Endothelial and vascular smooth muscle cell function on poly(lactic-co-glycolic acid) with nano-structured surface features

Biomaterials that successfully integrate into surrounding tissue should match not only the tissue's mechanical properties, but also its topography. The cellular response to a biomaterial may be enhanced in synthetic polymer formulations by mimicking the surface roughness created by the associated nano-structured extra-cellular matrix components of natural tissue. As a first step towards this endeavor, the goal of the present in vitro study was to use these design parameters to develop a synthetic, nano-structured, polymeric biomaterial that promotes cell adhesion and growth for vascular applications. In a novel manner, poly(lactic-co-glycolic acid) (PLGA) (50/50wt% mix) was synthesized to possess a range (from micron to nanometer) of surface features. Reduction of surface features was accomplished by treating conventional PLGA with various concentrations of NaOH for select periods of time. Results from cell experiments indicated that, compared to conventional PLGA, NaOH treated PLGA enhanced vascular smooth muscle cell adhesion and proliferation. However, PLGA prepared by soaking in NaOH decreased endothelial cell adhesion and proliferation compared to conventional PLGA. After further investigation, this finding was determined to be a result of chemical (and not topographical) changes during polymer synthesis. Surface chemistry effects were removed while retaining nano-structured topography by using polymer/elastomer casting methods. Results demonstrated that endothelial and smooth muscle cell densities increased on nano-structured cast PLGA. For these reasons, the present in vitro study provided the first evidence that nano-structured surface features can significantly improve vascular cell densities; such design criteria can be used in the synthesis of the next-generation of more successful tissue-engineered vascular grafts.     

Differential effects of agarose and poly(lactic-co-glycolic acid) on dendritic cell maturation

Application of biomaterials in combination products in which the biomaterial is presented to the host with a biological component prompts the need for understanding the biomaterial-associated adjuvant effect in the immune response against antigens associated with such a product. We have previously demonstrated that a polymer commonly used in tissue engineering and vaccine delivery, poly(lactic-co-glycolic acid) (PLGA), exerts an adjuvant effect in vivo, which was supported by PLGA-induced dendritic cell (DC) maturation in vitro. In this study, the effects of agarose and PLGA on DC maturation were compared in vitro to establish differential biomaterial effects. Human monocyte-derived DCs were treated with agarose or PLGA microparticles or films, and their maturation effect was measured as expression of costimulatory and MHC class II molecules, allostimulatory capacity, and proinflammatory cytokine secretion. Direct comparison of DC maturation phenotype indicated that PLGA was a stronger stimulus of DC maturation than agarose, and this maturation was not affected by microparticle phagocytosis. However, agarose-treated DCs showed higher activation of nuclear factor kappaB (NFkappaB) 24 h after the initial stimulation of DCs. Taken together, these results demonstrate differential biomaterial effects on DC maturation, substantiating the maturation effect of PLGA, and provide screening methods for biomaterial adjuvant effect for applications in combination products.     

Diazeniumdiolate-doped poly(lactic-co-glycolic acid)-based nitric oxide releasing films as antibiofilm coatings

Nitric oxide (NO) releasing films with a bilayer configuration are fabricated by doping dibutyhexyldiamine diazeniumdiolate (DBHD/N(2)O(2)) in a poly(lactic-co-glycolic acid) (PLGA) layer and further encapsulating this base layer with a silicone rubber top coating. By incorporating pH sensitive dyes within the films, pH changes in the PLGA layer are visualized and correlated with the NO release profiles (flux vs. time). It is demonstrated that PLGA acts as both a promoter and controller of NO release from the coating by providing protons through its intrinsic acid residues (both end groups and monomeric acid impurities) and hydrolysis products (lactic acid and glycolic acid). Control of the pH changes within the PLGA layer can be achieved by adjusting the ratio of DBHD/N(2)O(2) and utilizing PLGAs with different hydrolysis rates. Coatings with a variety of NO release profiles are prepared with lifetimes of up to 15 d at room temperature (23?°C) and 10 d at 37?°C. When incubated in a CDC flow bioreactor for a one week period at RT or 37?°C, all the NO releasing films exhibit considerable antibiofilm properties against gram-positive Staphylococcus aureus and gram-negative Escherichia coli. In particular, compared to the silicone rubber surface alone, an NO releasing film with a base layer of 30?wt% DBHD/N(2)O(2) mixed with poly(lactic acid) exhibits an ～98.4% reduction in biofilm biomass of S.?aureus and ～99.9% reduction for E.?coli at 37?°C. The new diazeniumdiolate-doped PLGA-based NO releasing coatings are expected to be useful antibiofilm coatings for a variety of indwelling biomedical devices (e.g., catheters).     

