High pH-Sensitive TRPA1 Activation in Odontoblasts Regulates Mineralization.

Calcium hydroxide and mineral trioxide aggregate are widely used for indirect and direct pulp capping and root canal filling. Their dissociation into Ca(2+) and OH(-) in dental pulp creates an alkaline environment, which activates reparative/reactionary dentinogenesis. However, the mechanisms by which odontoblasts detect the pH of the extracellular environment remain unclear. We examined the alkali-sensitive intracellular Ca(2+) signaling pathway in rat odontoblasts. In the presence or absence of extracellular Ca(2+), application of alkaline solution increased intracellular Ca(2+) concentration, or [Ca(2+)]i Alkaline solution-induced [Ca(2+)]i increases depended on extracellular pH (8.5 to 10.5) in both the absence and the presence of extracellular Ca(2+) The amplitude was smaller in the absence than in the presence of extracellular Ca(2+) Each increase in [Ca(2+)]i, activated by pH 7.5, 8.5, or 9.5, depended on extracellular Ca(2+) concentration; the equilibrium binding constant for extracellular Ca(2+) concentration decreased as extracellular pH increased (1.04 mM at pH 7.5 to 0.11 mM at pH 9.5). Repeated applications of alkaline solution did not have a desensitizing effect on alkali-induced [Ca(2+)]i increases and inward currents. In the presence of extracellular Ca(2+), alkaline solution-induced [Ca(2+)]i increases were suppressed by application of an antagonist of transient receptor potential ankyrin subfamily member 1 (TRPA1) channels. Ca(2+) exclusion efficiency during alkaline solution-induced [Ca(2+)]i increases was reduced by a Na(+)-Ca(2+) exchanger antagonist. Alizarin red and von Kossa staining revealed increased mineralization levels under repeated high pH stimulation, whereas the TRPA1 antagonist strongly reduced this effect. These findings indicate that alkaline stimuli-such as the alkaline environment inside dental pulp treated with calcium hydroxide or mineral trioxide aggregate-activate Ca(2+) mobilization via Ca(2+) influx mediated by TRPA1 channels and intracellular Ca(2+) release in odontoblasts. High pH-sensing mechanisms in odontoblasts are important for activating dentinogenesis induced by an alkaline environment.

Altered acetylation of proteins in patients with rheumatoid arthritis revealed by acetyl-proteomics.

OBJECTIVES: Post-translational modifications (PTMs) are often critical for the function of proteins as well as antigenicity of proteins. We here tried to elucidate alteration of PTMs in Rheumatoid arthritis (RA), focusing on acetylation. We applied acetyl-proteomics to peripheral blood mononuclear cells (PBMCs) to elucidate PTM difference between patients with RA and healthy donors. METHODS: Proteins, extracted from peripheral blood mononuclear cells (PBMCs) of 7 RA patients and 7 healthy donors, were separated by 2-dimansional electrophoresis. Acetylation ratios of each protein spot were estimated by the combination of Sypro Ruby staining and anti-acetylated lysine antibodies. Proteins highly acetylated in the RA group were identified by mass spectrometry. Focusing on α-enolase (ENO1), one of the identified proteins, involvement of histone deacetylases (HDACs) in the high acetylation was investigated. Furthermore, the effects of acetylation on the activity of ENO1 were investigated. RESULTS: In PBMCs from the patients with RA, 29 acetylated protein spots were detected. One of highly acetylated proteins in the RA patients was identified as ENO1. The acetylation of ENO1 was found to be regulated in part by HDAC1. The enzymatic activity of ENO1 was up-regulated by acetylation. CONCLUSIONS: Highly acetylated ENO1 may play roles in the pathophysiology of RA through the maintenance of activated lymphocytes by increasing glycolysis-derived energy supply.

Hypercoagulant states in malignant lymphoma.

The incidence of severe complications, such as disseminated intravascular coagulation (DIC) in malignant lymphoma, differs between clinical stages and histological types of the disease, but they occur frequently in stage IV or natural killer (NK) cell lymphoma. Patients with stage IV or NK cell lymphoma exhibit abnormal thrombotic and hemostatic states. One of the mechanisms in DIC might involve elevated cytokine expression by lymphoma cells stimulating the expression of tissue factor (TF) in blood cells or surrounding tissue. During chemotherapy for lymphoma, the white blood cell count was significantly reduced at days 1 and 3, but significantly increased at days 7 and 9. At day 7 of chemotherapy, leukocyte TF mRNA levels were significantly increased. Plasma concentrations of granulocyte elastase derived-XDP (GEXDP) levels correlated with D-dimer levels, suggesting that almost all elevated D-dimer is GE-XDP. C-reactive protein (CRP), GE-XDP and D-dimer were significantly elevated in patients with infection, DIC or acute respiratory distress syndrome (ARDS). Analysis of patients with DIC or ARDS revealed that TF mRNA correlated with D-dimer, and GE-XDP correlated with leukocyte count, CRP and D-dimer, suggesting that inflammatory changes due to thrombosis may cause the activation of leukocytes during chemotherapy.