- Email: [email protected]
6 mice
54
Poster Abstracts / Journal of Neuroimmunology 90 (1998) 13-105
293 Binding of Complement Component Clq to Mydin Oligodcndrocyte Glyeoprotein: A Novel M e c h a n i s m f o r R e g u l a t i n g C N S Inflammation T.G. Johns, C.C.A. Bernard, La Trobe Unwersity, Victoria, Australia
Myelin oligodendrocyte glyeopmtein (MOt3) is a myelin specific protein restricted to the cenwal nervous system (CNS). While M e G is considered a putative autoantigea in MS, its function(s) in myelin is unknown. As CNS myelin is able to activate the classical complemmt pathway, it must contain a Clq binding/activating prototn but the identity of this protein has not been reported. We now demon,strata that the extmc~tlular innonooglobulin-like domain of MOG specifically binds Clq in a dose dependent and ~turating manner. Using a series of overlapping pcptides representing the extracellalar don~in of M e G , we were able to identify a paptide that specifically itthibit~ the interaction between Clq and M e G . This peptide, however, did not bind Clq but to the extraccllular portion of MeG, suggesting that it constitutes of the dimer forming domain. Purified ?dOG -also inhibited the antibodydependent lysis of RBC by complement and is a target for C3 dispositino. Taken tognther these results demonstrate that MOQ may be the protein responsible for complement activation in myelin. This direct interaction between a myelin specific protein and CIq has significant implications for CNS inllanunotion and could be particularly importatu in dcmyetinatmg diseases such as multiple sclerosis.
294 IL-10 is Critical in the Regulation of Automimmune Encephalomyelitis as Demonstrated by Studies of IL-10 and IL-4 Deficient and T r a n s g e n i c Mice E. BettellL HarvardMedical School, USA, M.E Das, Millenium Pharmaceuticals, USA, E. Howard, H. Weiner, HarvardMedical School, USA, R- Sob el, VAMedical Center, USA, V. Kuohroo, Harvard Medical School, USA Experimental autoimmune encephalomyelitis (EAE) and a number of other experimental autoimmune diseases are induced by auto-antigen specific Thl cells. In contrast, transl~r of auto-antigen reactive Th2 cells that produce IL-4 and IL-10 can prevent and/or reverse EAE. The relative roles of these two Th2 cytokines in regulation of EAE has not been evaluated. Utilizing IL-4 and ILl0 deficient and transgenic mice, we demontrate that IL-10 deficient mice (1L10¢) are more suceptible and develop a more severe EAE when compared to IL4 deficient mice (IL-4 ~) or the wild type mice fWT). T cells from IL-10 ~ mice exhibit a stronger antigen specific proliferation, produce more Thl cytnkine (IFN-7 and TNF-cQ and induce very severe EAE upon transfered into wild type mice. In contrast, while IL-4 transgenic mice develop similar disease compared to their non-transgenic littermate, mice transgenic for IL-10 are completely resistant to the developmenl of EAE. Taken together our data suggest that LL10 plays a more critical role in the regulation of EAE by keeping autopathogenie T cell responses under check.
295 Inhibition of CNS Inflammation and Demyelination T. Bre~mer, Hadassah Medical Center, Israel, E. Poradosu, Hebrew University, Israel, D. Softer, C. Sicsic, HadassahMedical Center, Israel, G. Aviv, L. Alex, Hebrew University, Israel
Objectwe: The present study was designed to determine the effect of spocific Tyro~,ine Kinase(TK) inhibitors on CNS inflammation and demyelination. Methodology: TK blockers from the AG126/AG 556 tyrpho~tin family were tested for their ability to inhibit LPS-induced production of inflammatory nmdiators or to block the activation of stress kinases in primary rat glial ceil cultures In addition, the effect of these inhibitors on active EAE in SJL mice was evaluated. Results: The tyrphostins inhibited LPS-induced production of TNF~z. nitric oxide (NO) and Prostaglandin F,-z by gltal cells, but did not block the activation of JNK/SAPK or of p3g/HOG, However. AG 556 was efficient in inhibiting EAE: reduction of disease severity (g4%), incidence (60*/0). duration (55%), and inflanunatory lesions (76-97%). Conclusions: Our findings further suppo~ the important role of TNFct, NO and PGEz in CNS pathology and ,:onfinm the beneficial effects of AG 556 in th~ treatment of inflammatory conditions in general and EAE in particular. The biochemical target of AG 556 seems In be downstream to/NK or p38/HOG or in a parallel pathway, and is the subject of ongoing experiments.
