Noriko Tachikawa

GROWTH OF THE MALIGNANT AND NONMALIGNANT HUMAN SQUAMOUS CELLS IN A PROTEIN-FREE DEFINED MEDIUM

SummaryA novel protein-free synthetic medium has been developed for the culture of human squamous cell carcinoma cells. This medium, designated PF86-1, supports the serial subcultivation of six out of nine human squamous cell carcinoma cell lines in a protein-free, chemically defined condition without the adapting culture from serum-containing conditions. These cell lines growing in PF86-1 exhibited nearly equal potency to grow in massive culture without noticeable changes in morphology but presented a significantly decreased level of colony forming efficiency when compared with the cells cultured in serum-containing media, suggesting the implication of some autocrine mechanism. Interestingly, this medium supported the growth of normal human squamous cells of oral mucosa and skin for more than 2 mo. in the primary explant culture in spite of high levels of calcium ion concentration, where the overgrowth of fibroblasts as contaminant was not observed. These results suggest that PF86-1 supports the growth of cells derived from epidermal tissues selectively and provides the same defined condition for growth of malignant and nonmalignant human squamous cells. It seems, therefore, that PF86-1 allows investigations on the products of squamous cell carcinoma cells or on the differences of growth mechanisms between normal and neoplastic human squamous cells.

FGF18 accelerates osteoblast differentiation by upregulating Bmp2 expression

Fibroblast growth factor (FGF) signaling is involved in skeletal development. Among total 22 FGFs, it is suggested that FGF18 functions in promotion of osteoblast differentiation. In order to elucidate the mechanism of FGF18‐dependent acceleration of osteogenesis, we implanted rhFGF18 soaked beads over mouse fetal coronal sutures using ex‐utero surgery. The coronal suture area comprises the peripheries of the developing frontal and parietal bones, separated by the sutural mesenchyme. rhFGF18 accelerated osteogenesis by promoting connection of the frontal and parietal bone domains, resulting in elimination of the sutural mesenchyme. Expression of Fgf receptors, Fgfr1, ‐2 and ‐3 involved in skeletal development, was maintained or upregulated in the developing bone domains, consistent with enhanced osteogenesis. Bone morphogenetic protein (Bmp) 2 was specifically upregulated in the skeletogenic layer and the application of Bmp antagonist, rmNoggin, inhibited rhFGF18‐dependent upregulation of osteoblast markers. These results suggest that FGF18 accelerates osteogenesis by upregulation of Bmp2 as well as maintenance or upregulation of Fgfr1, ‐2 and ‐3 expression in osteoblasts.

Lateral Bone Window Closing Technique with Poly-L-Lactic Acid (PLLA) Membrane in the Augmentation of the Maxillary Sinus without Grafting Material: Evaluation of Bone Healing in a Rabbit Model.

BACKGROUND When augmenting the maxillary sinus without grafting material, the method used to cover the bony window is a subject of debate. PURPOSE The purpose of this study was to evaluate the poly-L-lactic acid (PLLA) membrane as closing material of the lateral window in a maxillary sinus augmentation without bone grafting. MATERIALS AND METHODS Augmentation of the maxillary sinus without grafting material and installation of titanium screws that fix the Schneiderian membrane were performed in 18 Japanese male white rabbits. The bony window was covered with a collagen membrane or PLLA membrane or no membrane. The animals were sacrificed at 4 or 8 weeks. New bone volume was calculated radiologically using microcomputed tomography (micro-CT). The samples were analyzed histologically after toluidine blue staining. RESULTS No significant differences were observed in the new bone volume, as measured by micro-CT. However, histomorphometric analysis demonstrated the superiority of the PLLA membrane in new bone formation compared with the collagen membrane. CONCLUSION The PLLA membrane is a suitable material to be applied for tissue regeneration in this animal model.

Involvement of CX3CL1 in the Migration of Osteoclast Precursors Across Osteoblast Layer Stimulated by Interleukin-1ß.

