Nanofiber technology in the ex vivo expansion of cord blood-derived hematopoietic stem cells

Umbilical cord blood (CB) can be used as an alternative source of hematopoietic stem cells (HSCs) for transplantation in hematological and non-hematological disorders. Despite several recognized advantages the limited cell number in CB one unit still restricts its clinical use. The success of transplantation greatly depends on the levels of total nucleated cell and CD34+ cell counts. Thus, many ex vivo strategies have been developed within the last decade in order to solve this obstacle, with more or less success, mainly determined by the degree of difficulty related with maintaining HSCs self-renewal and stemness properties after long-term expansion. Different research groups have developed very promising and diverse CB-derived HSC expansion strategies using nanofiber scaffolds. Here we review the state-of-the-art of nanofiber technology-based CB-derived HSC expansion.     

静电纺丝法制备丹参素-聚己内酯纳米纤维支架

目的 利用静电纺丝技术制备丹参素-聚己内酯（DSS-PCL）纳米纤维支架。方法 以二氯甲烷-N,N-二甲基甲酰胺（CH₂Cl₂-DMF）8：2为溶剂,利用静电纺丝仪制备载丹参素的纳米纤维,通过扫描电镜（SEM）观察纤维形貌,通过体外释放实验考察载药纳米纤维支架中丹参素的释放规律,通过MTT法和溶血率对载药纳米纤维支架的安全性进行初步评价。结果 CH₂Cl₂-DMF混合溶剂体系下药物和材料都能够很好地溶解并顺利电纺得到纳米纤维,平均纤维直径分别为210 nm和190 nm,体外释放实验表明药物的释放经过一个快速释放后进入缓慢释放。MTT法测得PCL及载有丹参素的纳米纤维对L929细胞存活率并无明显影响,溶血率实验测得各组纳米纤维的溶血率低于5%。结论 静电纺丝法制备DSS-PCL纳米纤维的工艺简单易行,易于推广。     

Substituent-Controlled Energetics and Barriers of Mechanochromic Spiropyran-Functionalized Poly(ɛ-caprolactone)

In a joint theoretical and experimental study, it is shown that the onset of the mechanically-induced spiropyran (SP) to merocyanine (MC) isomerization can be controlled by both the regiochemistry and the substitution pattern of SP. Four SP-based bifunctional initiators with consistently varied polymer chain anchor point and substituent are used to synthesize poly-ε-caprolactone (PCL). Theoretical calculations (1S and 3S COGEF methods) and in-situ visible light absorption measurements of films during uniaxial stress–strain experiments consistently show varying activation barriers of the force-induced ring-opening reaction of SP to give MC. SPs with PCL chains attached in ortho-position to the pyran oxygen isomerize at lower stress than their para-analogs. NO₂-substituted SP mechanophores exhibit a lower activation barrier compared with H-substituted ones, but only if the  NO₂ substituent is located in para-position relative to the O at the pyran half. These results are consistent with theoretical loading rate-dependent rupture forces required to break the C O bond of SP and may guide mechanophore design.     

Engineering calcium deposits on polycaprolactone scaffolds for intravascular applications using primary human osteoblasts

Total atherosclerotic occlusions often include significant calcium deposits. Current animal models do not mimic the pathology of gradual occlusion of arteries and lack cell‐mediated calcium. The primary goal of this project was to establish an animal model incorporating these features into chronic total occlusions, using biodegradable scaffolds. As the first step, this study sought to determine the optimal dosage of TGF‐β1 on polycaprolactone (PCL) scaffolds cultured with primary human osteoblasts (HOBs) to effectively induce in vitro calcification. HOBs were cultured in TGF‐β1 and dexamethsaone (Dex)‐supplemented medium in well plates. Calcium in the cultures was visualized using alizarin red. The highest calcification was observed in groups with both TGF‐β1 (0.02 ng/ml) and Dex (10^?10 M ) in the medium. Next, HOBs were cultured on PCL scaffolds with different loadings of TGF‐β1: 0 (control), 5, 10, 50 and 100 ng. These cultures were performed with or without Dex (10^?10 M ) in the medium. DNA content, ALP activity and the amount and distribution of calcium were examined at 7, 14, 21 and 28 days. TGF‐β1 appeared to have an inhibitory effect on scaffold calcification when grown in Dex‐supplemented medium. When cultured without Dex, the lower amount of TGF‐β1 loading (5 ng) showed the most calcification, high DNA synthesis and high ALP activity on scaffolds. This study demonstrates the potential of implanting a PCL–HOB construct in an animal artery to establish a model of atherosclerotic occlusion with calcification. Copyright © 2010 John Wiley & Sons, Ltd.     

电纺聚己内酯/Ⅰ型胶原蛋白/纳米锆酸钙复合支架的制备及其生物相容性的研究

目的:制备聚己内酯(PCL)/Ⅰ型胶原(COLI)/纳米锆酸钙(nCZ)复合支架用于骨组织再生,评价其性能及对人牙周膜细胞(PDLCs)生物相容性及成骨分化的影响.方法:用静电纺丝法制备PCL/COLI、PCL/COLI/纳米羟基磷灰石(nHA)和PCL/COLI/nCZ复合支架,通过扫描电子显微镜表征支架形貌,能量色...     

