Indirect osteoblast differentiation by liposomal clodronate.

Bisphosphonates impair function of osteoclasts and prevent bone resorption, the mechanism of which has been studied extensively. However, the possible effects of bisphosphonates on chondroblast differentiation and calcium deposition by osteoblasts have only been demonstrated recently. Moreover, cells from monocytic lineage are capable of stimulating osteoblast proliferation. Hence, susceptibility of osteoblasts to various factors requires further investigation. A primary culture of bone marrow-derived stromal cells was treated with liposomal clodronate (0.1, 0.5, or 1.0 mg/ml) or conditioned medium from liposomal clodronate. Liposomal clodronate (0.25 mg) was injected into mouse femur for in vivo experiments. The effects of liposomal clodronate were examined by alkaline phosphatase staining and/or activity assay, and real-time RT-PCR was used for studying the effect on osteogenic gene expression. Administration of liposomal clodronate to bone marrow-derived mesenchymal stromal cell culture enhanced alkaline phosphatase activity and mRNA levels of Runx2 and Dlx5. In addition, conditioned medium from liposomal clodronate also stimulated osteogenic characteristics similar to those of observed in vitro, and the number of exosomes in the conditioned medium was highest when pre-treated with liposomal clodronate. Western blot analysis revealed the presence of RANK proteins in exosomes collected from conditioned medium of liposomal clodronate. Identical observations were obtained in vivo, as liposomal clodronate-injected mouse femur showed increased alkaline phosphatase activity and Runx2 and Dlx5 mRNA expressions, even though the numbers of monocytes and macrophages were reduced. In conclusion, osteoblast differentiation was promoted via soluble RANK-containing exosomes in response to clodronates.

Unit Advancement Flap for Lower Lip Reconstruction.

BACKGROUND: Lower lip reconstruction requires consideration of esthetic and functional outcome in selecting a surgical procedure, and reconstruction with local tissue is useful. The authors reconstructed full-thickness defects with a unit advancement flap. METHODS: Reconstruction was performed using this method in 4 patients with lower lip squamous cell carcinoma in whom tumor resection with preservation of the mouth angle was possible. The lower lip resection width was 30 to 45 mm, accounting for 50% to 68% of the entire width of the lower lip. The flap was prepared by lateral extension from above the mental unit and matched with the potential wrinkle line of the lower lip in order to design a unit morphology surrounded by the anterior margin of the depressor labii inferioris muscle. It was elevated as a full-thickness flap composed of the orbicularis oris muscle, skin, and mucosa of the residual lower lip from the bilateral sides, and advanced to the defect. Flap transfer was adjusted by small triangular resection of the skin on the lateral side of the mental unit. RESULTS: The postoperative scar was inconspicuous in all patients and there was no impairment of the mouth opening-closing or articulation functions. CONCLUSIONS: This was a relatively simple surgical procedure. A blood supply of the flap was stable, and continuity of the orbicularis oris muscle was reconstructed by transferred the residual lower lip advancement flap from the bilateral sides. The postoperative mouth opening-closing function was sufficient, and dentures could be placed from an early phase in elderly patients. The postoperative scar was consistent with the lip unit morphology, being esthetically superior. This procedure may be applicable for reconstruction of defects approximately 1/3 to 2/3 the width of the lower lip where the mouth angle is preserved.

The Rintala flap: a versatile procedure for nasal reconstruction.

INTRODUCTION: For the nasal reconstruction, local flap using the adjacent tissue is selected from an esthetic viewpoint. The Rintala flap is a useful option, and reconstruction of the glabellar over the nasal tip with this flap is ideal, for which the procedure was modified to increase the blood flow to the Rintala flap to extend its clinical applications. METHODS: For defects of the glabellar over the middle one third of nasal dorsum, the Rintala flap is transferred employing the original design and technique dissected on supraperiosteal plane. For defects of the lower one third of nasal dorsum over the nasal tip, blood supply through the lateral nasal artery is added to the distal end of the flap, preparing a long flap with stable blood supply like Maruyama described in 1997. RESULTS: This procedure was applied for nasal reconstruction in 15 patients. The Rintala flap was applied in 12 and the modified Rintala flap with adding blood flow from the lateral nasal artery was applied in 3. Blood supply to the flap was very stable in all patients, and favorable outcomes were achieved. CONCLUSIONS: Using this procedure, the natural contour and morphology of the glabellar over the nasal tip may be reconstructed. The technique is simple and easy. Using this procedure, clinical applications of the Rintala flap can be extended, showing that it is a useful nasal reconstructive procedure.

