Masamichi Ohishi

Immunohistochemical evaluation of PCNA and Ki-67 in ameloblastoma

Thirty-two ameloblastoma tissues were immunohistochemically studied using monoclonal anti-proliferating cell nuclear antigen (PCNA) and anti-Ki-67 antibodies. Positive cells were evaluated and analyzed in relation to the WHO classification, cytological pattern of the outer layer cell, clinical appearance, tumor location, radiographic appearance and patients age. In regard to the cytological pattern of the outer layer cells, the basal cell type had significantly higher PCNA and Ki-67 (P<0.05) labeling indices than the cuboidal cell type. The solid type had significantly higher PCNA and Ki-67 (P<0.05) labeling indices than the cystic and the mixed type. The labeling index of the younger patient was found to be the lowest, the middle age was in the middle and the older patient was the highest. These results indicated that the proliferating activities of ameloblastomas are quite variable, and the evaluations of Ki-67 and PCNA seem to be good indicators to assess the proliferating activity of each type of ameloblastomas.

Marsupialization of Cystic Ameloblastoma : A Clinical and Histopathologic Study of the Growth Characteristics Before and After Marsupialization

PURPOSE This study evaluated the effectiveness of marsupialization for the treatment of cystic ameloblastoma. MATERIALS AND METHODS Clinical and histologic examination of 24 cystic ameloblastomas were performed before and after marsupialization. RESULTS Clinically and radiographically, the effect of marsupialization was evaluated as extremely effective (five cases), effective (11 cases), and noneffective (eight cases). Marsupialization was most effective in teenaged patients. In three cases, the tumor disappeared macroscopically, and further surgery was not done. Histologically, the growth characteristics of the tumor were classified into expansive and invasive types. Cytologic pattern was classified into three types according to the outer cell layer of the tumor parenchyme: cuboidal cell, columnar cell, and basal cell. Premarsupialization, the expansive pattern with the cuboidal cell type was dominant, whereas for postmarsupialization many cases that had been expansive changed to an invasive type showing columnar or basal cell types. CONCLUSION Marsupialization was found to be useful as a preliminary treatment of the cystic ameloblastoma. However, it must be borne in mind that following marsupialization the tumor still has a potential to infiltrate into the surrounding tissues.

Characterization of Dental Epithelial Progenitor Cells Derived from Cervical-loop Epithelium in a Rat Lower Incisor

Dental epithelial progenitor cells differentiate into various cell types during development of tooth germs. To study this mechanism, we produced immortalized dental epithelial progenitor cells derived from the cervical-loop epithelium of a rat lower incisor. The expression patterns of cytokeratin 14, nerve growth factor receptor p75, amelogenin, Notch2, and alkaline phosphatase were examined by immnohistochemistry in both lower and higher cell densities. The patterns of each were compared in the dental epithelium of rat lower incisors. The results demonstrated that these cells could produce ameloblast lineage cells, stratum intermedium cells, stellate reticulum, and outer enamel epithelium. Furthermore, fibroblast growth factor 10 stimulated proliferation of dental progenitor cells and subsequently increased the number of cells expressing alkaline phosphatase. These results suggest that fibroblast growth factor 10 plays a role in coupling mitogenesis of the cervical-loop cells and the production of stratum intermedium cells in rat incisors.

Establishment of Dental Epithelial Cell Line (HAT-7) and the Cell Differentiation Dependent on Notch Signaling Pathway

Rat incisors grow continuously throughout life. Producing a variety of dental epithelial cells is performed by stem cells located in the cervical loop of the incisor apex. To study the mechanisms for cell differentiation, we established a dental epithelial cell line (HAT-7) originating from a cervical loop epithelium of a rat incisor. Immunochemical studies showed that HAT-7 produced the cells expressing amelogenin, ameloblastin, or alkaline phosphatase (ALP). To illustrate a role of Notch signaling in the determinant of the cell fate, we examined expression patterns of Notch1 and Jagged1 in HAT-7 density dependently. At lower cell density, Notch1- or Jagged1-expressing cells were not seen. However, when they were fully confluent, cells began to express Notch1 or Jagged1 strongly. Some ALP-positive cells were almost consistent with Notch1-expressing cells but not Jagged1-expressing cells. These results suggested that the determinant of direction of differentiation was associated with Notch signaling pathway.

