Annemarie Henriksson

Reference values for CSF-IgM, CSF-IgM/S-IgM ratio and IgM index, and its application to patients with multiple sclerosis and aseptic meningoencephalitis

Immunoglobulin M (IgM) concentrations were determined in cerebrospinal fluid (CSF) and serum (S) by enzyme-linked immunosorbent assay (ELISA). In 52 reference subjects, the upper reference limit was 0.46 mg/l for CSF-IgM, 0.32 X 10(-3) for CSF-IgM/S-IgM ratio, and 0.061 for IgM index equal to CSF-IgM X S-albumin/S-IgM X CSF-albumin ratio. No correlation to age over 15-85 years was found for any of these variables. Among 22 patients with aseptic meningoencephalitis (AM) elevated values of CSF-IgM were found in 68%, CSF-IgM/S-IgM ratio in 73%, and IgM index in 73%. The corresponding values among 35 patients with multiple sclerosis (MS) were 66%, 60% and 63%. The differences in diagnostic sensitivities for the three IgM variables were not significant. Eleven of 16 AM patients and two of 22 with MS had elevated IgM index in the presence of normal IgG and IgA indices. Determination of IgM index should therefore be performed in suspected inflammatory nervous system disorders.

Comparison of Seven Formulae and Isoelectrofocusing for Determination of Intrathecally Produced IgG in Neurological Diseases

Seven different formulae and agarose isoelectrofocusing (AIF) using immunolabelling for IgG were compared for their ability to discriminate between intrathecally produced IgG and transudated IgG in cerebrospinal fluid. All reference limits were set to a specificity of 97·5% (reference group, n = 211). The probability of a positive test (p +) was evaluated for 112 patients with multiple sclerosis (MS), 42 with meningitis, 114 with noninflammatory diseases affecting the central nervous system (CNS), 23 with Guillain-Barré syndrome, and 56 with various diseases not affecting the CNS. Agarose isoelectrofocusing had the best diagnostic sensitivity (93%) for MS, combined with a low p + (0–19%) for other diseases. Among the formulae, the IgG extended index and Reibers hyperbolic formula were equivalent, giving high (75–79%) diagnostic sensitivity for MS combined with low p + (4–22%) for other diseases. All other formulae, although sensitive for MS, had a higher rate of false positive results.

Immunoglobulin-producing cells in CSF and blood from patients with multiple sclerosis and other inflammatory neurological diseases enumerated by Protein-A plaque assay

Using the Protein-A plaque assay, numbers of IgG + IgA + IgM producing cells determined in patients with multiple sclerosis (MS) were 0.1-5% in CSF and 0.1-0.7% in peripheral blood; interestingly, 7 of 11 MS patients had IgM producing cells in CSF. In patients with aseptic meningitis (AM), the corresponding values were 0.04-7.5% in CSF and 0.4-2.4% in peripheral blood. There were more Ig producing cells in peripheral blood from patients with AM and MS than in healthy subjects. A correlation between numbers of IgG producing cells in CSF and the concentrations of intrathecally produced IgG (CSF IgG index) was registered in patients with AM; the same was the true for IgA. The Protein-A plaque method, adopted for 20 X 10(3) lymphocytes, makes possible enumeration of Ig-producing cells in CSF and discrimination among cells secreting different Ig classes, thereby being a powerful tool for studying immune reactions in the CNS-CSF compartment.

In vitro synthesis of immunoglobulins and autoantibodies by lymphocytes from various body compartments during chronic relapsing experimental allergic encephalomyelitis.

Mononuclear cells extracted from regional lymph nodes, blood, spleen and central nervous system of guinea pigs with chronic relapsing experimental allergic encephalomyelitis (r-EAE), adjuvant immunized and untreated controls were cultured for 16 h in microtitre plates, and culture supernatants were then used to measure IgG and IgM, as well as IgG class anti-myelin antibody production by enzyme-linked immunosorbent assays. Increased synthesis of these immunoglobulins and antibodies was found during the course of r-EAE both in intra- and extrathecal compartments. Long-term cultures carried out for 7 days gave similar results but anti-myelin, anti-myelin basic protein and IgG synthesis was most pronounced intrathecally. Agarose isoelectric focusing of supernatants from these cultures showed oligoclonal IgG. These findings indicate in vivo synthesis of autoantibodies within the target for immune attack and a partial sequestration of the immune response to this compartment.

IgM and IgG responses during chronic relapsing experimental allergic encephalomyelitis (r-EAE) ☆

During chronic relapsing experimental allergic encephalomyelitis (r-EAE) in guinea pigs, serum IgM and IgG concentrations increased markedly early in disease. Serum IgM and IgG increased similarly in control animals immunized with Freunds incomplete adjuvant (FIA) and Mycobacterium tuberculosis (MT). In the chronic phase of r-EAE but not in control animals, elevated IgM was also found in central nervous system (CNS) extracts, suggesting intrathecal IgM synthesis. IgG antibodies against myelin and myelin basic protein (MBP) were regularly detected in r-EAE sera from day 21 post inoculation (p.i.), reaching maximum levels in the early chronic phase. IgG antibodies against galactocerebroside (GC) and galactose appeared in some r-EAE sera. Oligoclonal IgG bands were demonstrated in all r-EAE guinea pig sera 21-26 days p.i. The bands in serum decreased in number and strength in the chronic phase. They could be traced to antibodies against MT in 4 of 10 animals, but not to antibodies against myelin, MBP, GC or galactose. Oligoclonal IgG bands were also regularly visualized in r-EAE CNS 124 days p.i., suggesting persistent intrathecal IgG synthesis. They varied in number and migration between different regions of individual CNS. Oligoclonal CNS IgG was related to antibodies against MT in only one of 7 animals, and in no case to antibodies against myelin.

