骨髓基质干细胞与聚丙交酯/乙交酯/天冬氨酸/聚乙二醇体外复合培养的实验研究

目的探讨骨髓基质于细胞（marrow stromal stem cells，MSCs）与聚丙交酯／乙交酯／天冬氨酸／聚乙二醇[poly（lactic acid／glycolic acid／asparagic acid—co—polyethylene glycol），PLGA—ASP—PEG]的生物相容性，为构建组织工程骨进行骨缺损修复提供理论基础，以及为组织工程支架材料的优化提供实验依据。方法本体开环共聚法合成PLGA—ASP—PEG三嵌段共聚物。取4周龄新西兰大白兔骨髓，体外分离培养MSCs。将第3代MSCs以1．0×10^6／ml接种到PLGA—ASP—PEG材料上，测定细胞黏附力和黏附率；扫描电镜观察细胞的形态学特征；MTT法检测细胞增殖；流式细胞仪检测细胞周期、增殖指数、DNA指数及凋亡；通过考马斯亮蓝测定法和^3H-脯氨酸掺入实验观察细胞的蛋白合成和胶原合成情况。以PLGA支架材料作为对照。结果MSCs在PLGA—ASP—PEG支架材料上贴附、生长良好，其黏附、增殖和蛋白、胶原合成能力均显著高于PLGA组（P〈0.05），细胞凋亡率显著低于PLGA组（P〈0.05）。DNA指数显示两组细胞均为正常的二倍体细胞。结论PLGA—ASP—PEG的生物学性能与PLGA相比得到显著改善和提高，可作为MSCs的理想载体构建组织工程骨进行骨缺损修复研究。     

聚乳酸复合材料在整形外科的应用

随着组织工程技术的发展，生物高分子材料已经日益成为世界各国竞相研究的热门课题。由乳酸自行缩聚形成的高分子材料聚乳酸（poly lactic acid，PLA），不仅拥有优良的机械强度和化学稳定性，还具有良好的生物相容性，可吸收性和可降解性，是少数被美国食品和药物管理局（FDA）批准的生物降解性医用材料，     

浅谈丙交酯的研究进展

随着组织工程的发展和对环保生物材料需求的日益增加,人们不断开发出可降解的生物材料.聚乳酸(polylactic acid,PLA)及其共聚物就是其中应用最广泛的可降解的热塑性聚酯材料之一,它是以乳酸为单体采用化学方法合成的聚合物,是一种无毒、可生物降解吸收、强度高、易加工成型的合成类高分子材料.由于其良好的生物相容性和生物可降解性,已经在生物组织工程、农林渔业、食品包装以及服装等领域得到广泛的应用.     

生物活性PCL／PLA三维多孔块膜为载体的骺软骨本外实验研究

目的 探讨以生物活性聚己内酯／聚乳酸共聚物（PCL／PLA）为载体的兔骺软骨细胞体外培养的可行性。方法 应用具有三维多孔立体结构的生物活性支架材料（PCL／PLA）为细胞外基质替代物,体外进行兔骺软骨细胞培养。采用组织学、电镜、MTT比色法、免疫组化法等多种手段动态检测细胞生长情况。结果 兔骺软骨细胞生长良好,增殖活跃,合成大量软骨基质,并与材料形成细胞－－材料复合体,细胞充满于材料孔隙内。结论 生物活性可降解吸收材料PCL／PLA具有良好的组织相容性及表面活性,它所提供的三维立体空间结构有利于细胞生长繁殖。为探讨骨骺早闭的全新疗法奠定了基础。     

