[Show abstract][Hide abstract] ABSTRACT: To date there is no sufficient in vitro fat tissue engineering and a protocol has not been well established for this purpose. Therefore, we evaluated the in vitro influence of two different adipogenic growth media for their stimulation potential on different cell lineages to clearly define the most potent adipogenic growth media for future in vitro tissue engineering approaches. The samples for differentiation were composed of human adipogenic-derived stroma cells (hADSCs) and human bone marrow mesenchymal stroma cells (hMSCs). A normal adipogenic medium (NAM) and a specific adipogenic medium (SAM) were tested for their adipogenic stimulation potential. After 10 days and 21 days the relative gene expression was measured for the adipogenic marker genes PPARγ2, C/EBPα, FABP4, LPL, and GLUT4 detected through real time reverse transcriptase polymease chain reaction (RT-PCR). Other study variables were the comparison between NAM and SAM and between the used cells hADSCs and hMSCs. Additionally an Oil-Red staining was performed after 21 days. Our results revealed that only SAM was significantly (P<0.05) superior in the differentiation process in contrast to NAM for 10 days and 21 days. As well was SAM superior to differentiate the used cell lineages. This was evaluated by the detected marker genes PPARγ2, C/EBPα, FABP4, LPL, and GLUT4 through real time RT-PCR and by Oil-Red staining. In addition, the hMSCs proofed to be equal donor cells for adipogenic differentiation especially when stimulated by SAM. The results suggest that the SAM should be established as a new standard medium for a more promising in vitro adipogenic differentiation.
[Show abstract][Hide abstract] ABSTRACT: The changes in the surrounding soft tissues during long bone distraction in orthopedic surgery have been the subject of several reports, studies on changes in the craniofacial region, in which various tissues, including the skin, muscle, tendon, blood vessel, and gingiva are rare. Therefore, there is a need for studies on the soft tissue aspects of bone lengthening of the craniofacial region. The aim of this review was to address this issue by reviewing the literature about the distraction histogenesis of various tissues, including skin, muscle, blood vessel, nerve, and gingiva.
No preview · Article · Apr 2015 · Oral and Maxillofacial Surgery
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to assess the factors, mechanisms and the differences between periodontal ligament (PDL) cells and denta
l follicle (DF) progenitor cells towards the osteoblastic/cementoblastic differentiation and to investigate the effects of BMP-7 on developmental (DF) and mature tissue-derived (PDL) cells, respectively. Primary cell culture of PDL cells and DF progenitor cells was performed. Osteogenic differentiation was evaluated using von Kossa, Alizarin Red S and immuno-histo-chemistry staining of osteocalcin. Gene expression pattern was evaluated via real-time PCR. A series of CD surface marks were tested using flow cytometry and fluorescence-activated cell-sorting analysis was performed. Real-time RT-PCR demonstrated similar gene expression pattern of PDL cells and DF progenitor cells: the expression of OPN and OCN significantly was elevated when incubated with osteogenic components, Runx2 was unaffected, and Osteorix was hardly expressed whether in basic medium or induction medium. In addition, BMP-7 induced osteoblast/cementoblast differentiation of PDLSCs and DF progenitor cells in a dose- and time-dependent manner, as reflected by enhanced Runx2 and (OCN) mRNA transcript expression. BMP-7 triggers PDL cells and DF progenitor cells to differentiate towards an osteoblast/cementoblast phenotype.
[Show abstract][Hide abstract] ABSTRACT: A negative side effect of therapeutic irradiation is the radiation-induced bone loss which can lead, in long term, to pathological fractures. Until today, the detailed mechanism is unknown. If osteoclasts would mainly contribute to the pathological bone loss, bisphosphonates could potentially counteract the osteolytic process and possibly help to prevent long-term complications. The aim of this study was to evaluate the effect of zoledronic acid on the early radiation-induced degradation of bone collagen fibrils by monitoring the urinary excretion of hydroxylysylpyridinoline and lysylpyridinoline under radiotherapy.
A total of 40 patients with skeletal metastases were assigned for a local radiotherapy and bisphosphonate treatment. The patients were prospectively randomized into two treatment groups: group A (n = 20) received the first zoledronate administration after and group B (n = 20) prior to the radiotherapy. Urine samples were collected from each patient on the first day, in the middle, and on the last day of the radiation therapy. Measurement of the bone metabolites hydroxylysylpyridinoline and lysylpyridinoline was performed by high-performance liquid chromatography. Statistical analysis was performed using the Mann-Whitney U test.
