Ph.D. Thesis. Anti-Cholesterol Antibodies - ACHA - Anna Horváth M.D. Semmelweis University 2002.

Átírás

1 Ph.D. Thesis Anti-Cholesterol Antibodies - ACHA - Anna Horváth M.D. Semmelweis University 2002.

2 SUMMARY In animal experiments the protective role of anti-cholesterol antibodies (ACHA) has been demonstrated. ACHA is a natural antibody against 3B-OH group of cholesterol. Despite the fact that ACHA present in the serum of all healthy humans, no data on the occurrence of these antibodies in human vascular diseases are available. In our experiments we proved with a new antigen that the protective effects of immunization against arteriosclerosis in rabbits belongs to ACHA. ACHA binding to solid phase cholesterol may occur without contribution of serum cofactor. Human ACHA interact with cholesterol present in low-density lipoprotein fractions (LDL and VLDL). We showed that ACHA and anti-oxldl antibodies differ from each other. We measured ACHA levels in patients with various vascular diseases of atherosclerotic origin. ACHA levels were significantly lower in patients with peripheral atherosclerosis and cerebrovascular diseases than in healthy controls. ACHA levels were significantly higher in patients with coronary heart disease and in patients with hyperlipideme than in healthy controls. ACHA levels are higher in HIV positive patients than in controls, HAART induced a marked drop of ACHA concentration. From our results the hypothesis of ACHA are protective antibodies may be true, but high level of ACHA do not save us from atherosclerotis diseases. 2

3 AIMS 1. Influence of new cholesterol antigen immunisation on rabbits atherosclerosis induced by a high cholesterol diet? 2. What is the specificity of the human ACHA? 3. Do ACHA and oxldl antibody levels correlate? 4. Is there a co-factor dependence for ACHA? 5. What are the serum ACHA levels in diseases with atherosclerotic origin (Coronary Heart Diseases, Cerebro Vascular Diseases, Peripheral sclerosis)? 6. Detection of HIV pozitiv patients serum ACHA levels before and after HAART. 1. Immunisation of rabbits METHODS 2. Immunkomplex titer 3. RPR 4. Kolmer reaction 5. LDL és VLDL/LDL separation 6. ELISA (Enisme linked immunosorbent assay). 7. Statistical analyzises (Mann-Whitney test, Fisher s test, Wilcoxon test, Kruskal- Wallis test, Spearman correlation) 3

4 RESULTS Influence of new cholesterol antigen immunisation on rabbits atherosclerosis induced by a high cholesterol diet By immunisation with a new cholesterol antigen ACHA can be induced in rabbits. At the and of the 24th week there were significant difference among the serum cholesterol levels (1,3 0,3 vs. 14,43 1,3 mmol/l) (Mann-Whitney test: p<0.0001) in the immunised and non immunised groups. With pathológical examinations we did not find any plaque in the immunised groups annimals while the non-immunised but high cholesterol diet groups rabbits had severe atherosclerotic lasions on there aortas. Specificity study of the human ACHA On the cholesterol coated ELISA plates purified intravenosus immunglobulin (IVIG) showed dose-dependent binding. LDL dose-dependently inhiboted the binding of ACHA to solid phase cholesterol. Same did VLDL/LDL preperations. On the characterization of the specificity of human ACHA we compared coprostane, ergosterol and cholesterol. About the same amount of IgG binding was measured to cholesterol and ergosterol whereas only weak binding to the coprostanecoated wells was observed. Healthy human populations ACHA levels The levels of ACHA were measured in 218 healthy subjects. There were no significant difference between female and male donors in serum ACHA levels. A significant positive correlation (R=0.1924, p=0.0402) between the ages of the donors and the serum ACHA levels were observed. ACHA levels in diseases with atherosclerotic origin ACHA levels were determinated in the sera of 86 patients with femoro-popliteal reocclusion after reconstructive surgery. We found significantly (p<0,0001) lower 4

