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。     

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。     

聚乳酸-羟基乙酸输尿管支架植入后犬输尿管周围的组织学变化

背景：泌尿系统组织工程支架不仅需要生物相容性良好的生物材料,而且一定要利于组织周围细胞的生长。 目的：制备聚乳酸-羟基乙酸共聚物可降解输尿管支架,观察其植入后犬输尿管周围组织学变化。 方法：制备纳米聚乳酸-羟基乙酸共聚物输尿管支架,并以多聚赖氨酸对支架进行交联、改性,将交联后支架截成长约0.8 cm小段,植入犬损伤输尿管中进行体内观察实验。 结果与结论：①支架制备：支架具有纳米结构,孔隙率约90%,孔径(30±18) µm,多聚赖氨酸交联改性后纤维表面略显粗糙。②支架变化：支架植入30 d时已完全失去原始形态,与周边组织融合,可见裂解小块。③支架植入后输尿管周围组织学变化：植入后15 d炎症表现最为明显,主要是移行上皮脱落,肌层结构被破坏,固有层水肿明显;30 d后,炎症已经明显好转,但组织结构依然不规则;植入后45 d,输尿管全层组织基本恢复正常,组织结构成规则分布。说明聚乳酸-羟基乙酸共聚物输尿管支架具有良好的组织相容性,符合泌尿系统组织工程支架的要求。      

新型组织工程化骨材料植入动物体内的成血管效应

背景：以生长因子、种子细胞、载体支架为基础的骨组织工程研究取得的成功,向人们展示了再造骨组织器官的美好前景,然而在临床应用方面往往效果不理想。其中很重要一个原因是组织工程骨很大程度上受制于移植物血管网缺乏造成的细胞供养障碍而导致失效。 目的：新型组织工程骨修复材料植入新西兰兔桡骨缺损处观察其成血管作用。 方法：将聚乳酸-聚羟基乙酸共聚物包裹碱性成纤维细胞生长因子制备成微球囊,然后与磷酸钙骨水泥混合,并与体外培养的同种异体骨髓间充质干细胞共培养制备新型组织工程骨修复材料。60只成年新西兰兔建立15 mm桡骨缺损模型后随机分成2组,实验组植入新型组织工程骨修复材料,对照组植入复合骨髓间充质干细胞的聚乳酸-聚羟基乙酸共聚物与磷酸钙骨水泥的混合材料。于术后4、8、12周,通过组织细胞形态学观察、核素骨扫描等手段,观察各个时期血管形成情况。 结果与结论：光镜下组织形态学观察结果及核素骨扫描结果示血管化程度是实验组优于对照组。 结果显示聚乳酸-聚羟基乙酸共聚物-成纤维细胞生长因子/磷酸钙骨水泥材料复合骨髓间充质干细胞构建的新型组织工程骨修复材料在动物体内有较好的成血管效果。     

无纺网与多孔海绵状材料作为软骨组织工程支架的适用性及体内降解性

背景：聚羟基乙酸无纺网与聚羟基丁酸酯-聚羟基己酸酯共聚物多孔海绵具有良好的塑形适应性、生物降解性与生物相容性。 目的：观察聚羟基乙酸无纺网与聚羟基丁酸酯-聚羟基己酸酯共聚物多孔海绵作为软骨组织工程支架的适用性及体内降解性。 方法：分别制备乳兔软骨细胞-聚羟基乙酸无纺网复合物、乳兔软骨细胞-聚羟基丁酸酯-聚羟基己酸酯共聚物多孔海绵复合物。在实验组成年兔两侧背部皮下分别植入制备的两种复合物,在对照组成年兔两侧背部皮下分别植入聚羟基乙酸无纺网与聚羟基丁酸酯-聚羟基己酸酯共聚物。 结果与结论：组织学观察显示,以聚羟基乙酸无纺网获取的组织工程软骨,植入4 周时软骨细胞较小,软骨内有较多聚羟基乙酸纤维残留,8周时软骨细胞较成熟,包埋在陷窝内,聚羟基乙酸纤维消失,12周时软骨细胞成熟,基质分泌丰富,无聚羟基乙酸存留;以聚羟基丁酸酯-聚羟基己酸酯共聚物多孔海绵获取的组织工程软骨,植入4周时软骨细胞不成熟,软骨基质内似“杂质”样材料残留物较多,8周时软骨细胞较成熟,软骨基质内仍可见材料残留,12周时软骨基质材料残留基本消失。两组组织工程软骨特殊染色与免疫组织化学检测均显示再生软骨胶原与基质黏多糖生成良好,软骨中均检测出Ⅱ型胶原。表明两种材料作为软骨组织工程支架具有良好的适用性,其降解时间均达到组织工程软骨构建的要求。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;组织工程全文链接：     