The hydroxylysylpyridinoline and lysylpyridinoline excretion decreased significantly in the combined bisphosphonate and radiotherapy group (p = 0.02, p = 0.08). No significant change of the hydroxylysylpyridinoline and lysylpyridinoline excretion was determined in the patients that received solely irradiation.
The results indicate the ability of zoledronate to prevent the early radiation-induced bone collagen degradation suggesting that the radiation-induced bone loss is mainly caused by osteoclastic bone resorption rather than by a direct radiation-induced damage.
No preview · Article · Nov 2014 · Clinical and Translational Oncology
[Show abstract][Hide abstract] ABSTRACT: A variety of different growth factors, most notably bone morphogenetic proteins (BMPs), have been shown to stimulate the osteogenic differentiation of mesenchymal stromal cells (MSCs) in vitro. Yet, due to the lack of comparative studies it remains unclear which protocol is the most effective in the induction of osteogenesis in MSC cultures. The aim of this study was to compare the most potent growth factors in regard to their osteoinductive potential.Human MSCs were cultured for 10 days in the presence of BMP-2, BMP-6, BMP-9 + IGF-2 and BMP-2, -6, -9 (day 1 + 2: 50 ng/ml; days 3–6: 100 ng/ml; days 7–10: 200 ng/ml). The formation of the osteoblast phenotype was assessed by quantification of osteoblast-related marker genes using reverse transcription polymerase chain reaction (RT-PCR) and alkaline phosphatase (ALP) staining. Matrix mineralization was assessed by alizarin red S and von Kossa staining. Statistical analysis was carried out using the one-way analysis of variance (ANOVA) followed by Scheffe's post hoc procedure.Among the tested growth factors the combination of BMP-2 + BMP-6 + BMP-9 most effectively induced the upregulation of collagen type I, collagen type V, osteocalcin, alkaline phosphatase, RUNX2, BMP-2, osteonectin and DLX5 (p < 0.01) and resulted in a consistent matrix mineralization.The findings suggest the combined addition of BMP-2, BMP-6 and BMP-9 to the osteoinductive culture medium containing dexamethasone, β-glycerophosphate and ascorbate-2-phosphate produces more potent osteoblast differentiation of human MSCs in vitro.
No preview · Article · Nov 2014 · Journal of Cranio-Maxillofacial Surgery
[Show abstract][Hide abstract] ABSTRACT: AIM. This study aimed to clarify the relation between the angulation of the curved osteotome and fracture of the pterygoid plate during Le Fort I osteotomy. MATERIAL AND METHODS. Twenty-one specimens of hemisectioned Turkish skulls were used for the study. The maxilla was sectioned transversely on the floor of the pyriform aperture and posteriorly to the lateral pterygoid plate with a mechanical saw. The pterygomaxillary junction was separated with a curved osteotome by angulating the osteotome with, 0° and -30° to the occlusal plane. The undesired fractures of the lateral pterygoid plate were determined. Among 21 specimens, 7 pterygomaxillary junctions were separated with an angle of +30° , 7 with 0° and 7 with -30° to the occlusal plane. RESULTS. In group +30°, the undesired fracture occured in 6 of the cases. In group -30°, the undesired fracture was determines in one case. In cases where the separation was performed by placing the osteotome paralell to the occlusal plane all plates remained safe. CONCLUSION. Within the limited knowledge of the current study it can be concluded that the osteotome should be placed paralell to the occlusal plane.
Full-text · Article · Sep 2014 · Stomatologija / issued by public institution "Odontologijos studija" ... [et al.]
[Show abstract][Hide abstract] ABSTRACT: Adipose-derived stromal cells (ASCs) are mostly isolated by enzymatic digestion, centrifugation and adherent growth resulting in a very heterogeneous cell population. Therefore, other cell types in the cell culture can comprise the differentiation and proliferation potential of the ASC population. Recent studies indicated that an antibody-aided isolation of distinct ASC subpopulations provides advantages over the conventional method of ASC isolation. The aim of this study was to investigate the adipogenic differentiation potential of CD29-, CD71-, CD73- and CD90-selected ASCs in vitro.
