Food for Thought
Safety Assessment
Regina Kelder

Food for Thought

Mouse diets and cancer studies; watching our immune cells in action. Eureka's live coverage of SOT2016 continues. The third of a four-part series.

Last October, the International Agency for Research on Cancer, a division of the World Health Organization, shocked the global community by adding processed meats to a list of substances it considers cancer-causing. Its recommendation was based, partly, on studies from mice and rats that concluded nitrosamines might play a major role in the formation of colon cancer.

How did science learn this and are the findings reliable? For over 50 years, researchers have largely depended on male and female mice and rats to determine whether chemicals in foods, drugs or other products cause cancer. Two-year carcinogenicity studies in laboratory rodents are the primary method that drug developers use to determine if a test article, such as a drug, is hazardous to humans. These studies in rodents, along with epidemiology studies are the most reliable ways, currently, for identifying potential human hazards.

But for animal carcinogenicity studies to be informative the studies need to achieve their endpoints, and there can't be any confounding factors that cloud results. One of the known confounding factors in toxicology studies is chronic progressive nephropathy (CPN), a spontaneous renal disease in rats that causes tumors and which appears to be enhanced by the high protein diets that many vivariums feed their rodents. With these concerns in mind, a retrospective study led by Mark Morse, Director of Research at Charles River's Safety Assessment site in Spencerville, Ohio, set out to compare the effects of two certified rodent diets used in carcinogenicity studies, one containing a protein content of around 20% and the other with a content of around 14%. The findings are being presented today at SOT.

Morse's study looked at survival, tumor incidence, and the incidence of (CPN) in the rats and mice. He ultimately found that the tumor profiles were similar in the two groups, though the lower-protein diet was associated with greater survival rates in male rates and lower rates of CPN in both mice and rats. Further study is necessary to determine the influence the two different diets might be having on the outcomes of the carcinogenicity studies.

A separate study presented on Wednesday by Peter Mansell, Associate Scientific Director of the General Toxicology Department at Charles River's Safety Assessment site in Montreal, also dealt with carcinogenicity testing. With different animal models being added to the mix and studies becoming shorter, Mansell's poster, drawing from data collected over 20 years, nonetheless found the CD1 model used in two-year carcinogenicity studies still relevant, reliable and capable of delivering consistent results.

Live Action Shorts

Suppose you were that proverbial fly on the wall, able to spy on immune cells as they gobbled up bacteria in liver or lung endothelia?

That's the power of intravital microscopy (IVM), tools that grew out of the era of fluorescent protein technology and live cell microscopy and which have become essential tools in animal science for the study of subcellular processes, particularly in tumor biology, neurobiology and immunology. IVM is performed under conditions that closely resemble the natural environment.

One of the leading experts on IVM is Paul Kubes, Director of the Synder Institute for Chronic Diseases in Calgary, who is using spinning disc confocal fluorescence microscopy and other forms of IVM to study inflammation. Mouse transgenic and knockout technology can be used to delineate molecular mechanisms underlying the recruitment of leukocytes to areas of inflammation, leading Kubes to pursue a clearer understanding of the speed bumps that immune cells use to prevent bacteria from moving through the vasculature. His team devised a multi-channel fluorescence spinning-disk confocal microscopy device that has enabled them to image neutraphils, macrophages and other soldiers of the immune system that patrol the circulatory systems of mice.

A pre-recorded discussion of Kubes' work, which included short films of how different immune cells engage with platelets and bacteria at sites of inflammation in the lung and liver, were shown during Tuesday's exhibitor's session "Inflammation: Discovery to Toxicology." The session was led by Rana Samadfam, the Scientific Director of In Vivo Pharmacology at Charles River's Safety Assessment site in Montreal.

Spinning disk confocal systems use a rotating disk with a pattern of slits or holes installed in a microscope plane that is temporarily united with the specimen. The opaque areas of the disk block out-of-focus light from both the excitation and emission light paths. The spinning disks capture images much faster than standard confocal microscopy. IVM is also performed.

In 2011, Kubes was the first to show direct visualization of endogenous platelet recruitment to liver tissue and two years later he and his colleagues imaged the mouse liver to discover a patrolling role for blood platelets and their interaction with Kuppfer cells (KCs)—immune cells found in the liver sinusoid. Platelets, via the platelet-adhesion receptor GPIb, formed transient 'touch-and-go' interactions with von Willebrand factor (vWF) expressed on KC. Bacteria like methicillin-resistant staphyloccous aureus (MRSA) were rapidly caught by KCs and triggered platelets to switch from 'touch-and-go' adhesion to sustained GPIIb-mediated adhesion on the KC surface to encase the bacterium. Most recently, Kubes used IM of the peripheral knee vasculature to study transmigration of the Lyme spirochete in living Cd1d-/-mice.

How to cite:

McEnery, Regina. Food for Thought. Eureka blog. Mar 16, 2016. Available: