一般问题

20010694软骨细胞三维支架短期固定培养实验研究/吴海涛…//耳鼻咽喉一头颈外科一200〔},7(5)一298~301 目的:观察免肋软骨细胞聚怨基乙酸(PG八)三维支架短期固定培养对软骨细胞利用率、软骨细胞长满支架所需时间及软骨细胞支架复合体厚度等影响。方法:设PGA支架固定组和非固定组.固定组:先用鼠尾胶把PGA支架粘附在盖玻片[,再往固定后的PGA支架上滴加软骨细胞悬液.3~4天后支架与盖玻片分离。非固定组:把软骨细胞悬液直接滴加到P(子A支架上进行复合培养。固定组和非固定组软骨细胞PGA支架复合体培养2周后分别行同种异体皮下移植,3月后…     

软骨细胞三维支架短期固定培养实验研究

目的：观察兔肋软骨细胞聚羟基乙酸（ｐｏｌｙｇｌｙｃｏｌｉｃ ａｃｉｄ，ＰＧＡ）三维支架短期固定培养对软骨细胞利用率、软骨细胞长满支架所需时间及软骨细胞支架复合体厚度等影响。方法：设ＰＧＡ支架固定组和非固定组，固定组：先用鼠尾胶把ＰＧＡ支架粘附在盖玻片上，再往固定后的ＰＧＡ支架上滴加软骨细胞悬液，３￣４天后支架与盖玻片分离。非固定组：把软髓细胞悬液直接滴加到ＰＧＡ支架上进行复合培养。固定组和非固定组软骨细胞ＰＧＡ支架复合体培养２周后分别行同种异体皮下移植，３月后取出标本，光镜观察软内形成情况。结果：固定组中ＰＧＡ支架在加培养液过程中，不会在培养液内漂动，支架内软骨细胞无大量流失现象；相反，非固定组中ＰＧＡ支架在加培养液和移动培养皿过程中，会在培养液内飘动或晃动，支架内很多软骨细胞重新漂浮至培养液中。固定组ＰＧＡ支     

同种异体组织工程软骨修复兔气管软骨缺损实验研究

已有采用组织工程软骨修复裸鼠气管软骨缺损及同种异体组织工程软骨修复甲状软骨缺损等相关研究报道[1,2 ] ,在此基础上 ,我们采用同种异体组织工程软骨修复兔气管软骨缺损。一、材料和方法1 采用短期固定培养法构建兔软骨细胞 聚羟基乙酸(ｐｏｌｙｇｌｙｃｏｌｉｃａｃｉｄ ,ＰＧＡ)复合体[3 ] (图 1,2 ) ,复合培养 3周后行同种异体移植。2 软骨细胞 ＰＧＡ复合体同种异体皮下移植 :0 5～ 1ｋｇ新西兰兔 12只 ,每只兔背部皮下移植 3ｍｍ× 3ｍｍ的软骨细胞 ＰＧＡ复合体各 2块 ,移植术后 1、2、3和 4个月分批取出标本 ,每批 3只 (6块标…     

同种异体软骨细胞修复耳廓缺损的动物实验研究

目的探讨兔同种异体软骨细胞和自行制备生物材料高孔隙率聚乳酸(poly—DL-lactide,PDLLA)支架复合物修复兔耳廓软骨缺损的可行性。方法 18只大白兔分为软骨细胞/PDLLA复合物移植组、单纯PDLLA对照组和空白对照组,将体外培养的兔同种异体软骨细胞和自行制备的PDLLA支架形成复合物,修复兔耳廓软骨缺损。分别于植入后6周、12周、18周取材,HE和甲苯胺蓝染色了解兔耳廓缺损修复情况。结果术后18周,软骨细胞/PDLLA复合物移植组软骨缺损区为软骨组织修复,新生软骨与正常软骨问连接好,单纯PDLLA对照组和空白对照组为纤维软骨或纤维组织修复。结论同种异体软骨细胞/自制PDLLA复合物移植可较好地修复兔耳廓软骨缺损。     

