Facial cutaneous squamous cell carcinoma - microscopic safety margins and their impact on developing local recurrences.

OBJECTIVES: Surgical excision remains the treatment of choice for facial cutaneous squamous cell carcinoma (cSCC) despite there being no generally accepted diameter of clear margins. Therefore, the aim of this study was to evaluate the impact of microscopic clear margins diameter (mCMD) with respect to the development of local recurrences (LR). MATERIALS AND METHODS: The medical records of 99 patients with a total of 142 cases of facial cSCC, who underwent surgical treatment between January 2010 and December 2015, were reviewed for demographic data and clinicopathological features. RESULTS: 100 cases were diagnosed as primary cSCC and 42 cases as secondary cSCC. Of these, nine (6.3%) developed LR. Mean time to LR was 20 months, with the cheek as the predominant site 55.5% (n = 5). Wound closure was either primary (56%) or secondary (44%), depending on the site. Although no significant correlation between mCMD and LR was found (rPearson = 0.029; rPearson = 0.015), >4.1 mm was shown to be a negative cut-off-value (horizontally and vertically) without LR (100% vs 0%). CONCLUSIONS: Based on these results, however descriptive they are, the authors consider histological confirmation of clear margins to be necessary for reducing the formation of LR. Thus, consistent testing and histopathological reporting, in a multicentered effort, are needed to further clarify the role of mCMD in the development of cSCC-LR.

Is there an ideal way to close the donor site of radial forearm free flaps?

Radial forearm free flaps (RFFF) are the "workhorse" of reconstructive head and neck surgery, but have considerable morbidity at the donor site. The aim of this study was to review current publications about the incidence and type of morbidity and the different techniques used for closure of the site. We screened the MEDLINE database to find relevant papers using the terms "RFFF head and neck" and "RFFF donor site". Abstracts were filtered, and the full texts studied carefully. We found 1056 publications during the period 1982-2017 of which 389 were studied in full, and 39 studies were finally included in the review. We found four main methods of closure of the donor site: full-thickness skin grafts (FTSG); split-thickness skin grafts (STSG); modified techniques for raising the flap and closure of the wound by local flaps; and others (such as allografts, expanders, and vacuum bandages). For STSG and FTSG the preparation of the donor site seems to be a relevant factor. Special attention should be paid to the coverage of the flexor tendons. FTSG give better aesthetic results than STSG. Closure by local flaps may achieve primary closure of the donor site without a third surgical site, but the techniques are limited by the amount of tissue required at the site of the defect. The most common side effects are disorders of wound healing such as exposed tendons. To avoid exposure of the tendons, flexor tendons should be covered with muscle bellies when STSG are used. It is still not clear whether many other reported side effects (such as impairment of sensitivity) are induced by raising the flap or closing the donor site. There is an argument for closure of individual donor sites independently, but there is no one method of closure for all donor sites, because each has its specific disadvantages and complications.

The influence of geranylgeraniol on microvessel sprouting after bisphosphonate substitution in an in vitro 3D-angiogenesis assay.

OBJECTIVES: Recent studies focused on angiogenesis in the pathophysiology of bisphosphonate-associated osteonecrosis of the jaws (BP-ONJ) and identified geranylgeraniol (GGOH) as a feasible option for BP-ONJ therapy. This study investigated the influence of GGOH on microvessel sprouting after BP-incubation in vitro. MATERIALS AND METHODS: Ten experimental set-ups were randomly designed in an in vitro 3D-angiogenesis assay. Two groups included HUVEC cell spheroids with and without (±) GGOH substitution as controls and eight groups pairwise contained either clodronate or the nitrogen-containing bisphosphonates (N-BP) ibandronate, pamidronate, and zoledronate ± GGOH. The size of the cell spheroids including the outbranching sprouts (SpS) as well as the density (SpD) and length of the sprouts (SpL) were analyzed by a grid system after 0, 24, 48, and 72 h. RESULTS: For controls and NN-BP clodronate, no significant differences at any tested parameter and any point of measurement could be detected within the experimental set-ups ± GGOH (p each ≥0.05). For N-BP ibandronate, the experimental set-ups +GGOH showed a significantly increased SpS, SpD, and SpL after 48 and 72 h (p each ≤0.002) compared to the experimental set-ups -GGOH. For N-BPs pamidronate and zoledronate, the experimental set-ups + GGOH demonstrated a significantly increased SpS, SpD, and SpL after 24, 48, and 72 h (p each ≤0.001) compared to the experimental set-ups -GGOH. CONCLUSIONS: The strong negative influence of N-BPs on microvessel sprouting could be significantly reversed by GGOH. CLINICAL RELEVANCE: Since supportive therapeutic options for BP-ONJ are lacking, GGOH might be a promising substitute for BP-ONJ prevention and therapy.

