The use of poly(lactic-co-glycolic acid) microspheres as injectable cell carriers for cartilage regeneration in rabbit knees

The use of injectable scaffolding materials for in vivo tissue regeneration has raised great interest because it allows cell implantation through minimally invasive surgical procedures. Previously, we showed that poly(lactic-co-glycolic acid) (PLGA) microspheres can be used as an injectable scaffold to engineer cartilage in the subcutaneous space of athymic mice. The purpose of this study was to determine whether PLGA microspheres can be used as an injectable scaffold to regenerate hyaline cartilage in the osteochondral defects of rabbit knees. A full-thickness wound to the patellar groove of the articular cartilage was made in the knees of rabbits. Rabbit chondrocytes were mixed with PLGA microspheres and injected immediately into these osteochondral wounds. Both chondrocyte transplantations without PLGA microspheres and culture medium injections without chondrocytes served as controls. Sixteen weeks after implantation, chondrocytes implanted using the PLGA microspheres formed white cartilaginous tissues. Histological scores indicating the extent of the cartilaginous tissue repair and the absence of degenerative changes were significantly higher in the experimental group than in the control groups (P < 0.05). Histological analysis by a hematoxylin and eosin stain of the group transplanted with microspheres showed thicker and better-formed cartilage compared to the control groups. Alcian blue staining and Masson's trichrome staining indicated a higher content of the major extracellular matrices of cartilage, sulfated glycosaminoglycans and collagen in the group transplanted with microspheres than in the control groups. In addition, immunohistochemical analysis showed a higher content of collagen type II, the major collagen type in cartilage, in the microsphere transplanted group compared to the control groups. In the group transplanted without microspheres, the wounds were repaired with fibro-cartilaginous tissues. This study demonstrates the feasibility of using PLGA microspheres as an injectable scaffold for cartilage regeneration in a rabbit model of osteochondral wound repair.     

Poly(lactic-co-glycolic acid) microspheres as an injectable scaffold for cartilage tissue engineering

Injectable scaffold has raised great interest for tissue regeneration in vivo, because it allows easy filling of irregularly shaped defects and the implantation of cells through minimally invasive surgical procedures. In this study, we evaluated poly(lactic-co-glycolic acid) (PLGA) microsphere as an injectable scaffold for in vivo cartilage tissue engineering. PLGA microspheres (30-80 microm in diameter) were injectable through various gauges of needles, as the microspheres did not obstruct the needles and microsphere size exclusion was not observed at injection. The culture of chondrocytes on PLGA microspheres in vitro showed that the microspheres were permissive for chondrocyte adhesion to the microsphere surface. Rabbit chondrocytes were mixed with PLGA microspheres and injected immediately into athymic mouse subcutaneous sites. Chondrocyte transplantation without PLGA microspheres and PLGA microsphere implantation without chondrocytes served as controls. Four and 9 weeks after implantation, chondrocytes implanted with PLGA microspheres formed solid, white cartilaginous tissues, whereas no gross evidence of cartilage tissue formation was noted in the control groups. Histological analysis of the implants by hematoxylin and eosin staining showed mature and well-formed cartilage. Alcian blue/safranin O staining and Masson's trichrome staining indicated the presence of highly sulfated glycosaminoglycans and collagen, respectively, both of which are the major extracellular matrices of cartilage. Immunohistochemical analysis showed that the collagen was mainly type II, the major collagen type in cartilage. This study demonstrates the feasibility of using PLGA microspheres as an injectable scaffold for in vivo cartilage tissue engineering. This scaffold may be useful to regenerate cartilaginous tissues through minimally invasive surgical procedures in orthopedic, maxillofacial, and urologic applications.     

