Anne R. Moskovitz

Immune reactions to P2 protein in human inflammatory demyelinative neuropathies

The significance of immune reactions against peripheral nervous system antigens in the human inflammatory polyneuropathies is still uncertain. Using a very sensitive assay, we found greatly increased levels of anti-P2 antibodies in sera of animals with experimental allergic neuritis (EAN) but no increases in humans with acute Guillain-Barre syndrome (GBS), chronic relapsing polyneuritis (CRIP), axonal neuropathy, or normals. P2 protein and CNS basic protein did not induce any increased proliferation in lymphocytes of GBS or CRIP patients. We conclude that P2 and BP appear unlikely to be targets for humoral or cellular immune reactivity in GBS or CRIP.

Cellular inflammatory responses during immediate, developing, and established late-phase allergic cutaneous reactions: Effects of cetirizine

BACKGROUND In some previous studies, the antihistamine cetirizine has inhibited both developing (at 6 hours) and established (at 24 hours) gross late-phase skin reactions (LPR) to pollen antigens, possibly relevant to clinical drug effects. However, the effects of cetirizine at the histologic level require further definition. OBJECTIVE To characterize cetirizine effects on gross and histologic inflammatory events from 20 minutes to 24 hours after intradermal antigen challenge in sensitive patients. METHODS Gross and histologic responses to intradermal pollen antigen, codeine, histamine, and buffer diluent were assessed during randomized 7-day treatments with cetirizine and placebo. Accumulated neutrophils, eosinophils, activated (EG2+) eosinophils, and T lymphocytes were quantitated. The degrees of extracellular deposition of lactoferrin from neutrophils and eosinophilic cationic protein (ECP) from eosinophils were also assessed. RESULTS During placebo treatment, wheal-and-flare responses were significantly greater to antigen at 20 minutes (p < 0.01) and induration at 6 hours (p < 0.01) at antigen challenge sites than at buffer diluent sites. During cetirizine treatment, these wheal-and-flare responses to antigen were inhibited significantly (p < 0.01) but gross LPRs were not affected. During placebo treatment, significantly more cells per high-power field were found in antigen sites than in buffer sites of neutrophils at 20 minutes (p < 0.01) and 24 hours; than in eosinophils at 20 minutes, 6 hours, and 24 hours (p < 0.01 for each); than in EG2+ cells at 20 minutes (p = 0.004), 6 hours (p = 0.001), and 24 hours (p = 0.02); and at T lymphocyte sites at 24 hours (p = 0.001). Extracellular deposition of lactoferrin and ECP was significantly greater at antigen sites than at buffer sites at 6 and 24 hours. Cetirizine treatment had no significant effect on these responses. CONCLUSION Neutrophils, eosinophils, and T lymphocytes were persistently more common at antigen sites than at buffer sites through 24 hours. Many of these neutrophils and eosinophils were activated, releasing more lactoferrin and ECP into the extracellular dermis for at least 24 hours after antigen challenge. Cetirizine inhibited gross immediate responses to antigen, but not the gross LPR nor the cellular inflammatory responses seen in such LPR sites.

Thymic B‐cell activation in myasthenia gravis

We studied secretion of immunoglobulin (Ig) by freshly isolated and pokeweed mitogen (PWM)- stimulated thymus cells and blood mononuclear cells in patients with myasthenia gravis (MG) and control subjects undergoing elective cardiac surgery. We used a protein A reverse hemolytic plaque assay to enumerate cells secreting IgG, IgM, and IgA (IgSC), and an ELISA assay for measuring IgG secreted into culture supernatants. We found that freshly isolated suspensions of MG thymus cells, compared with control thymus cells, contained increased numbers of cells that spontaneously secreted immunoglobulin. Thymus mononuclear cells from control as well as MG patients appeared capable of B-cell differentiation responses when stimulated by PWM. PWM-induced responses were greater in thymic than in autologous blood mononuclear cells in some MG patients and controls, although B cells were much less frequent in suspensions of thymic cells than blood cells. Thus, the thymus provides a favorable milieu for differentiation of its few B cells. In MG, the thymus may be a site of accentuated in vivo B-cell activation, as evidenced by increased numbers of resident IgSC.

In vitro synthesis of antibodies to acetylcholine receptor by peripheral blood mononuclear cells of patients with myasthenia gravis

We studied the in vitro synthesis of antibodies to acetylcholine receptor (anti-AChR) by peripheral blood mononuclear cells (PBM) of patients with myasthenia gravis (MG) and normal subjects (NS). PBM from three of eight patients with generalized MG (MG-G) synthesized anti-AChR in vitro in the absence of pokeweed mitogen (PWM), and seven of eight did so in the presence of PWM. In individual subjects with MG-G, the levels of anti-AChR secreted in vitro by PBM correlated with serum anti-AChR antibody levels (r = 0.77) but not with the amount of IgG secreted in vitro (r = 0.44). No anti-AChR secretion was seen in culture of PBM from a patient with ocular MG, a patient with thymoma without MG, or six NS.

