Study On The Therapeutic Effect Of Yupingfeng San On Delayed-Type Hypersensitivity Mice And Its Mechanisms Based On The Pharmacology

Delayed-type hyper sensitivity (DTH) is the T-cell-mediated immunity response, T helper (Th) 1-type cells are major effector cells. Inflammation induced by DTH can cause tissue injury and monocytic infiltration. DTH usually take 24 to 48 hours to develop. DTH is one of the important challenges for public health. An estimated 15-20% of the general population suffers from DTH. Yupingfeng San is a Chinese medical formula that consists of Radix Astragali (RA, Huangqi), Rhizoma Atractylodis Macrocephalae (RAM, Baizhu) and Radix Saposhnikoviae (RS, Fangfeng). Yupingfeng San could regular immune system including specific immunity and nonspecific immunity. A number of researches have proved that Yupingfeng San could be applied in Delayed-type hypersensitivity, but its mechanism is rarely reported.Firstly, we constructed DTH-related proteins-proteins interactions network and YPFS bioactive ingredients-proteins interactions network. Then, we merged the two networks above mentioned and constructed DTH-related YPFS bioactive ingredients-protential targets interactions network. The network pharmacology analysis revealed that HSP90AA1, HSD17B1, ESR, NR3C1 and TTR could be key node proteins. The potential targets were significantly enriched in MAPK and NF-kB signaling pathway. The degree of HSP90AA1 was the highest, which indicated that the HSP90AA1 was a important potential targets. We further validated the associations of YPFS bioactive ingredients with potential targets by molecular docking. The results displayed that the hydroxyl group on the TRP139 of HPS90 could be the important hydrogen bridge donor and the phenyl ring on the PHE138 could be the important hydrophobic group.Next, 1-fluoro-2,4-dinitrobenzene (DNFB) was used to induce model of mice DTH, the level of sera IFN-7, the levels of spleen IL-1β and HSP90AA1 were tested to evaluate the pharmacological efficacy and potential targets of YPFS effective constituent in treating DTH. IFN-y, IL-1β and HSP90AA1 levels were increased on DTH mice. YPFS effective constituent significantly reduced the levels of IFN-y, IL-1β and HSP90AA1. This implied that HSP90AA1 could be the potential target of YPFS bioactive ingredients agaist DTH.Lastly, the protein levels of IRAK-1 and HSP90AA1 were increased in RAW 264.7 cells induced by IL-1β. When RAW 264.7 cells were stimulated with IL-ip for 48h, the production of IRAK-1 and HSP90AA1 were in the peak. RAW 264.7 cells were stimulated with 15ng/ml IL-1β for 48h, Prim-O-glucosylcimifugin (Pg) at 25mg/L reduced the levels of IRAK-1 and HSP90AA1 by western blotting. The combination of Astragaloside IV (AsIV), AtI, Atractylenolide I (AtI) and Prim-O-glucosylcimifugin (Pg) reduced the levels of IRAK-1 and HSP90AA1 with a final concentriation of 5mg/L AsIV,3.33mg/L At1 and 10mg/L Pg. This implied that HSP90 could regulate the stability of IRAK-1 in IL-1β/IRAK signaling pathway.So we inferred that HSP90AA1 could be the potential target of YPFS bioactive ingredients agaist DTH by regulate the stability of IRAK-1.