How to measure aging in vitro

Sydney Shall bafa1 at central.susx.ac.uk
Fri Jul 29 07:43:39 EST 1994

Chris Driver (drierac at deakin.edu.au) wrote:
: I have seen a number of papers in which substances will alter the lifespan of 
: cells in culture. One thing that appears to be a common feature in these 
: reports is that repeat runs of apparently identical conditions are not 
: particularly reproduceable. Robin Halliday in his paper comments on this and 
: suggests that a clonal succession may occur, where old cell lines are 
: dominated by clones arising from the odd cell that is rather less senescent 
: than their sibling cells.
We have done quite a lot of work on this.  Originally J. Smith et al
showed that it was true that successive waves of cell clones emerge
during the lifespan of human cells. He showed that in clonies that were
short-lived there were cells that could generate bigger colonies than
the parent clone.  We have subsequently confirmed this, and have
directly measured the fraction of growing cells in an ageing population;
we found that there is always a fraction of senescent cells, even in a
young population.  What changes with ageing is the fraction of
non-dividing, senescent cells in the population.  From the beginning of
the culture there are some senescent cells present; as the culture
divides the fraction of non-dividing, senescent cells increases until
the fraction of new-born cells which are non-dividers, that means
reproductively sterile, is greater than 50%.  At this point the culture
will inevitably begin to decline. Because although there are dividing
cells in the population, each set of new-born cells has fewer fertile
cells thatn the last generation. Clearly, eventually, the fraction of
new-born cells able to divide will eventually reach zero and the culture
will be entirely post-mitotic.

: In theory it should be possible to detect senescent cells arising in cultures 
: in about midpassage and quantify them. Has anyone any thoughts on this matter?

We have precisely this.  At the moment there is not available yet a
reliable easy marker for senescent cells, although recently a number of
potential candidates have come to light.  What we have done then is to
score all the cells in a population with antibodies to proteins which
are characteristic of dividing cells; or we have used bromo-deoxyuridine
incorporation (to simulate thymidine) to measure DNA synthesis.  In this
way we measured the fraction of cells throughtout the lifespan that were
dividers, and by difference the remainder of the population are
non-dividing, that is senescent cells.

: An alternative, but less useful approach may be to use markers for senescence 
: in the total cell mass. Has anyone any suggestions for good markers?

: Chris Driver

: Chris Driver, Ph D
: School of Biology and Chemistry, Rusden Campus
: Deakin University
: 662 Blackburn Rd
: Clayton, VIC, 3168
Sydney SHALL,
Laboratory of Cell and Molecular Biology,
Biology Building, University of Sussex, Brighton, East Sussex BN1 9QG, ENGLAND.
Telephone: +         FAX: +

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