Hiroto Kimura

Regulation of synthesis of osteoprotegerin and soluble receptor activator of nuclear factor-κB ligand in normal human osteoblasts via the p38 mitogen-activated protein kinase pathway by the application of cyclic tensile strain

Mechanical stress is thought to play an important role in bone remodeling. However, the correlation between mechanical stress and bone remodeling is poorly understood. In this context, using a model of cyclic tensile strain (CTS) toward human osteoblasts, synthesis of osteoprotegerin (OPG) and soluble receptor activator of nuclear factor-κB ligand (sRANKL), and the activation of mitogen-activated protein kinases (MAPKs) were examined. The application of 7%, 0.25-Hz CTS once a day for 4 h for 3 successive days simultaneously caused an increase of OPG synthesis and a decrease of sRANKL release and RANKL mRNA expression in osteoblasts. As for MAPKs activation in osteoblasts with the application of CTS, p38 MAPK was activated 10–20 min after the application of CTS, but extracellular signal-regulated kinase (ERK1/2) and c-Jun NH2-terminal kinase (JNK) were not activated by such application. Furthermore, when CTS was applied once a day for 4 h for 1, 2, or 3 successive days to osteoblasts, p38 MAPK activation was maintained during the 3-day period but ERK1/2 activation was downregulated from day to day, simultaneously. Then, when CTS was applied once a day for 4 h for 3 successive days to osteoblasts pretreated with the p38 MAPK inhibitor SB203580 for 1 h, OPG synthesis was dose-dependently suppressed and inhibition of sRANKL release and RANKL mRNA expression was abrogated. These results indicate that biological responses of OPG and sRANKL synthesis in osteoblasts to the application of CTS are regulated via the p38 MAPK pathway and suggest that CTS might modulate and regulate bone metabolism.

Characterization of Synergistic Induction of CX3CL1/Fractalkine by TNF-α and IFN-γ in Vascular Endothelial Cells: An Essential Role for TNF-α in Post-Transcriptional Regulation of CX3CL1

CX3CL1/Fractalkine, a chemokine specific to monocytes and NK cells, is induced synergistically by TNF-α and IFN-γ in vascular endothelial cells. However, the mechanism for this synergism remains unclear. This study explored the hypothesis that the CX3CL1 expression is regulated at a posttranscriptional level, which may responsible for the synergism between TNF-α and IFN-γ. Brief exposure of HUVECs to TNF-α led to a robust increase in IFN-γ–induced CX3CL1 production. We found that TNF-α stabilized CX3CL1 mRNA in HUVECs stimulated with IFN-γ. Cloning of 3′untranslated region (UTR) of CX3CL1 mRNA revealed the presence of a single copy of nonametric AU-rich element in its 3′UTR, and a luciferase reporter assay showed that a single AU-rich element is a crucial cis-element in the posttranscriptional regulation of CX3CL1. TNF-α treatment resulted in the phosphorylation of p38 MAPK and its downstream target, MAPK-activated protein kinase-2, but IFN-γ did not affect the levels of MAPK and MAPK-activated protein kinase-2 phosphorylation induced by TNF-α. Treatment of the cells with an inhibitor of p38 MAPK accelerated the decay of CX3CL1 mRNA induced by TNF-α or the combination of TNF-α and IFN-γ. Immunoprecipitation assay revealed that mRNA stabilizer HuR directly binds to 3′UTR of CX3CL1 mRNA. CX3CL1 expression is under control of posttranscriptional regulation, which is involved in the synergistic induction of CX3CL1 in response to the combined stimulation with TNF-α and IFN-γ.