电纺纳米纤维可降解输尿管支架肌肉埋植的降解性能*

背景：作者前期研究了电纺纳米纤维聚乳酸-羟基乙酸共聚物可降解输尿管支架材料的体外降解性能,发现80/20聚乳酸-羟基乙酸共聚物电纺纳米纤维材料在尿液中的降解时间可以满足临床需要。目的：观察80/20聚乳酸-羟基乙酸共聚物电纺纳米纤维输尿管支架的肌肉埋植降解性能。方法：采用静电纺丝法制备80/20聚乳酸-羟基乙酸共聚物纳米纤维输尿管支架,观察其在家兔脊柱旁肌肉中的降解情况。结果与结论：成功制备了电纺纳米纤维输尿管支架,扫描电镜见微观形貌良好。80/20的聚乳酸-羟基乙酸共聚物纳米纤维输尿管支架在体内降解至10周时,支架管降解至初始质量的60%左右,支架出现断裂和崩解,虽降解速度较体外降解稍慢,但其降解性能仍能够满足临床对可降解输尿管支架的需要。     

聚乳酸/壳聚糖纳米纤维/纳米羟基磷灰石支架与兔软骨细胞的相容性

背景：传统的支架材料存在疏水性强,材料表面缺乏细胞表面受体特异结合的生物活性分子,材料的酸性降解产物易引发无菌性炎性反应等不足。根据仿生原理及软骨真实结构和构成来选择和制备组织工程软骨支架能够获得理想效果。 目的：制备聚乳酸/壳聚糖纳米纤维/纳米羟基磷灰石支架,评价其与兔膝关节软骨细胞的生物相容性,探讨其应用于关节软骨组织工程的可行性。 方法：采用二次相分离技术制备聚乳酸/壳聚糖纳米纤维/纳米羟基磷灰石复合支架,将第3代新西兰兔软骨细胞接种至复合支架材料上复合培养,倒置相差显微镜下观察细胞生长情况。细胞-支架复合物在24孔板中培养5 d以后,将其植入裸鼠皮下8周。 结果与结论：聚乳酸/壳聚糖纳米纤维/纳米羟基磷灰石支架材料经化学合成后,具有合适的三维多孔结构,孔隙率为90%,孔径300~450 μm;植入裸鼠皮下8周后Ⅱ型胶原免疫组织化学染色和甲苯胺蓝染色显示细胞-支架复合物中的软骨细胞可以像天然软骨一样分泌黏多糖和Ⅱ型胶原。提示生物材料聚乳酸/壳聚糖纳米纤维/纳米羟基磷灰石对于兔软骨细胞有良好的生物相容性,可作为生物组织工程支架。     

静电纺丝制备聚乳酸乙醇酸共聚物-胶原皮肤组织工程支架的实验研究

制备具有良好生物相容性的组织工程皮肤支架。以聚乳酸乙醇酸共聚物和胶原为原料,利用静电纺丝的方法,制备组织工程皮肤支架,并观察支架的形貌、降解等物理性能,利用CCK-8、伊红染色等方法观察支架的生物相容性。支架具有良好的纤维形貌,直径均匀。支架在磷酸盐缓冲液中降解8周,质量损失为35%,表明支架具有良好的生物可降解性。人成纤维细胞种植在支架上后,生长情况良好。种植3 d后,细胞存活率为75%;种植14 d后,细胞存活率为90%,表明细胞支架没有明显的细胞毒性,具有良好的生物相容性。用静电纺丝制备的聚乳酸乙醇酸共聚物-胶原支架是一种良好的皮肤组织工程支架,可以构建具有生物学功能的组织工程皮肤。     