No preview · Article · May 2014 · Journal of Plastic Reconstructive & Aesthetic Surgery
[Show abstract][Hide abstract] ABSTRACT: The hypothesis of the present study was that topically applied bisphosphonate (BP) on a collagen membrane or alternatively bovine bone mineral prevents surface resorption of onlay bone grafts.
In eight adult pigs, bone blocks were harvested bilaterally from the mandible and fixed to the lateral cortex of the horizontal ramus to simulate a ridge augmentation. In a split-mouth study design, we used alendronate in aqueous solution (1 mg/ml) on the test-side in three different ways: on a collagen membrane (Bio-Gide®), soaked in bovine bone mineral granules (Bio-Oss®), or applied to the bone graft directly. The same materials without BP were used as controls on the contralateral side. After 3 months, the animals were sacrificed. The evaluation included sequential fluorochromic labeling and measurement of bone height in microradiography and toluidine blue staining.
In five cases, necrosis of the overlying periosteal tissues with BP was observed macroscopically. A statistically significantly lower loss in graft height was seen on the test-side for Bio-Gide® + alendronate (0.65 %) versus Bio-Gide® (1.52 %), p = 0.002; Bio-Oss® + alendronate (1.16 %) versus Bio-Oss® (4.20 %), p = 0.001; and bone graft + alendronate (1.25 %) versus bone graft alone (6.01 %), p = 0.006. An inhibitory effect on bone remodeling was observed by a statistically significantly lower number of resorption lacunae.
The hypothesis was accepted that a bisphosphonate-treated membrane reduced bone graft resorption; however, periosteal necrosis requires better adaptation of the dosage.
A bisphosphonate membrane could be a helpful tool to preserve augmentation height of onlay bone grafts.
No preview · Article · Feb 2014 · Clinical Oral Investigations
[Show abstract][Hide abstract] ABSTRACT: Adipose derived stromal cells (ASC’s) are mostly isolated by enzymatic digestion, centrifugation and adherent growth resulting in a very heterogeneous cell population. Therefore, other cell types in the cell culture can comprise the differentiation and proliferation potential of the ASC population. Recent studies indicated that an antibody-aided isolation of distinct ASC subpopulations provides advantages over the conventional method of ASC isolation. The aim of this study was to investigate the adipogenic differentiation potential of CD29-, CD71-, CD73- and CD90-selected ASC’s in vitro.
The stromal vascular fraction (SVF) was obtained from rat adipose tissue by enzymatic digestion and centrifugation. Subsequently, CD29+-, CD71+-, CD73+- and CD90+ cells were isolated by magnetic activated cell sorting (MACS), seeded into culture plates and differentiated into the adipogenic lineage. ASC’s isolated by adherent growth only served as controls. Adipogenic differentiation was assessed by Oil Red O staining and quantification of the Adiponectin and Leptin concentrations in the cell culture supernatants. Statistical analysis was carried out using one-way analysis of variance (ANOVA) followed by the Scheffe’s post hoc procedure.
The results showed that different subpopulations with different adipogenic differentiation potentials can be isolated by the MACS procedure. The highest adipogenic differentiation potential was determined in the CD29-selected ASC population followed by the unsorted ASC population. The CD71-, CD73- and CD90- selected cells exhibited significantly the lowest adipogenic differentiation potential.
In conclusion, the CD29-selected ASC’s and the unsorted ASC’s exhibited a similar adipogenic differentiation potential. Therefore, we don’t see a clear advantage in the application of an anti-CD29-based isolation of ASC’s over the conventional technique using adherent growth. However, the research on isolation / purification methods of adipogenic ASC’s should continue in order to make this stem cell source even more attractive for future adipose tissue engineering applications.
No preview · Article · Jan 2014 · Journal of Plastic Reconstructive & Aesthetic Surgery
[Show abstract][Hide abstract] ABSTRACT: Objectives
Injectable or implantable scaffolds seeded with autologous chondrogenic cells may represent a promising option for treatment of cartilage defects in the future. Current problems with the autologous chondrocyte implantation including dedifferentiation and the development of fibrocartilage suggest the use of alternative chondrogenic cell sources such as mesenchymal stromal cells (MSCs). The aim of this study was to compare the early effects of different scaffolds on the proliferation and metabolic activity of chondrogenic MSCs in vitro.
