Module 3R    Replacement Author: Peter Maier
Replace and Replacement A A A
Welcome !
Module 3R
Draize test replacement
Sensitization in-vitro?
Pain
Tox receptor
Implementation of the 3Rs
Pyrogen test in vitro
Replacement
in vivo
Receptor
Humans - Animals
Subcellular
Cells
with/without culture
Primary cells
Hepatocytes 24h
Hepatocytes 3d
HepG2
Established cell lines
Structure
Single cells
Interaction
Liver co-cultures
Brain co-culture
Aggregate culture of liver cells
Aggregate cultures of brain cells
Epithelium
Future
Receptor modeling
3R-Bologna declaration
Authors
Continuing Education
Contact
A replacement method can seldom directly replace animal testing. However one or multiple replacement methods in combination, already provide so much information, that the findings from the animal testing are no longer necessary.
Inhaltsverzeichnis
When is animal testing necessary ?
Whenever an answer cannot be provided by animal free testing methods (according to the law in Switzerland: Animal Protectiona Act and Animal Protection Regulations).
The computer replaces test animals?
With the help of computer simulated interactions, between test substances and biological structures, the effectiveness (therapeutic or toxic) of chemical substances can be estimated.
Investigations with humans - in vitro
The direct comparison between test animals and human beings is often only possible in vitro. With the use of similar types of testing arrangements, comparable results can be achieved.
Subcellular fractions in place of test animals
Experiments with cell homogenates or subcellular fractions, make it possible, to specifically analyse the sub-functions of cells or tissues.
Cells as a replacement for animals?
Intact cells are the smallest units, with which it is possible to answer questions concerning cellular structure (membranes, organelles, cell nucleus) and functional changes (transcription, translation, gen-expression, enzyme induction).
Freshly isolated or cultivated mammalian cells?
This free choice doesn’t exist. Isolation and cultivation is only possible with selected cells, tissues or organs. The methods and the experimental system have to be individually matched.
Freshly isolated cells, primary cultures
Investigations on cultivated, differentiated cells with organ specific structures and functions require specific professional knowledge. They are valuable additions to animal testing and make such testing in many cases unnecessary.
Hepatocytes after 24 hours in culture
Morphology and organisation almost like in the liver!
Hepatocytes and bile
Cultivated hepatozytes deliver comprehensive information: morphological changes, excretion products in the culture medium, biochemical investigations, immunohistochemical analysis.
Hepatocytes as established cell line
With transformed human hepatocytes from the cell line HepG2 selected metabolic functions can be investigated, but not many.
Established cell lines
Established cells are used, if fast growing cells is needed or a cell line has to be modified for a specific question (i.e. transgenic cell lines, gene silencing).
Monolayered cell cultures
In monolayered cell lawns structural changes (morphology), which are induced by a test substance or the culture conditions, can be qualitatively and quantitatively measured with high specificity and with many different methods.
Effecient single cell analysis
Monolayered cell cultures make it possible, to relate the effect which is to be measured to individual cells. This allows to draw quantative conclusions about the dose-effect relation. It is often possible to simultaneously collect various end points in one population of cells.
Cooperation and organisation in the liver
The structural organisation in living organisms, for instance in the liver is extremely efficient. Inter- and intracellular communication is the rule. Division of labor and structural organisation reach a maximum.
Co-cultures of cells from the same organ
In co-cultures two cell populations mutually support each other, in their vital and organ specific functions. The exchange of factors in an organ and their interaction can be studied.
Co-culture between cells of different organs
Interaction between different organs in an organism can be partially simulated i.e the effect of metabolites, which are excreted form co-cultured hepatozytes.
An approximation of parenchymal liver cells?
In freshyl isolated hepatocytes, co-cultured as aggregates with Kupffer cells, the metabolic activity can be maintained over a longer period (up to 14 days). However there are other problems.
Speroid cultures of brain cells
Aggregate cultures of brain cells make it possible, to simulate ischemic conditions, and to test substances which potentially relieve such damage.
Reconstructed epithelial human tissue
Cultivated skin, oral mucus, horny skin, lung epithelia, vaginal epithelium are the models of the future, to test the local interaction of substances, as well as bio-medical materials.
Cells and organs from stem cell cultures?
Stem cells could be used to replace animal and human cells in bio-medical experiments. But particular questions are still unsolved.