Basic Encephalitogen Research Articles

Lack of H-2 restriction of suppressor factor specific to myelin basic encephalitogen

A cell-free extract has been prepared from spleen cells of (SJL/J × BALB/c)F 1 mice which were rendered non-susceptible to EAE by treatment with mouse spinal cord homogenate in incomplete Freund's adjuvant. Such extract has been previously shown to have suppressive activity on the induction of EAE, as well as on the immune response in syngeneic mice towards the mouse basic protein. We have now demonstrated that this factor is as effective in several other strains of mice, of different H-2 haplotype. The activity of this factor was manifested both in vivo by inhibiting the DTH response to MBE in the various mouse strains, and in vitro, by blocking the MIF activity of the allogeneic lymphocytes. A suppressor factor was prepared by a similar procedure from the EAE-resistant BALB/c mice. This factor was as active as the factor from the (SJL/J × BALB/c)F 1 hybrid in blocking the anti-MBE antibody binding to MBE. It is also suppressive, and it inhibited the DTH response to MBE in both syngeneic and semi-allogeneic (SJL/J × BALB/c)F 1 mice. It is thus demonstrated that the MBE-specific soluble suppressor factor involved in the EAE system in mice shows no indication of H-2 restriction.

Unresponsiveness to experimental allergic encephalomyelitis in mice: replacement of suppressor cells by a soluble factor.

A soluble suppressor factor has been prepared from cells of mice rendered nonsusceptible to experimental allergic encephalomyelitis (EAE) by treatment with mouse spinal cord homogenate in incomplete Freund's adjuvant. The specific activity of this factor can be augmented by using a cell population enriched on plates coated with anti-mouse Fab and the specific antigen, mouse basic encephalitogen (MBE). The resultant suppressor factor had the same biologic activities as the cells from which it originated. Thus, it suppressed specifically the delayed-type hypersensitivity (DTH) response to MBE in vivo, and blocked in vitro the effector lymphocytes that adoptively transfer the DTH response. The suppressor factor reactivity was manifested also by the capacity to suppress the activity of macrophage migration inhibition factor produced by sensitized lymphocytes in the presence of the specific antigen MBE. The suppressor factor is antigen-specific and can bind the MBE in vitro and thus compete with its antibody binding. The most significant activity of the soluble suppressor factor is its ability to interfere with the induction of clinical EAE.

Cell‐mediated immunity to neural antigens in idiopathic polyneuritis and myeloradiculitis

Patients with peripheral nervous system disorders were tested for the presence of cellular hypersensitivity to peripheral and central nervous system antigens by means of the in vitro lymphocyte transformation technique. Lymphocytes sensitized to the neuritogenic peripheral nervous system P1L basic protein were found in pure polyradiculitis of the Guillain-Barré syndrome type. Lymphocytes from patients with myeloradiculitis underwent transformation by peripheral P2 basic protein and by central nervous system basic encephalitogenic protein. In cases of chronic relapsing polyneuropathy response was shown to the central nervous system basic encephalitogen and to both of the peripheral nerve basic proteins. Lymphocytes from patients with other neurologic conditions showed no response to any oth these antigens. These findings suggest that cell mediated immunity to specific basic proteins of the myelin plays a rolw in the pathogenesis of the above-mentioned demyelinating disorders and may lead to a new approach in their classification and diagnosis.

Inverse relationship between net electric charge on the antigen and that on the sensitized cell in cellular immune response: demonstration with basic encephalitogen of the brain.

An inverse relationship exists between the net-electrical charge of immunogens and the antibodies elicited (1). The cellular basis of the net charge phenomenon has been established for both positively and negatively charged immunogens, by cell separation techniques over columns of opposite charge (7, 8). To establish whether this phenomenon can be extended to include cell-mediated immunity, the response to basic encephalitogenic protein (BE) which induces experimental allergic encephalomyelitis (EAE) was now investigated. Lymph node cells from sensitized strain 13 guinea pigs were fractionated over positively and negatively charged columns and compared to unfractionated cell populations in two assay systems: (a) in vitro response to BE in terms of lymphocyte transformation and (b) the passive transfer of EAE to unsensitized syngeneic recipients. The response was found to be confined to the fraction of cells eluted from glass bead columns, namely, the more negative cells. Cells eluted from poly-L-lysine-coated glass bead columns (i.e., positive cells) were devoid of the capacity to respond to this antigen either in vivo or in vitro. It was previously established that thymocytes rather than bone marrow cells account for the inverse charge phenomenon as assayed by T-helper-cell function in in vivo antibody production (8). We have now extended the inverse charge effect to include cell-mediated immune response of the delayed hypersensitivity type.

Immunological comparison of basic encephalitogen and histone F2A1.

The extent of immunological cross-reaction between basic encephalitogen and histone F2A1 on both the humoral antibody level and on the cellular level has been established. The extent of humoral cross-reaction was tested by direct complement fixation employing both anti-histone F2A1 and antisera to basic encephalitogen, by inhibition of complement fixation, by radioimmunoassay and by passive cutaneous anaphylaxis. The data obtained failed to reveal immunological cross-reaction at the cellular level was tested by the lymphocyte stimulation technique in rabbits and guinea pigs, by inhibition of lymphocyte stimulation and by delayed hypersensitivity skin reactions. A slight but significant cross-reaction between the two proteins on the cellular level was detected by inhibition of lymphocyte stimulation and by the delayed hypersensitivity test. It is concluded that the immunological studies provide limited evidence that the two proteins share antigenic determinants.

