Myositis with sarcoplasmic inclusions in Nakajo-Nishimura syndrome: a genetic inflammatory myopathy.

AIMS: Nakajo-Nishimura syndrome (NNS) is an autosomal recessive disease caused by biallelic mutations in the PSMB8 gene that encodes the immunoproteasome subunit ß5i. There have been only a limited number of reports on the clinicopathological features of the disease in genetically confirmed cases. METHODS: We studied clinical and pathological features of three NNS patients who all carry the homozygous p.G201V mutations in PSMB8. Patients' muscle specimens were analysed with histology and immunohistochemistry. RESULTS: All patients had episodes of typical periodic fever and skin rash, and later developed progressive muscle weakness and atrophy, similar to previous reports. Oral corticosteroid was used for treatment but showed no obvious efficacy. On muscle pathology, lymphocytes were present in the endomysium surrounding non-necrotic fibres, as well as in the perimysium perivascular area. Nearly all fibres strongly expressed MHC-I in the sarcolemma. In the eldest patient, there were abnormal protein aggregates in the sarcoplasm, immunoreactive to p62, TDP-43 and ubiquitin antibodies. CONCLUSIONS: These results suggest that inflammation, inclusion pathology and aggregation of abnormal proteins underlie the progressive clinical course of the NNS pathomechanism.

Inhibition of PMA-induced, LFA-1-dependent lymphocyte aggregation by ADP ribosylation of the small molecular weight GTP binding protein, rho.

Botulinum C3 exoenzyme specifically ADP-ribosylates a group of ras-related small molecular weight GTP-binding proteins, rho, and inhibits their biological activity. Using this enzyme, we examined the function of rho in PMA-induced activation of lymphocyte function-associated antigen-1 (LFA-1) in a B lymphoblastoid cell line, JY. Northern blot analysis revealed that among the three rho genes, rhoA mRNA was predominantly expressed in JY cells. Consistently, only one [32P]ADP-ribosylated band was found when the lysate of the cells was subjected to ADP ribosylation by C3 exoenzyme. When the cells were cultured with C3 exoenzyme, this substrate was ADP-ribosylated in situ in a time- and concentration-dependent manner. Concomitant with this ADP ribosylation, PMA-induced LFA-1/intercellular adhesion molecule (ICAM)-1-dependent aggregation of JY cells was inhibited. This inhibition was blocked by prior treatment of the enzyme with an anti-C3 monoclonal antibody, and overcome by stimulation with higher concentrations of PMA. The C3 exoenzyme-induced inhibition was not affected by shaking of the cell suspension, while inhibition of aggregation by cytochalasin B was abolished by this procedure, suggesting that the inhibitory effect of the C3 exoenzyme treatment was not due to decrease in cell motility. The C3 exoenzyme treatment affected neither protein phosphorylation in JY cells before and after PMA stimulation, nor affected surface expression of LFA-1 and ICAM-1. These results suggest that rhoA protein works downstream of protein kinase C activation linking PMA stimulation to LFA-1 activation and aggregation in JY cells.

Prediction of postoperative clinical course by autologous tumor-killing activity in lung cancer patients.

Fifty patients with primary localized lung cancer were tested at the time of surgery for the ability of their lymphocytes to kill autologous, freshly isolated tumor cells, and the assay was evaluated for prognostic significance. Peripheral blood lymphocytes of 27 patients (54%) demonstrated significant autologous tumor-killing activity in 6-hour 51Cr-release assays. Twenty-three of the 27 patients with autologous tumor-killing activity remained tumor free and survived more than 5 years after curative surgery, while all 23 who were negative for autologous tumor-killing activity relapsed by 18 months after surgery and died within 42 months after surgery. The differences in survival curves for the two groups were highly significant (P less than .00003). Autologous tumor-killing activity was not correlated with natural killer (NK) cell activity against K562 human myeloid leukemia cells or proliferation of lymphocytes stimulated with autologous, freshly isolated tumor cells in mixed culture. There were no differences in total survival between patients with positive results and those with negative results in tests of NK cell activity and autologous mixed lymphocyte-tumor culture reaction. These results indicate that autologous tumor-killing activity is a meaningful prognostic indicator and provide evidence for immunological control of tumor growth and metastasis. According to our preliminary data, it is unlikely that lung cancer patients who remain tumor free after 60 months of follow-up will develop recurrence or die from the disease. We are conducting a study to determine whether induction of autologous tumor-killing activity before surgery, by treatment with biological response modifiers,can improve the clinical outcome in patients who do not naturally have this potential.

