The Role of Hedgehog Signaling in the Melanoma Tumor Bone Microenvironment.

A crucial regulator in melanoma progression and treatment resistance is tumor microenvironments, and Hedgehog (Hh) signals activated in a tumor bone microenvironment are a potential new therapeutic target. The mechanism of bone destruction by melanomas involving Hh/Gli signaling in such a tumor microenvironment is unknown. Here, we analyzed surgically resected oral malignant melanoma specimens and observed that Sonic Hedgehog, Gli1, and Gli2 were highly expressed in tumor cells, vasculatures, and osteoclasts. We established a tumor bone destruction mouse model by inoculating B16 cells into the bone marrow space of the right tibial metaphysis of 5-week-old female C57BL mice. An intraperitoneal administration of GANT61 (40 mg/kg), a small-molecule inhibitor of Gli1 and Gli2, resulted in significant inhibition of cortical bone destruction, TRAP-positive osteoclasts within the cortical bone, and endomucin-positive tumor vessels. The gene set enrichment analysis suggested that genes involved in apoptosis, angiogenesis, and the PD-L1 expression pathway in cancer were significantly altered by the GANT61 treatment. A flow cytometry analysis revealed that PD-L1 expression was significantly decreased in cells in which late apoptosis was induced by the GANT61 treatment. These results suggest that molecular targeting of Gli1 and Gli2 may release immunosuppression of the tumor bone microenvironment through normalization of abnormal angiogenesis and bone remodeling in advanced melanoma with jaw bone invasion.

Potential role of myeloid-derived suppressor cells in transition from reaction to repair phase of bone healing process.

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells with immunosuppressive functions; these cells play a key role in infection, immunization, chronic inflammation, and cancer. Recent studies have reported that immunosuppression plays an important role in the healing process of tissues and that Treg play an important role in fracture healing. MDSCs suppress active T cell proliferation and reduce the severity of arthritis in mice and humans. Together, these findings suggest that MDSCs play a role in bone biotransformation. In the present study, we examined the role of MDSCs in the bone healing process by creating a bone injury at the tibial epiphysis in mice. MDSCs were identified by CD11b and GR1 immunohistochemistry and their role in new bone formation was observed by detection of Runx2 and osteocalcin expression. Significant numbers of MDSCs were observed in transitional areas from the reactionary to repair stages. Interestingly, MDSCs exhibited Runx2 and osteocalcin expression in the transitional area but not in the reactionary area. And at the same area, cllagene-1 and ALP expression level increased in osteoblast progenitor cells. These data is suggesting that MDSCs emerge to suppress inflammation and support new bone formation. Here, we report, for the first time (to our knowledge), the role of MDSCs in the initiation of bone formation. MDSC appeared at the transition from inflammation to bone making and regulates bone healing by suppressing inflammation.

Comparing the Osteogenic Potential and Bone Regeneration Capacities of Dedifferentiated Fat Cells and Adipose-Derived Stem Cells In Vitro and In Vivo: Application of DFAT Cells Isolated by a Mesh Method.

BACKGROUND: We investigated and compared the osteogenic potential and bone regeneration capacities of dedifferentiated fat cells (DFAT cells) and adipose-derived stem cells (ASCs). METHOD: We isolated DFAT cells and ASCs from GFP mice. DFAT cells were established by a new culture method using a mesh culture instead of a ceiling culture. The isolated DFAT cells and ASCs were incubated in osteogenic medium, then alizarin red staining, alkaline phosphatase (ALP) assays, and RT-PCR (for RUNX2, osteopontin, DLX5, osterix, and osteocalcin) were performed to evaluate the osteoblastic differentiation ability of both cell types in vitro. In vivo, the DFAT cells and ASCs were incubated in osteogenic medium for four weeks and seeded on collagen composite scaffolds, then implanted subcutaneously into the backs of mice. We then performed hematoxylin and eosin staining and immunostaining for GFP and osteocalcin. RESULTS: The alizarin red-stained areas in DFAT cells showed weak calcification ability at two weeks, but high calcification ability at three weeks, similar to ASCs. The ALP levels of ASCs increased earlier than in DFAT cells and showed a significant difference (p < 0.05) at 6 and 9 days. The ALP levels of DFATs were higher than those of ASCs after 12 days. The expression levels of osteoblast marker genes (osterix and osteocalcin) of DFAT cells and ASCs were higher after osteogenic differentiation culture. CONCLUSION: DFAT cells are easily isolated from a small amount of adipose tissue and are readily expanded with high purity; thus, DFAT cells are applicable to many tissue-engineering strategies and cell-based therapies.

