1 Introduction

The development of metastasis is the main cause of death for prostate cancer patients and thus is a major obstacle to the successful treatment of these patients. However, the molecular aspect of  metastatic development is as yet poorly understood, mainly because metastasis is a highly complex process and involves a  variety of positive and negative factors^[1]. The KAI1 gene located in the p11.2 region of human chromosome 11 was first isolated by Dong et al in 1995^[2] as a prostate-specific tumor metastasis suppressor gene. CD82 was primarily identified by cDNA cloning as the R2 antigen in mitogen-activated human T cells and was subsequently cloned as IA4 and C33 antigens^[3-6]. The KAI1 gene product is identical to CD82 and is designated as KAI1/CD82, which has four hydrophobic and presumably transmembrane domains and one large extracellular N-glycosylated domain^[1]. It appears to function in cell-cell adhesion and cell-extracellular matrix interaction, thereby potentially influencing the ability of cancer cells to invade tissues and to metastasize. Down-regulation of KAI1/CD82 expression has been shown to be associated with metastasis or tumor progression in prostate^[7-9], lung^[10,11], breast^[12,13], bladder^[14], liver^[15,16] and pancreas cancer^[17].

The purpose of our study was to evaluate KAI1/CD82 expression in Chinese patients with benign prostatic hyperplasia (BPH) and late-stage carcinoma of prostate (CaP) by means of immunohistochemical analysis.

2 Materials and methods

2.1 Tissue samples

Prostatic tissue samples from 30 patients with  BPH, 55-87 years of age (mean: 67.5)  were obtained by suprapubic or transurethral prostatectomy.  Tissues from 34 patients with CaP, aged 32 to 89 years (mean: 68.1) were obtained from needle biopsy before anti-cancer treatment. The patients were recruited into the two hospitals between October 1996 and December 1998.

Tissues were stored at -80 before dealing with.  Sections (6 m) were cut on a cryostat, mounted on aminopropyl-triethoxysilane-coated slides, and air-dried. Slides were stained with hematoxylin and eosin and observed under the microscope. According to Gleason^[18] and the Prout GR^[19], the tumors of 11 patients were well differentiated (Gleason  Grades 2 to 4),  8 moderately differentiated (Grades 5 to 7), and 15 poorly differentiated (Grades 8 to 10). Fifteen patients were at stage C and nineteen at  stage D .

2.2  Immunohistochemistry

Immunohistochemical staining was performed as described by Ueda et al^[9] with minor modifications. The sections were fixed with acetone for 10 min at 4, and air-dried. All subsequent steps were performed at room temperature. They were incubated with methanol containing 0.3% hydrogen peroxide for 30 min to block endogenous peroxidase activity. After treatment with phosphate-buffered saline (PBS) containing 5% normal rabbit serum for 30 min, the sections were incubated with mouse monoclonal anti-human KAI1/CD82 antibody at a dilution of 1:200 in PBS for 2 h in a moist chamber. After  treatment with biotinylated rabbit anti-mouse immunoglobulin (Dako) diluted 1:400 in PBS for 30 min, the sections were incubated with biotin-streptavidin-peroxidase complex (Dako) for 30 min, followed by a 3,3'-diaminobenzidine tetrahydrochloride and hydrogen peroxide mixture. The sections were counterstained with hematoxylin, dehydrated in graded ethanol, cleared in xylene, and then mounted. As the negative control, non-immunized purified rat IgG2a  was used instead of the anti-human KAI1/CD82 antibody.

2.3 Specimen classification

Staining intensity in the cancer cells was estimated as positive when it appeared to be similar to that of glandular cells of BPH. In all specimen, adjacent noncancerous cells were used as internal positive controls for KAI1/CD82 exfpression. When the relationship between KAI1/CD82 expression and the Gleason grade were examined, the percentage of positively stained cells was calculated in 500 cells showing the same Gleason grade. When the relationship between the KAI1/CD82 expression and the clinical stage was estimated, the presence of 50% or more KAI1/CD82 positive cancer cells within the tumor tissue were classified as KAI1/CD82 positive, 5% to <50% as KAI1/CD82 reduced, and 0% to <5% as KAI1/CD82 negative. These judgements were counterchecked by three observers.

2.4 Statistical analysis

3 Results

3.1 Distribution and trait of KAI1/CD82 expression

In BPH tissues, KAI1/CD82 expression was all positive, and was located uniformly in the cellular membrane of glandular epithelial cells, especially at areas of cell-to-cell borders. KAI1/CD82 expression was not detected in the nucleus or cytoplasm (Figure 1). In late-stage CaP  tissues, KAI1/CD82 expression was either decreased or negative, which was also observed on the membrane at cell-to-cell borders,  but the immunostaining pattern along cell-to-cell borders was not continuous (Figure 2,3).

Figure 1. Immunohistochemical staining of KAI1/CD82 expression in BPH tissue. 300
Figure 2. Immunohistochemical staining of KAI1/CD82 expression in tissue of CaP. (well differentiated). 300
Figure 3. Immunohistochemical staining of KAI1/CD82 expression in tissue of CaP. (poorly   differentiated). 300

3.2 Correlation between KAI1/CD82 expression and pathologic grade and clinical stage

As shown in Table 1, a significant inverse correlation existed between KAI1/CD82 expression and Gleason Grade (P<0.05). There was no statistically significant inverse correlation between KAI1/CD82 expression and the clinical stage (Table 2).

Table 1.  Relationship between KAI1/CD82 expression and pathological grade  in CaP.

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