Fig 1: RImAb reprograms the tumor microenvironment towards a more inflammatory and antitumoral phenotype in a mouse model of LLC. a)Left: Representative immunohistochemistry images of MPO and TNF-a expression in tumor samples from control (PBS) and RImAb-treated groups. Right: quantification of the average stained area of five random images obtained from each group. b) Levels of TNF-a, GM-CSF, MIP-1a, and RANTES cytokines in tumor homogenates, expressed as pg cytokine/mg protein, in control (isotype IgG2a) and RImAb-treated mice (8 mice per group), measured by the Multiplex Assay. Bar graphs show means with SEM. c) Expression of Ctla4 and Pdcd1 mRNA was assessed by RT-qPCR in total tumor from control (isotype IgG2a) and RImAb-treated mice. d)Left: Representative immunohistochemistry images of BCL2 expression in tumor samples from the control (PBS) and RImAb-treated mice. Right: quantification of the average stained area of 5 random images obtained from each group. e) Expression of EMT markers Zeb1, Snail1, and Twist1 mRNA was assessed by RT-qPCR in total tumor from control (isotype IgG2a) and RImAb-treated mice. f)Left: Representative immunohistochemistry images of CD31 expression in the tumor samples from control (PBS) and RImAb-treated mice. Right: Quantification shows the number of blood vessels per field of five random images obtained from each group. g) Expression of Angpt2 mRNA was assessed by RT-qPCR in total tumor from control (isotype IgG2a) and RImAb-treated mice. For a), d) and f), pictures were quantified using Image J (color deconvolution) software. Scale bars represent 10 µm, and data are presented as the mean ± SEM (n = 8 per group). For c), e), and g), mRNA levels are relative to Actb, and fold induction levels were calculated by using as a reference the average level of expression of each gene in tumors from control (isotype IgG2a) mice, and are represented with a boxplot showing median with whiskers (min to max) (n = 8 per group). For all panels, data are presented as the mean ± SEM, and statistical significance was calculated using the Mann–Whitney t-test, *p < 0.05, **p < 0.01, ***p = 0.001.