The trigger for bone remodeling is bone resorption by osteoclasts. Osteoclast differentiation only occurs on the old bone, which needs to be repaired under physiological conditions. However, uncontrolled bone resorption is often observed in pro‐inflammatory bone diseases, such as rheumatoid arthritis. Mature osteoclasts are multinuclear cells that differentiate from monocyte/macrophage lineage cells by cell fusion. Although Osteoclast precursors should migrate across osteoblast layer to reach bone matrix before maturation, the underlying mechanisms have not yet been elucidated in detail. We herein found that osteoclast precursors utilize two routes to migrate across osteoblast layer by confocal‐ and electro‐microscopic observations. The osteoclast supporting activity of osteoblasts inversely correlated with osteoblast density and was positively related to the number of osteoclast precursors under the osteoblast layer. Osteoclast differentiation was induced by IL‐1ß, but not by PGE2 in high‐density osteoblasts. Osteoblasts and osteoclast precursors expressed CX3CL1 and CX3CR1, respectively, and the expression of CX3CL1 increased in response to interleukin‐1ß. An anti‐CX3CL1‐neutralizing antibody inhibited the migration of osteoclast precursors and osteoclast differentiation. These results strongly suggest the involvement of CX3CL1 in the migration of osteoclast precursors and osteoclastogenesis, and will contribute to the development of new therapies for bone diseases. J. Cell. Physiol. 232: 1739–1745, 2017.

Histological and Histomorphometrical Determination of the Biogradation of β-Tricalcium Phosphate Granules in Maxillary Sinus Floor Augmentation: A Prospective Observational Study

Introduction: We have recently used highly pure &bgr;-TCP (beta-tricalcium phosphate) as the bone grafting material to avoid highly invasive autogenous bone grafting. We evaluated the osseoconduction potential of highly pure &bgr;-TCP in sinus augmentation surgery treatment. Materials and Methods: The study group comprised 13 patients who underwent maxillary sinus floor augmentation with &bgr;-TCP alone. Seven patients underwent sinus augmentation and implant placement simultaneously. Six patients were treated with a staged approach. Six months after surgery, specimens were obtained from 7 patients (for lateral biopsy) and 6 patients (for vertical biopsy). Results: Histological and histomorphometrical analysis showed a mean bone proportion of 30.8% (vertical) and 12.0% (lateral) for new bone formation and good integration of the &bgr;-TCP. New bone formation was lower in the lateral biopsy specimens than in the vertical. Conclusion: Highly pure &bgr;-TCP is a safe bone-grafting material with superior osteoconductive properties. Histologic and radiographic examinations indicate that &bgr;-TCP is slowly resorbed, which results in unresorbed graft material remaining even 6 months after the procedure, and that new bone replacement occurs slowly for approximately 1 year.

Rehabilitation of a Bilateral Maxillectomy Patient with a Free Fibula Osteocutaneous Flap and with an Implant-Retained Obturator: A Clinical Report

A 47-year-old man underwent surgical resection and reconstruction with a fibula osteocutaneous flap. After the surgery, a surgical obturator was placed and adjusted. After flap healing, a conventional obturator was fabricated with polymethyl methacrylate resin and retained by the anatomical undercuts around the reconstructed fibula bone flap and the posterior part of the defect. As the defect shape changed with time, a second conventional obturator was fabricated and fitted. No further recurrence of myoepithelioma was observed for 2 years, and the patient was satisfied with the obturator during mastication and speech; however, despite having no major complaints, the patient found it difficult to chew on the right side, and the obturator was displaced slightly downward when the mouth was opened wide and shifted when chewing hard and sticky food. Thus, an implant-retained obturator was suggested to provide better retention and stability. Four dental implants were therefore placed into the fibula bone, although one did not osseointegrate because either primary stability was insufficient or overload was affected in the nonloaded implant environment and was replaced. After fitting custom abutments with a magnet, an implant-retained obturator was placed, and the patient was satisfied with the outcome. During 3 years of follow-up, no issues were noted with the implant bodies, abutments, obturator, or reconstructed site. The conventional obturator was displaced slightly downward when the patient opened his mouth wide, and it shifted when chewing hard and sticky food because there was limited fibula bone at the reconstruction site and more available posteriorly. For better retention and stability, the implant-retained obturator was fabricated with a custom abutment and magnetic retention. The patient was satisfied with the results, as improved implant retention increased the stability of the prosthesis. This clinical report describes the rehabilitation of a bilateral maxillectomy patient with a free fibula osteocutaneous flap and an implant-retained obturator. The patients oral functions were improved when the prosthesis was stabilized by means of dental implants and custom abutments.

