Three-Dimensional Spheroid Formation of Cryopreserved Human Dental Follicle-Derived Stem Cells Enhances Pluripotency and Osteogenic Induction Properties

BACKGROUND: Enhancement and maintenance of the stemness of mesenchymal stem cells (MSCs) is one of the most important factors contributing to the successful in vivo therapeutic application of these cells. In this regard, three-dimensional (3D) spheroid formation has been developed as reliable method for increasing the pluripotency of MSCs. Moreover, using a new protocol, we have previously shown that dental tissues of extracted wisdom teeth can be effectively cryopreserved for subsequent use as a source of autologous stem cells. The main purpose of this study is to analyze the stemness and in vitro osteogenic differentiation potential of 3D spheroid dental MSCs compared with conventional monolayer cultured MSCs. METHODS: In this study, MSC-characterized stem cells were isolated and cultured from long-term cryopreserved dental follicles (hDFSCs), and then 2D hDFSCs were cultured under 3D spheroid-forming conditions using a newly designed microchip dish. The spheroids (3D hDFSCs) thus produced were investigated and characterized with respect to stemness, MSC marker expression, apoptosis, cell cycle analysis, extracellular matrix (ECM) production, and osteogenic and adipogenic differentiation properties. RESULTS: In terms of MSC and senescence markers, spheroid cells showed no difference when compared with 2D hDFSCs; however, 3D hDFSCs were observed to have a higher proportion of cell cycle arrest and a larger number of apoptotic cells. Moreover, spheroids showed substantially increased levels of pluripotency marker (early transcription factors) and ECM protein expression. Compared with 2D hDFSCs, there was also a notable enhancement in the osteogenic induction potential of spheroids, although no differences were observed with respect to in vitro adipogenesis. CONCLUSION: To the best of our knowledge, this is the first study to demonstrate the application of a spheroid culture system for dental follicle-derived stem cells using a microchip dish. Although further studies are needed, including in vivo transplantation, the results obtained in this study indicate that spheroid hDFSCs derived from cryopreserved dental follicle tissues could be used as a valuable source of autologous stem cells for bone tissue regeneration.

Dental stem cells as a cell source for tissue engineering

No abstract available.

Tumor resection from retromolar trigone, posterolateral maxilla, and anterior mandibular ramus using lower cheek flap approach: a case report and review of literature

A surgical approach involving the retromolar trigone, posterolateral maxilla, and pterygoid region is the most challenging in the field of maxillofacial surgery. The upper cheek flap (Weber-Ferguson incision) with subciliary extension and the maxillary swing approach have been considered as alternatives; however, neither approach provides sufficient exposure of the pterygoid region and the anterior portion of the mandibular ramus. In this report, we describe two cases in which a lower cheek flap approach was used for complete tumor resection in the retromolar trigone and the anterior mandibular ramus. This approach allows full exposure of the posterolateral maxilla and the pterygoid region as well as the retromolar trigone without causing major sensory disturbances to the lower lip. A mental nerve anastomosis after tumor resection was performed in one patient and resulted in approximately 90% sensory recovery in the lower lip. The lower cheek flap approach provides adequate exposure of the posterolateral maxilla, including the pterygoid, retromolar trigone, and mandibular ramus areas. If the mental nerve can be anastomosed during flap approximation, postoperative sensory disturbances to the lower lip can be minimized.

