Journal: Scientific Reports

Article Title: Combined proteomics and metabolomics analyses revealed molecular signatures associated with proliferative diabetic retinopathy

doi: 10.1038/s41598-026-40551-1

Figure Lengend Snippet: ELISA quantification of CD5L ( A ), CLU ( B ), and SERPINF1 ( C ) protein levels in the vitreous humor of DR and Control patients. * p < 0.05, **** p < 0.0001.

Article Snippet: The C166 + CD5L group was supplemented with 400 pg/mL recombinant CD5L protein (MCE, USA, HY- P72717 ), while the control group received an equal volume of PBS.

Techniques: Enzyme-linked Immunosorbent Assay, Control

Journal: Scientific Reports

Article Title: Combined proteomics and metabolomics analyses revealed molecular signatures associated with proliferative diabetic retinopathy

doi: 10.1038/s41598-026-40551-1

Figure Lengend Snippet: Immunohistochemical analysis of CD5L, CLU, and SERPINF1 expression. Immunohistochemical analysis of CD5L, CLU, and SERPINF1 expression in whole-retinal and retinal nerve fiber layer from DR and control SD rats. Images were acquired at 40×magnification, full-field view; scale bar, 25 μm. Nuclei are counterstained blue, and positive staining for CD5L, CLU, and SERPINF1 appears brown. ** p < 0.01, *** p < 0.001.

Article Snippet: The C166 + CD5L group was supplemented with 400 pg/mL recombinant CD5L protein (MCE, USA, HY- P72717 ), while the control group received an equal volume of PBS.

Techniques: Immunohistochemical staining, Expressing, Control, Staining

Journal: Scientific Reports

Article Title: Combined proteomics and metabolomics analyses revealed molecular signatures associated with proliferative diabetic retinopathy

doi: 10.1038/s41598-026-40551-1

Figure Lengend Snippet: Prediction of Transcription factors (TFs) and modification sites in key proteins. ( A ) Transcription factor prediction of key proteins. ( B ) Prediction results of PTM sites of key protein APLP2. ( C ) Prediction results of PTM sites of key protein RBP3. ( D ) Prediction results of PTM sites of key protein SERPINF1. ( E ) Prediction results of acetylation of key protein CLU. ( F ) Prediction results of acetylation of key protein PSAP. ( G ) Prediction results of phosphorylation, acetylation, ubiquitination of key protein C5. ( H ) Prediction results of key protein CD5L.

Article Snippet: The C166 + CD5L group was supplemented with 400 pg/mL recombinant CD5L protein (MCE, USA, HY- P72717 ), while the control group received an equal volume of PBS.

Techniques: Modification, Phospho-proteomics, Ubiquitin Proteomics

Journal: Scientific Reports

Article Title: Combined proteomics and metabolomics analyses revealed molecular signatures associated with proliferative diabetic retinopathy

doi: 10.1038/s41598-026-40551-1

Figure Lengend Snippet: Prediction of drug candidates targeting key proteins and functional effects of exogenous CD5L on endothelial cells. ( A ) Molecular docking diagram of key protein RBP3 and drug VITAMIN A PALMITATE. ( B ) Molecular docking diagram of key protein RBP3 and drug VITAMIN A PALMITATE. The purple is the key protein, the iridescent is the drug, and the yellow dotted line is the hydrogen bond between the two, as well as the corresponding bond length and residue. ( C ) EdU assay showing the effect of CD5L on C166 cell proliferation.(Images are shown at 10×magnification, scale bar, 2 μm, proliferating cells are indicated by green fluorescence (EdU), ** p <0.01). ( D ) Scratch wound assay showing the effect of CD5L on C166 cell migration (* p <0.05).

Article Snippet: The C166 + CD5L group was supplemented with 400 pg/mL recombinant CD5L protein (MCE, USA, HY- P72717 ), while the control group received an equal volume of PBS.