生长因子缓释系统复合纳米支架材料修复犬下颌骨缺损

背景：支架材料联合细胞因子构建组织工程骨不受血管化和细胞培养因素的限制,这种构建模式可能诱导出较大体积的实用型组织工程骨。 目的：观察壳聚糖纳米微球/纳米羟基磷灰石/聚乳酸-羟基乙酸复合生长因子缓释支架修复犬下颌骨临界骨缺损的能力。 方法：取杂种犬12条,制作双侧下颌骨临界骨缺损模型,一侧植入复合生长因子骨形态发生蛋白2、转化生长因子β1及血管内皮生长因子165的壳聚糖纳米微球/纳米羟基磷灰石/聚乳酸-羟基乙酸缓释支架(实验组),另一侧植入壳聚糖纳米微球/纳米羟基磷灰石/聚乳酸-羟基乙酸缓释支架(对照组),术后4,8,12周取下颌骨标本行X 射线、组织学及免疫组织化学检查。 结果与结论：实验组术后不同时间点X射线灰度值及骨钙素积分吸光度值均高于对照组(P < 0.05),表明复合生长因子的支架材料修复骨缺损的成骨能力优于未复合生长因子的支架材料。组织学观察结果显示,实验组术后不同时间点成骨时间及效果均优于对照组,表明复合生长因子骨形态发生蛋白2、转化生长因子β1及血管内皮生长因子165的壳聚糖纳米微球/纳米羟基磷灰石/聚乳酸-羟基乙酸缓释支架可更快更有效地促进骨缺损修复。     

蚕丝/聚乳酸-羟基乙酸共聚物支架降解液与骨髓间充质干细胞的增殖

背景：前期研究表明,蚕丝/聚乳酸-羟基乙酸共聚物支架浸提液具有良好的细胞相容性,基本无细胞毒性。 目的：观察蚕丝/聚乳酸-羟基乙酸共聚物纤维细丝混合编织支架体外长期降解过程中降解液对兔骨髓间充质干细胞增殖活性的影响。 方法：将蚕丝/聚乳酸-羟基乙酸共聚物细丝混合编织支架材料置于完全培养基中体外降解14周,每周换液1次,测定各周支架降解液的pH值。将兔骨髓间充质干细胞分组培养,实验组加入各周支架降解液和新鲜完全培养基各100 µL,阴性对照组加入完全培养基200 µL,培养4 d。MTT法检测细胞增殖、生长情况。 结果与结论：①支架降解液pH值的变化：前3周下降缓慢,从7.00降到6.89;第4周起下降较快,6-11周较低,在5.16-5.67之间;12-14周呈上升趋势,回升到6.95。②骨髓间充质干细胞形态：实验组及阴性对照组细胞增殖生长及形态状况基本相似。降解7-10周支架降解液对细胞的生长有抑制作用,细胞数量相对较少、较疏,而其余各周支架降解液对细胞生长无明显抑制作用。③骨髓间充质干细胞的增殖：1-6周及11-14周的支架降解液对细胞增殖无显著影响,细胞相对增殖率均在92.1%以上,毒性分级为0或1级;7-10周的支架降解液虽对细胞增殖有抑制作用,但细胞相对增殖率为82.5%-87.9%,毒性分级为1级,为合格。表明蚕丝丝素/聚乳酸-羟基乙酸共聚物混合编织支架降解液具有良好的细胞相容性。     

Human dental pulp cell culture and cell transplantation with an alginate scaffold

Many studies on tissue stem cells have been conducted in the field of regenerative medicine, and some studies have indicated that cultured dental pulp mesenchymal cells secrete dentin matrix. In the present study we used alginate as a scaffold to transplant subcultured human dental pulp cells subcutaneously into the backs of nude mice. We found that when beta-glycerophosphate was added to the culture medium, dentin sialophosphoprotein mRNA coding dentin sialoprotein (DSP) was expressed. An increase in alkaline phosphatase, which is an early marker for odontoblast differentiation, was also demonstrated. At 6 weeks after implantation the subcutaneous formation of radio-opaque calcified bodies was observed in situ. Immunohistochemical and fine structure studies identified expression of type I collagen, type III collagen, and DSP in the mineralizing transplants. Isolated odontoblast-like cells initiated dentin-like hard tissue formation and scattered autolyzing apoptotic cells were also observed in the transplants. The study showed that subcultured dental pulp cells actively differentiate into odontoblast-like cells and induce calcification in an alginate scaffold.     

