Scientists have developed methods that can identify the genes involved in cholesterol and fat processing; these tools can also be used to discover new cholesterol-controlling drugs. Their results will help identify genes responsible for atherosclerosis, diseases associated with bile secretion in the liver, and some cancers.
The principal investigators, Steven Farber of Marnie Halpern's laboratory at the Carnegie Institution of Washington's Department of Embryology and Michael Pack of the University of Pennsylvania, report their results in today's issue of Science.
Currently, there are only a few ongoing genetic screens that study vertebrate physiology. The screening method devised in this study represents an important milestone for the real-time observations of lipid transport and processing in live animals.
The scientists used the tiny, clear zebrafish larva for their research because its transparency allows them to watch what happens when fat and cholesterol are digested.
They fed 5-day-old larvae a custom-synthesized phospholipid that emits a bright green fluorescence when digested. Phospholipids are fats that form cell membranes and potent cellular messengers. The labeled lipids are able to reveal lipid-processing enzymes that are produced by specific genes in the digestive system.
The researchers used the custom lipids to screen zebrafish families that carried a host of unknown mutations. The goal of the screen was to find larvae whose digestive organs exhibited an atypical fluorescence after eating the lipids. This indicates that an important gene for lipid processing is mutated.
In their Science paper, the team describes one such mutation, which they named fat free. Fish containing a mutation in the fat free gene appear to have normal digestive organs when examined under the microscope, but fail to accumulate the fluorescent lipid.
These mutant fish seem to process fat and cholesterol at a significantly lower rate, indicating that the fat free gene may be a regulator of dietary cholesterol use.
What does this mean for human lipid disorders?
The investigators suggest that drugs that interfere with the fat free gene may be a useful way to control high cholesterol in people. They also claim that zebrafish should be as good as mice for identifying genes that are involved in human lipid metabolism and disease. This claim is based on an experiment they conducted incubating fish with atorvastatin (Lipitor, by Parke-Davis).
Lipitor, a widely prescribed drug used to treat hypercholesterolemia, is an inhibitor of cholesterol synthesis. Fish on Lipitor were barely able to digest the green fluorescent phospholipid, suggesting that the gall bladder did not release bile, which is necessary for fat digestion.
The scientists believe that the drug works by inhibiting the synthesis of cholesterol-derived bile that is required for fat breakdown. A major effort to identify the molecular identity of the fat free gene is now underway in the Farber laboratory in the Kimmel Cancer Center at Thomas Jefferson University.
Farber notes, "These reagents used in conjunction with the transparent zebrafish larvae enable us to visualize lipid biochemistry in living animals and ask what genes are the key players. These kinds of experiments are exceedingly difficult to accomplish in other vertebrate systems."
This work is supported by the National Institutes of Health, the Deutscher Akademischer Austauschdienst, the March of Dimes, the Pew's Scholar Award and the Carnegie Institution of Washington.
Related website:
The Carnegie Institution of Washington
[Contact: Steven Farber, Michael Pack, Marnie Halpern, Tina McDowell]
18-May-2001