Techniques: Functional Assay, Residue, EdU Assay, Fluorescence, Scratch Wound Assay Assay, Migration

Journal: Frontiers in Immunology

Article Title: Endogenous CD5L controls the metabolic and inflammatory state of human macrophages

doi: 10.3389/fimmu.2025.1677948

Figure Lengend Snippet: CD5L -KO cells have reduced inflammatory state. Deletion of CD5L led to decrease in basal and LPS-induced cytokine mRNA levels. TNF and IL-1β levels in unstimulated cells were decreased nearly 300-fold, with average basal levels of 0.003 for knockout cells compared to the value of 1 set for the controls. A total of 2.5 × 10 5 cells in 24-well plates were differentiated using 50 ng/mL PMA for 48 hours. Stimulation was performed after 24 hours of rest in PMA-free media with 100 ng/mL LPS for 4 hours. Inflammatory cytokine expression levels were determined using qRT-PCR of RNAs isolated from cell lysates. Graphs are representative of at least three independent experiments. PMA, phorbol 12-myristate 13-acetate. (* p<0.05, ** p<0.01, *** p<0.001).

Article Snippet: The primary antibodies against CD5L (mouse monoclonal F1 clone, 1/1,000, sc-514283, Santa Cruz Biotechnology), GAPDH (1/2000, sc-47724, Santa Cruz Biotechnology, Santa Cruz, CA USA), RORA (1/1,000 PP-H3910-00, R&D Systems), and the HRP-conjugated goat anti-mouse secondary antibody (1/5000, #P0447, Dako) were diluted in 0.5× blocking buffer/PBS.

Techniques: Knock-Out, Expressing, Quantitative RT-PCR, Isolation

Journal: Frontiers in Immunology

Article Title: Endogenous CD5L controls the metabolic and inflammatory state of human macrophages

doi: 10.3389/fimmu.2025.1677948

Figure Lengend Snippet: RORα is epistatic to CD5L . Induction of inflammatory TNF (A) and IL-1β (B) gene expression in CD5L/RORA DKO cells is similar to that observed in RORA deletion cells (n = 3, ± SEM). Paired two-tailed t -test of three biological replicates has been used to evaluate statistical significance of differences between RORAko and CD5L/RORA DKO cell lines. Both RORAko and CD5L/RORA DKO cells produce similarly higher levels of TNF than the CD5Lko cells at 3 and 6 hours after LPS stimulation (C) . CD5Lko cells produce significantly lower amounts of IL-6 at both the 3- and 6-hour timepoints, while RORAko and CD5L/RORA DKO secrete similar levels of this cytokine (D) (n = 6, ± SEM, representative of two independent experiments * p<0.05, ** p<0.01).

Article Snippet: The primary antibodies against CD5L (mouse monoclonal F1 clone, 1/1,000, sc-514283, Santa Cruz Biotechnology), GAPDH (1/2000, sc-47724, Santa Cruz Biotechnology, Santa Cruz, CA USA), RORA (1/1,000 PP-H3910-00, R&D Systems), and the HRP-conjugated goat anti-mouse secondary antibody (1/5000, #P0447, Dako) were diluted in 0.5× blocking buffer/PBS.

Techniques: Gene Expression, Two Tailed Test

Journal: Frontiers in Immunology

Article Title: Endogenous CD5L controls the metabolic and inflammatory state of human macrophages

doi: 10.3389/fimmu.2025.1677948

Figure Lengend Snippet: RNA-seq analysis of differentiated CD5L deletion cells. (A) Volcano plot of ANOVA results. Vertical cutoff line is at FDR-adjusted p = 0.05. Vertical cutoff lines are at fold difference = 2. A total of 1,165 annotated genes were found to be differentially expressed between CD5L deletion and control cell lines using these criteria. The majority of differentially expressed genes (879) were downregulated (blue). (B) Gene Ontology terms enriched by the differentially expressed dataset include regulation of intracellular processes as well as cell-to-cell signaling characteristic of immune cells. (C) All five most significantly enriched KEGG pathways mediate immune signaling. (D) The magnitude of changes in expression levels of genes that contribute to the top five enriched KEGG pathways. For genes that are not part of a specific pathway, the values are shown in gray. Colors represent fold difference change in gene expression when compared to controls. FDR, false discovery rate.

