PHAR 303 Lecture Notes - Lecture 6: Ingrid Newkirk, Canada Council, Lymph Node
11 May 2018
School
Department
Course
Professor
Lecture 6 (Jan 25): In vitro Toxicology: Use of Cell Lines & Artificial Organs; the 3Rs
In vivo Toxicology
• Animal biology
• High doses are used
• Low throughput
• Very expensive 80-300 000$
• Time consuming
• Relatively large number of animals & ethical issues
• Only measure a few apical endpoints
The Drivers of In Vitro Toxicology
• Societal concerns
o People in the 70’s and 80’s put a lot of pressure on cosmetics &
pharmaceuticals to stop using animals for testing
• Knowledge – we now understand mechanisms better & can target specific receptors
• Money – animal test is very expensive compared to in vitro test that are more cheaper
• Speed – much faster
• Efficiency – done by robots
• Ethics – few animal use
Acceptance of Animals in Biomedical Research
• 75-80% people will accept animal use if it going to help human advance
• Decrease to less than 50% if pain is part of the protocol & the animal suffers
• In dealing with animal use is not just the animal but also the treatment of the animal,
the kind of food, toys in the cages, friends in the cages ( rats & mice like different
toys)
• The issue for the public is:
o Accountability for what we do with the animals
o No pain and distress of animals - On a yearly basis, the expectation of
eliminating pain and distress increases
• The issue for the scientific community:
o Increased effectiveness of IACUS’s (Institutional Animal Care & Use
Committees) – they regulate how we use animals
o Further enhance standard of care
Key events in in Vitro Toxicology
1824 – Society for the Prevention of Cruelty to Animals - oppose dissecting animals when
they were alive
1931 – Hall’s principles of ethical animal experimentation – first to formulize the idea that we
had to handle animals ethically.
1959 – Russell & Burch – Principle of Humane Experimental Technique – the basis of
everything we do today with respect with replacement, reduction.. all the issues about going
more towards in vitro is based on this book.
1980 – Henri Spira – Campaign against cosmetic companies – he pushed the cosmetics
industry to come up with ways of not using animals to test cosmetics. The cosmetics
companies put 1 million price to whoever found a better technique to stop using animal
testing – competition between colleges. This led to the creation of
1982 – Creation of Centre for Alternatives to Animal Testing & Johns Hopkins University
find more resources at oneclass.com
find more resources at oneclass.com
1985 – Institutional Animal Care Committees – every institutions had to create committees.
1989 – Creation of ECVAM : European Center for the Validation of Alternative Methods.
1994 - Creation of ICCVAM : Interagency Coordinating Committee on the Validation of
Alternative Methods
Legislation against cruelty
1876: Cruelty to Animals Act (UK)
1966: Animal Welfare Act (USA)
1986: EC Directive 86/609 (Europe)
In Canada, CCAC: The Canadian Council of Animal Care
• National organization responsible for setting and maintaining standards for the care
and use of animals in research, teaching and testing throughout Canada
• Council that has for mission to regulate the use & identifies standards.
• If you don’t follow their protocol, you are not certified for using animals & the
funding agencies cut your fund out and the funds of the entire university.
• Their guidelines are updated on regular basis and ethical considerations are based on
Marshall Hall principles regarding use of experimental animals – 1931
• An experiment should never be done if the necessary information can be obtained by
observation – make sure there’s a need of doing the study.
• No experiment should be done w/o a clearly defined & obtainable objective
• Scientists should be well informed to prevent duplication of previously done work.
• Experiments should be done with least infliction of suffering & use of least sentient
animals – stressed animals response differently than less stressed
o If hear another animal suffering or smells blood level of cortisol
• Every experiment should be done under conditions that provide the clearest possible
results – you know how to make predictions.
Concept of Surrogate Responsibility
To what extend should we be defending those who cannot defend themselves?
How far down the animal kingdom does that principle apply?
In some countries, birds are handle the same way as monkeys, in other it stops at mammals –
it really depends on the country regulations.
Confounding Sentimentalism
Ingrid Newkirk – President of People Ethical Treatment of Animals (extremist)
In McIntyre 14th floor, you can only enter if you have training on how to treat animals.
How do we balance the protection of animals, the health and wellbeing of man, concerns for
environment and safety for the future of the planet?
The 3R’s
• Refinement, Reduction, Replacement
• Developed by Russell & Burch in the Principles of Humane Experimental Tech.(1959)
• In you are going an experiment: refine to get the best results, reduce to a minimum nb.
of animals and if you can’t, replace the animals.
• Proper experimental design should consider techniques to:
find more resources at oneclass.com
find more resources at oneclass.com
1. Refine – experimental techniques to lessen pain or distress.
• People have done many studies to show that an animal that is in pain gives inaccurate
data (unless you study pain itself)
• Alternatives: Any modification of a procedure that potential pain or distress, or uses
animals lower in a phylogenetic category.
• If you can do the study on a rat and it gives similar results on cats (brain studies) –
better use rats.
• Experiments on dogs that study cardiovascular responses – the build of the dog’s
heart and the response to blood flow & electrical abnormalities are similar.
o Many studies that used to be on dogs are now done in mice and rats.
o Before going to human trials, then do it on the dog too.
o Some people argue that Drosophila & C.elegans (worms, flies) give us results
that are pretty similar to what happens in us.
o But, extrapolating from a fly to human is difficult.
Examples:
- Mouse local lymph node assay (allergic contact sensitization)
GPMT: Guinea pig maximization test
• In vivo test to screen for substances that cause human skin sensitization.
• Refine : time to perform assay reduced (32 days to 7 days)
number of animals (32-43 to 16-30)
Dermatitis induced & adjuvant(painful) required (initially yes, then no)
- Use of transgenic animals
Transgenic Animals:
• Specific target yields a decrease number of animals –knockouts genes,
• Decrease necessity for companion species
• Most transgenics require additional care.
- Non-invasive monitoring: MRI, light.
Different technologies: MRI, Ultrasound Imaging
CT: Computer Assisted Tomography – Anatomic information
• 3D X-ray technique 3D reconstruction and computer analysis
• 3D mouse images with a resolution of 100x100x100 microns obtained in a few min.
• Contrast generated by difference in tissue absorption.
PET: Positron Emission Tomography – Metabolic information
Optical Imaging
• Tag cell & gene, image, digitize, quantify, archieve.
• Camera in a box & can have any cells, markers, quantum dots, fluorescent (Luciferase
or green fluorescent protein) probe in the animal.
• Look at them in different times and see how they get metabolized & grow.
• Detection of tumors in vivo – see how it grows in response to a toxicant.
• Don’t have to kill the animal at each stage to see the growth.
find more resources at oneclass.com
find more resources at oneclass.com
