Macrophage depletion prevents leukocyte adhesion and disease induction in experimental melanin-protein induced uveitis.
The purpose was to study the effects of macrophage depletion with liposomal dichloromethylene-diphosphonate (Cl(2)MDP-lip) on inflammation and leukocyte-endothelium interaction in experimental melanin-protein induced uveitis (EMIU). Lewis rats (n = 48) were immunized with melanin-associated protein in complete Freund's adjuvant and pertussis toxin. Control groups received adjuvants without the antigen (n = 12) or no injection (n = 6). Animals received treatment with either CL(2)MDP-lip or empty liposomes (empty-lip) on day -2, 1, 4, 6 and 8. Leukocytes were stained with rhodamine 6G i.v. and intravital fluorescence microscopy (IVM) was performed on day 4, 6, 8 and 10 to quantify leukocyte rolling and arrest. After IVM, the cell count and protein concentration were determined in aqueous humor and plasma levels of TNF-alpha and IFN-gamma were measured by ELISA. In EMIU, leukocyte rolling increased on day 4 (10.0 +/- 1.2 cells min(-1)vs baseline of 5.7 +/- 0.7 cells min(-1), mean +/- S.E.(M.)) and peaked on day 8 (40.8 +/- 4.2 cells min(-1);P < or = 0.05). Leukocyte arrest was increased on day 8 (175.4 +/- 18.2 cells mm(-2)vs baseline of 59.7 +/- 7.1 cells mm(-2);P < or = 0.05) and day 10 (371.7 +/- 30.7 cells mm(-2)). CL(2)MDP-lip prevented leukocyte rolling (day 10: 16.6 +/- 1.8 cells min(-1)vs 30.7 +/- 2.9 cells min(-1); CL(2)MDP-lip vs untreated EMIU;P < or = 0.05) and arrest (day 8: 88.3 +/- 13 cells mm(-2); day 10: 128.5 +/- 12.9 cells mm(-2);P < or = 0.05). Empty-lip had no effect on leukocyte rolling (day 10: 34.8 +/- 4.2 cells min(-1)) or arrest (day 8: 159.3 +/- 12.9 cells mm(-2), day 10: 421.2 +/- 41.6 cells mm(-2)). CL(2)MDP-lip completely suppressed leukocyte emigration (11 +/- 2 cells microl(-1)vs 100 +/- 29 cells microl(-1); CL(2)MDP-lip vs empty-lip;P < or = 0.05) and protein extravasation into aqueous humor (2.7 +/- 0.3 mg ml(-1)vs 14.2 +/- 2.1 mg ml(-1); CL(2)MDP-lip vs empty-lip;P < or = 0.05), abrogated the TNF-alpha response (32.5 +/- 2.7 pg ml(-1)vs 954.9 +/- 216.3 pg ml(-1); CL(2)MDP-lip vs untreated EMIU;P < or = 0.05) and caused an attenuated and delayed elevation of IFN-gamma. CL(2)MDP-lip prevented the inflammatory reaction of EMIU and inhibited the increase of leukocyte-endothelium interaction in iris vessels. Our findings emphasize the pivotal role macrophages play in the initiation of autoimmune disease.
Baatz
H
H
Department of Ophthalmology, Johann Wolfgang Goethe-Universität, Theodor-Stern-Kai 7, D-60590 Frankfurt/Main, Germany. holger_baatz@hotmail.com
Puchta
J
J
Reszka
R
R
Pleyer
U
U
D000704:Analysis of Variance; D000818:Animals; D001082:Aqueous Humor / Q000737:chemistry; D001327:Autoimmune Diseases / Q000276:immunology*; D002448:Cell Adhesion / Q000502:physiology; D002452:Cell Count; D002464:Cell Migration Inhibition; D004002:Clodronic Acid / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D007091:Image Processing, Computer-Assisted; D007371:Interferon-gamma / Q000097:blood; D007962:Leukocytes / Q000502:physiology*; D008264:Macrophages / Q000502:physiology*; D008856:Microscopy, Fluorescence; D051381:Rats; D011917:Rats, Inbred Lew; D018709:Statistics, Nonparametric; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis; D014605:Uveitis / Q000276:immunology*
D014409:Tumor Necrosis Factor-alpha; D004002:Clodronic Acid; D007371:Interferon-gamma