Apatite-coated poly(lactic-co-glycolic acid) microspheres as an injectable scaffold for bone tissue engineering

Biodegradable polymer/ceramic composite scaffold could overcome limitations of biodegradable polymers or ceramics for bone regeneration. Injectable scaffold has raised great interest for bone regeneration in vivo, since it allows one for easy filling of irregularly shaped bone defects and implantation of osteogenic cells through minimally invasive surgical procedures The purpose of this study was to determine whether apatite-coated poly(lactic-co-glycolic acid) (PLGA) microspheres could be used as an injectable scaffold to regenerate bone in vivo. Apatite-coated PLGA microspheres were fabricated by incubating PLGA microspheres in simulated body fluid. The apatite that coated the PLGA microsphere surfaces was similar to apatite in natural bone, as demonstrated by scanning electron microscopy, X-ray diffraction spectra, energy-dispersive spectroscopy, and Fourier transformed-infrared spectroscopy analyses. Rat osteoblasts were mixed with apatite-coated PLGA microspheres and injected immediately into subcutaneous sites of athymic mice. Osteoblast transplantation with plain PLGA microspheres served as a control. Histological analysis of the implants at 6 weeks with hematoxylin and eosin staining, Masson's trichrome staining, and von Kossa staining revealed much better regeneration of bone in the apatite-coated PLGA microsphere group than the plain PLGA microsphere group. The new bone formation area and the calcium content of the implants were significantly higher in the apatite-coated PLGA microsphere group than in the plain PLGA microsphere group. This study demonstrates the feasibility of using apatite-coated PLGA microspheres as an injectable scaffold for in vivo bone tissue engineering. This scaffold may be useful for bone regeneration through minimally invasive surgical procedures in orthopedic applications.     

The use of poly(l-lactide) and RGD modified microspheres as cell carriers in a flow intermittency bioreactor for tissue engineering cartilage

The use of biodegradable microcarriers as initial supports for tissue engineering has been demonstrated to be advantageous for maintaining a differentiated cell phenotype; the high surface area also allows rapid cell expansion. Poly l-lactide (PLLA) is a significant member of a group of polymers regarded as bioresorbable and has been widely used for manufacturing 3D scaffolds for tissue engineering. In this study, the hypothesis that PLLA microspheres could be surface modified using RGD peptide sequences to improve the cell adhesion and function of those cells in contact with PLLA was tested. Using this type of approach it may be possible to generate larger structures that contain a high cell number relative to the amount of polymer, whilst remaining free from mass transport limitations. PLLA microspheres were prepared using an oil-in-water solvent-evaporation technique and then an RGD-motif was incorporated onto the microspheres surface by conjugation to improve cell attachment and function. Both PLLA and GRGDSPK modified PLLA microspheres were used as cell microcarriers for chondrocytes cultured in a flow intermittency bioreactor. At the same time, the degradation of the microspheres has been studied after 7, 14, 21, 28, 35, 49 and 56 days. The molecular weight of the PLLA microspheres was determined by Gel Permeation Chromatography. The morphology was assessed by scanning electron microscopy, and the thermal properties determined by Differential Scanning Calorimetry. It was demonstrated that the RGD modified and pure PLLA microspheres degraded gradually at a steady rate over the experimental period, which would provide a controlled degradation profile, both could serve as cell microcarriers because of their thermal and mechanical stabilities. The microspheres with RGD surface modification enhanced cell adhesion and increased the cell numbers in the microspheres aggregates.     