Promoted activation of matrix metalloproteinase (MMP)-2 in keloid fibroblasts and increased expression of MMP-2 in collagen bundle regions: implications for mechanisms of keloid progression.

AIMS: Keloid is characterized by excessive deposition of collagen, resulting from aberrant extracellular matrix (ECM) production and degradation. The aim was to investigate the role of matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases (TIMPs) in pathological wound healing in keloids. METHODS AND RESULTS: Semiquantitative analysis of 60 keloid tissue samples and 25 mature scar tissue samples demonstrated significantly increased expression of MMP-2, TIMP-2 and TIMP-3 in keloids compared with mature scars. Within keloid regions, MMP-2 expression was significantly higher in collagen bundle regions than in non-collagen bundle regions. Double immunofluorescence revealed that keloid fibroblasts between collagen bundles exhibited MMP-2, TIMP-2 and membrane-type 1 MMP (MT1-MMP) co-expression, whereas only MMP-2 expression was evident on the edge of collagen bundles. Western blot analysis and gelatin zymography of 13 keloid-derived fibroblasts (KFbs) and six normal skin dermal-derived fibroblasts (NFbs) demonstrated that unstimulated KFbs exhibited significantly increased MMP-2 activity and expression compared with NFbs under the same conditions. CONCLUSIONS: These results together indicate that MMP-2 activity can be promoted in keloid fibroblasts between collagen bundles in cooperation with TIMP-2 and MT1-MMP. This could contribute to remodelling of collagen bundle regions and invasion of fibroblasts into peripheral normal regions through promoted degradation of ECM.

Medial canthal reconstruction with multiple local flaps.

BACKGROUND: It is often difficult to reconstruct comparatively large defects in the medial canthal region. The authors have performed reconstruction with multiple local flaps in 4 medial canthal defects after resection of malignant skin tumors. METHOD: The defects exceeded the medial canthal region, extending to the upper eyelid and the lower eyelid. The medial canthal defect was covered by transposition of a glabellar flap, the upper eyelid defect was covered by an upper eyelid myocutaneous advancement flap, and the lower eyelid defect was covered by a cheek rotation flap. Then the donor site of the glabellar flap was covered by a Rintala flap. RESULTS: There was no recurrence in any of the cases, and good results were obtained. One case showed mild linear contracture, but the patient did not want corrective surgery. CONCLUSION: This method is somewhat complicated compared to reconstruction with a single flap, but it is a combination of standard local flaps and is a simple reconstructive procedure. By adding additional resection, the suture line is consistent with the border of the facial unit, so postoperative scarring is inconspicuous. This technique is aesthetically useful because of the continuity of colour and texture resulting from the use of adjacent flaps.

Osteogenic Potential of Adipose-Derived Macrospheroids Cocultured with CD11b+ Monocytes.