Expression of tumor necrosis factor-α and interleukin-6 in oral squamous cell carcinoma

To explore the role of cytokines in tumor development and clinical manifestations, we examined the expressions of tumor necrosis factor‐α (TNF‐α) and interleukin‐6 (IL‐6) in tumor tissues obtained from 57 patients with oral squamous cell carcinoma (OSCC) and their relationships to pathological grade and staging. Enzyme‐linked immunosorbent assay on the tumor tissues demonstrated elevated concentrations of TNF‐α and IL‐6 proteins and upregulated mRNA levels were detected by the reverse transcription‐polymerase chain reaction method when compared to those in normal control tissues. These cytokines and their transcripts were localized in stromal macrophages and in the tumor cells in particular of the front area of tumor issues, possibly indicating active synthesis of these cytokines by tumor cells. Larger‐sized tumors (T3, 4) contained significantly greater levels of IL‐6 proteins than small‐sized tumors (T1, 2) (P < 0.05). The levels of these cytokines were significantly reduced in cases with effective pre‐treatment with radiation or anti‐cancer agents compared to those in the less effective group (P < 0.05, grade IIa vs. grade IV for both TNF‐α and IL‐6). The present study thus demonstrated enhanced expression of cytokines in OSCC tissues.

Lidocaine added to a tracheostomy tube cuff reduces tube discomfort.

Purpose: To examine whether lidocaine diffusion across an endotracheal tube cuff affects tracheostomy tube discomfort.Methods: Two tracheostomy tube cuffs were inflated with 5 ml lidocaine 4% solution and air at 20 cmH2O, and then placed in 20 ml distilled water at 37°C. After vigorous stirring, 100 µl of this water was then sampled immediately then 1, 2, 4, 8, 24 hr later to measure lidocaine concentration by high-performance liquid chromatography. Sixteen patients undergoing tracheostomy following oral cancer resection were randomly assigned to two groups: lidocaine (n=8) and placebo (n=8). A tracheostomy tube cuff was inflated with 5 ml lidocaine 4% or saline 0.9% and air to a cuff pressure of 20 cmH2O, in the lidocaine and placebo groups respectively. Tube discomfort was evaluated using a visual analogue scale at 0, 0.5, 1, 2 and 4 hr after lidocaine or saline administration. Neither analgesics nor sedatives was given during the evaluation period.Results: Lidocaine time-dependently diffused across the tracheostomy tube cuff. Thirty and 60 min after cuff inflation lidocaine concentrations in the water bath reached approximately 8 and 17 µg·ml−1 representing 160 and 340 µg in 20 ml of water, respectively. The VAS decreased from 53.5 ± 10.6 to 25.1±9.8 mm (P<0.01) 0.5 hr following lidocaine administration which continued until the end of evaluation period. In the placebo group, VAS did not change.Conclusion: Lidocaine diffusion across the tracheostomy tube cuff reduces tube discomfort.RésuméObjectif: Vérifier si la diffusion de lidocaïne au travers du ballonnet du tube endotrachéal a un effet sur l’inconfort lié à la canule de trachéotomie.Méthode: Deux ballonnets de canules de trachéotomie ont été gonflés avec une solution contenant 5 ml de lidocaïne à 4 % et de l’air dans 20 cmH2O, et placées ensuite dans 20 ml d’eau distillée à 37 °C. Après avoir vigoureusement agité le mélange, on a immédiatement prélevé 100 µl de cette eau, puis 1, 2, 4, 8, 24 h plus tard, pour mesurer la concentration de lidocaïne par chromatographie à haute performance. Seize patients devant subir une trachéotomie à la suite de la résection d’un cancer oral ont été répartis au hasard en deux groupes: lidocaïne (n=8) et placebo (n=8). Une canule de trachéotomie à ballonnet a été gonflée avec 5 ml de lidocaïne à 4 % ou une solution salée à 0,9 % et de l’air jusqu’à une pression 20 cmH2O, chez les patients des groupes lidocaïne et placebo, respectivement. L’inconfort du tube a été évalué à l’aide de l’échelle visuelle analogique à 0; 0,5; 1; 2 et 4 h après l’administration de lidocaïne ou de solution salée. Aucun analgésique ou sédatif n’a été administré pendant la période d’évaluation.Résultas: La diffusion de la lidocaïne au travers du ballonnet de la canule de trachéotomie était fonction du temps. Trente et 60 min après le gonflement du ballonnet, les concentrations de lidocaïne avaient atteint environ 8 et 17 µg·ml−1, ce qui représentait 160 et 340 µg dans 20 ml d’eau, respectivement. Les valeurs de l’EVA ont diminué de 53,5±10,6 à 25,1±9,8 mm (P<0,01) 0,5 h après l’administration de lidocaïne, laquelle a été poursuivie jusqu’à la fin de la période d’évaluation. Dans le groupe placebo, l’EVA n’a pas montré de modification.Conclusion: La diffusion de la lidocaïne au travers du ballonnet de la canule de trachéotomie a réduit l’inconfort lié au tube.

