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3.P.362 Angiotensin II increases the cellular uptake of oxidized-LDL, but not of acetylated-LDL by mouse peritoneal macrophages
274
Wednesday 8 October 1997 : Posters Growth factors, cytokines and vaso-active peptides
Subclones from h-TNF transduced cells (RADT) with epithelioid (B, B4, C3,) spindle (A), elongated (B3) and mixed (A4) morphology were also obtained . As expected RAD (unstransduced) and RADTc (transduced with a vector lacking h-TNF cDNA sequence) did not express h-TNF mRNA measured by RT-PCR and Northern blot . Absence of h-TNF, in cell conditioned medium, measured by ELISA and bio-assay was also found . However, RADT cell line and some of their stable subclones (A, B, B3, B4, C3), independenly of their morphology, produce large amounts of h-TNFa (2000 to 5000 pg/ml) . Some clones (B1, B3) produce 300 to 500 pg/ml and others (A2 and A4) less than 100 pg/ml. Uncloned cultures are heterogeneous, but RADT cells proliferated more than RAD and RADTc, even with low serum (2 .5%) and in its absence. Differences in cell adhesion, spreading and cytoskeletal organization were found in comparison with controls . Interestingly, our h-TNF secretory clones of epithelioid type proliferate less than spindle or elongated ones . The expression of the intercellular adhesion molecule-1 (ICAM-1), measured by flow cytometry as relative mean fluorescence intensity (MFI), increased dose-dependently with recombinant rat TNFa (0 to 50 ng/ml) in RAD and RADTc cells (3 to 4-fold in MFI respect to untreated cells) but not in RADT which have similar basal expression than RADTc cells. Since RAD cells did not bind ' 25 1-h-TNF and h-TNF binds to the murine p55 receptor but not to the p75 receptor, we conclude that our model appears suitable to investigate the pleitropic effects of this cytokine and its receptors in SMC phenotypic modulation. Supported by FISS 96/2046 and Laboratorios Dr. Esteve
3 .P.359
sia subgroup were statistically significant by a non-parametric test (One-way Kruskal-Waliis test) . Plasma concentrations of the active form of TGFO-1 were significantly increased in all preeclampsia subgroups as well as in the total group (5 .63 ± 1 .68 ng/ml) compared to controls (4.67 ± 1 .33 ng/ml) . The outcomes of this study may suggest involvement of both parameters in the patophysiology of preeclampsia and substantiate the notion of a multifactorial etiology of the disease . To our knowledge, this is the first study attempting to elucidate the role of Lp(a) lipoprotein and its interaction with the active form of TGFO-1 in preeclampsia .
3 .R361
P. Fraunberger, M . Pfeiffer, B . Siegele, A .K . Walli, D . Seidel. Institute of Clinical Chemistry, Klinikum Grosshadern, University of Munich, 81377 Munich, Germany Inflammation is often associated with hypocholesterolemia, however the underlying mechanism is unclear. Previously we have shown that Tumor necrosis factor (TNF) and soluble TNF receptors correlate with hypocholesterolemia during sepsis. In order to test whether Interleukin-6 (IL-6) plays a role in lowering cholesterol during sepsis, serum levels of IL-6, cholesterol, apolipoprotein Al and B were measured in 32 septic patients . An inverse correlation between IL-6 and cholesterol and also Apo B and Al was found . In addition the effects of TNF and IL-6 on the uptake of cellulare 125 1-LDL as mediators for hypocholesterolemia in inflammatory disease was examined . Both of these cytokines increased the degradation and internalisation of LDL in HepG-2 cells at a concentration of (20 ng/ml) . However after increasing the concentration to 100 ng/ml no further increase could be noted for IL-6 but for TNF. Coincubation of HepG-2 cells with TNF and IL-6 (20 ng/ml respectively) led to stimulation of LDL which was higher than the additive effects of both cytokines . Antibodies to TNF abolished the stimulatory effect on LDL degradation of both TNF and also IL-6 suggesting, that IL-6 increases LDL uptake by induction of TNF and autokrine binding to TNF receptors . Our data substantiate the hypothesis that the hypocholesterolemia of septic patients is mediated by cytokine release . Whether IL-6 directly effects LDL receptor activity or induces TNF, remains to be determined .