Redox regulation of a src family protein tyrosine kinase p56lck in T cells.

Protein tyrosine phosphorylation was examined after T cells were exposed to oxidative stress in vitro to investigate the possible involvement of redox regulation in T-cell signaling. Oxidative reagents such as hydrogen peroxide (H2O2) and diamide, which oxidize the free sulfhydryl groups in the cells, markedly induced tyrosine phosphorylation of multiple cellular proteins, especially a 55-kDa protein, of cultured peripheral blood T lymphocytes (PBL blasts). The 55-kDa molecule phosphorylated by diamide turned out to be a src family protein tyrosine kinase, p56lck. The immune complex kinase assay showed that the kinase activity of p56lck of diamide-treated PBL blasts was enhanced. The tryptic peptide mapping of p56lck demonstrated that diamide induced the phosphorylation both at Tyr-394 (autophosphorylation site) and at Tyr-505 (negative regulatory site). Taken together, the tyrosine phosphorylation and presumably kinase activity of p56lck were swiftly enhanced by oxidative stress, indicating that T cells have a redox-sensitive signaling mechanism, which is partly mediated by the lymphocyte-specific protein tyrosine kinase p56lck.

Involvement of CD11b/CD18 in enhanced neutrophil adhesion by Fc gamma receptor stimulation.

Neutrophils showed a rapid and transient adhesion to immunoglobulin G (IgG)-coated plates compared with their adhesion to bovine serum albumin (BSA)-coated plates: the adhesion reached a peak after 15 min of incubation and then gradually returned to almost the basal state in 60 min. The addition of monomeric IgG or anti-Fc gamma RII monoclonal antibody (mAb) (IV.3) suppressed the increase in adhesion, whereas anti-Fc gamma RIII mAb (3G8) was hardly effective, indicating that the interaction of Fc gamma R, especially Fc gamma RII, with coated IgG is involved in the process. Adhesion was also blocked by cytochalasin B, suggesting that functional actin filament structures are crucial. Protein kinase inhibitors, erbstatin and genistein, inhibited the adhesion in a dose-dependent manner. The adhesion was inhibited by anti-CD11b (M1/70) and anti-CD18 (MHM23, TS1/18) mAbs. Moreover, neutrophils from a patient with complete leukocyte adhesion deficiency syndrome did not show increased adhesion to IgG-coated plates. The adhesion of neutrophils to fibrinogen- and BSA-coated plates was also increased when Fc gamma R was stimulated in the fluid phase with soluble aggregated IgG, which was also inhibited by anti-CD11b mAb. Stimulation of neutrophil Fc gamma R with soluble aggregated IgG enhanced the expression of CD11b in concert with the enhanced adhesion. These data collectively suggest that stimulation via Fc gamma R evokes a tyrosine kinase-dependent and actin filament-dependent intracellular signal that enhances the specific and nonspecific adhesive activity of neutrophils, presumably through the activation of CD11b/CD18.

Induction of plant gp91 phox homolog by fungal cell wall, arachidonic acid, and salicylic acid in potato.

