Squamous cell carcinoma of mandibular gingiva producing both parathyroid hormone-related protein and granulocyte colony-stimulating factor: a case report.

We describe a case of mandibular gingival carcinoma with hypercalcaemia and leukocytosis caused by tumour-derived parathyroid hormone-related protein (PTHrP) and granulocyte colony-stimulating factor (G-CSF). A 54-year-old man presented to our Department of Oral and Maxillofacial Surgery with a chief complaint of a left-sided mandibular gingival ulcer. A 42 mm × 20 mm sized ulcer was found on the left lower molar gingiva. Squamous cell carcinoma was pathologically diagnosed. The patient underwent a hemimandibulectomy, left-sided radical neck dissection, plate reconstruction, pectoralis major musculocutaneous flap reconstruction, and tracheostomy under general anaesthesia. Pathologically, two metastatic lymph nodes were identified. Residual tumour was suspected at the resection margins. Eight weeks after surgery, the patient started postoperative concurrent chemoradiotherapy (CCRT). Two weeks after CCRT, the patient developed hypercalcaemia. Serum levels of PTHrP and G-CSF increased in parallel with the progression of hypercalcaemia and leukocytosis. Immunohistochemical analysis of the surgical specimen showed positivity for G-CSF. Based on these clinical and pathological findings, the patient was diagnosed with hypercalcaemia and leukocytosis associated with malignancy and was treated with denosumab. Irradiation was terminated at 50 Gy because CT showed rapid disease progression. Chemotherapy was initiated, however, four weeks after the start of chemotherapy, a CT scan showed increased metastases and pleural dissemination. Therefore, chemotherapy was discontinued. One week after the chemotherapy was discontinued, the patient died of respiratory failure.

Application of mesenchymal stromal cell sheets to prevent medication-related osteonecrosis of the jaw with titanium implants in rats.

Medication-related osteonecrosis of the jaw (MRONJ) is an intractable adverse event. Dental implants are one of the triggering factors of MRONJ, and implant therapy with low MRONJ risk is required. This study aimed to investigate a rat model of MRONJ induced by extraoral placement of titanium materials and the use of mesenchymal stromal cell (MSCs) sheets to prevent MRONJ. Eight-week-old male rats were administered zoledronate and dexamethasone thrice weekly until killing. A week after drug initiation, a titanium screw and a plate were placed on the left buccal side of the mandible. Allogeneic bone marrow-derived MSC sheets were co-grafted with the titanium plates in the MSC sheet ( +) group. Six weeks after titanium placement, the rats were killed, and their excised mandibular bones were subjected to micro-computed tomography (CT) analysis. Histological analysis was performed after the titanium implants were removed. Empty lacunae visualized on hematoxylin and eosin staining were used as evidence of bone necrosis. Bone necrosis was reduced in the MSC sheet ( +) group. Tartrate-resistant acid phosphatase (TRAP) staining revealed a decreased number of TRAP-positive cells in areas with a large number of empty lacunae in the MSC sheet (-) group. Micro-CT analyses demonstrated that the bone volume fraction (BV/TV) was not significantly different between the MSC sheet (-) and ( +) groups. We conclude that MRONJ can be triggered by a titanium placement in rats, and grafting of allogeneic MSC sheets has the potential to prevent MRONJ.

Novel Cell Therapy Using Mesenchymal Stromal Cell Sheets for Medication-Related Osteonecrosis of the Jaw.