利福平/聚乳酸-聚羟基乙酸缓释微球的制备及特性

背景：虽然国内外有很多制备利福平/聚乳酸-聚羟基乙酸共聚物(poly lactic acid-glycolic acid copolymer,PLGA)微球的报道,但这些微球粒径多在10 μm左右,不适合与磷酸钙骨水泥复合制备成具有良好降解性的抗结核修复材料。 目的：制备大粒径利福平/PLGA缓释微球,观察其理化特性和体外缓释特性。 方法：以PLGA为载体,将利福平分散于PLGA的有机溶剂中,采用复乳溶剂挥发法制备利福平/ PLGA缓释微球。光镜和扫描电镜下观察微球的形态特征,测定微球平均直径和跨距,高效液相色谱法测定载药量和包封率,以溶出法和高效液相色谱法观察其体外释药特性,并拟合药物体外释放曲线建立曲线方程。 结果与结论：利福平/PLGA微球电镜观察呈圆球形,分散性好,粘连少,粒径分布集中,平均粒径(80.0±9.4) μm。载药量、包封率分别为(33.18±1.36)%,(54.79±1.13)%。体外缓释试验显示突释期内微球释放度为(14.66±0.18)%,前3 d累计释放度(18.09±0.45)%,到42 d体外累积释放度达到(92.17±1.23)%。提示利福平/PLGA微球具有良好的缓释效果,是一种较为理想的抗结核药物的载体材料和释放系统;PLGA是良好的药物缓释载体,可以用来制备载药缓释微球。     

聚乳酸/聚羟基乙酸共聚物修复髌股关节软骨缺损

背景：传统的方法修复软骨损伤,易发生退变。聚乳酸/聚羟基乙酸共聚物具有良好的生物相容性,可根据需要调节降解速度等性能,可能在修复软骨损伤方面具有应用前景。 目的：观察以聚乳酸/聚羟基乙酸共聚物为载体修复兔关节软骨缺损的可行性。 方法：选取2月龄新西兰兔骨髓培养,诱导间充质干细胞向软骨细胞分化。第3代细胞与聚乳酸/聚羟基乙酸共聚物共培养制成聚乳酸/聚羟基乙酸共聚物-细胞复合物。建立兔髌股关节股骨髁部缺损模型,在右侧36个膝关节植入聚乳酸/聚羟基乙酸共聚物-细胞复合物,左侧18膝植入聚乳酸/聚羟基乙酸共聚物,另18膝造成缺损后留作空白对照。术后4,8,12,24,36,48周取材,行大体及组织学观察,组织学评分。 结果与结论：聚乳酸/聚羟基乙酸共聚物-细胞复合物修复大鼠缺损后,软骨细胞分布较均一,色泽与正常软骨相似,与正常软骨界限消失,表面细胞平行于关节面,深层细胞排列紊乱,细胞呈团状,基质异染广泛,软骨下骨形成及潮线恢复正常,与周围正常软骨连接良好。而单纯植入聚乳酸/聚羟基乙酸共聚物或缺损后未处理大鼠缺损边缘细胞呈团块状增生,底部为纤维组织。提示骨髓基质细胞源性软骨细胞是修复关节软骨缺损较理想的种子细胞,聚乳酸/聚羟基乙酸共聚物适合作为组织工程修复关节软骨缺损的支架材料,具有良好的应用前景。     

Cell selective chitosan microparticles as injectable cell carriers for tissue regeneration

The detection, isolation and sorting of cells holds an important role in cell therapy and regenerative medicine. Also, injectable systems have been explored for tissue regeneration in vivo, because it allows repairing complex shaped tissue defects through minimally invasive surgical procedures. Here we report the development of chitosan microparticles with a size of 115.8 μm able to capture and expand a specific cell type that can also be regarded as an injectable biomaterial. Monoclonal antibodies against cell surface antigens specific to endothelial cells and stem cells were immobilized on the surface of the microparticles. Experimental results showed that particles bioconjugated with specific antibodies provide suitable surfaces to capture a target cell type and subsequent expansion of the captured cells. Primarily designed for an application in tissue engineering, three main challenges are accomplished with the herein presented microparticles: separation, scale-up expansion of specific cell type and successful use as an injectable system to form small tissue constructs in situ.     

