Developing biomimetic cartilaginous tissues that support locomotion while maintaining chondrogenic behavior is a major challenge in the tissue engineering field. Specifically, while locomotive forces ...demand tissues with strong mechanical properties, chondrogenesis requires a soft microenvironment. To address this challenge, 3D cartilage‐like tissue is fabricated using two biomaterials with different mechanical properties: a hard biomaterial to reflect the macromechanical properties of native cartilage, and a soft biomaterial to create a chondrogenic microenvironment. To this end, a bath composed of an interpenetrating polymer network (IPN) of polyethylene glycol (PEG) and alginate hydrogel (MPa order compressive modulus) is developed as an extracellular matrix (ECM) with self‐healing properties. Within this bath supplemented with thrombin, human mesenchymal stem cell (hMSC) spheroids embedded in fibrinogen are 3D bioprinted, creating a soft microenvironment composed of fibrin (kPa order compressive modulus) that simulate cartilage's pericellular matrix and allow a fast diffusion of nutrients. The bioprinted hMSC spheroids present high viability and chondrogenic‐like behavior without adversely affecting the macromechanical properties of the tissue. Therefore, the ability to locally bioprint a soft and cell stimulating biomaterial inside of a mechanically robust hydrogel is demonstrated, thereby uncoupling the micro‐ and macromechanical properties of the 3D printed tissues such as cartilage.
In this work, 3D bioprinting technology is used to develop a biomimetic cartilage‐like tissue with near‐paradoxical mechanical properties, being soft at the cellular level, due to the soft bioink composed of human bone marrow mesenchymal stem cells in the form of spheroids embedded in fibrinogen, and the stiff polyethylene glycol and alginate bath, showing great potential for cartilage regeneration studies.
The aim of this study was to evaluate the skin delivery and in vitro biological activity of trans-resveratrol (RES)-loaded solid lipid nanoparticles (SLNs). The SLNs were composed of stearic acid, ...poloxamer 407, soy phosphatidylcholine (SPC), an aqueous phase and 0.1% RES. The particle size, polydispersity index (PdI) and zeta potential were analyzed by dynamic light scattering (DLS). The SLNs were analyzed by scanning electron microscopy (SEM-FEG) and differential scanning calorimetry (DSC). In vitro RES-SLN skin permeation/retention assays were conducted, and their tyrosinase inhibitory activity was evaluated. An MTT reduction assay was performed on HaCat keratinocytes to determine in vitro cytotoxicity. The formulations had average diameter lower than 200 nm, the addition of SPC promoted increases in PdI in the RES-SLNs, but decreases PdI in the RES-free SLNs and the formulations exhibited zeta potentials smaller than -3 mV. The DSC analysis of the SLNs showed no endothermic peak attributable to RES. Microscopic analysis suggests that the materials formed had nanometric size distribution. Up to 45% of the RES permeated through the skin after 24 h. The RES-loaded SLNs were more effective than kojic acid at inhibiting tyrosinase and proved to be non-toxic in HaCat keratinocytes. The results suggest that the investigated RES-loaded SLNs have potential use in skin disorder therapies.
WAXS characterization of lipid materials to assess the crystallinity, polymorphism and lipid miscibility, of stearic acid or mixtures (70:30) of stearic acid–capric/caprylic triglycerides.
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► For the production of long-term stable lipid nanoparticles, analysis of HLB values allows anticipating the best solid to liquid lipid ratio. ► A suitable surfactant/co-surfactant combination needs to be reached to develop more homogeneous emulsions, with no foaming or phase separation. ► The perfect interaction of hydrophilic and lipophilic emulsifiers on the o/w interface of emulsion is a demand.