Influence of mechanical compression on human periodontal ligament fibroblasts and osteoblasts.

OBJECTIVES: The aim of this study was to investigate and compare the changes in human periodontal ligament fibroblasts (HPdLFs) and osteoblasts (HOBs) after the application of compressive force (CF) at two different strengths in vitro. MATERIALS AND METHODS: HPdLF and HOB were exposed to CF with various strengths (5 and 10 %) using a Flexercell Compression Unit for 12 h in vitro. Viability was detected via 3-(4.5-dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide (MTT) and apoptosis rate by transferase dUTP nick end labeling (TUNEL) assay. The gene expression of alkaline phosphatase (ALP), osteocalcin (OCN), osteoprotegerin (OPG), and receptor activator of NF-κB ligand (RANKL) was analyzed using reverse transcriptase polymerase chain reaction (RT-PCR). Osteopontin (OPN), matrix metalloproteinase-8 (MMP-8), and tissue inhibition of metalloproteinase-1 (TIMP-1) were quantified by an ELISA. RESULTS: Ten percent CF decreased viability, particularly in HOBs, but did not induce increased apoptosis. ALP gene expression increased the most after 5 % CF in HPdLFs and after 10 % CF in HOB. OCN was not affected by CF in either cell line. The highest RANKL/OPG ratio was measured after 5 % CF in both cell lines. OPN was upregulated in HOB by 5 %. HPdLFs showed an upregulation of MMP-8-synthesis and an increased MMP-8/TIMP-1 ratio. CONCLUSIONS: HOBs have a greater effect on bone remodeling through the upregulation of OPN, whereas HPdLFs facilitate orthodontic tooth movement by influencing the extracellular matrix via the MMP-8/TIMP-1 ratio. CLINICAL RELEVANCE: High CF in orthodontics should be avoided to prevent tissue damage, whereas moderate CF enables active tissue remodeling and tooth movement.

Influence of CAD/CAM zirconia for implant-abutment manufacturing on gingival fibroblasts and oral keratinocytes.

OBJECTIVES: This study investigated the influence of three CAD/CAM zirconia ceramics for implant-abutment manufacturing on cell viability, migration ability, and cytotoxicity of human gingival fibroblasts (HGF) and oral keratinocytes (HOK) in vitro. MATERIALS AND METHODS: HGF and HOK were cultured on zirconia ceramic disks (VITA In-Ceram YZ, Ivoclar IPS e.max ZirCAD, Sirona inCoris ZI) and on control disks made of tissue culture polystyrene. Cell viability was analyzed by a MTT assay. Migration ability was detected by a scratch assay. A ToxiLight assay was used to analyze the influence of the tested zirconia ceramics on adenylate kinase (ADK) release and cytotoxicity. RESULTS: At MTT assay, HGF showed an increased cell viability compared to the control after 9 and 12 days for all ceramics (p each ≤0.0002) while HOK demonstrated a decreased cell viability after 9 and 12 days for all ceramics (p each ≤0.0003). At scratch assay, HGF exhibited for all ceramics decreased relative distances of the scratch wound compared to the control from 24 to 48 h (p each <0.0001) with exception of VITA In-Ceram YZ after 48 h. HOK showed increased distances compared to the control for all ceramics after 48 h (p each <0.0001). At ToxiLight assay, a minimal cytotoxicity of the tested materials could be detected. CONCLUSIONS: Overall, significant influences of the investigated CAD/CAM zirconia ceramics on HGF and HOK could be shown. CLINICAL RELEVANCE: The analyzed zirconia ceramics could influence oral soft-tissue cells that might affect the esthetic outcome after implant placement using CAD/CAM zirconia abutments.