296 C y t o k i n e a n d C h e m o k i n e Profiles in E A E Mice Depleted of G a m m a Ddta T Cdls A.J. Rajan~Albert Einstein College ofMedicine, USA,V.C. Asensio, I.L. Campbell, The Scripps Research Institute, USA, C.F. Brosnalt, Alhert E inste in College ofMedicine, USA
We showed previously that depletion of gd T ceils using the mAb GL3 protected animals against MBP-induced EAE, We now report on the effect of gd T cell depletion on expression of the cytokines IL-1, IL-6, TNF, LT and IFNg and the chemokines MIP-la, MIP-lb, IP-10, MCP-1 and RANTES in spinal cord tissues. In control animals mRNA for II.-1 and IL-6 rose dramatically at disease onset and peaked prior to disease height whereas mRNA for TNF, LT and IFNg rose more slowly and peaked with peak of disease. In GL3-treated animals, a dramatic reduction in all five cytokines was noted at disease onset, but only IFNg remained significantly reduced at peak of disease. ELISA confirmed the reduced levels of IL-I and 11.-6. All of the ehemokJnes were markedly upregulated at disease Onset, peaked with height of disease and fell during the recovery phase which was delayed in animals treated with GL3. MRNA and protein for type II NOS was also reduced in GL3-treated animals. These results suggest that gd T ceils contribute to the pathogenesis of EAE by regulating the influx of inflammatory cells into the spinal cord and by augmenting the pro- inflammatory eytokine and chemokine profile of the inflammatory infiltrates.
297 Treatment with IGF-1 Fails to Enhance Remydination B. Cmmella, Albert Einstein College ofMedicine, USA, E. Capello,Laboratorio di Neuropatologia Genova-Italia~ C.S. Raine, Albert E i~'tein College oJMedicine, USA
Previous studies testing IGF-1 oo the clinical and histopathologic outcome of EAE have suggested beneficial effects (Liu et al 1995; Yao et al 1995; 1996; Li et al, 1996). In the present study, 7 experiments, each comprising two groups of SJL mice adoptively-sensitized for EAE with MBP-responsive lymphocytes were conducted. Each group was given a 14 day course of placebo or IGF-I (100 p.g s.c., 2 x daily) either during the acute or the chronic phase (l, 2 or 3 me pt). Acute treatment led to marked reduction in disease severity. CNS lesions were less inflammatory and demyelinative and in some cases, precocious remyelination was seen. However, with time. both treated and control groups developed comparable clinical and histopathologic scores. Groups treated with IGF-1 during the chronic phase showed slight clinical improvement over controls but no major pathologic differences. Detailed analysis of CNS remyelination revealed no significant differences related to 1GF-1 administration. We conclude that IGF-1 has a modest short-term beneficial effect upon EAE when administered prior to onset but little or no beneficial effect when given during the chronic phase. (We thank Genentech for providing IGF-I. Supported in part by NMSS RG 1001-I-9; NS 08952 and NS 11920.)
298 E x p e r i m e n t a l Allergic Encephalitis in B7-defieient C57BL/6 Mice T.T, Chang, A.H. Sharpe, V.K. Kuchroo, HmvardMedicalSchool, USA
In order to study the role of co-stimulation in the development of autoimmunity, we have induced experimental allergic encephalitis (EAE) in C57BL/6 mice deficient in B7.1, B7.2, and both B7.1 and B7.2 molecules. EAE was induced by active immunization with the peptide 35-55 of myelin oligodendrocyte glycoprotein (MEG) in adjuvant. In addition to disease course and pathology, we compared proliferation, cytokine secretion, and anti-MeG antibody production ill the three knock-out mice. We found a dramatic difference in EAE susceptibility between wild-type and B7.1/B7.2 double knock-out mice.