Stability of unilateral sagittal split ramus osteotomy for correction of facial asymmetry: long-term case series and literature review

Bilateral sagittal split ramus osteotomy is considered a standard technique in mandibular orthognathic surgeries to reduce unexpected bilateral stress in the temporomandibular joints. Unilateral sagittal split ramus osteotomy (USSO) was recently introduced to correct facial asymmetry caused by asymmetric mandibular prognathism and has shown favorable outcomes. If unilateral surgery could guarantee long-term postoperative stability as well as favorable results, operation time and the incidence of postoperative complications could be reduced compared to those in bilateral surgery. This report highlights three consecutive cases with long-term follow-up in which USSO was used to correct asymmetric mandibular prognathism. Long-term postoperative changes in the condylar contour and ramus and condylar head length were analyzed using routine radiography and computed tomography. In addition, prior USSO studies were reviewed to outline clear criteria for applying this technique. In conclusion, patients showing functional-type asymmetry with predicted unilateral mandibular movement of less than 7 mm can be considered suitable candidates for USSO-based correction of asymmetric mandibular prognathism with or without maxillary arch surgeries.

Cryopreservation of dental tissue and subsequent isolation of mesenchymal stem cells

No abstract available.

Cryopreservation of Human Wharton's Jelly-derived Mesenchymal Stem Cells Following Controlled Rate Freezing Protocol Using Different Cryoprotectants; A Comparative Study

OBJECTIVES: To compare the effect of three different cryoprotectants on basic stem cell characteristics for the possibility of using well defined, dimethyl sulfoxide (DMSO) and serum free freezing solutions to cryopreserve human Wharton's jelly-derived mesenchymal stem cells (WJMSCs) following controlled rate freezing protocol. METHODS: The mesenchymal stem cells isolated from human Wharton's jelly were cryopreserved using 10% DMSO, 10% polyvinylpyrrolidone (PVP) and a cocktail solution comprising of 0.05 M glucose, 0.05 M sucrose and 1.5 M ethylene glycol following controlled rate freezing protocol. We investigated the post-thaw cell viability, morphology, proliferation capacity, basic stem cell characteristics, in vitro differentiation potential and apoptosis-related gene expression profile before and after cryopreservation. RESULTS: The cryoprotectant 10% DMSO has shown higher post-thaw cell viability of 81.2+/-0.58% whereas 10% PVP and cocktail solution have shown 62.87+/-0.35% and 72.2+/-0.23%, respectively at 0 h immediately thawing. The cell viability was further reduced in all the cryopreserved groups at 24 h later post-thaw culture. Further, the complete elimination of FBS in cryoprotectants has resulted in drastic reduction in cell viability. Cryopreservation did not alter the basic stem cell characteristics, plasticity and multipotency except proliferation rate. The expression of pro-apoptotic BAX and p53 genes were higher whilst p21 was lower in all the cryopreserved groups when compare to the control group of WJMSCs. CONCLUSION: Although 10% DMSO has shown higher post-thaw cell viability compare to 10% PVP and cocktail solution, the present study indicates the feasibility of developing a well-defined DMSO free cryosolution which can improve storage and future broad range applications of WJMSCs in regenerative medicine without losing their basic stem cell characteristics.

Assessment of functional improvement with temporalis myofascial flap after condylectomy in elderly patients with anterior disc displacement without reduction and an erosive condylar surface / 대한악안면성형재건외과학회지

BACKGROUND: The purpose of this study was to investigate the functional effects of temporalis myofascial flap after condylectomy, with or without disc removal, in elderly patients with anterior disc displacement (ADD) without reduction and an erosive condylar surface of the temporomandibular joint (TMJ). METHODS: A total of 15 joints from 11 elderly patients (71-78 years old) were included. The patients had pain, mandibular dysfunction symptoms, and unilateral or bilateral ADD as well as an erosive condylar surface of the TMJ. All patients underwent temporalis myofascial flap reconstruction after condylectomy, with or without disc removal. If the maximal mouth opening (MMO) remained <35 mm after condylectomy, coronoidotomy was also performed. Self-assessed pain and mandibular function, including MMO and protrusive and lateral movements, were evaluated. RESULTS: No patient experienced serious complications. Most measurements improved significantly after surgery compared to preoperatively. Most patients achieved nearly-normal mouth opening at 4 weeks after surgery. Although most patients felt discomfort during active postoperative physiotherapy, no patient reported serious pain during the follow-up period. CONCLUSION: Although nonsurgical therapy is often the first treatment choice for ADD without reduction of the TMJ, surgical intervention involving condylectomy and temporalis myofascial flap reconstruction may be a reasonable first option for elderly patients with an erosive condylar surface of the TMJ.

