Safranin o fast green s o

After micro‐CT analysis, the knee joints of mice were soaked in a 10% EDTA (#1340, Biofroxx) solution for decalcification. The knee joint embedded in the paraffin block was cut into continuous coronal slides (5 µm thick) using a microtome (Thermo, Germany). The slices were detected by Safranin‐O/fast green (S.O.) (#G1371, Solarbio), haematoxylin (H&E) (#C0105M, Beyotime) staining, and Alcian blue (#G1560, Solarbio) staining to observe the integrity and thickness of articular cartilage. The severity of OA was assessed using the OARSI scoring system (0‐6 scale) by blinded observers. The highest OARSI score for each section was recorded and the average of all scores was calculated.

The mouse knee joints were harvested and decalcified by EDTA with a concentration of 10% (#1340, Biofroxx, Germany) solution, then were embedded in paraffin blocks. Following a previously reported procedure [35 (link)], mouse knee joints were prepared into continuous 5 μm coronal slides by a microtome (Thermo, Germany). After selecting every fifth of 2 slides, Safranin-O/fast green (S.O.) (#G1371, Solarbio) and H&E (#C0105S, Beyotime) staining were performed to assess the synovitis and cartilage degradation [36 ]. The synovitis score (0-3) and OARSI grading system (0-6) were used to assess inflammation and cartilage degeneration by 2 blinded observers [37 (link)]. The highest synovitis score and OARSI score were recorded and the averages of them were calculated. Moreover, H&E (#C0105S, Beyotime) staining of major organs (heart, liver, spleen, lung, and kidney) slides was performed to evaluate the biocompatibility of MNPs and MNPs-TRPV1 in vivo. The ROS levels and dead chondrocytes were evaluated by the tissue ROS detection assay (#BB-460522, Bestbio, China) and Fluorescein (FITC) Tunel Cell Apoptosis Detection Kit (#G1501, Servicebio) based on the manufacturer’s instructions, respectively.