Interdevice agreement of eight equivalent dental color measurement devices.

OBJECTIVES: The reproducibility of electronic color-measuring devices used in dentistry refers to the consistency of measurements of a single device and also that between identical devices. This in vitro study evaluated the short-term interdevice agreement of eight equivalent dental measurement devices. METHODS: The short-term repeatability of eight identical VITA Easyshade Advance color measurement devices was evaluated. Each device was used to quantify the Commission Internationale de l'Eclairage (CIE) L*a*b* color coordinates of ceramic disks, designed in accordance with guidelines set by the VITA Toothguide 3D-Master shade tab organization. Statistical analysis involved calculating the intraclass correlation coefficients (ICCs) and color differences (ΔE*) for corresponding measurements taken by the devices. In addition, the mean color difference from the mean (MCDM) was calculated for each color. RESULTS: All devices showed high ICCs for the color coordinates (L = 0.996, a = 0.993, b = 0.999). The ΔE* by shade ranged from 0.62 to 1.67. The MCDM by color-measuring device ranged from 0.11 to 0.23. CONCLUSION: This in vitro study showed that the color-measuring devices used were suitable for determining shade with high repeatability. CLINICAL RELEVANCE: The high interdevice agreement of the VITA Easyshade Advance means that dental professionals can use this device independently, without compromising the consistency and quality of color measurements.

Isoprenoid geranylgeraniol: the influence on cell characteristics of endothelial progenitor cells after bisphosphonate therapy in vitro.

OBJECTIVES: Geranylgeraniol (GGOH) has been reported as a potential treatment option for bisphosphonate-associated osteonecrosis of the jaws (BP-ONJ). The aim of this study was to analyze the effects of GGOH on endothelial progenitor cells (EPC) after bisphosphonate treatment in vitro. MATERIALS AND METHODS: EPC were incubated with different nitrogen (N-BPs: ibandronate, pamidronate, zoledronate) and a non-nitrogen-containing bisphosphonates (NN-BP: clodronate) with and without GGOH. Cell viability was measured by MTT and PrestoBlue assay. Migration ability was analyzed with a Boyden and Scratch assay. Apoptosis rates were determined by colony-forming, Tunel and ToxiLight assays. RESULTS: Negative effects of N-BPs on EPC were shown in all tests without GGOH. The substitution of GGOH demonstrated significantly increased cell viability (MTT: p each N-BP ≤0.004; PrestoBlue: p each N-BP <0.001) and migration ability (Boyden: p each N-BP <0.001; Scratch: p each N-BP <0.001). Concerning the apoptosis rates, increased EPC colony densities (p each N-BP ≤0.009), decreased numbers of apoptotic cells in the Tunel assay (p each N-BP <0.001), and a decreased adenylate kinase release in the ToxiLight assay (p each N-BP ≤0.03) were observed. For the clodronate-treated cells, no significant differences could be detected with or without GGOH in any assay (p each N-BP/NN-BP >0.05). CONCLUSIONS: GGOH cell treatment reversed the negative effects of bisphosphonates on EPC. CLINICAL RELEVANCE: These findings support the hypothesis that systemic or local GGOH treatment might lead to new therapeutic strategies for BP-ONJ.

Synchrotron-based X-ray tomographic microscopy for visualization of three-dimensional collagen matrices.