Degradation and Characterisation of Electrospun Polycaprolactone (PCL) and Poly(lactic-co-glycolic acid) (PLGA) Scaffolds for Vascular Tissue Engineering

The current study aimed to evaluate the characteristics and the effects of degradation on the structural properties of Poly(lactic-co-glycolic acid) (PLGA)- and polycaprolactone (PCL)-based nanofibrous scaffolds. Six scaffolds were prepared by electrospinning, three with PCL 15% (w/v) and three with PLGA 10% (w/v), with electrospinning processing times of 30, 60 and 90 min. Both types of scaffolds displayed more robust mechanical properties with increased spinning times. The tensile strength of both scaffolds with 90-min electrospun membranes did not show a significant difference in their strengths, as the PCL and PLGA scaffolds measured at 1.492 MPa ± 0.378 SD and 1.764 MPa ± 0.7982 SD, respectively. All membranes were shown to be hydrophobic under a wettability test. A degradation behaviour study was performed by immersing all scaffolds in phosphate-buffered saline (PBS) solution at room temperature for 12 weeks and for 4 weeks at 37 °C. The effects of degradation were monitored by taking each sample out of the PBS solution every week, and the structural changes were investigated under a scanning electron microscope (SEM). The PCL and PLGA scaffolds showed excellent fibre structure with adequate degradation, and the fibre diameter, measured over time, showed slight increase in size. Therefore, as an example of fibre water intake and progressive degradation, the scaffold’s percentage weight loss increased each week, further supporting the porous membrane’s degradability. The pore size and the porosity percentage of all scaffolds decreased substantially over the degradation period. The conclusion drawn from this experiment is that PCL and PLGA hold great promise for tissue engineering and regenerative medicine applications.     

Experimental Analysis of the Enzymatic Degradation of Polycaprolactone: Microcrystalline Cellulose Composites and Numerical Method for the Prediction of the Degraded Geometry

The degradation rate of polycaprolactone (PCL) is a key issue when using this material in Tissue Engineering or eco-friendly packaging sectors. Although different PCL-based composite materials have been suggested in the literature and extensively tested in terms of processability by material extrusion additive manufacturing, little attention has been paid to the influence of the fillers on the mechanical properties of the material during degradation. This work analyses the possibility of tuning the degradation rate of PCL-based filaments by the introduction of microcrystalline cellulose into the polymer matrix. The enzymatic degradation of the composite and pure PCL materials were compared in terms of mass loss, mechanical properties, morphology and infrared spectra. The results showed an increased degradation rate of the composite material due to the presence of the filler (enhanced interaction with the enzymes). Additionally, a new numerical method for the prediction of the degraded geometry was developed. The method, based on the Monte Carlo Method in an iterative process, adjusts the degradation probability according to the exposure of each discretized element to the degradation media. This probability is also amplified depending on the corresponding experimental mass loss, thus allowing a good fit to the experimental data in relatively few iterations.     

Functionalization of Polycaprolactone Electrospun Osteoplastic Scaffolds with Fluorapatite and Hydroxyapatite Nanoparticles: Biocompatibility Comparison of Human Versus Mouse Mesenchymal Stem Cells

A capability for effective tissue reparation is a living requirement for all multicellular organisms. Bone exits as a precisely orchestrated balance of bioactivities of bone forming osteoblasts and bone resorbing osteoclasts. The main feature of osteoblasts is their capability to produce massive extracellular matrix enriched with calcium phosphate minerals. Hydroxyapatite and its composites represent the most common form of bone mineral providing mechanical strength and significant osteoinductive properties. Herein, hydroxyapatite and fluorapatite functionalized composite scaffolds based on electrospun polycaprolactone have been successfully fabricated. Physicochemical properties, biocompatibility and osteoinductivity of generated matrices have been validated. Both the hydroxyapatite and fluorapatite containing polycaprolactone composite scaffolds demonstrated good biocompatibility towards mesenchymal stem cells. Moreover, the presence of both hydroxyapatite and fluorapatite nanoparticles increased scaffolds’ wettability. Furthermore, incorporation of fluorapatite nanoparticles enhanced the ability of the composite scaffolds to interact and support the mesenchymal stem cells attachment to their surfaces as compared to hydroxyapatite enriched composite scaffolds. The study of osteoinductive properties showed the capacity of fluorapatite and hydroxyapatite containing composite scaffolds to potentiate the stimulation of early stages of mesenchymal stem cells’ osteoblast differentiation. Therefore, polycaprolactone based composite scaffolds functionalized with fluorapatite nanoparticles generates a promising platform for future bone tissue engineering applications.     

Bone Tissue Engineering with Adipose-Derived Stem Cells in Bioactive Composites of Laser-Sintered Porous Polycaprolactone Scaffolds and Platelet-Rich Plasma

Three-dimensional porous polycaprolactone (PCL) scaffolds with consistent inter-pore channels, 83% porosity and 300–400 μm pore size were fabricated via selective laser sintering. The PCL scaffold was combined with platelet-rich plasma (PRP) to form a bioactive composite and studied for potential application in bone tissue engineering using porcine adipose-derived stem cells (PASCs). The PCL/PRP/PASCs construct showed enhanced cell seeding efficiency and synergistically increased the differentiation capability of PASCs in osteogenic medium toward the osteoblast lineage, judging from elevated alkaline phosphatase activity and up-regulated osteogenic genes expression. For in vivo study, a 3 cm × 3 cm mandible defect was created in pigs and reconstructed by implanting acellular PCL scaffolds or PCL/PRP/PASCs constructs. Both groups showed new bone formation, however, the new bone volume was 5.1 times higher for PCL/PRP/PASCs 6 months post-operation. The bone density was less and loose in the acellular PCL group and the Young’s modulus was only 29% of normal bone. In contrast, continued and compact bone formation was found in PCL/PRP/PASCs and the Young’s modulus was 81% that of normal bone. Masson’s trichrome stain, immunohistochemical analysis of osteocalcin and collagen type I also confirmed new bone formation.