Biomineralization is mediated by extracellular matrix (ECM) proteins with amino acid sequences rich in glutamic acid. The objective of this study was to investigate the effect of calcium phosphate deposition on aligned nanofibres surface-modified with a glutamic acid peptide on osteogenic differentiation of rat marrow stromal cells. Blend of EEGGC peptide (GLU) conjugated low molecular weight polylactide (PLA) and high molecular weight poly(lactide-co-glycolide) (PLGA) was electrospun to form aligned nanofibres (GLU-NF). The GLU-NF microsheets were incubated in a modified simulated body fluid for nucleation of calcium phosphate crystals on the fibre surface. To achieve a high calcium phosphate to fibre ratio, a layer-by-layer approach was used to improve diffusion of calcium and phosphate ions inside the microsheets. Based on dissipative particle dynamics simulation of PLGA/PLA-GLU fibres, > 80% of GLU peptide was localized to the fibre surface. Calcium phosphate to fibre ratios as high as 200%, between those of cancellous (160%) and cortical (310%) bone, was obtained with the layer-by-layer approach. The extent of osteogenic differentiation and mineralization of marrow stromal cells seeded on GLU-NF microsheets was directly related to the amount of calcium phosphate deposition on the fibres prior to cell seeding. Expression of osteogenic markers osteopontin, alkaline phosphatase (ALP), osteocalcin and type 1 collagen increased gradually with calcium phosphate deposition on GLU-NF microsheets. Results demonstrate that surface modification of aligned synthetic nanofibres with EEGGC peptide dramatically affects nucleation and growth of calcium phosphate crystals on the fibres leading to increased osteogenic differentiation of marrow stromal cells and mineralization.

Articular cartilage is organized into multiple zones including superficial, middle and calcified zones with distinct cellular and extracellular components to impart lubrication, compressive strength, and rigidity for load transmission to bone, respectively. During native cartilage tissue development, changes in biochemical, mechanical, and cellular factors direct the formation of stratified structure of articular cartilage. The objective of this work was to investigate the effect of combined gradients in cell density, matrix stiffness, and zone-specific growth factors on the zonal organization of articular cartilage. Human mesenchymal stem cells (hMSCs) were encapsulated in acrylate-functionalized lactide-chain-extended polyethylene glycol (SPELA) gels simulating cell density and stiffness of the superficial, middle and calcified zones. The cell-encapsulated gels were cultivated in medium supplemented with growth factors specific to each zone and the expression of zone-specific markers was measured with incubation time. Encapsulation of 60×106 cells/mL hMSCs in a soft gel (80 kPa modulus) and cultivation with a combination of TGF-β1 (3 ng/mL) and BMP-7 (100 ng/mL) led to the expression of markers for the superficial zone. Conversely, encapsulation of 15×106 cells/mL hMSCs in a stiff gel (320 MPa modulus) and cultivation with a combination of TGF-β1 (30 ng/mL) and hydroxyapatite (3%) led to the expression of markers for the calcified zone. Further, encapsulation of 20×106 cells/mL hMSCs in a gel with 2.1 MPa modulus and cultivation with a combination of TGF-β1 (30 ng/mL) and IGF-1 (100 ng/mL) led to up-regulation of the middle zone markers. Results demonstrate that a developmental approach with gradients in cell density, matrix stiffness, and zone-specific growth factors can potentially regenerate zonal structure of the articular cartilage.

Degradable, in situ gelling, inert hydrogels with tunable properties are very attractive as a matrix for cell encapsulation and delivery to the site of regeneration. Cell delivery is generally limited by the toxicity of gelation and degradation reactions. The objective of this work was to investigate by simulation and experimental measurement gelation kinetics and degradation rate of star acrylated polyethylene glycol (PEG) macromonomers chain-extended with short hydroxy acid (HA) segments (SPEXA) as a function of HA monomer type and number of HA repeat units. HA monomers included least hydrophobic glycolide (G), lactide (L), p-dioxanone (D), and most hydrophobic ε-caprolactone (C). Chain extension of PEG with short HA segments resulted in micelle formation for all HA types. There was a significant decrease in gelation time of SPEXA precursor solutions with HA chain-extension for all HA types due to micelle formation, consistent with the simulated increase in acrylate-acrylate (Ac-Ac) and Ac-initiator integration numbers. The hydrolysis rate of SPEXA hydrogels was strongly dependent on HA type and number of HA repeat units. SPEXA gels chain-extended with the least hydrophobic glycolide completely degraded within days, lactide within weeks, and p-dioxanone and ε-caprolactone degraded within months. The wide range of degradation rates observed for SPEXA gels can be explained by large differences in equilibrium water content of the micelles for different HA monomer types. A biphasic relationship between HA segment length and gel degradation rate was observed for all HA monomers, which was related to the transition from surface (controlled by HA segment length) to bulk (controlled by micelle equilibrium water content) hydrolysis within the micelle phase. To our knowledge, this is the first report on transition from surface to bulk degradation at the nanoscale in hydrogels.

