Overexpression of 15-lipoxygenase-1 in PC-3 human prostate cancer cells increases tumorigenesis

Abstract

The effect of overexpression of 15-lipoxygenase-1 (15-LO-1) was studied in the human prostate cancer cell line, PC-3. Stable PC-3 cell lines were generated by transfection with 15-LO-1-sense (15-LOS), 15-LO-1-antisense (15-LOAS) or vector (Zeo) and selection with Zeocin. After characterization by RT-PCR, western and HPLC, a PC3-15LOS clone was selected that possessed 10-fold 15-LO-1 enzyme activity compared with parental PC-3 cells. The PC3-15LOAS clone displayed little or no 15-LO-1 activity. These PC-3 cell lines were characterized for properties of tumorigenesis. The proliferation rates of the cell lines were as follows: PC3-15LOS > PC-3 = PC3-Zeo > PC3-15LOAS. Addition of a specific 15-LO-1 inhibitor, PD146176, caused a dose-dependent inhibition of proliferation in vitro. Overexpression of 15-LO-1 also caused [(3)H]thymidine incorporation to increase by 4.0-fold (P < 0.01). Compared with parental and PC-3-Zeo cells, PC3-15LOS enhanced whereas PC3-15LOAS reduced the ability of PC-3 cells to grow in an anchorage-independent manner, as assessed by colony formation in soft agar. These data suggested a pro-tumorigenic role for 15-LO-1 in PC-3 cells in vitro. Therefore, to clarify the role of 15-LO-1 in vivo, the effect of 15-LO-1 expression on the growth of tumors in nude mice was investigated. The PC-3 cell lines were inoculated subcutaneously into athymic nude mice. The frequency of tumor formation was increased and the sizes of the tumors formed were much larger in the PC3-15LOS compared with PC3-15LOAS, parental PC-3 and PC-3-Zeo cells. Immunohistochemistry for 15-LO-1 confirmed expression throughout the duration of the experiment. The expression of factor VIII, an angiogenesis marker, in tumor sections was increased in tumors derived from PC3-15LOS cells and decreased in those from PC3-15LOAS cells compared with tumors from parental or Zeo cells. These data further supported the evaluation by ELISA of vascular endothelial growth factor (VEGF) secretion by PC-3 cells in culture. Secretion of this angiogenic factor was elevated in PC3-15LOS cells compared with the other cell lines. These results support a role for 15-LO-1 in a novel growth-promoting pathway in the prostate.

Conditional Expression of 15-Lipoxygenase-1 Inhibits the Selenoenzyme Thioredoxin Reductase

“…Approximately 1 ϫ 10 4 cells were plated on an 85-mm dish coated with 1% agar, and maintained at 37°C. On day 10, cells were stained with 1 ml of 2-(p-isodophenyl)-3-(p-nitrophenyl)-5-phenyltetrazolium chloride hydrate (250 mg/l in PBS) for 24 h (24).…”

“…Expression of 15-LOX-1 correlates with tumorigenesis of PC-3 prostate cancer cells in vivo (24,29), yet exogenous 15(S)-HETE blocks the proliferation of these same cells in vitro (30,31). 15(S)-HETE arrests the growth of HT 29 colon cancer cells in some investigations (32) but not in others (33) arrests growth of CaCo-2, HT29, or RKO colon cancer cells.…”

Where is VEGF in the body? A meta-analysis of VEGF distribution in cancer

“…Studies with aberrant data (over a 10-fold difference than those reported in other papers) were also excluded. Based on these criteria, 12 studies were excluded from our collection (Liu et al, 1999;Haggstrom et al, 2000;Bhujwalla et al, 2001;Feldman et al, 2001;George et al, 2001;Huss et al, 2001;Kelavkar et al, 2001;Calvo et al, 2002;Mabjeesh et al, 2003;Cianchi et al, 2004;Singh et al, 2004;May et al, 2005) (Supplementary Table S6). …”

Inhibition of arachidonic acid metabolism and its implication on cell proliferation and tumour-angiogenesis

“…With regards to 15-LOX, there is evidence for opposing theories on correlation of expression levels with carcinogenesis where both over-and under-expression has been observed in cancerous cells [70,71]. However, an increasing number of investigations seem to indicate that 15-LOX-1 is positively, whilst 15-LOX-2 is negatively correlated with cell proliferation and carcinogenesis.…”