Ig-secreting cells pass the blood-brain barrier: studies on κ and λ light chain secreting cells in plasma cell dyscrasia

To study if immunoglobulin (Ig)-secreting cells actively pass the blood-brain barrier (BBB), 15 patients with monoclonal gammopathy underwent bonemarrow (BM) iliac crest aspiration biopsy, peripheral blood (PB) sampling and cerebrospinal fluid (CSF) analysis. With an enzyme-linked immunospot assay we investigated the number and ratio of mononuclear cells secreting Ig with kappa and lambda light chains in the three different compartments. A statistically significant (P < 0.05) correlation between the ratio of Ig kappa/lambda-secreting cells in CSF, PB and BM was found. The frequency of kappa and lambda (i.e. Ig in total) secreting mononuclear cells, of the same Ig class as the paraprotein, per 10(4) mononuclear cells was higher in BM (median 2.16%, range 0.43-9.28%) compared to CSF (median 0.44%, range 0.05-9.25%) and in CSF compared to PB (median 0.12%, range 0.02-0.96%). The proportion of all mononuclear cells with Ig kappa and lambda light chain (i.e. Ig) secretion was on average 5-fold greater in CSF compared to PB and 11-fold greater in BM compared to PB. The present study indicates that paraprotein-secreting cells preferentially pass from PB to CSF.

Cerebrospinal fluid lymphocytes from patients with multiple sclerosis do not increase immunoglobulin or measles antibody production after stimulation with pokeweed mitogen

Cerebrospinal fluid lymphocytes (CSF-L) and peripheral blood lymphocytes (PBL) from patients with multiple sclerosis (MS) and acute aseptic meningoencephalitis (AM) were cultured without and in the presence of pokeweed mitogen (PWM), a polyclonal B cell activator. IgG, IgA and IgM as well as measles IgG antibody production was measured in 7-day-culture supernatants by enzyme-linked immunosorbent assay. MS CSF-L did not respond with increased Ig production after PWM stimulation, in contrast to AM CSF-L which responded to PWM with a modest increase of production of all 3 Ig classes, especially IgG. PBL responded to PWM with a pronounced production of IgG, IgA and especially IgM, showing no difference between MS, AM and healthy controls. CSF-L from only 1 of 7 patients with MS showed increased measles IgG antibody production after PWM stimulation. The poor response of MS CSF-L might be due to maximal activation of B lymphocytes in vivo, thereby limiting further Ig production after stimulation in vitro.

Humoral and cellular immunity in patients with acute aseptic meningitis.

Publisher Summary Acute aseptic meningitis (AM) is mostly a self-limiting, self-healing disease of acute onset and short duration, characterized by an immune response within the central nervous system (CNS) cerebrospinal fluid (CSF) compartment, which is reflected by abnormalities in humoral as well as cell-mediated immune variables in the CSF. This chapter explains the spread of virus in the CNS and discusses experiments conducted over the last few years regarding the humoral and cell-mediated immune responses in AM. Host-dependent factors, including sensitized lymphocytes and antiviral antibodies, are responsible for inhibition of viral spread and elimination of the infection. Abnormalities occurring in the form of increased immunoglobulin concentrations and oligoclonal bands are observed. Careful dissection of the humoral and cell-mediated immune reactions within the CNS as reflected in CSF, compared to peripheral blood, in patients with AM, provides fundamental information about the localized immune response in the CNS–CSF, improving the ability to interpret and utilize new data about immune reactions.

Studies on B-Lymphocyte Function in Multiple Sclerosis

Among abnormalities reported in multiple sclerosis (MS) and considered to reflect derangements of the immune system, those of B-lymphocyte function within the CNS-CSF compartment are generally accepted as the most consistent and clear-cut. These abnormalities have recently been extensively reviewed (Trotter and Brooks, 1980; Brooks et al., 1983; Walsh and Tourtellotte, 1983; Walsh et al., 1983). They include elevation of the CSF-IgG/total protein ratio (Kabat et al., 1948) and the CSF-IgG index, equal to (CSF-IgG/serum-IgG): (CSF-albumin/serum-albumin) (Tibbling et al., 1977), accentuated predominance in CSF of IgGl (Palmer and Minard, 1976) and of IgG kappa (Link and Zettervall, 1970), and oligoclonal distribution of IgG on separation of CSF by electrophoresis or isoelectric focusing on suitable media such as agar or agarose, in about 90%. MS patients heterozygous to Glm (1) and Glm (3) allotypes of IgGl showed greatly increased concentrations in the CSF of Glm[l] in comparison with patients with other neurological disorders indicating that, in the heterozygous MS patient, most plasma cells in the CNS-CSF compartment preferentially synthesize Glm (1) IgGl molecules (Salier etal., 1981). Furthermore, unexpected Gm allotypes may be found in CSF but not serum from an occasional MS patient (Salier et al., 1983). A majority of patients with MS display evidence for intrathecal synthesis of IgG class antibodies against various viruses, especially measles (for review see Norrby, 1978). Simultaneous intrathecal production of antibodies against 11 different viral antigens in individual MS patients has been reported (Salmi et al., 1983).