Bone marrow derived progenitor cells have been widely studied for its multipotent property and have proofed to be an important resource in regenerative medicine. However, the propagation of murine bone marrow appeared to be a great challenge as compared to other mammalian species. In this study, various isolation techniques and the plasticity of the isolated cells were evaluated. Our result shows that magnetic sorting technique yielded the most viable cells and displayed wider differentiation capacity.
The repair of osteochondral defects requires a tissue engineering approach that aims at mimicking the physiological properties and structure of two different tissues (cartilage and bone) using specifically designed scaffold-cell constructs. Biphasic and triphasic approaches utilize two or three different architectures, materials, or composites to produce a multilayered construct. This article gives an overview of some of the current strategies in multiphasic/gradient-based scaffold architectures and compositions for tissue engineering of osteochondral defects. In addition, the application of finite element analysis (FEA) in scaffold design and simulation of in vitro and in vivo cell growth outcomes has been briefly covered. FEA-based approaches can potentially be coupled with computer-assisted fabrication systems for controlled deposition and additive manufacturing of the simulated patterns. Finally, a summary of the existing challenges associated with the repair of osteochondral defects as well as some recommendations for future directions have been brought up in the concluding section of this article.
A novel decellularization method using sonication treatment is described. Sonication treatment is the combination of physical and chemical agents. These methods will disrupt cell membrane and release cell contents to external environments. The cell removal was facilitated by subsequent rinsing of sodium dodecyl sulfate detergents. Sonication treatment is used in the preparation of complete decellularized bioscaffolds. The aim of this study is to confirm the usefulness of sonication treatment for preparation of biological scaffolds. In this study, samples of aortic tissues are decellularized by sonication treatment at frequency of 170 kHz in 0.1% and 2% sodium dodecyl sulfate detergents for 10-h treatment time. The relation between decellularization and sonication parameters such as dissolved oxygen concentration, conductivity, and pH is investigated. Histological analysis and biomechanical testing is performed to evaluate cell removal efficiency as well as changes in biomechanical properties. Minimal inflammation response elicit by bioscaffolds is confirmed by xenogeneic implantation and immunohistochemistry. Sonication treatment is able to produce complete decellularized tissue suggesting that these treatments could be applied widely as one of the decellularization method.
Biodegradable elastomers have clinical applicability due to their biocompatibility, tunable degradation and elasticity. The addition of bioactive glasses to these elastomers can impart mechanical properties sufficient for hard tissue replacement. Hence, a composite with a biodegradable polymer matrix and a bioglass filler can offer a method of augmenting existing tissue. This article reviews the applications of such composites for skeletal augmentation.
Stem cells are well-known to have prominent roles in tissue engineering applications. Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) can differentiate into every cell type in the body while adult stem cells such as mesenchymal stem cells (MSCs) can be isolated from various sources. Nevertheless, an utmost limitation in harnessing stem cells for tissue engineering is the supply of cells. The advances in biomaterial technology allows the establishment of ex vivo expansion systems to overcome this bottleneck. The progress of various scaffold fabrication could direct stem cell fate decisions including cell proliferation and differentiation into specific lineages in vitro. Stem cell biology and biomaterial technology promote synergistic effect on stem cell-based regenerative therapies. Therefore, understanding the interaction of stem cell and biomaterials would allow the designation of new biomaterials for future clinical therapeutic applications for tissue regeneration. This review focuses mainly on the advances of natural and synthetic biomaterials in regulating stem cell fate decisions. We have also briefly discussed how biological and biophysical properties of biomaterials including wettability, chemical functionality, biodegradability and stiffness play their roles.
To avoid tissue rejection during organ transplantation, research has focused on the use of tissue engineering to regenerate required tissues or organs for patients. The biomedical applications of hyperbranched, multivalent, structurally uniform, biocompatible dendrimers in tissue engineering include the mimicking of natural extracellular matrices (ECMs) in the 3D microenvironment. Dendrimers are unimolecular architects that can incorporate a variety of biological and/or chemical substances in a 3D architecture to actively support the scaffold microenvironment during cell growth. Here, we review the use of dendritic delivery systems in tissue engineering. We discuss the available literature, highlighting the 3D architecture and preparation of these nanoscaffolds, and also review challenges to, and advances in, the use dendrimers in tissue engineering. Advances in the manufacturing of dendritic nanoparticles and scaffold architectures have resulted in the successful incorporation of dendritic scaffolds in tissue engineering.
