High-Resolution Three-Dimensional Computed Tomography Analysis of the Clinical Efficacy of Cultured Autogenous Periosteal Cells in Sinus Lift Bone Grafting.

BACKGROUND AND PURPOSE: Sinus lift (SL) using cultured autogenous periosteal cells (CAPCs) combined with autogenous bone and platelet-rich plasma (PRP) was performed to evaluate the effect of cell administration on bone regeneration, by using high-resolution three-dimensional computed tomography (CT). MATERIALS AND METHODS: SL with autogenous bone and PRP plus CAPC [CAPC(+)SL] was performed in 23 patients. A piece of periosteum taken from the mandible was cultured in M199 medium with 10% fetal bovine serum (FBS) for 6 weeks. As control, 16 patients received SL with autogenous bone and PRP [CAPC(-)SL]. Three-dimensional CT imaging was performed before and 4 months and 1 year after SL, and stratification was performed based on CT numbers (HUs) corresponding to soft tissue and cancellous or cortical bone. RESULTS: The augmented bone in CAPC(+)SL revealed an increase in HUs corresponding to cancellous bone as well as a decrease in HUs corresponding to grafted cortical bone. In addition, HUs corresponding to cancellous bone in the graft bed were increased in CAPC(+)SL but were decreased in CAPC(-)SL. Insertion torque during implant placement was significantly higher in CAPC(+)SL. CONCLUSION: By promoting bone anabolic activity both in augmented bone and graft bed, CAPCs are expected to aid primary fixation and osseointegration of implants in clinical applications.

Diagnostic value of cyclin-dependent kinase/cyclin-dependent kinase inhibitor expression ratios as biomarkers of locoregional and hematogenous dissemination risks in oral squamous cell carcinoma.

The aim of the present study was to investigate the diagnostic value of cell cycle-related genes in oral squamous cell carcinoma (OSCC) by examining the expression of the following genes in 77 OSCC tissues by quantitative polymerase chain reaction: Cyclin genes (CCNA1, CCND1, CCND2 and CCNE1), cyclin-dependent kinase (CDK) genes (CDK1, CDK2 and CDK4), CDK inhibitor genes (CDKN2A, CDKN1A, CDKN1B and CDKN1C), and integrin and associated genes that we previously reported (ITGA3, ITGB4, CD9 and JUP). The expression ratios of 66 combinations of the 11 cell cycle-related genes were analyzed to examine their associations with major clinical events using Mann-Whitney U and log-rank tests. Three expression ratios (CDK1/CDKN1B, CDK2/CDKN1A and CCNE1/CDK2) showed associations on univariate analyses and their diagnostic value was re-analyzed with integrin gene expression biomarkers (ITGA3/CD9 and ITGB4/JUP) using the Cox proportional hazards model and Kaplan-Meier estimates. Lymph node metastasis occurred in >90% of double-positive cases (high-ITGA3/CD9 and high-CDK1/CDKN1B) irrespective of tumor size (P<0.0001). Primary site recurrence was found in >30% of double-positive cases (high-ITGA3/CD9 and high-CDK2/CDKN1A) with tumors >20 mm (P=0.003). Triple-positive (high-ITGB4/JUP, high-ITGA3/CD9 and high-CDK2/CDKN1A) was associated with distant metastasis (P<0.0001), but not with other clinical parameters. Disease-specific death occurred in 55% of double-positive cases (high-ITGA3/CD9 and high-CDK2/CDKN1A) (P<0.0001) and a positive surgical margin was a significant factor for fatality in these cases. Reliable prediction of locoregional and hematogenous dissemination risks in OSCC using the four CDK and integrin gene expression ratios is a promising biomarker system. Clinical use of these parameters may improve the control rate with the use of new therapeutic strategies.

Radiation-induced undifferentiated high-grade pleomorphic sarcoma (malignant fibrous histiocytoma) of the mandible: report of a case arising in the background of long-standing osteomyelitis with a review of the literature.

We report a rare case of radiation-induced undifferentiated high-grade pleomorphic sarcoma (UPS) (malignant fibrous histiocytoma, MFH) in the right mandible of a 44-year-old woman. The patient had suffered from osteomyelitis of the same region of the mandible for several years, which was considered to be due to radiotherapy for a malignant lymphoma in her right neck 19 years before. The tumor appeared as an exophytic and invasive growth in the molar region of the mandible. Histopathologically, the tumor consisted of an interlacing proliferation of vimentin-immunopositive spindle-shaped fibroblastic cells with bizarre nuclei with high Ki-67 labeling scores, and tumor cells showed storiform patterns mixed with pleomorphic cells. Taking the history of radiation into consideration, we diagnosed the lesion as radiation-induced MFH/UPS. Including the present case, there have been only 14 documented cases of radiation-induced UPS in the jawbone, and this is the first UPS case arising in the follow-up period of long-standing osteomyelitis.