Cognitive function and number of teeth in a community-dwelling population in Japan

BackgroundIt has been reported that oral health is poor in elderly populations and is associated with poor cognition and dementia. The objective of this study was to examine the association between tooth loss and cognitive function in a community-dwelling population in Japan.MethodsWe examined the association between tooth loss and cognitive function in 462 Japanese community-dwelling individuals. The Mini-Mental State Examination (MMSE) was employed to measure global cognitive status. A multiple logistic regression analysis, with both crude and adjusted conditions for confounding factors, was used to assess the relationship between poor cognition and the number of remaining teeth.ResultsThe overall prevalence of poor cognition (MMSE ≤ 23) in this study population was 5.6%. Subjects with poor cognition were significantly older, less educated, scored lower in intellectual activity, and had fewer remaining teeth than those with normal cognition. According to the multiple logistic regression analysis, a lower number of teeth (0–10) was found to be a significant independent risk factor (OR = 20.21, 95% confidence interval = 2.20 to 185.47) of cognitive impairment.ConclusionsThis cross-sectional study on a Japanese community-dwelling population revealed relationships between tooth loss and cognitive function. However, the interpretation of our results was hampered by a lack of data, including socioeconomic status and longitudinal observations. Future research exploring tooth loss and cognitive function is warranted.

Production of growth related oncogene protein-α in human umbilical vein endothelial cells stimulated with soluble interleukin-6 receptor-α: role of signal transducers, janus kinase 2 and mitogen-activated kinase kinase

Growth-related oncogene protein-alpha (GRO-alpha) is a member of the C-X-C chemokine family with a wide variety of biological activities. We studied the production of GRO-alpha by human umbilical vein endothelial cells (HUVEC) in response to the stimulation with soluble form of interleukin-6 receptor alpha (sIL-6R). sIL-6R stimulated HUVEC to express GRO-alpha mRNA and secrete GRO-alpha protein in concentration-and time-dependent manners. The sIL-6R-induced GRO-alpha expression was inhibited by the pretreatment of the cells with AG490, a janus kinase 2 (JAK2) inhibitor, or with U0126, a MAP kinase-ERK kinase (MEK) inhibitor. sIL-6R also induced the phosphorylation of both Src homology 2-protein tyrosine phosphatase-2 (SHP-2), signal transducer and activator of transcription 3 (STAT3) and MEK. AG490 pretreatment inhibited the MEK phosphorylation but did not affect the STAT3 phosphorylation. We conclude that sIL-6R induces GRO-alpha expression in HUVEC through the activation of JAK2 and MEK.

Influence of dentures in the initial occurrence site on the prognosis of bisphosphonate-related osteonecrosis of the jaws: a retrospective study.

OBJECTIVE This retrospective cohort study was conducted to investigate the influence of wearing dentures in the initial occurrence site of bisphosphonate-related osteonecrosis of the jaws (BRONJ). STUDY DESIGN A questionnaire regarding the prevalence, therapy, and outcome of jawbone lesions during 2006-08 was mailed to 248 medical institutions with an oral and maxillofacial surgery department in Japan. RESULTS Ninety-nine patients wearing dentures had significantly shorter duration until occurrence than 151 patients not wearing dentures. In addition, remission of BRONJ affecting the mandibular canine and premolar region in denture-wearing patients was significantly more difficult. Poor oral hygiene status was found to affect significantly the prognosis of BRONJ in denture-wearing patients. Alcohol habit also delayed remission, but high body mass index promoted remission. CONCLUSION Wearing a denture in the initial occurrence site of BRONJ was shown to influence the prognosis of BRONJ, especially in mandibular denture-wearing patients.