Accurately shaped tooth bud cell-derived mineralized tissue formation on silk scaffolds

Based on the successful use of silk scaffolds in bone tissue engineering, we examined their utility for mineralized dental tissue engineering. Four types of hexafluoroisopropanol (HFIP) silk scaffolds-(250 and 550 microm diameter pores, with or without arginine-glycine-aspartic acid (RGD) peptide) were seeded with cultured 4-day postnatal rat tooth bud cells and grown in the rat omentum for 20 weeks. Analyses of harvested implants revealed the formation of bioengineered mineralized tissue that was most robust in 550 microm pore RGD-containing scaffolds and least robust in 250 microm pore sized scaffolds without RGD. The size and shape of the silk scaffold pores appeared to guide mineralized tissue formation, as revealed using polarized light imaging of collagen fiber alignment along the scaffold surfaces. This study is the first to characterize bioengineered tissues generated from tooth bud cells seeded onto silk scaffolds and indicates that silk scaffolds may be useful in forming mineralized osteodentin of specified sizes and shapes.     

Isolated rat dental pulp cell culture and transplantation with an alginate scaffold

Many studies have been conducted on tissue stem cells in the field of regenerative medicine, and cultured dental pulp mesenchymal cells have been reported to secrete dentin matrix. In the present study we used alginate as a scaffold to transplant subcultured rat dental-pulp-derived cells subcutaneously into the back of nude mice. We found that when beta-glycerophosphate was added to the culture medium, the mRNA of the dentin sialophosphoprotein (DSPP) gene coding dentin sialoprotein (DSP) and dentin phosphoprotein (DPP) was expressed, and an increase in alkaline phosphatase, an early marker of odontoblast differentiation, was also demonstrated. Six weeks after implantation, subcutaneous formation of radiopaque calcified bodies was observed in situ. Immunohistochemical and fine structure studies identified expression of type I collagen, type III collagen, and DSP in the mineralizing transplants, and isolated odontoblast-like cells began to form dentin-like hard tissue formation. Scattered autolyzing apoptotic cells were also observed in the transplants. The study showed that subcultured rat dental-pulp-derived cells actively differentiate into odontoblast-like cells and induce calcification in an alginate scaffold.     

Models for the Study of Cementogenesis

Cementum is a mineralized tissue that acts to connect the periodontal ligament to the tooth root surface. Its composition is very much like bone, being comprised mainly of type I collagen, inorganic mineral and noncollagenous proteins, however the origin of the cells and factors necessary for cementum formation have yet to be elucidated.

Our laboratory has focused on the role that adhesion molecules, and their cell surface receptors, play in the formation of cementum and tooth root. In order to study this, we used a mouse molar as a model system. This system enabled us to study the formation of four distinct mineralized tissues; bone, cementum, dentin and enamel at various stages of their development. For these studies, we initiated experiments to examine potential cementoblast progenitor cells, in vitro. As a first step, we show that dental papilla and dental follicle cells, n vitro, obtained from molar tissues at day 21 of development, induce mineralized nodules, in vitro

In addition, we obtained tissues from mice where defects in root development may exist and determined bone sialoprotein (BSP) protein expression, a mineralized tissue specific adhesion molecule, in such tissues. As discussed here, we found that osteopetrotic (op/op) mice have delayed and/or defective root development and BSP does not localize in the dental tissues, at day 33 of development. In addition, dentin formation was defective and odontoblasts appeared immature, based on morphological examination. In contrast, the day 33 control molars demonstrated positive staining for BSP localized to root cementum, with normal formation of dentin.     

The performance of human dental pulp stem cells on different three-dimensional scaffold materials

The aim of this study was to investigate the in vitro and in vivo behavior of human dental pulp stem cells (DPSCs) isolated from impacted third molars, when seeded onto different 3-dimensional (3-D) scaffold materials: i.e. a spongeous collagen, a porous ceramic, and a fibrous titanium mesh. Scaffolds were loaded with DPSC, and subsequently divided into two groups. The first group was cultured in osteogenic differentiation medium in vitro for 4 weeks. The second group of samples was implanted subcutaneously in nude mice for 6 or 12 weeks. Samples cultured in vitro were analyzed by scanning electron microscopy and RT-PCR for dentin sialophosphoprotein (DSPP) expression. In vivo samples were evaluated by histology, RT-PCR and immunohistochemistry. The results indicated that in vitro, cells developed abundant deposition of mineralized extracellular matrix (ECM) with expression of DSPP in all 3-D materials. The simultaneous implantation experiment showed formation of tissue that was DSPP positive in all three scaffolds materials. However, the aspect of the formed tissues in all scaffolds resembled more connective tissue than a dentin-like tissue. Limited calcification of the ECM was only seen in the ceramic scaffold. In both experiments, no other differences could be attributed to the different materials used. In conclusion, the in vivo behavior of DPSC and their relations with 3-D scaffold materials should be further studied before clinical use can be considered.     