Intraoperative assessment of surgical margins of oral squamous cell carcinoma using frozen sections: a practical clinicopathological management for recurrences.

BACKGROUND: Local recurrence remains a challenging clinical issue for the treatment of oral squamous cell carcinoma (SCC). We analyzed retrospectively how effective the frozen section technique (FS) was against recurrences of oral SCC. METHODS: We screened 343 surgical samples from 236 patients who had oral SCC, carcinoma in situ (CIS), or epithelial dysplasia, and we followed up their clinical outcomes for at least 5 years. Histopathological states of surgical margins were compared between FS and surgical materials in relapse and relapse-free groups, respectively. RESULTS: Among the 236 patients, 191 were classified into the relapse-free group, and 45 into the relapse group. FS was more frequently performed in the relapse-free group (128/191) than in the relapse group (83/152). Histopathologically, moderate dysplasia or CIS (borderline malignancies) and SCC were recognized in 55 samples of the relapse-free group and in 57 of the relapse group. For those surgical margins with borderline malignancies, additional incisions were performed in 38 of the 55 relapse-free cases, which reduced to 20 from the 38 margins with borderline malignancies (47.4% reduction), and in 39 of the 57 relapse cases, which reduced to only 3 of 39 (7.7% reduction). CONCLUSIONS: The intraoperative assessment of surgical margins by FS is essential in preventing recurrences of oral mucosal malignancies.

ITGA3 and ITGB4 expression biomarkers estimate the risks of locoregional and hematogenous dissemination of oral squamous cell carcinoma.

BACKGROUND: Molecular biomarkers are essential for monitoring treatment effects, predicting prognosis, and improving survival rate in oral squamous cell carcinoma. This study sought to verify the effectiveness of two integrin gene expression ratios as biomarkers. METHODS: Gene expression analyses of integrin α3 (ITGA3), integrin ß4 (ITGB4), CD9 antigen (CD9), and plakoglobin (JUP) by quantitative real-time PCR were conducted on total RNA from 270 OSCC cases. The logrank test, Cox proportional hazards model, and Kaplan-Meier estimates were performed on the gene expression ratios of ITGA3/CD9 and ITGB4/JUP and on the clinicopathological parameters for major clinical events. RESULTS: A high rate (around 80%) of lymph node metastasis was found in cases with a high ITGA3/CD9 ratio (high-ITGA3/CD9) and invasive histopathology (YK4). Primary site recurrence (PSR) was associated with high-ITGA3/CD9, T3-4 (TNM class), and positive margin, indicating that PSR is synergistically influenced by treatment failure and biological malignancy. A high ITGB4/JUP ratio (high-ITGB4/JUP) was revealed to be a primary contributor to distant metastasis without the involvement of clinicopathological factors, suggesting intervention of a critical step dependent on the function of the integrin ß4 subunit. Kaplan-Meier curves revealed positive margin as a lethal treatment consequence in high-ITGA3/CD9 and YK4 double-positive cases. CONCLUSION: Two types of metastatic trait were found in OSCC: locoregional dissemination, which was reflected by high-ITGA3/CD9, and distant metastasis through hematogenous dissemination, uniquely distinguished by high-ITGB4/JUP. The clinical significance of the integrin biomarkers implies that biological mechanisms such as cancer cell motility and anchorage-independent survival are vital for OSCC recurrence and metastasis.

Ghost cell odontogenic carcinoma arising in the background of a benign calcifying cystic odontogenic tumor of the mandible.

Ghost cell odontogenic carcinoma (GCOC) is a rare malignant variant of odontogenic tumor with ghost cells; only 29 cases are documented. Our patient was a 68-year-old man with a painless, well-defined, radiolucent swelling of the mandibular gingiva in the right incisor-to-molar region. It was diagnosed as a benign calcifying cystic odontogenic tumor (CCOT) on fenestration biopsy. Eighteen years later, he returned with swelling in the same area. The lesion was excised, diagnosed as GCOC, and considered a secondary malignant manifestation of the benign CCOT. No adjuvant chemotherapy or radiotherapy was administered, and his postoperative course was uneventful for 48 months, with no recurrence or distant metastasis. Among the 30 reported cases of GCOC, the mean age at diagnosis was 40.3 years, 22 (73%) involved the maxilla. Twelve (40%) were secondary malignant manifestations of benign CCOTs or dentinogenic ghost cell tumors. Five patients died of recurrence or distant metastasis.

A clinical study of alveolar bone tissue engineering with cultured autogenous periosteal cells: coordinated activation of bone formation and resorption.

