含悬挂羧基手臂聚乳酸材料的合成与血小板粘附性研究

制备了乳酸-β-苹果酸共聚物,并在此基础上进一步修饰合成了含悬挂羟基(PLMAHE)以及悬挂羧基(PCA-PLA)的聚乳酸共聚物,利用原子力显微镜及环境扫描电镜,观察了聚合物膜的表面形貌以及粘附在聚合物膜上的血小板数量与形态.结果表明含悬挂羟基材料表面粘附血小板时发生聚集并有伪足生成,含悬挂羧基材料表面血小板粘附数量较少且形态正常,有望成为优良的抗凝血材料.     

新型含悬挂羧基聚乳酸的制备及其血液相容性和细胞粘附性

制备了乳酸-苹果酸共聚物(PLMA), 并在前期工作的基础上制备了悬挂羧基聚乳酸(PLMACA), 考察了手臂长度及端功能基团对改善聚乳酸的血液相容性及细胞粘附性的影响. 结果表明, PLMACA同时具有良好的血液相容性和细胞亲和性, 极有可能成为新一代血管(修复)材料.     

REDV/雷帕霉素复合涂层构建内皮细胞选择性功能界面

运用复合涂层的概念构建了兼具药物洗脱和内皮促进作用的载药涂层. 以载雷帕霉素(Rapamycin, RAP)的聚乙二醇甲基丙烯酸酯(PEGMA)-甲基丙烯酸丁酯(BMA)(PEGMA-BMA, PEGB)为内层, Arg-Glu-Asp-Val(REDV)多肽修饰的PEGBN为外层包裹载药涂层. 体外药物释放结果表明, 雷帕霉素可以维持缓慢稳定的长效释放, 释放过程中没有出现暴释现象. 表面细胞生长行为表明, 雷帕霉素可以有效地阻抗内皮细胞和平滑肌细胞的黏附, 抑制细胞活性; 随着药物释放的进行, 雷帕霉素浓度逐渐减低, 但涂层依然维持对平滑肌细胞的非特异性阻抗; 而REDV修饰的外涂层开始呈现内皮细胞的选择性黏附, 随着释放时间延长, 内皮细胞特异选择性也逐渐加强. 雷帕霉素和REDV多肽协同构建的复合涂层能够有效抑制平滑肌细胞的增殖, 获得内皮细胞选择性黏附.     

人工细胞外基质对血管内皮细胞生存的影响

通过物理吸附方法, 利用胶原、 聚赖氨酸和融合蛋白VEGF-Fc对聚苯乙烯培养板表面进行改性, 以研究细胞外基质材料对血管内皮细胞的影响. 结果表明, 3种蛋白显著提高了聚苯乙烯表面的亲水性. 内皮细胞的黏附、 增殖、 细胞骨架蛋白染色和血管性血友病因子(vWF)免疫染色实验结果表明, 胶原、 聚赖氨酸和VEGF-Fc基质均能有效提高血管内皮细胞的黏附, 其中胶原可与VEGF协同作用促进内皮细胞分化表型的表达; VEGF-Fc基质兼具了VEGF的生物学活性, 可促进内皮细胞的黏附和增殖以及vWF功能性蛋白的表达. 本研究为诱导材料表面内皮化和血管新生的生物活性材料的设计开发提供了新思路.     

基于聚乙二醇和不同多肽构建的生物涂层及其内皮细胞选择性研究

利用自由基聚合反应将甲基丙烯酸聚乙二醇酯(PEGMA)和甲基丙烯酸缩水甘油酯(GMA)的二元共聚物接枝在基材表面,并通过开环反应分别固定精氨酸-甘氨酸-天冬氨酸(RGD)、精氨酸-谷氨酸-天冬氨酸-缬氨酸(REDV)和酪氨酸-异亮氨酸-甘氨酸-丝氨酸-精氨酸(YIGSR)3种可特异性黏附内皮细胞的多肽.通过核磁共振检测合成的聚合物分子结构,并进一步通过X射线光电子能谱(XPS))以及原子力显微镜(AFM)的测试结果证明聚合物成功接枝在基材表面.利用紫外-可见吸收光谱(UV-Vis)对表面固定的3种多肽进行了定量表征.体外内皮细胞和平滑肌细胞黏附结果表明,3种不同多肽修饰的共聚物表面均能够有效阻抗平滑肌细胞的黏附,同时不同程度地促进内皮细胞的黏附,从而实现了基材表面内皮细胞的选择性黏附.其中与RGD和YIGSR多肽修饰的表面相比,REDV多肽修饰的表面呈现出更优异的内皮细胞选择性.这种具有内皮细胞特异选择性的界面在心血管支架涂层原位内皮化方面具有良好的应用前景.     

