T Kaku

Health Sciences University of Hokkaido, Tōbetsu, Hokkaido, Japan

Are you T Kaku?

Claim your profile

Publications (101)162.92 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: This study was conducted to examine the utility of the combined use of ascorbic acid (AsA) and radiation in clinical applications. We investigated cell survival, DNA fragmentation, and caspase activation after X-ray irradiation and AsA treatment of human leukemia HL60 cells. The number of living cells decreased after combined X-ray irradiation and AsA treatment (2 Gy + 5 mM) in comparison with that after X-ray irradiation (2 Gy) or AsA treatment (5 mM) alone. DNA fragmentation was more in the cells subjected to combined X-ray irradiation and AsA treatment than in those subjected to X-ray irradiation alone. Caspase-3, caspase-8, and caspase-9 were highly activated following combined X-ray irradiation and AsA treatment, but caspase-8 activity was not markedly increased after X-ray irradiation alone. Bax levels in the mitochondrial membrane fractions were increased after AsA treatment alone and after combined X-ray irradiation and AsA treatment. However, there was no apparent increase in the Bax levels after X-ray irradiation treatment alone. Thus, this study confirmed that supplementing X-ray irradiation with AsA treatment results in increased apoptosis in HL60 cells. With regard to the apoptosis-inducing factors, we hypothesized that Bax and caspase-8 were activated after combined X-ray irradiation and AsA treatment compared with either treatment alone.
    Journal of Radiation Research 02/2011; 52(2):229-37. · 1.45 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Human beta-defensins (hBDs), a group of antimicrobial peptides, are involved in the protective barrier of the oral epithelium. Nicotine induces periodontal and oral epithelial diseases. The purpose of the present study was to investigate the effect of nicotine on the expression pattern of hBD-2 in keratinocytes. HaCaT cells, a keratinocyte cell line, were incubated with 8, 15, 30, or 80 μM nicotine for 24 h. Expression of hBD-2 was observed by RT-PCR, qRTPCR, and ELISA assay. The cells were treated with inhibitors for intracellular pathways (p38MAP kinase, NF-κB, JNK, MAPK-ERK) and with nicotinic acetylcholine receptor (nAChR) inhibitors in a series of experiments. Data were analyzed using Student's t test. qRT-PCR revealed that the expression level of hBD-2 mRNA was significantly higher at 30 and 80 μM nicotine than the control without nicotine (P < 0.05). The 80 μM cell extraction contained significantly higher hBD-2 peptide levels than the control (P < 0.05). The p38MAP kinase inhibitor abolished the upregulated expression of hBD-2 by nicotine. Both nAChR inhibitors also abolished the upregulation of hBD-2 by nicotine. The present study demonstrated that nicotine causes upregulated expression of hBD-2 via the p38MAP kinase pathway in keratinocytes.
    Medical Molecular Morphology 12/2010; 43(4):204-10. · 1.17 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objectives: Although alteration of bone level and density around functional dental implants is well documented, there is little information about their alteration at other sites in dental implant procedures. The aim of the present study was to evaluate post-surgical bone level around submerged dental implants before secondary operation, and to evaluate the bone levels around natural teeth that oppose an implant. Methods: The radiographs taken for seventy-eight implants (32 implants of 3.25mm diameter, 46 implants of 4mm diameter; Spline, Zimmer dental) were used. The radiographs were obtained at the time immediately after the first surgery, and at 3 and 6 months after the surgery but before the second surgery. The radiographs taken for 80 natural molars that oppose an implant were used. The radiographs were obtained at 6, 9 and 12 months after the implant crowns were fixed. Contralateral molars in the same jaw that did not oppose an implant were used as controls. The images were manipulated by EMAGO/Advanced 5.x software (Oral Diagnostic system, Amsterdam, The Netherlands) and linear and logarithmic digital subtraction images were produced. The subtraction image was analyzed by NIH image. The data was analyzed using one-way ANOVA or F-test. Results: The implants of 4mm diameter showed significantly higher bone levels than the implants of 3.25 mm diameter both at 3 and 6 months after the first surgery (p<0.05). Significant bone loss or gain was often observed around the teeth that oppose implants at 6, 9 or 12 months after the implant crown was fixed. The variance of the data in the group of teeth that oppose an implant was significantly greater than that in the control group (p<0.05). Conclusions: The results indicate that it may be important to evaluate bone level and density in dental implant procedures as well as those around functional dental implants.