聚乳酸-骨基质明胶多孔复合材料制备及骨诱导活性研究

目的采用CO2超临界法制备一种新型的具有生物活性的聚乳酸（poly lactic acid，PLA）-骨基质明胶（bone matrix gelatin，BMG）多孔复合型骨支架材料，并检测骨诱导活性。方法选取健康成人供体皮质骨进行脱脂、脱钙和脱蛋白处理制备BMG，将按体积比3：1混合的BMG—PLA及纯PLA，分别置入超临界CO2反应装置中，同时加入粒径为100～200,am的NaCl颗粒作为制孔剂，反应制备多孔PLA—BMG和PLA复合材料。在含10％胎牛血清的DMEM高糖培养基中培养MC3T3-E1成骨前体细胞，按20μl／孔2×10^4／ml浓度的细胞悬液加入含不同材料的24孔培养板内共培养2周，其中PLA—BMG组：每孔含100μg粉碎的PLA—BMG复合材料；PLA组：每孔含100μg粉碎的PLA材料；DMEM高糖培养基组作为对照组；每组设6个复孔。通过茜素红S染色法染色钙化结节，定量分析钙化面积；收集共培养细胞并在1m10．2％的Nonidet P-40溶液中超声裂解，检测细胞内钙含量和ALP活性。结果超临界CO2法所制备PLA—BMG多孔复合材料中BMG与PLA混合均匀，孔隙大小为50～150μm，孔径联通性好；PLA—BMG组ALP活性、钙含量及钙化面积均值分别为325．59±70．40U／gprot、3．51±1．64mmol／gprot和42．98％±4．44％，均高于PLA组63．62±30．01U／gprot、1．04±0．21mmol／gprot和9．55％±1．94％，及对照组2．40±1．47U／gprot、0．70±0．24mmol／gprot和0．86％±0．41％，比较差异均有统计学意义（P〈0．05），且PLA组的ALP活性及钙化结节面积与对照组也有统计学意义（P〈0．05）。结论采用CO2超临界法制备的PLA—BMG多孔复合支架材料具有良好的骨诱导活性，有可能作为一种有前景的骨植入材料及骨组织工程支架材料。     

Lp-PLA2和心血管疾病

脂蛋白相关磷脂酶A2（1ipoproteinassociated phospholipase A2，Lp—PLA2）义称为血小板活化因子乙酰水化酶、ⅦA型磷脂酶A2。过去因为LP—PLA2能够水解灭火血小板活化因子（Dlatelet—activating factor,PAF）和低密度脂蛋白（LDL）中的氧化磷脂．曾经被认为具有抑制炎症．甚至抑制动脉粥样硬化形成作用。     

生物可降解材料聚乳酸/聚羟基乙酸复合壳聚糖在人工冠状动脉血管支架制备中的应用

第一代金属裸支架和第二代涂层支架介入治疗冠状动脉粥样硬化性心脏病(冠心病)已得到广泛应用。由于长期存在金属支架异物刺激及其携带的药物扰乱血管壁各层细胞生长,引起支架内再狭窄和血管栓塞,远期仍有较多的主要心血管不良事件发生和需要再血管化治疗。因此,由聚酯、聚碳酸酐及聚磷酸酯等高分子材料制备的完全可生物降解吸收支架及药物洗脱支架应运而生,其中聚乳酸(poly-lactic acid,PLA)、聚羟基乙酸(poly-glycolicacid,PGA)、壳聚糖、聚己内酯(poly-caprolactone,PCL)及一些共聚物如聚乳酸/聚羟基乙酸共聚物(poly-lactic-co-glycolic acid,PLGA)材料制备的心血管植入支架的安全性、组织及血液相容性已得到证实,然而这些支架具有各自的缺点,如PLA降解较慢质硬易断裂柔韧性不足,PGA降解较快质软支撑力不足,支架降解太快或者太慢,均难以达到有效支撑,支架植入后容易出现血管损伤、弹性回缩,导致血管再狭窄及血栓形成,远期效果不佳。通过优化组合不同摩尔比的PLA和PGA及壳聚糖涂层,可以获得具有更好的生物相容性、适度的降解速率(约3～6个月完全降解)、足够的机械强度、较低的炎症反应和伸展度良好的复合材料,从而为制备完全生物可降解冠状动脉支架奠定实验基础。     

脂蛋白相关磷脂酶A2活性与脑供血动脉粥样硬化关系探讨

目的探讨炎性反应标志物脂蛋白相关磷脂酶A2（Lp—PLA2）活性与脑供血动脉粥样硬化的关系。方法对148例脑供血动脉粥样硬化患者（脑供血动脉粥样硬化组）及40例健康体检者（对照组）的Lp-PLA2活性及传统炎性反应标志物和危险因素进行检测,并进行比较分析。结果脑供血动脉粥样硬化组Lp．PLA2活性显著高于对照组[（29．84±3．50）nmol／（min·ml）比（19．79±2．25）nmol／（min·ml）,P〈0．01）]。脑供血动脉粥样硬化患者血管狭窄程度与Lp．PIA2活性呈正相关（r=0．491,p〈0．01）;血管病变支数与Lp．PLA2活性呈正相关（r=0．249,P=0．026）。Logistic回归分析显示,Lp—PLA2活性升高是除高敏C反应蛋白、白细胞计数等传统非特异性炎性反应标志物以外脑供血动脉粥样硬化的独立危险因素。结论Lp—PLA2活性升高是脑供血动脉粥样硬化的独立危险因素。     

