Monitoring Cell Adhesion on Polycaprolactone-Chitosan Films with Varying Blend Ratios by Quartz Crystal Microbalance with Dissipation.

A detailed understanding of the cell adhesion on polymeric surfaces is required to improve the performance of biomaterials. Quartz crystal microbalance with dissipation (QCM-D) as a surface-sensitive technique has the advantage of label-free and real-time monitoring of the cell-polymer interface, providing distinct signal patterns for cell-polymer interactions. In this study, QCM-D was used to monitor human fetal osteoblastic (hFOB) cell adhesion onto polycaprolactone (PCL) and chitosan (CH) homopolymer films as well as their blend films (75:25 and 25:75). Complementary cell culture assays were performed to verify the findings of QCM-D. The thin polymer films were successfully prepared by spin-coating, and relevant properties, i.e., surface morphology, ζ-potential, wettability, film swelling, and fibrinogen adsorption, were characterized. The adsorbed amount of fibrinogen decreased with an increasing percentage of chitosan in the films, which predominantly showed an inverse correlation with surface hydrophilicity. Similarly, the initial cell sedimentation after 1 h resulted in lesser cell deposition as the chitosan ratio increased in the film. Furthermore, the QCM-D signal patterns, which were measured on the homopolymer and blend films during the first 18 h of cell adhesion, also showed an influence of the different interfacial properties. Cells fully spread on pure PCL films and had elongated morphologies as monitored by fluorescence microscopy and scanning electron microscopy (SEM). Corresponding QCM-D signals showed the highest frequency drop and the highest dissipation. Blend films supported cell adhesion but with lower dissipation values than for the PCL film. This could be the result of a higher rigidity of the cell-blend interface because the cells do not pass to the next stages of spreading after secretion of their extracellular matrix (ECM) proteins. Variations in the QCM-D data, which were obtained at the blend films, could be attributed to differences in the morphology of the films. Pure chitosan films showed limited cell adhesion accompanied by low frequency drop and low dissipation.

Surface analysis of (Ti,Mg)N coated bone fixation devices following the rabbit femur surgery.

BACKGROUND: Magnesium (Mg) enhances the bone regeneration, mineralization and attachment at the tissue/biomaterial interface. OBJECTIVE: In this study, the effect of Mg on mineralization/osseointegration was determined using (Ti,Mg)N thin film coated Ti6Al4V based plates and screws in vivo. METHODS: TiN and (Ti,Mg)N coated Ti6Al4V plates and screws were prepared using arc-PVD technique and used to fix rabbit femur fractures for 6 weeks. Then, mineralization/osseointegration was assessed by surface analysis including cell attachment, mineralization, and hydroxyapatite deposition on concave and convex sides of the plates along with the attachment between the screw and the bone. RESULTS: According to Scanning Electron Microscopy (SEM) and Energy Dispersive Spectroscopy (EDS) analyses; cell attachment and mineralization were higher on the concave sides of the plates from both groups in comparison to the convex sides. However, mineralization was significantly higher on Mg-containing ones. The mean gray value indicating mineralized area after von Kossa staining was found as 0.48 ± 0.01 and 0.41 ± 0.04 on Mg containing and free ones respectively. Similarly, Fourier Transform Infrared Spectroscopy (FTIR) and X-ray diffraction (XRD) analyses showed that hydroxyapatite growth was abundant on the Mg-containing and concave sides of the plates. Enhanced mineralization and strong attachment to bone were also detected in EDS and SEM analyses of Mg-containing screws. CONCLUSION: These findings indicated that (Ti,Mg)N coatings can be used to increase attachment at the implant tissue interface due to accelerated mineralization, cell attachment, and hydroxyapatite growth.

A methionine biomolecule-modified chromenylium-cyanine fluorescent probe for the analysis of Hg2+ in the environment and living cells.

The objective of the study is, for the first time, to construct a new near infrared (NIR) fluorophore, spectrophotometric, colorimetric, ratiometric, and turn-on probe (CSME) based on chromenylium cyanine platform decorated with methionine biomolecule to provide an efficient solution for critical shortcoming to be encountered for analysis of hazardous Hg2+ in environment and living cell. The CSME structure and its interaction with Hg2+ ion were evaluated by NMR, FTIR, MS, UV-Vis and fluorescence methods as well as Density Functional Theory (DFT) calculations. The none fluorescence CSME having spirolactam ring only interacted with Hg2+ in aqueous solution including competing ions. This interaction caused the fluorescence CSME with opened spirolactam form which exhibited spectral and colorimetric changes in the NIR region. The probe based on UV-Vis and fluorescence techniques respond in 90 s, has wide linear ranges (for UV-Vis: 6.29 × 10-8 - 1.86 × 10-4 M; for fluorescence: 9.49 × 10-9 - 1.13 × 10-5 M), and has a lower Limit of Detection (LOD) value (for fluorescence: 4.93 × 10-9 M, 0.99 ng/mL) than the value predicted by the US Environmental Protection Agency (EPA) organization. Hg2+ analysis was performed in drinking and tap water with low Relative Standard Deviation (RSD) values and high recovery. Smartphone and living cell applications were successfully performed for colorimetric sensing Hg2+ in real samples and 3T3 cells, respectively.

Silk Fibroin-Based Shape-Memory Organohydrogels with Semicrystalline Microinclusions.

Inspired by nature, we designed organohydrogels (OHGs) consisting of a silk fibroin (SF) hydrogel as the continuous phase and the hydrophobic microinclusions based on semicrystalline poly(n-octadecyl acrylate) (PC18A) as the dispersed phase. SF acts as a self-emulsifier to obtain oil-in-water emulsions, and hence, it is a versatile and green alternative to chemical emulsifiers. We first prepared a stable oil-in-water emulsion without an external emulsifier by dispersing the n-octadecyl acrylate (C18A) monomer in an aqueous SF solution. To stabilize the emulsions for longer times, gelation in the continuous SF phase was induced by the addition of ethanol, which is known to trigger the conformational transition in SF from random coil to ß-sheet structures. In the second step, in situ polymerization of C18A droplets in the emulsion system was conducted under UV light in the presence of a photoinitiator to obtain high-strength OHGs with shape-memory function, and good cytocompatibility. The incorporation of hydrophilic N,N-dimethylacrylamide and noncrystallizable hydrophobic lauryl methacrylate units in the hydrogel and organogel phases of OHGs, respectively, further improved their mechanical and shape-memory properties. The shape-memory OHGs presented here exhibit switchable viscoelasticity and mechanics, a high Young's modulus (up to 4.3 ± 0.1 MPa), compressive strength (up to 2.5 ± 0.1 MPa), and toughness (up to 0.68 MPa).

A new Fe3+-selective, sensitive, and dual-channel turn-on probe based on rhodamine carrying thiophenecarboxaldehyde: Smartphone application and imaging in living cells.

A new dual-channel probe based on rhodamine B derivative (MSB) was successfully designed, synthesized, characterized by Nuclear Magnetic Resonance (NMR) Spectroscopy, Fourier Transform Infrared Spectrophotometer (FTIR), Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS), X-ray Photoelectron Spectroscopy (XPS), and Single Crystal X-rayDiffraction, and the sensing abilities toward Fe3+ cation have been demonstrated and the probe was successfully utilized for fluorescence imaging of Fe3+ in living cells. The probe demonstrated quite fast, sensitive, and selective response to Fe3+ by causing an extreme enhancement in UV-vis and fluorescence spectroscopy techniques in the buffered aqueous media which makes MSB a dual-channel probe. While the color of MSB solution was initially light yellow, it turned pink in the presence of Fe3+, which provided highly selective naked-eye determination among several ions as alkaline, alkaline-earth, and transition metal ions. After that, the probe was easily applied to paper strips and real samples such as drinking waters and supplementary iron tablets for sensing Fe3+ in an aqueous solution. The detection limit (LOD) and the response time of the probe were determined as 4.85x10-9 M and 4 min, respectively, which are quite lower compared with other rhodamine based Fe3+ sensors in the literature. According to Job's plot, 1H NMR titration, MALDI-TOF MS, XPS, and DFT study techniques, the complexation ratio between MSB and Fe3+ was found as 1:1. Moreover, the spectral response was reversible with alternately addition of Fe3+ or Na2EDTA to the MSB solution. In addition, fluorescence imaging in NIH/3T3 mouse fibroblast cells and studies in real samples with a quite high recovery rate exhibited that the probe is qualified for detection of Fe3+ ion with multiple practical usages.

