Kingsley J. Micklem

Fewer thymic changes in MuSK antibody-positive than in MuSK antibody-negative MG

In generalized myasthenia gravis (MG) patients without detectable acetylcholine receptor (AChR) antibodies (SNMG), the thymus is often reported as “normally involuted.” We analyzed thymic compartments in 67 patients with generalized MG, with AChR antibodies (AChR+, n = 23), with muscle‐specific kinase (MuSK) antibodies (MuSK+, n = 14) or with neither (MuSK−, n = 30), and in 11 non‐MG controls. Four of 14 MuSK+ thymi had rare small germinal centers, but overall they were not different from age‐matched controls. However, approximately 75% MuSK− samples showed lymph node–type infiltrates similar to those in AChR+ patients, but with fewer germinal centers. These variations may explain some apparent differences in responses to thymectomy in SNMG. Ann Neurol 2005;57:444–448

Thymic myoid cells and germinal center formation in myasthenia gravis; possible roles in pathogenesis.

In early-onset myasthenia gravis (EOMG), the thymus usually shows medullary lymph node-type infiltrates, rearranged bands of hyperplastic epithelium and focal fenestrations in the intervening laminin borders. This resemblance to autoimmune target organs may reflect autoantigen expression by rare thymic myoid cells, long ago implicated circumstantially as agents provocateurs. In this quantitative study, they were frequently seen at the laminin fenestrations; if so, germinal centers (GC) were significantly commoner nearby, our most EOMG-specific finding (not seen in a distinct MG patient subset). As autoantibodies became detectable, myoid cell involvement apparently progressed. Our unifying hypothesis--that an early autoantibody attack on myoid cells provokes local GC formation--helps to resolve many puzzles.

A novel method for measuring intracellular pH and potassium concentration

The concentration of Na+and K+ and the pH in the cytoplasm of Lettré cells was measured by monitoring the net flux of H+, Na+, or K+ across the plasma membrane which had been rendered permeable to these ions by the action of Sendal virus. Ion flux was measured directly by analysis of cell composition, or indirectly by observing the change in membrane potential of cells treated with a specific ionophore. Cytoplasmic concentrations of cations were obtained by establishing the concentration of the cation in the medium at which addition of Sendai virus causes no change in cytoplasmic cation content. The value of Lettré-cell pH was confirmed by direct measurement employing 3tp nuclear magnetic resonance, and the values of Na+ and K+ concentration were confirmed by analysis of cell cation and water content. Lettré cells suspended at 32°C in Hepes-buffered saline at pH 7.3 maintain a cytosolic pH of 7.0 and contain 30 mM Na+ and 80 mM K+.

Protection against complement-mediated cell damage by Ca2+ and Zn2+.

Ca2+ and Zn2+ prevent antibody-dependent complement-induced permeability changes in tonsil lymphocytes and Lettre cells. Lactate dehydrogenase leaks out from Lettre cells at high complement:cell ratios, under which conditions higher concentrations of Ca2+ and Zn2+ are required for protection. Ca2+ and Zn2+ do not inhibit complement activation or C9 binding to Lettre cells, and prevent leakage through preformed lesions. It is concluded that the extent of complement-induced membrane damage depends on the concentration of extracellular Ca2+, and may be modulated by changes in extracellular Ca2+ or Zn2+.

The generation of active fragments of complement receptor type 2 by trypsin digestion

B lymphocytes and Raji cells express the complement receptor type 2 (CR2) of 145 kDa which recognises the C3d fragment of C3. When intact cells are treated with trypsin, CR2 is degraded. There is a parallel loss in C3d‐mediated rosetting and in proteins which bind to C3d‐Sepharose. Initially 97 and then 83 kDa fragments of CR2 are produced which retain C3d binding activity. These fragments are associated with the cell surface and mediate rosetting. Purified 125I‐labelled CR2, solubilised in detergent, produces fragments of apparently identical size on treatment with trypsin. The 83 kDa fragment produced by trypsin treatment closely resembles the major C3d binding protein spontaneously released into Raji cell culture medium.

Ion Permeability and Membrane Potential in Viral Disease

The ionic content of cells is an important determinant of their function. A raised intracellular Na+ in lens cells has been shown to be a direct cause of cataract (Piatigorsky, 1980) and changes in intracellular monovalent cations, as well as in Ca2+ have been implicated in diseases varying from hepatitis (Alam et al., 1978) to hypertension (Reid, 1982; Swales, 1982). Indeed, in many diseases in which the surface membrane of cells is primarily affected, it is an altered cation content that appears to be at fault (Griffin and Pasternak, 1982). Following our observation that certain viruses make cells permeable to low molecular weight compounds and ions without lysis (Pasternak and Micklem, 1973, 1974), we have been studying the effects of viruses on surface membrane function: our aim has been to determine to what extent such changes underly the processes of viral infection and disease. In this report we concentrate on changes in ion permeability and membrane potential.