基于聚多巴胺辅助自组装单分子层技术的PTFE表面修饰及其内皮细胞选择性黏附研究

利用聚多巴胺技术对PTFE进行表面改性,X射线光电子能谱(XPS)、椭偏、接触角以及石英晶体微天平(QCM-D)证实DOPA分子可以在PTFE表面自聚形成反应性的超薄膜功能涂层,并通过聚多巴胺辅助自组装单分子层(SAM)技术构建了活性多肽链段CGREDVDY的界面.细胞黏附实验反映活性链段CGREDVDY的修饰表面具备良好的内皮细胞选择性黏附能力.这种具有内皮细胞选择性黏附能力的界面有望实现材料在复杂生理环境中对内皮细胞的原位诱导,为制备具有血管内皮原位快速愈合功能的新型血液相容性人造血管提供新途径.     

表面接枝嵌段共聚物刷实现内皮细胞的选择性黏附

利用表面引发原子转移自由基聚合(SI-ATRP)技术将聚(甲基丙烯酸寡聚乙二醇酯)和聚(甲基丙烯酸缩水甘油酯)的双嵌段共聚物刷(POEGMA-b-PGMA)接枝在材料表面,并通过PGMA中丰富的环氧基团开环固定可特异性黏附内皮细胞的多肽GREDVY.静态水接触角、接枝层厚度、X射线光电子能谱(XPS)以及原子力显微镜(AFM)的测试结果证明了各步接枝反应的成功性.细胞培养研究表明这种连接有GREDVY的双嵌段共聚物表面能够有效的促进血管内皮细胞的黏附,同时排斥成纤维细胞的黏附,从而实现了内皮细胞的选择性黏附.     

1,3-PBO扩链改性端羧基聚乳酸的性能表征

以乳酸为原料、辛酸亚锡为催化剂,采用梯度升温法,在170℃、0.098 MPa条件下直接熔融缩聚合成端羧基共聚物P(LA/SA).将其用2,2-(1,3-亚苯基)-二噁唑啉(1,3-PBO)扩链,按n(—COOH)/n(—oxazoline)=1∶1.4比例加入1,3-PBO,在150℃,0.098 MPa条件下反应15 min制得聚酰胺酯(PEA).采用GPC、FTIR、1H-NMR、DSC、XRD、TGA、SEM等手段对聚合物的结构进行了表征和性能测试.结果表明,与P(LA/SA)相比,扩链产物相对分子质量大幅度提高,重均分子量达36×104;产物Tg比PLA升高,材料的刚性增强;产物热稳定性能提高,为一步分解;产物结晶度较P(LA/SA)降低,其柔韧性较P(LA/SA)增强,但相对于PLA有所降低.     

肾上腺髓质素微球复合PLGA/纳米轻基磷灰石支架材料的制备及生物活性评价

利用离子乳化交联法制备了负载肾上腺髓质素的壳聚糖微球,应用热致相分离法制备了乳酸和乙醇酸共聚物/纳米羟基磷灰石(PLGA/nHA)支架材料并在其中包覆载药微球.通过扫描电子显微镜、体外释放行为、材料溶血行为、碱性磷酸酶(ALP)活性的测定、支架材料表面细胞荧光染色和MTT[3-(4,5-二甲基噻唑-2)-2,5-二苯基...     

不同氨基酸直接改性聚乳酸的性能研究

分别以甘氨酸(Gly)和L-天冬氨酸(Asp)与L-乳酸(L-LA)直接熔融共聚制备改性聚乳酸(PLA),所得两种共聚物聚(乳酸-甘氨酸)[P(LA-co-Gly)]和聚(乳酸-天冬氨酸)[P(LA-co-Asp)]进行了表征。对P(LA-co-Gly),不同投料比时共聚物重均分子量(Mw)和分散度(Mw/Mn)随Gly投料量的增加而变小。与双官能团Gly直接改性的PLA相比,多官能团Asp直接改性的PLA具有一定的支链结构,分散度(Mw/Mn)二者接近或P(LA-co-Asp)的更高,但两种共聚物的Tg均比PLA的要大,亲水性有所提高。同时,所得共聚物均呈无定形态,且Mw都在2400到5600之间,能满足药物缓释对聚乳酸类高分子材料分子量的要求。     

