J. Ilonen

Lymphocyte blast transformation responses and viral antibodies in relation to HLA antigens in multiple sclerosis.

Fifty four clinically stable multiple sclerosis (MS) patients and 54 age- and sex-matched control subjects were HLA-typed, and their responses to herpes simplex, measles, mumps and rubella antigens were examined by the lymphocyte blast transformation test and by serum antibody titrations. Blast transformation response to purified tuberculin (PPD), mitogen phytohaemagglutinin (PHA), pokeweed mitogen (PWN) and concanavalin A (Con A) and spontaneous proliferation of lymphocytes were also studied. MS patients differed from controls by higher antibody levels to measles and rubella viruses and by lower specific blast transformation responses to rubella and measles antigens. When the relative strength of transformation responses was measured, mumps and herpes simplex responses were also lower in MS patients than in controls. In addition, spontaneous lymphocyte proliferation of MS patients in 6-day cultures was lower than that of control lymphocytes. In mitogen stimulations there were no differences between whole groups, but the oldest patients had lower responses to PHA and Con A than their matched controls. The frequency of HLA-Dw2 was 56.6% in MS patients and 32.1% in controls. The patients with and without Dw2 differed from each other only by a lower specific response to PPD in the Dw2-positive group. The immunological response of Dw2-positive controls resembled that of MS patients: low transformation response to viral antigens, low spontaneous proliferation and elevated measles antibodies. This finding supports the function of a genetically determined type of immune responsiveness with low cell-mediated immunity and high levels of certain viral antibodies as one susceptibility factor in multiple sclerosis.

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.

Stimulation of lymphocytes from subacute sclerosing panencephalitis patients by defined measles virus antigens

Abstract The lymphoproliferative response of human peripheral blood mononuclear cells to different measles virus antigen preparations was studied with lymphocytes from 38 measlesseropositive healthy donors and 4 subacute sclerosing panencephalitis patients. The response was very weak or absent in all of the controls and in three of the subacute sclerosing panencephalitis patients. The fourth subacute sclerosing panencephalitis patient had fluctuating levels of lymphocyte stimulation by measles antigens. The response was very strong for several months and during this time the parameters of the test system were characterized. It was discovered that a membrane preparation of measles-infected cells caused stimulation equal to that of highly purified virions. Purified measles ribonucleoprotein also induced specific stimulation, although lower than that seen with other types of measles antigens. Results of experiments on stimulation kinetics and antigen dose responses were compatible with antigen-specific stimulation. Enriched T cells were more vigorously stimulated than unfractionated peripheral blood mononuclear cells suggesting that this transformation test is specific for T cells.

Proliferation of multiple sclerosis cerebrospinal fluid lymphocytes after stimulation with measles virus antigens.

A method for measuring the proliferative response of cerebrospinal fluid (CSF) lymphocytes upon stimulation with mitogens and viral antigens has been developed. The number of lymphoid cells in DNA synthesis after in vitro stimulation is counted and the results expressed as a percentage of the total number of surviving lymphocytes. Preliminary results on four MS patients indicated that their CSF lymphocytes responded considerably more strongly to purified measles virus antigen than did corresponding peripheral blood lymphocytes (PBL). The responses to crude herpes virus antigen were at least as strong in PBL as in CSF lymphocyte cultures from the same patients. The CSF lymphocytes responded slightly stronger to phytohemagglutinin than PBL did.

Mitogen and antigen stimulation of multiple sclerosis cerebrospinal fluid lymphocytes in vitro

Responses of cerebrospinal fluid (CSF) and peripheral blood (PB) lymphocytes from 20 MS patients to phytohemagglutinin (PHA), measles, rubella, mumps and herpes simplex virus antigens were followed during periods of from 6 to 13 months. Up to 6 examinations, each with 1-5 stimulants, were performed with a lymphocyte blast transformation test. Most of the patients responded with their CSF cells to PHA (14/19) and at least to some of the viral antigens tested (15/20) during the follow-up. Although the maximal responses of CSF and PB cells to PHA and measles virus antigen were of the same magnitude, non-reactive or weakly responding lymphocytes were more common in CSF than in PB. In 7 of 15 patients having viral antigen responsive CSF cells simultaneous reactivity to several antigens could be shown. The stimulation results of CSF lymphocytes did not correlate with the numbers of CSF leukocytes or the intrathecal IgG synthesis. A negative correlation was observed between the strength of the CSF cellular response to PHA or measles virus antigen and the rate of intrathecal antibody synthesis to measles virus antigen, suggesting that the stimulated cells may at least partially represent suppressor cells.

