Fabrication of porous polycaprolactone/hydroxyapatite (PCL/HA) blend scaffolds using a 3D plotting system for bone tissue engineering

For tissue engineering and regeneration, a porous scaffold with interconnected networks is needed to guide cell attachment and growth/ingrowth in three-dimensional (3D) structure. Using a rapid prototyping (RP) technique, we designed and fabricated 3D plotting system and three types of scaffolds: those from polycaprolactone (PCL), those from PCL and hydroxyapatite (HA), and those from PCL/HA and with a shifted pattern structure (PCL/HA/SP scaffold). Shifted pattern structure was fabricated to increase the cell attachment/adhesion. The PCL/HA/SP scaffold had a lower compressive modulus than PCL and PCL/HA scaffold. However, it has a better cell attachment than the scaffolds without a shifted pattern. MTT assay and alkaline phosphatase activity results for the PCL/HA/SP scaffolds were significantly enhanced compared to the results for the PCL and PCL/HA scaffolds. According to their degree of cell proliferation/differentiation, the scaffolds were in the following order: PCL/HA/SP > PCL/HA > PCL. These 3D scaffolds will be applicable for tissue engineering based on unique plotting system.     

Immobilization of thrombocytes on PCL nanofibres enhances chondrocyte proliferation in vitro

Objectives: The aim of this study was to develop functionalized nanofibres as a simple delivery system for growth factors (GFs) and make nanofibre cell‐seeded scaffold implants a one‐step intervention. Materials and methods: We have functionalized polycaprolactone (PCL) nanofibres with thrombocytes adherent on them. Immobilized, these thrombocytes attached to nanofibre scaffolds were used as a nanoscale delivery system for native (autologous) proliferation and differentiation factors, in vitro. Pig chondrocytes were seeded on the thrombocyte‐coated scaffolds and levels of proliferation and differentiation of these cells were compared with those seeded on non‐coated scaffolds. Results: Immobilized thrombocytes on PCL nanofibres effectively enhanced chondrocyte proliferation due to time‐dependent degradation of thrombocytes and release of their GFs. Conclusions: These simply functionalized scaffolds present new possibilities for nanofibre applications, as smart cell scaffolds equipped with a GF delivery tool.     

Mesenchymal stem cell responses to bone-mimetic electrospun matrices composed of polycaprolactone, collagen I and nanoparticulate hydroxyapatite

The performance of biomaterials designed for bone repair depends, in part, on the ability of the material to support the adhesion and survival of mesenchymal stem cells (MSCs). In this study, a nanofibrous bone-mimicking scaffold was electrospun from a mixture of polycaprolactone (PCL), collagen I, and hydroxyapatite (HA) nanoparticles with a dry weight ratio of 50/30/20 respectively (PCL/col/HA). The cytocompatibility of this tri-component scaffold was compared with three other scaffold formulations: 100% PCL (PCL), 100% collagen I (col), and a bi-component scaffold containing 80% PCL/20% HA (PCL/HA). Scanning electron microscopy, fluorescent live cell imaging, and MTS assays showed that MSCs adhered to the PCL, PCL/HA and PCL/col/HA scaffolds, however more rapid cell spreading and significantly greater cell proliferation was observed for MSCs on the tri-component bone-mimetic scaffolds. In contrast, the col scaffolds did not support cell spreading or survival, possibly due to the low tensile modulus of this material. PCL/col/HA scaffolds adsorbed a substantially greater quantity of the adhesive proteins, fibronectin and vitronectin, than PCL or PCL/HA following in vitro exposure to serum, or placement into rat tibiae, which may have contributed to the favorable cell responses to the tri-component substrates. In addition, cells seeded onto PCL/col/HA scaffolds showed markedly increased levels of phosphorylated FAK, a marker of integrin activation and a signaling molecule known to be important for directing cell survival and osteoblastic differentiation. Collectively these results suggest that electrospun bone-mimetic matrices serve as promising degradable substrates for bone regenerative applications.     

