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Objectives: We aimed to investigate bone metastasis induced by Notch signalling pathway dysregulation and to demonstrate that SPARC is a potential therapeutic target in adenoid cystic carcinoma (AdCC) with Notch dysregulation. Materials and Methods: This retrospective study enrolled 144 AdCC patients. RNA‐sequencing and enrichment analyses were performed using 32 AdCC samples. Osteonectin/SPARC and the Notch activation indicator Notch intracellular domain (NICD) were detected using immunohistochemistry. Cell proliferation and migration assays were conducted using stably NICD over‐expressing cells. The effect of SPARC on osteoclast differentiation in NICD cells was investigated using western blotting, quantitative reverse transcription PCR, tartrate‐resistant acid phosphatase staining and resorption assays. Results: RNA‐sequencing analysis showed that genes down‐regulated in Notch‐mutant AdCCs, such as SPARC, were enriched in ossification and osteoblast differentiation. Most (75/110, 68.2%) Notch1‐wild‐type AdCCs showed SPARC over‐expression, whereas 30 out of 34 (88.2%) Notch1‐mutant tumours showed low SPARC expression. SPARC over‐expression was then found negatively to be correlated with NICD expression in 144 AdCCs. NICD over‐expression promoted cell growth, migration and osteoclast differentiation, which could be partly reversed by exogenous SPARC. Conclusions: Notch activation in AdCC contributes to bone metastasis through SPARC inhibition. The study results suggest that SPARC may represent a prognostic biomarker and potential therapeutic target. [ABSTRACT FROM AUTHOR]

Background: Familial gigantiform cementoma (FGC) is a rare tumor characterized by the early onset of multi‐quadrant fibro‐osseous lesions in the jaws, causing severe maxillofacial deformities. Its clinicopathological features overlap with those of other benign fibro‐osseous lesions. FGC eventually exhibits progressively rapid growth, but no suspected causative gene has been identified. Methods: In this study, three patients with FGC were recruited, and genomic DNA from the tumor tissue and peripheral blood was extracted for whole‐exome sequencing. Results: Results showed that all three patients harbored the heterozygous mutation c.1067G > A (p.Cys356Tyr) in the ANO5 gene. Furthermore, autosomal dominant mutations in ANO5 at this locus have been identified in patients with gnathodiaphyseal dysplasia (GDD) and are considered a potential causative agent, suggesting a genetic association between FGC and GDD. In addition, multifocal fibrous bone lesions with similar clinical presentations were detected, including five cases of florid cemento‐osseous dysplasia, five cases of polyostotic fibrous dysplasia, and eight cases of juvenile ossifying fibromas; however, none of them harbored mutations in the ANO5 gene. Conclusion: Our findings indicate that FGC may be an atypical variant of GDD, providing evidence for the feasibility of ANO5 gene testing as an auxiliary diagnostic method for complex cases with multiple quadrants. [ABSTRACT FROM AUTHOR]

Podoplanin is upregulated in the invasive front of oral squamous cell carcinoma (OSCC). Carcinoma-associated fibroblasts (CAFs) may mediate podoplanin expression. However, the role of podoplanin in OSCC cell and fibroblast interaction remains elusive. In the present study, we found that positive podoplanin expression in OSCC cells correlated with smooth muscle actin (α-SMA) expression in CAFs. Using CAFs and normal mucosal fibroblasts (NFs), we established indirect and direct co-culture systems mimicking the structure of OSCC. Podoplanin-overexpressing OSCC cells promoted NF activation; in direct co-culture, but not in indirect co-culture, podoplanin-overexpressing OSCC cells increased fibroblast invasion via matrix metalloproteinase 2 (MMP-2), MMP-14, and αv/β6 integrin receptor (ITGA5/ITGB6) signaling. CAFs also induced podoplanin expression through the transforming growth factor-β1 (TGF-β1)/Smad pathway. TGF-β1 increased the podoplanin-dependent activation of epidermal growth factor receptor (EGFR), AKT, and extracellular signal–regulated kinase (ERK) signaling. Additionally, CAFs promoted OSCC cell invasion by upregulating MMP-2 and MMP-14 expression in both indirect and direct co-culture. Taken together, our findings indicate that podoplanin regulates the interaction between OSCC cells and CAFs via the mutual paracrine effects of TGF-β1. [ABSTRACT FROM AUTHOR]