Poly(l-glutamic acid)/chitosan polyelectrolyte complex porous microspheres as cell microcarriers for cartilage regeneration

In this study a novel kind of porous poly(l-glutamic acid) (PLGA)/chitosan polyelectrolyte complex (PEC) microsphere was developed through electrostatic interaction between PLGA and chitosan. By adjusting the formula parameters chitosan microspheres with an average pore size of 47.5 ± 5.4 μm were first developed at a concentration of 2 wt.% and freeze temperature of −20 °C. For self-assembly of the PEC microspheres porous chitosan microspheres were then incubated in PLGA solution at 37 °C. Due to electrostatic interaction a large amount of PLGA (110.3 μg mg⁻¹) was homogeneously absorbed within the chitosan microspheres. The developed PEC microspheres retained their original size, pore diameters and interconnected porous structure. Fourier transform infrared spectroscopy, thermal gravimetric analysis and zeta potential analysis revealed that the PEC microspheres were successfully prepared through electrostatic interaction. Compared with microspheres fabricated from chitosan, the porous PEC microspheres were shown to efficiently promote chondrocyte attachment and proliferation. After injection subcutaneously for 8 weeks PEC microspheres loaded with chondrocytes were found to produce significant more cartilaginous matrix than chitosan microspheres. These results indicate that these novel fabricated porous PLGA/chitosan PEC microspheres could be used as injectable cell carriers for cartilage tissue engineering.     

载羟基喜树碱聚乳酸-羟基乙酸微球的制备及相关性能研究

目的制备一种载羟基喜树碱的聚乳酸-羟基乙酸(PLGA)缓释微球,并考察其相关性能。方法采用乳化-溶剂挥发法制备羟基喜树碱PLGA微球,用扫描电子显微镜观察载药微球表面形态,测定平均粒径及跨距,高效液相色谱检测包封率、载药率及体外释放情况,改良寇氏法计算小鼠半数致死量。结果制备的载药PLGA微球呈圆球形,表面光滑,无粘连,平均粒径30.8μm,跨距0.9,包封率为85.5%、载药率4.28%,在体外28 d累积释放药物81.4%。羟基喜树碱小鼠静脉注射的半数致死量为18.4 mg/kg,肌内注射半数致死量为71.3 mg/kg,而羟基喜树碱PLGA微球肌内注射的半数致死量为138.5 mg/kg。结论乳化-溶剂挥发法制备的羟基喜树碱PLGA微球粒径适宜,包封率、载药率高,缓释效果好,毒性低,具有潜在的临床应用价值。     

The release profiles and bioactivity of parathyroid hormone from poly(lactic-co-glycolic acid) microspheres

Poly(lactic-co-glycolic acid) (PLGA) microspheres containing bovine serum albumin (BSA) or human parathyroid hormone (PTH)(1-34) were prepared using a double emulsion method with high encapsulation efficiency and controlled particle sizes. The microspheres were characterized with regard to their surface morphology, size, protein loading, degradation and release kinetics, and in vitro and in vivo assessments of biological activity of released PTH. PLGA5050 microspheres degraded rapidly after a 3-week lag time and were degraded completely within 4 months. In vitro BSA release kinetics from PLGA5050 microspheres were characterized by a burst effect followed by a slow release phase within 1-7 weeks and a second burst release at 8 weeks, which was consistent with the degradation study. The PTH incorporated PLGA5050 microspheres released detectable PTH in the initial 24h, and the released PTH was biologically active as evidenced by the stimulated release of cAMP from ROS 17/2.8 osteosarcoma cells as well as increased serum calcium levels when injected subcutaneously into mice. Both in vitro and in vivo assays demonstrated that the bioactivity of PTH was maintained largely during the fabrication of PLGA microspheres and upon release. These studies illustrate the feasibility of achieving local delivery of PTH to induce a biologically active response in bone by a microsphere encapsulation technique.     