聚羟基乙酸负载软骨细胞修复同种异体甲状软骨缺损

目的　探讨聚羟基乙酸 ( polyglycolicacid ,PGA)负载软骨细胞在有免疫力动物修复同种异体甲状软骨缺损的可行性。方法　酶消化法获取 3天龄新西兰乳兔肋软骨和关节软骨细胞 ,采用组织工程技术制备软骨细胞 PGA复合物 ,体外共同培养 1周后用于修复同种异体成年新西兰白兔甲状软骨缺损 (实验组 7只 )。设单纯PGA材料修复组 (对照A组 4只 )和单纯软骨细胞修复组 (对照B组 4只 )作为对照实验。分别于术后不同时间取材 ,对甲状软骨缺损的修复效果进行大体和组织学评价。结果　体外培养阶段可见黏附于PGA纤维表面的细胞分泌出丰富的软骨基质 ,呈蜘蛛网状分布于PGA纤维之间。术后 4周大体观察 :实验组修复区呈淡黄色 ,与正常软骨界限分明 ;组织学检查 :修复区内有软骨细胞生成和基质分泌 ,但与正常软骨间存在界面无细胞区 ,未见明显炎细胞浸润。 8周 :实验组修复区色乳白 ,与正常软骨仍有界限 ;镜下见修复区软骨细胞较成熟 ,软骨基质含量丰富。1 2周 :实验组修复区呈瓷白色 ,界面区软骨细胞不明显 ,但修复区软骨细胞数量、形态和基质与正常软骨相似。各时间点对照组大体观察修复区均呈不同程度的凹陷 ,暗红色 ,部分软组织充填其中 ,质软 ;组织学及特殊染色检查未发现软骨样结构及其分泌的基质成分。新生软骨     

转化生长因子β1和同种异体软骨细胞及高孔隙率聚乳酸复合物修复兔耳廓软骨缺损

目的　观察加入转化生长因子 β1(transforminggrowthfactor beta 1,TGF β1)的同种异体软骨细胞 /高孔隙率聚乳酸 (poly DL lactide,PDLLA)支架复合物修复兔耳廓软骨缺损的效果。 方法18只大白兔随机分为同种异体软骨细胞 /PDLLA复合物加TGF β1组 (复合物组 )、同种异体软骨细胞 /PDLLA复合物组 (复合物对照组 )和不用任何修复材料的空白对照组 ,每组 6只。分别于 4、12、18周取材 ,行HE和甲苯胺蓝染色 ,观察各组兔耳廓软骨缺损修复情况。结果　术后 18周 ,TGF β1复合物组修复软骨缺损的效果无论从大体标本观察或是病理组织学检查都比复合物对照组的修复效果好 ,空白对照组为纤维组织修复。结论　同种异体软骨细胞 /PDLLA复合物移植可形成组织工程化软骨修复兔耳廓软骨缺损 ,TGF β1可提高同种异体软骨细胞 /PDLLA复合物移植修复兔耳廓软骨缺损的质量。     

转化生长因子β1和同种异体软骨细胞及高孔隙率聚乳酸复合物修复兔耳廓软骨缺损

目的 观察加入转化生长因子-β1(transforming growth factor-beta1，TGF-β1)的同种异体软骨细胞／高孔隙率聚乳酸(poly-DL-lactide，PDLLA)支架复合物修复兔耳廓软骨缺损的效果。方法 18只大白兔随机分为同种异体软骨细胞／PDLLA复合物加TGF-β1组(复合物组)、同种异体软骨细胞／PDLLA复合物组(复合物对照组)和不用任何修复材料的空白对照组，每组6只。分别于4、12、18周取材，行HE和甲苯胺蓝染色，观察各组兔耳廓软骨缺损修复情况。结果 术后18周，TGF-β1复合物组修复软骨缺损的效果无论从大体标本观察或是病理组织学检查都比复合物对照组的修复效果好，空白对照组为纤维组织修复。结论 同种异体软骨细胞／PDLLA复合物移植可形成组织工程化软骨修复兔耳廓软骨缺损，TGF-β1可提高同种异体软骨细胞／PDLLA复合物移植修复兔耳廓软骨缺损的质量。     