Investigation of dental pulp stem cells isolated from discarded human teeth extracted due to aggressive periodontitis

Recently, human dental pulp stem cells (DPSCs) isolated from inflamed dental pulp tissue have been demonstrated to retain some of their pluripotency and regenerative potential. However, the effects of periodontal inflammation due to periodontitis and its progression on the properties of DPSCs within periodontally compromised teeth remain unknown. In this study, DPSCs were isolated from discarded human teeth that were extracted due to aggressive periodontitis (AgP) and divided into three experimental groups (Groups A, B and C) based on the degree of inflammation-induced bone resorption approaching the apex of the tooth root before tooth extraction. DPSCs derived from impacted or non-functional third molars of matched patients were used as a control. Mesenchymal stem cell (MSC)-like characteristics, including colony-forming ability, proliferation, cell cycle, cell surface antigens, multi-lineage differentiation capability and in vivo tissue regeneration potential, were all evaluated in a patient-matched comparison. It was found that STRO-1- and CD146-positive DPSCs can be isolated from human teeth, even in very severe cases of AgP. Periodontal inflammation and its progression had an obvious impact on the characteristics of DPSCs isolated from periodontally affected teeth. Although all the isolated DPSCs in Groups A, B and C showed decreased colony-forming ability and proliferation rate (P < 0.05), the decreases were not consistent with the degree of periodontitis. Furthermore, the cells did not necessarily show significantly diminished in vitro multi-differentiation potential. Only DPSCs from Group A and the Control group formed dentin-like matrix in vivo when cell-seeded biomaterials were transplanted directly into an ectopic transplantation model. However, when cell-seeded scaffolds were placed in the root fragments of human teeth, all the cells formed significant dentin- and pulp-like tissues. The ability of DPSCs to generate dental tissues decreased when the cells were isolated from periodontally compromised teeth (P < 0.05). Again, increased periodontal destruction was not necessarily followed by a decrease in the amount of dentin- and pulp-like tissue formed. These findings provide preliminary evidence that periodontally compromised teeth might contain putative stem cells with certain MSC properties, as long as the vitality of the pulp has not been totally damaged. Whether these cells can serve as a source of autologous multipotent MSCs for clinical regenerative therapies warrants further investigation with larger sample sizes and various types of periodontitis.     

The induction of enamel and dentin complexes by subcutaneous implantation of reconstructed human and murine tooth germ elements

Tooth induction by xenogenic graft of reconstructed human tooth germ components has never been attempted. Here we report our first attempt at a transplantation of human tooth germ components, heterologously recombined with mouse dental epithelia, into immunocompromised animals. Human third molar tooth germs enucleated from young patients as prophylactic treatment for orthodontic reasons were collected. The whole or minced human dental papilla was reconstructed with human- or mouse molar enamel epithelium, and transplanted in the dorsal aspect of C.B-17/Icr-scid Jcl mice. The transplant of human dental papilla reconstructed with human enamel epithelium formed thin dentin and immature enamel layers by 3 to 4 weeks, but remained extremely small in quantity due to a shortage of epithelial components in the graft. The addition of E16 mouse molar enamel organs (n=10-12) to each graft augmented the formation of tooth germ-like structures, but the differentiation of mouse molar ameloblasts was suppressed. However, once a solid layer of mineralized dentin was established, mouse ameloblasts accelerated their differentiation, and completed the enamel matrix formation and maturation within the following 4 weeks, whereas human ameloblasts, which had interacted with human dental papilla, remained in the stage of matrix formation during the same period. These data imply that, in reconstructed transplants, the differentiation of mouse dental epithelia is restrained by putative suppressive factors derived from human dental papilla until they are separated by mineralized dentin layers that serve as a diffusion barrier. The mouse enamel organ nevertheless retains its own phenotypic characteristics and intrinsic timing of cell differentiation and function.     

