Biomechanical Behaviors and Degradation Properties of Multilayered Polymer Scaffolds: The Phase Space Method for Bile Duct Design and Bioengineering.

This article reports the electrospinning technique for the manufacturing of multilayered scaffolds for bile duct tissue engineering based on an inner layer of polycaprolactone (PCL) and an outer layer either of a copolymer of D,L-lactide and glycolide (PLGA) or a copolymer of L-lactide and ε-caprolactone (PLCL). A study of the degradation properties of separate polymers showed that flat PCL samples exhibited the highest resistance to hydrolysis in comparison with PLGA and PLCL. Irrespective of the liquid-phase nature, no significant mass loss of PCL samples was found in 140 days of incubation. The PLCL- and PLGA-based flat samples were more prone to hydrolysis within the same period of time, which was confirmed by the increased loss of mass and a significant reduction of weight-average molecular mass. The study of the mechanical properties of developed multi-layered tubular scaffolds revealed that their strength in the longitudinal and transverse directions was comparable with the values measured for a decellularized bile duct. The strength of three-layered scaffolds declined significantly because of the active degradation of the outer layer made of PLGA. The strength of scaffolds with the PLCL outer layer deteriorated much less with time, both in the axial (p-value = 0.0016) and radial (p-value = 0.0022) directions. A novel method for assessment of the physiological relevance of synthetic scaffolds was developed and named the phase space approach for assessment of physiological relevance. Two-dimensional phase space (elongation modulus and tensile strength) was used for the assessment and visualization of the physiological relevance of scaffolds for bile duct bioengineering. In conclusion, the design of scaffolds for the creation of physiologically relevant tissue-engineered bile ducts should be based not only on biodegradation properties but also on the biomechanical time-related behavior of various compositions of polymers and copolymers.

Angiogenic Modification of Microfibrous Polycaprolactone by pCMV-VEGF165 Plasmid Promotes Local Vascular Growth after Implantation in Rats.

Insufficient vascular growth in the area of artificial-material implantation contributes to ischemia, fibrosis, the development of bacterial infections, and tissue necrosis around the graft. The purpose of this study was to evaluate angiogenesis after implantation of polycaprolactone microfiber scaffolds modified by a pCMV-VEGF165-plasmid in rats. Influence of vascularization on scaffold degradation was also examined. We investigated flat microfibrous scaffolds obtained by electrospinning polycaprolactone with incorporation of the pCMV-VEGF-165 plasmid into the microfibers at concentrations of 0.005 ng of plasmid per 1 mg of polycaprolactone (0.005 ng/mg) (LCGroup) and 0.05 ng/mg (HCGroup). The samples were subcutaneously implanted in the interscapular area of rats. On days 7, 16, 33, 46, and 64, the scaffolds were removed, and a histological study with a morphometric evaluation of the density and diameter of the vessels and microfiber diameter was performed. The number of vessels was increased in all groups, as well as the resorption of the scaffold. On day 33, the vascular density in the HCGroup was 42% higher compared to the control group (p = 0.0344). The dose-dependent effect of the pCMV-VEGF165-plasmid was confirmed by enhanced angiogenesis in the HCGroup compared to the LCGroup on day 33 (p-value = 0.0259). We did not find a statistically significant correlation between scaffold degradation rate and vessel growth (the Pearson correlation coefficient was ρ = 0.20, p-value = 0.6134). Functionalization of polycaprolactone by incorporation of the pCMV-VEGF165 plasmid provided improved vascularization within 33 days after implantation, however, vessel growth did not seem to correlate with scaffold degradation rate.

Influence of γ-Radiation on Mechanical Stability to Cyclic Loads Tubular Elastic Matrix of the Aorta.

A significant drawback of the rigid synthetic vascular prostheses used in the clinic is the mechanical mismatch between the implant and the prosthetic vessel. When placing prostheses with radial elasticity, in which this deficiency is compensated, the integration of the graft occurs more favorably, so that signs of cell differentiation appear in the prosthesis capsule, which contributes to the restoration of vascular tone and the possibility of vasomotor reactions. Aortic prostheses fabricated by electrospinning from a blend of copolymers of vinylidene fluoride with hexafluoropropylene (VDF/HFP) had a biomechanical behavior comparable to the native aorta. In the present study, to ensure mechanical stability in the conditions of a living organism, the fabricated blood vessel prostheses (BVP) were cross-linked with γ-radiation. An optimal absorbed dose of 0.3 MGy was determined. The obtained samples were implanted into the infrarenal aorta of laboratory animals-Landrace pigs. Histological studies have shown that the connective capsule that forms around the prosthesis has signs of high tissue organization. This is evidenced by the cells of the fibroblast series located in layers oriented along and across the prosthesis, similar to the orientation of cells in a biological arterial vessel.

