CagA delivered from Helicobacter pylori into gastric epithelial cells undergoes tyrosine phosphorylation and induces host cell morphological changes. Here we show that CagA can interact with Grb2 both in vitro and in vivo, which results in the activation of the Ras/MEK/ERK pathway and leads to cell scattering as well as proliferation. Importantly, this ability of CagA is independent from the tyrosine phosphorylation, which occurs within the five repeated EPIYA sequences (PY region) of CagA. However, the PY region appears to be indispensable for the Grb2 binding and induction of the cellular responses. Thus, intracellular CagA via its binding to Grb2 may act as a transducer for stimulating growth factor-like downstream signals which lead to cell morphological changes and proliferation, the causes of H. pylori-induced gastric hyperplasia.

Attachment of Helicobacter pylori to gastric epithelial cells induces various cellular responses, including the tyrosine phosphorylation of an unknown 145-kD protein and interleukin 8 production. Here we show that this 145-kD protein is the cagA product of H. pylori, an immunodominant, cytotoxin-associated antigen. Epithelial cells infected with various H. pylori clinical isolates resulted in generation of tyrosine-phosphorylated proteins ranging from 130 to 145 kD in size that were also induced in vitro by mixing host cell lysate with bacterial lysate. When epithelial cells were infected with [35S]methionine-labeled H. pylori, a radioactive 145-kD protein was detected in the immunoprecipitates with antiphosphotyrosine antibody or anti-CagA (cytotoxin-associated gene A) antibody. Consistently, the 145-kD protein recognized by the anti-CagA and antiphosphotyrosine antibodies was induced in epithelial cells after infection of wild-type H. pylori but not the cagA::Km mutant. Furthermore, the amino acid sequence of the phosphorylated 145-kD protein induced by H. pylori infection was identical to the H. pylori CagA sequence. These results reveal that the tyrosine-phosphorylated 145-kD protein is H. pylori CagA protein, which may be delivered from attached bacteria into the host cytoplasm. The identification of the tyrosine-phosphorylated protein will thus provide further insights into understanding the precise roles of CagA protein in H. pylori pathogenesis.

Statins have a variety of myotoxic effects and can trigger the development of inflammatory myopathies or myasthenia gravis (MG) mediated by immunomodulatory properties. Autoantibodies to 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) have been identified in patients with statin-associated myopathy. The purpose of the present study is to develop an enzyme-linked immunosorbent assay (ELISA) of anti-HMGCR antibodies and to elucidate the clinical significance of anti-HMGCR antibodies in Japanese patients with inflammatory myopathies or MG.We enrolled 75 patients with inflammatory myopathies, who were all negative for anti-signal recognition particle and anti-aminoacyl transfer RNA synthetase antibodies. They were referred to Keio University and National Center of Neurology and Psychiatry between October 2010 and September 2012. We also studied 251 patients with MG who were followed at the MG Clinic at Keio University Hospital. Anti-HMGCR antibodies were detected by ELISA. We investigated demographic, clinical, radiological, and histological findings associated with anti-HMGCR antibodies.We established the anti-HMGCR ELISA with the recombinant protein. Protein immunoprecipitation detected autoantigens corresponding to HMGCR. Immunohistochemistry using muscle biopsy specimens revealed regenerating muscle fibers clearly stained by polyclonal anti-HMGCR antibodies and patients’ serum. Anti-HMGCR autoantibodies were specifically detected in 8 patients with necrotizing myopathy. The seropositivity rate in the necrotizing myopathy patients was significantly higher than those in the patients with other histological diagnoses of inflammatory myopathies (31% vs 2%, P = 0.001). Statins were administered in only 3 of the 8 anti-HMGCR-positive patients. Myopathy associated with anti-HMGCR antibodies showed mild limb weakness and favorable response to immunotherapy. All 8 patients exhibited increased signal intensities on short T1 inversion recovery of muscle MRI. Of the 251 patients with MG, 23 were administered statins at the onset of MG. One late-onset MG patient experienced MG worsening after 4-wk treatment with atorvastatin. However, anti-HMGCR antibodies were not detected in the 251 MG patients except for one early-onset MG patient with no history of statin therapy.Anti-HMGCR antibodies are a relevant clinical marker of necrotizing myopathy with or without statin exposure, but they are not associated with the onset or deterioration of MG.