The Effect Of CTNNAL-1on The Adhesion Of Human Bronchial Epithelial Cells

Numerous evidences have confirmed that adhesion molecules contributed to airway inflammation and damage repair. The adhesion molecules expressed by HBECs play an important role in the response to stress of the airway. The adhesion molecules maintain the structural integrity of epithelium, and involved in antioxidant, proliferation, migration, and damage repair of cells by the structural adhesion with matrix; they affect the inflammatory adhesion of HBECs and leukocyte so as to activate and transmit inflammation or stress signals.Our previous study indicated catenin alpha-like1(CTNNAL1), a member of catinin family, was downregulated in human peripheral blood and bronchial mucosa of asthma patients. This indicated that CTNNAL1was associated with the onset of asthma. But expression of CTNNAL1was significantly increased in animal models of acute stress and in wounded edge of the cultured human bronchial epithelial cells (HBECs) involving in the stress response of the airway. This phenomenon prompted that CTNNAL may be related with the stress response of the airway and participate in maintaining the integrity of the airway epithelium, whose downregulation may lead to epithelial functional defects and completeness of destruction. The aim of the present study is to observe the effect of CTNNAL1on adhesion of HBECs and explore its significance in the stress response of the airway.Objective:The aim of the present study is to investigate the effect of CTNNAL1on proliferation and adhesion, as well as on expression of some adhesion molecules in HBECs, preliminarily explore the significance of CTNNAL1in the stress response of the airway.Methods:(1) Construct pcDNA3.1/CTNNAL1over-expression plasmid and pGCU6/Neo/RFP/CTNNAL1silenced plasmid. Identify the efficiency of recombinant plasmid pcDNA3.1(-)-cyc-his/CTNNAL1and pGCU6/Neo/RFP/CTNNAL1silenced plasmid by Real-time PCR and flow cytometry (FCM).(2) Use FCM to detect cell cycle of transfected HBECs marked by PI which is a kind of cell nuclear fluorescent dye.(3) The adhesion rate of HBECs on rat-tail collagen was detected by MTT.(4) FCM was used to detect the adhesion of HBECs and human leukocyte whose CD45molecule was tagged by FITC.(5) The expression of the adhesion molecules E-cadherin and intercellular adhesion molecular1(ICAM-1) were observed by RT-PCR.(6) ELISA was used to detect the secretory volume of different HBECs lines. Results:(1) We successfully constructed the over-expression recombinant plasmid pcDNA3.1(-)-cyc-his/CTNNALl and the silenced plasmid pGCsilencerU6/Neo/RFP/CTNNAL1-RNAi, and transfected them into HBECs to establish the stably transfected cell lines. The expression of CTNNAL1mRNA and protein were significantly increased in pcDNA3.1/CTNNAL1stably transfected HBECs and were significantly decreased in pGCU6/Neo/RFP/CTNNAL1stably transfected HBECs.(2) The over-expression of CTNNAL1could promote the proliferation of HBECs, increase (G2+S)/G1phase of cell cycle and199.84%.(3) The adhesion ability of HBECs with rat tail collagen was promoted by the over-expression of CTNNAL1, and there were statistically significance at30min and120min.(4) The over-expression of CTNNAL1could reduce the adhesion percentage of HBECs with human total leukocyte by35.3%.(5) The mRNA expression of E-cadherin was increased56.5%while the mRNA expression of ICAM-1was reduced90.4%by the over-expression of CTNNAL1. Silenced CTNNAL1reduced the expression of E-cadherin (59.9%).(6) The CTNNAL1lessened the secretion of IL-8(53.87%).Conclusions: We successfully constructed CTNNAL1recombinant plasmids, and established there stably transfected cell lines. CTNNAL1can significantly promote the proliferation and the adhesion ability with matrix of HBECs. The mRNA expression of E-cadherin was increased by the over-expression of CTNNAL1. However, CTNNAL1weakened the inflammatory adhesion ability of HBECs with human leukocyte, and simultaneously decreased the expression of ICAM-1and the secretion of IL-8.