聚羟基乙酸负载软骨细胞修复同种异体甲状软骨缺损

目的 探讨聚羟基乙酸(polyglycolic acid，PGA)负载软骨细胞在有免疫力动物修复同种异体甲状软骨缺损的可行性。方法 酶消化法获取3天龄新西兰乳兔肋软骨和关节软骨细胞，采用组织工程技术制备软骨细胞-PGA复合物，体外共同培养1周后用于修复同种异体成年新西兰白兔甲状软骨缺损(实验组7只)。设单纯PGA材料修复组(对照A组4只)和单纯软骨细胞修复组(对照B组4只)作为对照实验。分别于术后不同时间取材，对甲状软骨缺损的修复效果进行大体和组织学评价。结果 体外培养阶段可见黏附于PGA纤维表面的细胞分泌出丰富的软骨基质，呈蜘蛛网状分布于PGA纤维之间。术后4周大体观察：实验组修复区呈淡黄色，与正常软骨界限分明；组织学检查：修复区内有软骨细胞生成和基质分泌，但与正常软骨间存在界面无细胞区，未见明显炎细胞浸润。8周：实验组修复区色乳白，与正常软骨仍有界限；镜下见修复区软骨细胞较成熟，软骨基质含量丰富。12周：实验组修复区呈瓷白色，界面区软骨细胞不明显，但修复区软骨细胞数量、形态和基质与正常软骨相似。各时间点对照组大体观察修复区均呈不同程度的凹陷，暗红色，部分软组织充填其中，质软；组织学及特殊染色检查未发现软骨样结构及其分泌的基质成分。新生软骨内未见血管生长。结论PGA负载软骨细胞能修复具有正常免疫功能同种异体兔甲状软骨缺损，但新生软骨与正常软骨间存在无细胞区界面，无明显免疫排斥。     

气管食管科学

20030418’同种异体组织工程软骨修复兔气管软骨缺损实验研究/吴海涛…//中华耳鼻咽喉科杂志一2002,37(4)一310 采用同种异体组织工程软骨修复兔气管软骨缺损,结果显示组织工程软骨同种异体移植术早期存在炎症细胞浸润,术后1个月最明显,2个月后明显减少,这与移植初期异体软骨细胞尚未被软骨基质完全包埋引起排异反应及与PGA降解时引起无菌性炎症反应有关;3~4个月后,异体软骨细胞已被软骨基质完全包埋,PGA也已完全降解、吸收,所以新软骨周围基本上无炎症细胞浸润,说明组织工程软骨同种异体移植后远期排异反应并不明显。图4参4(勤思)200304…     

一般问题

20 0 4 0 4 81　　聚羟基乙酸负载软骨细胞修复同种异体甲状软骨缺损 /孙安科… / /中华耳鼻咽喉科杂志 - 2 0 0 3;38(5 ) - 347～ 35 0目的 :探讨聚羟基乙酸 (PGA)负载软骨细胞在有免疫力动物修复同种异体甲状软骨缺损的可行性。方法 :酶消化法获取 3天龄新西兰乳兔肋软骨和关节软骨细胞 ,采用组织工程技术制备软骨细胞PGA复合物 ,体外共同培养 1周后用于修复同种异体成年新西兰白兔甲状软骨缺损 (实验组 7只 )。设单纯PGA材料修复组 (对照A组 4只 )和单纯软骨细胞修复组 (对照B组 4只 )作为对照实验。分别于术后不同时间取材 ,对甲…     