There is an increasing interest in lipid nanoparticles because of their suitability for several administration routes. Thus, it becomes even more relevant the physicochemical characterization of lipid materials with respect to their polymorphism, lipid miscibility and stability, as well as the assessment of the effect of surfactant on the type and structure of these nanoparticles. This work focuses on the physicochemical characterization of lipid matrices composed of pure stearic acid or of mixtures of stearic acid–capric/caprylic triglycerides, for drug delivery. The lipids were analyzed by Differential Scanning Calorimetry (DSC), Wide Angle X-ray Diffraction (WAXD), Polarized Light Microscopy (PLM) and hydrophilic–lipophilic balance (HLB) in combination with selected surfactants to determine the best solid-to-liquid ratio. Based on the results obtained by DSC and WAXD, the selected qualitative and quantitative composition contributed for the production of stable nanoparticles, since the melting and the tempering processes provided important information on the thermodynamic stability of solid lipid matrices. The best HLB value obtained for stearic acid–capric/caprylic triglycerides was 13.8, achieved after combining these lipids with accepted surfactants (trioleate sorbitan and polysorbate 80 in the ratio of 10:90). The proposed combinations were shown useful to obtain a stable emulsion to be used as intermediate form for the production of lipid nanoparticles.
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•Stearic acid SLN were optimized by a 22 full factorial design for skin administration.•Stearic acid SLN were of ca. 200 nm mean size, with 0.200 of PdI and ZP values of |26 ...mV|.•Stearic acid SLN were non-toxic to NIH3T3 fibroblasts.
The present study focuses on the characterization of the cytotoxic profile on NIH3T3 mouse embryonic fibroblasts of solid lipid nanoparticles (SLN) optimized by a 22 full factorial design for skin administration. To build up the surface response charts, a design of experiments (DoE) based on 2 independent variables was used to obtain an optimized formulation. The effect of the composition of lipid and water phases on the mean particle size (z-AVE), polydispersity index (PdI) and zeta potential (ZP) was studied. The developed formulations were composed of 5.0% of lipid phase (stearic acid (SA), behenic alcohol (BA) or a blend of SA:BA (1:1)) and 4.7% of surfactants (soybean phosphatidylcholine and poloxamer 407). In vitro cytotoxicity using NIH3T3 fibroblasts was performed by MTT reduction assay. This factorial design study has proven to be a useful tool in optimizing SLN (z-AVE ∼ 200 nm), which were shown to be non-cytotoxic. The present results highlight the benefit of applying statistical designs in the preparation and optimization of SLN formulations.
Agricultural resource derivatives (ARDs) such as hydrolysate soy protein concentrate (HSPC), whey protein concentrate (WPC), and cashew apple juice (CAJ) were studied with focus on the production of ...hyaluronic acid (HA) by
Streptococcus zooepidemicus. Supplementation of the media with corn steep liquor (CSL) was also evaluated. Synthetic medium containing glucose and yeast extract was used as control. CAJ was a promising medium for the production of HA. It produced the highest amount of HA (0.89
g
L
−1), similar to that of the control (0.86
g
L
−1). WPC and HSPC media were the most effective for the production of biomass. CSL did not influence the production of HA when HSPC and WPC were used. However, in the synthetic medium it doubled the yield of HA from glucose. The average molecular weight of HA ranged from 10
3 to 10
4
Da for the ARDs and 10
7
Da for the synthetic medium.
Fibrin networks are obtained through activation of platelet-rich plasma (PRP) for use in tissue regeneration. The importance of fibrin networks relies on mediation of release of growth factors, ...proliferation of tissue cells and rheological properties of the fibrin gels. Activation of PRP usually involves the decomposition of fibrinogen by agonists, in a wide range of concentrations. Therefore fibrin networks with a large structural diversity are formed, making comparative evaluations difficult. In order to standardize the fibrin networks, we used the statistical techniques central composite rotatable design and response-surface analysis, to correlate the radius of the fibers with the ratios between the agonists (autologous serum/calcium chloride) and agonist/PRP. From an individual and interactive analysis of the variables, architectures characterized by thick, medium and thin fibers were delineated on the response-surface. Furthermore, the architectures were correlated with coagulation time. This approach is valuable for standardizing the PRP preparation for clinical applications.