Heterogeneous pdgfrb+ cells regulate coronary vessel development and revascularization during heart regeneration.

Endothelial cells emerge from the atrioventricular canal to form coronary blood vessels in juvenile zebrafish hearts. We find that pdgfrb is first expressed in the epicardium around the atrioventricular canal and later becomes localized mainly in the mural cells. pdgfrb mutant fish show severe defects in mural cell recruitment and coronary vessel development. Single-cell RNA sequencing analyses identified pdgfrb+ cells as epicardium-derived cells (EPDCs) and mural cells. Mural cells associated with coronary arteries also express cxcl12b and smooth muscle cell markers. Interestingly, these mural cells remain associated with coronary arteries even in the absence of Pdgfrß, although smooth muscle gene expression is downregulated. We find that pdgfrb expression dynamically changes in EPDCs of regenerating hearts. Differential gene expression analyses of pdgfrb+ EPDCs and mural cells suggest that they express genes that are important for regeneration after heart injuries. mdka was identified as a highly upregulated gene in pdgfrb+ cells during heart regeneration. However, pdgfrb but not mdka mutants show defects in heart regeneration after amputation. Our results demonstrate that heterogeneous pdgfrb+ cells are essential for coronary development and heart regeneration.

Detection of cholera toxin with surface plasmon field-enhanced fluorescent spectroscopy.

In this work, a biosensor based on surface plasmon field-enhanced florescence spectroscopy (SPFS) method was successfully constructed to detect the truncated form of cholera toxin, that is, its beta subunit (CTX-B). CTX-B is a relatively small molecule (12 kDa) and it was chosen as model analyte for the detection of protein toxins originated from waterborne pathogens. Recognition layer was prepared on gold-coated LaSFN9 glasses modified with 11-mercaptoundecanoic acid (11-MUA). Biotin-conjugated anti-CTX-B polyclonal antibody (B-Ab) was immobilized on streptavidin (SA) layer constructed on the 11-MUA-modified surface. CTX-B amount was determined with direct assay using B-Ab in surface plasmon resonance (SPR) mode and with sandwich assay in SPFS mode using Cy5-conjugated anti-CTX-B polyclonal antibody. Minimum detected CTX-B concentrations were 10 and 0.01 µg/ml with SPR and SPFS, respectively, showing the sensitivity of the SPFS system over the conventional one. The detection was done in 2-6 h, which was faster than both culture and polymerase chain reaction (PCR)-based methods. Stability tests were performed with SA-coated sensors (excluding B-Ab). In this form, the layer was stable after 30 days of storage in phosphate-buffered saline (PBS; 0.01 M, pH = 7.4) at +4°C. B-Ab layer was formed immediately on them before each measurement.

Defining the Role of Nuclear Factor (NF)-κB p105 Subunit in Human Macrophage by Transcriptomic Analysis of NFKB1 Knockout THP1 Cells.

Since its discovery over 30 years ago the NF-ĸB family of transcription factors has gained the status of master regulator of the immune response. Much of what we understand of the role of NF-ĸB in immune development, homeostasis and inflammation comes from studies of mice null for specific NF-ĸB subunit encoding genes. The role of inflammation in diseases that affect a majority of individuals with health problems globally further establishes NF-ĸB as an important pathogenic factor. More recently, genomic sequencing has revealed loss of function mutations in the NFKB1 gene as the most common monogenic cause of common variable immunodeficiencies in Europeans. NFKB1 encodes the p105 subunit of NF-ĸB which is processed to generate the NF-ĸB p50 subunit. NFKB1 is the most highly expressed transcription factor in macrophages, key cellular drivers of inflammation and immunity. Although a key role for NFKB1 in the control of the immune system is apparent from Nfkb1-/- mouse studies, we know relatively little of the role of NFKB1 in regulating human macrophage responses. In this study we use the THP1 monocyte cell line and CRISPR/Cas9 gene editing to generate a model of NFKB1-/- human macrophages. Transcriptomic analysis reveals that activated NFKB1-/- macrophages are more pro-inflammatory than wild type controls and express elevated levels of TNF, IL6, and IL1B, but also have reduced expression of co-stimulatory factors important for the activation of T cells and adaptive immune responses such as CD70, CD83 and CD209. NFKB1-/- THP1 macrophages recapitulate key observations in individuals with NFKB1 haploinsufficiency including decreased IL10 expression. These data supporting their utility as an in vitro model for understanding the role of NFKB1 in human monocytes and macrophages and indicate that of loss of function NFKB1 mutations in these cells is an important component in the associated pathology.