鼻中隔软骨细胞组织工程法构建预定形态软骨

目的探讨利用人鼻中隔软骨细胞组织工程方法构建预定形态软骨的可能性。方法将人鼻中隔软骨细胞播种在聚乙醇酸（polyglycolicacid,PGA）无纺网支架材料上,制成片状和管状结构,埋入裸鼠体内,经4、6、8周后取材作大体及组织学观察。结果大体观察见裸鼠体内形成了预定的片状和管状软骨。组织学观察6周时软骨细胞基本成熟,Masson三色染色显示胶原形成,番红花-“O”染色证实其基质中存在糖氨多糖。对照组于6周时PGA纤维基本消失。结论人鼻中隔软骨细胞与PGA无纺网复合可在裸鼠体内形成预定形态软骨。     

Characteristics of tissue-engineered cartilage on macroporous biodegradable PLGA scaffold

BACKGROUND: The purpose of this study was to establish in vivo culture of chondrocytes on biodegradable, poly-D,L-lactic-co-glycolic acid (PLGA) scaffolds and to analyze the characteristics of the reconstructed cartilage. METHODS: In vitro cultured chondrocytes that were grown on a polyhydroxyethyl methacrylate (poly-HEMA) coated dish were seeded onto the PLGA scaffolds to make a cell-polymer construct before implantation. One cell scaffold construct was carefully implanted in the subcutaneous pocket of a nude mouse and another cell-free scaffold was implanted in the opposite side of the same nude mouse as the control. Morphologic, biochemical, and immunohistochemical characteristics of cells cultured within the PLGA constructs were examined after 8 weeks and 16 weeks of harvesting in the nude mouse. RESULTS: New cartilage began to be generated in the period of 8 weeks and the neocartilage formation was accomplished in 4 months with the exact dimensions of the original scaffold in this in vivo study. All the explants showed the irregular shape of viable chondrocytes within normal lacunae and a mature cartilaginous matrix, and they positively immunostained for collagen type II. CONCLUSION: The new tissue-engineered cartilage in vivo on PLGA scaffolds displayed the biochemical characteristics of cartilage tissue, and it showed chondrocyte-specific phenotypes and morphology that were similar to the native cartilage.     

Cartilage tissue engineering using cryogenic chondrocytes

OBJECTIVE: To generate in vitro hyaline cartilage from cryogenically preserved human septal chondrocytes in a simulated microgravity environment on a 3-dimensional biodegradable scaffolding material. METHODS: In this experiment, cryogenically frozen chondrocytes were thawed and cultured in a monolayer in serum-based chondrocyte media. They were seeded onto 3-dimensional biopolymer scaffolds in a spinner flask. The seeded constructs were then transferred to a bioreactor (an environment of solid-body rotation) for 6 weeks. Chondrocyte growth and extracellular matrix production in the constructs were confirmed by cell count, cell viability, and histologic analysis and by electron microscopy. RESULTS: Histologic sections stained with hematoxylin-eosin and Alcian blue (for acidic proteoglycans) confirmed the presence of hyaline cartilage in the cartilage constructs. Ultrastructural examination using transmission electron microscopy demonstrated matrix formation and chondrocyte viability. CONCLUSIONS: This study proves that chondrocytes that are cryogenically stored for extended periods can be used to grow cartilage in vitro. Cryogenically preserved chondrocytes retain their ability to grow in tissue culture, redifferentiate, and produce extracellular matrix.     

