The Mitochondrial Free Radical Theory of Aging - Supernova: Pliki

Some Testable Predictions of MiFRA
already been shown to be the case for succinate dehydrogenase. 14-16 Conversely, if the PMOR
is not up-regulated then we would expect to see an up-regulation of lactate dehydrogenase
(LDH), which is the enzyme that turns pyruvate into lactate for export from the cell if the
TCA cycle is not maintained. Assays for lactate dehydrogenase activity are routine. 17
The maintenance of the TCA cycle in anaerobic cells is not a vital component of the
mechanism described in Chapter 9, but it would certainly make that mechanism’s effects
much more severe, so it should be tested. Histochemical demonstration that the enzyme is
present is strong evidence that it is active, but falls short of proof. However, there are simple
direct tests of whether ρ 0 cells do this: the simplest are to ask whether they generate CO2,
and whether they can grow on non-fermentable carbon sources (such as, most simply, the
necessary exogenous pyruvate).
Likewise, there are well-developed assays for superoxide production; 18,19 but the rate
of single-electron reduction of extracellular oxygen by ρ 0 cells has, to my knowledge, been
established only under conditions where other electron acceptors are plentiful. 20 (One early
study 21 may point the way here: it measured reduction of oxygen but did not identify the
product.) Alternatively, the extracellular superoxide dismutase is predominantly bound to
endothelial cell surfaces. rather than free in plasma, 22a so any preferential colocalisation
with cytochrome c oxidase inactivity in tissue may well be directly visualisable. A new SOD
assay using cerium appears to allow accurate histochemical visualisation of all three SOD
activities and may be particularly useful in this regard. 22b
A third prediction is that chemicals which inhibit the PMOR should reduce the oxidative
modification of LDL in blood plasma. One chemical is known which inhibits the PMOR:
pCMBS, or p-chloromercuriphenylsulfonic acid. 23 This chemical is toxic, so it clearly cannot
be used as an anti-aging drug; but it may well be possible to assay its effect on LDL oxidation
in rodents before its toxic effects mask that. Alternatively, this may be studyable in vitro. The
oxidation of LDL by cultured aerobic cells is totally inhibited by physiological levels of vitamin
E or other antioxidants. 24 Incubation of LDL with ρ 0 cell lines in a physiologically realistic
medium should allow measurement of its rate of oxidation, which should be non-zero if
they are using the PMOR heavily.
A fourth prediction is that individuals with dysfunction in the machinery of LDL
import into cells will incur oxidative stress somewhat more slowly, and thence age more
slowly in general. This may be harder to test, however, because the import of cholesterol
into cells is a very definite requirement, so that a deficiency in the normal uptake pathway
is likely to be compensated for by a stimulation of some secondary pathway, or else by a
rise in the overall LDL level in the blood. (The latter is what is seen in sufferers from
familial hypercholesterolaemia, a genetic defect in the LDL receptor). 25 Either of these
would negate the retardation of oxidative stress that is predicted above. Again, however, in
vitro experiments may be more straightforward. There are numerous ways to quantify the
level of oxidative stress in cells: these include the concentrations of hydrogen peroxide, 26
of lipid peroxidation products, 27 and of oxidatively damaged proteins. 28 The level of
oxidation of LDL can also be quite accurately controlled in vitro. Thus, an approach to
testing the influence of LDL oxidation on intracellular oxidative stress would be to incubate
cells for an extended period in conditions where they were induced to import LDL at
physiological rates, and measure the dependence of one or more of the above indicators
on LDL oxidation levels.
12.3. Negligible Senescence: Predictions of MiFRA
As noted in Section 6.5.5, numerous species of cold-blooded vertebrate grow throughout
their lives, and some of these live exceptionally long—so long, in fact, that it is possible that
they do not senesce (their future life expectancy does not diminish with age) at all. It is,
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