Antimicrobial activity of ion-substituted calcium phosphates: A systematic review.

In this systematic review, the antimicrobial effect of ion-substituted calcium phosphate biomaterials was quantitatively assessed. The literature was systematically searched up to the 6th of December 2021. Study selection and data extraction was performed in duplo by two independent reviewers with a modified version of the OHAT tool for risk of bias assessment. Any differences were resolved by consensus or by a referee. A mixed effects model was used to investigate the relation between the degree of ionic substitution and bacterial reduction. Of 1016 identified studies, 108 were included in the analysis. The methodological quality of included studies ranged from 6 to 16 out of 18 (average 11.4). Selenite, copper, zinc, rubidium, gadolinium, silver and samarium had a clear antimicrobial effect, with a log reduction in bacteria count of 0.23, 1.8, 2.1, 3.6, 5.8, 7.4 and 10 per atomic% of substitution, respectively. There was considerable between-study variation, which could partially be explained by differences in material formulation, study quality and microbial strain. Future research should focus on clinically relevant scenarios in vitro and the translation to in vivo prevention of PJI.

Site-specific differences in osteoblast phenotype, mechanical loading response and estrogen receptor-related gene expression.

The osteoporosis-resistant nature of skull bones implies inherent differences exist between their cellular responses and those of other osteoporosis-susceptible skeletal sites. Phenotypic differences in calvarial and femoral osteoblastic responses to induction of osteogenesis, mechanical loading, estrogen, growth factor and cytokine stimulation were investigated. Primary rat calvarial and femoral adult male osteoblasts were cultured and osteoblastic mineralisation and maturation determined using Alizarin Red staining and expression of osteogenic marker genes assessed. Expression of known mechanically-responsive genes was compared between sites following loading of scaffold-seeded cells in a bioreactor. Cell proliferation and differentiation following growth factor and estrogen stimulation were also compared. Finally expression of estrogen receptors and associated genes during osteogenic differentiation were investigated. Calvarial osteoblasts exhibited delayed maturation (45d. vs 21d.) and produced less mineralised matrix than femoral osteoblasts when osteogenically induced. PDGF-BB and FGF2 both caused a selective increase in proliferation and decrease in osteoblastic differentiation of femoral osteoblasts. Mechanical stimulation resulted in the induction of the expression of Ccl2 and Anx2a selectively in femoral osteoblasts, but remained unchanged in calvarial cells. Estrogen receptor beta expression was selectively upregulated 2-fold in calvarial osteoblasts. Most interestingly, the estrogen responsive transcriptional repressor RERG was constitutively expressed at 1000-fold greater levels in calvarial compared with femoral osteoblasts. RERG expression in calvarial osteoblasts was down regulated during osteogenic induction whereas upregulation occurred in femoral osteoblasts. Bone cells of the skull are inherently different to those of the femur, and respond differentially to a range of stimuli. These site-specific differences may have important relevance in the development of strategies to tackle metabolic bone disorders.

Chitosan functionalized poly-ε-caprolactone electrospun fibers and 3D printed scaffolds as antibacterial materials for tissue engineering applications.

Tissue engineering (TE) approaches often employ polymer-based scaffolds to provide support with a view to the improved regeneration of damaged tissues. The aim of this research was to develop a surface modification method for introducing chitosan as an antibacterial agent in both electrospun membranes and 3D printed poly-ε-caprolactone (PCL) scaffolds. The scaffolds were functionalized by grafting methacrylic acid N-hydroxysuccinimide ester (NHSMA) onto the surface after Ar-plasma/air activation. Subsequently, the newly-introduced NHS groups were used to couple with chitosan of various molecular weights (Mw). High Mw chitosan exhibited a better coverage of the surface as indicated by the higher N% detected by X-ray photoelectron spectroscopy (XPS) and the observations with either scanning electron microscopy (SEM)(for fibers) or Coomassie blue staining (for 3D-printed scaffolds). A lactate dehydrogenase assay (LDH) using L929 fibroblasts demonstrated the cell-adhesion and cell-viability capacity of the modified samples. The antibacterial properties against S. aureus ATCC 6538 and S. epidermidis ET13 revealed a slower bacterial growth rate on the surface of the chitosan modified scaffolds, regardless the chitosan Mw.

Sprouty 2, an Early Response Gene Regulator of FosB and Mesenchymal Stem Cell Proliferation During Mechanical Loading and Osteogenic Differentiation.

Sprouty 2 (Spry2), an inhibitor of MAP kinase signaling was previously shown by our group to be induced during mechanical loading of mesenchymal stem cells (MSCs). Here, we studied the implication of Spry2 activation during mechanical loading and chemically induced MSC differentiation. Spry 2 expression showed an immediate early response during mechanical loading and chemical induction of osteogenic differentiation and followed the same pattern as osteogenic associated gene FosB and was necessary for the induction of FosB, as Spry 2 knock down also abrogated the upregulation of FosB expression. Spry 2 knock down was, also associated with an early response of the osteogenic genes Runx-2 and ALP. Neither the knock-down of Spry 2 nor the subsequent reduction in FosB had any effect on mid-late osteogenesis or mineralization but was associated with a significant increase in proliferation of MSC. These effects were possibly governed by negative regulation of MEK/Erk signaling as Spry 2 knock down resulted in an increase in phosphorylation of Erk1/2. In summary, our results shows the involvement of Spry2 in regulation of FosB and Runx2 genes, MAPK signaling and proliferation of MSC. Taken together these results suggest a possible role for Spry2 in regulation of MSC functions in response to mechanical loading and osteogenic differentiation. J. Cell. Biochem. 118: 2606-2614, 2017. © 2017 Wiley Periodicals, Inc.

