NR4A1 and NR4A3 expression in indolent and aggressive NHLs. (A) Depiction of NR4A1 and NR4A3 mRNA expression levels in indolent and aggressive NHL patients with reduced NR4A content and normal controls [reactive lymph nodes (LA), peripheral B-cells (CD19), and GCBs]. (B) Representative western blot analysis of NR4A1 and NR4A3 expression in peripheral B cells (B1-B4), DLBCL (DL1-12), and FLIII (FL1-12). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) western blot analysis served as a loading control. (C) Expression analysis of Fas-L, Trail, the 3 splice variants of BIM (BIM 1,6 and 9), and Puma as potential NR4A target genes in indolent and aggressive NHL patients with a reduced NR4A1 and NR4A3 content compared with GCBs and MC-Bs by RQ-PCR. (D) NR4A1 and NR4A3 expression in GCB-DLBCL, non–GCB-DLBCL, and FLIII by RQ-PCR. (E) Probability of cancer-specific survival in DLBCL patients stratified according to the NR4A1 mRNA expression level. Low NR4A1 expression is associated with poor survival. (F) Immunohistochemical analysis of NR4A1 in nonneoplastic lymph nodes (magnification ×200 for panel Fi, magnification ×400 for panel Fii), in GCB-DLBCL (magnification ×400 for panel Fiii), in non–GCB-DLBCL (magnification ×400 for panel Fiv), and in FLIII (magnification ×400 for panel Fv). Strong NR4A1 expression was found in almost all GCBs in nonneoplastic lymph nodes, whereas a weaker nuclear NR4A1 expression was detected in GCB-DLBCL and non–GCB-DLBCL and FLIII. All images were captured by using an Olympus BX51 microscope and an Olympus E-330 camera. mRNA expression levels were calculated as relative expression in comparison with peripheral mononucleated cells serving as a calibrator. Each bar represents the mean values of expression levels ± standard deviation (SD). The comparison of the expression levels was performed by using the Mann-Whitney U test; all significant associations were corrected for multiple testing by applying a Bonferroni correction. * indicates reduced expression compared with LA or CD19⁺ cells (P < .01); □, reduced expression compared with GCBs (P < .01); †, reduced expression compared with mantle cells (P < .01); Δ, reduced expression compared with GCB-DLBCL (P < .01); and ○, reduced expression compared with FLIII (P < .01). Number in or above columns indicates number of tested specimens.