Tomonori Uchimura

The N-terminal domain of thrombomodulin sequesters high-mobility group-B1 protein, a novel antiinflammatory mechanism

Thrombomodulin (TM) is an endothelial anticoagulant cofactor that promotes thrombin-mediated formation of activated protein C (APC). We have found that the N-terminal lectin-like domain (D1) of TM has unique antiinflammatory properties. TM, via D1, binds high-mobility group-B1 DNA-binding protein (HMGB1), a factor closely associated with necrotic cell damage following its release from the nucleus, thereby preventing in vitro leukocyte activation, in vivo UV irradiation-induced cutaneous inflammation, and in vivo lipopolysaccharide-induced lethality. Our data also demonstrate antiinflammatory properties of a peptide spanning D1 of TM and suggest its therapeutic potential. These findings highlight a novel mechanism, i.e., sequestration of mediators, through which an endothelial cofactor, TM, suppresses inflammation quite distinctly from its anticoagulant cofactor activity, thereby preventing the interaction of these mediators with cell surface receptors on effector cells in the vasculature.

Endocannabinoid, anandamide in gingival tissue regulates the periodontal inflammation through NF-κB pathway inhibition

Anandamide (AEA) exhibits anti‐inflammatory effects. However, its role in the periodontal field remains unknown. Here, we found that gingival crevicular fluid contained a detectable level of AEA. The cannabinoid receptors CB1 and CB2 were expressed by human gingival fibroblasts (HGFs), and markedly upregulated under pathological conditions. AEA significantly reduced the production of pro‐inflammatory mediators (IL‐6, IL‐8 and MCP‐1) induced by Porphyromonas gingivalis LPS in HGFs, and this effect was attenuated by AM251 and SR144528, selective antagonists of CB1 and CB2, respectively. Moreover, AEA completely blocked LPS‐triggered NF‐κB activation, implying that AEA may regulate hyperinflammatory reactions in periodontitis.

High Mobility Group Box Chromosomal Protein 1 in Patients with Renal Diseases

Background/Aim: The high mobility group box chromosomal protein 1 (HMGB1), a nuclear DNA-binding protein, has recently been recognized as a new proinflammatory cytokine. The purpose of this study was to examine the significance of HMGB1 in patients with renal diseases. Methods: HMGB1 concentrations in sera were measured by enzyme-linked immunosorbent assay, and antibodies against HMGB1 were examined by Western blotting in patients who underwent renal biopsies and in healthy controls. Immunohistochemistry for HMGB1 was also performed. Results: Serum HMGB1 was more likely to be positive in patients who underwent renal biopsies as compared with the controls. Patients with anti-neutrophil cytoplasmic antibody-related glomerulonephritis (ANCA-GN) and those with Henoch-Schönlein purpura nephritis showed a significantly higher tendency to be HMGB1 positive. The presence of anti-HMGB1 antibody was not associated with the presence of serum HMGB1. Immunohistochemistry revealed that HMGB1 was expressed in mononuclear cells in the interstitium or in the glomeruli of some patients with ANCA-GN or IgA nephropathy (IgAN). Subanalysis demonstrated that among patients with IgAN, those who had crescent formation showed a higher tendency to be HMGB1 positive than those who did not. Conclusions: HMGB1 was expressed in the sera of patients with renal diseases who underwent renal biopsies, especially among those who had vasculitis including ANCA-GN, Henoch-Schönlein purpura nephritis, and IgAN with glomerular crescents.

Endogenous cannabinoids are candidates for lipid mediators of bone cement implantation syndrome.

Acute hypotension, hypoxemia, cardiac arrhythmias, cardiac arrest, (or a combination of these), and sudden death are well-recognized complications of the cemented hip arthroplasty procedure. Collectively, these are known as the bone cement implantation syndrome (BCIS). The endogenous cannabinoids, anandamide (ANA) and 2-arachidonylglycerol (2-AG), are reported to be strong vasodilators and play a role in the hypotension associated with hemorrhagic and septic shock. In the present study, a potential role for the endogenous cannabinoids in influencing hemodynamic variables in BCIS was investigated. Thirty-five patients (35 hips) entered a prospective, randomized clinical trial. The patients were divided into two groups. Group 1 comprised 16 patients who had the component inserted using a conventional cementing technique, whereas group 2 consisted of 19 patients who had the femoral component inserted without cement. Blood samples were taken at six consecutive time points: before anesthesia, after reaming the femur, 2 min after insertion of stems with or without cement into the femur, and 10 min, 20, and 30 min after stem insertion. In group 1 (with cement), the mean levels of ANA and 2-AG significantly increased after stem insertion. In a comparison of each group after stem insertion, mean ANA and 2-AG levels in group 1 also significantly differed from those in group 2. By contrast, in group 2 (without cement) neither ANA nor 2-AG levels exhibited a significant increase or change at any point in time. In conclusion, we have shown for the first time that endogenous cannabinoids are candidates for lipid mediators of BCIS.

