Fig. 3

Impact of SLC7A5 intervention either by antibody blocking or siRNA knockdown on the expression levels of MMP3 and MMP13 in FLS from RA patients. a The mRNA expression of MMP13 and MMP3 in RA FLS, incubated with SLC7A5 monoclonal antibody or isotype IgG and stimulated with or without IL-1β. The cells were incubated with SLC7A5 monoclonal antibody or isotype IgG for 4 h and then treated with or without IL-1β (20 ng/mL) for 8 h. The mRNA levels were measured by RT-qPCR (n = 6). b The protein expression of MMP13 and MMP3 in RA FLS, incubated with SLC7A5 monoclonal antibody or isotype IgG and stimulated with or without IL-1β. The cells were incubated with SLC7A5 monoclonal antibody or isotype IgG 4 h and then treated with or without IL-1β (20 ng/mL) for 24 h. The protein levels were detected by Western blotting. The density of MMP13 and MMP3 immune-reactive bands was analyzed by using ACTB expression as a loading control (n = 3). c, d Optimization of the SLC7A5 RNAi efficiency in FLS from RA patients. The cells were transfected with siRNA (NC, siSLC7A5-1 or 2) for 24 h, and the protein and mRNA expression levels were detected by Western blotting (c) and RT-qPCR (d), respectively. e–g The expression of SLC7A5, MMP13, and MMP3 in FLS from RA patients transfected with siRNA (NC or siSLC7A5-2) and treated with or without IL-1β, detected at the mRNA level by RT-qPCR. The cells were first transfected with siRNA (NC or siSLC7A5-2) for 24 h and then treated with or without IL-1β (20 ng/mL) for 8 h without changing the culture medium. h The protein expression of SLC7A5, MMP13, and MMP3 in FLS from RA patients transfected with siRNA (NC or siSLC7A5-2) and treated with or without IL-1β. The cells were first transfected with siRNA (NC or siSLC7A5-2) for 24 h and then treated with or without IL-1β (20 ng/mL) for 24 h further, without changing the culture medium. The density of MMP13 and MMP3 immune-reactive bands was analyzed by using ACTB expression as a loading control (n = 3) (*p < 0.05)