Despite medication-related osteonecrosis of the jaw (MRONJ) being first reported in 2003, the optimal treatment and prevention modalities for MRONJ are not clear. As a result, dentistry, oral surgery, and departments involved in the treatment of cancer and bone diseases are struggling with the management of MRONJ. Several cases of MRONJ cannot be managed by conventional treatment strategies recommended in various position papers. Therefore, studies have been conducted to investigate the efficacy of novel therapies for MRONJ. However, the optimal treatment is unknown. Several cell therapies including autologous cell transplantation have been reported for MRONJ. Although the efficacy of cell therapy for MRONJ has been demonstrated, large, statistically accurate clinical trials are lacking. We have been investigating the efficacy of MRONJ treatment using mesenchymal stromal cell (MSC) sheets since 2013 and confirmed its efficacy through various experiments, wherein MSC sheets were transplanted in model rats and beagle dogs with MRONJ-like lesions. Based on these results, we are planning to conduct a clinical trial of MRONJ therapy using periodontal ligament-derived MSC sheets.

The progress of medication-related osteonecrosis of the jaw with conservative initial treatment: A 12-year retrospective study of 129 patients.

This retrospective study aimed to examine the course and prognosis of medication-related osteonecrosis of the jaw (MRONJ) initially treated conservatively and the effects of various factors affecting treatment outcomes. We evaluated 129 patients with MRONJ between January 2008 and December 2018 at a university hospital. The factors examined included sex, age, stage of MRONJ (1-3), type of bone modifying agents (bisphosphonate or denosumab), primary disease (osteoporosis or malignant tumor), medical history (diabetes and rheumatoid arthritis), use of corticosteroids, the trigger of MRONJ (teeth extraction or others), and separation of sequestrum, using logistic regression analysis. Patients with MRONJ were treated conservatively as the initial treatment in accordance with the position paper of the American Association of Oral and Maxillofacial Surgeons. Of the 129 patients, 59 (45.7%) were cured, and the condition of 70 (54.3%) remained unchanged or worsened. The overall cure rates at 12, 36, and 60 months were 25.8%, 50.8%, and 72.4% respectively. The cure rate of stage 1 was lower than that of stages 2 and 3 at 80 months. In multivariate analysis, it was found that 37 (64.9%) of 57 patients with osteoporosis as a primary disease were cured (odds ratio [OR], 7.7; 95% confidence interval [CI], 2.4-24.4). In addition, 40 (69.0%) of 58 patients with separation of sequestrum were cured (OR, 8.9; 95% CI, 3.4-23.5). The cure rate was significantly higher in patients with osteoporosis than in those with cancer when the treatment outcomes of primary disease were compared using the Kaplan-Meier method (p < 0.01). It was also significantly higher in patients who had separation of sequestrum than in those who did not (p < 0.05). Our results suggest that primary disease and separation of sequestrum were associated with favorable outcomes in patients with MRONJ initially treated conservatively. MRONJ had a poor prognosis with conventional treatment carried according to the stage of the disease. This was especially prominent when conservative treatment was employed for mild cases.

Allogeneic multipotent mesenchymal stromal cell sheet transplantation promotes healthy healing of wounds caused by zoledronate and dexamethasone in canine mandibular bones.

INTRODUCTION: Many cases of bisphosphonate-related osteonecrosis of the jaw (BRONJ), which is an intractable disease, have been reported. Although a general intravenous injection of multipotent mesenchymal stromal cells (MSCs) may be effective for treating BRONJ, it has some severe problems. Therefore, our aim was to develop a treatment of locally administered MSCs. In this study, we investigated the effect of MSC sheet transplantation in the mandibular bone healing in beagle dogs, which were administered zoledronate and dexamethasone. METHODS: MSCs isolated from subcutaneous fat were seeded onto temperature-responsive culture dishes to produce MSC sheets. Zoledronate and dexamethasone were administered to beagle dogs. Then, the parts of mandibular cortical bones were removed, and MSC sheets were transplanted to cover those bone defects (MSC sheet transplant side) or not (Control side). The specimens were evaluated in micro CT, histology, and immunohistochemistry. RESULTS: Four weeks after surgery, redness and swellings were observed in the mucosal wounds of the control sides of 2 of 3 dogs. In contrast, the mucosal wounds of the MSC sheet transplant sides of all dogs completely healed. Histological images showed some free sequestrums and many bacterial colonies, and Immunohistological analysis showed some cathepsin K-positive multinuclear cells detached from jaw bone surfaces in the control sides. CONCLUSIONS: MSC sheet transplantation promotes healthy healing of wounds caused by zoledronate and dexamethasone in canine mandibular bones. And the injured canine mandibular bones administered zoledronate and dexamethasone showed BRONJ-like findings.