Defining the structure of the NF-ĸB pathway in human immune cells using quantitative proteomic data.

The NF-ĸB transcription factor is a critical regulator of immune homeostasis and inflammatory responses and is a critical factor in the pathogenesis of inflammatory disease. The pathways to NF-ĸB activation are paradigms for signal-induced ubiquitination and proteasomal degradation, control of transcription factor function by subcellular localisation, and the control of gene transcription and physiological processes by signal transduction mechanisms. Despite the importance of NF-ĸB in disease, the NF-ĸB pathway remains unexploited for the treatment of inflammatory disease. Our understanding of NF-ĸB comes mostly from studies of transgenic mice and cell lines where components of the pathway have been deleted or over expressed. Recent advances in quantitative proteomics offer new opportunities to understand the NF-ĸB pathway using the absolute abundance of individual pathway components. We have analysed available quantitative proteomic datasets to establish the structure of the NF-ĸB pathway in human immune cells under both steady state and activated conditions. This reveals a conserved NF-κB pathway structure across different immune cell lineages and identifies important differences to the current model of the NF-ĸB pathway. These include the findings that the IKK complex in most cells is likely to consist predominantly of IKKß homodimers, that the relative abundancies of IκB proteins show strong cell type variation, and that the components of the non-canonical NF-ĸB pathway are significantly increased in activated immune cells. These findings challenge aspects of our current view of the NF-κB pathway and identify outstanding questions important for defining the role of key components in regulating inflammation and immunity.

Evaluation of natural gum-based cryogels for soft tissue engineering.

In this study, three natural biomaterials, Locust bean gum (LBG), Xanthan gum (XG), and Mastic gum (MG), were combined to form cryogel scaffolds. Thermal and chemical characterizations revealed the successful blend formation from LBG-XG (LX) and LBG-XG-MG (LXM) polymers. All blends resulted in macro-porous scaffolds with interconnected pore structures under the size of 400 µm. The swollen cryogels had similar mechanical properties compared with other polysaccharide-based cryogels. The mean tensile and compressive modulus values of the wet cryogels were in the range of 3.5-11.6 kPa and 82-398 kPa, respectively. The sustained release of the small molecule Kartogenin from varying concentrations and ratios of cryogels was in between 32 and 66% through 21 days of incubation. Physical, mechanical, and chemical properties make LX and LXM polysaccharide-based cryogels promising candidates for cartilage and other soft tissue engineering, and drug delivery applications.

Application potential of three-dimensional silk fibroin scaffold using mesenchymal stem cells for cardiac regeneration.

Cardiac tissue engineering focusing on biomaterial scaffolds incorporating cells from different sources has been explored to regenerate or repair damaged area as a lifesaving approach.The aim of this study was to evaluate the cardiomyocyte differentiation potential of human adipose mesenchymal stem cells (hAD-MSCs) as an alternative cell source on silk fibroin (SF) scaffolds for cardiac tissue engineering. The change in surface morphology of SF scaffolds depending on SF concentration (1-6%, w/v) and increase in their porosity upon application of unidirectional freezing were visualized by scanning electron microscopy (SEM). Swelling ratio was found to increase 2.4 fold when SF amount was decreased from 4% to 2%. To avoid excessive swelling, 4% SF scaffold with swelling ratio of 10% (w/w) was chosen for further studies.Biodegradation rate of SF scaffolds depended on enzymatic activity was found to be 75% weight loss of SF scaffolds at the day 14. The phenotype of hAD-MSCs and their multi-linage potential into chondrocytes, osteocytes, and adipocytes were shown by flow cytometry and immunohistochemical staining, respectively.The viability of hAD-MSCs on 3D SF scaffolds was determined as 90%, 118%, and 138% after 1, 7, and 14 days, respectively. The use of 3D SF scaffolds was associated with increased production of cardiomyogenic biomarkers: α-actinin, troponin I, connexin 43, and myosin heavy chain. The fabricated 3D SF scaffolds were proved to sustain hAD-MSCs proliferation and cardiomyogenic differentiation therefore, hAD-MSCs on 3D SF scaffolds may useful tool to regenerate or repair damaged area using cardiac tissue engineering techniques.