5 ACHA levels compared with the control group. We found a significant negative correlation (p=0.014, r=-0267) between serum concentrations of Lp(a) and ACHA. In the group of 146 patient with cerenrovascular disease significantly lower ACHA levels were found compared with healthy controls (p<0.0001). The 341 patients with severe coronary heart diseases had significantly (p<0.0001) higher ACHA levels compared with healthy controls. The newly diagnostised 37 patients with primer combined tipe hyperlipidaemie without any lipid lowering therapy had significantly (p=0.0085) higher ACHA levels compared with healthy controls. ACHA levels in patients with HIV infection ACHA levels were almost 4 times higher in the sera of 46 patients than in 110 controls. No difference in the specificity of ACHA was found between HIV seropositive and HIV seronegative sera. Binding of ACHA to cholesterol-cated plates from HIV seropositive serum was dose-dependly inhibited by preincubation with HIV-1 BA-L preparation. Serum concentration of ACHA was significantly higher in the patients with low seum sholesterol levels than in those with normal cholesterol levels. HAART induced a marked drop of ACHA concentration. 5

6 NEW FINDINGS The new cholesterol antigen can prevent rabbits from high cholesterol diet inducet atherosclerosis. The human ACHA binds the cholesterol without co-factors. Patients with femoro-popliteal reocclusion have lower ACHA levels than the control group. Patient with cerenrovascular disease significantly lower ACHA levels were found compared with healthy controls. Patients with severe coronary heart diseases had significantly higher ACHA levels compared with healthy controls. Patients with primer combined tipe hyperlipidaemie had significantly higher ACHA levels compared with healthy controls. The HIV seropozitiv patients had significantly higher ACHA levels compared with healthy controls. HAART induced a marked drop of ACHA concentration among HIV seropozitiv patients. 6

7 PUBLICATIONS [I.] István Horváth, B. Szende, Anna Horváth, I. Kocsis, A. Horváth: Influence of cholesterol liposome immunisation and immunostimulation on rabbits atherosclerosis induced by a high cholesterol diet, Med.Sci Monit, 1998; 4(3): [II.] Füst György, Harcos Péter, Duba Jenő, Kramer Judit, Prohászka Zoltán, Horváth Laura, Horváth Anna, Karádi István, Mahavir Singh, Császár Albert, Romics László: Hasonlóságok és különbségek a cardiovasculáris és cerebrovasculáris megbetegedések genetikai hátterében és immunológiai mechanizmusában, Magyar Belorvosi Archivum 1999;52: [III.] Anna Horváth, George Füst, István Horváth, Gábor Vallus Jenő Duba, Péter Harcos, Zoltán Prohászka, Éva Rajnavölgyi, Lívia Jánoskuti, Margit Kovács, Albert Császár, László Romics and István Karádi : Anti-cholesterol antibody (ACHA) levels in patients with different atherosclerotic vascular diseases. Characterization of human ACHA. Atherosclerosis 2001 May, 156: (IF: 2,877) [IV.] Anna Horváth, Dénes Bánhegyi, Adrien Bíró, Eszter Ujhelyi, Amarilla Veres, Laura Horváth, Zoltán Prohászka, Attila Bácsi, Vera Tarján, László Romics, István Horváth, Ferenc D. Tóth, George Füst; István Karádi: High Level of Anticholesterol Antibodies (ACHA) in HIV Patients. Normalization of Serum ACHA Concentration after Introduction of HAART. Immunbiology 2001;203, (IF: 2,416) [V.] Anna Horváth, Adrienn Bíró, George Füst, Margit Kovács, Bíborka Bereczky, Zsuzsanna Nébenfürer, Mónika Kleiber, István Horváth, István Karádi and László Romics: Anti-cholesterol antibodies (ACHA) and antibodies against oxidized low density lipoproteins (oxldl-ab) in the sera of patients with combined hyperlipidemia (send for publication to Chlinical Chemistry in 2002.) [VI.] István Horváth, Éva Puskás, Anna Horváth, Éva Balázs, Attila Horváth: Autoantitest, vagy immunológiai védekezés koleszterinnel szemben?, Orvosi Hetilap, 1994; 135(18): [VII.] György Nagy, Anna Horváth George Füst, László Romics, Péter Gergely and István Karádi: Anti-cholesterol antibodies in patients with systemic lupus erythematosus, Lupus , [VIII.] Amarilla Veres, George Füst, Matthew McQueen, Marek Smieja, Anna Horváth, Adrienn Bíró, Janice Pogue, László Romics, István Karádi, Mahavir Singh, Judy Gnarpe, Zoltán Prohászka, Salim Yusuf: Relationship of anti-60kda Heat Shock Protein and anti-cholesterol antibodies to cardiovascular events (send for publication to Circulation in 2002.) 7