    IADR General Session 2010; 07/2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objective: Drosomycin-like defensin (DLD) was recently found as a putative novel human defensin with high homology with drosomycin. It may be central in the defense against fungal infection. Expression of DLD in oral tissues has not been shown thus far. The present study investigated expression of DLD mRNA in oral cells and tissues. Methods: Normal oral keratinocytes (NOK), six kinds of oral squamous cell carcinoma cell (SCC) lines (SCC-9, SAS, HSC-2, OSC-19, BSC-OF, Ca-9), salivary duct cell line (HSY) and human gingival fibroblasts were used. The cells except NOK were cultured in DMEM supplemented with 10% FBS. NOK were cultures in keratinocyte basal medium (KBM, LONZA, Switzerland) containing 0.3mM Ca2+ supplemented with Ultroser G (Life sciences, NY). In order to stimulate the keratinocyte differentiation, the NOK were cultured for 4 days after the KBM was placed by a 1.8mM Ca2+ KBM. Expression of DLD mRNA was observed by RT-PCR and quantitative RT-PCR using TaqMan probes (Applied Biosystem, CA). The relative expression of each mRNA was calculated as the ΔΔCt using the formula described by Saitoh et al. (Med Mol Morph, 2007). The data was analyzed using one-way ANOVA. Differences between experimental groups were considered statistically significant at the p<0.05 levels. For in situ hybridization in human oral tissues, DIG-labeled RNA probes for DLD were used. Results: The expression of DLD mRNA was observed in the cells. The expression level of DLD in HSY was significantly higher than that in the other cells (p<0.01). There was no significant difference in the expression level of DLD in NOK cultured between 0.3 mM Ca2+ and 1.8mM Ca2+. By in situ hybridization, faint signals for DLD transcripts were detected in the whole layer of squamous epithelium and ciliated columnar epithelium. Conclusion: These results indicate that DLD may be primarily expressed in oral epithelium.
    IADR General Session 2010; 07/2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objective: Human beta-defensin(hBD)s are a group of antimicrobial peptides. Oral epithelium is exposed to many kinds of external stresses including bacteria, food and drink. Bacterial infection induces the upregulated expression of hBD-3. Food and drink causes osmotic stresses to oral epithelium. The present study investigated the effect of osmostic stress on the expression of hBD-3 in keratinocytes. Methods: Keratinocyte cell line, HaCaT cells, were cultured in keratinocyte growth media (KGM, Lonza, MD, USA). HaCaT cells were cultured with hyperosmotic KGM with 50, 100, 200, or 300mM sorbitol for 24, 48 or 72 h. No addition of sorbitol to KGM was used as a control. We observed the expression levels of hBD-3 mRNA and peptides by quantitative RT-PCR using TaqMan probes and ELISA, respectively. In order to examine whether epithelial growth factor receptor (EGFR) is involved in the responses, the cells were pretreated with a inhibitor for EGFR tyrosine kinase, AG1478 [1, 5, 10 or 20μM] (Sigma, St. Louis, MO, USA) or anti-EGFR, Ab-5 [0.1, 0.5, 1.0, 2.0μg/ml] (Calbiochem, San Diego, CA, USA). In order to elucidate the intracellular pathway involved in the expression of hBD-3, phosphorylation of ERK and p38 MAPK were evaluated by Western blotting. The statistical significance of the differentiation was analyzed using Student t-test (n=5). Results: Osmostic stress by means of addition of sorbitol upregulated hBD-3 in HaCaT cells at both mRNA and protein levels. The osmotic stress induced ERK and p38 MAPK phosphorylation. Both AG1478 and Ab-5 inhibited the upregulated expression of hBD-3 induced by the stimulation with osmostic stress (p<0.05). Conclusion: These findings indicate that osmotic stresses by food and drink may induce the upregulated expression of hBD-3 via EGFR, and this pathway is possibly involved in ERK and p38 MAPK.
