Comprehensive Analysis Identifies Ameloblastin-Related Competitive Endogenous RNA as a Prognostic Biomarker for Testicular Germ Cell Tumour

Simple Summary Testicular germ cell tumour is a common tumour in young males, and although it is one of the most curable cancers, many patients still experience recurrence after the chemotherapy. Tumour recurrence is not detected with high sensitivity by established blood tumour markers. Ameloblastin is identified as an extracellular matrix protein and has shown to be associated with tumour progression. We validated ameloblastin’s expression in testicular tissue, and used comprehensive bioinformatics analysis of 156 patients with testicular germ cell tumour to show that the level of ameloblastin was associated with the time of tumour recurrence after the first cure. In the analysis of ameloblastin differential genes in the tumour, a ceRNA (competing endogenous RNA) regulatory network associated with tumour diagnosis and an independent prognostic factor for the tumour, PELATON (Plaque Enriched LncRNA In Atherosclerotic And Inflammatory Bowel Macrophage Regulation), were identified, which could provide evidence for prediction of tumour prognosis. Abstract Testicular Germ Cell Tumour (TGCT) is one of the most common tumours in young men. Increasing evidence shows that the extracellular matrix has a key role in the prognosis and metastasis of various human cancers. This study analysed the relationship between the matrix protein ameloblastin (AMBN) and potential biological markers associated with TGCT diagnosis and prognosis. The relationship between AMBN and TGCT prognosis was determined by bioinformatic analysis using the expression profiles of three RNAs (long non-coding RNAs (lncRNAs), microRNAs (miRNAs) and mRNAs) from The Cancer Genome Atlas (TCGA) database, and available clinical information of the corresponding patients. Prediction and validation of competitive endogenous RNA (ceRNA) regulatory networks related to AMBN was performed. AMBN and its associated ceRNA regulatory network were found to be related to the recurrence of TGCT, and LINC02701 may be used as a diagnostic factor in TGCT. Furthermore, we identified PELATON (Plaque Enriched LncRNA In Atherosclerotic And Inflammatory Bowel Macrophage Regulation) as an independent prognostic factor for TGCT progression-free interval

A single amino acid deletion in the ER Ca2+ sensor STIM1 reverses the in vitro and in vivo effects of the Stormorken syndrome-causing R304W mutation

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Osteoblasts in a Perfusion Flow Bioreactor—Tissue Engineered Constructs of TiO2 Scaffolds and Cells for Improved Clinical Performance

Combining biomaterial scaffolds with cells serves as a promising strategy for engineering critical size defects; however, homogenous cellular growth within large scaffolds is challenging. Mechanical stimuli can enhance bone regeneration by modulating cellular growth and differentiation. Here, we compare dynamic seeding in a perfusion flow bioreactor with static seeding for a synthetic bone scaffold for up to 21 days using the cell line MC3T3-E1 and primary human osteoblast, confocal laser scanning microscopy, and real-time reverse transcriptase-polymerase chain reaction. The secretion of bone-related proteins was quantified using multiplex immunoassays. Dynamic culture improved cellular distribution through the TiO2 scaffold and induced a five-fold increase in cell number after 21 days. The relative mRNA expression of osteopontin of MC3T3-E1 was 40-fold enhanced after 7 and 21 days at a flow rate of 0.08 mL/min, and that of collagen type I alpha I expression was 18-fold after 21 days. A flow rate of 0.16 mL/min was 10-fold less effective. Dynamic culture increased the levels of dickkopf-related protein 1 (60-fold), osteoprotegrin (29-fold), interleukin-6 (23-fold), interleukin-8 (36-fold), monocyte chemoattractant protein 1 (28-fold) and vascular endothelial growth factor (6-fold) in the medium of primary human osteoblasts after 21 days compared to static seeding. The proposed method may have clinical potential for bone tissue engineering

Injectable biomaterials for Dental Tissue Regeneration

Injectable biomaterials scaffolds play a pivotal role for dental tissue regeneration, as such materials are more applicable in the dental field, particularly when compared to pre-formed scaffolds. The defects in maxilla-oral area are normally small, confined and sometimes hard to access. This narrative review described different types of biomaterials for dental tissue regeneration, also discussing the potential use of nanofibres for dental tissues. Various studies suggest that the tissue engineering approaches involving the use of injectable biomaterials have the potential of restoring not only dental tissue function but also their biological purpose