Human amniotic epithelial cells (hAECs) are potentially one of the key players in tissue engineering due to their easy availability. The aim of the present study was to develop an optimal isolation and transportation technique, as well as to determine the immunophenotype and epithelial gene expression of hAECs. Amnion was mechanically peeled off from the chorion and digested with trypsin-ethylenediaminetetraacetic acid. The isolated hAECs were cultured in medium containing 10 ng/mL epidermal growth factor until P4. The epithelial gene expression, cell surface antigen and protein expression of hAECs were analyzed by quantitative polymerase chain reaction, flow cytometry and immunocytochemistry. hAECs were also cultured in adipogenic, osteogenic and neurogenic induction media. The best cell yield of hAECs was seen in the digestion of 15 pieces of amnion (2 × 2 cm) and isolated 30 min after digestion with trypsin. F12:Dulbecco's modified eagle medium was the best medium for short term storage at 4 °C. hAECs expressed CD9, CD44, CD73 and CD90, and negligibly expressed CD31, CD34, CD45 and CD117. After serial passage, CK3, CK19 and involucrin gene expressions were upregulated, while p63, CK1 and CK14 gene expressions were downregulated. Sustained gene expressions of integrin β1 and CK18 were observed. At initial culture, these cells might have stem-like properties. However, they differentiated after serial passage. Nonetheless, hAECs have epithelial stem cell characteristics and have the potential to differentiate into corneal epithelial cells. Further investigations are still needed to elucidate the mechanism of differentiation involved and to optimize the culture condition for long term in vitro culture.
Nanotechnology advancements have led to the development of its allied fields, such as nanoparticle synthesis and their applications in the field of biomedicine. Nanotechnology driven innovations have given a hope to the patients as well as physicians in solving the complex medical problems. Nanoparticles with a size ranging from 0.2 to 100 nm are associated with an increased surface to volume ratio. Moreover, the physico-chemical and biological properties of nanoparticles can be modified depending on the applications. Different nanoparticles have been documented with a wide range of applications in various fields of medicine and biology including cancer therapy, drug delivery, tissue engineering, regenerative medicine, biomolecules detection, and also as antimicrobial agents. However, the development of stable and effective nanoparticles requires a profound knowledge on both physico-chemical features of nanomaterials and their intended applications. Further, the health risks associated with the use of engineered nanoparticles needs a serious attention.
Chitosan, collagen, gelatin, polylactic acid and polyhydroxyalkanoates are notable examples of biopolymers, which are essentially bio-derived polymers produced by living cells. With the right techniques, these biological macromolecules can be exploited for nanotechnological advents, including for the fabrication of nanocarriers. In the world of nanotechnology, it is highly essential (and optimal) for nanocarriers to be biocompatible, biodegradable and non-toxic for safe in vivo applications, including for drug delivery, cancer immunotherapy, tissue engineering, gene delivery, photodynamic therapy and many more. The recent advancements in understanding nanotechnology and the physicochemical properties of biopolymers allows us to modify biological macromolecules and use them in a multitude of fields, most notably for clinical and therapeutic applications. By utilizing chitosan, collagen, gelatin, polylactic acid, polyhydroxyalkanoates and various other biopolymers as synthesis ingredients, the 'optimal' properties of a nanocarrier can easily be attained. With emphasis on the aforementioned biological macromolecules, this review presents the various biopolymers utilized for nanocarrier synthesis along with their specific synthetization methods. We further discussed on the characterization techniques and related applications for the synthesized nanocarriers.