The effects of ethanol/broth proportions and the number of steps at varying pH in the presence or absence of sodium chloride (NaCl) were studied as precipitation strategies for the recovery and ...purification of high molar mass bio-hyaluronic acid (Bio-HA). Bio-HA was synthesized by
Streptococcus zooepidemicus
in a culture medium containing glucose and soy peptones. A single-step precipitation was more attractive than multistep precipitation in terms of recovery and purity as well as decreased use of ethanol. The best conditions in the absence and presence of salt were 2:1 ethanol/broth (
v
/
v
) at pH 4 (55.0 ± 0.2% purity and 85.0 ± 0.7% recovery) and 2:1 ethanol/broth (
v
/
v
) at pH 7 + 2 mol L
−1
NaCl (59.0 ± 0.9% purity and 82.0 ± 4.3% recovery). Dynamic light scattering (DLS) spectra showed different particle sizes as a consequence of the changes in the molecular structure of HA, mainly with changes in pH. Although slight changes in distribution were observed, the average HA molar mass was not affected by the precipitation strategy, remaining on the order of 10
5
Da. Therefore, pH and NaCl modulated the precipitation performance of HA. These findings are relevant to further optimizing the precipitation step, thus minimizing costs in the later stages of HA purification.
Resveratrol (RSV) is a natural antioxidant commonly found in grapes, berries, and nuts that has shown promising results in the treatment of a variety of degenerative and age-related diseases. Despite ...the proven beneficial results on reduction of reactive oxidant species (ROS) and on inflammatory process, RSV shows various limitations including low long-term stability, aqueous solubility, and bioavailability, restricting its applications in the medical-pharmaceutical area. To overcome these limitations, it has been applied in pharmaceutical formulations as nanostructured lipid carriers (NLC). Thus, the present study focuses on the optimization of the production process of NLC. NLC was produced by high shear homogenization (HSH) and ultrasound method (US) using Compritol® ATO C888 as solid lipid and Miglyol 812® as liquid lipid. In order to obtain an optimized formulation, we used a 22 full factorial design with triplicate of central point investigating the effects of the production process parameters; shear intensity and homogenization time, on the mean particle size (PS) and polydispersity index (PDI). Instability index, encapsulation efficiency, and production yield were also evaluated. As the PS and PDI values obtained with 6 min of shear at 19,000 rpm and 10 min of shear and 24,000 rpm were similar, the instability index (<0.1) was also used to select the optimal parameters. Based on the results of the experimental design and instability index, it was concluded that the shear rate of 19,000 rpm and the shear time of 6 min are the optimal parameters for RSV-loaded NLC production. Factorial design contributed therefore to optimize the variables of the NLC production process from a small number of experiments.
Chitosan cross-linked pentasodium tripolyphosphate particles were produced by ionotropic gelation. The aim of this study was to evaluate the influence of the molar mass and deacetylation degree of ...chitosan and of the concentration of pentasodium tripolyphosphate in the production of chitosan micro/nanoparticles. The obtained charge ratio (R±), mean particle size, surface electrical charge, polydispersity index, and tendency of particle aggregation were selected as dependent variables. Results demonstrated that stable particles exhibited a high zeta potential value, between +62 and +68 mV. Particles were produced in different size ranges controlling the R± between the positively charged chitosan and negatively charged pentasodium tripolyphosphate. Chitosan micro/nanoparticles were successfully prepared via the ionic gelation method controlling R±, therefore the association of an active ingredient to a micro/nanoparticle allows the molecule to intimately interact with specific structures, to overcome barriers and to prolong its residence time in the target. Chitosan cross-linked pentasodium tripolyphosphate particles are expected to be a good approach for active ingredients formulation in the agrofood sector and related industries.
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