Lymphocyte blast transformation responses in measles infection.

Lymphocyte blast transformation responses were studied in eight patients with natural measles infection. A specific response to purified measles virus antigen and a weaker response to crude measles virus antigen were found after infection. The response to purified measles antigen appeared 1–3 weeks after the beginning of the rash, when the great number of proliferating mononuclear cells found in the peripheral blood during the rash was declining. In the first weeks after onset of the rash also leucocyte and lymphocyte numbers were decreased, and specific responses to purified tuberculin (PPD) and to rubella and mumps virus antigens were suppressed. In mitogen stimulation tests there was no significant suppression of phytohaemagglutinin and concanavalin responses in this small series, but the response to pokeweed mitogen was decreased. The responses to antigens other than measles virus antigens recovered in parallel with the increase of the measles‐specific response.

Production of viral antibodies in vitro by CSF cells from mumps meningitis and multiple sclerosis patients

Cerebrospinal fluid (CSF) cells from 4 mumps meningitis and 11 multiple sclerosis (MS) patients were cultured in vitro for 7 days with and without pokeweed mitogen (PWM) stimulation. The cells produced varying amounts of IgG without stimulation and no significant increase of IgG synthesis was observed after PWM stimulation. Antibodies against mumps, measles, rubella, herpes simplex, and adeno viruses were measured in the supernatants of the cultures by a sensitive enzyme immunoassay. In the mumps meningitis patients, the largest amount of antibody was against mumps virus but low amounts of antibodies with other specificities were also synthesized by CSF cells of one patient. The most commonly detected specificities in MS patients were against measles and rubella viruses, whereas antibodies against adeno and mumps viruses were detected in only one CSF cell supernatant. No antibodies produced against herpes simplex virus in vitro were detected in any of the supernatants. The amounts of viral antibodies produced in vitro and intrathecally were only partially correlated.

Detection of Antigen‐specific Cellular Immune Response by the in Vitro Production of T‐Cell Growth Factor

The amount of T‐cell growlh factor (TCGF) produced in vitro in antigen‐stimulated lymphocyte cultures was measured by induction of growth in cell dependent on the factor. The method distinguishes eight herpes‐simplex‐seronegative subjects from ten seropositive ones, and these results correlated with those of the lymphocyte blast transformation (LBT) test. The LBT responses and TCGF amounts produced in cultures with purified tuberculin (PPD) were also in good correlation (r = 0.833, P<0.001). Considerable‐levels of the factor were already present 12 h after the beginning of the culture. The method offers rapid quantification of cell‐mediated immunity to specific antigens.

NK activity and NK-like non-specific cytolysis after PPD, rubella and measles antigen stimulation in multiple sclerosis☆

Blood samples were collected from 14 pairs of multiple sclerosis (MS) patients and from age- and sex-matched healthy controls. The cytotoxic activity of peripheral blood mononuclear cells was tested against the K562 cell line on the day of collection and again after 3 days in vitro culture with medium or with purified protein derivative of tuberculin (PPD), inactivated rubella virus or inactivated measles virus antigen. Lymphocytes from MS patients had a lower spontaneous cytotoxic activity compared to the controls both on the day of collection (P less than 0.025) and after 3 days in culture with medium alone (P less than 0.025). The activity decreased during in vitro culture in both groups, but the decrease was greater among MS patients (P less than 0.05). In cultures with antigens, a strong increase of NK-like cell-mediated cytolysis (NK-like CMC) was noted especially in cultures stimulated with PPD. There were no significant differences in the increase of the activity among MS patients and control subjects. A significant correlation between the increase of NK-like CMC and the lymphocyte blast transformation response induced by each antigen was found.