Assessment of bone healing using (Ti,Mg)N thin film coated plates and screws: Rabbit femur model.

Magnesium (Mg) based implants such as plates and screws are often preferred to treat bone defects because of the positive effects of magnesium in bone growth and healing. Their low corrosion resistance, however, leads to fast degradation and consequently failure before healing was completed. Previously, we developed Mg doped titanium nitrate (TiN) thin film coatings to address these limitations and demonstrated that <10 at% Mg doping led to enhanced mineralization in vitro. In the present study, in vivo performance of (Ti,Mg)N coated Ti6Al4V based plates and screws were studied in the rabbit model. Bone fractures were formed on femurs of 16 rabbits and then fixed with either (Ti,Mg)N coated (n = 8) or standard TiN coated (n = 8) plates and screws. X-ray imaging and µCT analyses showed enhanced bone regeneration on fracture sites fixed with (Ti,Mg)N coated plates in comparison with the Mg free ones. Bone mineral density, bone volume, and callus volume were also found to be 11.4, 23.4, and 42.8% higher, respectively, in accordance with µCT results. Furthermore, while TiN coatings promoted only primary bone regeneration, (Ti,Mg)N led to secondary bone regeneration in 6 weeks. These results indicated that Mg presence in the coatings accelerated bone regeneration in the fracture site. (Ti,Mg)N coating can be used as a practical method to increase the efficiency of existing bone fixation devices of varying geometry.

A functional coating to enhance antibacterial and bioactivity properties of titanium implants and its performance in vitro.

Bacterial infections and lack of osseointegration may negatively affect the success of titanium (Ti) implants. In the present study, a functional coating composed of chitosan (CS) microspheres and nano hydroxyapatite (nHA) was prepared to obtain antimicrobial Ti implants with enhanced bioactivity. First, the chitosan microspheres were fixed to Ti surfaces activated by alkali and heat treatment, then nHA coatings were precipitated onto these surfaces. Ciprofloxacin was loaded into the microspheres using two different procedures; encapsulation and diffusion. Scanning electron microscopy micrographs of the modified Ti surfaces showed that the coating was successfully deposited onto the Ti surfaces and stable for 30 days in PBS. The drug was completely released from free microspheres loaded by encapsulation in 21 days whereas only 89% release was observed after immobilization. The burst release also decreased from ca. 55% to ca. 35%. The release was further reduced following the nHA precipitation. The modified Ti surfaces showed antimicrobial activity based on the bacterial time-kill assay using S. aureus, but the efficiency was affected by both nHA precipitation and drug loading strategy. Highest antimicrobial activity was seen in the samples without nHA layer, and when the drug was loaded by diffusion. Fourier transform infrared spectroscopy and X-ray diffraction analyses revealed that nHA on the surface enhanced HA growth in simulated body fluid for 3 weeks, showing increased osseointegration potential. Therefore, the proposed coating may be used to prevent Ti implant failure originated from bacterial infection and/or low bioactivity.

Transcriptional dynamics of pluripotent stem cell-derived endothelial cell differentiation revealed by single-cell RNA sequencing.

