Elsevier

European Journal of Cancer

Volume 40, Issue 18, December 2004, Pages 2804-2811
European Journal of Cancer

Expression of heat-shock protein Hsp60 correlated with the apoptotic index and patient prognosis in human oesophageal squamous cell carcinoma

https://doi.org/10.1016/j.ejca.2004.08.013Get rights and content

Abstract

Cellular stress response and apoptosis are two highly conserved mechanisms for maintaining homeostasis. Hsp60 and Hsp90 have been shown to play pro- and anti-apoptotic roles, respectively. Our present study examined whether there is a correlation between the expression of Hsp60 and Hsp90, clinical parameters, the apoptotic index (AI), and the prognosis of patients with oesophageal squamous cell carcinoma (ESCC). We immunohistochemically stained cells for Hsp60, Hsp90, and single-stranded DNA (ssDNA), which acts as an apoptotic marker. In normal oesophageal epithelium tissue, Hsp60 and Hsp90 were expressed in the cytoplasm and membrane from the basal cell layer to the supra-basal cell layers. Hsp60 and Hsp90 positive stainings (+) were found in 63 of 123 cases (51%) and 62 of 123 cases (50%), respectively. There was no correlation between Hsp60 and Hsp90 expression levels and any of the clinical parameters examined. The five-year survival rate for ESCC patients with Hsp60 (+) expression was significantly higher than for those patients with Hsp60 (−) expression (P = 0.0371). Five-year survival rates of patients with Hsp60 (+) and (−) were 49% and 33%, respectively. By contrast, Hsp90 expression failed to predict patient prognosis (P = 0.7965). The high-AI group did not have a significantly better prognosis than the low-AI group (P = 0.2218). Statistical analysis showed a significant correlation between the expression of Hsp60 and AI in ESCC patients (P = 0.008). Thus, the five-year survival rate for the high-AI/Hsp60 (+) group was statistically significantly better than for the other groups (P = 0.0281). The results obtained in this study indicate that positive Hsp60 expression is a good prognostic indicator. This may be due to its role as a chaperone in contributing to the induction of apoptosis. These data suggest that Hsp60 expression correlates with the AI and patient prognosis in human ESCC.

Introduction

The cellular response to stress is represented at the molecular level by the induced synthesis of the heat-shock proteins (Hsp) as an essential defense mechanism for the protection of the cell from many harmful conditions, such as heat shock, alcohol, heavy metal, oxidative stress, fever or inflammation [1], [2]. During carcinogenesis, Hsp has been reported to show alteration of its expression level, either increasing or decreasing [3], [4]. Hsp has been classified into six major families according to the size of the molecule: small heat-shock protein, Hsp40, Hsp60, Hsp70, Hsp90, and Hsp100.

Hsp60 is abundant in most mammalian cells under normal conditions [5]. It has major roles in protein chaperoning and protein folding [6]. Hsp60 is a mitochondrial protein that is involved in the activation of apoptosis [7]. Its overexpression has been reported in exo-cervix [8] and colorectal carcinogenesis [9], suggesting that it may be involved in early carcinogenesis.

Hsp90 is associated with the folding of signal-transduction proteins, such as steroid hormone receptors and protein kinases. Hsp90 is an essential cytosolic protein; its expression in a wide variety of malignant tumours makes Hsp90 a candidate for pharmacological intervention [10]. Recently, several mechanisms by which Hsp90 acts as an anti-apoptotic factor have been reported [11]. Overexpression of Hsp90 in breast tumour, lung cancer, leukaemia, Hodgkin’s disease, pancreatic carcinoma, and gastric cancer [12], [13], [14], [15], [16], [17] has been observed.

In oesophageal cancer, the major risk factors for oesophageal squamous cell carcinoma (ESCC) are diet, tobacco use, and alcohol consumption [18], [19]. Hsp expression has been reported to correlate with prognosis and lymphocyte infiltration in ESCC [20], [21]. Based on these considerations, we performed an immunohistochemical study on Hsp60 and Hsp90, and used single-stranded DNA (ssDNA) as a marker for apoptosis. In this study, we examined whether there is a correlation between Hsp60 and Hsp90 expression levels, clinical parameters, apoptotic index (AI), and prognosis in patients with ESCC.

Section snippets

Patients

Surgical specimens were obtained from 123 patients (106 males and 17 females) who had ESCC and underwent potentially curative surgery at the Department of General Surgical Science, Gunma University, between 1983 and 2002. The patients’ age ranged from 40 to 79 years, with a mean of 61.2 years. Tumour stage and disease grade were classified according to the 5th edition of the TNM classification of the International Union against Cancer (UICC) [22]. The evaluation of tumour differentiation was

Hsp60, Hsp90, and ssDNA expression

In normal oesophageal epithelium tissue, Hsp60 and Hsp90 were expressed in the cytoplasm and membrane from the basal cell layer to the supra-basal cell layers (Figs. 1(a) and (b)). Immunostaining of Hsp60 and Hsp90 were seen in the cytoplasm and membrane of the cancer cells. Hsp60 expression (+) was observed as a diffuse staining of tumour cell cytoplasm and membrane as coarse granules (Fig. 1(c)). Hsp90 expression (+) was evident as diffuse cytoplasm staining (Fig. 1(d)). Positive staining for

Discussion

Heat-shock proteins can aid or inhibit the apoptotic machinery through their role as chaperones by affecting protein assembly, folding, and the ubiquitin degradation pathways. Hsp60 is a protein that primarily localises in the matrix of mitochondria. Kawanishi and colleagues [20] observed heterogenous/mosaic reduction of Hsp expression in a colony among cells that could not be distinguished morphologically. This may imply that their alteration is not due to genomic changes. One possibility for

Conflict of interest statement

None declared.

Acknowledgement

This work was supported by Grants-in-Aids from The Ministry of Education, Culture, Sports, Science and Technology, Japan for A.F.

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