Effect of Patient Clinical Variables in Osteoporosis Classification Using Hip X-rays in Deep Learning Analysis.

Background and Objectives: A few deep learning studies have reported that combining image features with patient variables enhanced identification accuracy compared with image-only models. However, previous studies have not statistically reported the additional effect of patient variables on the image-only models. This study aimed to statistically evaluate the osteoporosis identification ability of deep learning by combining hip radiographs with patient variables. Materials andMethods: We collected a dataset containing 1699 images from patients who underwent skeletal-bone-mineral density measurements and hip radiography at a general hospital from 2014 to 2021. Osteoporosis was assessed from hip radiographs using convolutional neural network (CNN) models (ResNet18, 34, 50, 101, and 152). We also investigated ensemble models with patient clinical variables added to each CNN. Accuracy, precision, recall, specificity, F1 score, and area under the curve (AUC) were calculated as performance metrics. Furthermore, we statistically compared the accuracy of the image-only model with that of an ensemble model that included images plus patient factors, including effect size for each performance metric. Results: All metrics were improved in the ResNet34 ensemble model compared with the image-only model. The AUC score in the ensemble model was significantly improved compared with the image-only model (difference 0.004; 95% CI 0.002-0.0007; p = 0.0004, effect size: 0.871). Conclusions: This study revealed the additional effect of patient variables in identification of osteoporosis using deep CNNs with hip radiographs. Our results provided evidence that the patient variables had additive synergistic effects on the image in osteoporosis identification.

Maxillofacial Trauma Surgery Patients With Titanium Osteosynthesis Miniplates: Remove or Not?

The authors examined the timing and causes of titanium miniplate removal after maxillofacial trauma surgery. The authors performed a retrospective study of maxillofacial fracture patients in whom maxillofacial osteosynthesis miniplates were inserted or removed at the Kagawa Prefectural Central Hospital, between 2008 and 2017. Predictive variables were age, sex, fracture site distribution, and time to miniplate removal with or without complications in relation to primary outcome variables. Among 185 patients, 440 miniplates were inserted and 272 miniplates were removed. In total, 116 patients (73.4%) had 282 miniplates (64.1%) removed, of which 4.8% fracture sites and 5.7% miniplates were removed because of complications. The mean time to miniplate removal was 630.9 and 258.0 days in patients with and without complications, respectively. There was a statistically significant difference in miniplate removal and miniplate retention relative to age and sex. This difference was not related to the presence or absence of sex- or age-related complications. The miniplates as osteosynthesis material were safe and useful for a long period of time with relatively few complications. Because complications requiring miniplate removal occurred within 1 or after 5 years postoperatively, osteosynthesis miniplate treatments should be decided while considering the patient's age and sex. Long-term follow-up is recommended for miniplates that remain implanted for >1 year.

Impact of the Stroma on the Biological Characteristics of the Parenchyma in Oral Squamous Cell Carcinoma.

Solid tumors consist of the tumor parenchyma and stroma. The standard concept of oncology is that the tumor parenchyma regulates the tumor stroma and promotes tumor progression, and that the tumor parenchyma represents the tumor itself and defines the biological characteristics of the tumor tissue. Thus, the tumor stroma plays a pivotal role in assisting tumor parenchymal growth and invasiveness and is regarded as a supporter of the tumor parenchyma. The tumor parenchyma and stroma interact with each other. However, the influence of the stroma on the parenchyma is not clear. Therefore, in this study, we investigated the effect of the stroma on the parenchyma in oral squamous cell carcinoma (OSCC). We isolated tumor stroma from two types of OSCCs with different invasiveness (endophytic type OSCC (ED-st) and exophytic type OSCC (EX-st)) and examined the effect of the stroma on the parenchyma in terms of proliferation, invasion, and morphology by co-culturing and co-transplanting the OSCC cell line (HSC-2) with the two types of stroma. Both types of stroma were partially positive for alpha-smooth muscle actin. The tumor stroma increased the proliferation and invasion of tumor cells and altered the morphology of tumor cells in vitro and in vivo. ED-st exerted a greater effect on the tumor parenchyma in proliferation and invasion than EX-st. Morphological analysis showed that ED-st changed the morphology of HSC-2 cells to the invasive type of OSCC, and EX-st altered the morphology of HSC-2 cells to verrucous OSCC. This study suggests that the tumor stroma influences the biological characteristics of the parenchyma and that the origin of the stroma is strongly associated with the biological characteristics of the tumor.