Modulatory effect of gliclazide on cytokine production in type II diabetic patients
A .C. Desfaits, O. Serri, G . Renier. Notre-Dame Hospital Research Center, University of Montreal, Canada Several studies have supported an important role of monocytes in the genesis of atherosclerotic lesions in diabetes . However, scanty informations are presently available on the diabetic monocyte function . In the present study, we measured in 8 type II diabetic patients the in vivo and in vitro monocyte cytokine production and the effect of gliclazide treatment on these variables . Poorly controlled, glyburide-treated diabetic patients and healthy control subjects were recruited . At the beginning of the study, glyburide was replaced for three months by an equivalent hypoglycemic dose of gliclazide . Our results show that with the exception of a slight but significant enhancement of tumor necrosis factor a (TNFa) and interleukin-6 (IL-6) in diabetic patients, serum levels of interleukin-1 beta (IL-l fi), insulin-like growth factor-I, vascular endothelial growth factor, fibroblast growth factor b and transforming growth factor beta did not differ in the two groups before gliclazide treatment . Although basal monocyte productions of TNFa, IL-10, IL-6 and IL-8 were similar in controls and diabetic patients, a marked increase in the lipopolysaccharide (LPS)-stimulated monocyte production of TNFa was observed in the diabetic group (4 .6 f 1 .1 vs 2.1 ± 0 .4 i g/ml, P < 0 .01) . At variance, LPS-stimulated monocyte production of IL-10, IL-6 and IL-8 did not differ between the two groups . Gliclazide treatment lowered LPS-stimulated TNFa production by diabetic monocytes to levels similar to those observed in control subjects . These results demonstrate that LPS-stimulated TNFa production by monocytes is enhanced in type II diabetic patients and that gliclazide reduces +k this hyperresponsivity . Supported by Servier.
3 .P.360
3 .P.362
Angiotensin II increases the cellular uptake of oxidized-LDL, but not of acetylated-LDL by mouse peritoneal macrophages
S . Keidar, J. Attias . The Lipid Research Laboratory, Rambam Med Center, The Bruce Rappaport Technion Faculty of Medicine, and the Rappaport Institute for research in the medical sciences, Haifa, Israel Macrophage cholesterol accumulation and the uptake of Ox-LDL by cells of the vessel wall are early events in atherosclerosis . Angiotensin II (Ang II) has been shown to have various atherogenic properties including the ability to induce cell mediated oxidation of LDL . In the present study we examined the effect of Ang II on the uptake of Ox-LDL by peritoneal macrophages from BALB/C mice (MPM). Intraperitoneal injection of Ang II (10 - M) twice a week for a period of 3 months to the mice, resulted in a 150% increment in the uptake of Ox-LDL, compared to macrophages from saline injected mice . No similar effect of Ang II could be demonstrated for the uptake of Ac-LDL by these cells . When Ang II was administrated once daily for a period of 2 days, the MPM exhibited increased uptake of Ox-LDL (20 pglml) by 30%, whereas the uptake of Ox-LDL was not changed . The uptake of Ox-LDL was Ang II dose dependent as studied by injection of 10-7 M-10 -6 M, Ang II to the mice. Similarly Ang II increased by two folds the binding of Ox-LDL to the MPM . Scatchard plot analysis demonstrated'that Ang lI increased the number of Ox-LDL receptors by 32%, and increased also the affinity of Ox-LDL to the receptors by 50% . We conclude that Ang II, in addition to its others known atherogenic properties also enhances the uptake of Ox-LDL by MPM, leading to foam cell formation and accelerated atherosclerosis .
Altered plasma levels of Lp(a) lipoprotein and TGF-$1 in preeclampsia
S . Diurovic, R. Schjetlein, F. Wislt ff, G. Haugen, H . Husby, K . Berg . Ullevdl University Hospital, University of Oslo and The National Hospital of Norway, Oslo, Norway This study was performed in order to investigate the possible association between preeclampsia and the plasma concentrations of Lp(a) lipoprotein and TGF,B-1, respectively. We examined 154 women with preeclampsia (preeclampsia group) and 76 healthy, pregnant normotensive women (control group) . The preeclampsia group was subdivided into : mild preeclampsia, severe preeclampsia and preeclampsia with fetal growth retardation . Plasma levels of Lp(a) lipoprotein were lower in the total preeclampsia group as well as in all preeclampsia subgroups than in the control group as determined by quantitative electroimmunoassay . Corresponding results were obtained with a radioimmunoassay (166 .03 ± 200.2 U/L in the total preeclampsia group vs . 229.18 ± 257 .7 U/L in controls). The differences between controls and the total preeclampsia group as well as each preeclamp-