INTRODUCTION: Three-dimensional collagen matrices (3D-CMs) may be visualized by cumbersome reconstructions of serial sections. We report here on the method of synchrotron-based X-ray tomographic microscopy (SRXTM) to image 3D-CMs in native tissue probes. MATERIAL AND METHODS: SRXTM of 3D-CMs (mucoderm®, mucograft®) was performed at the TOMCAT beamline of the Swiss Light Source (SLS) at the Paul Scherrer Institute (Villigen, Switzerland). RESULTS: SRXTM combines the advantages of high-resolution scanning electron microscopy (SEM) imaging with the low-resolution reconstructions of micro-CT (µCT) imaging. It may be used to non-destructively visualize and analyze structures within the 3D-CMs without the need of serial sectioning and reconstruction. CONCLUSION: High-resolution SRXTM is a useful tool in analyzing the topology and morphometry of structures in 3D-CMs. The outcome justifies the efforts in sophisticated data processing. CLINICAL RELEVANCE: SRXTM may help to understand the clinical characteristics of 3D-CMs in more detail.

The prevalence of human papilloma virus (HPV) infections in oral squamous cell carcinomas: a retrospective analysis of 88 patients and literature overview.

In addition to tobacco and alcohol consumption, the two main risk factors for oral squamous cell carcinoma (OSCC), recent studies have revealed infections with human papilloma virus (HPV) as an additional risk factor for OSCC development. In the field of head and neck malignancies, the prevalence of HPV infections in oropharyngeal cancer (OC) ranges in different studies up to 84%. While HPV infection is discussed as an independent risk factor in this region, its distinguished role in carcinogenesis of tumours localized to the oral cavity remains still uncertain. In this study, we analysed the HPV status in 88 consecutive patients with OSCCs localized anterior of the palatoglossal arch who were treated in the Department of Oral and Maxillofacial Surgery at the University Medical Center Mainz. The HPV status analysis was performed using DNA-PCR and immunostaining of p16 protein. The prevalence of HPV-positive OSCCs was about 6% (5 patients). In 3 patients the HPV subtypes 16/18 were found. No significant differences between the HPV positive and negative patients regarding age, gender, smoking and alcohol consumption, localization and TNM level could be detected. Contrary to other studies focussing on cancers of the lingual and palatine tonsil, the prevalence of HPV infections was much lower in the oral cavity. Therefore HPV infection might play a less important role in oral carcinogenesis.

Influence of CAD/CAM all-ceramic materials on cell viability, migration ability and adenylate kinase release of human gingival fibroblasts and oral keratinocytes.

OBJECTIVES: The aim of this study was to analyze the influence of four CAD/CAM all-ceramic materials on cell viability, migration ability and adenylate kinase (ADK) release of human gingival fibroblasts (HGF) and oral keratinocytes (HOK). MATERIALS AND METHODS: HGF and HOK were cultured on disc-shaped CAD/CAM all-ceramic materials (e.max CAD LT, e.max CAD HT, Empress CAD and Mark II) and on discs made of tissue culture polystyrene surface (TCPS) serving as control. Cell viability was analyzed by using an MTT assay, and migration ability was investigated by a scratch assay. A ToxiLight assay has been performed to analyze the effect of all-ceramic materials on ADK release and cell apoptosis. RESULTS: At MTT assay for HGF, no significant decrease of cell viability could be detected at all points of measurement (p each > 0.05), while HOK demonstrated a significant decrease in cell viability especially on Empress CAD and Mark II at each point of measurement (p each < 0.001). Scratch assay demonstrated an increased migration ability for HGF on e.max CAD HT, Empress CAD and Mark II (p each < 0.001), whereas HOK showed a significantly decreased migration ability on all tested materials at all points of measurement (between -36 % and -71 %; p each < 0.001). At ToxiLight assay, only small cytotoxic effects of the all-ceramic materials could be investigated. CONCLUSIONS: This study disclosed significant differences in cell viability and migration ability of HGF and HOK on CAD/CAM all-ceramic materials. CLINICAL RELEVANCE: CAD/CAM all-ceramic materials can influence oral cell lines responsible for soft tissue creation which may affect the esthetic outcome.

