X-Message-Number: 24552
Date: Tue, 24 Aug 2004 05:20:03 -0700 (PDT)
From: Doug Skrecky <>
Subject: telomere shortening may be the main cause of aging in some humans

[...at least in humans suffering from Werner's syndrome...
Could Atorvastatin be used to safely treat this? - see below]

Mouse Model of Rare Disease Offers Clues to Aging and Cancer Development
M. D. Anderson News Release 08/19/04
  Scientists have developed the first mouse model of a rare disease in
which people age rapidly and start developing cancers and other diseases
associated with the elderly when they are only about 30 years old.
The advance, reported in the August issue of Nature Genetics, is already
shedding light on a suspected strong link between aging and cancer by
suggesting that a single cellular protein can play a role in both
processes, according to researchers. "Given that most cancer occurs in the
elderly, aging is the biggest risk factor for developing cancer in
humans," says the study's lead author, Sandy Chang, M.D., Ph.D.,
assistant professor in the Department of Molecular Genetics at The
University of Texas M. D. Anderson Cancer Center. "Now, with
this animal model, we can look at the common pathways that unify aging
and cancer development." Working with Chang were researchers from Harvard
Medical School, Brigham and Women's Hospital in Boston, and the
Massachusetts Institute of Technology. Werner's syndrome, which only
strikes about one per million individuals worldwide, is the premier
example of an adult-onset premature aging syndrome. Individuals with the
disease have no symptoms for the first decade of life, but then begin to
rapidly develop signs of aging, including thinning hair, wrinkling skin,
cataracts, osteoporosis and diabetes, and they often die in their 40s of
cancer or heart disease. The root of the disease was found to be a failing
gene that produces the protein WRN, known to help maintain the stability
of the cell's genome. Patients with Werner's syndrome exhibit increased
chromosomal aberrations, with pieces missing or fused onto other
chromosomes, says Chang. WRN also has been implicated in telomere
maintenance. Telomeres are repeat sequences that cap the end of
chromosomes and are necessary for chromosomal stability. They also are
closely associated with the aging process - every time a cell divides,
telomeres lose some of their length, and once telomeres become too short,
the cell is programmed to stop dividing. Because the features of
accelerated aging in patients with Werner's syndrome only begins to be
seen after adolescence, the researchers hypothesized that telomere
malfunction produced by a loss of WRN contributes to disease progression.
In other words, when WRN is absent, telomeres shorten prematurely, leading
to a cessation of cell growth that becomes apparent much earlier in life,
says Chang. "We believe that loss of WRN accelerates telomere shortening
and promotes premature onset of aging phenotypes in mice," he says. "If
that is true, then studying this mutation may give us a handle to
understand what normally happens during the aging process." But in order
to test the idea that the symptoms of Werner's syndrome require telomere
shortening, the researchers had to create a mouse model of the disease.
The first attempt by others to develop a mouse without WRN failed; the
mice lived a long life. Scientists soon realized, however, that, in
regard to telomeres, mice differ from those in humans in an important
way. All cells in a mouse, they found, produce the enzyme telomerase that
prevents telomeres from shortening, whereas only a few human tissue
compartments typically "turn on" telomerase production. As a result,
telomeres in mouse cells are much longer than in human cells, and mouse
cells do not undergo the characteristic cessation of cell growth observed
in human cells. Knowing that, Chang and his colleagues bred mice without
WRN to mice engineered not to produce telomerase. They then allowed the
offspring to procreate through several generations in order to
progressively shorten the telomeres. In the first two generations, the
mice aged normally. In later generations, however, some of the mice - the
ones with the shortest telomeres - began to exhibit the classic symptoms
of Werner's syndrome. They also developed certain non-epithelial cell
cancers, including osteosarcomas, soft-tissues sarcomas and lymphoma.
They also died at a much younger age than normal. "The cancers these mice
developed are fairly rare in the general human population, but are common
in patients with Werner's syndrome," says Chang. Thus the mouse model
shows that a deficit in WRN can lead to genomic instability that both
accelerates aging and spurs cancer formation, according to the
researchers. Results of the study point to new avenues to explore, says
Chang. Genomic instability also is the hallmark of cancers that develop
in epithelial cells, which make up the majority of cancers in the general
population. And this instability may also involve telomere length, he
says. For example, the master tumor suppressor gene p53 senses when a
cell's telomeres are too short and tells the cell to shut down. If p53 is
dysfunctional, as it is in most cancers, the cell continues to divide.
Telomeres that are too short increase genomic instability by promoting
chromosomal fusions, a process which by itself is a risk factor for cancer
development. But a "would be" cancer cell must also possess the ability
to maintain their telomeres, otherwise they will die. Most human cancers
keep their telomeres intact by turning on production of telomerase, thus
ensuring that cells will remain immortal, dividing continually. "In fact,
90 percent of cancer cells use telomerase to keep the cells dividing,"
says Chang. "The common denominator in aging and cancer appears in part
to be the failure to maintain genomic stability, and how the p53 pathway
senses this instability will ultimately determine whether a cell is
programmed to stop dividing (aging) or progress to immortal growth
(cancer)," he says. "Now we have a mouse model that will help us
understand these links in detail."