Materials and methods
Multipotent stromal cells were isolated from rat bone marrow, phenotyped by flow cytometry, and differentiated into distinct lineages proved by lineage-specific staining and gene expression (RT-PCR) pattern. Cell proliferation on Tutodent® Membrane, Bio-Gide®, TissuFleece E, and Belotero® Soft was quantified by the MTT and WST-1 assay and direct determination of total cell numbers. Potential cytotoxic effects of eluates obtained from the materials were quantified by lactate dehydrogenase (LDH) and 5-bromo-2-deoxyuridine (BrdU) assay.
TissuFleece E displayed the best results regarding cell proliferation on the biomaterials and metabolic activity (MTT, WST-1) (p < 0.001). Yet, the eluates of TissuFleece E caused an increased LDH release and lower values in the BrdU test. Cell proliferations on Bio-Gide®, Tutodent® Membrane, and Belotero® Soft were similar to the control. The eluates of Belotero® Soft exhibited the highest LDH release and lowest values in the BrdU assay (p < 0.05).
Our results support the use of Tissufleece E as scaffold for chondrogenic rat MSCs. However, it should be prewashed with culture medium before seeding of the cells.
Tissufleece E may serve as a promising carrier material for chondrogenic MSCs for cartilage tissue engineering attempts.
Full-text · Article · Mar 2013 · Clinical Oral Investigations
[Show abstract][Hide abstract] ABSTRACT: Background
Adipose tissue derived stroma cells (ASC’s) offer for many advantages for tissue engineering strategies over mesenchymal stroma cells from other sources and ideal carrier materials have to be identified for them. The aim of this study was to demonstrate and to compare the effects of three clinically established biomaterials on proliferation and metabolic activity of rat ASC’s in vitro.
Materials and Methods
Rat adipose tissue derived stroma cells (ASC’s) were isolated and differentiated into distinct lineages proved by lineage specific staining and gene expression analysis (RT-PCR). The biomaterials Bio-Gide®, Tutodent® Membrane and Belotero® Soft were tested with rat ASC’s for their biocompatibility using scanning electron microscopy (SEM), cell vitality staining, cytotoxicity and proliferation tests (LDH, MTT, BrdU, WST-1).
The collagen membrane Bio-Gide® resulted in a significantly higher viability and proliferation (WST-1, BrdU) compared to Tutodent® Membrane. No significant difference was determined in the LDH and MTT test. The hyaluronic acid gel Belotero® Soft showed no cytoxicity (LDH, FDA/PI) and had no negative effects on metabolic activity (WST-1, MTT) or cell proliferation (BrdU) of ASC’s.
Our results indicate Bio-Gide® and Belotero® Soft as preferable carrier materials for ASC’s. For the further establishment of ASC’s-based treatment strategies, in vivo investigations on the tissue regeneration potential of these cell-biomaterial scaffolds should follow.
No preview · Article · Jan 2013 · Journal of Cranio-Maxillofacial Surgery
[Show abstract][Hide abstract] ABSTRACT: INTRODUCTION: Barrier membranes, both absorbable and non-absorbable, have been used in sinus augmentation for many years. Some years ago, a new autologous blood substrate called Platelet-Rich-Fibrin (PRF) was introduced, and to date, the supporting effect on bone regeneration has been controversial. This study aimed to evaluate the effect of PRF on bone regeneration when used as a barrier membrane at the lateral osteotomy site in sinus augmentation. MATERIAL AND METHODS: Twelve sinuses from six patients requiring bilateral sinus floor augmentation were treated with a two-stage surgical technique using sinus augmentation and implant placement after 5 months. The sinuses were grafted with autologous bone and bone-substitute material (Bio-Oss(®)) mixed in a 1:1 ratio and were covered in a randomized split-mouth design with a PRF or a conventional collagen membrane (Bio-Gide(®)), respectively. Five months later threaded titanium dental implants were inserted and bone specimens harvested with a trephine burr were evaluated histomorphometrically. RESULTS: Bone quality seemed to be equal at both sites of the grafted sinuses. Mean vital bone formation after 5 months was 17.0% and 17.2%, for the PRF and collagen sites, respectively. The mean of residual bone-substitute was 15.9% and 17.3% for PRF and collagen, respectively. No local complications, such as dehiscences or membrane exposures, were detected at either site in any of the treated patients. After 12 months all implants reached primary stability in the augmented maxillary sinus floor without any peri-implant tissue inflammation. CONCLUSIONS: Within the limits of the study the coverage of the lateral sinus window with two different absorbable membranes has been shown to result in a similar amount of vital bone formation and residual bone-substitute.