In ongoing clinical research into the use of cultured autogenous periosteal cells (CAPCs) in alveolar bone regeneration, CAPCs were grafted into 33 sites (15 for alveolar ridge augmentation and 18 for maxillary sinus lift) in 25 cases. CAPCs were cultured for 6weeks, mixed with particulate autogenous bone and platelet-rich plasma, and then grafted into the sites. Clinical outcomes were determined from high-resolution three-dimensional computed tomography (3D-CT) images and histological findings. No serious adverse events were attributable to the use of grafted CAPCs. Bone regeneration was satisfactory even in cases of advanced atrophy of the alveolar process. Bone biopsy after bone grafting with CAPCs revealed prominent recruitment of osteoblasts and osteoclasts accompanied by angiogenesis around the regenerated bone. 3D-CT imaging suggested that remodeling of the grafted autogenous cortical bone particles was faster in bone grafting with CAPCs than in conventional bone grafting. The use of CAPCs offers cell-based bone regeneration therapy, affording complex bone regeneration across a wide area, and thus expanding the indications for dental implants. Also, it enables the content of particulate autogenous bone in the graft material to be reduced to as low as 40%, making the procedure less invasive, or enabling larger amounts of graft materials to be prepared. It may also be possible to dispense with the use of autogenous bone altogether in the future. The results suggest that CAPC grafting induces bone remodeling, thereby enhancing osseointegration and consequently reducing postoperative waiting time after dental implant placement.

Immediate implant loading following computer-guided surgery.

PURPOSE: The aim of this study was to develop and apply a new method for easy intraoperative adjustment of a provisional fixed full-arch restoration, in order to allow immediate implant loading following computer-guided surgery, regardless of any implant positioning errors compared to the virtual planning. METHODS: In accordance with the NobelGuide™ protocol, a provisional restoration for immediate loading of six maxillary implants was prepared prior to surgery. Because small shifts between the planned and the actual implant positions were to be expected, the provisional restoration was not fabricated directly on temporary cylinders as a conventional one-piece superstructure, but was divided into two portions: six custom made abutments and a long span fixed restoration which were left unconnected. After implantation, the custom abutments were attached to the six implants to be immediately loaded, and the superstructure was cemented simultaneously to all abutments using dual cure resin cement. After the excess cement was cleaned and polished, the superstructure was then reseated. Passive fit was achieved between implants and the superstructure. CONCLUSION: The superstructure described in this article can be easily seated and adjusted to accommodate any possible shifts in implant positioning occurring during computer-guided surgery. Through this method uneventful immediate implant loading can be achieved in a reasonable operative time.

Relationship between oral health status and development of osteoradionecrosis of the mandible: a retrospective longitudinal study.

OBJECTIVE: Oral health status is a risk factor for postradiation bone complications (also known as osteoradionecrosis [ORN]), and oral health care is an important element in the prevention of this condition. Some authors recommend extracting teeth with a questionable prognosis and either gross mobility or periodontal disease. However, the criteria for making such decisions remain to be elucidated. In addition, the specific details of the association between oral health status and ORN have not yet been clearly demonstrated. The purpose of this study was to clarify the relationship between oral health status and the development of ORN. STUDY DESIGN: Thirty-nine head and neck cancer patients whose radiation fields included both the teeth and the mandible were followed for > or = 3 years after radiotherapy. Among these patients, 6 suffered from ORN (ORN group), and the other 33 did not experience the condition (non-ORN group). We analyzed the patient factors, radiation factors, and oral health factors related to the complication. RESULTS: The onset of ORN occurred from 18 months to 51 months after radiotherapy. The radiation dose in the oral cavity and the oral health status before radiotherapy were not significant risk factors for ORN. However, the oral health status at 1 year or 2 years after radiotherapy was significantly associated with the development of ORN. The oral health conditions that increased the risk of ORN were > 5 mm periodontal pocket depth, > 40% dental plaque score, > 60% alveolar bone loss level, and a grade 3 radiographic periodontal status. CONCLUSION: This is the first report to show the changes in the oral health status before and after radiotherapy regarding the development of ORN. The results support the periodontal status that almost all clinicians agree on as indications for preradiation teeth extraction (namely, periodontal pockets of > 5 mm) to prevent ORN. Good oral health status, especially after radiotherapy, is very important in the prevention of ORN, and we recommend periodical dental management and care by well trained dentists and dental hygienists to avoid the condition.

The 2G allele of promoter region of matrix metalloproteinase-1 as an essential pre-condition for the early onset of oral squamous cell carcinoma.

