EMT and Tumor Turning Point Analysis in 3D Spheroid Culture of HNSCC and Mesenchymal Stem Cells.

The prognosis, metastasis, and behavior of head and neck squamous cancer cells are influenced by numerous factors concerning the tumor microenvironment, intercellular communication, and epithelial-to-mesenchymal transition (EMT). The aim of this study was to examine the codependent interaction of the mesenchymal stroma with head and neck squamous cell carcinoma (HNSCC) in a 3D spheroid structure. To simulate stroma-rich and -poor 3D tumor microenvironments, cells of the established cell SCC-040 were cultured with human mesenchymal stromal cells (MSCs), forming 3D stroma-tumor spheroids (STSs). STSs were compared to uniform spheroids of SCC-040 and MSC, respectively. The expressions of CD24, ß-catenin, SNAI2, and ZEB2 were analyzed via RT-qPCR. The immunohistochemical expressions of E-cadherin, connexin 43, vimentin, and emmprin were analyzed, and protein expression pathways as well as Akt signaling were assessed via protein analysis. A promotive effect on the expressions of EMT markers ZEB2 (p = 0.0099), SNAI2 (p = 0.0352), and ß-catenin (p = 0.0031) was demonstrated in STSs, as was the expression of Akt pathway proteins mTOR (p = 0.007), Erk1/2 (p = 0.0045), and p70 S6 Kinase (p = 0.0016). Our study demonstrated a change in genetic expression patterns early on in tumor development, indicating a tumor turning point.

Accelerating the Front End of Medicine: Three Digital Use Cases and HCI Implications.

Digital applications in health care are a concurrent research and management question, where implementation experiences are a core field of information systems research. It also contributes to fighting pandemic crises like COVID-19 because contactless information flow and speed of diagnostics are improved. This paper presents three digital application case studies from emergency medicine, administration management, and cancer diagnosis with AI support from the University Medical Centers of Münster and Göttingen in Germany. All cases highlight the potential of digitalization to increase speed and efficiency within the front end of medicine as the crucial phase before patient treatment starts. General challenges for health care project implementations and human-computer interaction (HCI) concepts in health care are derived and discussed, including the importance of specific processes together with user analysis and adaption. A derived concept for HCI includes the criteria speed, accuracy, modularity, and individuality to achieve sustainable improvements within the front end of medicine.

Accuracy of intraoral real-time navigation versus static, CAD/CAM-manufactured pilot drilling guides in dental implant surgery: an in vitro study.

BACKGROUND: Nowadays, 3D planning and static for dynamic aids play an increasing role in oral rehabilitation of the masticatory apparatus with dental implants. The aim of this study is to compare the accuracy of implant placement using a 3D-printed drilling guide and an intraoral real-time dynamic navigation system. METHODS: A total of 60 implants were placed on 12 partially edentulous lower jaw models. 30 were placed with pilot drilling guides, the other half with dynamic navigation (DENACAM®). In addition, implant placement in interdental gaps and free-end situations were investigated. Accuracy was assessed by cone-beam computed tomography (CBCT). RESULTS: Both systems achieved clinically acceptable results, yet more accurate results regarding the offset of implant base and tip in several spatial dimensions were achieved using drilling guides (each p < 0.05). With regard to angulation, real-time navigation was more precise (p = 0.0016). Its inaccuracy was 3°; the template-guided systems was 4.6°. Median horizontal deviation was 0.52 mm at base and 0.75 mm at tip using DENACAM®. When using the pilot drill guide, horizontal deviation was 0.34 mm in the median and at the tip by 0.59 mm. Regarding angulation, it was found that the closer the drill hole was to the system's marker, the better navigation performed. The template did not show this trend (p = 0.0043; and p = 0.0022). CONCLUSION: Considering the limitations of an in vitro study, dynamic navigation can be used be a tool for reliable and accurate implantation. However, further clinical studies need to follow in order to provide an evidence-based recommendation for use in vivo.

Cotransplantation of mesenchymal stromal cells and endothelial cells on calcium carbonate and hydroxylapatite scaffolds in vivo.