Factors Affecting the Occurrence of Complications in the Early Stages After Dental Implant Placement: A Retrospective Cohort Study.

OBJECTIVE: To evaluate the background factors related to the occurrence of complications in the early stages after dental implant placement. MATERIALS AND METHODS: A total of 289 outpatients who received dental implants were retrospectively evaluated for the presence or absence of complications. Background factors, including age, sex, implant width, implant length, implant site, number of implants placed, Periotest values at the time of implant placement, presence/absence of systemic disease (particularly diabetes), and the use of anticoagulation therapy, were compared between patients with and without complications. Logistic regression analysis was performed to identify significant risk factors for the occurrence of complications after dental implant placement. RESULTS: Complications in the early stages after dental implant placement occurred in 25 (8.65%) patients. The patients with complications were older than those without complications (P = 0.003). In addition, the incidence of complications was significantly higher in patients with systemic diseases (P = 0.004) and in those receiving anticoagulation therapy (P = 0.005). Logistic regression analysis revealed that age was a significant risk factor (P = 0.025) for early-stage complications, whereas the number of implants, presence of diabetes, and the use of anticoagulation therapy were not significant risk factors. CONCLUSIONS: Our results show that age is a significant factor influencing the occurrence of complications in the early stages after dental implant placement. Therefore, clinicians should consider this factor when developing their treatment strategies.

Creation and Transplantation of an Adipose-derived Stem Cell (ASC) Sheet in a Diabetic Wound-healing Model.

Artificial skin has achieved considerable therapeutic results in clinical practice. However, artificial skin treatments for wounds in diabetic patients with impeded blood flow or with large wounds might be prolonged. Cell-based therapies have appeared as a new technique for the treatment of diabetic ulcers, and cell-sheet engineering has improved the efficacy of cell transplantation. A number of reports have suggested that adipose-derived stem cells (ASCs), a type of mesenchymal stromal cell (MSC), exhibit therapeutic potential due to their relative abundance in adipose tissue and their accessibility for collection when compared to MSCs from other tissues. Therefore, ASCs appear to be a good source of stem cells for therapeutic use. In this study, ASC sheets from the epididymal adipose fat of normal Lewis rats were successfully created using temperature-responsive culture dishes and normal culture medium containing ascorbic acid. The ASC sheets were transplanted into Zucker diabetic fatty (ZDF) rats, a rat model of type 2 diabetes and obesity, that exhibit diminished wound healing. A wound was created on the posterior cranial surface, ASC sheets were transplanted into the wound, and a bilayer artificial skin was used to cover the sheets. ZDF rats that received ASC sheets had better wound healing than ZDF rats without the transplantation of ASC sheets. This approach was limited because ASC sheets are sensitive to dry conditions, requiring the maintenance of a moist wound environment. Therefore, artificial skin was used to cover the ASC sheet to prevent drying. The allogenic transplantation of ASC sheets in combination with artificial skin might also be applicable to other intractable ulcers or burns, such as those observed with peripheral arterial disease and collagen disease, and might be administered to patients who are undernourished or are using steroids. Thus, this treatment might be the first step towards improving the therapeutic options for diabetic wound healing.

Cytological character of mini pig mesenchymal stromal cells from various tissues and the attempt of cell sheet formation.

