Fig. 6: LYN-dependent regulation of the ECM proteins like Thrombospondin-1 (THBS1) in stromal cells supported the viability of CLL cells.
a CLL cells (n = 8) were cultured on decellularized ECM from LYNWT and LYNKO HS-5 cells for 48 h. (i) illustration, (ii) CLL viability as Annexin V−/7AAD− per CD45+ cells. (mean, two-sided Wilcoxon test, p = 0.0078). b Immunoblot of BGN in LYNWT and LYNKO HS-5 cells (1 sample each). c HS-5 cells were transfected with siBGN or control siRNA (15 nM), CLL (n = 9) were co-cultured for 96 h (mean, two-sided Uncorrected Dunn’s test: without feeder vs. siCT p < 0.0001; without feeder vs. siBGN p = 0.0339; siCT vs. siBGN p = 0.0339). d–f THBS1 expression in HS-5 cells. d qRT-PCR analysis normalized to PPIA. e Immunoblot for THBS1. f Soluble THBS1 measured by ELISA (d: 2 clones per genotype, e, f: 3 clones per genotype; mean ± SEM; unpaired two-sided t-test: p = 0256). g CLL cells (n = 8) were cultured with recombinant human THBS1 and viability was assessed after 24 h (mean, two-sided Wilcoxon test compared to untreated, p = 0.0391). h THBS1-overexpressing and Mock HS-5 cells were cultured with CLL cells (n = 15, 1 clone per condition), (mean ± SEM; two-sided Wilcoxon test for Mock- vs. THBS1-overexpressing: 24 h p = 0.005371, 48 h p = 0.000366, 72 h p = 0.000427, 96 h p = 0.003906). i CLL cells (n = 6) were co-cultured with THBS1 knockdown-LYNKO or control HS-5 cells for 72 h (mean, two-sided Wilcoxon test p = 0.0312). j CLL cells (n = 10) were co-cultured with antibody-pretreated HS-5 cells for 48 h. (i) illustration, (ii) CLL cell viability was quantified by flow cytometry (mean, two-sided Wilcoxon test p = 0.0469). k Violin plot for mean THBS1 expression per segmented single cell in Imaging Mass Cytometry (Fig. 1d). HC-LN: n = 9 (6466 fibroblasts), CLL-LN: n = 12 (7543 fibroblasts). Boxplots represent median with the interquartile range, whiskers indicate adjacent values, violin represents kernel density estimation, independent two-sided t-tests (p = 1.522e−257). See also Fig. S4. Illustration was created with BioRender.com. Source data are provided as a Source Data file.
