Journal: bioRxiv

Article Title: CD5L insufficiency exacerbates skeletal joint damage in rheumatoid arthritis

doi: 10.64898/2025.12.16.694616

Figure Lengend Snippet:

Article Snippet: CD5L serum levels were quantified using the Mouse CD5L ELISA Pair Set (Sino Biological, SEK50020), according to the manufacturers’ instructions.

Techniques:

Journal: bioRxiv

Article Title: CD5L insufficiency exacerbates skeletal joint damage in rheumatoid arthritis

doi: 10.64898/2025.12.16.694616

Figure Lengend Snippet: Mice were immunized with chicken collagen on day 0 followed by a boost administration of the antigen on day 21. A , Incidence rates of WT and CD5L − animals were compared using the Chi 2 -statistic as detailed in table 3, * p < 0.05. B , Macroscopic assessment of arthritis was performed by scoring the swelling of each paw as follows: 0 = normal, 1 = mild swelling and/or erythema, 2 = pronounced swelling, 3 = deformity and 4 = ankylosis. RA severity score of each mouse was obtained by adding the score of the four paws. C , Weight variation after arthritis induction. Data represent weight variation only in mice that developed arthritis, shown as mean ± SEM (n = 10 WT; n = 20 CD5L⁻), pooled from five independent experiments. B, Pearson correlation between weight variation and disease score from WT (open circles) and CD5L⁻mice (filled circles).

Article Snippet: CD5L serum levels were quantified using the Mouse CD5L ELISA Pair Set (Sino Biological, SEK50020), according to the manufacturers’ instructions.

Techniques:

Journal: bioRxiv

Article Title: CD5L insufficiency exacerbates skeletal joint damage in rheumatoid arthritis

doi: 10.64898/2025.12.16.694616

Figure Lengend Snippet: Mice were immunized with chicken collagen on day 0 followed by a boost administration of the antigen on day 21. A , Quantification of CD5L by ELISA in the serum of mice after immunization in the indicated time points. Statistical comparisons were made using Kruskal-Wallis test with Dunn’s multiple comparisons. B, The frequency of the indicated populations was analyzed in different time points after immunization in the blood of WT and CD5L − mice by flow cytometry (frequencies within total live cells). Markers used for phenotyping: monocytes (Siglec-F − Ly6G − F4/80 + CD11b + ), inflammatory monocytes (Siglec-F − Ly6G − F4/80 + CD11b + Ly6C + ) and Ly6C − monocytes (Siglec-F − Ly6G − F4/80 + CD11b + Ly6C − ). Data shown are mean with SEM of n = 10 (WT) and n= 10 (CD5L − ) mice. Statistical comparisons were made using Šídák’s multiple comparisons test. C , The concentration of the indicated cytokines was analyzed in different time points after immunization in the sera of WT and CD5L − mice by ELISA. Statistical differences between groups were analyzed by Mann-Whitney test. The concentration of RANKL and cross-linked C-telopeptide of type I collagen (CTX-I) ( D ) was analyzed 56 days after immunization in the sera of WT and CD5L − mice by ELISA. E , RT-qPCR quantification of the indicated genes expression, normalized with Actb (β-actin), in total spleen cells from WT or CD5L − naïve mice. Mice per group: 5. Statistical differences between groups analyzed by two-tailed unpaired t-test with Welch’s correction. Data was pooled from 2 independent experiments except in c) where one representative experiment from two is depicted. Graphical representation of median and 25 th – 75 th quartiles in A ), C ), D ), E ). * p < 0.05, ** p < 0.01, ns: not significant.

Article Snippet: CD5L serum levels were quantified using the Mouse CD5L ELISA Pair Set (Sino Biological, SEK50020), according to the manufacturers’ instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Flow Cytometry, Concentration Assay, MANN-WHITNEY, Quantitative RT-PCR, Expressing, Two Tailed Test

Journal: bioRxiv

Article Title: CD5L insufficiency exacerbates skeletal joint damage in rheumatoid arthritis

doi: 10.64898/2025.12.16.694616

Figure Lengend Snippet: A , Representative histograms showing Pacific Blue-CD11b and APC-CD29 expression in human CD11b⁺CD14⁺CD4⁻ peripheral blood leukocytes stimulated with LPS and ConA for 48 h, with the addition of human recombinant CD5L (1 μg/ml) or recombinant CD5L plus human IgG (equimolar) during the final 24 h. Expression was assessed by flow cytometry. Box plots depict mean fluorescence intensity (MFI) of CD11b and CD29 in cultured cells (n = 8). P values were calculated using a paired Wilcoxon test. B , Column plots of CD16 expression on intermediate (IM, CD14 + CD16 + ) and non-classical (NCM, CD14 low CD16 + ) monocyte subsets. C , Protein levels of IL-10, PDL1, IFN-ψ and GAS6 in the supernatants of cell cultures. D , Heat maps showing Spearman correlation (π) values between IL-10 concentrations in culture supernatants and the expression intensity of CD11b and CD16, as measured by flow cytometry. Statistical significance is indicated as * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: CD5L serum levels were quantified using the Mouse CD5L ELISA Pair Set (Sino Biological, SEK50020), according to the manufacturers’ instructions.