Interleukin-6 correlates with hypocholesterolemia in inflammatory disease and stimulates LDL receptor activity in HepG-2 cells
1
3 .P.363
Complement activation during percutaneous transluminal coronary angioplasty (PTCA) procedure
J. Kowalski, L . Pawlicki, M . Kosmider, E . Glowacka, Z. Baj, L. Pokoca, E . Sibinska. Department of Lung Diseases Military School of Medicine, Lodz, Poland The aim of the study was to determine the concentration of complement components C3c, C4, C5 and complement hemolytic activity (CH5p) in the
11th International Symposium on Atherosclerosis, Paris, October 1997
Wednesday 8 October 1997 : Posters Growth factors, cytokines and vaso-active peptides
Subclones from h-TNF transduced cells (RADT) with epithelioid (B, B4, C3,) spindle (A), elongated (B3) and mixed (A4) morphology were also obtained . As expected RAD (unstransduced) and RADTc (transduced with a vector lacking h-TNF cDNA sequence) did not express h-TNF mRNA measured by RT-PCR and Northern blot . Absence of h-TNF, in cell conditioned medium, measured by ELISA and bio-assay was also found . However, RADT cell line and some of their stable subclones (A, B, B3, B4, C3), independenly of their morphology, produce large amounts of h-TNFa (2000 to 5000 pg/ml) . Some clones (B1, B3) produce 300 to 500 pg/ml and others (A2 and A4) less than 100 pg/ml. Uncloned cultures are heterogeneous, but RADT cells proliferated more than RAD and RADTc, even with low serum (2 .5%) and in its absence. Differences in cell adhesion, spreading and cytoskeletal organization were found in comparison with controls . Interestingly, our h-TNF secretory clones of epithelioid type proliferate less than spindle or elongated ones . The expression of the intercellular adhesion molecule-1 (ICAM-1), measured by flow cytometry as relative mean fluorescence intensity (MFI), increased dose-dependently with recombinant rat TNFa (0 to 50 ng/ml) in RAD and RADTc cells (3 to 4-fold in MFI respect to untreated cells) but not in RADT which have similar basal expression than RADTc cells. Since RAD cells did not bind ' 25 1-h-TNF and h-TNF binds to the murine p55 receptor but not to the p75 receptor, we conclude that our model appears suitable to investigate the pleitropic effects of this cytokine and its receptors in SMC phenotypic modulation. Supported by FISS 96/2046 and Laboratorios Dr. Esteve
3 .P.359
sia subgroup were statistically significant by a non-parametric test (One-way Kruskal-Waliis test) . Plasma concentrations of the active form of TGFO-1 were significantly increased in all preeclampsia subgroups as well as in the total group (5 .63 ± 1 .68 ng/ml) compared to controls (4.67 ± 1 .33 ng/ml) . The outcomes of this study may suggest involvement of both parameters in the patophysiology of preeclampsia and substantiate the notion of a multifactorial etiology of the disease . To our knowledge, this is the first study attempting to elucidate the role of Lp(a) lipoprotein and its interaction with the active form of TGFO-1 in preeclampsia .
3 .R361
P. Fraunberger, M . Pfeiffer, B . Siegele, A .K . Walli, D . Seidel. Institute of Clinical Chemistry, Klinikum Grosshadern, University of Munich, 81377 Munich, Germany Inflammation is often associated with hypocholesterolemia, however the underlying mechanism is unclear. Previously we have shown that Tumor necrosis factor (TNF) and soluble TNF receptors correlate with hypocholesterolemia during sepsis. In order to test whether Interleukin-6 (IL-6) plays a role in lowering cholesterol during sepsis, serum levels of IL-6, cholesterol, apolipoprotein Al and B were measured in 32 septic patients . An inverse correlation between IL-6 and cholesterol and also Apo B and Al was found . In addition the effects of TNF and IL-6 on the uptake of cellulare 125 1-LDL as mediators for hypocholesterolemia in inflammatory disease was examined . Both of these cytokines increased the degradation and internalisation of LDL in HepG-2 cells at a concentration of (20 ng/ml) . However after increasing the concentration to 100 ng/ml no further increase could be noted for IL-6 but for TNF. Coincubation of HepG-2 cells with TNF and IL-6 (20 ng/ml respectively) led to stimulation of LDL which was higher than the additive effects of both cytokines . Antibodies to TNF abolished the stimulatory effect on LDL degradation of both TNF and also IL-6 suggesting, that IL-6 increases LDL uptake by induction of TNF and autokrine binding to TNF receptors . Our data substantiate the hypothesis that the hypocholesterolemia of septic patients is mediated by cytokine release . Whether IL-6 directly effects LDL receptor activity or induces TNF, remains to be determined .
