The present study demonstrates that MMP-9 is overexpressed and TIMP-1 and RECK are underexpressed in malignant bladder tissues compared with normal bladder tissues. We demonstrated also that MMP-2 and its regulators (TIMP-2, MMP-14 and IL-8) are underexpressed in BC. Most importantly we found increased levels of MMP-9 and IL-8 in high grade, invasive and recurrent tumors.
Extensive studies have revealed that malignant invasion and metastasis require ECM degradation, mainly by MMPs [11
]. Excessive or inappropriate expression of MMPs may contribute to the pathogenesis of cancer in a wide variety of cases by facilitating tissue degradation. Currently, there are more than 20 identified MMPs that can be categorized by substrate specificity. Despite the clinical significance of the pathogenetic impact of MMPs in human cancer, including BC, only a few studies of MMPs are available in the literature, and those mainly analyze protein expression [12
In the present study, we have shown not only that MMP-9 is overexpressed in cancer tissue, but also that MMP-9 levels are significantly higher in patients with high grade and infiltrative tumors, two very important prognostics factors.
The same occurred considering IL-8 expression, besides the fact that IL-8 is underexpressed in cancer tissue, the relation between higher IL-8 expression and unfavorable prognostic characteristics could be related to the fact that IL-8 is important for tumor progression, but not for tumor initiation or promotion.
MMP-9 and IL-8 expression levels increased together with tumor invasiveness, suggesting that they are probably associated with a worse outcome in BC. This is the first time that a study has shown a correlation between high grade, invasive tumors and IL-8 mRNA expression in BC. Increased MMP-9 expression has also been shown to have an independent prognostic impact on operable non-small cell lung cancer [13
] and renal cell carcinoma [14
], and increased IL-8 expression has been associated with worse prognostic of colon cancer [15
Another important finding of our study was the relationship between MMP-9 and IL-8 higher levels and tumor recurrence during a long follow-up period (mean of 40
months). The mean expression of MMP-9 and IL-8 in patients with recurrence was greater than six times that observed in patients without recurrence. This result might reflect an important biological phenomenon that could be confirmed in larger studies and that could become a tool to identify patients likely to progress. MMPs are abundantly expressed in malignant tumors regardless of their origin, and a significant correlation between increased MMP expression and a poor prognosis in terms of survival has been demonstrated in several cases [16
]. As a result, the possibility of MMPs being used as tumor markers has been suggested.
At the post-translational level, all MMPs are under the control of specific tissue inhibitors (TIMPs) that bind proximal to the catalytic domain of MMPs, preventing substrate attachment. TIMPs are not simply regulators of MMP activity; they also have multifunctional roles that include cell growth promotion and inhibition of angiogenesis [18
]. Four TIMPs have been identified. They inhibit all MMPs, forming non-covalent complexes with the active forms. Among them, TIMP-1 selectively binds pro-MMP-9 and is considered the main inhibitor of MMP-9, TIMP-2 binds pro-MMP-2. The reversion-inducing cysteine-rich protein with Kazal motifs (RECK) gene was identified as an inducer of a flat morphology in v-Ki ras–transformed NIH3T3 cells [19
]. RECK expression is observed in normal human organs; however, several oncogenic factors, such as activated Ras [20
], EBV latent membrane protein 1 [21
], and HER-2/neu [22
] suppress the expression of RECK. Additionally, it has been reported that RECK overexpression decreases the amount of active MMP-2 and MMP-9 in conditioned medium and inhibits metastatic activity in vitro
] and in vivo
]. Considering activation of MMPs, MT1-MMP is a key enzyme in tumor angiogenesis and metastasis: it hydrolyzes a variety of ECM components, and is a physiological activator of pro-MMP-2 and MMP-9. IL-8 is involved in the transcription induction of MMP-2 increasing stromal invasion by tumor cells facilitating angiogenesis and development of metastasis.
Interestingly, our results demonstrate that there is an upregulation of MMP-9 and a downregulation of its specific inhibitors, TIMP-1 and RECK, in BC. Otherwise, we demonstrated that the low expression of TIMP-2, MMP-14 and IL-8 may be responsible for the decreased MMP-2 expression in BC tissue.