Inhibitor of PI3K/Akt Signaling Pathway Small Molecule Promotes Motor Neuron Differentiation of Human Endometrial Stem Cells Cultured on Electrospun Biocomposite Polycaprolactone/Collagen Scaffolds

Small molecules as useful chemical tools can affect cell differentiation and even change cell fate. It is demonstrated that LY294002, a small molecule inhibitor of phosphatidylinositol 3-kinase (PI3K)/Akt signal pathway, can inhibit proliferation and promote neuronal differentiation of mesenchymal stem cells (MSCs). The purpose of this study was to investigate the differentiation effect of Ly294002 small molecule on the human endometrial stem cells (hEnSCs) into motor neuron-like cells on polycaprolactone (PCL)/collagen scaffolds. hEnSCs were cultured in a neurogenic inductive medium containing 1 μM LY294002 on the surface of PCL/collagen electrospun fibrous scaffolds. Cell attachment and viability of cells on scaffolds were characterized by scanning electron microscope (SEM) and 3-(4,5-dimethylthiazoyl-2-yl)2,5-diphenyltetrazolium bromide (MTT) assay. The expression of neuron-specific markers was assayed by real-time PCR and immunocytochemistry analysis after 15 days post induction. Results showed that attachment and differentiation of hEnSCs into motor neuron-like cells on the scaffolds with Ly294002 small molecule were higher than that of the cells on tissue culture plates as control group. In conclusion, PCL/collagen electrospun scaffolds with Ly294002 have potential for being used in neural tissue engineering because of its bioactive and three-dimensional structure which enhances viability and differentiation of hEnSCs into neurons through inhibition of the PI3K/Akt pathway. Thus, manipulation of this pathway by small molecules can enhance neural differentiation.     

Supercritical fluid-assisted controllable fabrication of open and highly interconnected porous scaffolds for bone tissue engineering

Recently tremendous progress has been evidenced by the advancements in developing innovative three-dimensional(3 D)scaffolds using various techniques for addressing the autogenous grafting of bone. In this work, we demonstrated the fabrication of porous polycaprolactone(PCL) scaffolds for osteogenic differentiation based on supercritical fluid-assisted hybrid processes of phase inversion and foaming. This eco-friendly process resulted in the highly porous biomimetic scaffolds with open and interconnected architectures. Initially, a 2~3 factorial experiment was designed for investigating the relative significance of various processing parameters and achieving better control over the porosity as well as the compressive mechanical properties of the scaffold. Then, single factor experiment was carried out to understand the effects of various processing parameters on the morphology of scaffolds. On the other hand, we encapsulated a growth factor, i.e., bone morphogenic protein-2(BMP-2), as a model protein in these porous scaffolds for evaluating their osteogenic differentiation. In vitro investigations of growth factor loaded PCL scaffolds using bone marrow stromal cells(BMSCs) have shown that these growth factor-encumbered scaffolds were capable of differentiating the cells over the control experiments. Furthermore, the osteogenic differentiation was confirmed by measuring the cell proliferation, and alkaline phosphatase(ALP) activity, which were significantly higher demonstrating the active bone growth. Together, these results have suggested that the fabrication of growth factor-loaded porous scaffolds prepared by the eco-friendly hybrid processing efficiently promoted the osteogenic differentiation and may have a significant potential in bone tissue engineering.     

In vitro study of nano-HA/PLLA composite scaffold for rabbit BMSC differentiation under TGF-β1 induction

The aim of this study is to investigate the effects of differentiation of rabbit bone marrow mesenchymal stem cells (rBMSCs) into chondrocytes induced by transforming growth factor-beta1 (TCP-β1) composite poly-1actide-co-glycolic acid/nano-hydroxyapatite (PLLA/nano-HA) to the construction of biomimetic artificial cartilage in vitro. In the low-temperature extrusion preparation of PLLA/nano-HA composite porous scaffolds, rBMSCs were isolated and cultured to third generation in vitro, induced by TGF-β1-contained special inducing system into chondrocytes, 14 d later, identified by toluidine blue and type II collagen immunohistochemistry staining, and then the differential chondrocytes composite into the PLLA/nano-HA composite porous scaffolds, using scanning electron microscopy (SEM) to observe the growth conditions and cell attachment on the composite in the 7th,14th, and 21st day and to gather cells on composite in the 7th, 14th, and 21st day of cell. RT-PCR is used to detect the expression of aggrecan (Col2A1 in mRNA) and Western blot for detection of the expression of type II collagen of the attached cells. rBMSCs can differentiate into chondrocytes when induced, and the differentiation of chondrocytes secreting GAG by toluidine blue staining and type II collagen immunohistochemistry staining was positive; SEM confirm the cells distribution evenly, stretching well in composite. RT-PCR of aggrecan, Col2A1 in mRNA, and Western-blot of type II collagen expression in the differentiation of chondrocytes have different levels. Using TGF-β1 containing special inducing system induced rBMSCs into chondrocytes, then into compounds of PLLA/nano-HA composite porous scaffolds, and cell carrier complex proliferated well and secreted the chondrocyte-specific extracellular matrix stably, successfully constructing artificial bionic in vitro.     