    IADR General Session 2010; 07/2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Human beta-defensins (hBDs) are innate antimicrobial peptides produced in keratinocytes, which play an important role in the protective barrier of oral epithelium. Butyrate, the dietary histone deacetylase inhibitor, is produced by periodontal pathogens such as porphyromonas gingivalis and fusobacterium nucleatum. Objectives: The present study investigated how butyrate affected the expression patterns of hBD-2 and -3 in keratinocyte. Methods: HaCaT cells, a human keratinocyte cell line, were grown in DMEM supplemented with 10% fetal bovine serum. The cells were incubated with butyrate (Sigma) at the concentration of 0, 0.5, 2.5, 5mM for 0.5, 6, 24 or 48 hours. The expression levels of the hBDs mRNAs were evaluated by RT-PCR followed by assessment of transcript induction, and the level of hBDs within supernatants were measured by sandwich ELISA. The ELISA assays were performed using capture and biotinylated antibodies for hBD-2 and -3 (Peprotech). In order to elucidate the intracellular pathway involved in the expression of hBDs, cells were pretreated with inhibitors [AG1478 (EGFR), SB203580 (p38MAPK), or JNKII (JNK)] for an hour in some experiments. The phosphorylation of MAP-kinases family was detected by western blotting with p-p38MAPK, p-ERK, and p-JNK antibodies (Santa Cruz). The significance of the differences was analyzed by Student's t-test (n=5). Results: Butyrate induced upregulated expression of hBD-2 and -3 at both mRNA and protein levels in dose- and time-dependent manners. The expression level of hBD-2 was the highest in the keratinocyte stimulated with 5mM butyrate for 48hr. The JNK II abolished the upregulated expression of hBD-3 induced by the stimulation with butyrate. The JNK II and the AG1478 inhibited butyrate-induced JNK and ERK activation. Conclusion: These results suggest that butyrate induces upregulated expression of hBD-2 and -3 in the keratinocyte, and the upregulated expression is possibly involved in MAPK pathway.
    IADR General Session 2010; 07/2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objectives: Epithelial antimicrobial peptides play an important role in protective barrier of oral epithelium against bacterial infection. Alteration of expression of antimicrobial peptides including beta-defensin (hBD) s, cathelicidin/LL-37 (LL-37) and psoriasin (PS) is often observed in carcinoma cells. Epigenetic modifications are often involved in the transcriptional level of mRNAs. The present study investigated how epigenetic modifications are involved in expression of the antimicrobial peptide. Methods: Six oral SCC-established cell lines (SCC-9, SAS, HSC-2-3, OSC-19, BSC-OF), keratinocyte cell line, HaCaT were used. The expression level of hBD-1, -2, -3 and LL-37 was estimated by quantitative RT-PCR using TaqMan probes (Applied Biosystem, CA). In order to examine whether DNA hypermethylation and/or histone deacetylation are involved in the transcriptional levels, cells were treated with DNA methyltransferase inhibitor, 5-aza-dCyd (Sigma, MO) for 72hr or histone deacetylase inhibitor, Trihcostatin A (TSA: Sigma) for 24hr. No treatments were used as controls. The relative expression of each mRNA was calculated as the ΔΔCt using the formula described by Saitoh et al. (Med Mol Morph, 2007). Student's-t test was used to determine if the average expression level in experiment and control were different. Results: The expression levels of hBD-1, -2, -3, LL-37 and PS varied among the cell lines. Expression levels of hBD-1, -2, -3, LL37 and PS in HSC-3, OSC-19 and BSC-OF treated with 50μM of 5-aza-dCyd were significantly higher than each control (p<0.001). HSC-3, OSC-19 and BSC-OF showed more than 10-fold induction of hBD-1 and LL37 expression. Expression levels of hBD-2, -3, LL-37 and PS in SCC-9, OSC-19 and BSC-OF treated with 300nM of TSA were significantly higher than each control (p<0.001). Conclusions: The results indicate that epigenetic modification may affect transcriptional level of antimicrobial peptides in some oral carcinoma.