Immunohistochemistry of YAP and dNp63 and survival analysis of patients bearing precancerous lesion and oral squamous cell carcinoma.

Background: Yes-associated protein (YAP) is a candidate oncogene in various human cancers, and recently, it has been reported that YAP expression and its activity was enhanced by ΔNp63. However, the role of YAP and ΔNp63 expression in carcinogenesis and progression of oral squamous cell carcinoma (OSCC) has been unknown. Therefore, we investigated how YAP and ΔNp63 influence carcinogenesis and progression of OSCC. Methods: We performed immunohistochemical analyses in whole tissue samples to investigate YAP and ΔNp63 expression in normal oral mucosa, epithelial hyperplasia, oral epithelial dysplasia (OED; low/high grade), carcinoma in situ (CIS), and OSCC. Furthermore, in OSCC, we analyzed clinical significance by using Kaplan-Meier survival analysis. Results: In normal oral mucosa and epithelial hyperplasia, YAP expression was primarily confined to the basal and parabasal layers, but YAP expression was elevated in OED, CIS, and OSCC. In OED, YAP and ΔNp63 expression levels were markedly higher in high grade than in low grade. In OSCC groups, YAP and ΔNp63 expression patterns tended to differ according to histopathological differentiation of OSCC. Furthermore, the YAP high expression group, which showed YAP staining in >50% positive cells with strong cytoplasmic staining or >10% positive cells with nuclear reactivity, showed a tendency to have a poor survival rate. Conclusion: YAP and ΔNp63 expression levels correlated with grade of oral OED. Additionally, YAP expression was associated with OSCC survival rate. Our results suggested that YAP and ΔNp63 expression might serve as predictive markers to distinguish OSCC development and progression.

Feasible Advantage of Bioactive/Bioresorbable Devices Made of Forged Composites of Hydroxyapatite Particles and Poly-L-lactide in Alveolar Bone Augmentation: A Preliminary Study.

Purpose: We aimed to document the clinical usefulness of uncalcined and unsintered hydroxyapatite (u-HA) particles and poly-L-lactide (PLLA) composite materials and their advantageous properties. Methods: Between April 2016 and March 2018, five patients required anterior maxillary alveolar ridge augmentation using fixation with u-HA/PLLA screws for an onlay block bone graft harvested from the mandibular ramus at our institute. Bone biopsies were obtained from the dental implantation site following bone healing for histomorphometric and immunohistochemical (IHC) measurements. Results: Many stromal cells were positive for Osterix, RUNX2, and SOX9 but were negative for CD68. On cell counting, based on IHC staining for Osterix, RUNX2, SOX9 and CD68 from peripheral u-HA/PLLA screw or bone areas, both areas consistently showed no significant difference in terms of Osterix, RUNX2, and SOX9. Hematoxylin-eosin staining revealed direct bone connection to the biomaterials, and no inflammatory cells infiltrated the areas surrounding the bone or artificial material. Area between the bone and u-HA/PLLA screw was seamless with no boundary. Round small cells and immature fibroblasts were noted. The new bone showed the presence of bone lamellae, normal osteocytes, and osteoblasts. Conclusion: The u-HA/PLLA materials showed excellent biodegradability and bioactive osteoconductivity. In addition, this material induced no apparent inflammatory or foreign body reactions following implantation, and it directly bonded to the human bone. Therefore, this u-HA/PLLA material seems ideal and most suitable for use as a substitute for osteosynthesis.

Notch Signaling Affects Oral Neoplasm Cell Differentiation and Acquisition of Tumor-Specific Characteristics.