Fluoride Modification of Titanium Surfaces Enhance Complement Activation

Immediately after dental implant insertion, blood will be in direct contact and interact with the implant surface and activates inflammatory responses and complement cascades within seconds. The aim of the present study was to determine the ability of fluoride-modified titanium surfaces to activate complement cascades using the human buffy coat as model. The buffy coats were exposed to hydrofluoric acid-modified surfaces for a short time and its responses were compared to controls. Identification and quantification of complement cascade biomarkers were conducted using ELISA kits and multianalyte profiling using Luminex. A lower level of C3 at 30 min and increased levels of C4, MIP-4, CRP, and pigment epithelium-derived factor at 360 min were found on modified surfaces as compared to controls. We found no significant differences in the levels of C3a, C5a, C Factor H, α2M, ApoA1, ApoC3, ApoE, Prealbumin, α1AT, and SAP in modified surfaces in the buffy coats. We conclude that titanium surfaces treated with hydrofluoric acid modify the levels of specific biomarkers related to the complement cascade and angiogenesis and, thus, tissue growth, remodeling and repair, as this may play a role in the enhanced clinical performance of fluoride-modified Ti dental implants

General joint hypermobility in temporomandibular joint disease; clinical characteristics, biomarkers, and surgical aspects

Objectives: This study aimed at identifying biomarkers in the temporomandibular joint (TMJ) synovial tissue analysing 28 extra cellular matrix proteins in TMJ diseased patients, classified with either general joint hypermobility (GJH) or normal joint mobility (NJM), and to compile clinical and protein characterisation to reveal potential surgical predictive factors. Study design: A prospective observational cohort study including 97 consecutive patients scheduled for TMJ surgery was performed. Joint mobility and several other predefined clinical variables were recorded. Synovial tissue was harvested during surgery followed by examination using multi-analytic profiling. A multivariate quantile regression model was used for analysis purposes. Results: The GJH/NJM ratio was 2:5. The GJH cohort were younger (P = 0.001) and more likely to be women (P = 0.026) compared to the NJM cohort. None of the protein concentrations could be correlated to joint mobility in the multivariate regression model, but often to the variable TMJ diagnosis. The surgical outcome after the six-month follow-up were equal between GJH and NJM patients. Conclusions: GJH was more common in the study cohort compared to general population frequencies, but GJH was not a negative factor for surgical outcome. Young age and female gender correlated to GJH. No TMJ biomarkers were GJH specific, and the results suggested that TMJ diagnosis more strongly correlated to the protein profile compared to GJH and the other investigated variables

Mapping the global mRNA transcriptome during development of the murine first molar

The main objective of this study was to map global geneexpression in order to provide information about the populations of mRNA species participating in murine tooth development at 24 h intervals, starting at the eleventh embryonic day (E11.5) up to the seventh post-natal day (P7). The levels of RNA species expressed during murine tooth development were mesured using a total of 58 deoxyoligonucleotide microarrays. Microarray data was validated using real-time RT-PCR. Differentially expressed genes (p<0.05) were subjected to bioinformatic analysis to identify cellular activities significantly associated with these genes. Using ANOVA the microarray data yielded 4362 genes as being differentially expressed from the elleventh embryonic day (E11.5) up to seven days post-natal (P7), 1921 of these being genes without known functions. The remaining 2441 genes were subjected to further statistical analysis using a supervised procedure. Bioinformatic analysis results for each time-point studied suggests that the main molecular functions associated with genes expressed at the early pre-natal stages (E12.5-E18.5) studied were cell cycle progression, cell morphology, lipid metabolism, cellular growth, proliferation, senescence and apoptosis, whereas most genes expressed at post-natal and secretory stages (P0- P7) were significantly associated with regulation of cell migration, biosynthesis, differentiation, oxidative stress, polarization and cell death. Differentially expressed genes (DE) not described earlier during tooth murine tooth development; Inositol 1, 4, 5-triphosphate receptor 3 (Itpr3), metallothionein 1(Mt1), cyclin-dependent kinase 4 (Cdk4), cathepsin D (Ctsd), keratin complex 2, basic, gene 6a (Krt2-6a), cofilin 1, non-muscle (Cfl1), cyclin 2 (Ccnd2), were verified by real-time quantitative RT-PCR and showed good agreement with results obtained from microarray

