Fig. 4

Decreased RPS17 in monocytic OCPs from AS patients. a GSVA scores in several functions from monocyte subsets 0/1/2/5/7/9. Rows were normalized and clustered, and the transition of blue-white-red indicates an increase in the scores. b, c Histograms (red indicates upregulation while blue indicates downregulation) and heatmap showing DEGs for different translation levels in CD14⁺CD16⁻ monocytes between AS patients and HDs revealed by Ribo-seq. d Heatmap (left) showing differentially expressed sORFs and histograms (right; red indicates upregulation while blue indicates downregulation) showing differentially expressed dORFs (originating from 3’-UTR region) between two groups. e The expression distribution of RPS17 and CD14 across four groups. Circle size indicates PCT and color depth indicates average levels. f Western-blotting analysis of CD14 and RPS17 for CD14⁺CD16⁻ monocytes from four groups of subjects (n = 6/group; each 2 samples presented in one group). g H&E, SO-FG and IHC (TRAP) stainings in the hip-joint tissues from AS patients and non-AS patients. Scale bar: 400 µm. h Representative images of RPS17-RANK coimmunostaining in the hip-joint tissues from AS patients and non-AS patients. Scale bar: 50 or 25 μm. Data in g, h come from the collected joint samples from ANFH patients (n = 6) and AS patients (n = 6) during surgeries. i Western-blotting analysis of RPS17 for CD14⁺CD16⁻ monocytes at the 2^nd, 4^th, 8^th, and 16^th hours of osteoclastic induction. j Representative TEM images of ribosomes for RPS17-silenced CD14⁺CD16⁻ monocytes from HDs and AS patients (n = 3/group) at the 8^th hours of osteoclastic induction. The small black particles in the cytoplasm are ribosomes. Scale bar: 2 μm or 500 nm. Results are from three independent assays with unanimous trends. k Absorbance assays (260 nm) of ribosome extracts from RPS17-silenced CD14⁺CD16⁻ monocytes from the above subjects (n = 3/group) at the 8^th hours of osteoclastic induction. l Osteoclastogenic ability for RPS17-silenced CD14⁺CD16⁻ monocytes from the above subjects (n = 3/group). Scale bar: 100 µm. Data come from the mean values of osteoclasts in each three wells. m Western-blotting validation for RPS17-overexpressing CD14⁺CD16⁻ monocytes from HDs and AS patients. n Osteoclastogenic ability for RPS17-overexpressing CD14⁺CD16⁻ monocytes from the above subjects (n = 3/group). Scale bar: 100 µm. Data come from the mean values of osteoclasts in each three wells. *P < 0.05, **P < 0.01, ****P < 0.000 1 and ns (not significant) by one-way ANOVA (f, i). ***P < 0.001, ****P < 0.000 1 by Student’s t tests (h, j–n)