Polisetti Naresh, Agarwal Prasoon, Khan Imran, Kondaiah Paturu, Sangwan Virender S, Vemuganti Geeta K
C-TRACER, Hyderabad Eye Research Foundation, LV Prasad Eye Institute, Hyderabad, India.
Mol Vis. 2010 Jul 6;16:1227-40.
Limbal stem cell deficiency is a challenging clinical problem and the current treatment involves replenishing the depleted limbal stem cell (LSC) pool by either limbal tissue transplantation or use of cultivated limbal epithelial cells (LEC). Our experience of cultivating the LEC on denuded human amniotic membrane using a feeder cell free method, led to identification of mesenchymal cells of limbus (MC-L), which showed phenotypic resemblance to bone marrow derived mesenchymal stem cells (MSC-BM). To understand the transcriptional profile of these cells, microarray experiments were carried out.
RNA was isolated from cultured LEC, MC-L and MSC-BM and microarray experiments were carried out by using Agilent chip (4x44 k). The microarray data was validated by using Realtime and semiquntitative reverse transcription polymerase chain reaction.
The microarray analysis revealed specific gene signature of LEC and MC-L, and also their complementary role related to cytokine and growth factor profile, thus supporting the nurturing roles of the MC-L. We have also observed similar and differential gene expression between MC-L and MSC-BM.
This study represents the first extensive gene expression analysis of limbal explant culture derived epithelial and mesenchymal cells and as such reveals new insight into the biology, ontogeny, and in vivo function of these cells.
角膜缘干细胞缺乏是一个具有挑战性的临床问题，目前的治疗方法包括通过角膜缘组织移植或使用培养的角膜缘上皮细胞（LEC）来补充耗尽的角膜缘干细胞（LSC）库。我们使用无饲养层细胞方法在脱细胞人羊膜上培养LEC的经验，导致鉴定出角膜缘间充质细胞（MC-L），其表现出与骨髓来源的间充质干细胞（MSC-BM）相似的表型。为了了解这些细胞的转录谱，进行了微阵列实验。
从培养的LEC、MC-L和MSC-BM中分离RNA，并使用安捷伦芯片（4x44 k）进行微阵列实验。通过实时和半定量逆转录聚合酶链反应验证微阵列数据。
微阵列分析揭示了LEC和MC-L的特定基因特征，以及它们在细胞因子和生长因子谱方面的互补作用，从而支持了MC-L的滋养作用。我们还观察到MC-L和MSC-BM之间相似和差异的基因表达。
本研究首次对角膜缘外植体培养衍生的上皮细胞和间充质细胞进行了广泛的基因表达分析，从而揭示了这些细胞的生物学、个体发生和体内功能的新见解。