AIMS: Pluripotent stem cell-derived endothelial cell products possess therapeutic potential in ischaemic vascular disease. However, the factors that drive endothelial differentiation from pluripotency and cellular specification are largely unknown. The aims of this study were to use single-cell RNA sequencing (scRNA-seq) to map the transcriptional landscape and cellular dynamics of directed differentiation of human embryonic stem cell-derived endothelial cells (hESC-EC) and to compare these cells to mature endothelial cells from diverse vascular beds. METHODS AND RESULTS: A highly efficient directed 8-day differentiation protocol was used to generate a hESC-derived endothelial cell product (hESC-ECP), in which 66% of cells co-expressed CD31 and CD144. We observed largely homogeneous hESC and mesodermal populations at Days 0 and 4, respectively, followed by a rapid emergence of distinct endothelial and mesenchymal populations. Pseudotime trajectory identified transcriptional signatures of endothelial commitment and maturation during the differentiation process. Concordance in transcriptional signatures was verified by scRNA-seq analysis using both a second hESC line RC11, and an alternative hESC-EC differentiation protocol. In total, 105 727 cells were subjected to scRNA-seq analysis. Global transcriptional comparison revealed a transcriptional architecture of hESC-EC that differs from freshly isolated and cultured human endothelial cells and from organ-specific endothelial cells. CONCLUSION: A transcriptional bifurcation into endothelial and mesenchymal lineages was identified, as well as novel transcriptional signatures underpinning commitment and maturation. The transcriptional architecture of hESC-ECP was distinct from mature and foetal human EC.

A multifunctional long-term release system for treatment of hypothyroidism.

Hypothyroidism is an autoimmune disease associated with underactive thyroid gland. In this study, a dual effect polymeric system was designed to release Cepharanthine (CEP) to block T cell activation and Selenium (Se) to decrease the anti-thyroid peroxidase (TPOAb) concentration in order to treat hypothyroidism. For this purpose, poly(ethylene-vinyl acetate) (PEVA) and polyethylene glycol (PEG) nanoparticles (NPs) including CEP were synthesized by emulsion solvent evaporation method and they were loaded to polyurethane (PU)/PEG-PUSe-PEG block copolymer blends which were fabricated by particulate leaching technique as porous sponges. Fourier-Transform Infrared (FTIR), Raman, and Nuclear Magnetic Resonance (NMR) analysis showed successful synthesis of PEG-PUSe-PEG block copolymer. A long-term zero-order release profile was obtained for CEP. Se release rate from matrices showed an oxidative stress-mediated release which can be used to adjust Se amount. According to MTS results conducted by NIH 3T3 fibroblasts, both NPs and matrices have no adverse effect on cell viability. Fluorescence microscopy and SEM images confirm the MTS results. The dual release system has potential to be effectively used in long-term treatment of hypothyroidism by addressing both auto-immune response and hormone regulation.

The function of long non-coding RNAs in vascular biology and disease.

In recent years, it has been revealed that majority of the genome is transcribed in a cell- and context-specific manner into a vast array of RNA transcripts that do not encode proteins. Increasing evidence suggests that non-coding RNAs, especially long non-coding RNAs (lncRNAs) are essential regulators of gene expression and other cellular processes, including in the cardiovascular context. In this review, we discuss lncRNAs and their function during endothelial and vascular smooth cell differentiation, function and homeostasis as well as their role in vessel wall injury response and vascular disease pathophysiology. Although our understanding of lncRNAs is still emerging, these examples reveal important insights on how lncRNAs may ultimately be used in clinic as therapeutic targets for cardiovascular disease.

Peptide-functionalized supported lipid bilayers to construct cell membrane mimicking interfaces.

Supported lipid bilayers (SLB) functionalized with bioactive molecules can be effectively used to study the interaction of cells with different molecules for fundamental research or to develop biosynthetic systems for various biomedical applications. In this study, RGD and Osteocalcin mimetic (OSN) peptides were used as model molecules for functionalization of otherwise passive SLBs to evaluate cell-surface interactions via real-time monitoring in quartz crystal microbalance with dissipation. Similar platforms were also used in cell culture environment. It was seen that low density of mobile RGD peptides on SLB platforms preserved their biological activity and promoted cell adhesion more efficiently than high number of immobile, physisorbed peptides. Even though nonspecific protein and cell attachment was promoted, cells did not spread well on OSN-coated control surfaces. The stability of SLBs produced with different lipids were evaluated in various medium conditions. Enrichment with different lipids increased the stability of SLB to pure PC bilayer.