INTRODUCTION: Large animal experiments are important for translational research in regenerative medicine. Recently, mini pigs have been used in large animal studies and surgical training. The use of multipotent mesenchymal stromal cell (MSC) sheets for the treatment of many diseases is increasing. The purpose of the present study was to establish optimal methods for generating mini pig MSC sheets from various tissues and to compare the properties of MSCs in these sheets. METHODS: MSCs were isolated from the bone marrow, adipose, periodontal ligament, gingiva, or periosteum of mini pigs. The proliferation, markers, and mRNA expression of these MSCs were examined. Colony-forming and differentiation assays were performed. MSCs were seeded onto temperature-responsive culture dishes to develop MSC sheets. RESULTS: MSCs derived from bone marrow (BMSCs), adipose (ASCs), periodontal ligament (PDLCs), gingiva (GMSCs), and periosteum (PSCs) were positive for MSC-related markers. BMSCs and PSCs showed increased proliferation compared with other MSCs. The osteogenic potential of PDLCs and the adipogenic potential of PSCs were the highest among these MSCs. The expression levels of COL1A1 and COL3A1 in BMSCs and PSCs were significantly higher than those in other MSCs. The expression levels of FGF2, VEGFA, ICAM-1, and TIE-1 in GMSCs were significantly higher than those in other MSCs. PSCs showed the highest levels of TGF-ß1 and ANG-1 expression among all MSC types. We succeeded in developing MSC sheets from BMSCs, ASCs, and PSCs. CONCLUSIONS: We developed methods to generate MSC sheets from various tissues of mini pigs, and these methods are useful to pursue regenerative translational research using mini pigs.

Multipotent mesenchymal stromal cell sheet therapy for bisphosphonate-related osteonecrosis of the jaw in a rat model.

UNLABELLED: Bisphosphonates (BPs) inhibit bone resorption and are frequently used to treat osteoporosis, bone metastasis, and other conditions that result in bone fragility. However, numerous studies have reported that BPs are closely related to the development of osteonecrosis of the jaw (BRONJ), which is an intractable disease. Recent studies have demonstrated that intravenous infusion of multipotent mesenchymal stromal cells (MSCs) is effective for the treatment of BRONJ-like disease models. However, the stability of injected MSCs is relatively low. In this study, the protein level of vascular endothelial growth factor in BP-treated MSCs was significantly lower than untreated-MSCs. The mRNA expression levels of receptor activator of nuclear factor κ-B ligand and osteoprotegerin were significantly decreased in BP-treated MSCs. We developed a tissue-engineered cell sheet of allogeneic enhanced green fluorescent protein (EGFP)-labeled MSCs and investigated the effect of MSC sheet transplantation in a BRONJ-like rat model. The MSC sheet group showed wound healing in most cases compared with the control group and MSC intravenous injection group (occurrence of bone exposure: 12.5% compared with 80% and 100%, respectively). Immunofluorescence staining revealed that EGFP-positive cells were localized around newly formed blood vessels in the transplanted sub-mucosa at 2weeks after transplantation. Blood vessels were significantly observed in the MSC sheet group compared to in the control group and MSC intravenous injection group (106±9.6 compared with 40±5.3 and 62±10.2 vessels/mm(2), respectively). These results suggest that allogeneic MSC sheet transplantation is a promising alternative approach for treating BRONJ. STATEMENT OF SIGNIFICANCE: Bisphosphonates are frequently used to treat osteoporosis, bone metastasis of various cancers, and other diseases. However, bisphosphonate related-osteonecrosis of the jaw (BRONJ) is an intractable disease because it often recurs after surgery or is exacerbated following conservative treatment. Therefore, an alternative approach for treating BRONJ is needed. In this study, we developed a bone marrow-derived multipotent mesenchymal stromal cell (MSC) sheet to treat BRONJ and investigated the effect of MSC sheet transplantation in a rat model of BRONJ-like disease. The MSC sheet transplantation group showed wound healing in most cases, while only minimal healing was observed in the control group and MSC intravenous injection group. Our results suggest that the MSC sheet is a promising alternative approach for the treatment of BRONJ.