Alexa fluor 594 labelled goat anti rabbit igg h l

Melatonin, 3‐methyladenine (3‐MA), TBHP and the type II collagenases were from Sigma‐Aldrich (St. Louis, MO, USA). The Sirt1, RUNX‐2, p62, β‐actin, OCN primary antibodies were acquired from Abcam (Cambridge, UK). The LC‐3, Beclin‐1, p‐p53, p53, p21, cleaved caspase‐3, Bax and Bcl‐2 antibodies were obtained from Cell Signaling Technology (Danvers, MA, USA). Alexa Fluor^® 488‐ and Alexa Fluor^® 594 labelled goat anti‐rabbit IgG (H+L) secondary antibodies were purchased from Jackson ImmunoResearch (West Grove, PA, USA). The 4′,6‐diamidino‐2‐phenylindole (DAPI) was obtained from Beyotime (Shanghai, China). The cell culture reagents were purchased from Gibco (Grand Island, NY, USA), the 0.1% Alcian Blue 8GX was from Sigma‐Aldrich, and the 6‐chloro‐2,3,4,9‐tetrahydro‐1H‐carbazole‐1‐carboxamide (EX‐527) was from Tocris Bioscience (Ellisville, MO, USA).

Astilbin (purity > 98%) was purchased from BOC Sciences (New York, USA). Cell Counting Kit‐8 (CCK‐8) was purchased from Dojindo (Kumamoto, Japan). LPS, type II collagenase and dimethylsulphoxide (DMSO) were obtained from Sigma‐Aldrich (St Louis, MO, USA). The primary antibodies against TLR4, MyD88, TRAF6, Lamin B1 and GADPH were obtained from Abcam (Cambridge, UK); iNOS antibodies were obtained from Sigma‐Aldrich (St Louis, MO, USA); and MD2 antibody was purchased from eBioscience (eBioscience, SanDiego, CA). Antibodies against COX‐2, p65 and IκBα were purchased from Cell Signaling Technology (Danvers, MA, USA); anti‐IRAK1, goat anti‐rabbit and antimouse IgG‐HRP were from Bioworld (OH, USA). Recombinant human MD2 and TLR4 protein (rhMD2 and rhTLR4) were purchased from R&D Systems (Minneapolis, MN, USA). Biotin‐labelled LPS (Biotin‐LPS) was obtained from InvivoGen (San Diego, CA, USA). Alexa Fluor^®488 labelled and Alexa Fluor^®594 labelled Goat Anti‐Rabbit IgG (H + L) the second antibody were purchased from Jackson ImmunoResearch (West Grove, PA, USA). 4′,6‐Diamidino‐2‐phenylindole (DAPI) was purchased from Beyotime (Shanghai, China), and cell culture reagents were obtained from Gibco (Grand Island, NY, USA).