest that Pioglitazone has an inhibitory, effect on atherosclerosis in rabbits with hypercholesterolemia.
We report on the clinical long-term follow up of 13 hearttransplant (HTX) patients, which were treated with the Heparin-mediated Extracorporeal Low Density Precipitation (H.E.L.P.) for up to two years (n=500 treatments). Patients were suffering from severe hypercholesterolemia and were on treatment with 10-15 mg Simvastatin/day. The aim of the H.E.L,P.-therapy was to lower the LDLcholesterol levels below 110mg/dl and to reduce drastically lipoprotein(a) and fibrinogen in plasma. The table shows the effects of regular H.E.L.P.-therapy in these patients:
mg/dl; mean:t-S.E.M Total Cholesterol LDL-Cholesterol Lp(a) Fibrinogen
before H.E.L.P 251+11 161 + 11 44+9 4195:17
after H.E.L.P. 1405:9 765-7 155:2 1765:21
Mean Interval 190 114 26 265
The weekly/biweekly apheresis revealed no acute or chronic changes of the plasma levels of different cytokines. It did not influence the pharmakokinetic of cyclosporin A and had no effect on the number of rejection episodes. From our first clinical experiences the H.E.L.P.-apheresis may be a helpful tool to achieve long lasting transplantation and to retard the development of graft arteriosclerosis.
O18 CHOLESTEROL A N D PHOSPHOLIPID TRANSPORT 103
104
DIVERGENT EFFECTS OF APOPROTEIN All ON REMOVAL AND UPTAKE OF CHOLESTEROL IN CULTURED CELLS O. Stein ~, Y. Dabach t, G. HollandeP, M. Ben-Naim j, K. Oette:, Y Stein ~ ~Hadassah University Hospital, Jerusalem, Israel, -'University of Cologne. Cologne, Germany
EVIDENCE FOR THE INVOLVEMENT OF SPECIFIC REGIONS OF A1 APOLIPOPROTEIN IN MOBILISATION OF INTRACELLULAR CHOLESTEROL N.H. Fidge, D. Sviridov Baker Medical Research Institute, Melbourne, Australia
The role of apo All in reverse cholesterol transport (RCT) as promoted by apo AI. and rhapo AIV and E was studied Free cholesterol (FC) donors were mouse peritoneal macrophages (MP), bovine aortic smooth muscle (SMC) and human skin fibroblasts (HSF). labeled with ~H-FC. Acceptors of FC were dioleoyl phosphatidylcholine (DOPC) liposomes containing apo AI, AIV or E alone or together with apo All. When ~H-FC labeled MP were incubated for 2 or 4 h with equimolar apo A1, AII. A1V or E. the lowest efflux occurred with apo All. Exposure of 3H-FC MP to liposomes containing apo AI/AII at 1:2 M/M (50 ~g protein/ml), resulted in a lower 3H-FC efflux as compared to apo AI alone. Apo All added to apo E had no effect on FC efflux. However, when apo AI or A1V protein concen tration was kept constant and supplemented with apo AII. a lower ~H-FC efflux was found only at 1:3 M/M of apo AI:AII. With aortic SMC and HSF. no inhibitory effect of addition of apo All to apo AI or apo AIV on RCT was seen at apo AI/AII of 1:1 or 1:2 M/M When efflux media containing macrophage derived ~H-FC were added to HepG: cells and SMC. the 3H-FC in the cells was higher when apo All had been added to apo A1 or apo AIV than when the apoproteins were added alone. Thus the effect of apo All on RCT is cell-type dependent and its proatherogenic effect may be due to enhanced delivery of cholesterol to cells.
