CCR5 Antibody - #AF6339

Fig. 5 CCL5 promotes prostate cancer invasion and PCSCs self-renewal via activating the CCR5/β-catenin/STAT3 pathway. a–c Heatmaps of 94 metastasis-related DEGs (a), 42 stemness-related DEGs (b), as well as 30 STAT3 pathway-related DEGs (c). RNA-Seq analysis was conducted to characterize the cellular responses of PC3 cells to 40 ng/ml CCL5 treatment. Differential gene expression analysis was conducted to identify the DEGs (n = 3). d Venn diagram of the DEGs in the indicated groups. CTNNB1, also known as β-catenin, was the most significant one of them. e Western blotting assay indicated that the CD133+ PCSCs sorted from PC3 cells by MACS method exhibited increased expression of CCR5, β-catenin, and STAT3 when compared with the CD133- subpopulation (n = 3). f CCL5 treatment significantly elevated the promoter activity of STAT3 in PC3 cells while XAV-939, the specific inhibitor of β-catenin, partly abrogated that (n = 6). g Immunofluorescence assay indicated that CCL5 treatment (40 ng/ ml) could significantly induce the expression and nuclear translocation of β-catenin in prostate cancer cells. Scale bar, 10 μm. h CHIP assay suggested that β-catenin could bind to the promoter region of STAT3, while the specific inhibitor of β-catenin (XAV-939) partly decreased their binding activity. i CCR5 specific siRNAs could significantly abrogate the activation effect of CCL5 on β-catenin/STAT3 pathway (n = 3). All values are presented as the mean ± SD, *p < 0.05, **p < 0.01.

Fig. 4 Inhibiting the CCL5/CCR5 signaling pathway attenuates hyperalgesia in DRG PP2Cm-deficient mice a, Quantification for release of CCL5 in the L4-L5 DRGs of PP2Cm-ctrl and PP2Cm-cKO mice, as detected by ELISA. n = 4 mice/group. b, Illustrative cartoon depicting the intrathecal administration of an anti-CCL5 neutralizing antibody. c, Quantification for release of CCL5 in the L4-L5 DRGs of PP2Cm-cKO + IgG, PP2Cm-cKO + anti-CCL5 1d and PP2Cm-cKO + anti-CCL5 3d mice. n = 4 mice/group. d-f, Mechanical withdrawal thresholds (d), withdrawal latencies to heat (e) and cold stimulus (f) on day 0 (baseline, BL), 1, and 3 post-injection in PP2Cm-cKO mice treated with IgG or anti-CCL5 neutralizing antibody. n = 5–6 mice/group. g, The L4-L5 DRGs from PP2Cm-ctrl and PP2Cm-cKO mice were double stained with PP2Cm (green) and CCR5 (red). The corner image in white square is the zoomed-in image of the area in the smaller white square. Scale bar: 20 μm. h, i, Fluorescent quantifications for PP2Cm (h) and CCR5 (i) in DRG sections from PP2Cm-ctrl and PP2Cm-cKO mice. n = 3 sections from 3 mice/group. j, A representative cartoon illustrating the intrathecal injection of CCR5 antagonist maraviroc or Ccr5 small interfering RNA (siCcr5). k-m, Mechanical withdrawal thresholds (k), withdrawal latencies to heat (l) and cold stimulus (m) at BL, 1 h, and 24 h post-injection in PP2Cm-cKO mice treated with Vehicle or maraviroc. n = 6 mice/group. n, The mRNA levels of Ccr5 in the L4-L5 DRGs from PP2Cm-cKO + siCtrl, PP2Cm-cKO + siCcr5 2d, and PP2Cm-cKO + siCcr5 8d mice (to β-actin). n = 3 experimental repeats (6 mice)/group. o-q, Mechanical withdrawal thresholds (o), withdrawal latencies to heat (p) and cold stimulus (q) at BL, 2d, and 8d post-injection in PP2Cm-cKO mice treated with siCtrl or siCcr5. n = 5–6 mice/group. Data are shown as the mean ± SEM. Statistical tests used were unpaired two-tailed Student’s t-test (a, h, i), one-way ANOVA with Tukey’s multiple comparisons test (c, n), and two-way repeated-measures ANOVA with Bonferroni’s post hoc test (d-f, k-m, o-q). *p