Effects of ionizing irradiation and interface backscatter on human mesenchymal stem cells cultured on titanium surfaces

Radiotherapy to the head and neck region negatively influences the osseointegration and survival of dental implants. The effects of cobalt 60 (60Co) ionizing radiation and the impact of backscatter rays were investigated on human mesenchymal stem cells cultured on titanium surfaces. Bone marrow‐derived human mesenchymal stem cells were seeded on titanium (Ti), fluoride‐modified titanium (TiF), and tissue culture plastic. Cells were exposed to ionizing γ ‐radiation in single doses of 2, 6, or 10 Gy using a 60Co source. Density and distribution of cells were evaluated using confocal laser‐scanning microscopy, 21 d post‐irradiation. Lactate dehydrogenase concentration and the levels of total protein and cytokines/chemokines were measured in the cell‐culture medium on days 1, 3, 7, 14, and 21 post‐irradiation. Unirradiated cells were used as the control. Irradiation had no effect on cell viability, collagen and actin expression, or cell distribution, but induced an initial increase in the secretion of interleukin (IL )‐6, IL ‐8, monocyte chemotactic protein 1 (MCP ‐1), and vascular endothelial growth factor (VEGF ), followed by a decrease in secretion after 3 or 7 d. Irradiation resulted in secretion of a lower amount of all analytes examined compared with controls on day 21, irrespective of radiation dose and growth surface. Backscattering from titanium did not influence the cell response significantly, suggesting a clinical potential for achieving successful osseointegration of dental implants placed before radiotherapy

Methacrylation increase growth and differentiation of primary human osteoblasts for gelatin hydrogels

The role of gelatin methacrylate hydrogels with varying degree of methacrylation (69% and 84%) was accessed with FTIR, NMR, microCT and subsequently exposure to human osteoblasts. The cells responded positively to the degree of methacrylation and showed attachment, growth and proliferated on both hydrogels. The cell reacted differently to the degree of methacrylation with higher proliferation on higher substitution; however, cell differentiation behavior was improved for less substitution. The secretion of late osteogenic markers (Osteoprotegerin (OPG), Osteopontin (OPN) and Osteocalcein (OCN)) and angiogenic factor Vascular endothelial growth factor (VEGF) was increased for gelatin methacrylate hydrogels with 69% degree of methacrylation and thus would be the better candidate for future bone regenerative applications amongst the three tested hydrogel

Ameloblastin peptides modulates the osteogenic capacity of human mesenchymal stem cells

During amelogenesis the extracellular enamel matrix protein AMBN is quickly processed into 17 kDa (N-terminus) and 23 kDa (C-terminus) fragments. In particular, alternatively spliced regions derived by exon 5/6 within the N-terminus region are known to be critical in biomineralization. Human mesenchymal stem cells (hMSC) also express and secrete AMBN, but it is unclear if this expression has effects on the hMSC themselves. If, as suggested from previous findings, AMBN act as a signaling molecule, such effects could influence hMSC growth and differentiation, as well as promoting the secretion of other signaling proteins like cytokines and chemokines. If AMBN is found to modulate stem cell behavior and fate, it will impact our understanding on how extracellular matrix molecules can have multiple roles during development ontogenesis, mineralization and healing of mesenchymal tissues. Here we show that synthetic peptides representing exon 5 promote hMSC proliferation. Interestingly, this effect is inhibited by the application of a 15 aa peptide representing the alternatively spliced start of exon 6. Both peptides also influence gene expression of RUNX2 and osteocalcin, and promote calcium deposition in cultures, indicating a positive influence on the osteogenic capacity of hMSC. We also show that the full-length AMBN-WT and N-terminus region enhance the secretion of RANTES, IP-10, and IL-8. In contrast, the AMBN C-terminus fragment and the exon 5 deleted AMBN (DelEx5) have no detectable effects on any of the parameters investigated. These findings suggest the signaling effect of AMBN is conveyed by processed products, whereas the effect on proliferation is differentially modulated through alternative splicing during gene expression