Submucosal injection of poly(lactic-co-glycolic acid) microspheres in rabbit bladder as a potential treatment for urinary incontinence and vesicoureteral reflux: preliminary results

Endoscopic injection of bulking agents has been gaining attention as a therapy for urinary incontinence and vesicoureteral reflux because this therapy is simpler, less operation time-consuming and less painful than traditional surgical operations. The ideal bulking agent for the injection therapies must be easily injectable, biocompatible, volume-stable, non-antigenic and non-migratory. We evaluated poly(lactic-co-glycolic acid) (PLGA) microspheres as an injectable bulking agent for urologic injection therapies. To determine whether PLGA microspheres meet the requirements of an ideal bulking agent, PLGA microspheres were injected into the submucosal sites of a rabbit bladder wall. The microspheres were easily injectable. Two and five weeks post-implantation, histological examinations indicated that host cells from the surrounding bladder tissues migrated to the space between the injected microspheres and formed new hybrid tissue structures. Lymphocyte migration was noted around the implanted microspheres, but the inflammatory reaction diminished at 5 weeks. The hybrid tissue volume did not significantly decrease over time. There was no evidence of microsphere migration to the distant organs. Although long-term studies are needed to evaluate the therapeutic potential of this method, these preliminary results suggest the possibility of PLGA microspheres as a potentially useful injection material for urinary injection therapies.     

Submucosal injection of poly(lactic-co-glycolic acid) microspheres in rabbit bladder as a potential treatment for urinary incontinence and vesicoureteral reflux: preliminary results

Endoscopic injection of bulking agents has been gaining attention as a therapy for urinary incontinence and vesicoureteral reflux because this therapy is simpler, less operation time-consuming and less painful than traditional surgical operations. The ideal bulking agent for the injection therapies must be easily injectable, biocompatible, volume-stable, non-antigenic and non-migratory. We evaluated poly(lactic-co-glycolic acid) (PLGA) microspheres as an injectable bulking agent for urologic injection therapies. To determine whether PLGA microspheres meet the requirements of an ideal bulking agent, PLGA microspheres were injected into the submucosal sites of a rabbit bladder wall. The microspheres were easily injectable. Two and five weeks post-implantation, histological examinations indicated that host cells from the surrounding bladder tissues migrated to the space between the injected microspheres and formed new hybrid tissue structures. Lymphocyte migration was noted around the implanted microspheres, but the inflammatory reaction diminished at 5 weeks. The hybrid tissue volume did not significantly decrease over time. There was no evidence of microsphere migration to the distant organs. Although long-term studies are needed to evaluate the therapeutic potential of this method, these preliminary results suggest the possibility of PLGA microspheres as a potentially useful injection material for urinary injection therapies.     

Immune responses in mice of beta-galactosidase adsorbed or encapsulated in poly(lactic acid) and poly(lactic-co-glycolic acid) microspheres

The immune response induced in mice by beta-galactosidase (beta-gal) adsorbed or encapsulated on poly(lactic acid) (PLA) and poly(lactic-co-glycolic acid) (PLGA) microspheres was investigated. The encapsulated protein elicited higher antibody response than the protein adsorbed on the microspheres in the case of the PLA microspheres. However, the encapsulated protein elicited weaker antibody response than the adsorbed protein in the case of the PLGA (50:50) microspheres, probably because, in this case, the encapsulation process adversely affected protein immunogenicity. In the case of adsorbed beta-gal, higher antibody response was obtained with the PLA microspheres than with the PLGA (50:50) microspheres. This may be related to the lower rate of beta-gal desorption from the PLA microspheres. Based on the immunoglobulin G1/immunoglobulin G2a ratios and the stimulation indices for interferon-gamma and interleukin-4, beta-gal encapsulated or adsorbed on PLA microspheres induced a Th(1)-biased immune response whereas beta-gal encapsulated or adsorbed on PLGA (50:50) microspheres induced a Th(2)-biased immune response. The results obtained indicate that more potent immune responses are obtained when the protein is encapsulated than adsorbed on the microspheres, providing that the encapsulation process does not adversely affect protein immunogenicity. Also, the type of polymer used to prepare the microspheres, but not the method of protein association with the microspheres, may affect the type of immune response.     