8 [IX.] Horváth Anna, Bertók L., Horváth I., Hortoványi E., Lapis K., Hársing J., Kocsis I., Bozóky Z., Egri G. and Greskovits D.: Effects of Tolerin, a detoxified endotoxin on the immunisation against Atherosclerosis (send for publication to Patological Research Practice in 2001.) [X.] Horváth Anna, Prohászka Zoltán, Duba Jenő, Harcos Péter, Vallus Gábor, Horváth István, Romics László, Füst György, Karádi István: Anti-koleszterin antitestek összehasonlító vizsgálata atherosclerotikus érbetegekben Magyar Immunológiai Társaság XXVIII. Kongresszusa, Harkány 1998 Absztrakt füzet 30.o. [XI.] Anna Horváth, L. Varga, I. Horváth, M Kókai, I. Karádi, L. Romics, G. Füst: Complement activation in human sera by liposomes made from cholesterol, Mol. Immunology, 1998; 35(6-7):102 [XII.] Horváth Anna, Prohászka Zoltán, Duba Jenő, Harcos Péter, Vallus Gábor, Horváth István, Romics László, Füst György, Karádi István: Anti-koleszterin antitestek összehasonlító vizsgálata atherosclerotikus érbetegekben Magyar Atherosclerosis Társaság XIII. Kongresszusa, Sopron Absztrakt füzet 9/a. [XIII.] Horváth Anna: Anti-koleszterin antitest (ACHA) mint új marker az érelmeszesedésben, Doktoranduszok II. Országos konferenciája, Debrecen Abstrakt füzet 19.o. [XIV.] Horváth Anna., Prohászka Z., Duba J., Harczos P., Horváth I., Vallus G., Romics L., Füst Gy., Karádi I.: Anti-koleszterin antitest szintjének összehasonlító vizsgálata atherosclerotikus érbetegekben, Magyar Belorvosi Archivum 1998/3: 239. [XV.] Anna Horváth, Gábor Vallus, Zoltán Prohászka, István Horváth, Béla Dlustus, Árpád Baranyai, László Romics, George Füst, István Karádi: Anti-cholesterol antibody levels in healthy subjects and in patients with peripheral atherosclerotic disease, 1999, 70 th EAS Congress Abstract Book p.74. [XVI.] Anna Horváth, George Füst, István Horváth, Gábor Vallus Jenő Duba, Péter Harcos, Zoltán Prohászka, Éva Rajnavölgyi, Lívia Jánoskuti, Margit Kovács, Albert Császár, László Romics and István Karádi: Coronary heart disease differs from other atherosclerosic vascular diseases in one of the immunological aspects (anti-cholesterol antibody levels), European Journal of Internal Medicine 10, 1999 s167 [XVII.] Horváth Anna, Füst G., Horváth I., Prohászka Z., Rajnavölgyi É., Kovács M., Romics L., Karádi i: Characterization of human anti-cholesterol antibodies, Immunology Letters 73/ [XVIII.] Horváth Anna, Bánhegyi Dénes, Füst György, Újhelyi Eszter, Prohászka Zoltán, Horváth István, és Karádi István: Anti-koleszterin antitest szintek HIV fertőzött betegekben MIT XXIX. Kongresszusa, Bük 1999 Absztrakt füzet E- Sz-33. [XIX.] Horváth Anna, Bíró Adrienn, Füst György, Veres Amarilla, Kovács Margit, Horváth István és Karádi István: Anti-koleszterin antitest és oxldl ellenes 8