Role of synthetic extracellular matrix in development of engineered dental pulp

In cases of damaged oral tissues, traditional therapies, such as a root canal, replace the injured tissue with a synthetic material. However, while the materials currently used can offer structural replacement of the lost tissue, they are incapable of completely replacing the function of the original tissue, and often fail over time. This report describes a tissue engineering approach to dental pulp tissue replacement utilizing cultured cells seeded upon synthetic extracellular matrices. Human pulp fibroblasts were obtained and multiplied in culture. These cells were then seeded onto three different synthetic matrices: scaffolds fabricated from polyglycolic acid (PGA) fibers, a type I collagen hydrogel, and alginate in an effort to examine which matrix is most suitable for dental pulp tissue formation. In addition, methods previously developed for seeding and culturing pulp cells on PGA were optimized. Culturing cells on PGA resulted in a very high cell density tissue with significant collagen deposition. No cell proliferation was observed on alginate, and the growth of cells in collagen gels after 45 days was only moderate. These studies indicate dental pulp-like tissues can be engineered, and this may provide the first step to engineering a complete tooth.     

Regeneration of dentin-pulp complex with cementum and periodontal ligament formation using dental bud cells in gelatin-chondroitin-hyaluronan tri-copolymer scaffold in swine

The purpose of this study is to use a tissue engineering approach for tooth regeneration. The swine dental bud cells (DBCs) were isolated from the developing mandibular teeth, expanded in vitro, and cultured onto cylinder scaffold gelatin-chrondroitin-hyaluronan-tri-copolymer (GCHT). After culturing in vitro, the DBCs/GCHT scaffold was autografted back into the original alveolar socket. Hematoxylin and eosin (H&E) staining combined with immunohistochemical staining were applied for identification of regenerated tooth structure. After 36-week post-transplantation, tooth-like structures, including well-organized dentin-pulp complex, cementum, and periodontal ligament, were evident in situ in two of six experimental animals. The size of the tooth structure (1 x 0.5 x 0.5 cm(3) and 0.5 x 0.5 x 0.5 cm(3) size) appeared to be dictated by the size of the GCHT scaffold (1 x 1 x 1.5 cm(3)). The third swine was demonstrated with irregular dentin-bony like calcified tissue about 1 cm in diameter without organized tooth or periodontal ligament formation. The other three swine in the experimental group showed normal bone formation and no tooth regeneration in the transplantation sites. The successful rate of tooth regeneration from DBCs/GCHT scaffolds' was about 33.3%. In the control group, three swine's molar teeth buds were removed without DBCs/GCHT implantation, the other three swine received GCHT scaffold implants without DBCs. After evaluation, no regenerated tooth was found in the transplantation site of the control group. The current results using DBSs/GCHT scaffold autotransplantation suggest a technical breakthrough for tooth regeneration.     

Expression of neural cell adhesion molecule (NCAM) during the first molar development in the mouse

NCAM, the neural cell adhesion molecule, was immunolocalized in the mandibular first molar tooth germ of the mouse. NCAM was first detected in the tooth germ of the late bud stage, where only the cells in the outer part of the condensed mesenchyme (primitive dental follicle) exhibited faint immunoreactivity. The entire dental follicle was intensely immunostained for NCAM from cap stage to the stage when root formation started. During root formation, NCAM disappeared from the follicular tissue surrounding the cervical root as well as from the part covering the crown top. This loss of NCAM proceeded in the direction of the root apex, but even after the tooth had achieved functional occlusion, NCAM was still expressed by the mesenchymal cells adjacent to the root apex. On the other hand, NCAM was negative in the dental papilla until birth. After birth, NCAM-immunoreactivity appeared in the basal portion of the dental papilla, but this NCAM-positive area gradually diminished in width during the root elongation. Instead, another NCAM-positive zone appeared in the core of the pulp during root formation. Even in the tooth that had already erupted, the pulp core contained cells that were strongly positive for NCAM immunostaining. In addition to its expression in the above two mesenchymal cell lineages, NCAM was transiently expressed by epithelial components of the tooth germ, some of the cells of the dental lamina and the enamel organ. The results suggest that NCAM participates in several processes of tooth development.