Prevalence of Epstein–Barr virus in oral squamous cell carcinoma

Forty‐six samples of oral squamous cell carcinoma (OSCC) were evaluated for the prevalence of Epstein–Barr virus (EBV) infection by the polymerase chain reaction (PCR), Southern blot hybridization, and in situ hybridization (ISH). EBV DNA was detected in 7 (15·2 per cent) out of 46 samples by a combination of PCR and Southern blot hybridization methods. All seven positive samples showed well‐differentiated carcinoma, thus suggesting a possible relationship between EBV infection and the degree of differentiation of carcinoma tissue. Latent infection membrane protein 1 (LMP1) was detected immunohistochemically in six of the EBV‐positive OSCCs. However, no signal of the EBV‐encoded small RNA (EBER)‐1 was demonstrated by the ISH method. No significant relationship was observed between EBV infection and lymph node metastasis. A follow‐up study (range from 4·4 to 79 months; mean 34·9 months) showed no recurrence or death to occur in the EBV‐positive patients, which thus suggested a good prognosis for EBV‐positive OSCC patients. Copyright

Longitudinal Study on the Changes of Maxillary Arch Dimensions in Japanese Children with Cleft Lip and/or Palate: Infancy to 4 Years of Age

The focus of this study was an analysis of maxillary arch growth changes prior to the time of cheiloplasty up to 4 years of age. Serial dental casts were obtained and measured in 95 children with cleft lip and palate, or both: 7 unilateral cleft lip and alveolus (CLA), 52 unilateral cleft lip and palate (UCLP), 24 bilateral cleft lip and palate (BCLP), and 12 isolated cleft palate (CP). The children were treated at the Kyushu University Dental Hospital. The results are as follows: (1) Prior to cheiloplasty, the maxillary buccal segments in the subjects with cleft lip and palate showed lateral displacement. The premaxilla in BCLP subjects was protruded. (2) Cheiloplasty influenced maxillary anterior arch width, but not posterior width. The operation caused posterior displacement of the premaxilla in BCLP subjects. (3) Palatoplasty affected the growth of the maxillary arch in the transverse and anteroposterior dimensions. (4) A variety of growth patterns observed in the patients (e.g., increasing or decreasing of the maxillary arch dimensions) suggests that maxillary arch dimensions were affected not only by surgery, but also by other individual factors such as genetic facial pattern and severity of the cleft.

Platelet-derived growth factor and bone morphogenetic protein in the healing of mandibular fractures in rats.

We studied the effects of platelet-derived growth factor-B (PDGF-B) and bone morphogenetic protein-2 (BMP-2) during the healing of mandibular closed fractures in rats by immunohistochemical methods. Unilateral closed fractures were created in the mandibles of thirty 12-week-old rats. BMP-2 was expressed during all stages of healing, but PDGF-B was expressed mainly in the early and middle stages, and not in the later stage of the healing process. We conclude that PDGF-B was associated with the proliferation and migration of primitive mesenchymal cells. BMP-2 was related to the differentiation of mesenchymal cells into osteoblasts and chondroblasts. PDGF-B and BMP-2 both have distinct regulatory effects on the healing of fractures.

Two relatively distinct patterns of ameloblastoma: an anti‐apoptotic proliferating site in the outer layer (periphery) and a pro‐apoptotic differentiating site in the inner layer (centre)

Two relatively distinct patterns of ameloblastoma: an anti‐apoptotic proliferating site in the outer layer (periphery) and a pro‐apoptotic differentiating site in the inner layer (centre)

Age estimation in dental pulp DNA based on human telomere shortening

Age estimation based on evidence found in teeth has received considerable attention within the field of forensic science. We determined the terminal restriction fragment (TRF) length, as telomere length, to estimate age. Using dental pulp DNA we found the average TRF length showed a tendency to shortening with aging. Our findings show that telomere shortening, based on dental pulp DNA is a new and useful approach to estimate age of the subject at the time of death.

Endostatin inhibits lymph node metastasis by a down-regulation of the vascular endothelial growth factor C expression in tumor cells.

Anti-angiogenic therapy is a newly developed treatment method for malignant tumors. Endostatin has an anti-angiogenetic effect. Endostatin has also been shown to block the growth and metastasis of various cancers through the vascular system. However, there have so far been few reports on the relationship between endostatin and lymph node metastasis. In this study, we investigated the relationship between endostatin and the inhibition of lymph node metastasis. We first made recombinant adenovirus which expressed endostatin gene (Ad-end), and then performed the following experiments. Our findings showed Ad-end to inhibit the proliferation and tube formation of endothelial cells in vitro. In addition, Ad-end inhibited the growth of a human oral squamous cell carcinoma cell line (SQUU-B) implanted subcutaneously in the right flank of nude mice and orthotopically in the tongue of nude mice, and Ad-end also inhibited lymph node metastasis in orthotopic implantation. The number of CD31-positive blood vessels and 5’-nase-positive lymphatic vessels around Ad-end-infected tumors in tongue lesions was significantly lower than that in the control group. The down-regulation of vascular endothelial growth factor C (VEGF-C) in Ad-end-infected SQUU-B cells was recognized by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis. These findings suggested that endostatin inhibited lymphangiogenesis and lymph node metastasis by suppressing the production of VEGF-C in tumor cells.