Poster Abstracts / Journal of Neuroimmunology 90 (1998) 13-105
293 Binding of Complement Component Clq to Mydin Oligodcndrocyte Glyeoprotein: A Novel M e c h a n i s m f o r R e g u l a t i n g C N S Inflammation T.G. Johns, C.C.A. Bernard, La Trobe Unwersity, Victoria, Australia
Myelin oligodendrocyte glyeopmtein (MOt3) is a myelin specific protein restricted to the cenwal nervous system (CNS). While M e G is considered a putative autoantigea in MS, its function(s) in myelin is unknown. As CNS myelin is able to activate the classical complemmt pathway, it must contain a Clq binding/activating prototn but the identity of this protein has not been reported. We now demon,strata that the extmc~tlular innonooglobulin-like domain of MOG specifically binds Clq in a dose dependent and ~turating manner. Using a series of overlapping pcptides representing the extracellalar don~in of M e G , we were able to identify a paptide that specifically itthibit~ the interaction between Clq and M e G . This peptide, however, did not bind Clq but to the extraccllular portion of MeG, suggesting that it constitutes of the dimer forming domain. Purified ?dOG -also inhibited the antibodydependent lysis of RBC by complement and is a target for C3 dispositino. Taken tognther these results demonstrate that MOQ may be the protein responsible for complement activation in myelin. This direct interaction between a myelin specific protein and CIq has significant implications for CNS inllanunotion and could be particularly importatu in dcmyetinatmg diseases such as multiple sclerosis.
294 IL-10 is Critical in the Regulation of Automimmune Encephalomyelitis as Demonstrated by Studies of IL-10 and IL-4 Deficient and T r a n s g e n i c Mice E. BettellL HarvardMedical School, USA, M.E Das, Millenium Pharmaceuticals, USA, E. Howard, H. Weiner, HarvardMedical School, USA, R- Sob el, VAMedical Center, USA, V. Kuohroo, Harvard Medical School, USA Experimental autoimmune encephalomyelitis (EAE) and a number of other experimental autoimmune diseases are induced by auto-antigen specific Thl cells. In contrast, transl~r of auto-antigen reactive Th2 cells that produce IL-4 and IL-10 can prevent and/or reverse EAE. The relative roles of these two Th2 cytokines in regulation of EAE has not been evaluated. Utilizing IL-4 and ILl0 deficient and transgenic mice, we demontrate that IL-10 deficient mice (1L10¢) are more suceptible and develop a more severe EAE when compared to IL4 deficient mice (IL-4 ~) or the wild type mice fWT). T cells from IL-10 ~ mice exhibit a stronger antigen specific proliferation, produce more Thl cytnkine (IFN-7 and TNF-cQ and induce very severe EAE upon transfered into wild type mice. In contrast, while IL-4 transgenic mice develop similar disease compared to their non-transgenic littermate, mice transgenic for IL-10 are completely resistant to the developmenl of EAE. Taken together our data suggest that LL10 plays a more critical role in the regulation of EAE by keeping autopathogenie T cell responses under check.
295 Inhibition of CNS Inflammation and Demyelination T. Bre~mer, Hadassah Medical Center, Israel, E. Poradosu, Hebrew University, Israel, D. Softer, C. Sicsic, HadassahMedical Center, Israel, G. Aviv, L. Alex, Hebrew University, Israel
Objectwe: The present study was designed to determine the effect of spocific Tyro~,ine Kinase(TK) inhibitors on CNS inflammation and demyelination. Methodology: TK blockers from the AG126/AG 556 tyrpho~tin family were tested for their ability to inhibit LPS-induced production of inflammatory nmdiators or to block the activation of stress kinases in primary rat glial ceil cultures In addition, the effect of these inhibitors on active EAE in SJL mice was evaluated. Results: The tyrphostins inhibited LPS-induced production of TNF~z. nitric oxide (NO) and Prostaglandin F,-z by gltal cells, but did not block the activation of JNK/SAPK or of p3g/HOG, However. AG 556 was efficient in inhibiting EAE: reduction of disease severity (g4%), incidence (60*/0). duration (55%), and inflanunatory lesions (76-97%). Conclusions: Our findings further suppo~ the important role of TNFct, NO and PGEz in CNS pathology and ,:onfinm the beneficial effects of AG 556 in th~ treatment of inflammatory conditions in general and EAE in particular. The biochemical target of AG 556 seems In be downstream to/NK or p38/HOG or in a parallel pathway, and is the subject of ongoing experiments.