Tracheal reconstruction using tissue-engineered cartilage

OBJECTIVES: To determine whether rabbit cartilage can be tissue engineered using a polyglycolic acid (PGA) construct composed of PGA mesh, autologous chondrocytes, and alginate covalently linked with the cell adhesion sequence arginine-glycine-aspartic acid (RGD), and to investigate the feasibility of reconstructing tracheal defects using the PGA construct in conjunction with a bioabsorbable intratracheal stent. METHODS: Nineteen New Zealand White rabbits were used. Nine rabbits underwent subcutaneous implantation of 3 different PGA construct combinations: (1) PGA, autologous chondrocytes, and RGD-modified alginate; (2) PGA, autologous chondrocytes, and unmodified alginate; and (3) PGA and RGD-modified alginate. The remaining 10 animals underwent anterior tracheal reconstruction using fascia lata grafts and the complete PGA construct (PGA, autologous chondrocytes, and RGD-modified alginate). At the time of tracheal reconstruction, a poly-l-lactic acid intratracheal stent was placed in 5 of these latter animals. Rates of tracheal stenosis and mortality were compared with those of historical control animals. Histologic analysis was performed on the PGA constructs. RESULTS: In the subcutaneous implants, the PGA constructs made with chondrocytes (with and without RGD) demonstrated mature cartilage formation in 7 (78%) of the 9 animals. No cartilage was seen in PGA constructs made without chondrocytes. Two of the 10 animals that underwent tracheal reconstruction with the complete PGA construct survived to 20 weeks and demonstrated patent airways, 1 with a stent and 1 without a stent (80% overall mortality). Histologic analysis showed mature cartilage formation at the tracheal reconstruction site. Historical control animals that underwent reconstruction with fascia lata alone demonstrated the lowest overall mortality. CONCLUSIONS: Cartilage can be tissue engineered in rabbits using PGA mesh embedded with alginate-encapsulated autologous chondrocytes. It is also possible to reconstruct tracheal defects with this method of cartilage engineering, although the mortality rate in this study is high.     

Age dependence of cellular properties of human septal cartilage: implications for tissue engineering

BACKGROUND: The persistent need for cartilage replacement material in head and neck surgery has led to novel cell culture methods developed to engineer cartilage. Currently, there is no consensus on an optimal source of cells for these endeavors. OBJECTIVES: To evaluate human nasal cartilage as a potential source of chondrocytes and to determine the effect of donor age on cellular and proliferation characteristics. SUBJECTS: Nasal cartilage specimens were obtained after reconstructive surgery from 46 patients ranging in age from 15 to 60 years. METHODS: Specimens were weighed and chondrocytes were isolated by digestion in 0.2% collagenase type II for 16 hours. Cells were maintained in primary cultures until confluency, then seeded onto polylactic acid-polyglycolic acid scaffolds. Seeding efficiency was determined by quantification of DNA content of seeded constructs by means of Hoechst dye 33258. Specimen weights, cell yields, cell content, and doubling time were also measured and correlated to donor age. RESULTS: Mean (+/-SD) cartilage mass obtained (648 +/- 229 mg) is higher than from typical biopsy specimens of auricular cartilage, and the cellular characteristics show a higher proliferation rate than auricular chondrocytes. Cell yield increased with age, while doubling time decreased with age in samples from patients ranging from 15 to 60 years old. CONCLUSIONS: The use of nasal septal cartilage as a source of cells for tissue engineering may be valid over a wide range of patient ages. The large tissue yield and consequent cell yield make this tissue a potential starting source of chondrocytes for large-volume tissue-engineered implants.     

Fabrication of cartilage in predetermined shapes from human nasoseptal chondrocytes with tissue engineering method]

OBJECTIVE: To investigate the feasibility of fabricating a new cartilage in predetermined shapes with tissue engineering methods. METHODS: Human nasoseptal chondrocytes were seeded onto a nonwoven mesh of polyglycolic acid(PGA) to form a cell-PGA construct. The construction was then configured in sheet and tube shapes, and implanted subcutaneously into the dorsa of 11 athymic mice. The specimens were harvested 4, 6, 8 weeks after implantation and subjected to gross morphologic and histologic analysis. RESULTS: Gross observation showed that the predetermined sheet and tube shapes of new cartilage were formed. Histological observation demonstrated that new mature cartilages were formed in 6-week. A Masson's trichrome stain showed the interwining bands of collagen at the periphery of the cartilage. Staining of Safranin O evaluated that the new cartilage was bound of glycosaminoglycan. In the control group, the PGA of the specimens were completely absorbed at 6 weeks. CONCLUSION: Human nasoseptal chondrocytes-PGA construct could develop into a new cartilage in predetermined shapes in athymic mice.     