Podoplanin overexpression has been reported in various cancers, however, the precise mechanism for podoplanin to promote tumor progression remains elusive. In the present study, podoplanin overexpression was found associated with invasiveness both in OSCC tissues and cell lines. Moreover, the cell invasiveness increased with forced podoplanin expression and decreased when podoplanin was knockdown, indicating podoplanin-mediated cell invasion during OSCC progression. To further identify the role of podoplanin in tumor invasion, cell spreading and immunofluorescence assay were performed firstly. It was found that podoplanin knockdown caused an impaired cell spreading with reduced filopodia and the premature assembly of stress fibers while podoplanin overexpression induced an increase in cellular protrusions and stress fibers with extensive parallel bundles. Then, pull-down assays revealed forced podoplanin expression increased Cdc42 activity and reduced RhoA activity while podoplanin knockdown decreased Cdc42 and increased RhoA markedly. Moreover, a hierarchy of crosstalk between RhoA and Cdc42 was confirmed in podoplanin-mediated cell motility. On the other hand, a significant correlation between podoplanin and MT1-MMP expression in OSCCs was found both in vivo and in vitro, co-located in invasive cells and cellular protrusions. Furthermore, our data showed MT1-MMP knockdown significantly blocked the upregulation of cell motility by forced podoplanin expression, indicating that MT1-MMP played a role in podoplanin-mediated tumor invasion. To further confirm the interaction between RhoA/Cdc42 complex, MT1-MMP and podoplanin, co-precipitation experiments were performed. Both the co-precipitation of podoplanin with MT1-MMP and the podoplanin-induced specific binding of MT1-MMP to Cdc42 were found, and immunofluorescence revealed the co-location of podoplanin, MT1-MMP and Cdc42 at the plasma membrane and filopodia induced an increase in cellular protrusion and stress fibers formation. Moreover, MT1-MMP inhibition could partly rescue the increase of Cdc42 activity caused by forced podoplanin expression. Taken together, our data demonstrated a hierarchy of crosstalk between RhoA and Cdc42 was involved in podoplanin-mediated cytoskeleton remodeling and invasion; the co-location and co-ordination of podoplanin, Cdc42 and MT1-MMP in the invadopodia might induce cytoskeleton remodeling, ECM degradation and tumor invasion, while podoplanin-induced EMT may not be indispensible during OSCC progression.

Central mucoepidermoid carcinoma (MEC) is a rare neoplasm arising intraosseously in the jaws. To clarify the clinicopathologic profile and pathogenesis of central MEC, clinicopathologic findings and follow-up data of 39 cases were collected and analyzed. There were 16 male and 23 female patients (median age, 43 y). Sixteen cases affected the maxilla, and 23 occurred in the mandible. Radiographically, most cases (32 of 39) showed a unilocular or multilocular radiolucency with bone destruction, and 7 were found with scattered calcification. The margins of the lesions were ill defined or diffused in 14 cases and relatively well defined in 25 cases. Most cases (26 of 39) were classified as low-grade MECs, whereas 13 were moderate-to-high grade. Follow-up data were available for 35 patients with a median period of 36 months. All cases were found to be primary; local recurrence occurred in 8 cases, most (75.0%) of which were low-grade tumors. Four cases showed regional lymph node metastasis, and 1 developed distant metastasis. Of 11 cases with a clinical history of the jaw cyst, 8 initially showed a typical odontogenic cyst with local MEC-like proliferation. In summary, the most likely pathogenesis of central MEC is neoplastic transformation of the epithelial lining of an odontogenic cyst, diagnosis of which should be based on clinical, radiographic, and histopathologic findings. The immunohistochemical profile of keratins is helpful in differential diagnosis. Radical surgery is the treatment of choice, whereas the role of radiotherapy or chemotherapy is still controversial, and careful long-term follow-up is necessary.

Objective: To investigate the clinicopathologic features, differential diagnosis, treatment and prognosis of oncocytoma and oncocytic carcinoma of the salivary glands., Methods: The clinicopathologic features were studied in 23 cases of oncocytomas and 15 cases of oncocytic carcinomas, and immunohistochemical staining as well as electron microscopy were performed., Results: Most oncocytomas occurred in the parotid gland (95.6%) with no recurrence. The diagnosis was made based on histopathological features. Oncocytic carcinomas were high-grade tumors, mainly occurring in the parotid gland. The diagnosis was based not only on histopathology but also on ultrastructural findings, phosphotungstic acid hematoxylin (PTAH), and immunohistochemistry. Of the 14 cases with follow-up information, 7 cases recurred. Regional or distant metastases were found in 7 and 4 cases, respectively., Conclusion: Oncocytoma is a rare tumor with well prognosis, whereas oncocytic carcinoma of the salivary glands is a high-grade tumor, with frequent local recurrence, regional or distant metastasis, the diagnosis of which should be based on a combination of clinical and histopathological features. Immunohistochemistry for mitochondria is considered as a practical and helpful adjuvant for diagnosis.

Objective: To investigate the expression of Pin1, beta-catenin and cyclin D1 in salivary adenoid cystic carcinomas (SACC) and to evaluate the role of beta-catenin and Pin1 in SACC carcinogenesis., Methods: The expressions of Pin1, beta-catenin and cyclin D1 were examined in the specimens of 65 patients with SACC by immunohistochemistry, and Pin1 protein and mRNA expressions detected by Western blot and RT-PCR in four SACC cell lines., Results: Pin1 was overexpressed in 51 cases of ACC (78%), and 41 (63%) cases showed positive immunoreactivity for cyclin D1 protein in the nuclear fraction in tumor tissues. Fourteen (22%) cases showed positive immunoreactivity for beta-catenin protein in the nuclear/cytoplasmic fraction in tumor tissues, 6 of which exhibited quite evident expression of beta-catenin in nucleolus. The expression of membranous beta-catenin was down-regulated in most of the patients with lymph node metastasis (11/14)., Conclusions: The results suggest that Pin1 and beta-catenin signalling pathway were activated in SACC and might play a pivotal role in SACC carcinogenesis and metastasis.

The aims of this study were to investigate the expression levels of beta-catenin, Pin1 and cyclin D1 in salivary adenoid cystic carcinomas (SACC ) and to evaluate its clinical importance, furthermore, to elucidate whether beta-catenin expression was aberrant in SACC and whether Pin1 was involved in aberrant beta-catenin and cyclin D1 expression. The expression of Pin1, beta-catenin and cyclin D1 were examined in the specimens of 65 patients with SACC by immunohistochemistry, protein and mRNA expressions were detected by western blotting and RT-PCR in four SACC cell lines. Pin1 was overexpressed in 51 cases of SACC (78%), and high levels of Pin1 expression correlated with cyclin D1 positive expression (p = 0.02). Fourteen (22%) cases showed positive immunoreactivity for beta-catenin protein in the nuclear/cytoplasmic fraction in tumor tissues, which was defined as cytoplasm/nucleus staining, among which quite evident nuclear expression of beta-catenin was detected in six cases (9%), while cyclin D1 positive expression was detected in 41 cases of SACC (63%). Reduced membranous expression of beta-catenin was detected in the cases with metastasis (11/14). Theses results suggest that Pin1 and Wnt signalling pathway are activated in SACC and may play a pivotal role in SACC carcinogenesis and metastasis.