Article Snippet: The primary antibodies against CD5L (mouse monoclonal F1 clone, 1/1,000, sc-514283, Santa Cruz Biotechnology), GAPDH (1/2000, sc-47724, Santa Cruz Biotechnology, Santa Cruz, CA USA), RORA (1/1,000 PP-H3910-00, R&D Systems), and the HRP-conjugated goat anti-mouse secondary antibody (1/5000, #P0447, Dako) were diluted in 0.5× blocking buffer/PBS.

Techniques: RNA Sequencing, Control, Expressing, Gene Expression

Journal: Frontiers in Immunology

Article Title: Endogenous CD5L controls the metabolic and inflammatory state of human macrophages

doi: 10.3389/fimmu.2025.1677948

Figure Lengend Snippet: Expression of genes in the TNF signaling pathway is impacted by deletion of CD5L . Deletion of CD5L results in downregulation (green) of transcription of nearly half of the genes comprising the TNF signaling pathway. Expression of signal propagating kinases is also affected by p38, and JNK is downregulated at the transcriptional level (adapted from KEGG TNF signaling pathway, hsa04668 6/25/18).

Article Snippet: The primary antibodies against CD5L (mouse monoclonal F1 clone, 1/1,000, sc-514283, Santa Cruz Biotechnology), GAPDH (1/2000, sc-47724, Santa Cruz Biotechnology, Santa Cruz, CA USA), RORA (1/1,000 PP-H3910-00, R&D Systems), and the HRP-conjugated goat anti-mouse secondary antibody (1/5000, #P0447, Dako) were diluted in 0.5× blocking buffer/PBS.

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: Endogenous CD5L controls the metabolic and inflammatory state of human macrophages

doi: 10.3389/fimmu.2025.1677948

Figure Lengend Snippet: Lipid remodeling function of CD5L. (A) Total levels of lipid classes analyzed in mutant and control cells. (B) Proportion of side acyl chains classified according to the extent of their saturation. MUFA: one or two double bonds in a 3-acyl chain lipid or one double bond in a 2-acyl chain lipid. MUFA1: number of double bonds and acyl chains is equal. PUFA2, PUFA3, and PUFA4: number of double bonds is two-, three-, and fourfold higher compared to number of acyl chains in the lipid, respectively. (C) Deletion of CD5L significantly (*** p < 0.001, Mann–Whitney test) increases levels of highly polyunsaturated [PUFA(3,4,5)] fatty acids. (D) CD5L deletion results in an increase of free cholesterol in mutant cells. PUFA, polyunsaturated fatty acid. (E) Corrected Total Cell Fluorescence (CTCF) values calculated for 10 randomly selected cells for each cell line (* p = 0.014 unpaired two-tailed t -test).

Article Snippet: The primary antibodies against CD5L (mouse monoclonal F1 clone, 1/1,000, sc-514283, Santa Cruz Biotechnology), GAPDH (1/2000, sc-47724, Santa Cruz Biotechnology, Santa Cruz, CA USA), RORA (1/1,000 PP-H3910-00, R&D Systems), and the HRP-conjugated goat anti-mouse secondary antibody (1/5000, #P0447, Dako) were diluted in 0.5× blocking buffer/PBS.

Techniques: Mutagenesis, Control, MANN-WHITNEY, Fluorescence, Two Tailed Test

Journal: Frontiers in Immunology

Article Title: Endogenous CD5L controls the metabolic and inflammatory state of human macrophages

doi: 10.3389/fimmu.2025.1677948

Figure Lengend Snippet: FASN activity in knockout and control cells. Deletion of CD5L does not significantly change the overall FASN activity of human macrophages. Lysates from 10 6 cells were used to measure FASN activity by monitoring NADPH oxidation over 15 minutes. Activity is presented as an average of replicates from three independent experiments. FASN, fatty acid synthase.

Article Snippet: The primary antibodies against CD5L (mouse monoclonal F1 clone, 1/1,000, sc-514283, Santa Cruz Biotechnology), GAPDH (1/2000, sc-47724, Santa Cruz Biotechnology, Santa Cruz, CA USA), RORA (1/1,000 PP-H3910-00, R&D Systems), and the HRP-conjugated goat anti-mouse secondary antibody (1/5000, #P0447, Dako) were diluted in 0.5× blocking buffer/PBS.