Bonding performance of experimental bioactive/biomimetic self-etch adhesives doped with calcium-phosphate fillers and biomimetic analogs of phosphoproteins.

OBJECTIVES: This study examined the bonding performance and dentin remineralization potential of an experimental adhesive containing calcium-phosphate (Ca/P) micro-fillers, and self-etching primers doped with phosphoprotein biomimetic analogs (polyacrylic acid-(PAA) and/or sodium trimetaphosphate-(TMP)). METHODS: Experimental self-etching primers doped with biomimetic analogs (PAA and/or TMP), and an adhesive containing Ca(2+), PO4(-3)-releasing micro-fillers (Ca/P) were formulated. Sound human dentin specimens were bonded and cut into sticks after aging (24h or 6 months) under simulated pulpal pressure (20cm H2O), and tested for microtensile bond strength (µTBS). Results were analyzed using two-way ANOVA and Tukey's test (p<0.05). Interfacial silver nanoleakage was assessed using SEM. Remineralization of EDTA-demineralized dentin was assessed through FTIR and TEM ultrastructural analysis. RESULTS: Application of the Ca/P-doped adhesive with or without dentin pre-treatments with the primer containing both biomimetic analogs (PAA and TMP) promoted stable µTBS over 6 months. Conversely, µTBS of the control primer and filler-free adhesive significantly decreased after 6 months. Nanoleakage decreased within the resin-dentin interfaces created using the Ca/P-doped adhesives. EDTA-demineralized dentin specimens treated the Ca/P-doped adhesive and the primer containing PAA and TMP showed phosphate uptake (FTIR analysis), as well as deposition of needle-like crystallites at intrafibrillar level (TEM analysis). SIGNIFICANCE: The use of Ca/P-doped self-etching adhesives applied in combination with analogs of phosphoproteins provides durable resin-dentin bonds. This approach may represent a suitable bonding strategy for remineralization of intrafibrillar dentin collagen within the resin-dentin interface.

Di-Calcium Phosphate and Phytosphingosine as an Innovative Acid-Resistant Treatment to Occlude Dentine Tubules.

The present investigation evaluated the ability of an experimental di-calcium phosphate (DCP) desensitising agent used alone or combined with phytosphingosine (PHS) to occlude dentine tubules and resist a citric acid (CA) or artificial saliva (AS) challenge. Three groups of human dentine specimens (DS) were treated with the following: (1) PHS alone, (2) DCP or (3) a combination of PHS and DCP. Dentine hydraulic conductance of DS was evaluated using a digital flow sensor at 6.9 kPa. The average fluid volume for each of the treated DS was used to calculate the total dentine permeability reduction (%P) prior to and following CA immersion for 1 min or AS immersion for 4 weeks. The treated DS were subjected to both scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectroscopy analysis. Statistically significant differences (%P) were identified between the groups by ANOVA and Fisher's multiple comparison test (p < 0.05), respectively. Interestingly, both PHS and DCP appeared to work synergistically. DS treated with DCP or PHS/DCP demonstrated a significant reduction (%P) prior to and following CA or AS challenge (p < 0.05). Both the SEM and FTIR analyses showed consistent brushite crystals occluding the dentine tubules. Conversely, the application of PHS alone failed to demonstrate any significant reduction of dentine permeability (p > 0.05) or show any evidence of occlusion of the dentine tubules. DCP can be used alone or combined with PHS to decrease the dentine permeability as well as to resist a CA and AS challenge. These results would, therefore, suggest that DCP may be a suitable treatment option for dentine hypersensitivity.

In vitro osteoinductive potential of porous monetite for bone tissue engineering.

Tissue engineering-based bone grafts are emerging as a viable alternative treatment modality to repair and regenerate tissues damaged as a result of disease or injury. The choice of the biomaterial component is a critical determinant of the success of the graft or scaffold; essentially, it must induce and allow native tissue integration, and most importantly mimic the hierarchical structure of the native bone. Calcium phosphate bioceramics are widely used in orthopaedics and dentistry applications due to their similarity to bone mineral and their ability to induce a favourable biological response. One such material is monetite, which is biocompatible, osteoconductive and has the ability to be resorbed under physiological conditions. The osteoinductive properties of monetite in vivo are known; however, little is known of the direct effect on osteoinduction of human mesenchymal stem cells in vitro. In this study, we evaluated the potential of monetite to induce and sustain human mesenchymal stem cells towards osteogenic differentiation. Human mesenchymal stem cells were seeded on the monetite scaffold in the absence of differentiating factors for up to 28 days. The gene expression profile of bone-specific markers in cells on monetite scaffold was compared to the control material hydroxyapatite. At day 14, we observed a marked increase in alkaline phosphatase, osteocalcin and osteonectin expressions. This study provides evidence of a suitable material that has potential properties to be used as a tissue engineering scaffold.