Histopathological findings of oral neoplasm cell differentiation and metaplasia suggest that tumor cells induce their own dedifferentiation and re-differentiation and may lead to the formation of tumor-specific histological features. Notch signaling is involved in the maintenance of tissue stem cell nature and regulation of differentiation and is responsible for the cytological regulation of cell fate, morphogenesis, and/or development. In our previous study, immunohistochemistry was used to examine Notch expression using cases of odontogenic tumors and pleomorphic adenoma as oral neoplasms. According to our results, Notch signaling was specifically associated with tumor cell differentiation and metaplastic cells of developmental tissues. Notch signaling was involved in the differentiation of the ductal epithelial cells of salivary gland tumors and ameloblast-like cells of odontogenic tumors. However, Notch signaling was also involved in squamous metaplasia, irrespective of the type of developmental tissue. In odontogenic tumors, Notch signaling was involved in epithelial-mesenchymal interactions and may be related to tumor development and tumorigenesis. This signaling may also be associated with the malignant transformation of ameloblastomas. Overall, Notch signaling appears to play a major role in the formation of the characteristic cellular composition and histological features of oral neoplasms, and this involvement has been reviewed here.

The Role of Sonic Hedgehog Signaling in the Tumor Microenvironment of Oral Squamous Cell Carcinoma.

Sonic hedgehog (SHH) and its signaling have been identified in several human cancers, and increased levels of SHH expression appear to correlate with cancer progression. However, the role of SHH in the tumor microenvironment (TME) of oral squamous cell carcinoma (OSCC) is still unclear. No studies have compared the expression of SHH in different subtypes of OSCC and focused on the relationship between the tumor parenchyma and stroma. In this study, we analyzed SHH and expression of its receptor, Patched-1 (PTCH), in the TME of different subtypes of OSCC. Fifteen endophytic-type cases (ED type) and 15 exophytic-type cases (EX type) of OSCC were used. H&E staining, immunohistochemistry (IHC), double IHC, and double-fluorescent IHC were performed on these samples. ED-type parenchyma more strongly expressed both SHH and PTCH than EX-type parenchyma. In OSCC stroma, CD31-positive cancer blood vessels, CD68- and CD11b-positive macrophages, and α-smooth muscle actin-positive cancer-associated fibroblasts partially expressed PTCH. On the other hand, in EX-type stroma, almost no double-positive cells were observed. These results suggest that autocrine effects of SHH induce cancer invasion, and paracrine effects of SHH govern parenchyma-stromal interactions of OSCC. The role of the SHH pathway is to promote growth and invasion.

The Role of Bone Marrow-Derived Cells during Ectopic Bone Formation of Mouse Femoral Muscle in GFP Mouse Bone Marrow Transplantation Model.

Multipotential ability of bone marrow-derived cells has been clarified, and their involvement in repair and maintenance of various tissues has been reported. However, the role of bone marrow-derived cells in osteogenesis remains unknown. In the present study, bone marrow-derived cells during ectopic bone formation of mouse femoral muscle were traced using a GFP bone marrow transplantation model. Bone marrow cells from C57BL/6-Tg (CAG-EGFP) mice were transplanted into C57BL/6 J wild type mice. After transplantation, insoluble bone matrix (IBM) was implanted into mouse muscle. Ectopic bone formation was histologically assessed at postoperative days 7, 14, and 28. Immunohistochemistry for GFP single staining and GFP-osteocalcin double staining was then performed. Bone marrow transplantation successfully replaced hematopoietic cells with GFP-positive donor cells. Immunohistochemical analyses revealed that osteoblasts and osteocytes involved in ectopic bone formation were GFP-negative, whereas osteoclasts and hematopoietic cells involved in bone formation were GFP-positive. These results indicate that bone marrow-derived cells might not differentiate into osteoblasts. Thus, the main role of bone marrow-derived cells in ectopic osteogenesis may not be to induce bone regeneration by differentiation into osteoblasts, but rather to contribute to microenvironment formation for bone formation by differentiating tissue stem cells into osteoblasts.

Effects of the Geometrical Structure of a Honeycomb TCP on Relationship between Bone / Cartilage Formation and Angiogenesis.

A number of biomaterials have been developed, some of which already enjoy widespread clinic use. We have devised a new honeycomb tricalcium phosphate (TCP) containing through-and-through holes of various diameters to control cartilage and bone formation. However, the way in which the geometric structure of the honeycomb TCP controls cartilage and bone tissue formation separately remains unknown. In addition, an association has been reported between bone formation and angiogenesis. Therefore, in the present study, we investigated the relationship between angiogenesis and various hole diameters in our honeycomb TCP over time in a rat ectopic hard tissue formation model. Honeycomb TCPs with hole diameters of 75, 300, and 500 µm were implanted into rat femoral muscle. Next, ectopic hard tissue formation in the holes of the honeycomb TCP was assessed histologically at postoperative weeks 1, 2, and 3, and CD34 immunostaining was performed to evaluate angiogenesis. The results showed that cartilage formation accompanied by thin and poor blood vessel formation, bone marrow-like tissue with a branching network of vessels, and vigorous bone formation with thick linear blood vessels occurred in the TCPs with 75-µm, 300-µm, and 500-µm hole diameters, respectively. These results indicated that the geometrical structure of the honeycomb TCP affected cartilage and bone tissue formation separately owing to the induced angiogenesis and altered oxygen partial pressure within the holes.

Characterization and potential roles of bone marrow-derived stromal cells in cancer development and metastasis.

Background: The tumor microenvironment and its stromal cells play an important role in cancer development and metastasis. Bone marrow-derived cells (BMDCs), a rich source of hematopoietic and mesenchymal stem cells, putatively contribute to this tumoral stroma. However their characteristics and roles within the tumor microenvironment are unclear. In the present study, BMDCs in the tumor microenvironment were traced using the green fluorescent protein (GFP) bone marrow transplantation model. Methods: C57BL/6 mice were irradiated and rescued by bone marrow transplantation from GFP-transgenic mice. Lewis lung cancer cells were inoculated into the mice to generate subcutaneous allograft tumors or lung metastases. Confocal microscopy, immunohistochemistry for GFP, α-SMA, CD11b, CD31, CD34 and CD105, and double-fluorescent immunohistochemistry for GFP-CD11b, GFP-CD105 and GFP-CD31 were performed. Results: Round and dendritic-shaped GFP-positive mononuclear cells constituted a significant stromal subpopulation in primary tumor peripheral area (PA) and metastatic tumor area (MA) microenvironment, thus implicating an invasive and metastatic role for these cells. CD11b co-expression in GFP-positive cells suggests that round/dendritic cell subpopulations are possibly BM-derived macrophages. Identification of GFP-positive mononuclear infiltrates co-expressing CD31 suggests that these cells might be BM-derived angioblasts, whereas their non-reactivity for CD34, CD105 and α-SMA implies an altered vascular phenotype distinct from endothelial cells. Significant upregulation of GFP-positive, CD31-positive and GFP/CD31 double-positive cell densities positively correlated with PA and MA (P<0.05). Conclusion: Taken together, in vivo evidence of traceable GFP-positive BMDCs in primary and metastatic tumor microenvironment suggests that recruited BMDCs might partake in cancer invasion and metastasis, possess multilineage potency and promote angiogenesis.

Parenchyma-stromal interactions induce fibrosis by secreting CCN2 and promote osteoclastogenesis by stimulating RANKL and CD68 through activated TGF-ß/BMP4 in ameloblastoma.

BACKGROUND: Tumor parenchyma-stromal interactions affect the properties of tumors and their dynamics. Our group previously showed that secreted frizzled related protein (sFRP)-2 impairs bone formation and promotes bone invasion in ameloblastoma. However, the effects of the secreted growth factors CCN2, TGF-ß, and BMP4 on stromal tissues in ameloblastoma remain unclear. MATERIALS AND RESULTS: Thirty-five paraffin-embedded ameloblastoma cases, ameloblastoma-derived cell lines (AM-1), and primary cultures of ameloblastoma stromal fibroblasts (ASF) were used. Immunohistochemistry, MTT assay, Western blotting, and RT-PCR were performed on these samples. Parenchyma-stromal CCN2 overexpression correlated significantly with fibrous-type stroma, but not with myxoid-type stroma, suggesting a role of CCN2 in fibrosis (P < 0.05). Recombinant CCN2 induction of enhanced ASF proliferation in AM-1 medium supports this view. Conversely, BMP4 and TGF-ß were expressed in myxoid-type fibroblasts, but little expression was found in parenchyma. RANKL-positive and CD68-positive stromal cell populations were significantly greater in myxoid-type tumor areas than in fibrous-type tumor areas, while a higher Ki-67 labeling index was recorded in ameloblastoma with fibrous-type stroma. These data suggest that stromal properties influence bone resorption-related activities and growth rates, respectively. CONCLUSIONS: These results suggest that the effects of secreted growth factors are governed by ameloblastoma parenchyma-stromal interactions. CCN2 promotes fibrogenesis independent of TGF-ß signaling. Absence of CCN2 expression is associated with a phenotypic switch to a myxoid-type microenvironment that is conducive for TGF-ß/BMP4 signaling to promote osteoclastogenesis.

Efficacy of Honeycomb TCP-induced Microenvironment on Bone Tissue Regeneration in Craniofacial Area.

Artificial bone materials that exhibit high biocompatibility have been developed and are being widely used for bone tissue regeneration. However, there are no biomaterials that are minimally invasive and safe. In a previous study, we succeeded in developing honeycomb ß-tricalcium phosphate (ß-TCP) which has through-and-through holes and is able to mimic the bone microenvironment for bone tissue regeneration. In the present study, we investigated how the difference in hole-diameter of honeycomb ß-TCP (hole-diameter: 75, 300, 500, and 1600 µm) influences bone tissue regeneration histologically. Its osteoconductivity was also evaluated by implantation into zygomatic bone defects in rats. The results showed that the maximum bone formation was observed on the ß-TCP with hole-diameter 300µm, included bone marrow-like tissue and the pattern of bone tissue formation similar to host bone. Therefore, the results indicated that we could control bone tissue formation by creating a bone microenvironment provided by ß-TCP. Also, in zygomatic bone defect model with honeycomb ß-TCP, the result showed there was osseous union and the continuity was reproduced between the both edges of resected bone and ß-TCP, which indicated the zygomatic bone reproduction fully succeeded. It is thus thought that honeycomb ß-TCP may serve as an excellent biomaterial for bone tissue regeneration in the head, neck and face regions, expected in clinical applications.

Efficacy of Cisplatin-CXCR4 Antagonist Combination Therapy in Oral Cancer.

Cisplatin is a platinum-based compound that is widely used for treating inoperable oral squamous cell carcinoma (OSCC) in Japan; however, resistance to cisplatin presents a challenge and innovative approaches are required. We aimed to investigate the therapeutic potential of targeting the chemokine receptor CXCR4, which is involved in angiogenesis and tumor progression, using the CXCR4 inhibitor AMD3100, in combination with cisplatin. AMD3100 induced necrosis and bleeding in OSCC xenografts by inhibiting angiogenesis. We investigated the combined ability of AMD3100 plus cisplatin to enhance the antitumor effect in cisplatin-resistant OSCC. An MTS assay identified HSC-2 cells as cisplatin-resistant cells in vitro. Mice treated with the cisplatin-AMD combination exhibited the most significant reduction in tumor volume, accompanied by extensive hemorrhage and necrosis. Histological examination indicated thin and short tumor vessels in the AMD and cisplatin-AMD groups. These results indicated that cisplatin and AMD3100 had synergistic antitumor effects, highlighting their potential for vascular therapy of refractory OSCC. Antitumor vascular therapy using cisplatin combined with a CXCR4 inhibitor provides a novel strategy for addressing cisplatin-resistant OSCC.

Central dentinogenic ghost cell tumor of the maxilla: a case report with new imaging findings and review of the literature.

A dentinogenic ghost cell tumor (DGCT) is a rare benign odontogenic tumor that commonly shows characteristics of solid proliferation and has a relatively high risk of recurrence after surgical treatment. We herein report a case of a central DGCT that occurred in the maxilla and resulted in bone expansion. This study highlights new imaging findings (particularly magnetic resonance imaging) along with histopathological observations. In addition, we conducted a review of the existing literature on this rare tumor. A 37-year-old man developed swelling around the right cheek. A benign odontogenic tumor such as ameloblastoma was suspected based on the imaging examination findings (including bone expansion and the internal characteristics of the tumor) on panoramic imaging, computed tomography, and magnetic resonance imaging. The lesion was surgically excised from the right maxilla. Postoperative histopathological examination led to a definitive diagnosis of central DGCT. The tumor comprised epithelial neoplastic islands, resembling ameloblastoma, inside tight fibroconnective tissue; masses of ghost cells and formation of dentin were also observed. We had suspected that the minute high-density region around the molars on the imaging examinations represented alveolar bone change; however, it represented dentin formation. This led to difficulty diagnosing the lesion. Although DGCT may present characteristic findings on imaging examinations, its occurrence is infrequent, and in some cases, the findings may include the presence or absence of an impacted tooth without obvious calcification. The present case suggests that we should consider the possibility of an odontogenic tumor with calcification when high-density structures are observed inside the lesion.