Tissue-engineered human nasal septal cartilage using the alginate-recovered-chondrocyte method

OBJECTIVES: Tissue engineering of nasal septal cartilage has numerous potential applications in craniofacial reconstruction. Chondrocytes suspended in alginate gel have been shown to produce a substantial cell-associated matrix. The objective of this study was to determine whether cartilage tissue could be generated using the alginate-recovered-chondrocyte (ARC) method, in which chondrocytes are cultured in alginate as an intermediate step in tissue fabrication. METHODS: Nasal septal chondrocytes from five patient donors were isolated by enzymatic digestion, then expanded in monolayer culture. At confluency, a portion of those cells were seeded at high density onto a semipermeable membrane and cultured for 14, 21, or 28 days (monolayer group). The remaining cells were suspended in alginate and cultured until a cell-associated matrix was observed (10-17 days). Cells and their associated matrix were released from alginate (ARC group), seeded onto a semipermeable membrane, and cultured as already described. DNA (Hoechst 33258 Assay), glycosaminoglycan (GAG; dimethylmethylene blue assay), and collagen (hydroxyproline assay) were analyzed biochemically. Immunohistochemistry was performed to assess expression of collagens type I and type II. Histochemistry was performed to localize cells accumulating sulfated GAG (Alcian blue stain). RESULTS: The ARC constructs, in contrast to the monolayer constructs, had substantial structural stability and the histologic and gross appearance of cartilaginous tissue. ARC constructs demonstrated significantly greater GAG and collagen accumulation than monolayer constructs (P <.05). Histologic analysis revealed substantial GAG and collagen type II production and only moderate collagen type I production. The composition of the matrix was thus similar to that of native human septal cartilage. CONCLUSIONS: Tissue-engineered human nasal septal cartilage using the ARC method has the histologic and gross appearance of native cartilage and has biochemical composition more like that of native cartilage than monolayer constructs. This is the first report of human nasal septal neocartilage formation without the use of biodegradable scaffolds.     

Human septal chondrocyte redifferentiation in alginate,polyglycolic acid scaffold,and monolayer culture

OBJECTIVES/HYPOTHESIS: Tissue engineering laboratories are attempting to create neocartilage that could serve as an implant material for structural support during reconstructive surgery. One approach to forming such tissue is to proliferate chondrocytes in monolayer culture and then seed the expanded cell population onto biodegradable scaffolds. However, chondrocytes are known to dedifferentiate after this type of monolayer growth and, as a result, decrease their production of cartilaginous extracellular matrix components such as sulfated glycosaminoglycans. The resultant tissue lacks the biomechanical properties characteristic of cartilage. The objective of the study was to determine whether different culture systems could induce monolayer-expanded human septal chondrocytes to redifferentiate and form extracellular matrix. STUDY DESIGN: Laboratory research. METHODS: Chondrocytes were isolated from human nasal septal cartilage of five donor patients (age, 35.8 +/- 9.3 y). Cell populations were seeded at low density (30,000 cells/cm2) into monolayer culture and expanded for 4 to 6 days. Following trypsin release, chondrocytes were placed into three different systems for neocartilage formation: alginate beads, polyglycolic acid scaffolds, and monolayer. After 7 and 14 days of growth, neocartilage was analyzed using histological and quantitative biochemical assessment of cellularity (Hoechst 33258 assay) and sulfated glycosaminoglycan content (dimethyl methylene blue assay). RESULTS: Histologically, alginate beads contained spherical chondrocytes surrounded by dense extracellular matrix, an appearance similar to that of native cartilage. In contrast, polyglycolic acid scaffolds and monolayer cultures contained elongated cells with scant staining for matrix sulfated glycosaminoglycans, which are features that are characteristic of dedifferentiated chondrocytes. Biochemical analysis demonstrated a lower level of cell proliferation (P <.001) in scaffolds (+52% over baseline) and alginate (+96% over baseline) than in monolayer (+366% over baseline), as well as a higher content of sulfated glycosaminoglycans per cell (P <.001), after 14 days of growth in alginate culture than in either polyglycolic acid scaffolds (19-fold difference) or monolayer (98-fold difference). CONCLUSIONS: Of the systems compared, monolayer-expanded human septal chondrocytes demonstrated the greatest accumulation of sulfated glycosaminoglycans per cell when grown in alginate beads. Future research on cartilage tissue engineering may use alginate culture for reverting dedifferentiated cells back to the chondrocytic phenotype.     

