POS0401 THE APPLICATION OF MTX-LOADING DNA TETRAHEDRON IN TREATING COLLAGEN-INDUCED ARTHRITIS MICE VIA REGULATING MACROPHAGE

Rheumatoid arthritis (RA) is a systemic autoimmune disease which mainly affect joints. [1]. Macrophages often infiltrate in the inflammatory joints. Activated macrophages release TNF-α, IL-1β to accelerate tissue damage, is one of the most important targets for RA intervention. The traditional drugs currently used commonly have some disadvantages cannot be bypassed[2], while DNA nanostructure is a new type of drugs have precise design, and likewise takes biological effect together[3]. We synthesized a DNA tetrahedron loaded with MTX and conjugated with HA which targeted to macrophage.To verify whether MTX-loading DNA tetrahedron can regulate the apoptosis and polarization of macrophage and finally improve the condition of CIA model mice by while decrease the side effect of MTX.DNA TET was synthesized by mixing signal strand DNA in TM buffer and heated to 95 °C for 10 min, then cooling to 4 °C. Electrophoresis was applied to confirm the formation of TET. The absorbance of MTX solution was detected by microplate reader to analyze the loading efficiency of MTX into TET. Fluorescence microscope was used to observe the intake of TET into cells. CCK8 experiment was applied to measure the vitality of cells. Flow cytometry was used to detect the apoptosis and polarization. CIA model was established based on DBA/1 mice. Mice were randomly divided into five groups: normal group injected with NS; after established CIA model, CIA group injected with NS, MTX group injected with MTX solution, MTX-TET group injected with MTX-TET NP.We synthesized DNA tetrahedron(A) and used 8% PAGE electrophoresis to confirmed the successfully synthesis(B). Then We found that when TET concentration fixed, the loading MTX concentration gradually increased and saturated at 190μM(C). While completely loading needed at least 4 hours(D). Fluorescence showed that single DNA strand cannot be taken by RAW, while TET can be easily taken by RAW(E). CCK8 showed that empty TET had no obvious effect on cells, while MTX and MTX-TET with equivalent concentration can obviously suppress the vitality(F). Similarly, the apoptosis trial showed that TET can slightly decrease the apoptosis of RAW, MTX and MTX-TET can significantly promote the apoptosis(G). Flow cytometry showed that the MTX-TET can decrease the expression of M1 marker CD80 (H). At last, we treat mice with NS, TET, MTX and MTX-TET once a week after CIA model established, and found that TET have no significantly effect on mice, while MTX and MTX-TET can alleviate the inflammation symptom of paws(I). Conclusions: We synthesized MTX-loading DNA tetrahedron conjugated with HA, and found that the MTX-TET NP have the excellent ability of promote RAW apoptosis and relieve proinflammatory M1 polarization. while also can alleviate the symptom of CIA mice. [1]Aletaha D, Smolen JS: Diagnosis and Management of Rheumatoid Arthritis: A Review. JAMA 2018, 320(13):1360-1372. [2]Smolen JS, Aletaha D, McInnes IB: Rheumatoid arthritis. Lancet 2016, 388(10055):2023-2038. [3]Hu Q, Li H, Wang L, Gu H, Fan C: DNA Nanotechnology-Enabled Drug Delivery Systems. Chem Rev 2019, 119(10):6459-6506.Figure 1.None declared