9 antitestek összehasonlító vizsgálata hypertrigliceridaemiás és egészséges egyének szérumában, MIT XXX. Kongresszusa, Budapest Absztrakt füzet E/24 [XX.] Horváth Anna, Bíró Adrienn, Füst György, Veres Amarilla, Kovács Margit, Horváth István és Karádi István: Anti-koleszterin antitest és oxldl ellenes antitestek összehasonlító vizsgálata hypertrigliceridaemiás és egészséges egyének szérumában, Magyar Atherosclerosis Társaság XIII. Kongresszusa, Sopron [XXI.] Anna Horváth, Adrienn Bíró, George Füst, Amarilla Veres, Margit Kovács, Zsuzsa Nebenfürer, Bíborka Bereczky, István Horváth István Karádi and László Romics: The level of anti-cholesterol antibody and anti-oxidized LDL antibody in serum of patients with Hypertrigliceridaemia and of healthy controls 3rd EFIM (European Federation of Internal Medicine) Congress Edinburgh 2001 [XXII.] Anna Horváth, T. Glasz, E. Hortoványi, G. Füst, I. Karádi, A. Kádár, F. Horkay, G. Mózes, A. Szik, I. Sziller, B. Nagy, Z. Bán, A. Tóth, I. Kassai, G. Dudás: Infection of restenotic coronary bypass grafts by Chlamydia pneumoniae and anti-cholesterol antibodies (ACHA) in the serum of patients with severe coronary artery disease, Virchows Archiv 439/3:251 [XXIII.] István Horváth, Anna Horváth, Gábor Vallus, Zoltán Prohászka, B. Dlustus, Á. Baranyai, László Romics, George Füst and István Karádi: From the lues screening to the anti-cholesterol antibody studies of healthy subjects and patients with peripheral atherosclerotic disease, Acta Microbiologica et Immonologica Hungarica, 47 (2000) 126. [XXIV.] G. Füst, Anna Horváth, D.Bánhegyi, E.Ujhelyi, A.Veres, L.Horváth, Z.Prohászka, V.Tarján, L.Romics, I.Horváth, I.Karádi. High level of anticholesterol antibodies (ACHA) in HIV patients. Normalization of ACHA upon highly active antiretroviral therapy (HAART), Magyar Venerológiai Archivum 2000, 4/37. [XXV.] Bíró Adrienn, Horváth Anna, Füst György, Cervenak László, Veres Amarilla, Bereczki Biborka, Kovács Margit és Karádi István: Anti-koleszterin antitest szint változása hypertrigliceridaemiás betegek szérumában gemfibrozil kezelés hatására, MIT XXX. Kongresszusa, Budapest Absztrakt füzet P/58. [XXVI.] E. Ujhelyi, D. Bánhegyi, Anna Horvath, G. Füst, Z. Prohaszka, I. Horvath: Anti-cholesterol antibody level changes in HIV positive patients after therapy, 2000, XIII. International AIDS Conference CD: TuPeA3057 Absztrakt füzet 3057 [XXVII.] Bíró A, Horváth A, Varga L, Nemesánszky E, Csepregi A, Dávid K, Tolvaj Gy, Ibrányi E, Telegdy L, Pár A, Romics L, Karádi I, Köles A, Bálint T, Horányi M: A koleszterin-ellenes autoantitestek (ACHA) szérumszintjének sorozatos mérése IFN-a-2b-vel kezelt krónikus hepatitis C-s (CHC) betegekben, MIT Kongresszus, Eger Absztrakt füzet P/11 9

10 [XXVIII.] Horváth Laura, Fekete Béla, Czirják László, Kalabay László, Veres Amarilla, Fónyad Gábor, Horváth Anna, Viczián Ágnes, Hoffer Izabella, Füst György, Romics László, Prohászka Zoltán: Helicobacter pylori fertőzöttség és 60kD-os hősokkfehérjék elleni antitestek kapcsolata különböző autoimmun betegségekben MIT Kongresszus, Eger Absztrakt füzet P26 10