    IADR General Session 2010; 07/2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objective: Human beta-defensin(hBD)s belong to a group of antimicrobial peptides expressed in epithelial cells. Diabetes is a risk factor in oral infections including periodontitis. The hyperglycemia, insulin and adiponectin deficiency/insensitivity may affect hBDs expression. Therefore, the present study investigated the effect of glucose, insulin and adiponectin on the expression of hBD-1, -2 and -3. Methods: Keratinocyte cell line, HaCaT cells were grown in DMEM supplemented with 10% fetal bovine serum. To examine the effect of the combination of glucose, insulin and adiponectin on the expression of hBDs mRNAs and proteins, the cells were incubated with 5.5, 10, 15 and 50mM of glucose and/or with 10 and 20μU/ml of insulin and/or with 3, 5, 10 and 30μg/ml of adiponectin for 24hr. The cells incubated without glucose, insulin, or adiponectin were used as controls. Expression of hBD1~3 in HaCaT cells were observed by RT-PCR and quantitative RT-PCR using TaqMan probes (Applied Biosystem, CA). The data was analyzed using one-way ANOVA. To evaluate peptide expression level of hBDs, an ELISA assay was carried out. We combined a sandwich ELISA with the best commercially available capture and detection antibodies (Peprotech, NJ). Fold induction represents the mean SD of triplicate experiments. P values were calculated by student's-t test. Results: No significant differences are observed in the expression of hBDs between any concentrations of glucose. Expression levels of hBD-2 in 200nM of insulin were significantly higher than those in the controls. Expression levels of hBD-1 in 30μg/ml of adiponectin were significantly higher than those in the controls. Combination of 50mM of glucose, 20μU/ml of insulin and 5μg/ml adiponectin stimulated the highest levels of the hBDs in each experiment. Conclusion: The results indicated that optimal concentration of glucose insulin and adiponectin might be needed to maintain a high expression level of hBDs.
    IADR General Session 2010; 07/2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objective: Diabetes increases susceptibility to infection, possibly leading to delayed wound healing. The differentiation and proliferation pattern of the keratinocytes during the process in diabetes is, however, still unclear. The present study investigated the pattern of differentiation and proliferation of keratinocytes during oral wound healing in diabetic mice. Method: Female non-obese diabetic (NOD) mice (CLEA, Japan) and BALB mice (CLEA) were used as animal models of diabetes and controls, respectively. Superficial circular punch biopsy wound measuring ~ 2.0mm was made in the middle of the tongue using a 1.0mm Biopsy Punch (Ft.Lauderdale, FL) by ablating the epithelial layer without damage to the underlying muscle (Nagy et al. Diabetes, 2001). The animals were euthanized at 1, 2, 3, 4 and 6 days after the wound were made. The wound size was calculated using the formula A=LWp/4. Frozen sections were taken from the tongues dissected from the animals. Immunohistochemical staining was carried out using anti-Keratin 10, 14 and 16 antibodies and anti-Ki67 as primary antibodies. Immunodetection kit, peroxidase was used to detect the immunoreactions according to manufacture's protocol (Elite ABC; Vector Lab. CA). The data was analyzed using ANOVA. Results: The size of wound was significantly bigger in NOD than in BALB mice at 4 and 6days (p<0.05). The number of positive staining for Ki67 was significantly more at epithelial edges of the wound healing in BALB than in NOD mice (p<0.05). In NOD mice at 6days, the staining for K14 was limited at basal layer and the staining for K10 appeared at suprabasal layer of the epithelium. Conversely, In BALB mice, K14 immunostaining was observed throughout the entire epithelial edges, whereas no positive staining for K10 was observed at the epithelial edges. Conclusions: The results indicate that both keratinocyte proliferation and differentiation may be delayed during the wound healing in diabetics.
    IADR General Session 2010; 07/2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Oral epithelium plays an important role in protecting the oral cavity by virtue of its barrier function, and has an ability to produce antimicrobial and immunoregulatory peptides such as human β-defensins (hBDs). Butyrate, the dietary histone deacetylase inhibitor, is produced by periodontal pathogens such as porphyromonas gingivalis and fusobacterium nucleatum. Objectives: The present study investigated how butyrate affected the expression patterns of hBD-2, -3 in keratinocyte. Methods: HaCaT cells, a human keratinocyte cell line, were grown in DMEM supplemented with 10% Fetal bovine serum. The cells were incubated with butyrate (Sigma) at the concentration of 0, 0.5, 2.5, 5mM for 0.5, 6, 24 or 48 hours. The expression levels of the hBDs mRNAs were evaluated by RT-PCR followed by assessment of transcript induction, and the level of hBDs within supernatants were measured by sandwich ELISA. The ELISA assays were performed using capture and biotinylated antibodies for of hBD-2 and -3 (Peprotech). In order to elucidate the intracellular pathway involved in the expression of hBDs, cells were pretreated with inhibitors [AG1478 (EGFR), SB203580 (p38MAPK), or JNK II (JNK) inhibitor] for an hour in some experiments. The significance of differences was analyzed by Student's t-test (n=5). Results:Butyrate induced upregulated expression of hBD-2 and -3 at both mRNA and protein levels in dose- and time-dependent manners. The expression level of hBD-2 was the highest in the keratinocyte stimulated with 5mM butyrate for 48hr. The JNK II inhibitor abolished the upregulated expression of hBD-3 induced by the stimulation with butyrate. Conclusion: These results suggest that butyrate induces upregulated expression of hBD-2 and -3 in the keratinocyte.
