Reijo Salonen

HLA class II associated risk and protection against multiple sclerosis—a Finnish family study

We analyzed the HLA class II haplotypes in 249 Finnish nuclear families and compared the frequencies of parental haplotypes transmitted or non-transmitted to multiple sclerosis (MS) patients. The most important predisposing haplotype was DRB1*15-DQB1*0602 (P<10(-6)) as expected and a weak predisposing effect of DRB1*04-DQB1*0302 was revealed after the elimination of DRB1*15-DQB1*0602. HLA-DRB1*01-DQB1*0501 and DRB1*13-DQB1*0603 were negatively associated with MS in transmission disequilibrium test, but only the DRB1*13-DQB1*0603 association remained significant (P=0.008) after the elimination of DRB1*15-DQB1*0602 haplotypes. Based on this study HLA class II haplotypes exhibit both predisposing and protective effects in MS.

Therapy with antibody against leukocyte integrin VLA-4 (CD49d) is effective and safe in virus-facilitated experimental allergic encephalomyelitis

Experimental allergic encephalomyelitis (EAE) is facilitated in resistant BALB/c mice by intraperitoneal infection with an avirulent Semliki Forest virus (SFV-A7). Viral infection increases the incidence of EAE from 15-30% to 60-90% and speeds up appearance of paralysis from 24 to 14 days. In this paper, we describe treatment of virus-facilitated EAE with monoclonal antibodies (mAbs) against leukocyte and/or endothelial cell adhesion molecules. Therapy with mAb against ICAM-1 (intercellular adhesion molecule-1) had a modest effect, but caused hemorrhagic brain and spinal cord lesions. Therapy with mAb against Mac-1 (alpha M beta 2-integrin) was well tolerated but had no effect. Therapy with mAb against VLA-4 (alpha 4 beta 1-integrin) was safe, diminished both clinical and histopathological signs of EAE, decreased induction of VCAM-1 (vascular cell adhesion molecule-1) on brain vessels and diminished infiltration of VLA-4+ cells into the brain. The amount of viral antigen in the brain was not altered. We conclude that facilitation of leukocyte entry into the brain is a major mechanism for viral facilitation of EAE in the BALB/c mouse, and that facilitation can be inhibited by anti-adhesion therapy. This may have implications for treatment of relapses triggered by viral infections in multiple sclerosis.

Interferon-β downregulates expression of VLA-4 antigen and antagonizes interferon-γ-induced expression of HLA-DQ on human peripheral blood monocytes

We have studied the effect of recombinant human IFN-beta on the basal and IFN-gamma-induced expression of adhesion molecules and class II MHC antigens on human peripheral blood monocytes and on ICAM-1 (intercellular adhesion molecule-1) expression of a human umbilical vein endothelial cell line (EAhy 926). We show that IFN-beta downregulates both basal and IFN-gamma-induced expression of VLA-4 (very late activation antigen-4) antigen on monocytes, but has no effect on the expression of CD11a, CD11b, CD11c, L-selectin, CD18, ICAM-1, beta 1-integrin or CD44 on monocytes or ICAM-1 on EAhy 926 cells. We also show that IFN-beta antagonizes the IFN-gamma-induced expression of HLA-DQ-antigen, but not HLA-DR or HLA-DP antigens on monocyte surface. These findings may partially explain the beneficial effect of IFN-beta in multiple sclerosis, since VLA-4-antigen is critical for leukocyte recruitment into inflamed brain and downregulation of HLA-class II expression diminishes antigen presenting capacity of monocytes.

Blood-brain barrier breakdown and increased intercellular adhesion molecule (ICAM-1/CD54) expression after Semliki Forest ( A7) virus infection facilitates the development of experimental allergic encephalomyelitis

This report describes two mechanisms by which virus infection can facilitate demyelinating autoimmune inflammation in the murine CNS. In the BALB/c mouse model of experimental allergic encephalomyelitis (EAE), peripheral infection with an avirulent strain (A7) of Semliki Forest virus (SFV) increased the morbidity to EAE by infecting endothelial cells and damaging the blood-brain barrier (BBB). An influx of hematogenous CD18+ (LFA-1+ and MAC-1+) cells into the CNS compartment was followed by a local increase in intercellular adhesion molecule 1 (ICAM-1) expression on the vascular endothelium. Although SFV A7 infection without EAE induction caused multifocal cerebral vascular endothelial cell infection and BBB damage followed by cellular infiltration and transient increase of ICAM-1, inflammation and demyelination of CNS white matter with classical clinical signs of EAE was observed only in EAE-induced BALB/c mice, whereas the control mice remained neurologically healthy. The upregulation of ICAM-1 after virus infection was detected after the CD18+ (LFA-1+ and MAC-1+) cells had infiltrated the CNS both after EAE induction and also in nonsensitized control mice. The observed increase in ICAM-1 expression was transient in nonsensitized SFV A7 infected mice just as in the cellular infiltrates in the CNS, but EAE induction resulted in prolongation in both the cellular infiltrates and upregulation of ICAM-1. Thus, SFV A7 infection causes BBB damage and prolongs increased ICAM-1 expression on brain endothelium. This results in increased and more rapid morbidity to EAE in mice which have been sensitized with neuroantigen. However, SFV A7-infected mice without neuroantigen sensitization remain neurologically healthy.