Evaluation of biological protein-based collagen scaffolds in cartilage and musculoskeletal tissue engineering--a systematic review of the literature

The term tissue engineering is the technology that combines cells, engineering and biological/synthetic material in order to repair, replace or regenerate biological tissues such as bone, muscle, tendons and cartilage. The major human applications of tissue engineering are: skin, bone, cartilage, corneas, blood vessels, left mainstem bronchus and urinary structures. In this systematic review several criteria were identified as the most desirable characteristics of an ideal scaffold. These state that an ideal scaffolds needs to be biodegradable, possess mechanical strength, be highly porous, biocompatible, non-cytotoxic, non antigentic, stuitable for cell attachment, proliferation and differentiation, flexible and elastic, three dimensional, osteoconductive and support the transport of nutrients and metabolic waste. Subsequently, studies reporting on the various advantages and disadvantages of using collagen based scaffolds in musculoskeletal and cartilage tissue engineering were identified. The purpose of this review is to 1) provide a list of ideal characteristics of a scaffold as identified in the literature 2) identify different types of biological protein-based collagen scaffolds used in musculoskeletal and cartilage tissue engineering 3) assess how many of the criteria each scaffold type meets 4) weigh different scaffolds against each other according to their relative properties and shortcomings. The rationale behind this approach is that the ideal scaffold material has not yet been identified. Hence, this review will define how many of the identified ideal characteristics are fulfilled by natural collagen-based scaffolds and address the shortcomings of its use as found in the literature.     

Three-dimensional hierarchical composite scaffolds consisting of polycaprolactone, β-tricalcium phosphate, and collagen nanofibers: fabrication, physical properties, and in vitro cell activity for bone tissue regeneration

β-Tricalcium phosphate (β-TCP) and collagen have been widely used to regenerate various hard tissues, but although Bioceramics and collagen have various biological advantages with respect to cellular activity, their usage has been limited due to β-TCP's inherent brittleness and low mechanical properties, along with the low shape-ability of the three-dimensional collagen. To overcome these material deficiencies, we fabricated a new hierarchical scaffold that consisted of a melt-plotted polycaprolactone (PCL)/β-TCP composite and embedded collagen nanofibers. The fabrication process was combined with general melt-plotting methods and electrospinning. To evaluate the capability of this hierarchical scaffold to act as a biomaterial for bone tissue regeneration, physical and biological assessments were performed. Scanning electron microscope (SEM) micrographs of the fabricated scaffolds indicated that the β-TCP particles were uniformly embedded in PCL struts and that electrospun collagen nanofibers (diameter = 160 nm) were well layered between the composite struts. By accommodating the β-TCP and collagen nanofibers, the hierarchical composite scaffolds showed dramatic water-absorption ability (100% increase), increased hydrophilic properties (20%), and good mechanical properties similar to PCL/β-TCP composite. MTT assay and SEM images of cell-seeded scaffolds showed that the initial attachment of osteoblast-like cells (MG63) in the hierarchical scaffold was 2.2 times higher than that on the PCL/β-TCP composite scaffold. Additionally, the proliferation rate of the cells was about two times higher than that of the composite scaffold after 7 days of cell culture. Based on these results, we conclude that the collagen nanofibers and β-TCP particles in the scaffold provide good synergistic effects for cell activity.     

