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A CRONie noted:

>>From Dr. Edward J Masoro, Dept Physiology, Univ Texas
> Quote from Dr. Ed's Laboratory Research paper below:
>
> ----------
> "Recently, methionine restriction has received wide
> acclaim including speculation that it may play a role in
> FR-induced life extension. In my opinion, work on rats done
> in our laboratory makes it unlikely that this speculation is
> correct, at least for that species (4).
>
> We found that a long term 40% reduction in food intake
> without a reduction in methionine intake extends the life
> of rats to the same extent as a 40% reduction of both food
> and methionine intake does (Table 1). Thus, it is clear
> that methionine intake need not be decreased for FR to
> markedly extend the life of rats."
>
> ---------
> Masoro EJ.
> Caloric Restriction and Aging: Controversial Issues.
> J Gerontol A Biol Sci Med Sci. 2006 Jan;61(1):14-19.
> PMID: 16456190

Of further interest may be the paper below. The data of Fig. 1
are fairly well delineated in Table 1, but the graphs and
the data for the
yeast versus casein, with the latter having a significantly
greater
methionine content, were informative. The shapes and ages
for the survivor
curves over time differed with the different casein levels,
the yeast versus
casein levels, and the males versus females. For this
figure alone, the pdf
says much more than can be captured in numeric description.
The rates of
aging and the longevity varied much. The 4% casein diet
reduced female
versus male longevity. The 2% casein diet increased male
versus female
longevity. The 0.5 and 1% casein female curves overlapped
and were distinct
in their slower rate of aging. Had the animals more
resembled humans, the
results may have carried greater import, but, with respect
to fruit flies
relying more on sugars for nutrition, the emphasis clearly
was on proteins
and their methionine content.

Min KJ, Tatar M.
Restriction of amino acids extends lifespan in Drosophila
melanogaster.
Mech Ageing Dev. 2006 Jul;127(7):643-6. Epub 2006 Apr 17.
PMID: 16616772

Dietary restriction extends adult Drosophila melanogaster
life span when the
concentration of dietary yeast is diluted in a media with
abundant
carbohydrates. Here we vary the concentration of casein as a
source of amino
acids in adult diet to uncover a quality of nutrient yeast
responsible for
longevity control. Longevity is maximized upon diet with
intermediary levels
of casein. Differences in survival are not caused by elevated
age-independent mortality; the longevity maximum at
intermediate casein does
not arise because casein is non-specifically harmful at higher
concentrations. Furthermore, fecundity increases when the
level of dietary
casein is elevated. The demographic phenotypes of adult
Drosophila
maintained on intermediate levels of casein resemble their
response to
limited dietary yeast. Dietary restriction through dilution
of yeast may
extend longevity because this limits the intake of amino acids.

Reduction of food intake without starvation extends life
span in many
organisms including yeast, nematode, fruit fly and rodents
(Bertrand et al.,
1999; and reviews in Masoro, 2000, Koubova and Guarente,
2003, Partridge et
al., 2005 and Tatar, in press). These manipulations are
referred to as
caloric restriction (CR) when applied to rodents because
longevity may be
extended through control of calorie intake alone (Masoro,
2000). Studies
with non-vertebrates, in contrast, typically dilute all
components of the
diet and thus vary all nutrients in an otherwise constant
volume of inert
media (e.g., Lin et al., 2002, Johnson et al., 1990,
Chippindale et al.,
1993, Chapman and Partridge, 1996 and Mair et al., 2003). These
manipulations are referred to as dietary restriction (DR)
because they do
not directly regulate the intake of calories, and it remains
generally
unknown whether calories or a specific component of the
adult diet is
responsible for longevity extension.