Non-woven bilayered biodegradable chitosan-gelatin-polylactide scaffold for bioengineering of tracheal epithelium.

OBJECTIVES: The conversion of tissue engineering into a routine clinical tool cannot be achieved without a deep understanding of the interaction between cells and scaffolds during the process of tissue formation in an artificial environment. Here, we have investigated the cultivation conditions and structural features of the biodegradable non-woven material in order to obtain a well-differentiated human airway epithelium. MATERIALS AND METHODS: The bilayered scaffold was fabricated by electrospinning technology. The efficiency of the scaffold has been evaluated using MTT cell proliferation assay, histology, immunofluorescence and electron microscopy. RESULTS: With the use of a copolymer of chitosan-gelatin-poly-l-lactide, a bilayered non-woven scaffold was generated and characterized. The optimal structural parameters of both layers for cell proliferation and differentiation were determined. The basal airway epithelial cells differentiated into ciliary and goblet cells and formed pseudostratified epithelial layer on the surface of the scaffold. In addition, keratinocytes formed a skin equivalent when seeded on the same scaffold. A comparative analysis of growth and differentiation for both types of epithelium was performed. CONCLUSIONS: The structural parameters of nanofibres should be selected experimentally depending on polymer composition. The major challenges on the way to obtain the well-differentiated equivalent of respiratory epithelium on non-woven scaffold include the following: the balance between scaffold permeability and thickness, proper combination of synthetic and natural components, and culture conditions sufficient for co-culturing of airway epithelial cells and fibroblasts. For generation of skin equivalent, the lack of diffusion is not so critical as for pseudostratified airway epithelium.

In vitro assessment of electrospun polyamide-6 scaffolds for esophageal tissue engineering.

Artificial tissue-engineered grafts offer a potential alternative to autologous tissue grafts for patients, which can be traumatic. After decellularizing Papio hamadryas esophagus and studying the morphology and physical properties of the extracellular matrix (ECM), we generated electrospun polyamide-6 based scaffolds to mimic it. The scaffolds supported a greater mechanical load than the native ECM and demonstrated similar 3D microstructure, with randomly aligned fibers, 90% porosity, 29 µm maximal pore size, and average fiber diameter of 2.87 ± 0.95 µm. Biocompatibility studies showed that human adipose- and bone marrow-derived mesenchymal stromal cells (AD-MSC and BMD-MSC) adhered to the scaffold surface and showed some proliferation: scaffold cell coverage was 25% after 72 h of incubation when seeded with 1000 cells/mm2 ; cells elongated processes along the polyamide-6, although they flattened 1.67-4 times less than on cell culture plastic. Human umbilical vein endothelial cells, however, showed poor adherence and proliferation. We thus provide in vitro evidence that polyamide-6 scaffolds approximating the esophageal biomechanics and 3D topography of nonhuman primates may provide a biocompatible substrate for both AD-MSC and BMD-MSCs, supporting their adhesion and survival to some degree. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 107B: 253-268, 2019.

Control on rheological behavior of collagen 1 dispersions for efficient electrospinning.

For efficient manufacturing of fibrous collagen-based materials by electrospinning, the search on optimal rheological parameters is of the great importance. Rheological characteristics and denaturation of collagen in aqueous dispersions were studied as a function of shear rate and acetic acid concentration in the range of 3-9% w/w at temperature from 20 to 40°C. It was shown that an increase in temperature, acetic acid concentration of the collagen dispersion leads to a significant decrease in its viscosity. It was found that helical conformation of the collagen macromolecules is preserved up to 31°C. An increase in acetic acid concentration leads to a reduction of denaturation temperature. The complex viscosity of collagen dispersions exhibits a sharp drop, followed by a rapid growth of damping factor in the temperature range from 22 to 35°C. Both storage (G') and loss (G″) moduli increase with frequency and collagen concentration. It was revealed that optimal parameters for electrospinning of highly concentrated collagen dispersions can be achieved by adjusting of the concentration of acetic acid, temperature, and stirring speed. As a result, collagen nonwoven materials with diameter from 100 to 700 nm were obtained. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 312-318, 2019.