Artesunate interacts with Vitamin D receptor to reverse mouse model of sepsis-induced immunosuppression via enhancing autophagy

Shenglan Shang; Jiaqi Wu; Xiaoli Li; Xin Liu; Pan Li; Chunli Zheng; Yonghua Wang; Songqing Liu; Jiang Zheng; Hong Zhou

Microlon ELISA Plates were coated with antibody against VDR (anti-VDR antibody) (Cell Signaling Technology, Boston, MA, USA) overnight at 4 ï‚°C, followed by three washes with PBS. The cell lysate containing the total proteins, including the VDR protein, were added into the plates. After incubating at 37 ï‚°C for 2 h, the VDR protein was captured by anti-VDR antibody coated on the plates. Subsequently, the plates were washed three times and V D3 (Selleck, Houston, TX, USA) was added. After incubating at 37 ï‚°C for 2 h, the plates were washed three times and fluorophore-conjugated AS (designed and synthesized by Chongqing Canbipharma Technology, Chongqing, China) was added. After incubating at 37 ï‚°C for 2 h, the plates were washed three times and the fluorescence intensity was observed using a Spectra Max i3x Multifunctional enzyme marker (Molecular Devices, USA) with an excitation wavelength of 360 nm and an emission wavelength of 455 nm. AS with fluorophore 12-(7-oxycoumarinyl-ethoxy) dihydro-artemisinin was named AS I, and AS with 12-(-1H-benzo [de] isoquinoline-1, 3(2H)-dione-2-ethoxy) dihydro-artemisinin was named AS II.