Scaffolds supplemented with naturally derived materials seem to be a good choice in bone tissue engineering. This study aims to develop polyurethane (PU) nanofibers added with ylang ylang (YY) and zinc nitrate (ZnNO3) using the electrospinning method. Field emission scanning electron microscopy (FESEM) images showed that the diameter of the PU nanofibers (869 ± 122 nm) was reduced with the addition of YY and ZnNO3 (PU/YY-467 ± 132 nm and PU/YY/ZnNO3-290 ± 163 nm). Fourier transform infrared (FTIR), a thermal gravimetric analysis (TGA) and an X-ray diffraction (XRD) analysis confirmed the interactions between PU with YY and ZnNO3. In addition, a thermal gravimetric analysis (TGA) study revealed the improved thermal stability for PU/YY and a slight reduction in the thermal stability for PU/YY/ZnNO3. A tensile test indicated that the addition of YY and ZnNO3 (PU/YY-12.32 MPa and PU/YY/ZnNO3-14.90 MPa) improved the mechanical properties of the pristine PU (6.83 MPa). The electrospun PU/YY (524 nm) and PU/YY/ZnNO3 (284 nm) showed a reduced surface roughness when compared with the pristine PU (776 nm) as depicted in the atomic force microscopy (AFM) analysis. The addition of YY and ZnNO3 improved the anticoagulant and biocompatibility nature of the pristine PU. Furthermore, the bone mineralization study depicted the improved calcium deposition in the fabricated composites (PU/YY-7.919% and PU/YY/ZnNO3-10.150%) compared to the pristine PU (5.323%). Hence, the developed composites with desirable physico-chemical properties, biocompatibility and calcium deposition can serve as plausible candidates for bone tissue engineering.
The development of nano-sized scaffolds with antibacterial properties that mimic the architecture of tissue is one of the challenges in tissue engineering. In this study, polycaprolactone (PCL) and PCL/gelatine (Ge) (70:30) nanofibrous scaffolds were fabricated using a less toxic and common solvent, formic acid and an electrospinning technique. Nanofibrous scaffolds were coated with silver (Ag) in different concentrations of silver nitrate (AgNO3) aqueous solution (1.25, 2.5, 5, and 10 %) by using dipping method, drying and followed by ultraviolet (UV) photoreduction. The PCL/Ge (70:30) nanofibrous scaffold had an average fibre diameter of 155.60 ± 41.13 nm. Characterization showed that Ag was physically entrapped in both the PCL and PCL/Ge (70:30) nanofibrous scaffolds. Ag(+) ions release study was performed and showed much lesser release amount than the maximum toxic concentration of Ag(+) ions in human cells. Both scaffolds were non-toxic to cells and demonstrated antibacterial effects towards Gram-positive Bacillus cereus (B. cereus) and Gram-negative Escherichia coli (E. coli). The Ag/PCL/Ge (70:30) nanofibrous scaffold has potential for tissue engineering as it can protect wounds from bacterial infection and promote tissue regeneration.
Conductive materials are potential candidates for developing bone tissue engineering scaffolds as they are nontoxic and can enhance bone tissue regeneration. Their bioactivity can be enhanced by depositing biomineralization in simulated body fluid (SBF). In the current study, a composite electrospun membrane made up of poly(lactic) acid, poly(3-hydroxybutyrate-co-3-hydroxyvalerate), and hydroxyapatite was fabricated using an electrospinning method. The fabricated membranes were dip-coated with a conductive polymer solution, poly(3,4-ethylenedioxythiophene) poly(4-styrenesulfonate), to induce conductivity. Characterization of the membranes based on characteristics such as morphology, chemical bonding, and wettability was conducted using scanning electron microscopy, field emission scanning electron microscopy, energy-dispersive X-ray spectroscopy, attenuated total reflectance Fourier transform infrared spectroscopy, and contact angle measurement. From the results, biomineralization of both coated and noncoated composite membranes was observed on the surface of nanofibers after 21 days in SBF. The membranes provide a superhydrophilic surface as shown by the contact angle. In conclusion, this biomimetic electrospun composite membrane could be used to further support cell growth for bone tissue engineering application.
Tissue engineering seeks strategies to design polymeric scaffolds that allow high-cell-density cultures with signaling molecules and suitable vascular supply. One major obstacle in tissue engineering is the inability to create thick engineered-tissue constructs. A pre-vascularized tissue scaffold appears to be the most favorable approach to avoid nutrient and oxygen supply limitations as well as to allow waste removal, factors that are often hurdles in developing thick engineered tissues. Vascularization can be achieved using strategies in which cells are cultured in bioactive polymer scaffolds that can mimic extracellular matrix environments. This review addresses recent advances and future challenges in developing and using bioactive polymer scaffolds to promote tissue construct vascularization.