The traffic in cholesterol between ceils and lipoproteins, particularly high density lipoprotein (HDL), plays an important role in cholesterol homeostasis in the body. Perturbations of this system may influence the development of atherosclerosis. Recent evidence suggests that HDL may not only act as an acceptor of cell cholesterol, but may also influence the redistribution of cholesterol between cellular pools as a prerequisite to cholesterol efflux. These events may involve cellular signalling and the participation of putative HDL receptors. Because liver membranes have high affinity sites for apoAl (kd = 3x10-gM) we probed apoAl for structural sites specifically involved in cholesterol efflux. HepG2 cells labelled with 14C-cholesterol predominantly in the endoplasmic reticulum (ER) or plasma membrane (PM) were incubated with plasma, HDL or apoAI-PL as cholesterol acceptors in the presence or absence of monoclonal antibodies (Fab fragments) recognising epitopes at various sites, e.g. AI-I [27-48], AI-3, [140-147], AI-4.1 [211-222] and AI~i.2 [149-150]. With plasma as acceptor none of the antibodies affected the efflux of cholesterol from the PM pool, whereas the addition of McAbs A1-3 or AI-4.2 reduced efflux of cholesterol from the ER by 35%. In combination, AI-3 and AI-4.2 McAbs produced a 60% inhibition of cholesterol efflux from the ER. AI-I and AI-4.1 had no effect on intracellular mobilisation of cholesterol. When HDL, not plasma, was added as acceptor, the addition of AI-3 produced a 65% inhibition of efflux. These data suggest that a specific structural site in apoAl near residues 140-150 stimulates cholesterol effiux by a signalling mechanism that involves mobilisation of intracellular sterol. By contrast, no specific AI sites are involved in the transfer of cholesterol from PM to HDL.
Oral session abstracts / Atherosclerosis 115 (Suppl.) (1995) $3-$42 017 Treatment of Hyperlipidaemia
101
102
EFFECTS OF PIOGLITAZONE ON SERUM LIPIDS AND ATHEROSCLEROS1S IN RABBITS WITH HYPERCHOLESTEROLEMIA J~ Sato, T. Inamura, H. Nishimura, S. Shirabe, M. Kanazawa, Y. Notoya, T. Hayashi, H. Ito Dept. of Internal. Med., Tokyo Med. Coll.
CLINICAL UTILITY OF THE H.E.L.P.-TREATMENT IN HEART TRANSPLANT PATIENTS U. BrandP, C. Engelschalk2, B. Meiser ~, K. Wenke ~, J. Thiery2, D. SeideF, B. Reichart ~ Clinic for Heart Surgery' and Institute of Clinical Chemistry~, University Hospital GroBhadern, Munich, FRG
A new oral agent, Pioglitazone has been developed for the treatment of NIDDM. In the present study, we administered Pioglitazone to rabbits with hypercholesterolemia, in order to determine its effects on serum lipids and atherosclerosis. Methods: Male Japan White rabbits (2.5 Kg) were used. The animals were fed with diet containing 0.5% cholesterol (group HC) or 0.5% cholesterol and 300 p.p.m. Pioglitazone (group HP) at 200 g/day. Serum lipids were measured at the start and after 10 weeks of treatment. After 10 weeks, the descending aorta was removed under anesthesia, and the lipid deposit lesion area in the intima was measured. Results: As for serum lipids, T-CHO significantly increased from the pretreatment level in the HC group at 10 weeks, whereas HDL-C and T-G significantly decreased. T-CHO significantly increased from the pretreatment level in the HP group at 10 weeks; HDL-C decreased slightly, but not significantly; T-G significantly decreased. There were no significant differences in T-CHO, T-G or LDL-C between the two groups at 10 weeks, but HDL-C was significantly higher, and the lipid deposit lesion area in the intima was significantly lower in the HP group. Sununary: These results suggest that Pioglitazone has an inhibitory, effect on atherosclerosis in rabbits with hypercholesterolemia.
We report on the clinical long-term follow up of 13 hearttransplant (HTX) patients, which were treated with the Heparin-mediated Extracorporeal Low Density Precipitation (H.E.L.P.) for up to two years (n=500 treatments). Patients were suffering from severe hypercholesterolemia and were on treatment with 10-15 mg Simvastatin/day. The aim of the H.E.L,P.-therapy was to lower the LDLcholesterol levels below 110mg/dl and to reduce drastically lipoprotein(a) and fibrinogen in plasma. The table shows the effects of regular H.E.L.P.-therapy in these patients:
mg/dl; mean:t-S.E.M Total Cholesterol LDL-Cholesterol Lp(a) Fibrinogen
before H.E.L.P 251+11 161 + 11 44+9 4195:17
after H.E.L.P. 1405:9 765-7 155:2 1765:21
Mean Interval 190 114 26 265
The weekly/biweekly apheresis revealed no acute or chronic changes of the plasma levels of different cytokines. It did not influence the pharmakokinetic of cyclosporin A and had no effect on the number of rejection episodes. From our first clinical experiences the H.E.L.P.-apheresis may be a helpful tool to achieve long lasting transplantation and to retard the development of graft arteriosclerosis.