The oxidative burst has been suggested to be a primary event responsible for triggering the cascade of defense responses in various plant species against infection with avirulent pathogens or pathogen-derived elicitors. The molecular mechanisms of rapid production of active oxygen species (AOS), however, are not well known. We isolated homologs of gp91 phox, a plasma membrane protein of the neutrophil NADPH oxidase, from a potato cDNA library. Molecular cloning of the cDNA showed that there are two isogenes, designated StrbohA and StrbohB, respectively. The RNA gel blot analyses showed that StrbohA was constitutively expressed at a low level, whereas StrbohB was induced by hyphal wall components (HWC elicitor) from Phytophthora infestans in potato tubers. Treatment of potato tubers with HWC elicitor caused a rapid but weak transient accumulation of H2O2 (phase I), followed by a massive oxidative burst 6 to 9 h after treatment (phase II). Diphenylene iodonium (DPI), an inhibitor of the neutrophil NADPH oxidase, blocked both bursts, whereas pretreatment of the protein synthesis inhibitor cycloheximide with the tuber abolished only the second burst. These results suggest that the expression of StrbohA and StrbohB contributes to phase I and II bursts, respectively. The same is true for arachidonic acid, a lipid component of P. infestans-stimulated biphasic oxidative burst, whereas an endogenous signaling molecule, salicylic acid, only induced a weak phase II burst. Both molecules induced the StrbohB expression, which is in agreement with the second burst. To characterize the signal transduction pathway leading to the oxidative burst, we examined the role of protein phosphorylation in HWC-stimulated StrbohB gene expression. K252a and staurosporine, two protein kinase inhibitors, blocked the transcript accumulation. Two inhibitors of extracellular Ca2+ movement, however, did not abolish the transcript accumulation of StrbohB, suggesting that certain calcium-independent protein kinases are involved in the process of StrbohB gene expression. Additionally, we examined a causal relationship between the oxidative burst and expression of defense genes induced by the HWC elicitor. The transcript accumulation of genes related to sesquiterpenoid phytoalexin synthesis (lubimin and rishitin) and phenylpropanoid pathway was inhibited slightly by the DPI treatment, suggesting that the oxidative burst is not essential to activate these genes. Interestingly, the concomitant presence of DPI with the elicitor resulted in an increase in lubimin accumulation and a decrease in rishitin accumulation. Because it is known that lubimin is metabolized into rishitin via oxylubimin, we propose that AOS mediates the synthesis of rishitin from lubimin.

Ullrich disease: collagen VI deficiency: EM suggests a new basis for muscular weakness.

Ullrich disease is a form of congenital muscular dystrophy characterized clinically by generalized muscle weakness, contractures of the proximal joints, and hyperflexibility of the distal joints from birth or early infancy. Recently, mutations of the collagen VI gene have been associated with Ullrich disease. The authors report on a boy with Ullrich disease who has complete deficiency of collagen VI and harbors compound heterozygous mutations in the collagen VI alpha 2 gene. Absence of microfibrils on EM, together with normal collagen fibrils and basal lamina, suggests that loss of a link between interstitium and basal lamina may be a new molecular pathomechanism of muscular dystrophy.

Distal myopathy with rimmed vacuoles is allelic to hereditary inclusion body myopathy.

BACKGROUND: Distal myopathy with rimmed vacuoles (DMRV) is an autosomal-recessive disorder with preferential involvement of the tibialis anterior muscle that starts in young adulthood and spares quadriceps muscles. The disease locus has been mapped to chromosome 9p1-q1, the same region as the hereditary inclusion body myopathy (HIBM) locus. HIBM was originally described as rimmed vacuole myopathy sparing the quadriceps; therefore, the two diseases have been suspected to be allelic. Recently, HIBM was shown to be associated with the mutations in the gene encoding the bifunctional enzyme, UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE). OBJECTIVE: To determine whether DMRV and HIBM are allelic. METHODS: The GNE gene was sequenced in 34 patients with DMRV. The epimerase activity in lymphocytes from eight DMRV patients was also measured. RESULTS: The authors identified 27 unrelated DMRV patients with homozygous or compound-heterozygous mutations in the GNE gene. DMRV patients had markedly decreased epimerase activity. CONCLUSIONS: DMRV is allelic to HIBM. Various mutations are associated with DMRV in Japan. The loss-of-function mutations in the GNE gene appear to cause DMRV/HIBM.

Clinicopathological features of genetically confirmed Danon disease.

BACKGROUND: Danon disease is due to primary deficiency of lysosome-associated membrane protein-2. OBJECTIVE: To define the clinicopathologic features of Danon disease. METHODS: The features of 20 affected men and 18 affected women in 13 families with genetically confirmed Danon disease were reviewed. RESULTS: All patients had cardiomyopathy, 18 of 20 male patients (90%) and 6 of 18 female patients (33%) had skeletal myopathy, and 14 of 20 male patients (70%) and one of 18 female patients (6%) had mental retardation. Men were affected before age 20 years whereas most affected women developed cardiomyopathy in adulthood. Muscle histology revealed basophilic vacuoles that contain acid phosphatase-positive material within membranes that lack lysosome-associated membrane protein-2. Heart transplantation is the most effective treatment for the otherwise lethal cardiomyopathy. CONCLUSIONS: Danon disease is an X-linked dominant multisystem disorder affecting predominantly cardiac and skeletal muscles.