Gene expression responses to mechanical stimulation of mesenchymal stem cells seeded on calcium phosphate cement.

INTRODUCTION: The aim of the study reported here was to investigate the molecular responses of human mesenchymal stem cells (MSC) to loading with a model that attempts to closely mimic the physiological mechanical loading of bone, using monetite calcium phosphate (CaP) scaffolds to mimic the biomechanical properties of bone and a bioreactor to induce appropriate load and strain. METHODS: Human MSCs were seeded onto CaP scaffolds and subjected to a pulsating compressive force of 5.5±4.5 N at a frequency of 0.1 Hz. Early molecular responses to mechanical loading were assessed by microarray and quantitative reverse transcription-polymerase chain reaction and activation of signal transduction cascades was evaluated by western blotting analysis. RESULTS: The maximum mechanical strain on cell/scaffolds was calculated at around 0.4%. After 2 h of loading, a total of 100 genes were differentially expressed. The largest cluster of genes activated with 2 h stimulation was the regulator of transcription, and it included FOSB. There were also changes in genes involved in cell cycle and regulation of protein kinase cascades. When cells were rested for 6 h after mechanical stimulation, gene expression returned to normal. Further resting for a total of 22 h induced upregulation of 63 totally distinct genes that were mainly involved in cell surface receptor signal transduction and regulation of metabolic and cell division processes. In addition, the osteogenic transcription factor RUNX-2 was upregulated. Twenty-four hours of persistent loading also markedly induced osterix expression. Mechanical loading resulted in upregulation of Erk1/2 phosphorylation and the gene expression study identified a number of possible genes (SPRY2, RIPK1, SPRED2, SERTAD1, TRIB1, and RAPGEF2) that may regulate this process. CONCLUSION: The results suggest that mechanical loading activates a small number of immediate-early response genes that are mainly associated with transcriptional regulation, which subsequently results in activation of a wider group of genes including those associated with osteoblast proliferation and differentiation. The results provide a valuable insight into molecular events and signal transduction pathways involved in the regulation of MSC osteogenic differentiation in response to a physiological level of mechanical stimulation.

Preparation of multicompartment sub-micron particles using a triple-needle electrohydrodynamic device.

Control over the size and morphology of polymeric carriers for drug delivery systems is essential to optimize their functionality. In the current study, we demonstrate the feasibility of using an electrohydrodynamic process with a triple-needle device to prepare nearly mono-dispersed, spherical, tri-layered sub-micron particles. Three biocompatible polymer solutions of poly (lactic-co-glycolic acid) (PLGA), polycaprolactone (PCL) and polymethylsilsesquioxane (PMSQ) were used to prepare particles with three distinct layers. Optimized particles were shown to be spherical with an average size ranging from 320 nm (±80 nm) to 220 (±8 nm), which varied with a change in the working distance in the electrohydrodynamic processing. The surface and internal structure and morphology were studied using confocal, transmission and scanning electron microscopy combined with focused ion beam sectioning. Cytotoxicity was shown to be negligible in an in vitro assay. The ability to fabricate such multilayered particles in a single step, under ambient conditions has considerable potential for a range of applications in particular controlled release drug delivery system.

Remineralisation properties of innovative light-curable resin-based dental materials containing bioactive micro-fillers.

Innovative dental restorative materials should be highly bioactive to induce therapeutic effects at the bonding interface during intimate contact with biological fluids. This study aimed at evaluating the remineralisation properties of innovative light-curable resin-based dental materials containing bioactive calcium-phosphosilicates micro-fillers. The apatite formation ability was assessed by ATR-FTIR, XRD, calcium-chelation dye-assisted confocal microscopy (CLSM) and SEM analysis after soaking in SBF. Changes in transition glass temperature (Tg), water sorption/solubility, alkalinising activity (pH) Knoop micro-hardness (KHN), were also evaluated. Four experimental resin-based materials containing various silicate-base micro-fillers and a control filler-free resin blend were formulated. Disc-shaped specimens were polymerised and submitted to pH/alkalinising activity and water sorption/solubility analyses. The bioactivity and the apatite precipitation induced by the tested materials were evaluated through ATR/FTIR vibrational analysis, SEM and XRD analyses subsequent to SBF storage. Knoop micro-hardness (KHN) and differential scanning calorimetry (DSC) were also performed. The experimental resins containing the bioactive micro-fillers were able to induce apatite precipitation subsequent to prolonged SBF storage (30-90 days). However, the resins containing the bioactive ZnO/polycarboxylated micro-fillers (BAG-Zn; ßTCS-Zn) showed the lowest water sorption and solubility mass changes (P < 0.05). A significant increase of the KHN and Tg subsequent prolonged SBF storage was also observed. The use of such light-curable bioactive materials in restorative dentistry might represent a potentially therapeutic approach to increase the longevity of the restorations via apatite deposition the mineral-depleted tissues at the bonding.