Expression and role of Notch signalling in the regeneration of rat tracheal epithelium.

OBJECTIVES

This study is to explore the role of Notch signalling during the regeneration of rat tracheal epithelium after injury induced by 5-fluorouracil (5-FU).

MATERIALS AND METHODS

We developed an ex vivo model of rat tracheal epithelial regeneration using 5-FU to induce injury. Expression levels of members of the Notch signalling pathway, ABCG2, CK19, and proliferating cell nuclear antigen (PCNA) were examined by reverse transcription-polymerase chain reaction, Western blot, and immunofluorescence. One group of tracheas were cultured in the medium with a gamma-secretase inhibitor or Jag-1 peptide after 5-FU treatment and another group were pre-treated with the gamma-secretase inhibitor or Jag-1 peptide before 5-FU treatment. The expression changes of ABCG2, CK19, and PCNA were examined by Western blot or immunofluorescence and the morphologic changes were observed by haematoxylin and eosin stain during the recovery process.

RESULTS

Expression levels of Notch3, Jagged1, and Hey1 were increased in rat tracheal epithelial cells after treatment with 5-FU. During injury recovery, disruption of Notch signalling by treatment with the gamma-secretase inhibitor reduced expression of ABCG2 and PCNA, but promoted expression of CK19, while persistent activation of Notch signalling promoted expression of ABCG2 and PCNA, but reduced expression of CK19. Under both conditions, recovery from injury was reduced. However, blocking Notch signalling prior to 5-FU treatment led to the complete blockage of recovery, while activating Notch signalling before 5-FU treatment promoted recovery.

CONCLUSIONS

During tracheal epithelial regeneration, Notch signalling maintains an undifferentiated state and promotes proliferation among a population of tracheal epithelial cells.