296 C y t o k i n e a n d C h e m o k i n e Profiles in E A E Mice Depleted of G a m m a Ddta T Cdls A.J. Rajan~Albert Einstein College ofMedicine, USA,V.C. Asensio, I.L. Campbell, The Scripps Research Institute, USA, C.F. Brosnalt, Alhert E inste in College ofMedicine, USA
We showed previously that depletion of gd T ceils using the mAb GL3 protected animals against MBP-induced EAE, We now report on the effect of gd T cell depletion on expression of the cytokines IL-1, IL-6, TNF, LT and IFNg and the chemokines MIP-la, MIP-lb, IP-10, MCP-1 and RANTES in spinal cord tissues. In control animals mRNA for II.-1 and IL-6 rose dramatically at disease onset and peaked prior to disease height whereas mRNA for TNF, LT and IFNg rose more slowly and peaked with peak of disease. In GL3-treated animals, a dramatic reduction in all five cytokines was noted at disease onset, but only IFNg remained significantly reduced at peak of disease. ELISA confirmed the reduced levels of IL-I and 11.-6. All of the ehemokJnes were markedly upregulated at disease Onset, peaked with height of disease and fell during the recovery phase which was delayed in animals treated with GL3. MRNA and protein for type II NOS was also reduced in GL3-treated animals. These results suggest that gd T ceils contribute to the pathogenesis of EAE by regulating the influx of inflammatory cells into the spinal cord and by augmenting the pro- inflammatory eytokine and chemokine profile of the inflammatory infiltrates.
297 Treatment with IGF-1 Fails to Enhance Remydination B. Cmmella, Albert Einstein College ofMedicine, USA, E. Capello,Laboratorio di Neuropatologia Genova-Italia~ C.S. Raine, Albert E i~'tein College oJMedicine, USA
Previous studies testing IGF-1 oo the clinical and histopathologic outcome of EAE have suggested beneficial effects (Liu et al 1995; Yao et al 1995; 1996; Li et al, 1996). In the present study, 7 experiments, each comprising two groups of SJL mice adoptively-sensitized for EAE with MBP-responsive lymphocytes were conducted. Each group was given a 14 day course of placebo or IGF-I (100 p.g s.c., 2 x daily) either during the acute or the chronic phase (l, 2 or 3 me pt). Acute treatment led to marked reduction in disease severity. CNS lesions were less inflammatory and demyelinative and in some cases, precocious remyelination was seen. However, with time. both treated and control groups developed comparable clinical and histopathologic scores. Groups treated with IGF-1 during the chronic phase showed slight clinical improvement over controls but no major pathologic differences. Detailed analysis of CNS remyelination revealed no significant differences related to 1GF-1 administration. We conclude that IGF-1 has a modest short-term beneficial effect upon EAE when administered prior to onset but little or no beneficial effect when given during the chronic phase. (We thank Genentech for providing IGF-I. Supported in part by NMSS RG 1001-I-9; NS 08952 and NS 11920.)
298 E x p e r i m e n t a l Allergic Encephalitis in B7-defieient C57BL/6 Mice T.T, Chang, A.H. Sharpe, V.K. Kuchroo, HmvardMedicalSchool, USA
In order to study the role of co-stimulation in the development of autoimmunity, we have induced experimental allergic encephalitis (EAE) in C57BL/6 mice deficient in B7.1, B7.2, and both B7.1 and B7.2 molecules. EAE was induced by active immunization with the peptide 35-55 of myelin oligodendrocyte glycoprotein (MEG) in adjuvant. In addition to disease course and pathology, we compared proliferation, cytokine secretion, and anti-MeG antibody production ill the three knock-out mice. We found a dramatic difference in EAE susceptibility between wild-type and B7.1/B7.2 double knock-out mice.