Wayne K. Nelson and Dr. Merrill Christensen, Food Science and Nutrition
According to the National Cancer Institute, prostate cancer is the most frequently diagnosed nonskin cancer in men. In 1997 alone, it is estimated that 209,900 men were diagnosed as having the disease. Prostate cancer is also the second leading cause of cancer related death in the United States and, in 1997, claimed the lives of an estimated 41,800 people (1).
It has recently been discovered that selenium supplementation in men reduces the incidence of prostate cancer (2). This has led researchers to study the metabolism of selenium in the human body to find possible mechanisms of action for cancer prevention. For years our laboratory has studied selenium regulation of gene expression and has made exciting discoveries in the field of cancer research.
NF-κB is a combination of five transcription factors that code for apoptosis (programmed cell death), cell proliferation, and responses to stress. They are found in the cytoplasm of prostate cells bound to inhibitory proteins which prevent NF-κB from entering the nucleus and turning on the expression of DNA – including the expression of anti-apoptotic proteins. As long as NF-κB is bound, the cells are safe from its effects. However, when the inhibitory proteins are exposed to oxidative stress by pro-oxidants, they are phosphorlized. Then protein-bound NF-κB is released and goes into the nucleus to turn on the anti-apoptotic genes. Anything that inhibits apoptosis, like NF-κB, will induce cancer-like, unregulated cell growth and division. Indeed, many types of cancer are characterized by a loss of apoptosis.
Selenium-dependant glutathione peroxidase is an outstanding anti-oxidant that provides protection from pro-oxidants that produce oxidative stress on the cell. Consequently, as the amount of anti-oxidants in a cell increases, the proportion of NF-κB that is bound should increase as well. In this manner gene expression is inhibited. By this mechanism, it is probable that selenium inhibits NF-κB from entering the nucleus in prostate cancer cells (LNCaP Cells). It is important to note that the effects of selenium supplementation on NF-κB transcription factors have never been studied.
I set out to show that Selenium’s effect on the reduction of pro-oxidants changes NF-κB binding in prostate cancer cells and is partly responsible for selenium’s chemopreventive effects. The study was designed to demonstrate:
• That the amount of NF-κB bound is dependant on selenium availability.
• That selenium affects the binding of NF-κB in cells that are growing in traditional medium as well as cells that are growing in medium that has pro-oxidants added to it.
• That selenium regulation of NF-κB binding is dependant on the form of selenium supplemented.
When doing research, it is essential that work done on cell lines is relevant to a human model. In research performed previously, we have found that Se levels of 100ng/ml in vitro is the highest level of Se that can be added to medium without the LNCaP cells showing signs of toxicity. This correlates well with levels found in the blood of people supplementing their diet with Se. It was more challenging however, to find the appropriate adequate level of Se for the control groups in the study so we could compare them to the treatment groups receiving high levels of Se supplementation.
Of all the proteins in the human body that contain Se, glutathione peroxidase or GPX is most sensitive marker of Se availability. It is the first have its levels decline in Se deficiency and the last to reappear again when adequate Se is provided. With this knowledge, a complex experiment was set up using low levels of Se. The cells were collected and the GPX levels in each group were determined. From the data we decided that a level of 4ng/ml would be used in the main experiment as a control.
With treatment and control levels determined, we seeded 28 groups of 2 flasks, let them seed for 48 hours, then replaced the medium with medium that had Se supplementation (one of two forms) in one of the two levels. In addition we added the pro-oxidants hydrogen peroxide and TNF to some of the groups. The cells were treated an additional 24 hours after which they were removed from the medium and frozen.
The nuclear proteins from each group have been extracted and are currently undergoing analysis by way of an electrophretic mobility shift assay. The early results are promising but not enough information has been gathered to warrant a conclusion. When that is possible, the results of this experiment will be submitted for scientific review and publication.
References
- Recent Trends in Prostate Cancer Incidence and Mortality. National Cancer Institute.http://cancernet.nci.nih.gov/cgibin/srchcgi.exe?DBID=pdq&TYPE=search&SF MT=pdq_statement/1/0/0 &ZUI=600630.
- Clark LC, Dalkin B, Kongrad A, et al. Decreased incidence of prostate cancer with selenium supplementation: results of a double-blind cancer prevention trial. Br J Urol 1998; 81:730-4.