Limiting calories, in fact, may not be the feature of
nutrition responsible
for extended life span in Drosophila melanogaster. Longevity
can be extended
in males and females maintained on a carbohydrate media
supplemented with a
relatively low concentration of live yeast (Chippindale et
al., 1993).
Likewise, lifespan is increased by reducing the
concentration of inactive
dry yeast in constant base medium of sugar and cornmeal (Min
and Tatar,
2006). Variation in yeast may be sufficient to modulate
aging but because
caloric intake is increased on yeast-rich medium, yeast
concentration and
total caloric intake can be positively correlated. Mair et
al. (2005) have
recently manipulated both dietary yeast and sugar to
disentangle these
nutrient effects. Reduction of either yeast or sugar alone
improved survival
in females but flies were markedly more sensitive to
variation in yeast, and
across diets longevity did not correlate with the food
energetic value.
Since feeding rate was thought to be constant across these
diets at least in
terms of proboscis extension (but see Carvalho et al., 2005
and Min and
Tatar, 2006), the increase in longevity with restricted diet
was argued to
be independent of net caloric intake. The data of
Chippindale et al. (1993)
and of Mair et al. (2005) suggest that limiting a specific
nutrient
component of the yeast mediates longevity in Drosophila.

Dietary yeast is a complex source of nutrition, containing
essential and
non-essential amino acids and fatty acids, nucleic acids,
minerals, vitamins
and carbohydrates. Although any of these components could
influence
longevity, here we focus on amino acids, which are the most
abundant
nutrient metabolite of dry yeast (45%) (Schulze, 1995). In
rodents,
restriction of the methionine alone is sufficient to extend
life span and to
retard the progression of age-dependent degenerative
pathology (Zimmerman et
al., 2003 and Miller et al., 2005). Amino acids are also
implicated in
recent molecular studies in Drosophila and C. elegans where
longevity is
extended by repressing the cellular amino acid responsive
Target of
Rapamycin (TOR) pathway (Kapahi et al., 2004, Vellai et al.,
2003 and
Kaeberlein et al., 2005).

Amino acids as a potential mediator of dietary restriction
in D.
melanogaster have been investigated in two early studies
through
manipulation of casein as a dietary nutrient. Hollingsworth
and Burcombe
(1970) compared survival on an adult diet of sugar alone
relative to sugar
plus casein. The addition of amino acids improved longevity.
Van Herrewege
(1974) compared survivorship across several casein
concentrations in a base
medium with sugar, vitamins, nucleic acids and essential
lipids. Mean
lifespan was maximized at intermediate concentrations of
casein but these
data did not address whether mortality and reproduction
changed in ways
consistent with slow aging as induced by dietary
restriction. In particular,
high levels of a refined nutrient such as casein may be
toxic and contribute
to mortality that is independent of aging. Here we follow-up
on these early
reports and describe not only the overall survivorship
across varied casein
diets but also the trajectories of mortality and the
patterns of
age-specific fecundity.

Larvae of the Canton-S strain were grown on standard medium
(5.2 g cornmeal,
11 g sugar, 11 g yeast, 0.79 g agar per 100 ml water)
supplemented with
several grains of live yeast (Elgin and Miller, 1980). Newly
elose adults
were collected over 48 h and were randomly assigned to three
1 l demography
cages to a final density of 150 individuals (mixed sex).
Food vials were
affixed to each cage and changed every 2 days, at which time
dead flies were
removed, sexed and recorded.

Adults were short-lived (median life span: males 18 days,
females 22 days)
on a sucrose-only diet (11 g sugar and 1.1 g agar per 100 ml
water) as
expected (Good and Tatar, 2001), and the addition of 10%
yeast strongly
increased survival (Fig. 1A). Likewise, the addition of
casein to the
sucrose diet increased lifespan. In males, casein at levels
of 0.5%, 1% and
2% produced survival similar to the diet with 10% yeast and
significantly
greater than the survival on 4% casein (Log rank test, 1%
versus 4%, chi^2 =
233.82, p < 0.0001). Male survival is reduced 38% upon 4%
diet compared to
one on 1% casein diet. In females survival was greatest when
maintained on
0.5% or 1% casein, intermediate on 2% casein and lowest on
4% casein (Log
rank test, 1% versus 2%, vhi^2 = 42.195, p < 0.0001; 1%
versus 4%, chi^2 =
261.37, p < 0.0001). Thus, as previously observed, D.
melanogaster longevity
is extended when dietary amino acids are limited.