No preview · Article · Dec 2012 · Journal of cranio-maxillo-facial surgery: official publication of the European Association for Cranio-Maxillo-Facial Surgery
[Show abstract][Hide abstract] ABSTRACT: Objectives
The augmentation of the alveolar ridge using iliac cortico-spongeous bone grafts is routinely used. However, bone grafts show a substantial degree of resorption, which may negatively affect the long-term success of dental implants in the augmented area. The aim of this study was to evaluate the effect of a deproteinized bovine bone matrix coverage on the resorption of iliac bone grafts. Material and methodsTwo cohorts consisting of 40 patients who received a vertical augmentation of the alveolar ridge with onlay grafts from the iliac crest were prospectively investigated over a period of 2 years. In half of the patients (n=40), the grafts were covered by a thin layer of deproteinized bovine bone matrix (DBBM cohort). The other 40 patients received the identical surgical procedure without a DBBM coverage (non-DBBM cohort). The graft height/resorption was radiographically determined immediately after surgery, 6 months, 1 year, and 2 years postoperatively. ResultsThe height of the bone graft 6months after surgery accounted 92.15% of the initial value in the DBBM cohort and 87.76% in the non-DBBM cohort. One year after augmentation, the height reduced to 83.95% in the DBBM cohort and 72.92% in the non-DBBM cohort. Two years after surgery, the resorption slowed down and the height of the grafts accounted 81.27% in the DBBM cohort and 71.43% in the non-DBBM cohort. The difference was statistically significant. Conclusion
Deproteinized bovine bone matrix reduces the postoperative resorption of iliac bone block grafts and may therefore enhance the long-term implant prognosis in the augmented area.
No preview · Article · Nov 2012 · Clinical Oral Implants Research
[Show abstract][Hide abstract] ABSTRACT: Radiotherapy can lead to a reduction of bone density with an increased risk of pathological fractures. Bisphosphonates may represent a preventive treatment option by increasing the density of anorganic bone mineral. Yet it is unknown how bisphosphonates act on irradiated collagen cross-links, which play an essential role for the mechanical stability of bone. The aim of this study was to evaluate the effects of zoledronate on bone collagens and their cross-links after irradiation. The right femur of 37 rats was irradiated with a single dose of 9.5 Gy at a high dose rate using an afterloading machine. Half of the rats (n = 18) received additionally a single dose zoledronate (0.1 mg/kg body weight). Fourteen and 100 days after irradiation the femora were collected for histologic evaluation and determination of the collagen cross-links lysylpyridinoline, hydroxylysylpyridinoline, and hydroxyproline. The collagen types were characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Fourteen days after treatment the lysylpyridinoline levels of all treatment groups were significantly lower compared to the untreated control. After 100 days, in the combined radiotherapy + zoledronate group significantly lower lysylpyridinoline values were determined (p = 0.009). Radiotherapy and/or zoledronate did not change significantly the level of hydroxylysylpyridinoline. The concentration of hydroxyproline was 14 days after irradiation significantly higher in the combined treatment group compared to the control. No significant differences were observed 100 days after treatment. Zoledronate does not have the ability to restore the physiological bone collagen cross-link levels after radiotherapy. However, this would be necessary for regaining the physiological mechanical stability of bone after irradiation and therefore to prevent effectively radiation-induced fractures.