Techniques: Expressing, Recombinant, Flow Cytometry, Fluorescence, Cell Culture

Journal: bioRxiv

Article Title: CD5L insufficiency exacerbates skeletal joint damage in rheumatoid arthritis

doi: 10.64898/2025.12.16.694616

Figure Lengend Snippet: A , Box plots showing CD5L, IL-10, PD-L1, IFN-ψ, and GAS6 protein levels in supernatants from human peripheral blood leukocyte cultures (n = 14) stimulated with LPS and ConA (5 μg/mL and 0.625 μg/m, respectively) for 48 h, with the addition of a histone deacetylase inhibitor (HDACi, 50 μg/ml) during the final 24 h. B , Heat maps displaying Spearman correlation (π) values between CD5L and IL-10 levels in supernatants, CD16 expression intensity measured by flow cytometry, IFN-γ production, the relative size of the non-classical monocyte (NCM) subset, and the mean fluorescence intensity (MFI) of CD16 and CD11b. Statistical significance is indicated as * p < 0.05; ** p < 0.01; *** p < 0.001. C, Box plots illustrating changes in classical (CM; CD14⁺CD16⁻), intermediate (IM; CD14⁺CD16⁺), and non-classical (NCM; CD14 low CD16⁺) monocyte subsets in stimulated human leukocyte cultures (n = 14), treated as described in panel ( A ). D, Representative histograms of Pacific Blue-CD11b expression in human CD11b⁺CD14⁺CD4⁻ blood leukocytes stimulated as indicated and analyzed by flow cytometry. Box plots summarize CD11b mean fluorescence intensity (MFI) across conditions. P values were calculated using a paired Wilcoxon test.

Article Snippet: CD5L serum levels were quantified using the Mouse CD5L ELISA Pair Set (Sino Biological, SEK50020), according to the manufacturers’ instructions.

Techniques: Histone Deacetylase Assay, Expressing, Flow Cytometry, Fluorescence

Journal: bioRxiv

Article Title: CD5L insufficiency exacerbates skeletal joint damage in rheumatoid arthritis

doi: 10.64898/2025.12.16.694616

Figure Lengend Snippet: CD14⁺ cells were isolated from peripheral blood leukocyte cultures of 35 RA patients, and CD5L levels in culture supernatants were measured by ELISA. Cultures with CD5L protein levels above 0.6 pg/mL (double detection limit) were classified as CD5L hi , and the remaining as CD5L low . A, Box plots showing protein levels of CD5L, IL-10, and PD-L1 in supernatants, as well as patient white blood cell (WBC) and platelet counts, and serum IFN-ψ levels, comparing CD5L hi (n = 10) and CD5L low (n = 25) CD14⁺ cells. B, Box plots of normalized gene expression levels in CD14⁺ cells measured by RNA-seq. P values were calculated using DESeq2; nominal p values are indicated.

Article Snippet: CD5L serum levels were quantified using the Mouse CD5L ELISA Pair Set (Sino Biological, SEK50020), according to the manufacturers’ instructions.

Techniques: Isolation, Enzyme-linked Immunosorbent Assay, Gene Expression, RNA Sequencing

Journal: bioRxiv

Article Title: CD5L insufficiency exacerbates skeletal joint damage in rheumatoid arthritis

doi: 10.64898/2025.12.16.694616

Figure Lengend Snippet: CD14⁺ cells were isolated from peripheral blood leukocyte cultures of 35 RA patients, activated with LPS for 2 h, and subjected to RNA sequencing (RNA-seq, Illumina). A, Bubble plot showing mean normalized expression of gene markers for four monocyte clusters identified in human blood leukocytes. B, Bar plot representing the distribution of monocyte clusters based on the sum of cluster-specific gene expression. C, Heat map of gene expression changes (log₂ fold change) in non-classical and IFN-primed monocyte clusters in regression to serum CD5L levels and joint skeletal damage quantified by vdH-Sharp score. P values were calculated using DESeq2; nominal P values are indicated as * p < 0.05; ** p < 0.01; *** p < 0.001. D, Heat map of gene expression changes (log₂ fold change, FC) for efferocytosis markers (1), resolution mediators (2), and immunoregulatory macrophage program genes (3) in regression to serum CD5L levels and vdH-Sharp score. E, Schematic representation of a monocyte conditioned by CD5L.

Article Snippet: CD5L serum levels were quantified using the Mouse CD5L ELISA Pair Set (Sino Biological, SEK50020), according to the manufacturers’ instructions.

Techniques: Isolation, RNA Sequencing, Expressing, Gene Expression

Journal: bioRxiv

Article Title: CD5L insufficiency exacerbates skeletal joint damage in rheumatoid arthritis

doi: 10.64898/2025.12.16.694616

Figure Lengend Snippet: A , Box plots of CD5L protein levels in serum of RA patients and healthy controls (1), paired serum and synovial fluid samples from RA patients (2), and anti-CD5L antibodies in serum of RA patients and controls (3). B, Heat map of Spearman correlations (ρ) between serum CD5L levels or vdH-Sharp joint damage scores and serum cytokine and growth factor levels. Statistical significance is indicated as * p < 0.05; ** p < 0.01; *** p < 0.001. C, Dot plots showing correlations between serum CD5L levels and radiographic joint damage in 80 RA patients (1), and correlation of a CD5L-dependent gene signature in non-classical monocytes with vdH-Sharp score (2). Enrichment with osteoclast-related markers enhanced the correlation, whereas inclusion of IFN-sensitive genes abolished it. D, Bubble plot of the percentage frequency of CD5L⁺ cells within synovial myeloid clusters. E, UMAP of the scaled expression intensity sum of the CD5L-dependent signature in synovial myeloid cells from single-cell transcriptomics. Further enrichment distinguishes IFN-primed versus osteoclast-fostering macrophages. F, Heat map of a CD5L-dependent osteoclastogenic signature in blood CD14⁺ cells, showing a positive correlation with vdH-Sharp joint damage scores.

Article Snippet: CD5L serum levels were quantified using the Mouse CD5L ELISA Pair Set (Sino Biological, SEK50020), according to the manufacturers’ instructions.

Techniques: Expressing, Single-cell Transcriptomics