乳酸-羟基乙酸共聚物缓控释微球的制备及突释成因与影响因素

背景：乳酸-羟基乙酸共聚物是一种生物可降解高分子材料,以乳酸-羟基乙酸共聚物为原料制备的载药微球和纳米粒既可提高药物的稳定性,又能实现缓释、控释和靶向释放。 目的：分析乳酸-羟基乙酸共聚物缓控释微球的制备方法以及突释的成因、影响因素和改进方法。 方法：应用计算机检索1990/2010中国期刊全文数据库和PubMed数据库与乳酸-羟基乙酸共聚物缓控释微球的制备及突释联系紧密的文章。 结果与结论：目前乳酸-羟基乙酸共聚物缓释微球制备方法主要有单凝聚法、乳化-固化法、喷雾干燥法。造成其突释的原因首先是药物分子和聚合物分子之间的相互作用太弱,导致药物很容易从微球进入释放递质中,其次是在微球释放初期,药物从微球中的孔洞和缝隙中释放出来导致突释。影响突释程度的具体因素有乳酸-羟基乙酸共聚物的相对分子质量、浓度、微球载药量、主药理化性质、微球制备方法及制备参数等。虽然国内外对突释机制以及控制突释措施的研究都还处于初步阶段,通过对各影响因素加以适当优化与控制,可在一定程度上减少微球的突释率,突释问题应该能够得到解决和控制。     

Preparation and properties of an injectable scaffold of poly(lactic-co-glycolic acid) microparticles/chitosan hydrogel

Hydrogels are more and more attractive in biomedical fields, since they can be used as injectable scaffolds, drugs and gene carriers and smart sensors. The highly hydrated hydrogels, however, generally have low mechanical strength. In this work, a composite chitosan hydrogel was prepared by blending water soluble and crosslinkable chitosan derivative (CML) with poly(lactic-co-glycolic acid) (PLGA) particles whose surfaces were grafted with double carbon bonds containing gelatin (GM), following gelation under UV irradiation. The as-prepared composite hydrogel showed lower swelling ratio than that of the CML hydrogel, and higher elastic stiffness (i.e. storage modulus) than that of the CML hydrogel and the hydrogel filled with the same amount of PLGA particles or gelatin modified PLGA particles. Moreover, the storage modulus of the composite hydrogel was increased with the amount of GM modified PLGA particles. In vitro chondrocyte culture revealed that viability of the cells co-cultured with the GM modified PLGA particles was higher than that of the cells co-cultured with the unmodified PLGA particles. The composite hydrogel blended with the GM modified PLGA particles also showed higher cytoviability than that of the original CML hydrogel after 9d culture.     

Porous poly(lactic-co-glycolic acid) microsphere as cell culture substrate and cell transplantation vehicle for adipose tissue engineering

Tissue engineering often requires ex vivo cell expansion to obtain a large number of transplantable cells. However, the trypsinization process used to harvest ex vivo expanded cells for transplantation interrupts interactions between cultured cells and their extracellular matrices, facilitating apoptosis and consequently limiting the therapeutic efficacy of the transplanted cells. In the present study, open macroporous poly(lactic-co-glycolic acid) (PLGA) microspheres were used as a cell culture substrate to expand human adipose-derived stromal cells (ASCs) ex vivo and as a cell transplantation vehicle for adipose tissue engineering, thus avoiding the trypsinization necessary for transplantation of ex vivo expanded cells. Human ASCs cultured on macroporous PLGA microspheres in stirred suspension bioreactors expanded 3.8-fold over 7 days and differentiated into an adipogenic lineage. The apoptotic activity of ASCs cultured on microspheres was significantly lower than that of trypsinized ASCs. ASCs cultured on microspheres survived much better than trypsinized ASCs upon transplantation. The implantation of ASCs cultured on microspheres resulted in much more extensive adipose tissue formation than the implantation of ASCs cultured on plates, trypsinized, and subsequently mixed with microspheres. Ex vivo cell expansion and transplantation using this system would improve the therapeutic efficacy of cells over the current methods used for tissue engineering.     