Angiokeratoma circumscriptum of the buccal mucosa: a case report and literature review

Angiokeratoma is a benign cutaneous lesion of the capillaries, presenting as dilated vessels in the upper part of the dermis. Although this disorder is classified into various types and has been occasionally reported in the skin of the scrotum or extremities, the involvement of the oral cavity mucosa has been rarely reported. The present study reports a case of angiokeratoma circumscriptum in the buccal mucosa. The expression of vascular endothelial growth factor (VEGF) and both of its receptors (VEGFR-1 and VEGFR-2) was demonstrated by immunohistochemistry in the endothelial cells lining the dilated vessels. The expression of VEGFR-2 was higher than that of VEGFR-1 in the endothelial cells in the lesion, indicating an increased rate of endothelial cell proliferation within the lesion. Interestingly, some of the endothelial cells co-expressed VEGF and its two receptors. These results suggest that endothelial cells in the pathologically dilated vessels possess VEGF autocrine growth activity involved in vasculogenesis and maintenance in angiokeratoma lesions. To our knowledge, this is the second report published on isolated oral angiokeratoma confined to the buccal mucosa and the first case report on angiokeratoma circumscriptum involving the buccal mucosa.

Successful Localization of Intraoral Foreign Body with C-arm Fluoroscopy / 대한악안면성형재건외과학회지

During surgical procedures, unexpected material, including surgical instruments and tissue segments, may get lost in the surgical field. Most of these should be immediately removed to prevent further complications, such as vital organ irritation, infection, and inflammatory pseudo-tumor formation. However, it is not always easy to define the exact location of the foreign body, especially if the item is very small and/or it is embedded in the soft tissue of the head and neck region. Intraoperative real-time radiological imaging with C-arm fluoroscopy can be useful to trace the three-dimensional location of small and embedded foreign bodies in the oral and maxillofacial area. We describe an unusual case of an embedded micro-screw in the intrinsic tongue muscle that had been dropped into the sublingual space during a lower alveolar bone graft procedure. The lost foreign body was accurately identified with C-arm fluoroscopy and safely removed without any further complications.

Expansile keratocystic odontogenic tumor in the maxilla: immunohistochemical studies and review of literature

Keratocystic odontogenic tumors (KCOT) - previously termed odontogenic keratocysts (OKC) - are characterized by aggressive behavior and a high rate of recurrence. Histopathologically, the basal layer of KCOT shows a higher cell proliferation rate and increased expression of anti-apoptosis genes. Clinically, KCOT is frequently involved in the mandibular posterior region but is not common in the posterior maxilla. However, it should be noted that due to its expansive characteristics, KCOT involved near the maxillary sinus could easily expand to an enormous size and occupy the entire maxilla. To achieve total excision of these expanded cystic tumors in the maxilla, a more aggressive approach would be needed. In this report, we describe two cases of expansile KCOT involving the entire unilateral maxilla and maxillary sinus; they were completely excised using the Weber-Ferguson approach, showing no evidence of recurrence during the follow-up period of more than two years. In immunohistochemical analyses of the tumor specimens, p53 and p63 showed strong expression, and B-cell lymphoma 2 (BCL2) and MKI67 (Ki-67) showed moderate or weak expression, however, detection of BCL2-associated X protein (BAX) was almost negative. These data indicate that expansile KCOT possesses increased anti-apoptotic activity and cell proliferation rate but decreased apoptosis. These properties of KCOT may contribute to tumor enlargement, aggressive behavior, and high recurrence rate.

Dental tissues as adult stem cell source

No abstract available.