Marked Increases in Macrophage Colony-Stimulating Factor and Interleukin-18 in Maintenance Hemodialysis Patients: Comparative Study of Advanced Glycation End Products, Carboxymethyllysine and Pentosidine

Background: Recent studies have demonstrated the crucial involvement of advanced glycation end products (AGEs) in major complications of long-term hemodialysis (HD) patients. HD, in a clinical setting, is characterized by increased production of proinflammatory cytokines. AGEs and cytokines are presumed to be responsible for the development of major complications in long-term HD. We therefore investigate here the relationship between a newly identified cytokine, interleukin-18 (IL-18), and two AGEs, carboxymethyllysine-hemoglobin (CML-Hb) and pentosidine. Methods: CML-Hb, pentosidine macrophage colony-stimulating factor (M-CSF), and IL-18 were evaluated in 35 patients undergoing stable maintenance HD. CML-Hb and pentosidine were measured by a dot blot and competitive ELISA. Cytokines were measured with a cytokine-specific ELISA. Results: Circulating levels of CML-Hb and pentosidine were elevated in HD patients as compared to controls. The serum values of M-CSF and IL-18 were significantly increased in the HD patients in comparison to controls. Moreover, these two AGEs and serum values of M-CSF, M-CSF and IL-18 showed significant correlation by simple and multiple regression analysis. Conclusion: Elevation of circulating IL-18 levels was demonstrated in maintenance HD patients relative to controls. A correlative increase in M-CSF and IL-18 suggests the presence of a primed state of monocytes/macrophages in HD patients.

Potential value of CML-Hb in predicting the progression of bone cysts in dialysis-related amyloidosis.

Background/Aim: Carboxymethyllysine is one of the common advanced glycation end products in vivo. In a previous report, we were the first to describe the increase of circulating carboxymethyllysine-hemoglobin (CML-Hb) levels in hemodialysis patients, particularly in patients with dialysis-related amyloidosis (DRA). The aim of this study was to investigate the predictive value of CML-Hb in the progression of DRA using computed tomography images of 2-year or 3-year follow-up periods of patients with amyloid bone cysts at the hip joint. Methods: Circulating CML-Hb levels were measured as previously reported, and computed tomography scanning was conducted from 1996 to 1998 or 1999 in 57 hemodialysis patients whose original renal disease had been confirmed to be nondiabetic. Patients who showed a new growth of cysts or a growth rate of 30% or more were classified as progressive cases, while the other patients were classified as nonprogressive cases. Results: (1) The circulating CML-Hb levels showed a strong correlation with the DRA score by multiple regression analysis, and (2) patients with progression of amyloid cysts showed a significantly higher circulating CML-Hb level than patients without progression. Conclusion: Based upon these results, we conclude that measurement of circulating CML-Hb levels has a potential value in both judgment of the clinical state and prediction of progression of DRA in hemodialysis patients.

Epinephrine prevents nitric oxide/peroxynitrite induced apoptosis of neuronal cells through ?-adrenergic receptor activation

In the present study, we demonstrate that the NO-donors sodium nitroprusside (SNP), S-nitroso-N-acetylpenicillamine (SNAP) and 3-morpholino sydnonimine (SIN-1) dose-dependently caused rat pheochromocytoma (PC-12) and human neuroblastoma (NB-1) cells death, determined by tryphan blue dye exclusion. That death was apoptosis, as determined by terminal-deoxynucleotide transferase-mediated dUTP-digoxigenin nick end-labeling (TUNEL) method and propidium iodide (PI) staining using fluorescence-activated cells sorting (FACS). Epinephrine exhibited pronounced inhibitory effect on this NO donors-induced cell death. Similar protecting effect was also found when cells were pretreated with β-adrenergic receptor agonist isoproterenol, adenylate cyclase activator forskolin, nerve growth factor (NGF) or antioxidant N-acetyl-L-cysteine (NAC). However, cycloheximide the inhibitor of protein synthesis, the transcription inhibitor actinomycin D (act D) or anti-NGF abolished the rescuing effect of epinephrine. Furthermore, SNP and SIN-1, which form peroxynitrite in the culture media were more effective to retract neurites of NB-1 cells compared to SNAP. These results suggest that NGF plays pivotal role in epinephrine-supported cytoprotection, and probably peroxynitrite is involved in the retraction of neurites of the neuronal cells.

Two advanced glycation molecules, carboxymethyllysine and pentosidine, and dialysis membrane in dialysis-related amyloidosis

Studies on advanced glycation end products will provide new scientific clues to the pathogenesis of dialysis-related amyloidosis, a crucial problem in dialysis patients. We previously reported our data onNε-(carboxymethyl)lysine-hemoglobin from maintenance hemodialysis patients. Further detailed data onNε-(carboxymethyl)lysine-hemoglobin and serum pentosidine are presented herein and discussed in relation to dialysis-related amyloidosis and dialysis membranes.Clinical features of dialysis-related amyloidosis and the history of the dialysis membrane used were thoroughly reviewed in 66 patients undergoing hemodialysis. Circulating levels ofNε-(carboxymethyl)lysine-hemoglobin were assayed by the dot-blotting method, and serum levels of pentosidine were measured by enzyme-linked immunosorbent assay.Circulating concentrations ofNε-(carboxymethyl)lysine-hemoglobin were increased to about twice those present in healthy persons, and serum levels of pentosidine were elevated to as high as 2402.8±1132.1 pmol/ml in 66 patients, as compared with 220.0±39.4 pmol/ml in healthy persons. Circulating levels ofNε-(carboxymethyl)lysine-hemoglobin correlated significantly with both the total length of dialysis history and the severity of the clinical features of dialysis-related amyloidosis. On the other hand, serum levels of pentosidine showed no correlation with either of them.These results strongly suggest that cellulosic dialysis membranes have a critical role in the development of dialysis-related amyloidosis. In addition, quite a large difference in the circulating levels of two advanced glycation end-product molecules,Nε-(carboxymethyl)lysine-hemoglobin and pentosidine, was found in the clinical setting of dialysis, and thus difference deserves further investigation.