The guidance of endothelial cell organization into a capillary network has been a long-standing challenge in tissue engineering. Some research efforts have been made to develop methods to promote capillary networks inside engineered tissue constructs. Capillary and vascular networks that would mimic blood microvessel function can be used to subsequently facilitate oxygen and nutrient transfer as well as waste removal. Vascularization of engineering tissue construct is one of the most favorable strategies to overpass nutrient and oxygen supply limitation, which is often the major hurdle in developing thick and complex tissue and artificial organ. This paper addresses recent advances and future challenges in developing three-dimensional culture systems to promote tissue construct vascularization allowing mimicking blood microvessel development and function encountered in vivo. Bioreactors systems that have been used to create fully vascularized functional tissue constructs will also be outlined.
Polyhydroxyalkanoates (PHAs) are biodegradable polyesters produced by microorganisms, under unbalanced growth conditions, as a carbon storage compound. PHAs are composed of various monomers such as 3-hydroxybutyrate (3HB) and 3-hydroxyhexanoate (3HHx). Silk fibroin (SF) derived from Bombyx mori cocoons, is a widely studied protein polymer commonly used for biomaterial applications. In this study, non-woven electrospun films comprising a copolymer of 3HB and 3HHx [P(3HB-co-3HHx)], SF and their blends were prepared by electrospinning technique. The growth and osteogenic differentiation of human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) were studied using different types of fabricated electrospun films. The differentiation study revealed that electrospun P(3HB-co-3HHx)/SF film supports the differentiation of hUC-MSCs into the osteogenic lineage, confirmed by histological analysis using Alizarin Red staining, energy dispersive X-ray (EDX) and quantitative real-time PCR analysis (qPCR). Electrospun P(3HB-co-3HHx)/SF film up-regulated the expression of osteogenic marker genes, alkaline phosphatase (ALP) and osteocalcin (OCN), by 1.6-fold and 2.8-fold respectively, after 21 days of osteogenic induction. In conclusion, proliferation and osteogenic differentiation of hUC-MSCs were enhanced through the blending of P(3HB-co-3HHx) and SF. The results from this study suggest that electrospun P(3HB-co-3HHx)/SF film is a promising biomaterial for bone tissue engineering.
Aloe vera (AV) and tetracycline hydrochloride (TCH) exhibit significant properties such as anti-inflammatory, antioxidant and anti-bacterial activities to facilitate skin tissue engineering. The present study aims to develop poly-ε-caprolactone (PCL)/ AV containing curcumin (CUR), and TCH loaded hybrid nanofibrous scaffolds to validate the synergistic effect on the fibroblast proliferation and antimicrobial activity against Gram-positive and Gram-negative bacteria for wound healing. PCL/AV, PCL/CUR, PCL/AV/CUR and PCL/AV/TCH hybrid nanofibrous mats were fabricated using an electrospinning technique and were characterized for surface morphology, the successful incorporation of active compounds, hydrophilicity and the mechanical property of nanofibers. SEM revealed that there was a decrease in the fiber diameter (ranging from 360 to 770 nm) upon the addition of AV, CUR and TCH in PCL nanofibers, which were randomly oriented with bead free morphology. FTIR spectra of various electrospun samples confirmed the successful incorporation of AV, CUR and TCH into the PCL nanofibers. The fabricated nanofibrous scaffolds possessed mechanical properties within the range of human skin. The biocompatibility of electrospun nanofibrous scaffolds were evaluated on primary human dermal fibroblasts (hDF) by MTS assay, CMFDA, Sirius red and F-actin stainings. The results showed that the fabricated PCL/AV/CUR and PCL/AV/TCH nanofibrous scaffolds were non-toxic and had the potential for wound healing applications. The disc diffusion assay confirmed that the electrospun nanofibrous scaffolds possessed antibacterial activity and provided an effective wound dressing for skin tissue engineering.
Selections of collagen available commercially were tested for their biocompatibility as scaffold to promote cell growth in vitro via simple collagen fast test and cultivation of mammalian cells on the selected type of collagen. It was found that collagen type C9791 promotes the highest degree of aggregation as well as cells growth. This preliminary study also indicated potential use of collagen as scaffold in engineered tissue.
Porous calcium phosphate ceramics have found enormous use in biomedical applications including bone tissue regeneration, cell proliferation, and drug delivery. In bone tissue engineering it has been applied as filling material for bone defects and augmentation, artificial bone graft material, and prosthesis revision surgery. Their high surface area leads to excellent osteoconductivity and resorbability providing fast bone ingrowths. Porous calcium phosphate can be produced by a variety of methods. This paper discusses briefly fundamental aspects of porous calcium phosphate for biomedical applications as well as various techniques used to prepare porous calcium phosphate.