A novel simple,sensitive fluorescence immunosensing method based on aptamer-plasmid complex amplification was developed. This method utilized the specific recognition between antibody and antigen as well as aptamer-plasmid complex and the intercalation of fluorescence dye SYBR Green Ⅰ in the groove of duplex plasmid DNA in detection of Platelet-Derived Growth Factor BB (PDGF-BB). The immunoassay was performed in the microtiter wells in which rabbit anti PDGF-BB antibody was immobilized. The PDGF BB analyte was captured by the primary antibody and then sandwiched by the aptamer-plasmid DNA complex. The introduction of fluorescence dye SYBR Green Ⅰ allows for the detection of the sandwiched immunocomplex of antibody/anigen/aptamer-plasmid complex. Under the optimized conditions of salt concentration,ratio of aptamer to PUC19,and SYBR Green Ⅰ concentration,the proposed method offers a linear detection range from 0.2 μg/L to 200 μg/L with a detection limit of 0.1μg/L.

Osteoarthritis (OA), in which M1 macrophage polarization in the synovium exacerbates disease progression, is a major cause of cartilage degeneration and functional disabilities. Therapeutic strategies of OA designed to interfere with the polarization of macrophages have rarely been reported. Here, we report that SHP099, as an allosteric inhibitor of src-homology 2-containing protein tyrosine phosphatase 2 (SHP2), attenuated osteoarthritis progression by inhibiting M1 macrophage polarization. We demonstrated that M1 macrophage polarization was accompanied by the overexpression of SHP2 in the synovial tissues of OA patients and OA model mice. Compared to wild-type (WT) mice, myeloid lineage conditional Shp2 knockout (cKO) mice showed decreased M1 macrophage polarization and attenuated severity of synovitis, an elevated expression of cartilage phenotype protein collagen II (COL2), and a decreased expression of cartilage degradation markers collagen X (COL10) and matrix metalloproteinase 3 (MMP3) in OA cartilage. Further mechanistic analysis showed thatSHP099 inhibited lipopolysaccharide (LPS)-induced Toll-like receptor (TLR) signaling mediated by nuclear factor kappa B (NF-κB) and PI3K-AKT signaling. Moreover, intra-articular injection of SHP099 also significantly attenuated OA progression, including joint synovitis and cartilage damage. These results indicated that allosteric inhibition of SHP2 might be a promising therapeutic strategy for the treatment of OA.