Double-faced CX3CL1 enhances lymphangiogenesis-dependent metastasis in an aggressive subclone of oral squamous cell carcinoma.

Because cancer cells have a genetically unstable nature, they give rise to genetically different variant subclones inside a single tumor. Understanding cancer heterogeneity and subclone characteristics is crucial for developing more efficacious therapies. Oral squamous cell carcinoma (OSCC) is characterized by high heterogeneity and plasticity. On the other hand, CX3C motif ligand 1 (CX3CL1) is a double-faced chemokine with anti- and pro-tumor functions. Our study reported that CX3CL1 functioned differently in tumors with different cancer phenotypes, both in vivo and in vitro. Mouse OSCC 1 (MOC1) and MOC2 cells responded similarly to CX3CL1 in vitro. However, in vivo, CX3CL1 increased keratinization in indolent MOC1 cancer, while CX3CL1 promoted cervical lymphatic metastasis in aggressive MOC2 cancer. These outcomes were due to double-faced CX3CL1 effects on different immune microenvironments indolent and aggressive cancer created. Furthermore, we established that CX3CL1 promoted cancer metastasis via the lymphatic pathway by stimulating lymphangiogenesis and transendothelial migration of lymph-circulating tumor cells. CX3CL1 enrichment in lymphatic metastasis tissues was observed in aggressive murine and human cell lines. OSCC patient samples with CX3CL1 enrichment exhibited a strong correlation with lower overall survival rates and higher recurrence and distant metastasis rates. In conclusion, CX3CL1 is a pivotal factor that stimulates the metastasis of aggressive cancer subclones within the heterogeneous tumors to metastasize, and our study demonstrates the prognostic value of CX3CL1 enrichment in long-term monitoring in OSCC.

Novel Artificial Scaffold for Bone Marrow Regeneration: Honeycomb Tricalcium Phosphate.

Bone marrow is complex structure containing heterogenetic cells, making it difficult to regenerate using artificial scaffolds. In a previous study, we succeeded in developing honeycomb tricalcium phosphate (TCP), which is a cylindrical scaffold with a honeycomb arrangement of straight pores, and we demonstrated that TCP with 300 and 500 µm pore diameters (300TCP and 500TCP) induced bone marrow structure within the pores. In this study, we examined the optimal scaffold structure for bone marrow with homeostatic bone metabolism using honeycomb TCP. 300TCP and 500TCP were transplanted into rat muscle, and bone marrow formation was histologically assessed. Immunohistochemistry for CD45, CD34, Runt-related transcription factor 2 (Runx2), c-kit single staining, Runx2/N-cadherin, and c-kit/Tie-2 double staining was performed. The area of bone marrow structure, which includes CD45(+) round-shaped hematopoietic cells and CD34(+) sinusoidal vessels, was larger in 300TCP than in 500TCP. Additionally, Runx2(+) osteoblasts and c-kit(+) hematopoietic stem cells were observed on the surface of bone tissue formed within TCP. Among Runx2(+) osteoblasts, spindle-shaped N-cadherin(+) cells existed in association with c-kit(+)Tie-2(+) hematopoietic stem cells on the bone tissue formed within TCP, which formed a hematopoietic stem cell niche similar to as in vivo. Therefore, honeycomb TCP with 300 µm pore diameters may be an artificial scaffold with an optimal geometric structure as a scaffold for bone marrow formation.

Polymeric Materials, Advances and Applications in Tissue Engineering: A Review.

Tissue Engineering (TE) is an interdisciplinary field that encompasses materials science in combination with biological and engineering sciences. In recent years, an increase in the demand for therapeutic strategies for improving quality of life has necessitated innovative approaches to designing intelligent biomaterials aimed at the regeneration of tissues and organs. Polymeric porous scaffolds play a critical role in TE strategies for providing a favorable environment for tissue restoration and establishing the interaction of the biomaterial with cells and inducing substances. This article reviewed the various polymeric scaffold materials and their production techniques, as well as the basic elements and principles of TE. Several interesting strategies in eight main TE application areas of epithelial, bone, uterine, vascular, nerve, cartilaginous, cardiac, and urinary tissue were included with the aim of learning about current approaches in TE. Different polymer-based medical devices approved for use in clinical trials and a wide variety of polymeric biomaterials are currently available as commercial products. However, there still are obstacles that limit the clinical translation of TE implants for use wide in humans, and much research work is still needed in the field of regenerative medicine.