Effect of acidic degradation products of poly(lactic-co-glycolic)acid on the apatite-forming ability of poly(lactic-co-glycolic)acid-siloxane nanohybrid material

The effect of poly(lactic-co-glycolic) acid (PLGA) degradation products on the apatite-forming ability of a PLGA-siloxane nanohybrid material were investigated. Two PLGA copolymer compositions with low and high degradability were used in the experiment. The PLGA-siloxane nanohybrid materials were synthesized by end-capping PLGA with acid end-groups using 3-isocyanatopropyl triethoxysilane following the sol-gel reaction with calcium nitrate tetrahydrate. Two nanohybrid materials that had different degradability were exposed to simulated body fluid (SBF) for 1-28 days at 36.5 degrees C. The low degradable PLGA hybrid showed apatite-forming ability within 3 days of incubation while the high degradable one did not within 28 days testing period. The results were explained in terms of the acidity of the PLGA degradation products, which could directly influence on the apatite dissolution.     

Platelet adhesion studies on nanostructured poly(lactic-co-glycolic-acid)-carbon nanotube composite

Design of blood-compatible surfaces is required to minimize platelet-surface interactions and increase the thromboresistance of foreign surfaces. Poly(lactic-co-glycolic-acid)-carbon nanotube (PLGA-CNT) composite is studied as a building material to fabricate artificial blood prostheses. This nanocomposite-based biomaterial is prepared by an electrostatic Layer-by-Layer (LbL) deposition technique, in which layers of CNTs are adsorbed onto a PLGA film. Before incubation in nonstimulated platelet-rich plasma (PRP) for platelet studies, fibrinogen is immobilized on PLGA-CNT composite. Interactions between the plasma proteins, e.g. fibrinogen and PRP, are investigated on the prepared PLGA-CNT composite. Contact angle measurements on the PLGA-CNT composite displayed a good resistance of platelets adhesion on a hydrophilic surface with an angle of 64.94 degrees as compared to pristine PLGA control with an angle of 93.43 degrees . A significant reduction of adhesion is observed on the PLGA-CNT composite, as well as the absence of platelet activation. On the contrary, both platelet adhesion and activation are observed on control samples. We inferred this suppression in secretion of granule contents in the platelet by the presence of the CNTs that resulted in the absence of platelet activation and its subsequent inhibition in the release of adhesive membrane receptors on the PLGA-CNT composite.     

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。     

Nanomechanical properties of poly(lactic-co-glycolic) acid film during degradation

Despite the potential applications of poly(lactic-co-glycolic) acid (PLGA) coatings in medical devices, the mechanical properties of this material during degradation are poorly understood. In the present work, the nanomechanical properties and degradation of PLGA film were investigated. Hydrolysis of solvent-cast PLGA film was studied in buffer solution at 37 °C. The mass loss, water uptake, molecular weight, crystallinity and surface morphology of the film were tracked during degradation over 20 days. Characterization of the surface hardness and Young’s modulus was performed using the nanoindentation technique for different indentation loads. The initially amorphous films were found to remain amorphous during degradation. The molecular weight of the film decreased quickly during the initial days of degradation. Diffusion of water into the film resulted in a reduction in surface hardness during the first few days, followed by an increase that was due to the surface roughness. There was a significant delay between the decrease in the mechanical properties of the film and the decrease in the molecular weight. A sudden decline in mechanical properties indicated that significant bulk degradation had occurred.     