Semliki Forest virus infection leads to increased expression of adhesion molecules on splenic T-cells and on brain vascular endothelium

Semliki Forest virus A7 (SFV-A7) is a neurotropic alphavirus that leads to an asymptomatic encephalitis in adult immunocompetent mice. We studied the expression of leukocyte and endothelial cell adhesion molecules in the spleen and in the central nervous system (CNS) during SFV-A7 infection. Kinetics of the expression of LFA-1 alpha/CD11a, LFA-1 beta/CD18, Mac-1/CD11b, VLA-4/CD49d, ICAM-1/CD54 and L-selectin/CD62L was determined on splenic CD4+ and CD8+ T-cells and macrophages by flow cytometry. Time course of the expression of these antigens and VCAM-1/CD106 as well as viral antigens in the CNS was studied by immunoperoxidase staining. In the spleen, a sustained increase in LFA-1-expression and a temporary increase at day 7 in the expression of VLA-4, Mac-1 and ICAM-1 were detected on CD8+ T-cells. L-selection was down-regulated on CD4+ cells. Adhesion molecules on macrophages remained unchanged. In the CNS, expression of Mac-1+, VLA-4+ and LFA-1+ cells increased in parallel with the kinetics of the expression of their ligands ICAM-1 and VCAM-1 on brain vessels. Upregulation of adhesion of molecules peaked between days 5-8 and was most prominent in the cerebellar and brain stem white matter where viral antigens were most abundant. We conclude that the adhesion molecules profile of splenic T cells is altered during SFV-A7 infection which may influence their homing into the CNS. Macrophages are probably recruited non-specifically as a consequence of activation of the brain vascular endothelium in the inflamed areas of the brain.

Defective production of interferon-α associated with HLA-Dw2 antigen in stable multiple sclerosis☆

Interferon (IFN) production by peripheral blood mononuclear cells after induction with one purified and three crude viral antigens was studied in 29 patients with stable multiple sclerosis (MS) and 29 healthy controls. Antiviral substance produced was characterized as interferon-alpha. MS patients produced significantly less IFN-alpha after induction with mumps and purified measles virus antigens and the same tendency was seen after induction with rubella virus antigen. However, when herpes simplex virus antigen was used as the stimulating agent, no difference was seen between MS patients and controls. The decreased ability to produce IFN-alpha was associated with the histocompatibility antigen Dw2. Control subjects positive for Dw2 also produced less IFN-alpha than Dw2-negative controls. In conclusion, we suggest that the observed impaired interferon-alpha production in MS is at least partially due to a high prevalence of Dw2 antigen in this disease.

Selective downregulation of Th1 response by Linomide reduces autoimmunity but increases susceptibility to viral infection in BALB/c and SJL mice

Susceptibility to autoimmunity has been associated with polarization of Th1/Th2 balance in immune system towards the Th1-type of reactivity. We report here that orally administered quinoline-3-carboxamide (Linomide) selectively downregulates Th1 response in BALB/c and SJL mice, leading to reduction of autoimmunity in the BALB/c and SJL models of experimental allergic encephalomyelitis (EAE). This was shown by prevention of EAE in Th1 responding SJL mice and partial downregulation of EAE in Th2-prone BALB/c mice. In a BALB/c model of EAE, in which infection with Semliki Forest A7 virus (SFV-A7) is used for enhancement of autoimmunity, clinical signs of EAE were reduced while mortality due to viral infection in the CNS was enhanced. Selective downregulation of the Th1 response by Linomide also rendered initially resistant SJL mice susceptible to SFV-A7 CNS infection. This was shown by immunohistochemical detection of extensive deposits of viral antigen in numerous perivascular foci within the CNS and abolished virus antigen-specific lymphocyte reactivity in Linomide-treated SJL mice. In addition, analysis of spleen cell cytokine mRNA production profile revealed decreased number of IFN-gamma producing cells in both SJL and BALB/c mice, reduced number of IL-12p40 producing cells in SJL and increased number of 12p40 producing cells in BALB/c mice along with slightly increased IL-4 production in both strains of mice. These results indicate that oral treatment with Linomide induces selective downregulation of Th1 reactivity causing reduction of autoimmunity and increased susceptibility to SFV-A7 CNS infection. Selective downregulation of Th1 response is a desired effect in the treatment of autoimmune diseases but our results suggest that the benefits have to be balanced against the possible loss in immunoprotection against pathogens.

