Effect of Dlk1 and Jagged1 gene on the proliferation and transdifferentiation of the primary type II alveolar epithelial cells

Objective To study the effects of the Notch ligands Dlk1 and recombinant human nucleu factor-κB(Jagged1) on the proliferation and transdifferentiation of the type Ⅱ alveolar epithelial cells when the Notch signaling pathway activated. Methods The primary type Ⅱ alveolar epithelial cells (AECⅡ) cultured with recombinant protein Dlk1 and recombinant human nucleu factor-κB(rhNF-κB)(activator of Jagged1), respectively, and then cultured with DMEM(containing 120 mL/L FBS) as controls.Proliferation and differentiation conditions of the AECⅡ were observed at 48 h, 72 h, 96 h time point by the light microscope and electron microscopes separately.Cell number was counted with hemacytometer; the proliferation rate was measured by methyl thiazolyl tetrazolium(MTT); Immunofluo-rescence double standard method was used to detect the AEC Ⅱ specific surfactant protein C (SP-C) and AECⅠ specific protein aquaporin5(AQP5); the expression of SP-C, AQP5, Dlk1, Jagged1, Notch1 and Hes1 mRNA were detected by real time-PCR. Results The cell population and proliferation: compared with control group, AECⅡ proliferation was promoted in the Dlk1 group [cell numbers (×109/L) 9.05±0.45 vs 7.95±0.65, 11.68±0.43 vs 8.68±0.52, 11.55±0.17 vs 8.73±0.48, all P<0.05; MTT results (value A) 0.699±0.050 vs 0.462±0.080, 0.912±0.080 vs 0.535±0.040, 0.726±0.050 vs 0.540±0.020, all P<0.05] and decelerated AECⅡ transdifferentiation into AECⅠ; while AECⅡ proliferation was inhibited in rhNF-κB group [cell numbers (×109/L) 4.95±0.33 vs 7.95±0.65, 4.73±0.71 vs 8.68±0.52, 4.04±0.11 vs 8.73±0.48, all P<0.05; MTT results (value A) 0.398±0.030 vs 0.462±0.080, 0.402±0.070 vs 0.535±0.040, 0.380±0.110 vs 0.540±0.020, all P<0.05] and accelerated AECⅡ transdifferentiation into AECⅠ.One-Way ANOVA showed that the difference among the 3 groups had statistical significance (cell numbers: F=486.73, P=0.02; cell proliferation: F=37.16, P=0.02). The mRNA expression: compared with control group, the expression of SP-C mRNA of Dlk1 group was significantly higher(P<0.05) while the expression of AQP5 mRNA was remarkably lower and delayed(P<0.05), the expression of Jagged1 mRNA was weak or little, Dlk1 and Notch1 mRNA were up-regulated (P<0.05), and the Hes1 mRNA was reduced(P<0.05); the expression of SP-C mRNA of rhNF-κB group was significantly reduced(P<0.05), while the AQP5 mRNA expressed ahead of time and increased (P<0.05), Jagged1, Hes1 and Notch1 mRNA were higher(P<0.05), and the Dlk1 mRNA was weak.One-Way ANOVA showed that the difference in the expressions of SP-C, AQP5, Dlk1, Jagged1, Hes1 and Notch1 mRNA among the 3 groups had statistical significance (F=96.80, P=0.01; F=82.55, P=0.01; F=269.80, P=0.00; F=312.34, P=0.00; F=169.17, P=0.01; F=19.85, P=0.02). Conclusions There are varied effects on proliferation and differentiation of the AECⅡ when the Notch signaling is activated by different ligands: Dlk1 promoted proliferation and inhibited differentiation, while Jagged1 inhibited proliferation and promoted transdifferentiation. Key words: Type Ⅱ alveolar epithelial cells; Notch; Dlk1; Jagged1; Newborn respiratory distress syndrome