Circulating tumor cells (CTCs) are promising biomarkers for diagnosis and therapy in systemic cancer. However, their infrequent and unreliable detection, especially in nonmetastatic cancer, currently ...impedes the clinical use of CTCs. Because leukapheresis (LA) targets peripheral blood mononuclear cells, which have a similar density to CTCs, and usually involves processing the whole circulating blood, we tested whether LA could substantially increase CTC detection in operable cancer patients. Therefore, we screened LA products generated from up to 25 L of blood per patient in two independent studies, and found that CTCs can be detected in more than 90% of nonmetastatic breast cancer patients. Interestingly, complete white blood cell sampling enabled determining an upper level for total CTC numbers of about 100,000 cells (median, 7,500 CTCs) per patient and identified a correlation of CTC numbers with anatomic disease spread. We further show that diagnostic leukapheresis can be easily combined with the US Food and Drug Administration-approved CellSearch system for standardized enumeration of CTCs. Direct comparison with 7.5 mL of blood revealed a significantly higher CTC frequency in matched LA samples. Finally, genomic single-cell profiling disclosed highly aberrant CTCs as therapy-escaping variants in breast cancer. In conclusion, LA is a clinically safe method that enabled a reliable detection of CTCs at high frequency even in nonmetastatic cancer patients, and might facilitate the routine clinical use of CTCs as in the sense of a liquid biopsy. Combined with technologies for single-cell molecular genetics or cell biology, it may significantly improve prediction of therapy response and monitoring of early systemic cancer.
The aim of this retrospective, single-center study was to analyze long-term results after marginal and segmental mandibulectomies in patients with oral squamous cell carcinoma (OSCC).
The study ...included 259 patients treated for OSCC with mandibulectomy between 1996 and 2010. Data acquisition consisted of analysis of operation reports, re-evaluation of histologic bone specimens, and collection of clinical follow-up data.
Of the included patients, 86.5% had received segmental and 13.5% marginal mandibulectomies. Patients who received segmental mandibulectomy generally displayed a higher TNM (tumor–node–metastasis) stage; 47% of patients who received segmental mandibulectomy and 14% of those receiving marginal mandibulectomy showed bone infiltration (pT4 a). Of all patients with bone infiltration, 49% showed an invasive histologic infiltration pattern, and 35% showed an erosive histologic infiltration pattern. We found healthy residual crestal bone height in 43% of all segmental mandibulectomies. Only 8% of all patients were prosthodontically rehabilitated. With regard to prognostic parameters, there was no significant difference between patients receiving marginal mandibulectomy and those receiving segmental mandibulectomy.
Because healthy residual crestal bone height was found in 43% of all patients who had received segmental mandibulectomies, it is conceivable that a significant number of patients would profit from marginal mandibulectomy, at least in cases of absent or erosive bone infiltration pattern, because the residual crestal bone is functionally stable.
PEMT and epithelial differentiating gene expression signatures are comparable to previously published EMT signatures. (C) Heatmap for scaled, log-normalized gene expression of all cell types from ...patient #1 and their top 10 DEGs (rows) against all other cells. PEMT and epithelial differentiating gene expression signatures are comparable to previously published EMT signatures. (B) Average log2 fold change of gene expression (x-axis) and differences in cellular fractions expressing the respective gene (y-axis) between pEMT and epithelial differentiated cell clusters. On the right side, the correlation coefficients between GSVA scores of EMT signatures from the EMTome database and of epithelial differentiation and pEMT signatures from patient #1 (right) are shown and next to it, annotated as”EXPR_perc”, is the fraction of genes with non-zero expression and the size of the respective EMT signature in log10 scale with the respective number next to it.
Oral squamous cell carcinoma (OSCC), an HPV-negative head and neck cancer, frequently metastasizes to the regional lymph nodes but only occasionally beyond. Initial phases of metastasis are ...associated with an epithelial-mesenchymal transition (EMT), while the consolidation phase is associated with mesenchymal-epithelial transition (MET). This dynamic is referred to as epithelial-mesenchymal plasticity (EMP). While it is known that EMP is essential for cancer cell invasion and metastatic spread, less is known about the heterogeneity of EMP states and even less about the heterogeneity between primary and metastatic lesions.
To assess both the heterogeneity of EMP states in OSCC cells and their effects on stromal cells, we performed single-cell RNA sequencing (scRNAseq) of 5 primary tumors, 9 matching metastatic and 5 tumor-free lymph nodes and re-analyzed publicly available scRNAseq data of 9 additional primary tumors. For examining the cell type composition, we performed bulk transcriptome sequencing. Protein expression of selected genes were confirmed by immunohistochemistry.
