Expressions of inducible nitric oxide synthase and matrix metalloproteinase-9 and their effects on angiogenesis and progression of hepatocellular carcinoma

Abstract

Aim: To determine the expressions of inducible nitric oxide synthase (iNOS) and matrix metalloproteinase-9 (MMP-9) in hepatocellular carcinoma (HCC) and to investigate the relationship between iNOS and MMP-9 expression and their effects on angiogenesis and progression of HCC.

Methods: In this study, we examined iNOS, MMP-9, and CD34 expression in specimens surgically removed from 32 HCC patients and 7 normal liver tissues by immunohistochemical staining. Meanwhile, microvessel density (MVD) was determined as a marker of angiogenesis by counting CD34-positive cells.

Results: The positive rates of iNOS and MMP-9 expression were 71.88% (23/32) and 78.13% (25/32) in HCC. MMP-9 expression was significantly correlated with tumor size, capsule status, TNM stage, and risk of HCC recurrence (P = 0.032, P = 0.033, P = 0.007, and P = 0.001, respectively). There was also a significant relationship between iNOS expression and capsule status and risk of HCC recurrence (P = 0.049 and P = 0.004, respectively), but no correlation between iNOS expression and tumor size and TNM stage. There was a positive association between MVD and TNM stage and risk of HCC recurrence (P = 0.037 and P = 0.000, respectively). The count of MVD was significantly different in different iNOS and MMP-9 immunoreactivity groups (F = 17.713 and 17.097, P = 0.000 and P = 0.000, respectively). The examination of Spearman's rank correlation coefficient showed that there was a significant positive correlation between MVD and iNOS, MMP-9 immunoreactivity (r = 0.754 and 0.751, P = 0.000 and P=0.000, respectively). There was also a significant association between MMP-9 and iNOS expression in HCC (P = 0.010).

Conclusion: Nitric oxide (NO) produced by iNOS could modulate MMP-9 production and therefore contribute to tumor cell angiogenesis and invasion and metastasis in HCC. The strong expression of iNOS and MMP-9 in HCC may be helpful in evaluating the recurrence of HCC, predicting poor prognosis. For patients with strong expression of MMP-9 and iNOS, the optimal treatment scheme needs to be selected.

Figure 1

Immunohistochemical staining for CD34 in HCC (×100).

Figure 2

Immunohistochemical staining for MMP-9. A: MMP-9 in cytoplasm and cytoplasmic membranes in HCC (×400); B: MMP-9 in vascular endothelial cells, bile ducts, HCC cells, and stromal fibroblasts (arrow heads from top to bottom) within HCC (×200); C: MMP-9 expression in neighboring capsule of HCC (×200); and D: weak expression of MMP-9 in central areas of HCC tissues (×200).

Figure 3

Immunohistochemical staining for iNOS. A: iNOS expression in tumor cytoplasm (×100); B: strong expression of iNOS in neighboring capsule of HCC tissues (×400); and C: weak expression of iNOS in central areas of HCC tissues (×400).