BACKGROUND: Matrix metalloproteinase (MMP) is known to be involved in the initial and progressive stages of cancer development, and in the aggressive phenotypes of cancer. This study examines the association of single nucleotide polymorphisms in promoter regions of MMP-1 and MMP-3 with susceptibility to oral squamous cell carcinoma (OSCC). METHODS: We compared 170 Japanese OSCC cases and 164 healthy controls for genotypes of MMP-1 and MMP-3. RESULTS: The frequency of the MMP-1 2G allele was higher and that of the 1G homozygote was lower in the OSCC cases (p = 0.034). A multivariate logistic regression analysis revealed that subjects who were 45 years old or older had a significantly increased (2.47-fold) risk of OSCC (95%CI 1.47-4.14, p = 0.0006), and those carrying the MMP-1 2G allele had a 2.30-fold risk (95%CI 1.15-4.58, p = 0.018), indicating independent involvement of these factors in OSCC. One of the key discoveries of this research is the apparent reduction of the MMP-1 1G/1G and 1G/2G genotype distributions among the early onset OSCC cases under the ages of 45 years. It should be noted that the tongue was the primary site in 86.2% of these early onset cases. This could suggest the specific carcinogenic mechanisms, i.e. specific carcinogenic stimulations and/or genetic factors in the tongue. CONCLUSION: Since the 2G allele is a majority of the MMP-1 genotype in the general population, it seems to act as a genetic pre-condition in OSCC development. However this report suggests a crucial impact of the MMP-1 2G allele in the early onset OSCC.

Identification of potential biomarkers of lymph node metastasis in oral squamous cell carcinoma by cDNA microarray analysis.

We surveyed the expression of 557 cancer-related genes in 15 cases of well-differentiated OSCC by cDNA microarray analysis. To identify potential biomarkers for lymph node metastasis, all microarray data were compared by the Mann-Whitney test and the significance analysis of microarrays between OSCCs with and those without lymph node metastasis. The tissues of OSCCs with lymph node metastasis exhibited increased expression levels of MMP-1, MMP-3, uPA, integrin-alpha3, paxillin, tenascin C and IL-6 transcripts. All of these genes were included in common clusters on the Cluster/TreeView analysis, implying that functional gene groups of proteolytic enzymes and integrin-related molecules are involved in cervical lymph node metastasis. The results of RTQ-PCR for differentially expressed genes were in accord with those of cDNA microarray analyses, suggesting that the data obtained by microarray gene expression analyses were valid. Consistent with cooperative expression patterns, immunohistochemical analyses demonstrated that products of MMP-1, MMP-3 and uPA were colocalized to components of the neoplastic stroma, particularly mononuclear inflammatory cells with well-developed eosinophilic cytoplasm. Our results suggest that expression levels of molecules involved in tissue remodeling and cell-ECM adhesion, especially MMP-1 and integrin-alpha3, can provide an accurate biomarker system for predicting the risk of cervical lymph node metastasis in OSCC.

The clinical significance of follow-up sonography in the detection of cervical lymph node metastases in patients with stage I or II squamous cell carcinoma of the tongue.

OBJECTIVE: We sought to assess the reliability of repeated sonography in the detection of cervical lymph node metastases at the earliest stage during a follow-up period in patients with stage I or stage II carcinoma of the tongue. STUDY DESIGN: Eighteen consecutive patients with stage I or II squamous cell carcinoma of the tongue were included. When possible, every patient was examined with sonography approximately every 2 weeks during the follow-up period. RESULTS: With repeated sonography on 18 patients, 7 metastatic nodes of 7 patients (39%) meeting our criteria were found. With the use of computed tomography, we diagnosed 10 nodes (including the 7 nodes observed with sonography) in these 7 patients as metastatic. A histopathologic examination revealed that 12 nodes in the same 7 patients had metastatic foci. For sonography, the sensitivity per node was 58%, whereas that of computed tomography was 83%. CONCLUSIONS: Follow-up sonography enabled the detection of all patients who had subsequent subclinical lymph node metastases. However, the sensitivity of sonography in the detection of smaller metastatic nodes was lower than that of computed tomography.

The basement membrane-type heparan sulfate proteoglycan (perlecan) in ameloblastomas: its intercellular localization in stellate reticulum-like foci and biosynthesis by tumor cells in culture.

The localization and biosynthesis of basement membrane-type heparan sulfate proteoglycan (HSPG), known as perlecan, were studied in ameloblastomas using surgical tissue sections and cells in primary culture to demonstrate the existence of extracellular matrix (ECM) molecules in the intercellular space of epithelial tissue. HSPG was immunolocalized in the intercellular spaces of stellate reticulum-like cells and small vacuolar structures between basal cells in tumor cell nests as well as in myxofibrous stroma. By means of in-situ hybridization, mRNA signals for the HSPG core were intensely demonstrated in the cytoplasm of basal and parabasal cells of parenchyma. Furthermore, the in-vitro biosynthesis of HSPG core protein by ameloblastoma cells was confirmed using immunofluorescence, immunoprecipitation, and reverse-transcriptase polymerase chain reaction (RT-PCR). The results indicated that ameloblastoma cells synthesize HSPG and deposit it in their intercellular space. The intercellular HSPG might act as a carrier for transport of nutrients to tumor cells within ameloblastomatous foci.