Slimming of Lower Face by Contouring of the Mandibular Body in Orthognathic Surgery Patients

Isolation of human mesenchymal stem cells from the skin and their neurogenic differentiation in vitro

OBJECTIVES: This aim of this study was to effectively isolate mesenchymal stem cells (hSMSCs) from human submandibular skin tissues (termed hSMSCs) and evaluate their characteristics. These hSMSCs were then chemically induced to the neuronal lineage and analyzed for their neurogenic characteristics in vitro. MATERIALS AND METHODS: Submandibular skin tissues were harvested from four adult patients and cultured in stem cell media. Isolated hSMSCs were evaluated for their multipotency and other stem cell characteristics. These cells were differentiated into neuronal cells with a chemical induction protocol. During the neuronal induction of hSMSCs, morphological changes and the expression of neuron-specific proteins (by fluorescence-activated cell sorting [FACS]) were evaluated. RESULTS: The hSMSCs showed plate-adherence, fibroblast-like growth, expression of the stem-cell transcription factors Oct 4 and Nanog, and positive staining for mesenchymal stem cell (MSC) marker proteins (CD29, CD44, CD90, CD105, and vimentin) and a neural precursor marker (nestin). Moreover, the hSMSCs in this study were successfully differentiated into multiple mesenchymal lineages, including osteocytes, adipocytes, and chondrocytes. Neuron-like cell morphology and various neural markers were highly visible six hours after the neuronal induction of hSMSCs, but their neuron-like characteristics disappeared over time (24-48 hrs). Interestingly, when the chemical induction medium was changed to Dulbecco's Modified Eagle Medium (DMEM) supplemented with fetal bovine serum (FBS), the differentiated cells returned to their hSMSC morphology, and their cell number increased. These results indicate that chemically induced neuron-like cells should not be considered true nerve cells. CONCLUSION: Isolated hSMSCs have MSC characteristics and express a neural precursor marker, suggesting that human skin is a source of stem cells. However, the in vitro chemical neuronal induction of hSMSC does not produce long-lasting nerve cells and more studies are required before their use in nerve-tissue transplants.

In vivo Osteogenesis of Cultured Human Periosteal-derived Cells and Polydioxanone/Pluronic F127 Scaffold

Use of Peristeum as a Source of Endothelial-like Cells

Effects of Neuromuscular Electrical Stimulation on Distracted Bone after Mandibular Distraction Osteogenesis in Canine Model

Sialadenoma papilliferum: a case report and immunohistochemical study review

Sialadenoma papilliferum (SP) is a rare benign neoplasm that normally arises from the minor salivary glands, particularly in the palate. SP is normally encountered in older men with an exophytic papillary surface growth. In the present study, an SP of the hard palate of a 69-year-old woman was examined immunohistochemically. Myoepithelial cell markers, such as S-100, smooth muscle actin and vimentin, were observed in the basal or luminal layer of tumor cells, indicating that myoepithelial cells participate in the pathogenesis of SP. In addition, cytokeratin 7 was also strongly detected in the tumor cells, suggesting that excretory ductal epithelial cells have a role in its histogenesis. A review of the literature of immunohistochemical studies on SP showed that the expression and co-expression of cytokeratins and myoepithelial cell markers have been reported in tumor cells. These results suggested that excretory duct cells and myoepithelial cells participate in the pathogenesis of SP.