Repair and regeneration of osteochondral defects in the articular joints

People suffering from pain due to osteoarthritic or rheumatoidal changes in the joints are still waiting for a better treatment. Although some studies have achieved success in repairing small cartilage defects, there is no widely accepted method for complete repair of osteochondral defects. Also joint replacements have not yet succeeded in replacing of natural cartilage without complications. Therefore, there is room for a new medical approach, which outperforms currently used methods. The aim of this study is to show potential of using a tissue engineering approach for regeneration of osteochondral defects. The critical review of currently used methods for treatment of osteochondral defects is also provided. In this study, two kinds of hybrid scaffolds developed in Hutmacher's group have been analysed. The first biphasic scaffold consists of fibrin and PCL. The fibrin serves as a cartilage phase while the porous PCL scaffold acts as the subchondral phase. The second system comprises of PCL and PCL-TCP. The scaffolds were fabricated via fused deposition modeling which is a rapid prototyping system. Bone marrow-derived mesenchymal cells were isolated from New Zealand White rabbits, cultured in vitro and seeded into the scaffolds. Bone regenerations of the subchondral phases were quantified via micro CT analysis and the results demonstrated the potential of the porous PCL and PCL-TCP scaffolds in promoting bone healing. Fibrin was found to be lacking in this aspect as it degrades rapidly. On the other hand, the porous PCL scaffold degrades slowly hence it provides an effective mechanical support. This study shows that in the field of cartilage repair or replacement, tissue engineering may have big impact in the future. In vivo bone and cartilage engineering via combining a novel composite, biphasic scaffold technology with a MSC has been shown a high potential in the knee defect regeneration in the animal models. However, the clinical application of tissue engineering requires the future research work due to several problems, such as scaffold design, cellular delivery and implantation strategies.     

Ovine bone marrow mesenchymal stem cells: isolation and characterization of the cells and their osteogenic differentiation potential on embroidered and surface-modified polycaprolactone-co-lactide scaffolds

The current study was undertaken with the goal being isolation, cultivation, and characterization of ovine mesenchymal stem cells (oMSC). Furthermore, the objective was to determine whether biological active polycaprolactone-co-lactide (trade name PCL) scaffolds support the growth and differentiation of oMSC in vitro. The oMSC were isolated from the iliac crest of six merino sheep. Three factors were used to demonstrate the MSC properties of the isolated cells in detail. (1) Their ability to proliferate in culture with a spindle-shaped morphology, (2) presence of specific surface marker proteins, and (3) their capacity to differentiate into the three classical mesenchymal pathways, osteoblastic, adipogenic, and chondrogenic lineages. Furthermore, embroidered PCL scaffolds were coated with collagen I (coll I) and chondroitin sulfate (CS). The porous structure of the scaffolds and the coating with coll I/CS allowed the oMSC to adhere, proliferate, and to migrate into the scaffolds. The coll I/CS coating on the PCL scaffolds induced osteogenic differentiation of hMSC, without differentiation supplements, indicating that the scaffold also has an osteoinductive character. In conclusion, the isolated cells from the ovine bone marrow have similar morphologic, immunophenotypic, and functional characteristics as their human counterparts. These cells were also found to differentiate into multiple mesenchymal cell types. This study demonstrates that embroidered PCL scaffolds can act as a temporary matrix for cell migration, proliferation, and differentiation of oMSC. The data presented will provide a reliable model system to assess the translation of MSC-based therapy into a variety of valuable ovine experimental models under autologous settings.     

Electrospun Poly(L-lactide)/Poly(ε-caprolactone) Blend Nanofibrous Scaffold: Characterization and Biocompatibility with Human Adipose-Derived Stem Cells

