Fabrication of hybrid scaffolds obtained from combinations of PCL with gelatin or collagen via electrospinning for skeletal muscle tissue engineering.

The creation of skeletal muscle tissue in vitro is a major topic of interest today in the field of biomedical research, due to the lack of treatments for muscle loss due to traumatic accidents or disease. For this reason, the intrinsic properties of nanofibrillar structures to promote cell adhesion, proliferation, and cell alignment presents an attractive tool for regenerative medicine to recreate organized tissues such as muscle. Electrospinning is one of the processing techniques often used for the fabrication of these nanofibrous structures and the combination of synthetic and natural polymers is often required to achieve optimal mechanical and physiochemical properties. Here, polycaprolactone (PCL) is selected as a synthetic polymer used for the fabrication of scaffolds, and the effect of protein addition on the final scaffolds' properties is studied. Collagen and gelatin were the proteins selected and two different concentrations were analyzed (2 and 4 wt/vol%). Different PCL/protein systems were prepared, and a structural, mechanical and functional characterization was performed. The influence of fiber alignment on the properties of the final scaffolds was assessed through morphological, mechanical and biological evaluations. A bioreactor was used to promote cell proliferation and differentiation within the scaffolds. The results revealed that protein addition produced a decrease in the fiber size of the membranes, an increase in their hydrophilicity, and a softening of their mechanical properties. The biological study showed the ability of the selected systems to harbor cells, allow their growth and, potentially, develop musculoskeletal tissues.

Electrical Modeling of the Growth and Differentiation of Skeletal Myoblasts Cell Cultures for Tissue Engineering.

In tissue engineering, of utmost importance is the control of tissue formation, in order to form tissue constructs of clinical relevance. In this work, we present the use of an impedance spectroscopy technique for the real-time measurement of the dielectric properties of skeletal myoblast cell cultures. The processes involved in the growth and differentiation of these cell cultures in skeletal muscle are studied. A circuit based on the oscillation-based test technique was used, avoiding the use of high-performance circuitry or external input signals. The effect of electrical pulse stimulation applied to cell cultures was also studied. The technique proved useful for monitoring in real-time the processes of cell growth and estimating the fill factor of muscular stem cells. Impedance spectroscopy was also useful to study the real-time monitoring of cell differentiation, obtaining different oscillation amplitude levels for differentiated and undifferentiated cell cultures. Finally, an electrical model was implemented to better understand the physical properties of the cell culture and control the tissue formation process.

Proof of concept, design, and manufacture via 3-D printing of a mesh with bactericidal capacity: Behaviour in vitro and in vivo.

Currently, hernia treatment involves implantation of a mesh prosthesis, usually made of polypropylene, and the primary complication is infection of the device, which leads to an exponential increase in morbidity. Three-dimensional printing offers a method of dealing with complications of this magnitude. Therefore, in this study, the bactericidal properties and effectiveness of three-dimensional-printed meshes with polycaprolactone (PCL) and gentamicin were evaluated in vitro in Escherichia coli cultures, and their histological behaviour was examined in vivo. Different PCL meshes were implanted into four groups of rats, with 10 rats in each group: PCL meshes, PCL meshes with alginate and calcium chloride, PCL meshes with gentamicin, and PCL meshes with alginate and gentamicin. Thirty-six microporous meshes were manufactured, and their bactericidal properties were assessed. When the meshes did not include an antibiotic, an inhibition halo was not observed; when the gentamicin was free, an asymmetric inhibition area of 5.65 ± 0.46 cm2 was present; when the gentamicin was encapsulated, a rectangular area of 5.40 ± 0.38 cm2 was observed. In the rats, macroporous and microporous mesh implants produced mild inflammation and substantial fibrosis with collagen and neovascular foci. A significant difference was observed in fibroblastic activity between the PCL with alginate group and the PCL with alginate and gentamicin group microporous meshes (p = .013) and in collagen deposits between the macroporous and microporous meshes in the PCL mesh group (p = .033). The feasibility of manufacturing drug-doped printed PCL meshes containing alginate and gentamicin was verified, and the meshes exhibited bactericidal effects and good histopathological behaviour.

Electrical pulse stimulation of skeletal myoblasts cell cultures with simulated action potentials.

Electrical pulse stimulation has an important effect on skeletal muscle development and maturation. However, the methodology for controlling these stimulation parameters to develop in vitro functional skeletal muscle tissues remains to be established. In this work, we have studied the effect of simulated action potentials on the growth and differentiation of skeletal myoblast cell cultures. A circuit simulating action potentials of 0.15 and 0.3 V/mm, at a frequency of 1 Hz and with a 4-ms pulse width, is proposed. Results show an important improvement of the growth rate and differentiation of myoblasts at a voltage of 0.15 V/mm. Parameters such as electrodes geometry or type of signals must be considered in the development of in vitro skeletal muscle.

Local Mesenchymal Stem Cell Therapy in Experimentally Induced Colitis in the Rat.

BACKGROUND: Multipotent mesenchymal stem cells (MSCs) have been used in inflammatory bowel diseases because of their immunomodulatory and regenerative properties. We investigated their local use in an experimental model of colitis in the rat. MATERIALS AND METHODS: Colitis was induced into 20 Wistar rats with local TNBS instillation. Allogeneic stem cells were derived from rat adipose tissue and labeled with PKH2 linker dye with creation of a control and a second group treated by a local injection into the rectal wall of 2×106 allogeneic adipose tissue-derived stem cells (ADSCs). The thicknesses of different components of the rectum were measured with comparisons made in different parts of the colon of the Hunter inflammatory score. PKH2-dyed ADSCs were detected by fluorescence microscopy. RESULTS AND CONCLUSIONS: Total colitis was induced in 19/20 rats with homing of fluorescent ADSCs. to the crypt base and perivascular space of the submucosa. There were no differences in component rectal wall thicknesses with a higher Hunter score in the treated group compared with the controls, in the rectum (3.8±2.74 vs. 1.5±2.37, respectively; p=0.017) and in right colon (2.5±1.08 vs. 0.20±0.42, respectively; p=0.0001). Local colonic injection of allogeneic adipose stem cells. in experimental colitis is feasible and safe. There is demonstrable homing of cells in chemically-induced colitis both to the treated region and parts of the colon distant to the MSC treatment site. Such cells readily proliferate in vitro and could potentially be a source for future treatment of resistant disease.