Models of Disease 1 Flashcards
What does in vitro mean ?
Cells growing in culture outside the normal biological context
Describe the basics of the theory behind in vitro cell culture
Cell isolation and growth - primary cells are isolated directly from tissue
Culture medium with nutrients
Sterile environment
Controlled conditions - temperature, pH
Substrate for growth
Immortalized cells – adults cells with ability to divide indefinitely either through random mutation (cancer) or deliberate modification
What are the advantages of in vitro cell culture ?
Relatively cheap and easy to do
High level of control over the environment of the cell
Can test drugs easily and cheaply
Potential to investigate disease models & cell interactions with less dependence on animals
Reproducibility
What are the disadvantages of in vitro cell culture ?
Lack of physiological relevance
Loss of cellular heterogeneity
Define in vivo and ex vivo:
In vivo - cells growing in their normal biological context
Ex vivo - from the body but now outside the body – any experimentation using recently excised tissue in conditions as close to physiological as normal
What is passage ?
Each increase in flask number = passage
To passage, cells are removed from the bottom of the dish, counted and specific numbers added (seeded/plated) into each new flask/disk
Passage no is always noted for primary cells but not immortalized cells
What is Replicative Senescence in Primary Cells ?
Primary cells in culture undergo senescence according to their Hayflick limit
Defined as the mean number of doubling times before division ceases
Evidence shows that the telomeres associated with each cell’s DNA will get slightly shorter with each new cell division until they shorten to a critical length
What are the limitations of tissue culture ?
Potential for cross species contamination or introduction of viruses or prions from FBS - alternative is to use human platelet lysate (hPL).
Animals vary in genetics just as much as humans – need 6-15 animals to take account for variability
Loss or changes in differentiated features of cells including phenotype and gene expression profile
What are some questions that should be asked during cell culturing ?
Do the cells replicate the in vivo situation being investigated?
Should they be cycling or quiescent?
Do they require special growth supplements or attachment matrix or additional cells types to function properly?
Do they show differentiated morphological features and/or positive staining with immunotypic markers?