The objective of this work was to investigate the effect of chemical composition and segment number (n) on gelation, stiffness, and degradation of hydroxy acid-chain-extended star polyethylene glycol acrylate (SPEXA) gels. The hydroxy acids included glycolide (G,), L-lactide (L), p-dioxanone (D) and -caprolactone (C). Chain-extension generated water soluble macromers with faster gelation rates, lower sol fractions, higher compressive moduli, and a wide-ranging degradation times when crosslinked into a hydrogel. SPEGA gels with the highest fraction of inter-molecular crosslinks had the most increase in compressive modulus with n whereas SPELA and SPECA had the lowest increase in modulus. SPEXA gels exhibited a wide range of degradation times from a few days for SPEGA to a few weeks for SPELA, a few months for SPEDA, and many months for SPECA. Marrow stromal cells and endothelial progenitor cells had the highest expression of vasculogenic markers when co-encapsulated in the faster degrading SPELA gel.

Optimization of nanofiber (NF) surface properties is critical to achieve an adequate cellular response. Here, the impact of conjugation of biomimetic aspartic acid (ASP) and glutamic acid (GLU) templated peptides with poly(lactic-co-glycolic acid) (PLGA) electrospun NF on osteogenic differentiation of human bone marrow-derived mesenchymal stem cells (hMSCs) was evaluated. Cold atmospheric plasma (CAP) was used to functionalize the NF surface and thus to mediate the conjugation. The influence of the CAP treatment following with peptide conjugation to the NF surface was assessed using water contact angle measurements, Fourier-Transform Infrared Spectroscopy (FTIR) and X-ray Photoelectron Spectroscopy (XPS). The effect of CAP treatment on morphology of NF was also checked using Scanning Electron Microscopy (SEM). Both the hydrophilicity of NF and the number of the carboxyl (-COOH) groups on the surface increased with respect to CAP treatment. Results demonstrated that CAP treatment significantly enhanced peptide conjugation on the surface of NF. Osteogenic differentiation results indicated that conjugating of biomimetic ASP templated peptides sharply increased alkaline phosphatase (ALP) activity, calcium content, and expression of key osteogenic markers of collagen type I (Col-I), osteocalcin (OC), and osteopontin (OP) compared to GLU conjugated (GLU-pNF) and CAP treated NF (pNF). It was further depicted that ASP sequences are the major fragments that influence the mineralization and osteogenic differentiation in non-collagenous proteins of bone extracellular matrix.

The aim of this study was to investigate the synergistic effect of cold atmospheric plasma (CAP) treatment and RGD peptide coating for enhancing cellular attachment and proliferation over titanium (Ti) surfaces. The surface structure of CAP-treated and RGD peptide-coated Ti discs were characterized by contact angle goniometer and atomic force microscopy. The effect of such surface modification on human bone marrow derived mesenchymal stem cells (hMSCs) adhesion and proliferation was assessed by cell proliferation and DNA content assays. Besides, hMSCs' adhesion and morphology on surface modified Ti discs were observed via fluorescent and scanning electron microscopy. RGD peptide coating following CAP treatment significantly enhanced cellular adhesion and proliferation among untreated, CAP-treated and RGD peptide-coated Ti discs. The treatment of Ti surfaces with CAP may contribute to improved RGD peptide coating, which enables increased cellular integrations with the Ti surfaces.

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Surgical site infections have a remarkable impact on morbidity, extended hospitalization and mortality. Sutures strongly contribute to development of surgical site infections as they are considered foreign material in the human body. Sutures serve as excellent surfaces for microbial adherence and subsequent colonization, biofilm formation and infection on the site of a surgery. Various antimicrobial sutures have been developed to prevent suture-mediated surgical site infection. However, depending on the site of surgery, antimicrobial sutures may remain ineffective, and antimicrobial agents on them might have drawbacks. Plasma, defined as the fourth state of matter, composed of ionized gas, reactive oxygen and nitrogen species, free radical and neutrals, draws attention for the control and prevention of hospital-acquired infections due to its excellent antimicrobial activities. In the present study, the efficacy of non-thermal atmospheric plasma treatment for prevention of surgical site infections was investigated. First, contaminated poly (glycolic-co-lactic acid), polyglycolic acid, polydioxanone and poly (glycolic acid-co-caprolactone) sutures were treated with non-thermal atmospheric plasma to eradicate contaminating bacteria like Staphylococcus aureus and Escherichia coli. Moreover, sutures were pre-treated with non-thermal atmospheric plasma and then exposed to S. aureus and E. coli. Our results revealed that non-thermal atmospheric plasma treatment effectively eradicates contaminating bacteria on sutures, and non-thermal atmospheric plasma pre-treatment effectively prevents bacterial colonization on sutures without altering their mechanical properties. Chemical characterization of sutures was performed with FT-IR and XPS and results showed that non-thermal atmospheric plasma treatment substantially increased the hydrophilicity of sutures which might be the primary mechanism for the prevention of bacterial colonization. In conclusion, plasma-treated sutures could be considered as novel alternative materials for the control and prevention of surgical site infections.