Enzyme prodrug therapy achieves site-specific, personalized physiological responses to the locally produced nitric oxide

Nitric oxide (NO) is a highly potent but short-lived endogenous radical with a wide spectrum of physiological activities. In this work, we developed an enzymatic approach to the site-specific synthesis of NO mediated by biocatalytic surface coatings. Multilayered polyelectrolyte films were optimized as host compartments for the immobilized β-galactosidase (β-Gal) enzyme through a screen of eight polycations and eight polyanions. The lead composition was used to achieve localized production of NO through the addition of β-Gal–NONOate, a prodrug that releases NO following enzymatic bioconversion. The resulting coatings afforded physiologically relevant flux of NO matching that of the healthy human endothelium. The antiproliferative effect due to the synthesized NO in cell culture was site-specific: within a multiwell dish with freely shared media and nutrients, a 10-fold inhibition of cell growth was achieved on top of the biocatalytic coatings compared to the immediately adjacent enzyme-free microwells. The physiological effect of NO produced via the enzyme prodrug therapy was validated ex vivo in isolated arteries through the measurement of vasodilation. Biocatalytic coatings were deposited on wires produced using alloys used in clinical practice and successfully mediated a NONOate concentration-dependent vasodilation in the small arteries of rats. The results of this study present an exciting opportunity to manufacture implantable biomaterials with physiological responses controlled to the desired level for personalized treatment

Bioinspired Nanofeatured Substrates: Suitable Environment for Bone Regeneration.

Bone mimicking coatings provide a complex microenvironment in which material, through its inherent properties (such as nanostructure and composition), affects the commitment of stem cells into bone lineage and the production of bone tissue regulating factors required for bone healing and regeneration. Herein, a bioactive mineral/biopolymer composite made of calcium phosphate/chitosan and hyaluronic acid (CaP-CHI-HA) was elaborated using a versatile simultaneous spray coating of interacting species. The resulting CaP-CHI-HA coating was mainly constituted of bioactive, carbonated and crystalline hydroxyapatite with 277 ± 98 nm of roughness, 1 μm of thickness, and 2.3 ± 1 GPa of stiffness. After five days of culture, CaP-CHI-HA suggested a synergistic effect of intrinsic biophysical features and biopolymers on stem cell mechanobiology and nuclear organization, leading to the expression of an early osteoblast-like phenotype and the production of bone tissue regulating factors such as osteoprotegerin and vascular endothelial growth factor. More interestingly, amalgamation with biopolymers conferred to the mineral a bacterial antiadhesive property. These significant data shed light on the potential regenerative application of CaP-CHI-HA bioinspired coating in providing a suitable environment for stem cell bone regeneration and an ideal strategy to prevent implant-associated infections.journal article2017 Apr 122017 03 30importe

Modulation of Cellular Colonization of Porous Polyurethane Scaffolds via the Control of Pore Interconnection Size and Nanoscale Surface Modifications.

Full-scale cell penetration within porous scaffolds is required to obtain functional connective tissue components in tissue engineering applications. For this aim, we produced porous polyurethane structures with well-controlled pore and interconnection sizes. Although the influence of the pore size on cellular behavior is widely studied, we focused on the impact of the size of the interconnections on the colonization by NIH 3T3 fibroblasts and Wharton's jelly-derived mesenchymal stem cells (WJMSCs). To render the material hydrophilic and allow good material wettability, we treated the material either by plasma or by polydopamine (PDA) coating. We show that cells weakly adhere on these surfaces. Keeping the average pore diameter constant at 133 μm, we compare two structures, one with LARGE (52 μm) and one with SMALL (27 μm) interconnection diameters. DNA quantification and extracellular matrix (ECM) production reveal that larger interconnections is more suitable for cells to move across the scaffold and form a three-dimensional cellular network. We argue that LARGE interconnections favor cell communication between different pores, which then favors the production of the ECM. Moreover, PDA treatment shows a truly beneficial effect on fibroblast viability and on matrix production, whereas plasma treatment shows the same effect for WJMSCs. We, therefore, claim that both pore interconnection size and surface treatment play a significant role to improve the quality of integration of tissue engineering scaffolds.journal article2019 Jun 052019 05 24importe

Harnessing Wharton's jelly stem cell differentiation into bone-like nodule on calcium phosphate substrate without osteoinductive factors.

