목적 : 각막 상피조직의 손상에 의해 형성되는 미세환경 변화가 중배엽 줄기세포의 특성 혹은 분화에 미치는 영향을 알아보고자 하였다. 대상과 방법 : 각막 상피세포를 단일 층으로 배양한 후, 인위적으로 각막 상피세포 층에 손상을 가하고, 중배엽 줄기세포를 손상부위에 접종하여 공배양하였다. 공배양된 중배엽 줄기세포의 형태학적 변화를 관찰하였다. 중배엽 줄기세포의 각막 간질세포 또는 상피세포로의 분화 여부를 확인하기 위하여 α-smooth muscle actin, keratin-3/-12, E-cadherin의 발현을 면역형광염색을 통해 알아보았다. 결과 : 인위적으로 손상된 각막 상피세포와 공배양한 중배엽 줄기세포는 손상 근접 부위에서 세포 부착이 증진되었고, 접종 6일 후 가지세포와 유사한 형태적 변화를 확인할 수 있었다. 중배엽 줄기세포의 면역형광염색 결과, 중배엽 줄기세포의 표지인자로 알려진 α-smooth muscle actin의 발현이 각막 상피세포의 손상된 접촉 부분에서 현저하게 감소하였으나, 각막 상피세포 표지인자인 keratin-3/-12, E-cadherin의 발현 변화는 확인할 수 없었다. 특히, 각막 상피세포의 표지인자인 keratin-3/-12의 발현이 손상 각막 상피세포와 공배양한 중배엽 줄기세포에서 관찰되었다. 정상 각막 상피세포 층에 중배엽 줄기세포를 접종한 경우, 부착이 진행되지 않았고, α-smooth muscle actin의 발현 변화 또한 나타나지 않았다. 결론 : 손상 각막 상피세포는 중배엽 줄기세포의 형태 변화를 유도하며, α-smooth muscle actin의 발현 감소를 확인하였다. 이를 통해 손상 각막 상피세포가 중배엽 줄기세포의 특성 변화 및 분화에 영향을 미침을 알 수 있었다.

Purpose: To identify the effects of microenvironmental changes caused by human corneal epithelial damages to characteristics or differentiation of human mesenchymal stem cells (hMSCs). Methods: Artificial corneal damage was induced onto a cultured monolayer of human corneal epithelial cells. hMSCs were then co-cultured with damaged human corneal epithelial cells (dIHCE). Morphological changes in the co-cultured hMSCs were observed. To elucidate the differentiation of hMSCs into corneal keratocytes or epithelial cells, the expressions of α-smooth muscle actin, keratin-3/-12, and E-cadherin were confirmed by immunofluorescence. Results: hMSCs co-cultured with dIHCE showed enhanced adherence in the neighborhood of dIHCE and morphological change into dendritic shapes at 6 days post-seeding. Although the expression of α-smooth muscle actin, known as hMSCs marker, significantly decreased at the dIHCE-contacted site of hMSCs; there were no expressional changes on keratin-3/-12 and E-cadherin, the markers of corneal epithelial cells. Interestingly, positive expression of corneal epithelial marker keratin-3/-12 was observed in dIHCE co-cultured hMSCs. hMSCs co-cultured with normal human corneal epithelial cells (nIHCE) were unable to attach, and showed no change in the expression of α-smooth muscle actin. Conclusions: It is proposed that dIHCE causes a morphological change in hMSCs, and decreased expression of α-smooth muscle actin. These results suggest that dIHCE can affect a change in the characteristics and differentiation of hMSCs.

Purpose: To identify the effects of microenvironmental changes caused by human corneal epithelial damages to characteristics or differentiation of human mesenchymal stem cells (hMSCs). Methods: Artificial corneal damage was induced onto a cultured monolayer of human corneal epithelial cells. hMSCs were then co-cultured with damaged human corneal epithelial cells (dIHCE). Morphological changes in the co-cultured hMSCs were observed. To elucidate the differentiation of hMSCs into corneal keratocytes or epithelial cells, the expressions of α-smooth muscle actin, keratin-3/-12, and E-cadherin were confirmed by immunofluorescence. Results: hMSCs co-cultured with dIHCE showed enhanced adherence in the neighborhood of dIHCE and morphological change into dendritic shapes at 6 days post-seeding. Although the expression of α-smooth muscle actin, known as hMSCs marker, significantly decreased at the dIHCE-contacted site of hMSCs; there were no expressional changes on keratin-3/-12 and E-cadherin, the markers of corneal epithelial cells. Interestingly, positive expression of corneal epithelial marker keratin-3/-12 was observed in dIHCE co-cultured hMSCs. hMSCs co-cultured with normal human corneal epithelial cells (nIHCE) were unable to attach, and showed no change in the expression of α-smooth muscle actin. Conclusions: It is proposed that dIHCE causes a morphological change in hMSCs, and decreased expression of α-smooth muscle actin. These results suggest that dIHCE can affect a change in the characteristics and differentiation of hMSCs.

Mesenchymal stem cell, Cornea epithelial cell, Coculture, -smooth muscle actin