Preparation of orthogonally functionalized surface using micromolding in capillaries technique for the control of cellular adhesion

This study presents a simple method for the fabrication of an orthogonal surface that can be applied for cell patterning without the need to immobilize specific adhesive peptides, proteins, or extracellular matrix (ECM) for cell attachment. Micromolding in capillaries (MIMIC) produced two distinctive regions. One region contained poly(ethylene glycol)–poly(d,l-lactide) diblock copolymer (PEG–PLA) designed to provide a biological barrier to the nonspecific binding of proteins and fibroblast cells. The other region was coated with polyelectrolyte (PEL) to promote the adhesion of biomolecules including proteins and cells. Resistance to the adsorption of proteins increased with the length of PEG and PLA chains because the longer PEG chain increased the PEG layer thickness and the longer PLA chain induced stronger interaction with the PEL surface. The PEG5k–PLA2.5k (20 mg/ml) was the most efficient candidate for the prevention of protein adhesion among the PEG–PLA copolymers examined. The orthogonal functionality of prepared surfaces having PEL regions and background PEG–PLA regions resulted in rapid patterning of biomolecules. Fluorescein isothiocyanate-tagged bovine serum albumin (FITC-BSA) and fibroblast cells successfully adhered to the exposed PEL surfaces. Although methods for cell patterning generally require an adhesive protein layer on the desired area, these fabricated surfaces without adhesive proteins provide a gentle microenvironment for cells. In addition, our proposed approach could easily control patterns, sizes, and shapes at micron scale.     

Synthesis and characterization of nitric oxide-releasing sol-gel microarrays

Diazeniumdiolate-modified sol-gel microarrays capable of releasing low levels of nitric oxide are reported as a viable means for improving the blood compatibility of a surface without fully modifying the underlying substrate. Several parameters are characterized including: (1) NO surface flux as a function of sol-gel composition and microarray geometry; (2) microstructure dimensions and spacing for optimal blood compatibility; and (3) the effect of sol-gel surface modification on analyte accessibility to platinum electrodes. The sol-gel microarrays release biologically relevant levels of NO under physiological conditions for >24 h. In vitro platelet adhesion assays indicate that a NO surface flux of 2.2 pmol cm(-2) s(-1) effectively reduces platelet adhesion to glass substrates modified with sol-gel microstructures separated by 50 microm. The blood compatibility observed for these micropatterned surfaces is comparable to NO-releasing sol-gel films. When the separation between NO-releasing microstructures is reduced to 10 microm, the NO surface flux required to reduce platelet adhesion is lowered to 0.4 pmol cm(-2) s(-1). Finally, the oxygen response of platinum electrodes modified with NO-releasing sol-gel microarrays indicates that selective modification via micropatterning enhances analyte accessibility to the sensor surface.     

Hybrid Pericardium with VEGF‐Loaded Hyaluronic Acid Hydrogel Coating to Improve the Biological Properties of Bioprosthetic Heart Valves

Bioprosthetic heart valves (BHVs) used in the clinic are mostly fixed by glutaraldehyde and the lack of endothelialization is a major problem for glutaraldehyde‐fixed pericardia. Hyaluronic acid is a major glycosaminoglycan that exists in native heart valves. Coupled with its inherent biocompatibility, it may enhance endothelial adhesion and proliferation when associated with vascular endothelial growth factor (VEGF). In this study, an optimized system is developed to improve the endothelialization of glutaraldehyde‐fixed pericardium. A hybrid pericardium with VEGF‐loaded hyaluronic acid hydrogel coating is developed by the crosslinking of 1,4‐butanediol diglycidyl ether. The adhesion and growth potential of human umbilical vein endothelial cells (HUVECs) on pericardia, platelet adhesion, and calcification by an in vivo rat subdermal implantation model are investigated. The results show improved HUVEC adhesion and proliferation, less platelet adhesion, and less calcification for hybrid pericardium by introducing the coating of VEGF‐loaded hyaluronic acid hydrogel. Thus, the coating of VEGF‐loaded hyaluronic acid hydrogel on pericardium is a promising approach to obtain bioprosthetic valves for clinical applications with increased endothelialization and antithrombotic and anticalcification properties.     

In vitro model on glass surfaces for complex interactions between different types of cells

This report establishes an in vitro model on glass surfaces for patterning multiple types of cells to simulate cell-cell interactions in vivo. The model employs a microfluidic system and poly(ethylene glycol)-terminated oxysilane (PEG-oxysilane) to modify glass surfaces in order to resist cell adhesion. The system allows the selective confinement of different types of cells to realize complete confinement, partial confinement, and no confinement of three types of cells on glass surfaces. The model was applied to study intercellular interactions among human umbilical vein endothelial cells (HUVEC), PLA 801 C and PLA801 D cells.     