The essence of tissue engineering is the fabrication of autologous cells or induced stem cells in naturally derived or synthetic scaffolds to form specific tissues. Polymer is thought as an appealing source of cell-seeded scaffold owing to the diversity of its physicochemical property and can be electrospun into nano-size to mimic natural structure. Poly (L-lactic acid) (PLLA) and poly (ε-caprolactone) (PCL) are both excellent aliphatic polyester with almost “opposite” characteristics. The controlling combination of PLLA and PCL provides varying properties and makes diverse applications. Compared with the copolymers of the same components, PLLA/PCL blend demonstrates its potential in regenerative medicine as a simple, efficient and scalable alternative. In this study, we electrospun PLLA/PCL blends of different weight ratios into nanofibrous scaffolds (NFS) and their properties were detected including morphology, porosity, degradation, ATR-FTIR analysis, stress-stain assay, and inflammatory reaction. To explore the biocompatibility of the NFS we synthesized, human adipose-derived stem cells (hASCs) were used to evaluate proliferation, attachment, viability and multi-lineage differentiation. In conclusion, the electrospun PLLA/PCL blend nanofibrous scaffold with the indicated weight ratios all supported hASCs well. However, the NFS of 1/1 weight ratio showed better properties and cellular responses in all assessments, implying it a biocompatible scaffold for tissue engineering.     

牙源性干细胞复合微渠多孔羟基磷灰石支架成骨性能研究*

目的: 探讨牙源性干细胞复合微渠多孔羟基磷灰石支架(grooved porous hydroxyapatite scaffolds, HAG支架)的成骨性能,为骨缺损修复治疗提供新手段。方法: 从健康成人第三磨牙中提取牙周膜干细胞(periodontal ligament stem cells, PDLSCs)及牙髓干细胞(dental pulp stem cells, DPSCs)分别接种于HAG支架上,进行多向分化鉴定及碱性磷酸酶(alkaline phosphatase,ALP)活性测定;并通过CCK-8检测细胞增殖能力;逆转录聚合酶链反应(qRT-PCR)检测骨形态发生蛋白2(bone morphogenetic protein 2, BMP-2)、骨钙素(osteocalcin, OCN)和骨桥蛋白(osteopontin, OPN)等成骨相关基因的表达。体内研究中将搭载PDLSCs和DPSCs的HAG支架移植到裸鼠的背部皮下,8周后取材,组织切片后采用苏木精-伊红(HE)染色观察新骨形成,提取组织蛋白采用Western blot检测ALP、OCN等成骨相关蛋白的表达。结果: 体外研究中DPSCs复合HAG支架组的细胞增殖能力、ALP活性,以及成骨相关基因ALP、BMP2、OCN等的表达均高于PDLSCs复合HAG支架组。体内研究中HE染色显示,PDLSCs复合HAG支架组及DPSCs复合HAG支架组均较空白HAG支架组有更多细胞生长区、纤维细胞增生及骨基质形成,且DPSCs复合HAG支架组的骨基质面积更大,成纤维细胞数量更多;PDLSCs复合HAG支架组及DPSCs复合HAG支架组成骨相关蛋白的表达量均高于空白HAG组,且DPSCs复合HAG支架组中ALP蛋白表达量显著高于PDLSCs复合HAG支架组。结论: PDLSCs、DPSCs复合HAG支架在体内外均表现出良好的成骨性能,其中DPSCs复合HAG支架的成骨性能更为优异。     

Combinatorial polymer electrospun matrices promote physiologically-relevant cardiomyogenic stem cell differentiation

Myocardial infarction results in extensive cardiomyocyte death which can lead to fatal arrhythmias or congestive heart failure. Delivery of stem cells to repopulate damaged cardiac tissue may be an attractive and innovative solution for repairing the damaged heart. Instructive polymer scaffolds with a wide range of properties have been used extensively to direct the differentiation of stem cells. In this study, we have optimized the chemical and mechanical properties of an electrospun polymer mesh for directed differentiation of embryonic stem cells (ESCs) towards a cardiomyogenic lineage. A combinatorial polymer library was prepared by copolymerizing three distinct subunits at varying molar ratios to tune the physicochemical properties of the resulting polymer: hydrophilic polyethylene glycol (PEG), hydrophobic poly(ε-caprolactone) (PCL), and negatively-charged, carboxylated PCL (CPCL). Murine ESCs were cultured on electrospun polymeric scaffolds and their differentiation to cardiomyocytes was assessed through measurements of viability, intracellular reactive oxygen species (ROS), α-myosin heavy chain expression (α-MHC), and intracellular Ca(2+) signaling dynamics. Interestingly, ESCs on the most compliant substrate, 4%PEG-86%PCL-10%CPCL, exhibited the highest α-MHC expression as well as the most mature Ca(2+) signaling dynamics. To investigate the role of scaffold modulus in ESC differentiation, the scaffold fiber density was reduced by altering the electrospinning parameters. The reduced modulus was found to enhance α-MHC gene expression, and promote maturation of myocyte Ca(2+) handling. These data indicate that ESC-derived cardiomyocyte differentiation and maturation can be promoted by tuning the mechanical and chemical properties of polymer scaffold via copolymerization and electrospinning techniques.     