From the 23 OSCC lesions, the single cell transcriptomes of a total of 7263 carcinoma cells were available for in-depth analyses. We initially focused on one lesion to avoid confounding inter-patient heterogeneity and identified OSCC cells expressing genes characteristic of different epithelial and partial EMT stages. RNA velocity and the increase in inferred copy number variations indicated a progressive trajectory towards epithelial differentiation in this metastatic lesion, i.e., cells likely underwent MET. Extension to all samples revealed a less stringent but essentially similar pattern. Interestingly, MET cells show increased activity of the EMT-activator ZEB1. Immunohistochemistry confirmed that ZEB1 was co-expressed with the epithelial marker cornifin B in individual tumor cells. The lack of E-cadherin mRNA expression suggests this is a partial MET. Within the tumor microenvironment we found immunomodulating fibroblasts that were maintained in primary and metastatic OSCC.
This study reveals that EMP enables different partial EMT and epithelial phenotypes of OSCC cells, which are endowed with capabilities essential for the different stages of the metastatic process, including maintenance of cellular integrity. During MET, ZEB1 appears to be functionally active, indicating a more complex role of ZEB1 than mere induction of EMT.
Successful treatment for any type of carcinoma largely depends on understanding the patterns of invasion and migration. For oral squamous cell carcinoma (OSCC), these processes are not entirely ...understood as of now. Invadopodia and podosomes, called invadosomes, play an important role in cancer cell invasion and migration. Previous research has established that cortactin (CTTN) is a major inducer of invadosome formation. However, less is known about the expression patterns of CTTN and other genes related to it or invadopodia formation in OSCC during tumor progression in particular. In this study, gene expression patterns of CTTN and various genes (n = 36) associated with invadopodia formation were analyzed to reveal relevant expression patterns and give a comprehensive overview of them. The genes were analyzed from a whole genome dataset of 83 OSCC samples relating to tumor size, grading, lymph node status, and UICC (Union for Internatioanl Cancer Control). The data revealed significant overexpression of 18 genes, most notably CTTN, SRC (SRC proto-onocogene, non-receptor tyrosine kinase), EGFR (epidermal growth factor receptor), SYK (spleen associated tyrosine kinase), WASL (WASP like actin nucleation promotion factor), and ARPC2 (arrestin beta 1) due to their significant correlation with further tumor parameters. This study is one of the first to summarize the expression patterns of CTTN and related genes in a complex group of OSCC samples.
Objective
This study was performed to determine whether the results of prevailing in vivo and in vitro studies offer a reliable model for investigation of medication-related osteonecrosis of the jaw ...(MRONJ).
Methods
Embase, Medline, and the Cochrane Library were searched for articles published from September 2003 to June 2017 involving experimental approaches to the pathogenesis of MRONJ. In vivo and in vitro trials were analyzed with respect to the scientific question, study design, methodology, and results.
Results
Of 139 studies, 87, 46, and 6 conducted in vivo, in vitro, and both in vivo and in vitro experiments, respectively. Rats, mice, dogs, minipigs, sheep, and rabbits were the preferred animal models used. Osteoblasts, osteoclasts, fibroblasts, keratinocytes, macrophages, and human umbilical vein endothelial cells were the preferred cell types. Zoledronate, alendronate, ibandronate, and risedronate were the most frequent bisphosphonates used. MRONJ was most reliably induced in minipigs because of the close relationship with human bone physiology. In vitro studies showed that reduced viability, growth, and migration of cells in the bone and soft tissues were causative for MRONJ. Other than exposed jawbone after tooth extraction, no reliable cofactors were found.
Conclusion
The minipig is the most suitable animal model for MRONJ.
Sufficient vertical and lateral bone supply and a competent osteogenic healing process are prerequisities for the successful osseointegration of dental implants in the alveolar bone. Several ...techniques including autologous bone grafts and guided bone regeneration are applied to improve quality and quantity of bone at the implantation site. Depending on the amount of lacking bone one- or two-stage procedures are required. Vertical bone augmentation has proven to be a challenge particularly in terms of bone volume stability. This study focuses on the three dimensional vertical bone generation in a one stage procedure in vivo. Therefore, a collagenous disc-shaped scaffold (ICBM = Insoluble Collagenous Bone Matrix) containing rhBMP-2 (Bone Morphogenetic Protein-2) and/or VEGF (Vascular Endothelial Growth Factor) was applied around the coronal part of a dental implant during insertion. RhBMP-2 and VEGF released directly at the implantation site were assumed to induce the generation of new vertical bone around the implant.
One hundred eight titanium implants were inserted into the mandible and the tibia of 12 mini pigs. Four experimental groups were formed: Control group, ICBM, ICBM + BMP-2, and ICBM + BMP-2 + VEGF. After 1, 4 and 12 weeks the animals were sacrificed and bone generation was investigated histologically and histomorphometrically.
After 12 weeks the combination of ICBM + rhBMP2 + VEGF showed significantly more bone volume density (BVD%), a higher vertical bone gain (VBG) and more vertical bone gain around the implant (PVBG) in comparison to the control group.
