ctx ii  (Novus Biologicals)

Journal: Nature Communications

Article Title: Regulation of senescence-associated secretory phenotypes in osteoarthritis by cytosolic UDP-GlcNAc retention and O-GlcNAcylation

doi: 10.1038/s41467-024-55085-1

Figure Lengend Snippet: a Twelve-week-old WT C57BL/6 J mice underwent DMM surgery followed by weekly IA injection of vehicle or thiamet-G. Joint sections were stained with safranin O, fast green, and hematoxylin, with insets magnified below and on the right. b Cartilage destruction and synovial inflammation were scored ( n = 8, 9 respectively). c The percentage of weight placed on the DMM-operated limb versus the contralateral limb of the WT mice injected with vehicle or thiamet-G, determined using a static weight bearing test ( n = 8, 9 respectively). d GSEA of the “Upregulated genes in OA” gene set in chondrocytes treated with bleomycin for 24 h followed by vehicle or ST045849 treatment for five days. e Twelve-week-old WT C57BL/6 J mice received DMM surgery followed by weekly IA injection of vehicle or ST045849 using a hydrogel-based drug delivery system. Joint sections were stained with safranin O, fast green, and hematoxylin, with insets magnified on the right. f – h Immunohistochemistry of ( f ) GATA4 and cartilage matrix neoepitopes (C-telopeptide of type II collagen, CTX-II and aggrecan neoepitope, NITEGE), ( g ) MMP3, MMP13, IL-6, ( h ) senescence markers (p16 INK4a and HMGB1), and HGF in cartilage sections from DMM-operated WT mice followed by IA injection with vehicle or ST045849 ( n = 3). i Cartilage destruction, subchondral bone sclerosis, osteophyte formation, and synovial inflammation were scored ( n = 10). j Percentage of weight placed on the DMM-operated limb versus the contralateral limb of WT mice injected with vehicle or ST045849, determined using a static weight bearing test ( n = 10). k Electronic Von Frey test in ipsilateral hindpaw of sham- or DMM-operated WT mice IA-injected with vehicle or ST045849 using a hydrogel-based drug delivery system ( n = 10). Scale bars: a, e , 200 μm, f – h , 25 μm. b, c, i – k Data represent means ± s.e.m. P values are from two-tailed Mann–Whitney U test ( b, i ), two-tailed t test ( c, j, k ), or one-tailed permutation test corrected for multiple comparisons with FWER ( d ). Source data are provided as a Source Data file.

Article Snippet: Primary antibodies used for immunohistochemical staining were as follows: O-GlcNAc (clone RL2, Abcam, ab2739, 1:50–100), p16 INK4a (Proteintech, 10883-1-AP, 1:100; against human p16 INK4a ), GATA4 (clone G-4, Santa Cruz, sc-25310, 1:50–100), IL-6 (clone 1, Santa Cruz, sc-130326, 1:50–100), MMP3 (clone EP1186Y, Abcam, ab52915, 1:50–100), MMP13 (EP1263Y, Abcam, ab51072, 1:50–100), p16 INK4a (clone EPR20418, Abcam, ab211542, 1:50–100; against mouse p16 INK4a ), HMGB1 (Abcam, ab18256, 1:100), HGF (Novus, AF2207, 1:50), CTX-II (Novus, NBP2-59386, 1:100), NITEGE (clone BC-13, MD Bioproducts, 1042003, 1:100), and ADAMTS5 (Abcam, ab41037, 1:100).

Techniques: Injection, Staining, Immunohistochemistry, Two Tailed Test, MANN-WHITNEY, One-tailed Test