The Maxillary Sinus Floor Elevation Using a Poly-L-Lactic Acid Device to Create Space Without Bone Graft: Case Series Study of Five Patients

Maxillary sinus floor elevation using autologous or alloplastic bone grafting is often performed for implant treatment of maxillary molars; however, issues related to the donor site and complications such as infection have been reported. We performed maxillary sinus floor elevation using poly-L-lactic acid (PLLA) as a space-making material in patients with an insufficient bone mass (<3 mm) for simultaneous implantation between the alveolar crest and floor of the maxillary sinus and evaluated the newly formed bone. Conventional antrostomy of the maxillary sinus from the lateral wall was performed, and PLLA was placed on the floor of the maxillary sinus after elevating the sinus membrane. Six months after surgery, the bone mass and density were measured using quantitative computed tomography, and histological evaluation was performed. No complications were recorded. Radiological findings showed a bone-like radiopaque appearance, and histological examination revealed new bone formation in all patients. In cases with insufficient bone mass prior to simultaneous implant placement, this method of maxillary sinus augmentation allows for sufficient bone augmentation without bone grafting.

Aesthetical Reconstruction of an Anterior Tooth in an Alveolar Cleft Site: A Case Report and 3-Year Follow-up Findings

Abstract Abstract An alveolar cleft often displays the furrowed shape of the buccal site because of the lack of alveolar bone and gingival contracts. A 35-year-old woman presented with a repaired cleft lip and a non-grafted alveolar cleft associated with a missing lateral incisor on the left side of the maxilla. The central incisor had been an abutment tooth of a bridge for 15 years; however, it fractured and was extracted. Orthodontic treatment was antecedent to implant reconstruction because the edentulous space was not adequate for a single tooth replacement. As the palate was closed by the mucosa, bone grafting was not performed at the cleft site beforehand. Dental implant placement was performed within the narrowed, beamlike bone, and hydroxyapatite particles were used to augment the ridge. A relaxing incision and expansion of the invaginated mucosa improved the buccal shape of the gum. The usage of a narrow-type implant and bone substitutes facilitated the minimization of surgical stress, even though she did not undergo autologous bone-grafting surgery. Additionally, the aesthetical reconstruction of the anterior region significantly improved her quality of life at the 3-year follow-up.

Cross-sectional imaging of extracted jawbone of a pig by optical coherence tomography

Dental implantation has become popular in dental treatments. Although careful planning is made to identify vital structures such as the inferior alveolar nerve or the sinus, as well as dimensions of the bone, prior to commencement of surgery, dental implantation is not fully free from risks. If a diagnostic tool is available to objectively measure bone feature before surgery and dimensions during surgery, considerable fraction of the risks may be avoided. Optical coherence tomography (OCT) is a candidate for the purpose, which enables cross-sectional imaging of bone. In this work, we performed in vitro cross-sectional imaging of extracted pigs jawbone with swept source OCT using superstructure-grating distributed Bragg reflector (SSG-DBR) laser as the source. The relatively long wavelength range of 1600nm of the laser is suitable for deeper bone imaging. We confirmed an image penetration depth of about 3 mm in physical length, which satisfies one of the criterions to apply OCT for in vivo diagnosis of bone during surgery.

Guided bone regeneration using a hydrophilic membrane made of unsintered hydroxyapatite and poly(L-lactic acid) in a rat bone-defect model

The effectiveness of a previously developed unsintered hydroxyapatite (uHA) and poly(L-lactic acid) (PLLA) hydrophilic membrane as a resorbable barrier for guided bone regeneration (GBR) was evaluated. Critical-size 8-mm diameter bone defects were surgically generated in the parietal bones of 24 12-week-old male Wistar rats, which were then divided into three groups in which either a uHA/PLLA or a collagen membrane or no membrane (control) was placed onto the bone defect. Following sacrifice of the animals 2 or 4 weeks after surgery, bone defects were examined using microcomputed tomography and histological analysis. Bone mineral density, bone mineral content, and relative bone growth area values 2 or 4 weeks after surgery were highest in the uHA/PLLA group. Four weeks after surgery, the relative bone growth area in the uHA/PLLA group was larger than that in the collagen group. The resorbable uHA/PLLA membrane is thus potentially effective for GBR.