Interleukin-1β enhances the angiotensin-induced expression of plasminogen activator inhibitor-1 through angiotensin receptor upregulation in human astrocytes

Plasminogen activator inhibitor-1 (PAI-1) regulates not only fibrinolysis but extracellular matrix remodeling, and angiotensin II is known to play an important role in controlling the expression of PAI-1 in astrocytes. We have studied the effect of interleukin-1beta (IL-1beta), one of major cytokines also active in the nervous system, on the angiotensin II-induced expression of PAI-1 in human astrocytes. Cultures of normal human astrocytes were stimulated with IL-1beta and angiotensin II, and the expression of mRNAs for angiotensin II type 1 receptor (AT1) and PAI-1 was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) or real-time quantitative PCR. PAI-1 protein in astrocyte-conditioned medium was measured by enzyme-linked immunosorbent assay (ELISA). IL-1beta enhanced the expression of AT1 in astrocytes in time- and concentration-dependent manners. After 24-h stimulation, 10 ng/ml IL-1beta and 10 nM angiotensin II increased the levels of PAI-1 protein in astrocyte-conditioned medium by 1.9-fold and 1.8-fold of the basal value, respectively. There was no synergistic effect when the cells were stimulated simultaneously with IL-1beta and angiotensin II. When the cells were stimulated, with angiotensin II, 16 h after the stimulation with IL-1beta, the production of PAI-1 was enhanced by 1.4-fold as compared to the cells stimulated only with IL-1beta. CV-11794, an AT1 antagonist, inhibited the enhanced PAI-1 production in response to angiotensin II. We conclude that IL-1beta increases angiotensin II-induced PAI-1 secretion by astrocytes through the induction of AT1, and the enhanced secretion of PAI-1 may modulate functions of plasminogen activators in the nervous system.

Interleukin-1 induces the expression of vascular endothelial growth factor in human pericardial mesothelial cells

Vascular endothelial growth factor (VEGF) is a mitogen for endothelial cells. We have studied the production of VEGF by human pericardial mesothelial cells. Mesothelial cells were separated by scraping the pericardial surface during cardiac surgery and cultured. When stimulated with interleukin (IL)-1α, pericardial mesothelial cells expressed VEGF mRNA and protein in concentration- and time-dependent manners. Hypoxia was also found to enhance mesothelial VEGF mRNA expression. The cells expressed mRNA for Flt-1 (VEGF receptor 1) and Flk-1 (VEGF receptor 2), and exogenous VEGF was found to have migration-promoting activity on cultured cells. We conclude that pericardial mesothelial cells express VEGF, which may serve as an autocrine growth-regulatory mechanism.

Tissue specific expression and differential regulation by 1α, 25-dihydroxyvitamin D3 of the calcium-sensing receptor (CaSR) gene in rat kidney, intestine, and calvaria

The calcium-sensing receptor (CaSR) plays a critical role incalcium (Ca2+) homeostasis, and it exists in Ca2+ regulatory tissues such as parathyroid, kidney and intestine. As changes in the quality and quantity of CaSR mRNA may have an effect on sensing of extracellular Ca2+ concentration,we analyzed the ontogeny and regulation by 1α,25-dihydroxyvitamin D3 (1α,25(OH)2D3) of CaSR mRNA expression in the kidney, intestine, and bone. In 6-week-old rats, CaSR mRNA was expressed as a majortranscript of 8.5 kb and as minor transcripts of 4.8 and 2.5 kb in the kidney, whereas it appeared as faint transcripts of 8.5, 4.0 and 2.5 kb in the intestine and calvaria.These results showed that CaSR mRNAs were expressed indifferent structures among these organs. Moreover, the levelof CaSR mRNA increased in the kidney from the embryo to theadult. In contrast, the CaSR mRNA level decreased in theintestine during this transition, and the level of it did notchange in the calvaria. Moreover, 1α,25(OH)2D3 up-regulated the level of CaSR mRNA in thekidneys in 6-week-old rats. On the other hand, the 1α,25(OH)2D3 did not affect the CaSR mRNA expression in the intestine or calvaria. We concluded that different transcripts of CaSR were expressed in rat kidney, intestine, and calvaria and that the level of CaSR mRNA was different atvarious developmental stages in the kidney and intestine. Morever, 1α,25(OH)2D3 regulated the expression of CaSR mRNA only in the kidney.