An important aim of bone regenerative medicine is to design biomaterials with controlled chemical and topographical features to guide stem cell fate towards osteoblasts without addition of specific osteogenic factors. Herein, we find that sprayed bioactive and biocompatible calcium phosphate substrates (CaP) with controlled topography induce, in a well-orchestrated manner, Wharton's jelly stem cells (WJ-SCs) differentiation into osteoblastic lineage without any osteogenic supplements. The resulting WJ-SCs commitment exhibits features of native bone, through the formation of three-dimensional bone-like nodule with osteocyte-like cells embedded into a mineralized type I collagen. To our knowledge, these results present the first observation of a whole differentiation process from stem cell to osteocytes-like on a synthetic material. This suggests a great potential of sprayed CaP and WJ-SCs in bone tissue engineering. These unique features may facilitate the transition from bench to bedside and the development of successful engineered bone.Designing materials to direct stem cell fate has a relevant impact on stem cell biology and provides insights facilitating their clinical application in regenerative medicine. Inspired by natural bone compositions, a friendly automated spray-assisted system was used to build calcium phosphate substrate (CaP). Sprayed biomimetic solutions using mild conditions led to the formation of CaP with controlled physical properties, good bioactivity and biocompatibility. Herein, we show that via optimization of physical properties, CaP substrate induce osteogenic differentiation of Wharton's jelly stem cells (WJ-SCs) without adding osteogenic supplement factors. These results suggest a great potential of sprayed CaP and WJ-SCs in bone tissue engineering and may facilitate the transition from bench to beside and the development of clinically successful engineered bone.journal articleresearch support, non-u.s. gov't2017 022016 11 22importe

Polysaccharide films built by simultaneous or alternate spray: a rapid way to engineer biomaterial surfaces.

We investigated polysaccharide films obtained by simultaneous and alternate spraying of a chitosan (CHI) solution as polycation and hyaluronic acid (HA), alginate (ALG), and chondroitin sulfate (CS) solutions as polyanions. For simultaneous spraying, the film thickness increases linearly with the cumulative spraying time and passes through a maximum for polyanion/CHI molar charge ratios lying between 0.6 and 1.2. The size of polyanion/CHI complexes formed in solution was compared with the simultaneously sprayed film growth rate as a function of the polyanion/CHI molar charge ratio. A good correlation was found. This suggests the importance of polyanion/polycation complexation in the simultaneous spraying process. Depending on the system, the film topography is either liquid-like or granular. Film biocompatibility was evaluated using human gingival fibroblasts. A small or no difference is observed in cell viability and adhesion between the two deposition processes. The CHI/HA system appears to be the best for cell adhesion inducing the clustering of CD44, a cell surface HA receptor, at the membrane of cells. Simultaneous or alternate spraying of CHI/HA appears thus to be a convenient and fast procedure for biomaterial surface modifications.journal articleresearch support, non-u.s. gov't2012 Jun 052012 05 23importe

O2 Level Controls Hematopoietic Circulating Progenitor Cells Differentiation into Endothelial or Smooth Muscle Cells