    AADR Annual Meeting 2010; 03/2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Junctional epithelium, a nonkeratinized stratified epithelium, extends apically in apposition to the surface of the enamel to form a seal between the epithelium and the tooth. Desmosomes and gap junctions adhere to the junctional epithelium through cell-cell contact, but no evidence of tight junctions has been found. Recently, tight junction hallmark proteins and tight junction-related structures have been identified in stratified squamous epithelium. The present study examined whether tight junction proteins were expressed in the junctional epithelium. We used immunohistochemical techniques to observe expression of claudin-1, -4, -5, -7, and occludin in porcine gingival junctional epithelium. Claudin-4 exhibited immunoreactivity in the intercellular spaces of all layers of the oral epithelium and the junctional epithelium. Stronger expression was observed in junctional epithelial cells adjacent to the inner and outer basal laminae than in the inner cell layers. Immunohistochemical positivity for claudin-7 was clearly observed in the junctional epithelium, but only a faint positivity was observed in the basal layer of the oral epithelium. No immunohistochemical positivity for claudin-1, -5, or occludin was observed in the junctional epithelium. RT-PCR assay confirmed expression of porcine claudin-4 and -7 mRNAs in the junctional epithelium. These findings indicate that claudin-4 and -7 may play a role in the junctional epithelium even in the absence of tight junctions.
    Medical Molecular Morphology 12/2009; 42(4):212-5. · 1.17 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objective: Human beta-defensin(hBD)s belong to a group of antimicrobial peptides expressed in epithelial cells. Diabetes is a risk factor in oral infections including periodontitis. The hyperglycemia and insulin deficiency/insensitivity may affect hBDs expression. Therefore, the present study investigated the effect of glucose and insulin on the expression of hBD-1,-2 and -3. Methods: HSY cells, from a human salivary duct cell line, were grown in DMEM supplemented with 10% fetal bovine serum. To examine the effect of glucose and/or insulin on the expression of hBDs mRNAs and proteins, the cells were incubated with 5.5, 10, 15 and 50mM of glucose and/or with 100, 150 and 200nM of insulin for 6hr. The cells incubated without glucose or insulin were used as controls. Expression of hBD1~3 in HSY cells were observed by RT-PCR and quantitative RT-PCR using TaqMan probes. The data was analyzed using one-way ANOVA. To evaluate peptide expression level of hBDs, an ELISA assay was carried out. Cell free supernatant and cell lysate from each of cultures and un-stimulated control culture were analyzed for the level of hBD-1, hBD-2 and hBD-3 peptides, using an ELISA. We combined a sandwich ELISA, the best commercially available capture and detection antibodies (Peprotech, NJ). Fold induction represent means SD of triplicate experiments. P values were calculated by student's t test. Results: No significant differences are observed in the expression of hBDs between any concentrations of glucose. Expression levels of hBDs in 200nM of insulin were significantly higher than those in the controls. Combination of 5.5mM of glucose and 200nM of insulin stimulated the highest levels of the hBDs in each experiment. Conclusion: The results indicated that optimal concentration of glucose and insulin might be needed to maintain a high expression level of hBDs.