Cellular inflammatory responses and mediator release during early developing late-phase allergic cutaneous inflammatory responses: Effects of cetirizine

BACKGROUND Events in developing cutaneous late-phase allergic reactions can be characterized by a combination of skin chamber and biopsy approaches. In some previous studies, cetirizine reportedly inhibited mediator release and/or inflammatory cell responses in late-phase reactions. OBJECTIVE This study was carried out to determine the effects of cetirizine on early late-phase reactions by using skin chamber and skin biopsy specimens. METHODS Skin chamber responses during a 6-hour challenge with pollen antigens were assessed in 15 sensitive subjects during randomized, crossover treatment with cetirizine (20 mg/day) or placebo for 7-day periods with measurements of humoral and cellular components. Biopsy specimens of the underlying dermis were obtained. RESULTS During cetirizine treatment, there was significant (p < 0.01) inhibition of immediate wheal and flare reactions to pollen antigens (34, 46%), codeine (41, 65%), and histamine (38, 68%). However, gross late-phase reactions at 6 hours were unaffected. During both cetirizine and placebo treatment, there was significantly greater accumulation at antigen sites in: (1) skin chamber levels of histamine, total cells, lactoferrin, and eosinophil cationic protein; (2) eosinophils (total and activated) on appended cover glasses; (3) deposition of lactoferrin and eosinophil cationic protein in the underlying dermis. However, these responses were not significantly different during cetirizine treatment compared with placebo treatment periods. CONCLUSION A persistent pattern of inflammatory cell accumulation with release of granule proteins during early late-phase reactions was unaffected by cetirizine treatment.

In vitro synthesis of antibodies to acetylcholine receptor by peripheral blood cells Role of suppressor T cells in normal subjects

Peripheral blood mononuclear cells of 15 of 20 patients with generalized myasthenia gravis synthesized antibodies to acetylcholine receptor (AChR) when the cells were stimulated in vitro with pokeweed mitogen. In contrast, mononuclear cells of 1 of 16 normal subjects synthesized detectable AChR antibodies. Peripheral blood mononuclear cells of five normal subjects were studied before and after putative suppressor T cells(OKT8+) were removed by a fluorescent activated cell sorter. Depletion ofOKT8+ cells did not result in production of AChR antibodies, but pokeweed mitogen-induced polyclonal IgG synthesis and activation of B cells to form immunoglobulin-secreting cells (reverse hemolytic plaque assay) were increased. Therefore, failure of blood mononuclear cells of normal subjects to synthesize detectable anti-AChR in response to pokeweed mitogen is not due to suppression by OKT8+ cells.

Circulating T-cell subsets in Guillain-Barré syndrome

We compared the distribution of circulating T-cell subsets within 2 weeks of onset of symptoms in 14 patients with acute Guillain-Barré syndrome (GBS) and 37 normal controls. The levels of OKT4+ (putative helper-inducer) cells was definitely abnormal (decreased) in 3/13 tested. The levels of OKT8+ (putative suppressor-cytotoxic) cells were elevated in 3 and decreased in 2 of the 14 tested. Abnormal OKT4/OKT8 ratios were detected in 5 (2 elevated and 3 decreased) patients. Four of the 5 GBS patients with abnormal OKT4+/OKT8+ ratios were studied sequentially at least 4 times over 1-10 months; there was a return towards a normal ratio in all. Serial studies in 3 other GBS patients showed consistently normal values. In comparison, sequential studies over 5-24 months in 7 normals showed no abnormal OKT4+/OKT8+ ratios. Thus, abnormalities in OKT4+/OKT8+ ratios appear to be a marker of systemic events during symptomatic phases of GBS. It is not as yet known whether this is related to the cause or is secondary to the clinical manifestations of GBS.

Antibodies to P2 and P1 myelin antigens in experimental allergic neuritis and allergic encephalomyelitis

We confirmed earlier observations that experimental allergic neuritis (EAN) in Lewis rats induced by injection of bovine peripheral nerve myelin in complete Freunds adjuvant is not accompanied by development of experimental allergic encephalomyelitis. However, sera of these animals contained elevated titers of antibodies against central nervous system myelin basic protein (BP), likely induced by peripheral nerve P1 protein. Anti-BP antibodies were not seen in sera of rats with EAN induced by peripheral nerve P2 protein. Lack of encephalogenicity of bovine myelin in Lewis rats does not reflect simply lack of immune responses induced against BP.

Thymic lymphocyte subpopulations in myasthenia gravis

We compared the percentages of thymic mononuclear cells (TMC) that bind monoclonal antibodies to T-cell subpopulations in patients with myasthenia gravis (MG) and non-MG patients undergoing cardiac surgery (CS). There were no significant differences in percentages of OKT3+, OKT4+, OKT6+, or OKT8+ cells or the OKT4:OKTB ratio. There was an increase in the percentage of Ia+ (immune response gene-associated antigen) TMC in MG compared with CS but no significant differences in B cells or phagocytic cells. The Ia+ cells could be abnormal B cells, activated T cells, or thymic dendritic cells.

Polyclonal B‐cell activity in myasthenia gravis

Using a protein A-reverse hemolytic plaque assay, we found that some patients with myasthenia gravis have increased numbers of circulating immunoglobulin secreting cells (IgSC). This pattern was not related to drug therapy, age, sex, duration of symptoms, thymectomy, or serum levels of AChR antibody, although elevated IgSC values tended to occur in patients with active symptoms. The responses of peripheral blood mononuclear cells to pokeweed mitogen were normal. These data suggest increased in vivo polyclonal B-cell activation in some myasthenic patients, although in vitro polyclonal B-cell activation is normal.