Maxillary sinus floor elevation using autogenous skin-derived mesenchymal stem cells in miniature pigs

INTRODUCTION: In our previous studies, we isolated porcine skin-derived mesenchymal stem cells (pSDMSCs) from the ears of adult miniature pigs and evaluated the pluripotency of these pSDMSCs based on expressions of transcription factors, such as Oct-4, Sox-2, and Nanog. Moreover, the characteristic of mesenchymal stem cells was revealed by the expression of various mesenchymal stem cell markers, including CD29, CD44, CD90, and vimentin. The aim of this study was to evaluate in vivo osteogenesis after maxillary sinus lift procedures with autogenous pSDMSCs and scaffold. MATERIALS AND METHODS: The autogenous pSDMSCs were isolated from the 4 miniature pigs, and cultured to 3rd passage with same methods of our previous studies. After cell membranes were labeled using a PKH26, 1x10(7) cells/100 microliter of autogenous pSDMSCs were grafted into the maxillary sinus with a demineralized bone matrix (DBM) and fibrin glue scaffold. In the contralateral control side, only a scaffold was grafted, without SDMSCs. After two animals each were euthanized at 2 and 4 weeks after grafting, the in vivo osteogenesis was evaluated with histolomorphometric and osteocalcin immunohistochemical studies. RESULTS: In vivo PKH26 expression was detected in all specimens at 2 and 4 weeks after grafting. Trabecular bone formation and osteocalcin expression were more pronounced around the grafted materials in the autogenous pSDMSCs-grafted group compared to the control group. Newly generated bone was observed growing from the periphery to the center of the grafted material. CONCLUSION: The results of the present study suggest that autogenous skin-derived mesenchymal stem cells grafting with a DBM and fibrin glue scaffold can be a predictable method in the maxillary sinus floor elevation technique for implant surgery.

Squamous cell carcinoma occurring with aspergillosis in the maxillary sinus: a case report and histological study

The coexistence of aspergillosis and squamous cell carcinoma (SCC) in the maxillary sinus was very rare. To our knowledge, this is the second report of coexistent SCC and aspergillosis in the maxillary sinus. A 58-year-old man underwent surgery for unilateral maxillary sinus infection with oroantral fistula. In the surgical specimen, SCC and aspergillosis were co-detected with routine and immunohistochemical stainings. Moreover, human papillomavirus 18 (HPV-18) was detected by polymerase chain reaction in the sinus specimen. The patient was re-operated with subtotal maxillectomy and has been followed up for two years without any evidence of recurrence or metastasis. Although it is not understood how aspergillosis could induce carcinoma formation, the chronic inflammation caused by prolonged fungal infection might be carcinogenic. Moreover, HPV-16 and -18 were another causative pathogens of SCC in the head and neck region. We recommend careful examination, including preoperative cytology, in patients with maxillary sinus fungal infections because of the potential for cancer development.

Evaluation of osteogenic activity of periosteal-derived cells treated with inflammatory cytokines

INTRODUCTION: Skeletal homeostasis is normally maintained by the stability between bone formation by osteoblasts and bone resorption by osteoclasts. However, the correlation between the inflammatory reaction and osteoblastic differentiation of cultured osteoprogenitor cells has not been fully investigated. This study examined the effects of inflammatory cytokines on the osteoblastic differentiation of cultured human periosteal-derived cells. MATERIALS AND METHODS: Periosteal-derived cells were obtained from the mandibular periosteum and introduced into the cell culture. After passage 3, the periosteal-derived cells were further cultured in an osteogenic induction Dulbecco's modified Eagle's medium (DMEM) medium containing dexamethasone, ascorbic acid, and beta-glycerophosphate. In this culture medium, tumor necrosis factor (TNF)-alpha with different concentrations (0.1, 1, and 10 ng/mL) or interleukin (IL)-1beta with different concentrations (0.01, 0.1, and 1 ng/mL) were added. RESULTS: Both TNF-alpha and IL-1beta stimulated alkaline phosphatase (ALP) expression in the periosteal-derived cells. TNF-alpha and IL-1beta increased the level of ALP expression in a dose-dependent manner. Both TNF-alpha and IL-1beta also increased the level of alizarin red S staining in a dose-dependent manner during osteoblastic differentiation of cultured human periosteal-derived cells. CONCLUSION: These results suggest that inflammatory cytokines TNF-alpha and IL-1beta can stimulate the osteoblastic activity of cultured human periosteal-derived cells.