Interlink between cholesterol & cell cycle in prostate carcinoma

Abstract

Background & objectives: Earlier reports have shown hypocholesterolaemia in cancer patients and high number of lipid rafts in cancer cells. The primary objective of this study was to compare the intracellular cholesterol turnover in non-cancerous (benign) prostatic hyperplasia (BPH) and carcinoma prostate (CAP) with normal prostate cells obtained from patients undergoing radical cystectomy for carcinoma bladder (sham control).

Methods: ELISA-based estimation of prostate-specific antigen (PSA), evaluation of expression of low-density lipoprotein receptor (LDLR), peripheral-type benzodiazepine receptor (PBR) and cyclin E, immunohistochemistry and confocal microscopy, measurement of integrated optical density of the diaminobenzidine (DAB)-stained immunohistograms, isolation of nucleus and cell cytoplasm from prostate tissue by ultracentrifugation followed by estimation of cholesterol spectrophotometrically in isolated nuclear and cytoplasmic fractions were performed.

Results: Seventy five individuals, 25 for each group (BPH n=25; CAP n=25 and sham control n=25), were included in the study. Cholesterol was increased in the cytoplasm and nucleus of the prostate cancer cells along with elevated expression of LDLR. Increased cholesterol concentration in the cell nucleus was found comparable with the increased expression of cholesterol transporter viz. PBR in the prostate tumour tissues as compared to its expression in normal prostate cells obtained from individuals undergoing radical cystectomy for carcinoma bladder. Cell cycle protein cyclin E was also highly expressed in cancer tissues.

Interpretation & conclusions: The present findings along with increased expression of cell cycle protein cyclin E in the cell nucleus of the tumour tissue suggested the possibility of an intriguing role of cholesterol in the mechanism of cell cycle process of prostate cell proliferation.

Keywords: Cholesterol - cyclin E - low-density lipoprotein receptor - peripheral-type benzodiazepine receptor - prostate cancer.

Fig. 1

Low-density lipoprotein receptor (LDLR) protein expression by immunohistochemistry (magnification ×200). DAB stain (diaminobenzidine; brown colour) depicts a significant increase in LDLR expression in carcinogenic and non-carcinogenic (benign) tumours. Arrows indicate areas of higher expression. In the graph each bar shows the mean±standard deviation of 25 samples, represented as IOD (integral optical density) from the respective category. ***P<0.001 compared to sham control.

Fig. 2

Peripheral-type benzodiazepine receptor (PBR) protein expression by immunohistochemistry (magnification ×200). DAB stain (brown colour) depicts a significant increase in PBR expression in carcinogenic and non-carcinogenic (benign) tumours. Arrows indicate areas of higher expression. In the graph each bar shows the mean±standard deviation of 25 samples, represented as IOD. ***P<0.001 compared to sham control.

Fig. 3

Marker assay in the isolates of ultracentrifugation. (A) DNA (nuclear marker), and (B) lactate dehydrogenase, LDH (cytosolic marker) were measured to determine the purity of the fractions. Each bar shows the mean±standard deviation of 25 samples. P*<0.05, **<0.01, ***<0.001 compared to respective control.

Fig. 4

Measurement of cholesterol content. The concentration of cholesterol in cell nucleus and cytoplasm was found higher in cancer prostate tissue as compared to control prostate. The data show the mean±standard deviation (mg cholesterol/g prostate tissue) of 25 samples. *P<0.05 compared to respective control. Nuclear-cholesterol ratio in cancer prostate tissue to control prostate: 1.8.

Fig. 5

Nuclear localization of cyclin E by confocal microscopy. Cyclin E was labelled by fluorescein isothiocyanate (green) and nucleus was stained with propidium iodide (red) (Magnification ×400). The merged image (yellow-coloured) shows the nuclear localization of cyclin E.

Fig. 6

Expression of cyclin E on prostate tissue detected by immunohistochemistry (magnification ×200) of cyclin E protein. The IOD of DAB stain taken by cyclin E protein shows a significant increase of cyclin E expression in carcinogenic and non-carcinogenic (benign) tumours. Increased expression was observed in the carcinoma prostate group. Arrows indicate areas of higher expression. Each bar shows the mean±standard deviation of 25 samples. P**<0.01, ***<0.001 compared to sham control.