No preview · Article · Nov 2012 · Calcified Tissue International
[Show abstract][Hide abstract] ABSTRACT: The ideal treatment of the nasolabial fold, the tear trough, the labiomandibular fold and the mentolabial sulcus is still discussed controversially. The detailed topographical anatomy of the fat compartments may clarify the anatomy of facial folds and may offer valuable information for choosing the adequate treatment modality. Nine non-fixed cadaver heads in the age range between 72 and 89 years (five female and four male) were investigated. Computed tomographic scans were performed after injection of a radiographic contrast medium directly into the fat compartments surrounding prominent facial folds. The data were analysed after multiplanar image reconstruction. The fat compartments surrounding the facial folds could be defined in each subject. Different arrangement patterns of the fat compartments around the facial rhytides were found. The nasolabial fold, the tear trough and the labiomandibular fold represent an anatomical border between adjacent fat compartments. By contrast, the glabellar fold and the labiomental sulcus have no direct relation to the boundaries of facial fat. Deep fat, underlying a facial rhytide, was identified underneath the nasolabial crease and the labiomental sulcus. In conclusion, an improvement by a compartment-specific volume augmentation of the nasolabial fold, the tear trough and the labiomandibular fold is limited by existing boundaries that extend into the skin. In the area of the nasolabial fold and the mentolabial sulcus, deep fat exists which can be used for augmentation and subsequent elevation of the folds. The treatment of the tear trough deformity appears anatomically the most challenging area since the superficial and deep fat compartments are separated by an osseo-cutaneous barrier, the orbicularis retaining ligament. In severe cases, a surgical treatment should be considered. By contrast, the glabellar fold shows the most simple anatomical architecture. The fold lies above one subcutaneous fat compartment that can be used for augmentation.
No preview · Article · May 2012 · Journal of Plastic Reconstructive & Aesthetic Surgery
[Show abstract][Hide abstract] ABSTRACT: Many different materials are proposed for reconstruction of traumatic orbital floor defects. Donor-site morbidity of autologous transplants and infections or extrusions of nonresorbable implants lead to a widespread use of resorbable, alloplastic materials such as polydioxanone (PDS). The goal of this study was to evaluate the prevalence of orbital floor fracture-related problems after surgical treatment using PDS. Ophthalmologic and clinical examinations were performed at 194 patients before orbital floor reconstruction, 14 days and 6 months after surgery (approximate defect sizes: <1 cm², n=50; 1-2 cm², n=97; >2 cm², n=47). Clinical findings including the ocular motility, the sensibility of the infraorbital nerve, and the position of the globe were evaluated. For statistical analysis of categorical data, confidence intervals of percentages were determined. Linear relationships between 2 variables were assessed with Pearson correlation analysis. A reduced ocular motility was diagnosed in 60 patients (31%) before surgery; in 14 patients (7%), 2 weeks; and in 10 patients (5%), 6 months after surgery. Infraorbital hypesthesia was found in 120 patients (62%) before surgery; in 47 patients (24%), 2 weeks; and in 35 patients (18%), 6 months after surgery. An enophthalmos was present in 10 patients (5%) before surgery, and in 4 patients (2%), 6 months after surgery. Our data suggest that PDS is a suitable implant for orbital floor reconstruction with acceptable low rates of infraorbital hypesthesia, bulbus motility disturbances, and enophthalmos. Polydioxanone can also be used for orbital floor defects exceeding 2 cm².
No preview · Article · Jan 2012 · The Journal of craniofacial surgery
[Show abstract][Hide abstract] ABSTRACT: The restoration of a natural volume distribution is a major goal in facial rejuvenation. The aims of this study were to establish a radiographic method enabling effective measurements of the midfacial fat compartments and to compare the anatomy between human cadavers of younger versus older age.
Data from computed tomographic scans of 12 nonfixed cadaver heads, divided into two age groups (group 1, 54 to 75 years, n = 6; and group 2, 75 to 104 years, n = 6), were analyzed. For evaluation of the volume distribution within a specific compartment, the sagittal diameter of the upper, middle, and lower thirds of each compartment was determined. For evaluation of a "sagging" of the compartments, the distance between the cephalad border and the infraorbital rim was determined.
Computed tomography enables a reproducible depiction of the facial fat compartments and reveals aging changes. The distance between the fat compartments and the infraorbital rim was higher in group 2 compared with group 1. The sagittal diameter of the lower third of the compartments was higher, and the sagittal diameter of the upper third was smaller in group 2 compared with group 1. The buccal extension of the buccal fat pad was shown to be an independent, separate compartment.
This study demonstrates an inferior migration of the midfacial fat compartments and an inferior volume shift within the compartments during aging. Additional distinct compartment-specific changes (e.g., volume loss of the deep medial cheek fat and buccal extension of the buccal fat pad) contribute to the appearance of the aged face.
No preview · Article · Sep 2011 · Plastic and Reconstructive Surgery