Nat Genet. 2004 Aug;36(8):877-82. Epub 2004 Jul 04
Essential role of limiting telomeres in the pathogenesis of Werner syndrome.
  Mutational inactivation of the gene WRN causes Werner syndrome, an
autosomal recessive disease characterized by premature aging, elevated
genomic instability and increased cancer incidence. The capacity of
enforced telomerase expression to rescue premature senescence of cultured
cells from individuals with Werner syndrome and the lack of a disease
phenotype in Wrn-deficient mice with long telomeres implicate telomere
attrition in the pathogenesis of Werner syndrome. Here, we show that the
varied and complex cellular phenotypes of Werner syndrome are
precipitated by exhaustion of telomere reserves in mice. In
late-generation mice null with respect to both Wrn and Terc (encoding the
telomerase RNA component), telomere dysfunction elicits a classical
Werner-like premature aging syndrome typified by premature death, hair
graying, alopecia, osteoporosis, type II diabetes and cataracts. This
mouse model also showed accelerated replicative senescence and
accumulation of DNA-damage foci in cultured cells, as well as increased
chromosomal instability and cancer, particularly nonepithelial
malignancies typical of Werner syndrome. These genetic data indicate that
the delayed manifestation of the complex pleiotropic of Wrn deficiency
relates to telomere shortening.

Ann N Y Acad Sci. 2004 Jun;1019:186-90
Telomerase expression is differentially regulated in birds of differing
life span.
  Cellular senescence caused by telomere shortening has been suggested as
one potential causal agent of aging. In some tissues, telomeres are
maintained by telomerase; however, telomerase promotes tumor formation,
suggesting a trade-off between aging and cancer. We predicted that
telomerase activity should vary directly with life span. We determined
telomerase activity in bone marrow in cross-sectional samples from two
short-lived bird species and two long-lived bird species. The two
short-lived species had high telomerase activity as hatchlings but showed
a sharp downregulation in both the young and old adults, whereas the two
long-lived species had relatively high telomerase activity in bone marrow
that did not decrease with age. In zebra finches, the age-related change
in telomerase activity varied in different tissues. Telomerase activity
increased late in life in skeletal muscle, liver, and gonad, but not in
blood or bone marrow.

Gut. 2004 Jul;53(7):1001-9.
Prevention of critical telomere shortening by oestradiol in human normal
hepatic cultured cells and carbon tetrachloride induced rat liver fibrosis.
  BACKGROUND AND AIM: Significant telomere shortening of hepatocytes is
associated with replicative senescence and a non-dividing state in
chronic liver disease, resulting in end stage liver failure and/or
development of hepatocellular carcinoma. To prevent critical telomere
shortening in hepatocytes, we have focused on oestrogen dependent
transactivation of the human telomerase reverse transcriptase (hTERT)
gene as a form of telomerase therapy in chronic liver disease. METHODS:
We examined expression of hTERT mRNA and its protein, and telomerase
activity (TA) in three human normal hepatic cell lines (Hc-cells, h-Nheps,
and WRL-68) before and after treatment with 17beta-oestradiol. The
effects of exogenous oestradiol administration were examined in a carbon
tetrachloride (CCl(4)) induced model of liver fibrosis in rats. RESULTS:
Expression of hTERT mRNA and its protein was upregulated by
oestradiol treatment. Telomere length decreased in Hc-cells and h-Nheps
with accumulated passages whereas with long term oestradiol exposure it
was greater than without oestradiol. The incidence of beta-galactosidase
positive cells, indicating a state of senescence, decreased significantly
in oestradiol treated cells in comparison with non-treated cells (p<0.05).
TA in both male and female rats with CCl(4) induced liver fibrosis was
significantly higher with oestradiol administration than without
(p<0.05). Long term oestradiol administration markedly rescued the
hepatic telomere from extensive shortening in both male and female rats.
CONCLUSION: These results suggest that oestradiol acts as a positive
modulator of the hTERT gene in the liver. Oestrogen dependent
transactivation of the hTERT gene is a new strategy for slowing the
progression of chronic liver disease.