Facilitation of experimental allergic encephalomyelitis by irradiation and virus infection: role of inflammatory cells

Abstract Infection with an avirulent strain of Semliki Forest virus (SFV-A7) facilitates the development of experimental allergic encephalomyelitis (EAE) in a genetically resistant BALB/c mouse strain. Irradiation which is necessary for EAE induction caused a decrease in the total number of lymphocytes and an increase in CD4+/CD8+ T cell ratio in the spleen of BALB/c mice. EAE induction increased the ratio further until clinical and histological signs of EAE appeared. Entry of perivascular CD4+ and CD8+ cells preceded the onset of clinical signs and the appearance of MAC-1+ cells in the central nervous system (CNS). In the acute phase of EAE, cellular infiltrates, which were sparse, consisted mainly of MAC-1+ cells and a few CD4+ and CD8+ cells. Inflammatory cells gradually disappeared during the recovery phase. SFV-A7 infection after irradiation and EAE induction did not significantly change the CD4+/CD8+ ratio in the spleen or in the CNS infiltrates but enhanced the entry of inflammatory cells into the CNS. Similar perivascular cell influx was also seen in untreated mice infected with SFV-A7. We conclude that observed rapid reduction of splenic mononuclear cells and increase of the CD4+/CD8+ T cell ratio caused by irradiation prior EAE induction are early crucial events in disease induction in this resistant strain of mice. SFV-A7 infection, which further facilitates the development of EAE, does not induce immunoregulatory changes but provides its effect by enhancing the entry of inflammatory cells into the CNS. The combination of these two mechanisms thus effectively breaks the natural resistance against EAE in this genetically resistant mouse strain.

Intrathecal Immune Responses in Mumps Meningitis Patients

The in vitro 3H‐thymidine incorporation of peripheral blood (PB) and cerebrospinal fluid (CSF) lymphocytes from 11 mumps meningitis patients was studied after stimulation with non‐specific mitogens and microbial antigens. Although corresponding viral antibodies were always found in the CSF of seropositive patients by a sensitive radioimmunoassay, their intrathecal synthesis was directed mainly against mumps virus. Most of the patients had PB lymphocytes that reacted on stimulation with phytohaemagglutinin (PHA) or pokeweed mitogen (PWM), but a smaller number of them had reactive CSF cells. Only four patients showed stronger responses to PHA in the CSF than in PB. Most patients had mumps‐reactive lymphocytes in PB but only two of them in the CSF. In contrast, these patients more often had increased CSF cell reactivity when tested with purified protein derivative, measles, and herpes simplex virus antigens. The results could not be explained by a damaged blood—brain barrier alone but may reflect the immunological status of the brain compartment in these patients.

Measles virus hemagglutinin mediates monocyte aggregation and increased adherence to measles-infected endothelial cells

Abstract The effect of measles virus (MV) infection on monocyte adhesion was studied using human peripheral blood monocytes and monocytic and endothelial cell lines. The infection of monocytic U-937 cells led to the formation of large cellular aggregates. Aggregation was independent of intercellular adhesion molecule-1 (ICAM-1)/ lymphocyte function-associated antigen-1 (LFA-1), but could be inhibited by monoclonal antibodies (mAb) against the MV hemagglutinin glycoprotein (MV-H). mAb against the MV receptor, CD46, also blocked aggregation. No significant changes in the cell surface expression of adhesion molecules CD11a, CD11b, CD11c, CD18, CD54, CD44, CD49d (α4-integrin) and CD62L (L-selectin) were observed on MV-infected monocytes. Infection of a human endothelial cell line, EAhy 926 (HEC), with MV led to a two-fold increase in ICAM-1 expression and a two-fold increase in monocyte adherence to the HEC (from 22±1.6% to 42±4.8%). However, ICAM-1 mAb reduced monocyte adhesion to the control and MV-infected HEC to a similar degree, whereas anti-MV-H antibodies abolished the difference between binding to infected and control HEC. We conclude that MV hemagglutinin mediated both the homotypic aggregation in infected monocyte cultures and increased monocyte adherence to the infected endothelial cells.