Fractionation of Functional Lymphocytes Sensitized to Basic Encephalitogen on Derivatized Collagen and Gelatin Gels

The lymph node cells of basic encephalitogen (BE)-sensitized guinea pigs were fractionated on derivatized collagen and gelatin gels. The population of cells specifically reactive to this antigen can be isolated from derivatized gelatin gels and retain their viability and functionality as assayed in vitro. The specific binding of BE-sensitized cells to BE-derivatized gels comprised between 1 and 2% of the cells applied per plate. The ratio of sensitized cells bound to non-sensitized cells bound ranged between 4 and 6. The viability and functionality of adherent cells detached from collagen gels after enzymatic degradation were impaired. In contrast, the responses obtained with the adherent cell population released from the gelatin gels, by melting at 37 degrees C, were equal or greater than those of the original unfractionated lymph node cell cultures. Furthermore, it was possible to obtain a nonadherent cell population which was virtually completely depleted of the capacity to respond to the sensitizing antigen.

In vivo and in vitro immunological cross‐reactions between basic encephalitogen and synthetic basic polypeptides capable of suppressing experimental allergic encephalomyelitis

AbstractA significant extent of immunological cross‐reactivity has been demonstrated between the basic encephalitogenic protein of bovine origin and several synthetic amino acid copolymers which have suppressive effect on experimental allergic encephalomyelitis (EAE). This cross‐reactivity has been conclusively established on the cellular level, both in vivo by means of delayed hypersensitivity skin tests and in vitro using transformation of sensitized lymphocytes, as measured by incorporation of radioactive thymidine. The in vitro experiments have been conducted with lymph node cells from guinea pigs of both random bred and inbred strains, and on spleen and lymph node cells from rabbits. Definite cross‐reactivity was observed between the basic encephalitogen and all the synthetic copolymers which were previously shown effective in suppression of EAE, whereas ineffective copolymers or unrelated proteins did not show any cross‐reactivity. In the case of strain 2 guinea pigs and rabbit lymph node cells the cross‐reactivity in vitro was manifested by direct cross‐stimulation of the lymphocytes, whereas in random‐bred or strain 13 guinea pigs and rabbit spleen cells, the cross‐reaction was detected only by means of specific inhibition of the homologous stimulation by the heterologous antigen.A limited extent of cross‐reactivity was observed on the humoral level as well; antibodies provoked in guinea pigs against the synthetic copolymer Cop 1 cross‐reacted in the passive cutaneous anaphylaxis assay with the bovine basic encephalitogen.

Suppression by several synthetic polypeptides of experimental allergic encephalomyelitis induced in guinea pigs and rabbits with bovine and human basic encephalitogen.

AbstractThe basic copolymer of alanine, glutamic acid, lysine and tyrosine, denoted Cop 1, is an efficient suppressor of experimental allergic encephalomyelitis (EAE) induced with the bovine basic encephalitogen in guinea pigs and rabbits. The suppression was very effective also when the experimental disease was induced with the basic protein from human source. Two related polymers, one in which glutamic acid was replaced with aspartic acid, and one devoid of tyrosine, were less effective in the suppression of the experimentally induced disease.The route of administration of Cop 1 plays an important role in its suppressive effectiveness, with intravenous injections being the most efficient. The injection schedule and dose of injection are also of great importance; thus, repeated doses of Cop 1 starting five days after the administration of the basic protein are suppressive, whereas a single dose of 5 mg given 72 h after challenging with the basic protein does not suppress EAE. Cop 1 is not a nonspecific immunosuppressant, as it does not affect the immune response towards a variety of antigens, including the rejection of a skin graft.When EAE was induced with the human encephalitogen, the histological changes observed included demyelination and fibrosis in the guinea pig brain.

Correlation between strain differences in susceptibility to experimental allergic encephalomyelitis and the immune response to encephalitogenic protein in inbred guinea pigs.

Differences were observed between guinea pigs of inbred strains 2 and 13 in both the susceptibility to the induction of experimental allergic encephalo-myelitis (EAE), and the immune response to the basic encephalitogen. Strain 2 guinea pigs, which are resistant to the induction of the disease, produce humoral antibodies against the encephalitogen in a titer 40–50-fold higher than guinea pigs of strain 13, which are sensitive to EAE.

Protection against Experimental Allergic Encephalomyelitis

EXPERIMENTAL allergic encephalomyelitis (EAE) is an acute neurological autoimmune disease induced in laboratory animals such as guinea-pigs, rabbits, rats and monkeys by administration in complete Freund's adjuvant of a basic encephalitogenic protein (BE) isolated from the brain or spinal cord of several animal species1. We have recently shown2 that the disease may be suppressed by intravenous injection of a synthetic amino-acid copolymer (denoted Cop 1). This copolymer is composed of alanine, glutamic acid, lysine and tyrosine, in a residue molar ratio of 6.0 : 1.9 : 4.7 : 1.0, and has an average molecular weight of 23,000. This basic polymer is not encephalitogenic2 and is not a general non-specific immunosuppressive agent (our unpublished results). Whereas it efficiently suppresses the disease when administered after the basic encephalitogen, it cannot prevent the disease when injected intravenously in saline before the basic brain protein2.

A conformation change induced in the basic encephalitogen by lipids

Using ORD/CD we have studied the conformation of the basic encephalitogenic protein isolated from normal human myelin. Using the curve fitting method we have found this protein to be essentially random. When sodium dodecyl sulphate was added to a solution of the protein the random curve obtained by ORD was altered to an α-helical type. Addition of sodium dodecyl sulphate, sodium oleate or triphosphoinostide to the protein, all had the effect of changing the CD curve from random to α-helical type. The most effective agent was sodium dodecyl sulphate since it produced the largest rotation. Positively charged lipids, cetyltrimethylammonium bromide and lysolecithin did not affect the random CD curve at pH 2.0. However, at pH 5.0 where lysolecithin is a zwitterion, the CD curve appeared to be α-helical.