O18 CHOLESTEROL A N D PHOSPHOLIPID TRANSPORT 103
104
DIVERGENT EFFECTS OF APOPROTEIN All ON REMOVAL AND UPTAKE OF CHOLESTEROL IN CULTURED CELLS O. Stein ~, Y. Dabach t, G. HollandeP, M. Ben-Naim j, K. Oette:, Y Stein ~ ~Hadassah University Hospital, Jerusalem, Israel, -'University of Cologne. Cologne, Germany
EVIDENCE FOR THE INVOLVEMENT OF SPECIFIC REGIONS OF A1 APOLIPOPROTEIN IN MOBILISATION OF INTRACELLULAR CHOLESTEROL N.H. Fidge, D. Sviridov Baker Medical Research Institute, Melbourne, Australia
The role of apo All in reverse cholesterol transport (RCT) as promoted by apo AI. and rhapo AIV and E was studied Free cholesterol (FC) donors were mouse peritoneal macrophages (MP), bovine aortic smooth muscle (SMC) and human skin fibroblasts (HSF). labeled with ~H-FC. Acceptors of FC were dioleoyl phosphatidylcholine (DOPC) liposomes containing apo AI, AIV or E alone or together with apo All. When ~H-FC labeled MP were incubated for 2 or 4 h with equimolar apo A1, AII. A1V or E. the lowest efflux occurred with apo All. Exposure of 3H-FC MP to liposomes containing apo AI/AII at 1:2 M/M (50 ~g protein/ml), resulted in a lower 3H-FC efflux as compared to apo AI alone. Apo All added to apo E had no effect on FC efflux. However, when apo AI or A1V protein concen tration was kept constant and supplemented with apo AII. a lower ~H-FC efflux was found only at 1:3 M/M of apo AI:AII. With aortic SMC and HSF. no inhibitory effect of addition of apo All to apo AI or apo AIV on RCT was seen at apo AI/AII of 1:1 or 1:2 M/M When efflux media containing macrophage derived ~H-FC were added to HepG: cells and SMC. the 3H-FC in the cells was higher when apo All had been added to apo A1 or apo AIV than when the apoproteins were added alone. Thus the effect of apo All on RCT is cell-type dependent and its proatherogenic effect may be due to enhanced delivery of cholesterol to cells.
The traffic in cholesterol between ceils and lipoproteins, particularly high density lipoprotein (HDL), plays an important role in cholesterol homeostasis in the body. Perturbations of this system may influence the development of atherosclerosis. Recent evidence suggests that HDL may not only act as an acceptor of cell cholesterol, but may also influence the redistribution of cholesterol between cellular pools as a prerequisite to cholesterol efflux. These events may involve cellular signalling and the participation of putative HDL receptors. Because liver membranes have high affinity sites for apoAl (kd = 3x10-gM) we probed apoAl for structural sites specifically involved in cholesterol efflux. HepG2 cells labelled with 14C-cholesterol predominantly in the endoplasmic reticulum (ER) or plasma membrane (PM) were incubated with plasma, HDL or apoAI-PL as cholesterol acceptors in the presence or absence of monoclonal antibodies (Fab fragments) recognising epitopes at various sites, e.g. AI-I [27-48], AI-3, [140-147], AI-4.1 [211-222] and AI~i.2 [149-150]. With plasma as acceptor none of the antibodies affected the efflux of cholesterol from the PM pool, whereas the addition of McAbs A1-3 or AI-4.2 reduced efflux of cholesterol from the ER by 35%. In combination, AI-3 and AI-4.2 McAbs produced a 60% inhibition of cholesterol efflux from the ER. AI-I and AI-4.1 had no effect on intracellular mobilisation of cholesterol. When HDL, not plasma, was added as acceptor, the addition of AI-3 produced a 65% inhibition of efflux. These data suggest that a specific structural site in apoAl near residues 140-150 stimulates cholesterol effiux by a signalling mechanism that involves mobilisation of intracellular sterol. By contrast, no specific AI sites are involved in the transfer of cholesterol from PM to HDL.