Modulatory effect of gliclazide on cytokine production in type II diabetic patients
A .C. Desfaits, O. Serri, G . Renier. Notre-Dame Hospital Research Center, University of Montreal, Canada Several studies have supported an important role of monocytes in the genesis of atherosclerotic lesions in diabetes . However, scanty informations are presently available on the diabetic monocyte function . In the present study, we measured in 8 type II diabetic patients the in vivo and in vitro monocyte cytokine production and the effect of gliclazide treatment on these variables . Poorly controlled, glyburide-treated diabetic patients and healthy control subjects were recruited . At the beginning of the study, glyburide was replaced for three months by an equivalent hypoglycemic dose of gliclazide . Our results show that with the exception of a slight but significant enhancement of tumor necrosis factor a (TNFa) and interleukin-6 (IL-6) in diabetic patients, serum levels of interleukin-1 beta (IL-l fi), insulin-like growth factor-I, vascular endothelial growth factor, fibroblast growth factor b and transforming growth factor beta did not differ in the two groups before gliclazide treatment . Although basal monocyte productions of TNFa, IL-10, IL-6 and IL-8 were similar in controls and diabetic patients, a marked increase in the lipopolysaccharide (LPS)-stimulated monocyte production of TNFa was observed in the diabetic group (4 .6 f 1 .1 vs 2.1 ± 0 .4 i g/ml, P < 0 .01) . At variance, LPS-stimulated monocyte production of IL-10, IL-6 and IL-8 did not differ between the two groups . Gliclazide treatment lowered LPS-stimulated TNFa production by diabetic monocytes to levels similar to those observed in control subjects . These results demonstrate that LPS-stimulated TNFa production by monocytes is enhanced in type II diabetic patients and that gliclazide reduces +k this hyperresponsivity . Supported by Servier.
3 .P.360
3 .P.362
Angiotensin II increases the cellular uptake of oxidized-LDL, but not of acetylated-LDL by mouse peritoneal macrophages
S . Keidar, J. Attias . The Lipid Research Laboratory, Rambam Med Center, The Bruce Rappaport Technion Faculty of Medicine, and the Rappaport Institute for research in the medical sciences, Haifa, Israel Macrophage cholesterol accumulation and the uptake of Ox-LDL by cells of the vessel wall are early events in atherosclerosis . Angiotensin II (Ang II) has been shown to have various atherogenic properties including the ability to induce cell mediated oxidation of LDL . In the present study we examined the effect of Ang II on the uptake of Ox-LDL by peritoneal macrophages from BALB/C mice (MPM). Intraperitoneal injection of Ang II (10 - M) twice a week for a period of 3 months to the mice, resulted in a 150% increment in the uptake of Ox-LDL, compared to macrophages from saline injected mice . No similar effect of Ang II could be demonstrated for the uptake of Ac-LDL by these cells . When Ang II was administrated once daily for a period of 2 days, the MPM exhibited increased uptake of Ox-LDL (20 pglml) by 30%, whereas the uptake of Ox-LDL was not changed . The uptake of Ox-LDL was Ang II dose dependent as studied by injection of 10-7 M-10 -6 M, Ang II to the mice. Similarly Ang II increased by two folds the binding of Ox-LDL to the MPM . Scatchard plot analysis demonstrated'that Ang lI increased the number of Ox-LDL receptors by 32%, and increased also the affinity of Ox-LDL to the receptors by 50% . We conclude that Ang II, in addition to its others known atherogenic properties also enhances the uptake of Ox-LDL by MPM, leading to foam cell formation and accelerated atherosclerosis .
Altered plasma levels of Lp(a) lipoprotein and TGF-$1 in preeclampsia
S . Diurovic, R. Schjetlein, F. Wislt ff, G. Haugen, H . Husby, K . Berg . Ullevdl University Hospital, University of Oslo and The National Hospital of Norway, Oslo, Norway This study was performed in order to investigate the possible association between preeclampsia and the plasma concentrations of Lp(a) lipoprotein and TGF,B-1, respectively. We examined 154 women with preeclampsia (preeclampsia group) and 76 healthy, pregnant normotensive women (control group) . The preeclampsia group was subdivided into : mild preeclampsia, severe preeclampsia and preeclampsia with fetal growth retardation . Plasma levels of Lp(a) lipoprotein were lower in the total preeclampsia group as well as in all preeclampsia subgroups than in the control group as determined by quantitative electroimmunoassay . Corresponding results were obtained with a radioimmunoassay (166 .03 ± 200.2 U/L in the total preeclampsia group vs . 229.18 ± 257 .7 U/L in controls). The differences between controls and the total preeclampsia group as well as each preeclamp-
Interleukin-6 correlates with hypocholesterolemia in inflammatory disease and stimulates LDL receptor activity in HepG-2 cells
1
3 .P.363
Complement activation during percutaneous transluminal coronary angioplasty (PTCA) procedure
J. Kowalski, L . Pawlicki, M . Kosmider, E . Glowacka, Z. Baj, L. Pokoca, E . Sibinska. Department of Lung Diseases Military School of Medicine, Lodz, Poland The aim of the study was to determine the concentration of complement components C3c, C4, C5 and complement hemolytic activity (CH5p) in the
11th International Symposium on Atherosclerosis, Paris, October 1997