Reduced survival upon diet with elevated casein is not
caused by elevated
age-independent mortality. We assessed the relative
goodness-of-fit between
mortality with and without the Makeham parameter (c), which
estimates the
magnitude of age-independent mortality. The Makeham
parameter did not
significantly differ from zero in any cohort of casein-fed
males or females;
most cohort were best described by the Gompertz (µx = A
exp(Bx)) or
Gompertz-Logistic (µx = A exp(Bx) + f(s)) mortality model
(Table 1). These
patterns are clear in Fig. 1B where there is no trend of early
age-independent mortality; the observed early death rates
correspond to
observations expected at the 'left-hand boundary' of a
mortality plot (see
Promislow et al., 1999), and mortality rates decelerate
among the oldest-old
(Vaupel et al., 1998). Reduced amino acids increases
lifespan because this
nutrient state consistently reduces the age-dependent
trajectory of
mortality, it retards demographic aging.

Table 1. Mortality statistics of the five different cohort
flies fed
different concentration of casein, with best-fit mortality
models
===================================================================
Best-fit model a (×10^-3) (LCI, UCI) b* (×10-^2) (LCI,
UCI) s* (LCI,
UCI) c* (×10-3^) (LCI, UCI) Med LS
===================================================================
Males
0% L 0.11 (0.03, 0.47) 44.45 (35.36, 55.87) 1.19 (0.72,
1.98) - 18
0.5% G 0.71 (0.42, 1.2) 11.29 (10.08, 12.64) - - 40
1% M 0.12 (0.04, 0.38) 15.09 (12.80, 17.78) - 3.10 (1.52,
6.32) 44
2% G 0.28 (0.15, 0.55) 13.54 (12.07, 15.17) - - 42
4% L 0.78 (0.3, 2.02) 21.33 (16.50, 27.56) 0.98 (0.54,
1.77) - 26

Females
0% L 0.09 (0.01, 0.53) 37.40 (27.68, 50.55) 1.45 (0.77,
2.71) - 22
0.5% L 0.19 (0.05, 0.77) 17.84 (13.31, 23.92) - - 38
1% L 0.28 (0.07, 1.09) 16.08 (11.70, 22.09) 0.60 (0.19,
1.82) - 38
2% LM 0.01 (0.01, 0.11) 30.85 (23.67, 40.19) 1.28 (0.77,
2.10) 2.05 (0.8,
4.77) 34
4% L 0.21 (0.06, 0.40) 30.88 (25.14, 37.92) 0.71 (0.4,
1.24) - 24
===================================================================
Best-fit models and parameter estimates were obtained
using WinModest
Software (Pletcher, 1999).
*a Data were fitted to among Gompertz (G, µx = a e^bt
where µx represents
mortality rate at age t).
*b the change in mortality rate with age.
*c the age-independent mortality risk.
Median life span (Med LS, days).

Fecundity was measured by locating single female with two
males for 10 days.
Females laid very few eggs when maintained on sugar-only
medium (Fig. 2A).
Sugar medium supplemented with 10% yeast elicited normal
oviposition;
females laid 30 eggs/day/female by day 5 and stabilized at
18-25
eggs/day/female thereafter (mean among female fecundity for
10 days: 160 ±
11.7). Females maintained on casein produced a small number
of eggs each day
(Fig. 2A), with a peak of 5-10 eggs at early ages.
Importantly, the total
number of eggs from casein-fed females generally increased
with the
concentration of the amino acid nutrient (Fig. 2B, one-way
ANOVA on all data
points, F(3,54) = 12.77, p < 0.0001; one-way ANOVA on data
points after day
4, F(3,54) = 32.44, p < 0.001). Although our casein diets
were not optimal
for egg production, the increase in reproduction with high
casein represents
a physiological trade-off between fecundity and lifespan.
Consistent with
our interpretation of the mortality patterns, flies on
reduced casein were
not relatively long-lived because flies on high casein
suffered a
pharmacological artifact which may cause decrease in fecundity.

Several protocols increase lifespan D. melanogaster by
restricting dietary
yeast (Chippindale et al., 1993, Mair et al., 2005 and Min
and Tatar, 2006).
We now confirm that restricting the source of dietary amino
acids in media
with abundant carbohydrate is sufficient to increase
longevity. Importantly,
this effect is consistent with physiological and demographic
patterns
induced by limiting the level of dietary yeast. Amino acids
are at least one
specific nutrient through which dietary restriction
modulates aging.