Bioinspired mineralization and cell adhesion on surface functionalized poly(vinyl alcohol) films

Poly(vinyl alcohol) (PVA) films, when surface functionalized by phosphorylation, induced biomimetic nucleation and growth of calcium phosphate in a simulated physiological environment. The surface phosphorylation on PVA was ensured by attenuated total reflectance infrared spectroscopy. The morphology of the calcium phosphate phase grown on surface-phosphorylated PVA (PPVA) was analysed using scanning electron microscopy coupled with an energy-dispersive X-ray detector. The primary nucleation of calcium phosphate occurs in 3 days and secondary nucleation occurs after 10 days. The energy-dispersive X-ray analysis shows that the Ca/P ratio of the coating increases with time of exposure to the simulated physiological fluid and reaches 1.67 at 10 days. The PPVA supports in vitro cell adhesion and promotes in vitro biomineralization in the presence of cells, evaluated using human osteosarcoma cells.     

In vitro evaluation of biodegradation of poly(lactic-co-glycolic acid) sponges

Evaluation of the degradability of porous scaffolds is very important for tissue engineering. A protocol in which the condition is close to the in vivo pH environment was established for in vitro evaluation of biodegradable porous scaffolds. Degradation of PLGA sponges in phosphate-buffered solution (PBS) was evaluated with the protocol. The PLGA sponges degraded with incubation time. For the first 12 weeks, the weight loss increased gradually and then remarkably after 12 weeks. In contrast, the number-average molecular weight (Mn) decreased dramatically for the first 12 weeks and then less markedly after 12 weeks. Thermal analysis showed that the glass transition temperatures (Tg) decreased rapidly for the first 12 weeks, and the change became less evident after 12 weeks. These results suggest that the degradation mechanism of PLGA sponges was dominated by autocatalyzed bulk degradation for the first 12 weeks and then by surface degradation after 12 weeks. Physical aging was observed during incubation at 37 degrees C. The heterogeneous structure caused by physical aging might be one of the driving forces that induced autocatalyzed bulk degradation. The degradation mechanism was further supported by the data of pH change and the morphology of the degraded PLGA sponges. The autocatalyzed acidic products flooded out after 8 weeks, the pH dropped, and the walls of the sponges became more porous. The increase of the pore surface area facilitated surface degradation after 12 weeks. The pH was in the range between 7.43 and 7.24 during the entire incubation time. The protocol suppressed extreme changes of the pH and will be useful in the biodegradation evaluation of porous scaffolds for tissue engineering.     

Biocompatibility of electrospun halloysite nanotube-doped poly(lactic-co-glycolic acid) composite nanofibers

Organic/inorganic hybrid nanofiber systems have generated great interest in the area of tissue engineering and drug delivery. In this study, halloysite nanotube (HNT)-doped poly(lactic-co-glycolic acid) (PLGA) composite nanofibers were fabricated via electrospinning and the influence of the incorporation of HNTs within PLGA nanofibers on their in vitro biocompatibility was investigated. The morphology, mechanical and thermal properties of the composite nanofibers were characterized by scanning electron microscopy (SEM), tensile test, differential scanning calorimetry and thermogravimetric analysis. The adhesion and proliferation of mouse fibroblast cells cultured on both PLGA and HNT-doped PLGA fibrous scaffolds were compared through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay of cell viability and SEM observation of cell morphology. We show that the morphology of the PLGA nanofibers does not appreciably change with the incorporation of HNTs, except that the mean diameter of the fibers increased with the increase of HNT incorporation in the composite. More importantly, the mechanical properties of the nanofibers were greatly improved. Similar to electrospun PLGA nanofibers, HNT-doped PLGA nanofibers were able to promote cell attachment and proliferation, suggesting that the incorporation of HNTs within PLGA nanofibers does not compromise the biocompatibility of the PLGA nanofibers. In addition, we show that HNT-doped PLGA scaffolds allow more protein adsorption than those without HNTs, which may provide sufficient nutrition for cell growth and proliferation. The developed electrospun HNT-doped composite fibrous scaffold may find applications in tissue engineering and pharmaceutical sciences.     

Fabrication and characterization of hydrophilic poly(lactic-co-glycolic acid)/poly(vinyl alcohol) blend cell scaffolds by melt-molding particulate-leaching method

Porous PLGA/PVA scaffolds were fabricated by blending poly(lactic-co-glycolic acid) (PLGA) with polyvinyl alcohol (PVA) to improve the hydrophilicity and cell compatibility of the scaffolds for tissue engineering applications. PLGA/PVA blend scaffolds with different PVA compositions up to 20wt% were fabricated by a melt-molding particulate-leaching method (non-solvent method). The prepared scaffolds were investigated by scanning electron microscopy (SEM), mercury intrusion porosimetry, the measurements of water contact angles and bi-axial tensile strengths, etc. for their surface and bulk characterizations. The scaffolds exhibited highly porous and open-cellular pore structures with almost same surface and interior porosities (pore size, 200-300 microm; porosity, about 90%). The PLGA/PVA blend scaffolds with PVA compositions more than 5% were easily wetted in cell culture medium without any prewetting treatments, which is highly desirable for tissue engineering applications. In vitro cell compatibility of the control hydrophobic PLGA and hydrophilized PLGA/PVA (5wt%) blend scaffolds was compared by the culture of human chondrocytes in the scaffolds and the following analyses by MTT assay and SEM observation. It was observed that the PLGA/PVA blend scaffold had better cell adhesion and growth than the control PLGA scaffold. For in vivo evaluation of tissue compatibility, the scaffolds were implanted into the skull defects of rabbits. The results were evaluated by histology examinations. The PLGA/PVA (5wt%) blend scaffold showed better bone ingrowth into the scaffold and new bone formation inside the scaffold than the PLGA scaffold. It seems that 5% addition of PVA to PLGA to fabricate PLGA/PVA blend scaffolds is enough for improving the hydrophilicity and cell compatibility of the scaffolds.     

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。     

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.     

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.     

纳米可降解聚乳酸-羟基乙酸尿道支架的制备及性能

背景：聚乳酸-羟基乙酸可作为尿道替代物进行组织缺损的修复。 目的：观察电纺丝法制备聚乳酸-羟基乙酸共聚物可降解尿道支架的可行性,并评价支架管的体外降解性能。 方法：采用电纺丝技术制备纳米聚乳酸-羟基乙酸共聚物(摩尔比80∶20)尿道支架管,并以戊二醛对支架进行交联、改性,将交联后支架截成长约1 cm小段并浸于尿液中进行体外降解实验。 结果与结论：支架管具有纳米结构,孔隙率约89%,孔径(32±19) µm;交联后可见纤维表面变粗糙,但纤维丝直径、孔径及孔隙率与交联前差异无显著性意义(P > 0.05),但交联后支架管力学性能显著提高。支架降解初期速度相对较快,中后期降解速度减慢,至8周时材料质量损失约50%,第10周完全崩解。材料在体内降解过程中相对分子质量的变化趋势与质量损失大体相同,降解早期相对分子质量下降相对较快,后期下降速度减慢并趋于平稳。表明采用电纺丝技术制备的纳米聚乳酸-羟基乙酸共聚物尿道支架可满足尿道组织工程支架的要求。     

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。     

Modulation of protein adsorption and cell adhesion by poly(allylamine hydrochloride) heparin films

Electrostatic layer-by-layer film assembly is an attractive way to non-covalently incorporate proteins and bioactive moieties into the surface of conventional biomaterials. Selection of polycationic and polyanionic components and deposition conditions can be used to control the interfacial properties, and through them protein adsorption, cell adhesion, and tissue development. In this study the polycation was poly(allylamine hydrochloride) (PAH), which is a weak base and consequently adsorbs at interfaces in a pH-dependent manner, and the polyanion was heparin, which is capable of interacting with many adhesion ligands and growth factors. PAH/heparin multilayer films were formed using PAH solutions of pH 6.4, 7.4, 8.4, and 9.4. Film thickness increased both with the number of PAH/heparin bilayers and the pH of the PAH solution. Films consisting of 10 bilayers with heparin topmost exhibited similar bulk atomic compositions and penetration of PAH into the heparin top layer. Finally, fibronectin adsorption and cell adhesion were maximal at an intermediate pH (pH 8.4>pH 9.4>pH 7.4). These results demonstrate that heparin-containing electrostatic films support cell adhesion and protein adsorption in a manner sensitive to film deposition conditions.     