This study investigated the cotransplantation of bone marrow mesenchymal stromal cells (BMSC) and human umbilical cord endothelial cells (HUVEC), and evaluated their contribution to vascular and bone tissue engineering in vivo. To evaluate the success of osteogenic differentiation and timely vascularization of different osteoconductive scaffolds in vivo, we transferred BMSC and HUVEC pre-cultivated calcium carbonate (CaCO3) and hydroxylapatite (HA) matrices into immunocompromised RNU-rats, and analyzed mineralization, expression of osteopontin, and vascular integration via new vessel formation. After in vivo transplantation, pre-cultivated scaffolds demonstrated overall improved mineralization of 44% for CaCO3 (p = 0.01, SD ± 14.3) and 34% for HA (p = 0.001, SD ± 17.8), as well as improved vascularization of 5.6 vessels/0.1 mm2 on CaCO3 (p < 0.0001, SD ± 2.0) and 5.3 vessels/0.1 mm2 on HA (p < 0.0001, SD ± 2.4) compared with non-pre-cultivated controls. However, no significant differences between the implantation of BMSC-only, HUVEC-only, or BMSC + HUVEC cocultures could be observed. There is an increasing demand for improved bone regeneration in tissue engineering. Cotransplantation of mesenchymal stromal cells and endothelial cells often demonstrates synergistic improvements in vitro. However, the benefits or superiority of cotransplantation was not evident in vivo and so will require further investigation.

Bone tissue engineering in the greater omentum is enhanced by a periosteal transplant in a miniature pig model.

AIM: Reconstruction of bone defects with autologous grafts has certain disadvantages. The aim of this study is to introduce a new type of living bioreactor for engineering of bone flaps and to evaluate the effect of different barrier membranes. MATERIALS & METHODS: Scaffolds loaded with bone morphogenetic proteins and bone marrow aspirate wrapped with either a collagen membrane or a periosteal flap were implanted in the greater omentum of miniature pigs. RESULTS: Both histological and radiographic evaluation showed proven bone formation and increased density after 8 and 16 weeks, with an enhanced effect of the periosteal transplant. CONCLUSION: The greater omentum is a suitable bioreactor for bone tissue engineering. Endocultivation is both an innovative and promising approach in regenerative medicine.

Removable thermoplastic appliances modified by incisal cuts show altered biomechanical properties during tipping of a maxillary central incisor.

BACKGROUND: The present study aimed to evaluate the force delivery of removable thermoplastic appliances (RTAs), modified by different sized incisal cuts, during tipping of a maxillary central incisor in palatal and vestibular direction. METHODS: Forty-five RTAs from three different materials (Biolon®, Erkodur®, Ideal Clear®) of the same thickness (1 mm) were used. Analysis was performed on a separated maxillary central incisor which was part of a resin model with a complete dentition. In 15 RTAs, of different material, a cut was inserted at the incisal edge of tooth 11. In 15 other appliances, the cut was extended to teeth 12 and 21. Fifteen aligners remained uncut. The experimental tooth was tipped starting from the zero position in 0.05° steps to a maximal deflection of ± 0.42° of the incisal edge in vestibular and palatal direction, after positioning the RTA onto the model. RESULTS: The horizontal (Fx) and the vertical (Fz) force components were decreased by approximately half with increasing cut size. Fz values changed during palatal tipping from a weak intrusive force, for aligners without cut, to an extrusive force with increasing cut size. Compared to both other materials used (Erkodur® and Ideal Clear®), the Biolon® aligners showed significantly higher Fx and Fz values (p < 0.0001, respectively). CONCLUSIONS: RTAs modified by different sized incisal cuts show altered biomechanical properties and an inversion of the vertical force component, during tipping of a maxillary central incisor.

Bone Morphogenetic Protein-2 Hybridized with Nano-Anchored Oligonucleotides on Titanium Implants Enhances Osteogenic Differentiation In Vivo.

PURPOSE: Previous in vitro studies have shown that DNA oligonucleotides (ODN) can be successfully used as anchor strands for the binding and retarded release of biologically active recombinant human bone morphogenetic protein 2 (rhBMP-2). The aim of the present study was to test the hypothesis that rhBMP-2 bound to the surface of titanium implants through hybridization with nano-anchored ODN strands is biologically active and can enhance the induction of osteogenic markers in peri-implant bone in vivo. MATERIALS AND METHODS: Custom-made, surface acid-etched (SAE) titanium discs and implants were coated with ODN anchor strands and subsequently hybridized with complementary ODN strands conjugated to rhBMP-2 (AS_CS_BMP-2). Discs/implants with SAE surface, ODN-coated surface (AS), and ODN-coated surface with nonconjugated rhBMP-2 (AS_BMP-2) served as controls. Release of rhBMP-2 from the coated discs was evaluated in vitro using enzyme-linked immunosorbent assay (ELISA), and bone-specific activity was assessed through pNPP turnover by induced alkaline phosphatase (AP) up to a period of 56 days. In vivo expression of bone-specific markers was analyzed after bilateral placement of coated implants into the tibiae of 36 Wistar rats (72 tibiae total). Immunostaining for AP and runt-related transcription factor 2 (Runx2) was carried out after 1, 4, and 13 weeks. RESULTS: Release from the AS_CS_ BMP-2-coated titanium surfaces was significantly retarded compared to surfaces loaded with AS_BMP-2. The in vitro biologic activity of the released rhBMP-2 conjugates measured by AP induction was equivalent to released nonconjugated rhBMP-2. Immunostaining revealed a significant increase in the in vivo induction of AP around AS_CS_BMP-2 implants compared to the controls after 1 and 4 weeks. CONCLUSION: Titanium AS_CS_BMP-2 implants can significantly enhance osteogenic differentiation in vivo in peri-implant bone in early periods of osseointegration.

