RESUMO
Aim To predict the potential mechanism of ophiopogonin D (OPD) against pulmonary fibrosis by network pharmacology, and further verify it by experiment in vivo. Methods This study found that ophiopogon was the most frequently used drug in the treatment of pulmonary fibrosis with deficiency of Qi and Yin through data mining. In order to explore its material basis, network pharmacology analysis was carried out. A model of pulmonary fibrosis was established by bleomycin, and different concentrations of ophiopogonin D were administered to verify the results of the pharmacological network. Results Firstly, through network pharmacology analysis, it was found that mitophagy might be the potential target for ophiopogon to exert anti-pulmonary fibrosis effect. Meanwhile, network topology analysis showed that OPD had the greatest relationship with mitophagy. Animal experiments showed that OPD could relieve pulmonary fibrosis and reduce collagen deposition in mice. At the same time, the detection of mitophagy related proteins showed that the compound could increase the expression of PINK1 and Parkin proteins, reduce the content of P62 protein in lung tissue, and reduce the intracellular ROS level. Conclusions OPD can improve mitochondrial function and play an anti-pulmonary fibrosis role by promoting PINKl/Parkin dependent mitophagy in lung tissue.
RESUMO
The full length of chicken interferon alpha (ChIFN-alpha) gene was amplified by the polymerase chain reaction (PCR) from total liver genome of Sanhuang meat-chicken and sequenced. The amplified gene was about 582bp. The coding region for mature protein (489bp) was subcloned into pET-28a(+). The recombinant plasmid pET-28a(+)-IFNalpha was identified by enzyme digestion and DNA sequencing. Data of SDS-PAGE and Western-blot indicated that a 22kD fusion protein was expressed in the form of inclusion bodies with good immunity. The purity of inclusion bodies was above 70% and that of protein purified by nickel affinity chromatography was 95%. The recombinant protein could inhibit H9N2 avian influenza virus (H9N2 AIV) replication on chick embryo fibroblast. 2 microg of recombinant IFN-alpha could completely protect Chick embryo from H9N2 AIV infection. The recombinant IFN-alpha can also delay Newcastle disease virus (NDV) replication on chick embryo for 12 approximately 48h. Chicken administered recombinant IFN-alpha can resist the H9N2 AIV infection. The bioactivities of recombinant IFN-alpha purified by affinity chromatograph were 20 times higher than that of inclusion bodies.