Control of cell adhesion on poly(methyl methacrylate)

Keratoprostheses have been constructed from a wide variety of transparent materials, including poly(methyl methacrylate) (PMMA). However, the success of keratoprosthesis has been plagued by numerous shortcomings that include the weakening of the implant-host interface due to weak cell adhesion and opaque fibrous membrane formation over the inner surface of the implant due to fibroblast attachment. An effective solution requires a surface modification that would selectively allow enhanced cell attachment at the implant-host interface and reduced cell attachment over the interior surface of the implant. Here, we have developed a novel and simple peptide conjugation scheme to modify PMMA surfaces, which allowed for region-specific control of cell adhesion. This method uses di-amino-PEG, which can be grafted onto PMMA using hydrolysis or aminolysis method. PEG can resist cell adhesion and protein adsorption. The functionalization of grafted di-amino-PEG molecules with RGD peptide not only restored cell adhesion to the surfaces, but also enhanced cell attachment and spreading as compared to untreated PMMA surfaces. Long-term cell migration and micropatterning studies clearly indicated that PEG-PMMA surfaces with and without RGD conjugation can be used to differentiate cell adhesion and control cell attachment spatially on PMMA, which will have potential applications in the modification of keratoprostheses.     

Novel gradient casting method provides high-throughput assessment of blended polyester poly(lactic-co-glycolic acid) thin films for parameter optimization

Pure polymer films cannot meet the diverse range of controlled release and material properties demanded for the fabrication of medical implants or other devices. Additives are added to modulate and optimize thin films for the desired qualities. To characterize the property trends that depend on additive concentration, an assay was designed which involved casting a single polyester poly(lactic-co-glycolic acid) (PLGA) film that blends a linear gradient of any PLGA-soluble additive desired. Four gradient PLGA films were produced by blending polyethylene glycol or the more hydrophobic polypropylene glycol. The films were made using a custom glass gradient maker in conjunction with a 180 cm film applicator. These films were characterized in terms of thickness, percent additive, total polymer (PLGA+additive), and controlled drug release using drug-like fluorescent molecules such as coumarin 6 (COU) or fluorescein diacetate (FDAc). Material properties of elongation and modulus were also accessed. Linear gradients of additives were readily generated, with phase separation being the limiting factor. Additive concentration had a Pearson's correlation factor (R) of >0.93 with respect to the per cent total release after 30 days for all gradients characterized. Release of COU had a near zero-order release over the same time period, suggesting that coumarin analogs may be suitable for use in PLGA/polyethylene glycol or PLGA/polypropylene glycol matrices, with each having unique material properties while allowing tuneable drug release. The gradient casting method described has considerable potential in offering higher throughput for optimizing film or coating material properties for medical implants or other devices.     

Retinal pigment epithelium cell culture on thin biodegradable poly(DL-lactic-co-glycolic acid) films

Thin films of 50 : 50 and 75 : 25 poly(DL-lactic-co-glycolic acid) (PLGA) were manufactured with a controlled thickness of less than 10 μm. The effect of PLGA copolymer ratio on in vitro cell attachment, proliferation, morphology, and tight junction formation was evaluated using a human D407 retinal pigment epithelium (RPE) cell line. Almost complete cell attachment was achieved on both PLGA films after 8 h of cell seeding, which was comparable to that on tissue culture polystyrene (TCPS) controls. The initial cell seeding density affected attachment, and the optimal value for 50: 50 PLGA was 25 000 cells cm^-2. After 7 days of in vitro culture, cell density on 50 : 50 and 75 : 25 PLGA films increased 45 and 40 folds, respectively, and a 34-fold increase was observed on TCPS. The RPE cells cultured on PLGA films at confluence had a characteristic cobblestone morphology. Confluent RPE cells also developed normal tight junctions in vitro which were concentrated mainly at the apical surfaces of cell-cell junctions. These results demonstrated that thin biodegradable PLGA films can provide suitable substrates for human RPE cell culture, and may serve as temporary carriers for subretinal implantation of organized sheets of RPE.