Techniques: Activity Assay, Knock-Out, Control

Journal: Frontiers in Immunology

Article Title: Endogenous CD5L controls the metabolic and inflammatory state of human macrophages

doi: 10.3389/fimmu.2025.1677948

Figure Lengend Snippet: Differences in gene expression in CD5L and RORA deletion cells. (A) Self-organizing map (SOM) clustering identified six distinct patterns of changes in gene expression between respective mutant and control cells. (B) GO term analysis of genes in highlighted divergently regulated clusters identifies terms characteristic of immune response. (C) Eight out of 10 KEGG pathways enriched by genes from highlighted clusters are central to shaping the inflammatory signaling of immune cells. Genes contributing to the TNF (D) and NF-κB (E) signaling pathways are downregulated in CD5L mutant cells and upregulated in RORA mutants. GO, Gene Ontology.

Article Snippet: The primary antibodies against CD5L (mouse monoclonal F1 clone, 1/1,000, sc-514283, Santa Cruz Biotechnology), GAPDH (1/2000, sc-47724, Santa Cruz Biotechnology, Santa Cruz, CA USA), RORA (1/1,000 PP-H3910-00, R&D Systems), and the HRP-conjugated goat anti-mouse secondary antibody (1/5000, #P0447, Dako) were diluted in 0.5× blocking buffer/PBS.

Techniques: Gene Expression, Mutagenesis, Control, Protein-Protein interactions

Journal: Frontiers in Immunology

Article Title: Endogenous CD5L controls the metabolic and inflammatory state of human macrophages

doi: 10.3389/fimmu.2025.1677948

Figure Lengend Snippet: CD5L deletion induces transcriptional changes in undifferentiated monocytes. (A) Deletion of CD5L in undifferentiated monocytic THP-1 cells significantly changed expression of 52 genes more than twofold up and 95 genes twofold down (in color). (B) Expression of atherogenesis-related genes involved in interaction of monocytes with ECM is changed in CD5Lko cells. (C) CD52 expression is significantly increased in undifferentiated (FDR = 3e−14) and differentiated (FDR = 2.9e−35) CD5Lko cells. (D) Cellular CD52 protein levels are increased in undifferentiated CD5Lko THP-1 monocytes. ECM, extracellular matrix; FDR, false discovery rate.

Article Snippet: The primary antibodies against CD5L (mouse monoclonal F1 clone, 1/1,000, sc-514283, Santa Cruz Biotechnology), GAPDH (1/2000, sc-47724, Santa Cruz Biotechnology, Santa Cruz, CA USA), RORA (1/1,000 PP-H3910-00, R&D Systems), and the HRP-conjugated goat anti-mouse secondary antibody (1/5000, #P0447, Dako) were diluted in 0.5× blocking buffer/PBS.

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: Endogenous CD5L controls the metabolic and inflammatory state of human macrophages

doi: 10.3389/fimmu.2025.1677948

Figure Lengend Snippet: Model for the role of CD5L in mediating inflammatory state of human macrophages. (A) CD5L induces changes in macrophages’ lipid content, reducing the availability of RORα ligands and subsequent RORα inactivation. Thus, no inhibitory effect of RORα is applied on inflammatory signaling. (B) In the absence of CD5L, there are sufficient levels of RORα ligands driving its activation and downregulation of inflammatory signaling.

Article Snippet: The primary antibodies against CD5L (mouse monoclonal F1 clone, 1/1,000, sc-514283, Santa Cruz Biotechnology), GAPDH (1/2000, sc-47724, Santa Cruz Biotechnology, Santa Cruz, CA USA), RORA (1/1,000 PP-H3910-00, R&D Systems), and the HRP-conjugated goat anti-mouse secondary antibody (1/5000, #P0447, Dako) were diluted in 0.5× blocking buffer/PBS.

Techniques: Activation Assay

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

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