Delivery of platelet-derived growth factor as a chemotactic factor for mesenchymal stem cells by bone-mimetic electrospun scaffolds

The recruitment of mesenchymal stem cells (MSCs) is a vital step in the bone healing process, and hence the functionalization of osteogenic biomaterials with chemotactic factors constitutes an important effort in the tissue engineering field. Previously we determined that bone-mimetic electrospun scaffolds composed of polycaprolactone, collagen I and nanohydroxyapatite (PCL/col/HA) supported greater MSC adhesion, proliferation and activation of integrin-related signaling cascades than scaffolds composed of PCL or collagen I alone. In the current study we investigated the capacity of bone-mimetic scaffolds to serve as carriers for delivery of an MSC chemotactic factor. In initial studies, we compared MSC chemotaxis toward a variety of molecules including PDGF-AB, PDGF-BB, BMP2, and a mixture of the chemokines SDF-1α, CXCL16, MIP-1α, MIP-1β, and RANTES. Transwell migration assays indicated that, of these factors, PDGF-BB was the most effective in stimulating MSC migration. We next evaluated the capacity of PCL/col/HA scaffolds, compared with PCL scaffolds, to adsorb and release PDGF-BB. We found that significantly more PDGF- BB was adsorbed to, and subsequently released from, PCL/col/HA scaffolds, with sustained release extending over an 8-week interval. The PDGF-BB released was chemotactically active in transwell migration assays, indicating that bioactivity was not diminished by adsorption to the biomaterial. Complementing these studies, we developed a new type of migration assay in which the PDGF-BB-coated bone-mimetic substrates were placed 1.5 cm away from the cell migration front. These experiments confirmed the ability of PDGF-BB-coated PCL/col/HA scaffolds to induce significant MSC chemotaxis under more stringent conditions than standard types of migration assays. Our collective results substantiate the efficacy of PDGF-BB in stimulating MSC recruitment, and further show that the incorporation of native bone molecules, collagen I and nanoHA, into electrospun scaffolds not only enhances MSC adhesion and proliferation, but also increases the amount of PDGF-BB that can be delivered from scaffolds.     

Promotion of human mesenchymal stem cell differentiation on bioresorbable polycaprolactone/biphasic calcium phosphate composite scaffolds for bone tissue engineering

An artificial construct mimicking the intrinsic properties of the natural extracellular matrix in bones has been considered an ideal platform for bone tissue engineering, as it can present an appropriate microenvironment and regulate cell behaviours. In this report, we introduce biodegradable composite scaffolds consisting of polycaprolactone (PCL) and biphasic calcium phosphate (BCP). The scaffolds were fabricated by a salt-leaching process, and the ability of the scaffolds to facilitate osteogenic differentiation was investigated using human mesenchymal stem cells (hMSCs). The scaffolds had an inter-connected porous structure with quadrilateral pores of approximately 200 ～ 500 μm in width. The mechanical properties of the scaffolds changed as the BCP content was increased in the starting mixture. In the hMSC experiment, although we found that hMSCs adhered to the surface, as well as the inside, of the scaffolds, the incorporated BCP did not increase the proliferation of the hMSCs over 7 days in culture. Interestingly, the alkaline phosphatase (ALP) activity was 4 times higher on the PCL/BCP composite scaffold (0.12 ± 0.03 nmol/min/μg protein) thanon the PCL scaffold (0.03 ± 0.01 nmol/min/μg protein), suggesting that BCP can aid in generating a local environment that promotes bone regeneration. Therefore, a strategy combining polymers and ceramics can be considered a useful platform for bone tissue engineering.     

