Multi-layered periodontal pocket epithelium reconstituted in vitro: Histology and cytokeratin profiles
Multi-layered periodontal pocket epithelium reconstituted in vitro: Histology and cytokeratin profiles
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DOI：
10.1902/jop.1999.70.6.668
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发表时间：
1999-06-01
影响因子：
4.3
通讯作者：
Quirynen, M
中科院分区：
文献类型：
--
作者：
Papaioannou, W;Cassiman, JJ;Quirynen, M
背景资料：为了研究个体间的差异，在细菌粘附/牙周组织的侵袭，在体外培养多层口袋上皮细胞无饲养层或基质的等价物（包括其细胞角蛋白的档案的评价）的模型developed.Methods：口袋上皮细胞收集和生长，直到汇合在猎鹰烧瓶中使用角质形成细胞无血清培养基（KSFM），没有饲养层。在第二次传代中，使用透明聚酯（transwell-clear [TCL]）或胶原（transwell-coI [TCO]）膜作为培养表面，在2隔室系统中使口腔角质形成细胞再生长。第一周后，将钙浓度升高至1.2 mM，并在一半的威尔斯孔中，向KSFM中补充10%胎牛血清（FCS）。组织学和免疫组织化学进行后1，2，和3周的额外growth.Results：在一般情况下，所有条件下导致在结构化的上皮细胞组成的3至5层，但重要的差异之间的膜类型和媒体之间观察。CK 4很少表达，仅轻度表达，而CK 18和19（结合上皮的特征）在较旧（2周和3周）培养物中表达非常强烈。CK 13和14（任何可分层上皮细胞的特征）也倾向于随时间增加;在KSFM与FCS中，CK 13似乎更强，而CK 14则相反。TCL/KSFM +10%FCS的组合创建的多层最像一个交界上皮，而生长在TCO没有FCS模仿沟epithelium.Conclusions：它似乎可以创建一个组织型培养类似于牙周袋或交界上皮，而不使用基质等价物或饲养层，使这种方法更繁琐。这种多层培养提供了一个模型，以研究袋上皮的渗透性和细菌在明确定义的环境条件下的粘附和渗透。
Background: In order to study inter-individual differences in bacterial adhesion/invasion of periodontal tissues, an in vitro model for culturing multi-layered pocket epithelium without feeder layers or stromal equivalents (including the evaluation of their cytokeratin profiles) was developed.Methods: Pocket epithelium was collected and grown until confluent in Falcon flasks using keratinocyte-serum free medium (KSFM), without a feeder layer. In the second passage, oral keratinocytes were re-grown in a 2 compartment system using either a clear polyester (transwell-clear [TCL]) or a collagen (transwell-coI [TCO]) membrane as culture surface. After the first week, the calcium concentration was raised to 1.2 mM and in half the wells, the KSFM was supplemented with 10% fetal calf serum (FCS). Histology and immunohistochemistry were performed after 1, 2, and 3 weeks of additional growth.Results: In general, all conditions resulted in a structured epithelium consisting of 3 to 5 layers, but important differences were observed between the membrane types and between the media. CK4 was rarely and only lightly expressed while CK18 and 19 (characteristic of junctional epithelium) were very strongly expressed in the older (2 and 3 weeks) cultures. CK13 and 14 (characteristic of any stratifiable epithelial cell) also tended to increase over time; CK13 seemed to be stronger in KSFM with FCS while the contrary was true for CK14. The multi-layer created by the combination TCL/KSFM + 10% FCS resembled a junctional epithelium most, while that grown on TCO without FCS mimicked the sulcular epithelium.Conclusions: It seems possible to create a histiotypic culture resembling either periodontal pocket or junctional epithelium without the use of stromal equivalents or feeder layers which make this approach more cumbersome. This multi-layered culture offers a model to investigate the permeability of pocket epithelium and the adhesion and penetration of bacteria under well-defined environmental conditions.