慢性胰腺炎患者磷酯酶A_2族水平变化及其意义

磷酯酶A2（PLA2）异构体中PLA2－Ⅰ和PLA2－Ⅱ在急性胰腺炎和慢性胰腺炎（CP）急性发作时水平升高，动物实验也得到证实。据此作者推测PLA2族在CP中也可能升高。按此设想作者用免疫级化、Northenblot和原位杂交技术对15例CPPLA2族中PLA2－Ⅰ、PLA2－Ⅱ和PLA2－Ⅳ进行了测定，并以16例健康者为对照。结果发现，正常胰腺组织中PLA2－Ⅰ和弹力蛋白酶mRNA水平明显高于PLA2－Ⅱ和PLA2－Ⅳ（后两者不能测出）。相反，15例CP患者中11例PLA2－Ⅰ和弹力蛋白酶mRNA水平低于正常，而PLA2－Ⅱ和PLA2－ⅣmRNA水平反而增高，其中…     

不同比例的PLA—PGA支架对软骨细胞复合的影响

目的研究经聚乳酸（PLA）包埋后的不同比例PLA-聚羟基乙酸（PGA）组织工程支架对软骨细胞复合的影响及原因。方法分离培养小猪软骨细胞，并以5×10^7cells／mL的密度接种在经PLA包埋的不同比例PGA—PLA支架材料上，4h后检测软骨细胞在支架材料上的流失率。24h后检测软骨细胞在支架材料上的粘附率，并对支架材料进行电镜扫描，分析原因。结果随着支架材料中PLA比例的提高，软骨细胞的接种流失率上升，支架材料的接种负荷下降，24h时软骨细胞粘附率无明显差异。扫描电镜证实其原因。结论PLA包埋的PGA支架材料会降低对软骨细胞的接种负荷，但细胞粘附率无变化。     

卵磷脂、多聚赖氨酸和PLA包埋PGA与软骨细胞体外培养的实验研究

组织工程技术中细胞培养支架的选择是研究的焦点之一。以ＰＬＡ包埋的ＰＧＡ无纺网是应用较为广泛的支架之一。但其亲水性差，对细胞吸附力弱，是其不足之处。此实验选择了卵磷脂和多聚赖氨酸来分别和共同包埋ＰＧＡ＋ＰＬＡ，来观察其亲水性和对细胞吸附力的改变，及对细胞功能的影响。本实验证明卵磷脂具有增强支架亲水性作用；而多聚赖氨酸除具有增加支架对细胞的吸附力外，还具有促进细胞功能的作用。以卵磷脂和多聚赖氨酸共同包埋支架使细胞均匀地分布于支架纤维之间和支架表面，充分利用了支架空间，从而更好地发挥细胞的功能。所以本实验将为组织工程技术中细胞培养支架的选择提供一条新的途径。     

骨髓基质细胞与生物活性玻璃陶瓷和聚乳酸生物相容性的实验研究

目的 探讨骨髓基质细胞与生物活性玻璃陶瓷（ＢＧＣ）和聚乳酸（ＰＬＡ）的生物相容性,为骨组织工程中生物材料的选择提供依据。方法 将骨髓基质细胞与ＢＧＣ、ＰＬＡ复合体培养,进行形态学观察、细胞增殖、蛋白质含量及酶学测定。结果 骨髓基质细胞能在ＢＧＣ、ＰＬＡ上贴附、繁殖,其生长及功能不受影响,并且ＢＧＣ具有一定的促细胞增殖作用。结论 ＢＧＣ、ＰＬＡ具有良好的细胞相容性,有可能作为骨髓基质细胞的载体应用于     

明胶杂化三维聚乳酸支架研究

目的 克服聚乳酸材料的疏水性,制备表面特性和孔隙结构均符合组织工程需要的细胞支架.方法 应用改进的溶液浇注/粒子沥滤技术,制备孔隙完全连通的三维聚乳酸细胞支架,碱液预处理支架表面,浸润明胶溶液,戊二醛蒸汽交联支架上的明胶,获得杂化改性支架;用扫描电镜等对改性后的支架进行结构与性能表征.结果 圆柱形多孔聚乳酸支架,在杂化改性后吸水率增加约2倍,由改性前的(1164.2±172.9)%上升到(2637.7±527.8)%,亲水性显著增强;杂化改性后,明胶均匀分布在支架孔隙表面,支架孔隙形态、连通性基本没有变化.结论 明胶杂化能显著提高聚乳酸细胞支架的亲水性,改性过程不影响支架的孔隙结构和形态,可以制备出亲水性好、内部结构可控、孔隙率高、孔隙连通性好的组织工程细胞支架.     

