Probing into the Mechanism of Alkaline Citrus Extract Promoted Apoptosis in Pulmonary Fibroblasts of Bleomycin-Induced Pulmonary Fibrosis Mice

الملخص EN

We extracted the primary pulmonary fibroblasts of the normal and bleomycin-induced pulmonary fibrosis mice and investigated the functioning mechanism of citrus alkaline extract (CAE) in the induction of pulmonary fibroblast apoptosis.

The expression intensity of vimentin of the pulmonary fibroblasts in the model mice was higher than that in the normal mice.

Meanwhile, the positive expression rate and expression intensity of alpha smooth muscle actin (α-SMA) of the pulmonary fibroblasts in the model mice were higher than those in the normal mice.

Results of MTT showed that pulmonary fibroblast activity of the normal and model mice has been significantly inhibited by CAE in a concentration-dependent manner.

The results of flow cytometer analysis showed that the proportion of pulmonary fibroblast apoptosis in the model mice has been profoundly increased by CAE treatment in a dosage-dependent manner.

Besides we found that the expression of Cleaved-Caspase 3, Cleaved-Caspase 8, Cleaved-poly-ADP-ribose polymerase (Cleaved-PARP), and Fas and Fas Ligand (FasL) was markedly increased after CAE treatment.

A further study showed that the expression of Cyclooxygenase-2 (COX-2) and prostaglandin E receptor 2 (EP2) was dependant on the concentration of CAE, indicating that CAE-regulated receptor apoptosis of Fas was probably related to COX-2.

The results of fluorescence detection of oxidative stress showed that the level of oxidative stress was significantly increased after CAE treatment.

Furthermore, the results of Western Blot showed that the phosphorylation level of p38 (p-p38) was markedly increased, suggesting that CAE probably has regulated COX-2 through increased p-p38 following oxidative stress.

Our results therefore suggest that CAE can effectively induce pulmonary fibroblast apoptosis of the normal and model mice, and its functioning mechanism is probably related to the p38/COX-2/Fas signaling pathway regulated by oxidative stress.