Supportive angiogenic and osteogenic differentiation of mesenchymal stromal cells and endothelial cells in monolayer and co-cultures.

Sites of implantation with compromised biology may be unable to achieve the required level of angiogenic and osteogenic regeneration. The specific function and contribution of different cell types to the formation of prevascularized, osteogenic networks in co-culture remains unclear. To determine how bone marrow-derived mesenchymal stromal cells (BMSCs) and endothelial cells (ECs) contribute to cellular proangiogenic differentiation, we analysed the differentiation of BMSCs and ECs in standardized monolayer, Transwell and co-cultures. BMSCs were derived from the iliac bone marrow of five patients, characterized and differentiated in standardized monolayers, permeable Transwells and co-cultures with human umbilical vein ECs (HUVECs). The expression levels of CD31, von Willebrand factor, osteonectin (ON) and Runx2 were assessed by quantitative reverse transcriptase polymerase chain reaction. The protein expression of alkaline phosphatase, ON and CD31 was demonstrated via histochemical and immunofluorescence analysis. The results showed that BMSCs and HUVECs were able to retain their lineage-specific osteogenic and angiogenic differentiation in direct and indirect co-cultures. In addition, BMSCs demonstrated a supportive expression of angiogenic function in co-culture, while HUVEC was able to improve the expression of osteogenic marker molecules in BMSCs.

Common molecularcytogenetic alterations in tumors originating from the pineal region.

Tumors of the pineal region (PR) are rare and can be subdivided into four main histomorphological groups: Pineal-parenchymal tumors (PPT), germ cell tumors (GCT), glial tumors and miscellaneous tumors. The appropriate pathological classification and grading of these malignancies is essential for determining the clinical management and prognosis. However, an early diagnosis is often delayed due to unspecific clinical symptoms, and histological support is not always decisive to identify the diversity of tumors of the PR. The present study aimed to characterize 18 tumors of the PR using comparative genomic hybridization. All the tumors were primarily surgically resected without any previous irradiation or chemotherapy. In addition to chromosomal aberrations in PPT and different GCTs of the PR, the present study described, for the first time, the chromosomal changes in a few rare entities (solitary-fibrous and neuroendocrine tumors) of the PR. The tumors in the study, regardless of histology and World Health Organization grade, were characterized by frequent gains at 7, 9q, 12q, 16p, 17 and 22q, and losses at 13q. While the detection of chromosomal aberrations in these tumors appears not to be indicative enough of histological entities and their grade of malignancy, the present data may be of use to select genes of interest for higher resolution genomic analyses.

The submental island flap for the treatment of intraoral tumor-related defects: No effect on recurrence rates.

OBJECTIVES: The submental island flap has become increasingly popular in the treatment of intraoral defects following tumor ablation. However, there was concern that the elevation of the pedicled flap might interfere with the efficiency of level I-lymph node dissection and decrease the oncologic prognosis of the patients. MATERIALS AND METHODS: In a prospective clinical study over five years the outcome of 45 consecutive patients with intraoral cancer of various T-stages treated with submental island flaps was evaluated and compared to 45 patients with a T-status analogous oral cancer treated with free radial forearm flaps. RESULTS: All submental island flaps beside three were successful (93.3%). The obtained functional results were pleasing and the donor morbidity low. Patients treated with submental island flaps exhibited no enhanced risk of local tumor recurrence or lymph node metastasis (p<0.86). In contrast, the operation time, time of intensive care and hospitalization were reduced (p<0.001). DISCUSSION: We conclude that the submental island flap is an effective and predictable option of small and medium-sized oral defect treatment. It is a valuable alternative to free flap soft tissue reconstructions such as radial forearm or perforator flaps. It seems particularly beneficial to patients with relevant comorbidities as often present in the oral cancer population. The application of the submental island flap does not reduce the oncological prognosis of oral cancer patients.