Magnesium doping on TiN coatings affects mesenchymal stem cell differentiation and proliferation positively in a dose-dependent manner.

BACKGROUND: In vitro evaluation of cell-surface interactions for hard tissue implants have mostly been done using osteoblasts. However, when an implant is placed in the body, mesenchymal stem cells (MSCs) play a major role in new bone formation. Therefore, using MSCs in cell-surface investigations may provide more reliable information on the prediction of in vivo behavior of implants. OBJECTIVE: In this study, Mg doped TiN coatings ((Ti,Mg)N) were prepared and tested for their effect on MSC differentiation and mineralization. METHODS: MSCs were isolated from rat bone marrow (rBMSCs) and seeded onto bare Ti, TiN and Mg containing (Ti,Mg)N surfaces. Cell proliferation, osteogenic differentiation (collagen type 1, alkaline phosphatase activity), calcium phosphate deposition (von Kossa staining, Scanning Electron Microscopy) analysis were conducted. RESULTS: Differentiation towards osteoblast lineage was significantly improved with the increment in Mg presence. Collagen type I deposition, mineralization, and the ALP activity were higher on high Mg containing (>10 at% Mg) surfaces but differentiation of rBMSCs were found to be delayed. CONCLUSIONS: Mg presence affected rBMSCs proliferation and differentiation positively in a dose-dependent manner. However, high Mg amounts delayed both proliferation and differentiation.

Role of STRO-1 sorting of porcine dental germ stem cells in dental stem cell-mediated bone tissue engineering.

Stem cells of dental origin emerged as a new source for the regeneration of tissues with advantages mainly including non-invasive collection procedures and lack of ethical contraversies with their harvest or use. In this study, porcine TGSCs (pTGSCs) were isolated from mandibular third molar tooth germs of 6-month-old domestic pigs. This is the first study that reports the isolation and characterization of TGSCs from porcine third molars and their differentiation depending on STRO-1 expression. PTGSCs were sorted according to their STRO-1 expression as STRO-1(+) and STRO-1(-). Sorted and unsorted heterogenous cells (US) were characterized by their osteogenic, chondrogenic and adipogenic differentiation capabilities. STRO-1(+) cells exhibited a higher proliferation rate owing to their clonogenic properties. All three groups of cells were found differentiated into osteogenic lineage as shown by ALP activity, calcium deposition assay, detection of osteogenic mRNAs and, proteins and mineralization staining. According to differentiation analysis, STRO-1(+) cells did not show a better performance for osteogenesis compared to STRO-1(-) and US cells. This might indicate that STRO-1(+) cells might require a heterogeneous population of cells including STRO-1(-) in their niche to perform their proposed role in osteogenesis.

Quartz crystal microbalance with dissipation as a biosensing platform to evaluate cell-surface interactions of osteoblast cells.

Quartz crystal microbalance with dissipation (QCM-D) is one of the powerful techniques, which allow real time, quantitative and noninvasive analysis of the interaction of different cell types with various modified surfaces. In this study, the dynamic adhesion behavior of human fetal osteoblastic bone (hfOB) cell lines was first monitored on untreated and hydrophilically treated gold sensor surfaces as reference substrates. Adhesion was also observed under light microscopy to facilitate the evaluation. Cells increased their surface contact area and spread more on hydrophilic surfaces, and showed distinct profile with an increased rigidity at the interfacial layer, which is assigned to extracellular matrix remodeling. Further, the adhesion strength and kinetics were characterized on cell adhesive (poly-l-lysine and fibronectin) and repellent (bovine serum albumin) surfaces. The overall results indicated that protein-mediated specific interactions contributed mostly to the dissipation changes (ΔD) or acoustic ratio (ΔD/Δf). Finally, the potential of QCM-D to distinguish healthy and cancerous cells were evaluated by comparing the results of hfOB cells with that of SaOS-2 (osteosarcoma) cancerous cells. Cancerous cells interacted more strongly and showed more viscoelastic characteristic than the healthy cells.