骨髓基质干细胞复合纳米羟基磷灰石/聚乳酸构建组织工程骨

目的：了解骨髓基质干细胞（MSCs）复合纳米羟基磷灰石／聚乳酸（n-HA／PLA）构建组织工程骨的异位成骨作用。方法：选择6只新西兰白兔，实验组于动物脊柱左侧肌肉内植入MSCs复合n-HA／PLA构建的组织工程骨，对照组于右侧植入n-HA／PLA生物材料。术后4、8周取材，行溴脱氧尿嘧啶（Brdu）标记细胞检测和组织学观察。结果：术后4周，实验组材料边缘均可检测到Brdu标记的MSCs。术后4、8周，实验组材料内部有新骨形成，随着时间推移，新骨量增多，编织骨向板层骨过渡。对照组材料未见新骨形成。结论：MSCs复合n-HA／PLA构建的组织工程骨具有很好的异位成骨作用。     

包埋后的几丁质与软骨细胞体外培养的实验研究

目的 探讨几丁质作为组织工程技术中细胞培养支架的可行性。方法 采用聚乳酸、卵磷脂及多聚赖氨酸分别或工同包埋几丁质与软骨细胞体外培养,观察其产亲水性的改变、对细胞吸附力和细胞功能的影响。结果 以聚乳酸包埋的几丁质对细胞的生长有抵制作用;以卵磷脂包埋的几柄质亲水性增强;以多聚赖氨酸包埋的几丁质对细胞吸附力增强;以卵磷脂和多聚赖氨酸共同包埋的几丁质具有良好的亲水性和对细胞吸附力,并可使细胞更好地发挥功能     

Human fetal bone cells associated with ceramic reinforced PLA scaffolds for tissue engineering

Fetal bone cells were shown to have an interesting potential for therapeutic use in bone tissue engineering due to their rapid growth rate and their ability to differentiate into mature osteoblasts in vitro. We describe hereafter their capability to promote bone repair in vivo when combined with porous scaffolds based on poly(l-lactic acid) (PLA) obtained by supercritical gas foaming and reinforced with 5 wt.% beta-tricalcium phosphate (TCP). Bone regeneration was assessed by radiography and histology after implantation of PLA/TCP scaffolds alone, seeded with primary fetal bone cells, or coated with demineralized bone matrix. Craniotomy critical size defects and drill defects in the femoral condyle in rats were employed. In the cranial defects, polymer degradation and cortical bone regeneration were studied up to 12 months postoperatively. Complete bone ingrowth was observed after implantation of PLA/TCP constructs seeded with human fetal bone cells. Further tests were conducted in the trabecular neighborhood of femoral condyles, where scaffolds seeded with fetal bone cells also promoted bone repair. We present here a promising approach for bone tissue engineering using human primary fetal bone cells in combination with porous PLA/TCP structures. Fetal bone cells could be selected regarding osteogenic and immune-related properties, along with their rapid growth, ease of cell banking and associated safety.     

个性化外鼻形态组织工程支架材料的制作

目的探索应用计算机辅助设计与快速成型技术制作个性化精确外鼻形状组织工程支架材料的可行性。方法通过计算机辅助设计技术,对患者头颅CT数据进行三维重建,并对外鼻进行分割,得到外鼻的三维重建模型,并建立外鼻模型数据库。利用快速成型技术,打印出两块与外鼻模型对应的树脂阴模。将非编织聚羟基乙酸(PGA)置入树脂阴模内压制成型,再通过聚乳酸(PLA)包埋,提高支架的强度,最终制作出个性化外鼻形态PGA-PLA支架。结果该外鼻形状支架具有精确的三维立体结构,符合组织工程化外鼻支架材料制作的形态需求。结论应用计算机辅助设计与快速成型技术,能精确制作个性化组织工程外鼻形状支架材料。     

Development of in vivo muCT evaluation of neovascularisation in tissue engineered bone constructs