Highly porous large poly(lactic-co-glycolic acid) microspheres adsorbed with palmityl-acylated exendin-4 as a long-acting inhalation system for treating diabetes

A porous large poly(lactic-co-glycolic acid) (PLGA) microspheres (MS) adsorbed with palmityl-acylated exendin-4 (Ex4-C(16)) was devised as an inhalation delivery system. The porous MS was prepared by a single o/w emulsification/solvent evaporation method using extractable Pluronic F68/F127, and its fabrication and formulation conditions were carefully optimized. Results show that the prepared MS was in the appropriate size range for inhalation and contained many surfaces and internal pores meaning low aerodynamic density. Ex4-C(16) was more efficiently adsorbed onto porous PLGA MSs than native exendin-4, and an approximately 5% loading of Ex4-C(16) onto this porous MS (RG504H) was achieved. This optimized porous MS was found to be efficiently deposited throughout the entire lungs of mice including alveoli region. Furthermore, this porous MS adsorbed with Ex4-C(16) (approx. 100?μg/mouse) displayed much protracted hypoglycemic efficacy in non-fasted type 2 diabetic db/db mice. Porous PLGA MS with adsorbed Ex4-C(16) showed the dual-advantages of (i) sustained release and acceptable drug-loading due to strong hydrophobic interaction and (ii) longer in vivo pulmonary hypoglycemic duration due to albumin-binding by the palmityl group. We consider that this new prototype of porous PLGA MS has considerable pharmaceutical potential as a type 2 anti-diabetic inhalation treatment.     

Development of biodegradable poly(propylene fumarate)/poly(lactic-co-glycolic acid) blend microspheres. I. Preparation and characterization

We developed poly(propylene fumarate)/poly(lactic-co-glycolic acid) (PPF/PLGA) blend microspheres and investigated the effects of various processing parameters on the characteristics of these microspheres. The advantage of these blend microspheres is that the carbon-carbon double bonds along the PPF backbone could be used for their immobilization in a PPF scaffold. Microspheres containing the model drug Texas red dextran were fabricated using a double emulsion-solvent extraction technique. The effects of the following six processing parameters on the microsphere characteristics were investigated: PPF/PLGA ratio, polymer viscosity, vortex speed during emulsification, amount of internal aqueous phase, use of poly(vinyl alcohol) (PVA) in the internal aqueous phase, and PVA concentration in the external aqueous phase. Our results showed that the microsphere surface morphology was affected most by the viscosity of the polymer solution. Microspheres fabricated with a kinematic viscosity of 39 centistokes had a smooth, nonporous surface. In most microsphere formulations, the model drug was dispersed uniformly in the polymer matrix. For all fabricated formulations, the average microsphere diameter ranged between 19.0 and 76.9 microm. The external PVA concentration and vortex speed had most effect on the size distribution. Entrapment efficiencies varied from 60 to 98% and were most affected by the amount of internal aqueous phase, vortex speed, and polymer viscosity. Overall, we demonstrated the ability to fabricate PPF/PLGA blend microspheres with similar surface morphology, entrapment efficiency, and size distribution as conventional PLGA microspheres.     

构建聚乳酸-羟基乙酸电纺丝-壳聚糖电喷微球牙周仿生膜

文题释义： 离子交联法制备壳聚糖微球：离子交联法利用酸性环境下壳聚糖呈阳离子性,向壳聚糖溶液中边搅拌边滴加三聚磷酸钠,带负电的磷酸根离子与壳聚糖分子链上带正电荷的氨基通过静电吸附,从而形成微球。 牙周膜：是连接牙齿和牙槽骨之间具有方向性的结缔组织,宽度为0.15-0.38 mm,其内含具有一定方向性胶原纤维束,其一端埋入牙骨质,另一端伸入牙槽骨内,具有固定牙根和缓解咀嚼时所产生压力的作用,又称牙周关节;牙周膜能形成牙槽骨及牙骨质,被破坏后能重建。由此可见,牙周膜不是普通的纤维结缔组织,它具有方向性、附着点、可再生牙周组织(牙骨质、牙周膜、牙槽骨)。 背景：当牙齿脱离牙槽窝后,牙周膜断裂,残留在脱位牙根表面的牙周膜由三维变成二维,丧失了支架膜的作用,导致脱位牙再植后根骨粘连。如何研发一种能黏附牙根表面具有一定厚度及强度的三维缓释支架材料,是脱位牙牙周膜再生成功的关键之一。 目的：构建可黏附脱位牙根表面的缓释生长因子的三维仿生膜。 方法：采用静电纺丝技术制备聚乳酸-羟基乙酸电纺膜,研究电纺溶剂二氯甲烷与二甲基甲酰胺混合溶液、六氟异丙醇、三氯甲烷对电纺膜的影响,筛选最佳的电纺溶剂。采用电喷技术与离子交联法制备壳聚糖微球,研究壳聚糖相对分子质量(5万、10万)与质量浓度(10,20 g/L)、接受液三聚磷酸钠浓度(2%,5%,10%)、电压(14,28 kV)对壳聚糖微球的影响,筛选最佳的参数。构建含基质细胞衍生因子1壳聚糖微球(最优参数设计),检测其体外释放基质细胞衍生因子1α的速率。首先将聚乳酸-羟基乙酸电纺膜裹在牙齿根表面,然后在其表面滴加壳聚糖微球,在其外层裹一层薄薄的聚乳酸-羟基乙酸电纺膜,从而形成聚乳酸-羟基乙酸-壳聚糖微球-聚乳酸-羟基乙酸膜。 结果与结论：①利用电纺溶剂六氟异丙醇制备的聚乳酸-羟基乙酸电纺膜平均直径最小、空隙率最大;②当壳聚糖相对分子质量为5万、质量浓度为20 g/L时,微球的大小基本一致,平均直径366.6 μm,单分散性好、饱满、稳定;28 kV电压下形成的壳聚糖微球更符合脱位牙仿生膜的要求;利用5%三聚磷酸钠制备的壳聚糖微球表面微观结构孔径居中,最有利于临床牙周膜再生;壳聚糖微球可持续释放基质细胞衍生因子1α 1个月左右;③实验创建了一种黏附牙齿表面的具有缓释效能的聚乳酸-羟基乙酸-壳聚糖微球-聚乳酸-羟基乙酸三维仿生膜并筛选出构建此仿生膜的最佳参数,可在此模型基础上进一步研究组织工程手段对脱位牙再植的效果及机制。 ORCID: 0000-0003-3957-3423(封小霞) 中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程     

Poly(D,L lactic-co-glycolic acid) microspheres as biodegradable microcarriers for pluripotent stem cells

The pluripotent nature and proliferative capacity of embryonic stem cells makes them an attractive cell source for tissue engineering and regeneration. In our study we investigated the use of poly(D,L-lactic-co-glycolic acid) (PLGA) microspheres as biodegradable microcarriers of pluripotent cells and as delivery systems of bioactive factors, which influence cell differentiation. The pluripotent P19 embryonal carcinoma cell line was used as a model to study cell attachment, growth and differentiation of pluripotent stem cells on PLGA microspheres. Retinoic acid (RA) was encapsulated in the PLGA microcarriers to influence cell differentiation-more specifically, to induce P19 cell differentiation into neurons. The results revealed that P19 cells attach and grow on the surface of the RA loaded PLGA microspheres. Moreover, the RA loaded PLGA microspheres were shown to be as effective as soluble RA at inducing P19 cell differentiation into neurons. Hence, the results of these ex vivo studies clearly demonstrate the capacity of PLGA microspheres to serve a dual role as both delivery systems of bioactive factors and as scaffolds for pluripotent cells. More importantly, our study demonstrates the potential use of PLGA microspheres as transplantation matrices of pluripotent stem cells for tissue engineering and regeneration.