The regeneration/ proliferation potential of mouse lacrimal gland epithelium following duct ligation procedure

Abstract

Purpose: We previously reported that adult mouse lacrimal glands (LGs) have a strong regeneration potency after atrophy by duct ligation as a result of inflammation. The purpose of this study is to observe regeneration and proliferation of adult mouse lacrimal gland epithelium in vivo and in vitro.

Methods: We used previously reported the injury model. Right LGs were subjected to the main excretory duct-ligation procedure, and right side skin cut as a sham, LGs were removed at day 3,7,14 after operation. The Tissue was prepared for RNA extraction, primary culture and immunohistochemical analysis. Isolated lacrimal gland epithelial cells were seeded onto tissue culture treated wells in Cnt-07 medium and Matrigel to assess Colony-forming efficiency (CFE) and Sphere-forming efficiency (SFE). Cultured cells were evaluated for the expression of K14, α-SMA, P63α and Nestin.

Results: Real-time PCR results showed that the expression of α-SMA was significantly higher at 3 days after injury compared with other groups (p<0.01). The expression of AQP5 at 3, 7 and 14 days after injury was significantly decreased compare with the sham (p<0.01). In the morphology study, the number of Nestin positive cells was significantly increased at 3, 7, and 14 days after injury. The expression of K14 and α-SMA showed strong staining at 7 and 14 days after injury. Cultured cells at7 and 14 days after injury formed a monolayer faster than control, and CFE at 14 days after injury was significantly higher than 3 and 7 days (p<0.01) SFE at 14 days after injury showed significantly higher compared with 3 and 7 days (p<0.01). The immunocytochemistry showed the α-SMA and Nestin positive cells had a tendency to increase at 7 and 14 days after injury.

Conclusions: This study showed that, after injury the LG suffered a pattern of proliferation. This high proliferation population could be isolated and proliferate in-vitro with the characteristic of duct-like cells.

Keywords: 576 lacrimal gland • 486 cornea: tears/tear film/dry eye • 687 regeneration