Human mesenchymal stromal cells from adipose tissue of the neck.

Mesenchymal stromal cells (MSC) have been introduced into the field of tissue-engineered airway transplantation. Since patients with extensive tracheal defects often require an open tracheotomy, this study investigated if MSC could be obtained from the adipose tissue of the neck during this procedure. Cells were isolated by plastic adherence from the adipose tissue of 8 patients. Cell isolates were analyzed for (i) proliferation, (ii) the expression of CD marker molecules and (iii) multilineage differentiation. The isolated spindle-shaped cells showed a high proliferation capacity and the flow cytometric analysis revealed a distinct population meeting the criteria for MSC. Using classical MSC cultivation protocols the characterized cells showed adipogenic, chondrogenic and osteogenic differentiation for all analyzed cell isolates. This study was able to demonstrate that sufficient amounts of stem/progenitor cells can be easily isolated from adipose tissue of the neck obtained during open tracheotomy. These cells may be a source for future tracheal replacement therapies.

Murine mesenchymal progenitor cells from different tissues differentiated via mesenchymal microspheres into the mesodermal direction.

BACKGROUND: Because specific marker molecules for phenotypical identification of mesenchymal stem and progenitor cells are missing, the assessment of the in vitro-differentiation capacity is a prerequisite to characterize these cells. However, classical differentiation protocols are often cell-consuming and time intensive. Therefore, the establishment of novel strategies for differentiation is one topic of current efforts in stem cell biology. The goal of this study was to demonstrate the practicability of a new differentiation test using plastic adherent cell isolates from different tissues. RESULTS: We introduced the mesenchymal microsphere method as a feasible time- and cell saving screening method to analyse multilineage differentiation properties of adult progenitor cells in a three-dimensional system. For this purpose we isolated, characterized and analyzed new sources of adult murine mesenchymal progenitor cells from perirenal adipose tissue and mediastinal stromal tissue in comparison to bone marrow progenitor cells. The proliferation capacity of the cells was demonstrated by determination of the daily doubling index. Although the flow cytometry analysis of undifferentiated cells revealed differences in the expression of CD marker molecules, all isolates have the capacity for multilineage differentiation following the mesenchymal microsphere protocol as well as the classical "micro mass body" protocol for chondrogenic and the monolayer cultivation protocol for osteogenic and adipogenic differentiation. Differentiation was characterized using histochemical and immunhistochemical staining as well as RT-PCR. CONCLUSIONS: We were able to show that the mesenchymal microsphere method is an efficient test system for chondro-, osteo- and adipogenic differentiation of adult progenitor cells. The advantage of this system in comparison to classical protocols is that approximately 7 times lower cell numbers are necessary. Since classical culture procedures are time intensive because high cell numbers have to be obtained, the new differentiation method may also save cells and time in future clinical applications using human mesenchymal stromal cells.

Isolation and characterization of adult stem cells from human salivary glands.

Currently, adult stem cells are attracting significant interest in regenerative medicine and tissue engineering. These cells have been isolated from various tissue sources; however, in most cases, adult stem cells useful for tissue engineering and regeneration are present at a low frequency. High numbers of stem cells with an effective and reliable potential for differentiation are needed for clinical applications. Thus, the identification of new stem cell sources and the establishment of optimized cell culture conditions that allow for the amplification of stem cells are of utmost relevance. In addition, the isolation procedure should ideally be minimally invasive and possibly be performed under local anesthesia. We report here for the first time on the identification of adult stem cells with mesenchymal characteristics in human parotid gland tissue. Cells were isolated from freshly resected specimens of parotid glands using enzymatic digestion and plastic adhesion protocols. Following an initial proliferation period and short-term culture for four passages, immunophenotyping revealed the presence of mesenchymal stem cell markers. In the presence of tissue-specificinduction medium, stem cells could be differentiated into adipogenic, osteogenic, and chondrogenic cell types. Tissue-specific differentiation was confirmed by histochemical and immunocytochemical staining as well as by RT-PCR for defined marker genes. This study is, to the best of our knowledge, the first report on the isolation and differentiation of stem cells from adult human parotid glands. Although isolated from an endodermal tissue source, these stem cells share many characteristics with MSCs. Easy accessibility and a high differentiation potential make salivary gland-derived stem cells a promising source for future applications in regenerative medicine.