By using collagenous disc-shaped matrices in combination with rhBMP-2 and VEGF vertical bone can be generated in a one stage procedure without donor site morbidity. The results of the presenting study suggest that the combination of rhBMP-2 and VEGF applied locally by using a collagenous carrier improves vertical bone generation in vivo. Further research is needed to establish whether this technique is applicable in clinical routines.
Detailed examination of tumor components is leading‐edge to establish personalized cancer therapy. Accompanying research on cell‐free DNA, the cell count of circulating tumor cells (CTCs) in patient ...blood is seen as a crucial prognostic factor. The potential of CTC analysis is further not limited to the determination of the overall survival rate but sheds light on understanding inter‐ and intratumoral heterogeneity. In this regard, commercial CTC isolation devices combining an efficient enrichment of rare cells with a droplet deposition of single cells for downstream analysis are highly appreciated. The Liquid biopsy platform CTCelect was developed to realize a fully‐automated enrichment and single cell dispensing of CTCs from whole blood without pre‐processing. We characterized each process step with two different carcinoma cell lines demonstrating up to 87 % enrichment (n = 10) with EpCAM coupled immunomagnetic beads, 73 % optical detection and dispensing efficiency (n = 5). 40 to 56.7 % of cells were recovered after complete isolation from 7.5 ml untreated whole blood (n = 6). In this study, CTCelect enabled automated dispensing of single circulating tumor cells from HNSCC patient samples, qPCR‐based confirmation of tumor‐related biomarkers and immunostaining. Finally, the platform was compared to commercial CTC isolation technologies to highlight advantages and limitations of CTCelect. This system offers new possibilities for single cell screening in cancer diagnostics, individual therapy approaches and real‐time monitoring.
Alloplastic and xenogeneic bone grafting materials are frequently used for bone augmentation. The effect of these materials on precursor cells for bone augmentation is yet to be determined. The aim ...of this study was to ascertain, in vitro, how augmentation materials influence the growth rates and viability of human unrestricted somatic stem cells. The biocompatibility of two xenogeneic and one alloplastic bone graft was tested using human unrestricted somatic stem cells (USSCs). Proliferation, growth, survival and attachment of unrestricted somatic stem cells were monitored after 24 h, 48 h and 7 days. Furthermore, cell shape and morphology were evaluated by SEM. Scaffolds were assessed for their physical properties by Micro-CT imaging. USSCs showed distinct proliferation on the different carriers. Greatest proliferation was observed on the xenogeneic carriers along with improved viability of the cells. Pore sizes of the scaffolds varied significantly, with the xenogeneic materials providing greater pore sizes than the synthetic inorganic material. Unrestricted somatic stem cells in combination with a bovine collagenous bone block seem to be very compatible. A scaffold’s surface morphology, pore size and bioactive characteristics influence the proliferation, attachment and viability of USSCs.
Objectives
Sialolithiasis is the most common cause of chronic sialadenitis of the submandibular gland (SMG). Symptomatic superficial lobe stones are often treated by submandibulectomy. A gland ...preserving operation allows for transoral stone removal through endoscopically assisted sialolithotomy. Herein, we provide clinical and sonographical follow-up data in patients who underwent sialolithotomy under general anesthesia.
Materials and methods
Sixty patients treated at the Department of Oral and Maxillofacial Surgery at Düsseldorf University Hospital for superficial lobe sialolithiasis of SMG were included in this study. All received transoral sialolithotomy under general anesthesia. Follow-up was conducted via standardized patient questionnaires, clinical examination, and B-mode and color Doppler sonography.
Results
Mean patient age was 48.9 years. 56.6% of right and 43.4% of left SMG were affected. Mean follow-up was 45 months. Fifty-five of 59 detected stones could be removed. Mean operation time was 71 min. 3.3% of patients reported recurrent episodes of postoperative pain and 10% felt recurrent episodes of gland swelling. Persistent postoperative lingual nerve hypesthesia was described in one patient. No facial nerve damages occurred. Salivary flow rates remained reduced in most of the affected glands upon stone removal. Sonographical follow-up data of the previously affected SMG after intraoral endoscopy-assisted sialolithotomy showed a regular gland size in 70.8% of cases, a parenchyma free of inflammation in 93.8%, and without signs of fibrosis in 72.9% of cases. 68.7% of patients showed a regular structure of Wharton’s duct at time of follow-up. In total, 89.6% of patients were diagnosed stone-free within both glands on follow-up. No case required subsequent submandibulectomy.
Conclusions
Sialolithotomy of Wharton’s duct for removal of stones from the SMG’s superficial lobe is a promising alternative to submandibulectomy.
Clinical relevance
Reduction of postoperative morbidity through endoscopically assisted sialolithotomy for removal of superficial lobe stones from SMG.
Trial registration
Ethics Committee of Heinrich-Heine-University Düsseldorf (no. 5586)
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