BACKGROUND:Recent studies showed that progenitor cells could differentiate into mature vascular cells. The main physiological factors implicated in cell differentiation are specific growth factors. We hypothesized that simply by varying the oxygen content, progenitor cells can be differentiated either in mature endothelial cells (ECs) or contractile smooth muscle cells (SMCs) while keeping exactly the same culture medium. METHODOLOGY/PRINCIPAL FINDINGS:Mononuclear cells were isolated by density gradient were cultivated under hypoxic (5% O2) or normoxic (21% O2) environment. Differentiated cells characterization was performed by confocal microscopy examination and flow cytometry analyses. The phenotype stability over a longer time period was also performed. The morphological examination of the confluent obtained cells after several weeks (between 2 and 4 weeks) showed two distinct morphologies: cobblestone shape in normoxia and a spindle like shape in hypoxia. The cell characterization showed that cobblestone cells were positive to ECs markers while spindle like shape cells were positive to contractile SMCs markers. Moreover, after several further amplification (until 3(rd) passage) in hypoxic or normoxic conditions of the previously differentiated SMC, immunofluorescence studies showed that more than 80% cells continued to express SMCs markers whatever the cell environmental culture conditions with a higher contractile markers expression compared to control (aorta SMCs) signature of phenotype stability. CONCLUSION/SIGNIFICANCE:We demonstrate in this paper that in vitro culture of peripheral blood mononuclear cells with specific angiogenic growth factors under hypoxic conditions leads to SMCs differentiation into a contractile phenotype, signature of their physiological state. Moreover after amplification, the differentiated SMC did not reverse and keep their contractile phenotype after the 3rd passage performed under hypoxic and normoxic conditions. These aspects are of the highest importance for tissue engineering strategies. These results highlight also the determinant role of the tissue environment in the differentiation process of vascular progenitor cells

Geopolymers from Algerian metakaolin. Influence of secondary minerals

The influence of secondary phases (illite, quartz) on the geopolymerization reaction of metakaolin has been investigated by comparing two metakaolins, one prepared from a pure kaolinite and the other from illite- and quartz-containing Algerian kaolin from the Tamazert region, respectively. Geopolymerization was achieved by mixing the metakaolins with an alkaline sodium silicate solution at room temperature and curing at 50 °C. The products were characterized by X-ray diffraction and 29Si and 27Al MAS-NMR. The results show that the secondary phases, at the concentration used in this work, do not prevent the geopolymerization reactio

Thermal behaviour of kaolin of tamazert (Algeria) deposit

Kaolin from Tamazert deposit in the north Algeria was used in this study. The physical and chemical properties were determined by the performance of several analyses as well as the X-ray fluorescence, X-ray diffraction, thermal behaviour (ATD, TG), dilatometric and granulometric distribution. The study of ceramic behaviour as function of firing temperature has been investigated by shrinkage, adsorption both density and mechanical tests after gradually increasing from room temperature to 1500 °C. The modification of the structure of the raw material samples has been observed after fining and the crystallization of mullite and amorphous silica phase were confirmed by scanning microscopy SEM and evaluated by XRD. The suitability of kaolin for ceramics process was discussed from the linear shrinkage, both density and open porosity. The amorphous phase content is varied from 27-34 % and the tests of flexion have been carried out as depending of temperature, revealed a suitable mechanical properties and the relationship between amorphous phase and sample properties. The adsorption rate was found to be about 35

Structural characterization of mullite formed from heated kaolin of tamazert deposit (Algeria)

A quantitative analysis of the mullite phase obtained after sintering of the kaolin has been performed by treatment of X ray diagrams. Kaolin is treated in the range of 950-1400 *deg;C. The oxide NiO is added as internal standard after cooling to sintered sample kaolin. Bragg Brentano diffractometer, scanning electron microscopy and differential thermal analysis were used. Analyses of selected directions profile of mullite were carried out by using the adjustment of program, available in the software Highscore of Pan Analytical. The mullite phase that formed from kaolin appears at 1000 °C, observed by XRD and tallies with DTA. The primary mullite crystal showed a plate-like morphology. Two kinds of morphology corresponding to primary (elongated grains) and secondary (equiaxed grains) mullite were observed. A bimodal crystallite size distribution was detected through XRD microstructural analysis from 1300 °C. The apparent sizes obtained of crystallites are determined for mullite with directions-dependent (anisotropy