    IADR General Session 2009; 04/2009
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objectives: Toll-like receptors (TLR) play a fundamental role in the activation of the innate immune system during infection. Recent papers suggested that TLRs expression is involved in the development of autoimmune diseases. There is little information on the expression profiles of TLRs in the autoimmune sialoadenitis. Non-obese diabetic (NOD) mice spontaneously develop sialoadenitis, producing a condition that resembles Sjogren's syndrome. The present study analyzed the expression patterns of TLR-1,2,-3-4,-5,-6-7,-8 and -9 during the development of sialoadenitis in NOD mice. Method: Female NOD mice (ClEAJapan) were used in the present study. Submandibular glands were dissected from the mice at 4,8,10,12,14 and 16 weeks of age (n=5 in each group). Expression of and TLR-1,-2,-3-4,-5,-6-7,-8 and -9, and myeloid differentiation factor 88 (MyD88) was observed by RT-PCR and quantitative RT-PCR using TaqMan probes (Applied Biosystem, CA). Several experiments were performed, each in triplicate. The relative expression of each mRNA was calculated as the ΔCt using the formula described by Saitoh et al.(Med Mol Morphol 2007). The data was analyzed using one-way ANOVA. Differences between experimental groups were considered statistically significant at the p<0.05 levels. Immunohistochemical staining for TRL-7 and -9 was carried out using anti-TLR-3, -TLR7 and TLR-9 antibodies (Hycult Biotech.) Results: The submandibular glands show no indication of inflammatory reactions in 4- and 8-week-old mice. Ten-week-old mice generally had one focus of lymphatic infiltration per lobule. Expression levels of TLR-3, -7 and-9 at 14 and 16 weeks were significantly higher than at 4 or 8 weeks (p<0.01). Immunohistochemical positives for TLR-7 and -9 were observed in the foci of lymphatic infiltration. Conclusion: The results indicate that upregulated expression of TRL-7 and -9 may be involved in the development of autoimmune sialoadenitis.
    IADR General Session 2009; 04/2009
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objective: Human beta-defensin(hBD)s are a group of antimicrobial peptides. Oral epithelium is exposed to many kinds of external stresses including bacteria, food and drink. Bacterial infection induces upregulated expression of hBD-2 and -3. Food and drink causes osmotic stresses to oral epithelium. The present study investigated whether osmotic stress altered expression of hBDs in keratinocytes. Method: Keratinocyte derived from normal foreskin (NHK) were cultured in keratinocyte growth media (KGM, Lonza MD) and used as control. In order to add osmotic stresses to the cells, NHKs were treated with hypersomotic KGM with 50, 100, 150, or 200mM sorbitol for 1, 2 or 3days. Expression of and hBD-1, 2 and -3 was observed by RT-PCR and quantitative RT-PCR using TaqMan probes (Applied Biosystem, CA). The relative expression of each mRNA was calculated as the ΔCt using the formula described by Saitoh et al.(Med Mol Morphol 2007). The data was analyzed using one-way ANOVA. Differences between experimental groups were considered statistically significant at the p<0.05 levels. To evaluate peptide expression level of hBDs, an ELISA assay was carried out. Cell free supernatant and cell lysate from each of the cultures and un-stimulated control culture were analyzed for the level of hBD-1, hBD-2 and hBD-3 peptides, using an ELISA. We combined a sandwich ELISA, the best commercially available capture and detection antibodies (Peprotech, NJ). Fold induction represent means SD of triplicate experiments. P values were calculated by student's t test. Results: The expression level of hBD-1, -2 and -3 mRNAs in keratinocytes cultured in KGM with 150mM sorbitol is significantly higher than the control. The ELISA assay confirmed increased expression of hBDs at peptide levels. Conclusion: The results indicate that osmotic stresses by food and drink may induce upregulated expression of hBDs.
