Coexpression of CD44-positive/CD133-positive cancer stem cells and CD204-positive tumor-associated macrophages is a predictor of survival in pancreatic ductal adenocarcinoma

Abstract

Background: The interactions between cancer stem cells (CSCs) and tumor-associated macrophages (TAMs) can promote tumor progression, maintain the CSCs population, and reduce therapeutic effects. The objective of this study was to investigate the coexpression of CSCs and TAMs and its clinical significance in pancreatic ductal adenocarcinoma (PDAC).

Methods: Ninety-six patients with PDAC were included in this study. Tissue microarrays were constructed for immunostaining of the CSCs markers CD44 and CD133 and the TAMs marker CD204. Correlations between the expression of CSCs and TAMs markers and clinicopathologic characteristics or disease progression were analyzed.

Results: Expression levels of CD44/CD133 and CD204 were significantly higher in tumor tissues than in normal tissues (P < .0001). The variables associated with survival were high coexpression of CD44/CD133 (P = .000), high expression of CD204 (P = .011), and tumor grade (P = .014). There was a positive correlation between CD44/CD133 and CD204 expression (r = 0.294; P = .004). Survival analysis indicated that high coexpression of CD44/CD133 and CD204 was associated significantly with shorter overall survival (P = .000) and disease-free survival (P = .003). Multivariate analysis revealed that high CD44/CD133 expression was an independent prognostic factor for disease-free survival, whereas high CD204 expression was an independent predictor for both overall and disease-free survival.

Conclusions: Coexpression of CD44/CD133 and CD204 is a useful survival prediction marker for patients with PDAC.

Keywords: CD133; CD204; CD44; cancer stem cells; pancreatic ductal adenocarcinoma; tissue microarrays; tumor-associated macrophages.

Figure 1

Expression of the cancer stem cells markers CD44 and CD133 is observed in tissue microarrays (TMAs) and in corresponding full sections of pancreatic ductal adenocarcinoma. (A) Two different TMA cores from the same tumor have a similar pattern of low CD44/CD133 expression; and (B) Two different TMA cores from the same tumor have a similar pattern of high CD44/CD133 expression. White arrows indicate CD44-positive/CD133-positive cells (original magnification ×4 in A and B; scale bars = 200 μm). (C) Part of a slide from the donor block of the same tumor is shown. Normal pancreatic tissues were obtained from the same patient as a control. Red indicates CD44 staining; green, CD133 staining. Nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI) (blue). Yellow indicates the colocalization of CD44 and CD133 (original magnification ×400; scale bar = 20 μm). (D) CD44/CD133 expression levels (mean ± standard error) were compared between pancreatic cancer specimens and their matched normal tissues (n = 11 pairs; P < .0001). (E) CD44/CD133 expression levels (mean ± standard error) were compared between pancreatic cancer tissues and adjacent normal tissues (P < .0001).

Figure 2

Expression of the tumor-associated macrophages marker CD204 is observed in tissue microarrays (TMAs) and corresponding full sections of pancreatic ductal adenocarcinoma tissue. (A) Two different TMA cores from the same tumor have a similar pattern of low CD204 expression, and (B) Two different TMA cores from the same tumor have a similar pattern of high CD204 expression. White arrows indicate CD204-positive cells (original magnification ×4 in A and B; scale bars = 200 μm). (C) Part of a slide from the donor block of the same tumor is shown at ×200 original magnification (scale bar = 50 μm), and at ×400 magnification on the right (scale bar = 20 μm). The paraffin-embedded patient tissue samples were stained with anti-CD204 antibody (green) and with 4′,6-diamidino-2-phenylindole (DAPI) for cell nuclei (blue). (D) CD204 expression was compared between pancreatic cancer specimens and their matched normal tissues (n = 11 pairs; P < .0001). Values indicate the mean ± standard error. (E) CD204 expression levels (mean ± standard error) were compared between pancreatic cancer tissues and adjacent normal tissues (P < .0001).

Figure 3

The correlation between CD44-positive/CD133-positive cancer stem cells (CSCs) and CD204-positive tumor-associated macrophages (TAMs) in pancreatic ductal adenocarcinoma is illustrated. (A) CD44/CD133 colocalized with CD68/CD204 in pancreatic cancer specimens. White arrows indicate CD44-positive/CD133-positive or CD68-positive/CD204-positive cells. Red indicates CD44 or CD68 staining; green, CD133 or CD204 staining. Nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI) (blue) (original magnification ×400; scale bar = 50 μm). Charts illustrate correlation analyses of CSCs marker expression and macrophages marker (B) or TAMs marker (C) expression level (Pearson test). (D,E) The presence of CD44-positive/CD133-positive CSCs and CD204-positive TAMs had a positive correlation with overall survival and disease-free survival in patients with pancreatic ductal adenocarcinoma.