Due to an increasing aging population the need for innovative approaches to aid skeletal repair and reconstruction is a significant socio-economic increasing problem. The emerging discipline of tissue engineering has sort to augment the growth and repair of bone loss particularly in areas of trauma, degeneration and revision surgery. However, the initiation and development of a fully functional vascular network are critical for bioengineered bone to repair large osseous defects, whether the material is osteosynthetic (poly (d,l)-lactic acid, PLA) or natural bone allograft. Quantification and three-dimensional visualization of new vessel networks remain a problem in bone tissue engineering constructs. A novel technique utilising a radio-opaque dye and micro-computed tomography (muCT) has been developed and applied to study angiogenesis in an impaction bone graft model. Tissue-engineered constructs combining human bone marrow stromal cells (HBMSC) with natural allograft and synthetic grafts (PLA) were impacted and implanted into the subcutis of MF-1 nu/nu mice for a period of 28 days. Microfil consisting of radio-opaque polymer was perfused through the mice and scanned using a Bench Top CT system for micro-computed tomography. Analysis of three-dimensional muCT reconstructions demonstrated an increase in vessel volume and vessel number in the impacted scaffolds/HBMC compared to scaffolds alone. Vessel volume: allograft/HBMSC=0.57 mm(3)+/-0.19; allograft=0.04 mm(3)+/-0.04; PLA/HBMSC=1.19 mm(3)+/-0.31; and PLA=0.12 mm(3)+/-0.01. Penetrating vessel number: allograft/HBMSC=22.33+/-3.21; allograft=3.67+/-1.153; PLA/HBMSC=32.67+/-8.33; and PLA=7.67+/-3.06. Type 1 collagen and von Willebrand factor immunohistochemistry in scaffold/HBMSC constructs indicated the osteogenic cell phenotype, and new blood vessel formation respectively. Contrast-enhanced 3D reconstructions facilitated the visualization and quantification of neovascularisation. This novel technique has been used to demonstrate neovascularisation in impacted tissue engineered constructs providing a facile approach with wide experimental application.     

Orthopaedic applications for PLA-PGA biodegradable polymers

Biodegradable polymers, especially those belonging to the family of polylactic acid (PLA) and polyglycolic acid (PGA), play an increasingly important role in orthopaedics. These polymers degrade by hydrolysis and enzymatic activity and have a range of mechanical and physical properties that can be engineered appropriately to suit a particular application. Their degradation characteristics depend on several parameters including their molecular structure, crystallinity, and copolymer ratio. These biomaterials are also rapidly gaining recognition in the fledging field of tissue engineering because they can be fashioned into porous scaffolds or carriers of cells, extracellular matrix components, and bioactive agents. Although their future appears to be bright, several questions regarding the biocompatibility of these materials linger and should be addressed before their wide-scale use. In the context of musculoskeletal tissue, this report provides a comprehensive review of properties and applications of biodegradable PLA/PGA polymers and their copolymers. Of special interest are orthopaedic applications, biocompatibility studies, and issues of sterilization and storage of these versatile biomaterials. Also discussed is the fact that terms such as PLA, PGA, or PLA-PGA do not denote one material, but rather a large family of materials that have a wide range of differing bioengineering properties and concomitant biological responses. An analysis of some misconceptions, problems, and potential solutions is also provided.Arthroscopy 1998 Oct;14(7):726-37     

Experimental study on the adhesion, migration and three-dimensional growth of Schwann cells on absorbable biological materials

Objective: To study the adhesion, migration and three-dimentional growth of Schwann cells on PLA(polylactic acid) nonspinning fibre cloth and polyglycolic/polylactic acid (PLGA) fibres. Methods: Schwann cells/ECM gel solution and PLA nonspinning fibre cloth and PLGA fibres pretreated by collagen, polylysine and ECM were co-cultured. Then the migration and three-dimensional growth of Schwann cells on the fibres were observed under phase contrast microscope and laser scanning confocal microscope. Results : Schwann cell/ECM solution was compounded with PLA nonspinning fibre cloth. With formation of gel, most Schwann cells resided in the fibre net holes, and adhered to the fibres to form a multiplayerarranged Schwann cell column like Biingner band.Schwann cells could adhere to PLGA fibres and grew and migrated along the fibres. ECM gel could significantly increase the adhering and migrating cell number. Conclusions: ECM gel can facilitate the adhesion,growth and migration of Schwann cells on the seteroframe.It is a good integrating material for tissue engineering bioartificial nerve.