    IADR General Session 2009; 04/2009
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objective: Previously, we described the decreased expression of human beta-defensins (hBD), a family of antimicrobial peptides, in oral squamous cell carcinoma (SCC) cell lines (Abiko,Y. et al. Cancer Lett., 1999). hBD-1 has been identified as a tumor suppressor gene in renal and prostate cancers. In addition, decreased gene expression of hBD-1 in the development of oral SCC was demonstrated. In the present study, we investigated the decrease of hBD-1 transcription in oral SCC. Methods: Eight oral SCC-established cell lines (SAS, HSC-2,-3,-4, SCC-9, OSC-19, BSC-OF, HAC) and normal keratinocytes were used. The expression level of hBD-1 mRNA was estimated by quantitative RT-PCR. Total DNA was extracted and the promoter region of hBD-1 up to -900 was directly sequenced using an ABI PRISM 310 Genetic Analyzer (Applied Biosystems, CA). In order to examine whether hypermethylation is involved in the change in transcriptional levels, cells were treated with the DNA methyltransferase inhibitor, 5-aza-dCyd (Sigma), and the promoter activities were analyzed by luciferase reporter assay after transfection of the constructs into HaCaT cells. Several experiments were performed, each in triplicate. The data was analyzed using one-way ANOVA. Results: The RT-PCR assay revealed that the expression level of hBD-1 mRNA in SAS and OSC-19 cell lines was less than one-fifth the level observed in normal keratinocytes. Neither SAS nor OSC-19 showed changes in the expression level of hBD-1 mRNA after 5-aza-dCyd treatment. By direct sequencing, polymorphisms were revealed at -688(C/G) in SAS, and at -20(C/T) and -52(T/C) in OSC-19. The luciferase activity in SAS with -688(G) was one-fifth of that in the wild-type. The activity in OSC-19 with both -20(T) and -52(C) was one-third of that in the wild-type. Conclusion: These results indicate that genetic polymorphism sites at -20,-52,and -688 may be crucial for the decreased expression of hBD-1 mRNA in oral SCC.
    IADR General Session 2009; 04/2009
  • [Show abstract] [Hide abstract]
    ABSTRACT: Objectives: Although the bone levels around natural teeth that oppose an implant may be altered, little information has been reported. The aim of the present study was to evaluate the bone levels around natural teeth that oppose an implant using digitized conventional radiographs and subtraction images. Methods: Radiographs taken for 80 natural molars that oppose an implant were used. The radiographs were obtained at 6, 9 and 12 months after the implant crowns were fixed (HA coated implants of 4mm diameter, Spline, Zimmer Dental). Contralateral molars in the same jaw that did not oppose an implant were used as controls. The radiographs were digitized into a personal computer using a scanner. The images were manipulated by EMAGO/Advanced 5.x software (Oral Diagnostic system, Amsterdam, The Netherlands), and linear and logarithmic digital subtraction images were produced. The logarithmic subtraction was enhanced with the use of a filter. The bone levels around natural teeth that oppose an implant and around control teeth were assessed in the subtraction images. The subtraction image was analyzed using NIH image. The statistical difference between the bone levels was analyzed using F-test. Differences between the group of teeth that oppose implants and the control group were considered statistically significant at p<0.05 levels. Results: Significant bone loss or gain was often observed around the teeth that oppose implants at 6, 9 or 12 months after the implant crown was fixed. Little bone loss was observed around the control teeth. The variance of the data in the group of teeth that oppose an implant was significantly greater than that in the control group. Conclusions: Our results suggest that the bone levels around teeth that oppose implants may be altered when compared to bone levels around teeth that do not oppose an implant.
    IADR General Session 2009; 04/2009
  • [Show abstract] [Hide abstract]
    ABSTRACT: Two cases of basalold-squamous cell carcinoma (BSC) of the oral mucosa are described. The first case occurred at the floor of the mouth in a 58-year-old man, and the second case occurred at the mandibular gingiva in a 79-year-old woman. The laboratory data of the first case showed a positive response to hepatitis C virus antibody. in the first case, the tumor mass measured 4 times 4 cm in size, and was i-texl at the lingual side of the median mandible beside the sublingual gland. In the second case, the tumor mass measured 25 times 15 mm In size, and was located in the alveolar mucosa of the right mandible. Histologically, both tumors showed a neoplastic epithelium arranged in a solid pattern with evidence of peripheral palisading, central necrosis, and some squamous differentiation. The pro-ilferathfe activities of the BSC were compared with conventional squamous cell carcinomas (SCC) in the oral floor and gingiva, respectively, by employing a sensitive argy-rophillc nuclear organizer region (AgNOR) staining method. The number of AgNOR per nucleus of the BSC was higher than that of any other SCC cases. The results support the opinion that BSC of the oral mucosa has a worse prognosis than conventional SCC.