Circ Res. 2004 Apr 2;94(6):768-75. Epub 2004 Feb 12
Antioxidants inhibit nuclear export of telomerase reverse transcriptase
and delay replicative senescence of endothelial cells.
  Aging is associated with a rise in intracellular reactive oxygen species
(ROS) and a loss of telomerase reverse transcriptase activity. Incubation
with H2O2 induced the nuclear export of telomerase reverse transcriptase
(TERT) into the cytosol in a Src-family kinase-dependent manner.
Therefore, we investigated the hypothesis that age-related increase in
reactive oxygen species (ROS) may induce the nuclear export of TERT and
contribute to  endothelial cell senescence. Continuous cultivation of
endothelial cells resulted in an increased endogenous formation of ROS
starting after 29 population doublings (PDL). This increase was
accompanied by mitochondrial DNA damage and preceded the onset of
replicative senescence at PDL 37. Along with the enhanced formation of
ROS, we detected an export of nuclear TERT protein from the nucleus into
the cytoplasm and an activation of the Src-kinase.
Moreover, the induction of premature senescence by low concentrations of
H2O2 was completely blocked with the Src-family kinase inhibitor PP2,
suggesting a crucial role for Src-family kinases in the induction of
endothelial cell aging. Incubation with the antioxidant N-acetylcysteine,
from PDL 26, reduced the intracellular ROS formation and prevented
mitochondrial DNA damage. Likewise, nuclear export of TERT protein, loss
in the overall TERT activity, and the onset of replicative senescence
were delayed by incubation with N-acetylcysteine. Low doses of the
statin, atorvastatin (0.1 micromol/L), had also effects similar to those
of N-acetylcysteine. We conclude that both antioxidants and statins can
delay the onset of replicative senescence by counteracting the increased
ROS production linked to aging of endothelial cells.

Circ Res. 2003 May 16;92(9):1049-55. Epub 2003 Apr 03.
HMG-CoA reductase inhibitors reduce senescence and increase proliferation
of endothelial progenitor cells via regulation of cell cycle regulatory genes.
  Endothelial progenitor cells (EPCs) play an important role in postnatal
neovascularization of ischemic tissue. Ex vivo expansion of EPCs might be
useful for potential clinical cell therapy of myocardial ischemia.
However, cultivation of primary cells leads to cellular aging
(senescence), thereby severely limiting the proliferative capacity.
Therefore, we investigated whether statins might be able to prevent
senescence of EPCs. EPCs were isolated from peripheral blood and
characterized. After ex vivo cultivation, EPCs became senescent as
determined by acidic beta-galactosidase staining. Atorvastatin or
mevastatin dose-dependently inhibited the onset of  EPC senescence in
culture. Moreover, atorvastatin increased proliferation of EPCs as
assessed by BrdU incorporation and colony-forming capacity. Whereas
geranylgeranylpyrophosphate or farnesylpyrophosphate reduced the
senescence inhibitory effect of atorvastatin, NO synthase inhibition,
antioxidant s, or Rho kinase inhibitors had no effect. To get further
insights into the underlying downstream effects of statins, we measured
telomerase activity and determined the expression of various cell cycle
regulatory genes by using a microarray assay. Whereas telomerase activity
did not change, atorvastatin modulated expression of cell cycle genes
including upregulation of cyclins and downregulation of the cell cycle
inhibitor p27Kip1. Taken together, statins inhibited senescence of EPCs
independent of NO, reactive oxygen species, and Rho kinase, but dependent
on geranylgeranylpyrophosphate. Atorvastatin-mediated prevention of EPC
senescence appears to be mediated by the regulation of various cell cycle
proteins. The inhibition of EPC senescence and induction of EPC
proliferation by statins in vitro may importantly improve the functional
activity of EPCs for potential cell therapy.

Clin Exp Pharmacol Physiol. 2004 Jul;31(7):407-13.
Oxidized low-density lipoprotein induces endothelial progenitor cell
senescence, leading to cellular dysfunction.
1. Recent studies have revealed an association between coronary risk
factors and both the number and function of bone marrow-derived
endothelial progenitor cells (EPC). We investigated the effect of
oxidized low-density lipoprotein (ox-LDL) on the senescence of EPC,
leading to cellular dysfunction.
2. Endothelial progenitor cells were isolated from human peripheral blood
and characterized. The exposure of cultured EPC to ox-LDL (10 microg/mL)
significantly accelerated the rate of senescence compared with control
during 20 days in culture as determined by acidic beta-galactosidase
staining. Oxidized LDL-induced EPC senescence was significantly inhibited
by pretreatment with either lectin-like ox-LDL receptor-1 (LOX-1)
antibody (Ab) or atorvastatin (P < 0.01). 3. Because cellular senescence
is critically influenced by telomerase, which elongates telomeres, we
measured telomerase activity using a polymerase chain reaction-ELISA-based
assay. Oxidized LDL significantly diminished telomerase activity to
approximately 50%, an effect that was significantly abolished by
pretreatment with either LOX-1 Ab or atorvastatin (P < 0.01). 4. We
examined whether ox-LDL-induced EPC senescence translates into EPC
dysfunction. An MTS assay disclosed an inhibitory effect of ox-LDL on EPC
proliferation. In a Matrigel assay, EPC treated with ox-LDL were less
likely to participate in network formation compared with controls. 5. In
conclusions, ox-LDL accelerates the onset of EPC senescence, which may be
related to telomerase inactivation. Oxidized LDL-induced EPC senescence
leads to the impairment of proliferative capacity and network formation.

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