Osteogenic differentiation of human placenta-derived mesenchymal stem cells (PMSCs) on electrospun nanofiber meshes

Numerous challenges remain in the successful clinical translation of cell-based therapeutic studies for skeletal tissue repair, including appropriate cell sources and viable cell delivery systems. Poly(ethylene glycol)-poly(ε-caprolactone) (PEG-PCL) amphiphilic block copolymers have been extensively explored in microspheres preparation. Due to the introduction of hydrophilic PEG segments into PCL backbones, these copolymers have shown much more potentials in carrying protein, lipophilic drugs or genes than commonly used poly (ε-caprolactone) (PCL) and poly (lactic acid). The aim of this study is to investigate the attachment and osteogenic differentiation of human placenta derived mesenchymal stem cells (PMSCs) on PEG-PCL triblock copolymers nanofiber scaffolds. Here we demonstrated that PMSCs proliferate robustly and can be effectively differentiated into osteogenic-like cells on nanofiber scaffolds. This study provides evidence for the use of nanofiber scaffolds as an ideal supporting material for in vitro PMSCs culture and an in vivo cell delivery vehicle for bone repair.     

Different Porosities of Chitosan Can Influence the Osteogenic Differentiation Potential of Stem Cells

Scaffolds porosity has an important role in in vitro and in vivo differentiation process of stem cells with given the amount of space available to the cells to proliferate and differentiate. In the present study, chitosan with three porosities including 10%, 15%, and 20% that created by gelatin were used for investigation of the proliferation and osteogenic differentiation potential of adipose‐derived stem cells (ADSCs). In order to be more like the scaffold to natural bone tissue, freeze‐drying method was used in the scaffold preparation. Scaffold morphology, cell attachment, and toxicity were evaluated using scanning electron microscopy and MTT assay. Then, osteogenic differentiation potential of ADSCs cultured on chitosan with different porosities was evaluated by common osteogenic markers such as Alizarin red staining, ALP activity, calcium content, and osteogenic‐related genes expression via real‐time RT‐PCR. Although all scaffolds supported the proliferation and differentiation of ADSCs, but 10% scaffold demonstrated higher amount of osteogenic markers in comparison with the other porosities and control groups. Taking together, it can be concluded that osteogenic differentiation well done in the scaffolds with lower porosity because density of the cells will increase by forcing resulted from the scaffold, so osteogenic differentiation of the stem cells have an inverse association with scaffold porosity. J. Cell. Biochem. 119: 625–633, 2018. © 2017 Wiley Periodicals, Inc.     

The effects of chemokine, adhesion and extracellular matrix molecules on binding of mesenchymal stromal cells to poly(l-lactic acid)

Abstract Background aims. Mesenchymal stromal cells (MSC) are pluripotent adult stem cells capable of osteogenesis and chondrogenesis to form bone and cartilage. This characteristic gives them the potential for bone and cartilage regeneration. Synthetic polymers have been studied to examine whether they could be used as a scaffold for tissue engineering. In the current study a two-dimensional (2-D) poly(l-lactic acid) (PLLA) scaffold was treated with chemokine, adhesion and extracellular matrix molecules with the aim of using biologic molecules to improve the attachment of human MSC. Methods. MSC were isolated from human bone marrow and applied to a 2-D PLLA scaffold. Chemokines ligand (CXCL12 and CXCL13), adhesion molecules [P-selectin, vascular cell adhesion molecule (VCAM)-1 and heparin] and extracellular matrix molecules (fibronectin and type IV collagen) were coated on the scaffold and their effects on the number of MSC that adhered were recorded. Results. When used alone CXCL12 and CXCL13 enhanced MSC adhesion, as did VCAM-1, P-selectin, fibronectin and collagen, but not heparin. The effects of VCAM-1, P-selectin and heparin were enhanced by the addition of CXCL12. Incubation of MSC with antibodies to integrins α4 and α5β1 inhibited their adhesion to VCAM-1 and fibronectin-treated PLLA respectively, suggesting that these integrins were involved in the MSC interactions. Conclusions. The use of certain chemokines and adhesion and extracellular matrix molecules, alone or in combination, is beneficial for the attachment of MSC to PLLA, and may be helpful as natural molecules in scaffolds for regenerative medicine.