The decay of mitochondria with age due to oxidation of RNA/DNA, proteins,
and lipids is a major interest; we are making progress in reversing some
of this decay in old rats by feeding them normal mitochondrial metabolites
at high levels and are extending the work to humans. Another major interest
is determining optimum micronutrient intakes for minimizing human DNA damage
as an aid in the prevention of cancer and other degenerative diseases associated
Oxidative mitochondrial decay is a major contributor to aging [1-5]. In
old rats (vs. young rats) mitochondrial membrane potential, cardiolipin
level, respiratory control ratio, and cellular O2 uptake are lower; oxidants/02,
neuron RNA oxidation, and mutagenic aldehydes from lipid peroxidation are
higher [3, in prep.] Ambulatory activity and cognition declines with age.
Feeding old rats the normal mitochondrial metabolites acetyl carnitine and
lipoic acid for a few weeks restores mitochondrial function; lowers oxidants,
neuron RNA oxidation, and mutagenic aldehydes; and increases rat ambulatory
activity and cognition (as assayed with the Skinner box and Morris water
maze) [6-9, 10, in prep.]. With age, increased oxidative damage to protein
and mitochondrial membranes and the loss of cardiolipin causes a deformation
of structure of key enzymes, with a consequent lessening of affinity (Km)
for the enzyme substrate ; increasing the level of the substrate (acetyl
carnitine) restores the velocity of the reaction, membranes, Km for acyl
carnitine transferase, and function [in prep.].
Apurinic/apyrimidinic (AP) sites are common DNA lesions that arise from
spontaneous depurination or by base excision repair (BER) of modified bases.
A biotin-containing aldehyde-reactive probe (ARP) is used to measure AP
sites in living cells . The assay was applied to living cells and nuclei.
The number of AP sites in old human fibroblasts (IMR90 cells) was about
two to three times higher than that in young cells, and the number in human
leukocytes from old donors was about seven times that in young donors. The
repair of AP sites was slower in senescent cells compared with young IMR90
cells. An age-dependent decline is shown in the activity of the glycosylase
that removes methylated bases in IMR90 cells and in human leukocytes. The
decline in excision of methylated bases from DNA suggests an age-dependent
decline in 3-methyladenine DNA glycosylase, a BER enzyme responsible for
removing alkylated bases.
Approximately forty micronutrients are required in the human diet. Deficiency
of vitamins B12, folic acid, B6, niacin, C, or E, or iron, or zinc, appears
to mimic radiation in damaging DNA by causing single- and double-strand
breaks, oxidative lesions, or both . The percentage of the U.S. population
that has a low intake (<50 percent of the RDA) for each of these eight
micronutrients ranges from 2 percent to 20+ percent; half of the population
may be deficient in at least one of these micronutrients . We have shown
 that folate deficiency breaks chromosomes due to massive incorporation
of uracil in human DNA (4 million/cell) with subsequent single strand breaks
in DNA formed during base excision repair: two nearby single strand breaks
on opposite strands cause the chromosome to fall apart. The level of folate
where we see high uracil and breaks was present in 10percent of the U.S.
population and close to half of poor urban minorities, due to poor diets.
Vitamin B12 (14 percent elderly) and B6 (10 percent of U.S.) deficiencies
also cause high uracil in human DNA and chromosome breaks as indicated by
our new evidence and as expected from mechanistic considerations. We are
currently attempting to determine the level of these three vitamins that
minimizes both nuclear and mitochondrial DNA damage in humans. We have evidence
that inadequate iron (19 percent of women of menstruating age in the U.S.)
causes oxidative damage to mtDNA in rats. Micronutrient deficiency may explain,
in good part, why the quarter of the population that eats the fewest fruits
and vegetables (five portions a day is advised) has about double the cancer
rate for most types of cancer when compared to the quarter with the highest
intake . A number of other degenerative diseases of aging are also associated
with low fruit and vegetable intake. Eighty percent of American children
and adolescents and 68 percent of adults do not eat five portions a day
. Common micronutrient deficiencies are likely to damage DNA by the
same mechanism as radiation and many chemicals, appear to be orders of magnitude
more important, and should be compared for perspective [11, 14, 15].
We are studying the role of micronutrient inadequacy, particularly folate,
B12, B6, iron, and zinc on neuronal damage and cognitive function in rats.
Considerable epidemiological evidence in humans shows an association between
low intakes and cognitive dysfunction. Much circumstantial evidence supports
the plausibility of a direct causal relationship.
We are investigating the effect of inadequate micronutrient intake on genetic
damage to sperm . We have shown that folic acid deficiency decreases
the sperm count in the rat by 90 percent, and that uracil is found in the
sperm DNA of men on low fruit and vegetable diets. Our recent work in humans
 shows an inverse association between the level of the non-methyl THF
pool, but not the methyl-THF pool, with both sperm count and quality, consistent
with a uracil misincorporation mechanism. We had previously shown that men
with low vitamin C intake had more oxidative damage to their sperm DNA and
that male smokers (smoking depletes the vitamin C level markedly) had more
oxidative damage to their sperm . Recent epidemiology supports the notion
that smoking males have more offspring with childhood cancer.
Tuning-up human metabolism, which varies with genetic constitution and changes
with age, is likely to be a major way to minimize DNA damage, improve health,
and prolong healthy lifespan.