Genistein activates endothelial nitric oxide synthase in broiler pulmonary arterial endothelial cells by an Akt-dependent mechanism

Deregulation of endothelial nitric oxide synthase (eNOS) plays an important role in the development of multiple cardiovascular diseases. Our recent study demonstrated that genistein supplementation attenuates pulmonary arterial hypertension in broilers by restoration of endothelial function. In this study, we investigated the molecular mechanism by using broiler pulmonary arterial endothelial cells (PAECs). Our results showed that genistein stimulated a rapid phosphorylation of eNOS at Ser(1179) which was associated with activation of eNOS/NO axis. Further study indicated that the activation of eNOS was not mediated through estrogen receptors or tyrosine kinase inhibition, but via a phosphatidylinositol 3-kinase (PI3K)/Akt-dependent signaling pathway, as the eNOS activity and related NO release were largely abolished by pharmacological inhibitors of PI3K or Akt. Thus, our findings revealed a critical function of Akt in mediating genistein-stimulated eNOS activity in PAECs, partially accounting for the beneficial effects of genistein on the development of cardiovascular diseases observed in animal models.     

Platelet and bacterial repellence on sulfonated poly(ethylene glycol)-acrylate copolymer surfaces

Novel poly(ethylene glycol) (PEG) and sulfonated PEG (PEG-SO₃) acrylate copolymers have been prepared and characterized to apply as coating and blending materials for biomedical applications. The modified surfaces using acrylate copolymers demonstrated increased hydrophilicity, possibly due to the hypothesized reorientation of PEG/PEG-SO₃ chains into water phase. All copolymer surfaces demonstrated less platelet adhesion than control. In addition, platelet adhesion on copolymer surfaces decreased as the chain length of PEG and sulfonated PEG in copolymers increases. All copolymer surfaces reduced bacterial adhesion significantly and the adhesion level differs depending on surfaces as well as media. The obtained results attest to the usefulness of these copolymers as a coating or additive material to improve the blood compatibility of blood contacting devices.     

Prostaglandin E2 stimulates angiogenesis by activating the nitric oxide/cGMP pathway in human umbilical vein endothelial cells

Prostaglandin E2 (PGE2), a major product of cyclooxygenase, has been implicated in modulating angiogenesis, vascular function, and inflammatory processes, but the underlying mechanism is not clearly elucidated. We here investigated the molecular mechanism by which PGE2 regulates angiogenesis. Treatment of human umbilical vein endothelial cells (HUVEC) with PGE2 increased angiogenesis. PGE2 increased phosphorylation of Akt and endothelial nitric oxide synthase (eNOS), eNOS activity, and nitric oxide (NO) production by the activation of cAMP-dependent protein kinase (PKA) and phosphatidylinositol 3-kinase (PI3K). Dibutyryl cAMP (DB-cAMP) mimicked the role of PGE2 in angiogenesis and the signaling pathway, suggesting that cAMP is a down-stream mediator of PGE2. Furthermore, PGE2 increased endothelial cell sprouting from normal murine aortic segments, but not from eNOS-deficient ones, on Matrigel. The angiogenic effects of PGE2 were inhibited by the inhibitors of PKA, PI3K, eNOS, and soluble guanylate cyclase, but not by phospholipase C inhibitor. These results clearly show that PGE2 increased angiogenesis by activating the NO/cGMP signaling pathway through PKA/PI3K/Akt-dependent increase in eNOS activity.     

表面微图案化聚乳酸膜的制备及其细胞亲和性评价

利用溶剂-非溶剂法(SNS)制备表面具有微孔图案的聚乳酸(PLA)膜和聚苯乙烯(PS)膜,并以微孔PS膜为模板,构建表面具有微岛图案的PLA膜.以此为基础,对所制备的微图案表面对PLA膜亲/疏水性及成骨细胞粘附与增殖性能的影响进行研究.结果显示微图案的存在显著增强了PLA膜的表面疏水性(水接触角90°);成骨细胞在微图案表面具有良好的铺展性,其黏附数量明显高于光滑PLA膜,但细胞的生长曲线相对较平缓,显示微图案表面虽有利于细胞在PLA膜表面的粘附与铺展,但对促进细胞的增殖无贡献.