PURPOSE: Among potential cell-based therapies, adipose-derived stem cells (ASCs) have been proposed as a promising source of stem cells for tissue regeneration. Although many recent clinical trials have investigated the use of adipose tissue or ASCs in transplantation, analysis of the microstructures of outgrowing macrosized spheroids (macrospheroids) or three-dimensional coculture of ASC spheroids and monocyte/macrophage lineages has not been performed. The aim of this study was to analyze the microstructures of murine-derived ASC macrospheroids and the growth and osteogenic potential of these macrospheroids in a three-dimensional environment and after calcification induction by coculture with monocytes. MATERIALS AND METHODS: The histologic structures of murine-derived ASC macrospheroids and the expression of marker genes for multipotency within these macrospheroids were analyzed by hematoxylin and eosin staining and in situ hybridization. ASC macrospheroid microstructures were observed by transmission electron microscopy, and cell proliferation in the spheroids was analyzed. Additionally, the growth and osteogenic potential of these macrospheroids were assessed in two-dimensional and three-dimensional environments and after calcification induction by coculture with monocytes. RESULTS: The expression of Oct3/4, Nanog, and Sox2 was detected even in the deep zone of spheroids, although higher expression was observed at the surface. Cell proliferation was detected within the spheroid centers. Observation of spheroid microstructure revealed extracellular matrix production within the spheroid architecture. Transplantation of a spheroid on the hydroxyapatite disc resulted in three-dimensional cell growth, filling the disc. Coculture of the spheroids with monocytes led to the formation of many osteoclast-like, multinucleated cells, and calcification was observed after 3 weeks of coculture. CONCLUSION: ASC spheroids exhibited high capacity for dynamic three-dimensional growth and osteogenic differentiation. Furthermore, ASC spheroids promoted monocyte differentiation into osteoclast-like cells, which may enhance the osteogenic potential of ASC spheroids.

Histological and micro-computed tomographic observations after maxillary sinus augmentation with porous hydroxyapatite alloplasts: a clinical case series.

BACKGROUND: It is important to visualize the phenomenon which occurs in actual clinical cases to decide the timing of implant placement subsequently after sinus graft. Although several clinical cases of bone augmentation using xenograft have been evaluated, the number of those reports which have described bone remodeling by alloplasty are not sufficient. FINDINGS: In the present report, to investigate bone remodeling histologically after maxillary sinus augmentation with porous hydroxyapatite alloplast, bone cores from the sinus floor of three female nonsmoking patients (aged 64-73 years) were collected 6 months after sinus lift surgery, embedded in methyl methacrylate resin, and prepared by conventional methods. Bone architecture and graft residues were evaluated by micro-computed tomography of the same specimens. Hematoxylin-eosin and Villanueva-Goldner staining revealed mature osteoblasts and multinucleated osteoclasts on the grafted sinus floor and surface of residual hydroxyapatite particles. The particulate interspace was partially filled with osteoid and calcified tissue and showed active vascularization. CONCLUSION: The results suggested that bone regeneration and angiogenesis within and between porous hydroxyapatite particles were sufficiently found after 6 month histologically in the grafted sinus floor.

Medial canthal reconstruction with glabellar combined Rintala flaps.

BACKGROUND: In medial canthal reconstruction, the continuity of color and texture and the reproduction of natural external appearance are required, and reconstruction with a flap adjacent to the defect is frequently performed as the first choice. The authors have performed reconstruction with a glabellar flap combined with a Rintala flap in five patients. METHODS: A glabellar flap, which is rectangular according to the unit principle, was elevated, and after thinning and trimming to the defect shape, was transferred to the medial canthal defect. The donor site of the glabellar flap was closed with a Rintala flap. RESULTS: Reconstruction by means of this procedure was performed in five patients with defects after resection of medial canthal basal cell carcinoma. In one patient with extension of the defect to the upper and lower eyelids, the tip of the glabellar flap was divided into two portions and transferred to the defective site. No recurrence was observed in any patient, and satisfactory results including aesthetic results were obtained in all patients. CONCLUSIONS: This technique can be performed readily and is applicable to the reconstruction of relatively large defects and is also aesthetically excellent because of a postoperative suture line that is consistent with the topographic curve. This technique may be useful for medial canthal reconstruction.

Nasal augmentation using calcium phosphate cement.

This report describes a simple technique for augmentation rhinoplasty using calcium phosphate cement paste for postoperative and posttraumatic nasal deformities. This biomaterial was used to correct traumatic nasal bone deformity in one patient and for augmentation rhinoplasty after removal of the nasal implant in two patients. These patients were followed for a minimum 6 months and a maximum of 26 months. The esthetic results were satisfactory with no complication. Calcium phosphate cement consisting of alpha-tricalcium phosphate, dicalcium phosphate dibasic, and tetracalcium phosphate monoxide has been used for bone replacement and augmentation because of its good biocompatibility and osteoconductivity. This technique is easy and safe. It seems to be a suitable material for augmentation rhinoplasty in the Orient. Its long-term safety and reliability require proof with longer follow-up periods, however.