    Pathology International 12/2008; 48(6):460 - 466. · 1.72 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: It is unknown which genes are differentially expressed in cultured epithelial cells derived from the epithelial rests of Malassez (ERM) in periodontal ligament and oral gingival epithelium (OGE). This study analysed the different gene expression of OGE and ERM cells using a DNA microarray technique. Epithelial cells from ERM and OGE were isolated from porcine periodontal ligament and oral gingival epithelium. Each RNA sample extracted from the cells was reverse transcribed into cDNA and labelled with either cytidine 5-dUTP (Cy5) or cytidine 3-dUTP (Cy3). These labelled cDNA probes were then mixed and simultaneously hybridised to the Pig 13K microarray plate bearing 13,295 different genes (Operon, AL). Cellular enzyme-linked immunosorbent assay (CELISA) was performed to confirm the expression at protein level. There were nine genes common to the triplicate microarrays in ERM cells and one in OGE cells. Four of the nine genes including tissue factor (TF), FAT cadherin (FAT) and two unknown genes were expressed at levels more than threefold higher in ERM cells than in OGE cells. The protein levels of both TF and FAT in ERM cells were significantly higher than those in OGE. TF and FAT may act as markers to distinguish ERM cells from OGE cells in vitro.
    Archives of Oral Biology 06/2008; 53(5):437-42. · 1.55 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Oral squamous cell carcinoma and the most common oral pre-malignancies appear to be related to the habit of betel-quid chewing in Sri Lanka. Although hypermethylation of the tumour suppressor genes in oral cancer have been well documented, little information has been available concerning hypermethylation in oral pre-cancerous lesions. In the present study, we investigated the hypermethylation of p14, p15 and p16 in pre-cancerous lesions including epithelial dysplasia and submucous fibrosis. All samples were obtained from patients with a betel-quid chewing habit in Sri Lanka. Sixty-four patients were clinically diagnosed with leukoplakia, and histopathologically diagnosed with mild or severe dysplasia. Ten patients were diagnosed with submucous fibrosis without epithelial dysplasia. CpG island hypermethylation was assessed by a methylation-specific PCR method. Immunohistochemical staining was performed using anti-p53 antibodies. A high frequency of hypermethylation of p14, p15 and p16 was detected in the pre-cancerous lesions, although no hypermethylation was found in normal epithelium. The frequency of hypermethylation was higher than that of positive staining for p53 mutation except in the case of p16 in mild dysplasia. No significant correlation was observed between p53-positive reactions and hypermethylation in any lesions. The hypermethylation was highly detectable even in p53-negative lesions, suggesting that hypermethylation of p14, p15 and p16 occur regardless of whether the lesions have p53 mutations or not. The present study indicates that hypermethylation may be involved in the pathogenesis of oral pre-cancerous lesions associated with betel-quid chewing in Sri Lanka.
    Journal of Oral Pathology and Medicine 03/2008; 37(8):475-9. · 2.06 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The oral epithelium functions as a mechanical and protective barrier to resist bacterial infection. beta-Defensins are a group of antimicrobial peptides mainly produced by epithelial cells of many organs including skin, lung, kidney, pancreas, uterus, eye, and nasal and oral mucosa. This review focuses on beta-defensins (BDs) in oral epithelia and discusses their importance in oral epithelial health and disease. BDs exhibit antimicrobial activity against oral microbes including periodontitis-related bacteria, Candida, and papilloma virus. Alterative expression of BDs was observed in oral epithelial diseases, including oral inflammatory lesions with and without microbial infection and oral cancer. BDs may be useful in the treatment of oral infectious diseases, ulcerative lesions, and cancer. BDs play an important role in protection against oral microbes and may be used in clinical applications.
    Medical Molecular Morphology 01/2008; 40(4):179-84. · 1.17 Impact Factor

Publication Stats

884 Citations
162.92 Total Impact Points

Institutions

  • 1994–2010
    • Health Sciences University of Hokkaido
      • • School of Dentistry
      • • Department of Pediatric Dentistry
      • • Department of General Dental Sciences
      Tōbetsu, Hokkaido, Japan
  • 2003
    • Osaka Dental University
      Hirakata, Ōsaka, Japan
  • 2002
    • Kanagawa Dental University
      Йокосука, Kanagawa, Japan
  • 1984
    • Higashi Nippon International University
      Sapporo, Hokkaidō, Japan