In vitro and in vivo characterization of porcine acellular dermal matrix for gingival augmentation procedures.

BACKGROUND AND OBJECTIVE: Recently, porcine acellular dermal matrix (PADM) has been proposed as a possible alternative to autogenous grafts in periodontal plastic surgery. The aim of the present study was to investigate the in vitro responses of four different oral cell lines cultured on a novel PADM. Furthermore, tissue reaction to PADM was evaluated histologically after subcutaneous implantation in mice. MATERIAL AND METHODS: Human gingival fibroblasts (HGF), human osteoblast-like cells, human umbilical vein endothelial cells and human oral keratinocytes (HOK) were cultured and transferred on to the PADM. A tissue culture polystyrene surface served as the control. The viability of all tested cell lines on PADM was measured by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide colorimetric assay and PrestoBlue(®) reagent. The ToxiLight(®) assay was performed to analyze the effect of PADM on adenylate kinase release. PADM was implanted into nude mice subcutaneously and subjected to histological analysis after 21 d. RESULTS: Using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide colorimetric assays, all tested cell lines cultured on PADM demonstrated a significant increase of viability compared to the control group (each p < 0.001) with the exception of HGF and HOK after 3 d (each p > 0.05). According to the PrestoBlue(®) analysis, all cell lines demonstrated a significant increase of viability compared to the control group at the particular points of measurement after 18 h (HGF p < 0.01; human osteoblast-like cells, human umbilical vein endothelial cells, HOK each p < 0.001). No significant cytotoxic effects of PADM on the tested cell lines could be observed, as assessed by changes in adenylate kinase release. Subcutaneous implantation of PADM into nude mice demonstrated good integration with surrounding tissues and significant revascularization of its collagen structure. CONCLUSION: Overall, the results suggest that PADM is a promising substitute for autogenous soft tissue grafts in periodontal surgery.

Bisphosphonates' antiangiogenic potency in the development of bisphosphonate-associated osteonecrosis of the jaws: influence on microvessel sprouting in an in vivo 3D Matrigel assay.

OBJECTIVES: Bisphosphonate-associated osteonecrosis of the jaw (BP-ONJ) is an adverse side effect of long-term bisphosphonate treatment. One theory of BP-ONJ etiology suggests a negative influence of these agents on angiogenesis and vascularization. This in vivo study analyzed the effects of bisphosphonates on angiogenesis in a 3D Matrigel assay. MATERIALS AND METHODS: Matrigel plugs were implanted into fifty 6-8-week-old female nude mice. Ten animals each were treated either with clodronate, ibandronate, pamidronate, zoledronate, or carrier solution as controls. The microvessel density (MVD), microvessel area (MVA), and microvessel size (MVS) in Matrigel plugs were analyzed after 21 days of treatment by immunohistochemistry and exemplary 3D microvascular corrosion castings. RESULTS: All bisphosphonates induced a statistically significant decrease of MVD (p each <0.001), whereby the nitrogen-containing bisphosphonates (N-BPs) demonstrated a clearly stronger effect than non-nitrogen-containing bisphosphonates (NN-BP) clodronate (control 166, clodronate 99, ibandronate 48, pamidronate 47, zoledronate 35 microvessels/mm(2)). Referring to MVA, similar results could be detected. MVS was significantly increased especially by ibandronate (103 %) compared to control group (p < 0.001). Scanning electron microscope scans of the corrosion castings confirmed these results. CONCLUSIONS: The stronger influence on MVD by N-BPs compared to the NN-BP clodronate may explain for the lack of BP-ONJ after treatment with NN-BPs. CLINICAL RELEVANCE: Ibandronate induced a strong increase of MVS. In combination with the reduced MVD, this could result only in a fractional reduced perfusion which might be an explanation for the lower occurrence of BP-ONJ in patients receiving ibandronate compared to patients receiving pamidronate or zoledronate.