Internal support of tissue-engineered cartilage

BACKGROUND: Auricles previously created by tissue engineering in nude mice used a biodegradable internal scaffold to maintain the desired shape of an ear. However, the biodegradable scaffold incited a compromising inflammatory response in subsequent experiments in immunocompetent animals. OBJECTIVE: To test the hypothesis that tissue-engineered autologous cartilage can be bioincorporated with a nonreactive, permanent endoskeletal scaffold. MATERIALS AND METHODS: Auricular elastic cartilage was harvested from Yorkshire swine. The chondrocytes were isolated and suspended into a hydrogel (Pluronic F-127) at a cell concentration of 5 x 10(7) cells/mL. Nonbiodegradable endoskeletal scaffolds were formed with 1 of 5 polymers: (1) high-density polyethylene, (2) soft acrylic, (3) polymethylmethacrylate, (4) extrapurified Silastic, and (5) conventional Silastic. Three groups were studied: (1) a control group using only the 5 polymers, (2) the 5 polymers enveloped by Pluronic F-127 only, and (3) the implants coated with Pluronic F-127 seeded with chondrocytes. All constructs were implanted subdermally; implants containing cells were implanted into the same animal from which the cells had been islolated. The implants were harvested after 8 weeks of in vivo culture and histologically analyzed. RESULTS: Only implants coated by hydrogel plus cells generated healthy new cartilage. With 3 polymers (high-density polyethylene, acrylic, and extrapurified Silastic), the coverage was nearly complete by elastic cartilage, with minimal fibrocartilage and minimal to no inflammatory reaction. The Food and Drug Administration-approved conventional Silastic implants resulted in fragments of fibrous tissue mixed with elastic cartilage plus evidence of chronic inflammation. The polymethylmethacrylate implant was intermediate in the amount of cartilage formed and degree of inflammation. CONCLUSIONS: This pilot technique combining tissue-engineered autologous elastic cartilage with a permanent biocompatible endoskeleton demonstrated success in limiting the inflammatory response to the scaffold, especially to high-density polyethylene, acrylic, and extrapurified Silastic. This model facilitates the potential to generate tissue of intricate shape, such as the human ear, by internal support. Arch Otolaryngol Head Neck Surg. 2000;126:1448-1452     

组织工程化兔耳廓软骨的实验研究

目的 探讨采用表面修饰过的壳聚糖（chitosan)-聚乳酸（polylacticacid,PLA)－聚乙内酯（polycrylactone,PLC)及牛跟腱胶原海绵作为支架培养组织工程化耳廓软骨的可行性，比较动态与静肪两种不同培养方法对软骨生长的影响。方法 将4周龄新西兰大耳白兔耳廓软骨细胞接种在20个壳聚－PLA－PCL及牛跟腱I型胶原海绵支架上，将细胞支架复合物分为2组，动态组（样本量为10）采用旋转培养，静态组（样本量为10）采用静置培养。所有标本体外培养1周，裸鼠或兔皮下培养8周。分别于体外1周，体内4周及8周时取样行扫描电镜检查，大体及组织学观察以及免疫组化分析，测定软骨细胞数及细胞外基质中的Ⅱ型胶原含量。结果 表面修饰过的壳聚糖支架上软骨细胞贴壁率高，细胞分化与增殖好；每个时段动态组生成的软骨细胞量及Ⅱ型胶原均较静态组多，两组间差异有显著性（P＜0．05）；2种生物支架上均长出了组织工程软骨；裸鼠皮下较兔皮下形成的软骨更接近天然软骨。结论 壳聚糖－PLA－PCL及牛跟腱胶原海绵是较理想的组织工程耳廓软骨的支架材料，动态培养更有利于新生软骨组织的生成，兔皮下仅能形成软骨样组织。