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1.  Mitochondria and Cardiovascular Aging 
Circulation research  2012;110(8):10.1161/CIRCRESAHA.111.246140.
Old age is a major risk factor for cardiovascular diseases. Several lines of evidence in experimental animal models have indicated the central role of mitochondria both in lifespan determination and cardiovascular aging. In this article we review the evidence supporting the role of mitochondrial oxidative stress, mitochondrial damage and biogenesis as well as the crosstalk between mitochondria and cellular signaling in cardiac and vascular aging. Intrinsic cardiac aging in the murine model closely recapitulates age-related cardiac changes in humans (left ventricular hypertrophy, fibrosis and diastolic dysfunction), while the phenotype of vascular aging include endothelial dysfunction, reduced vascular elasticity and chronic vascular inflammation. Both cardiac and vascular aging involve neurohormonal signaling (e.g. renin-angiotensin, adrenergic, insulin-IGF1 signaling) and cell-autonomous mechanisms. The potential therapeutic strategies to improve mitochondrial function in aging and cardiovascular diseases are also discussed, with a focus on mitochondrial-targeted antioxidants, calorie restriction, calorie restriction mimetics and exercise training.
PMCID: PMC3867977  PMID: 22499901
2.  Measuring telomere length for the early detection of precursor lesions of esophageal squamous cell carcinoma 
BMC Cancer  2013;13:578.
Esophageal cancer is the sixth leading cause of cancer death worldwide; current early detection screening tests are inadequate. Esophageal balloon cytology successfully retrieves exfoliated and scraped superficial esophageal epithelial cells, but cytologic reading of these cells has poor sensitivity and specificity for detecting esophageal squamous dysplasia (ESD), the precursor lesion of esophageal squamous cell carcinoma (ESCC). Measuring telomere length, a marker for chromosomal instability, may improve the utility of balloon cytology for detecting ESD and early ESCC.
We examined balloon cytology specimens from 89 asymptomatic cases of ESD (37 low-grade and 52 high-grade) and 92 age- and sex-matched normal controls from an esophageal cancer early detection screening study. All subjects also underwent endoscopy and biopsy, and ESD was diagnosed histopathologically. DNA was extracted from the balloon cytology cells, and telomere length was measured by quantitative PCR. A receiver operating characteristic (ROC) curve was plotted for telomere length as a diagnostic marker for high-grade dysplasia.
Telomere lengths were comparable among the low- and high-grade dysplasia cases and controls, with means of 0.96, 0.96, and 0.92, respectively. The area under the ROC curve was 0.55 for telomere length as a diagnostic marker for high-grade dysplasia. Further adjustment for subject characteristics, including sex, age, smoking, drinking, hypertension, and body mass index did not improve the use of telomere length as a marker for ESD.
Telomere length of esophageal balloon cytology cells was not associated with ESCC precursor lesions. Therefore, telomere length shows little promise as an early detection marker for ESCC in esophageal balloon samples.
PMCID: PMC3882883  PMID: 24308314
Esophageal squamous cell carcinoma; Esophageal squamous dysplasia; Early detection; Screening; Balloon cytology; Telomeres
3.  Mitochondrial oxidative stress mediates Angiotensin II-induced cardiac hypertrophy and Gαq overexpression-induced heart failure 
Circulation research  2011;108(7):837-846.
Mitochondrial dysfunction has been implicated in several cardiovascular diseases; however, the roles of mitochondrial oxidative stress and DNA damage in hypertensive cardiomyopathy are not well understood.
We evaluated the contribution of mitochondrial reactive oxygen species (ROS) to cardiac hypertrophy and failure by using genetic mouse models overexpressing catalase targeted to mitochondria and to peroxisomes.
Methods and Results
Angiotensin II increases mitochondrial ROS in cardiomyocytes, concomitant with increased mitochondrial protein carbonyls, mitochondrial DNA deletions, increased autophagy and signaling for mitochondrial biogenesis in hearts of Angiotensin II treated mice. The causal role of mitochondrial ROS in Angiotensin II-induced cardiomyopathy is shown by the observation that mice that overexpress catalase targeted to mitochondria, but not mice that overexpress wild-type peroxisomal catalase, are resistant to cardiac hypertrophy, fibrosis and mitochondrial damage induced by Angiotensin II, as well as heart failure induced by overexpression of Gαq. Furthermore, primary damage to mitochondrial DNA, induced by zidovudine administration or homozygous mutation of mitochondrial polymerase gamma, is also shown to contribute directly to the development of cardiac hypertrophy, fibrosis and failure.
These data indicate the critical role of mitochondrial ROS in cardiac hypertrophy and failure and support the potential use of mitochondrial-targeted antioxidants for prevention and treatment of hypertensive cardiomyopathy.
PMCID: PMC3785241  PMID: 21311045
mitochondria; reactive oxygen species; angiotensin; cardiomyopathy; heart failure
4.  Mitochondrial targeted antioxidant peptide ameliorates hypertensive cardiomyopathy 
We investigated the effect of reducing mitochondrial oxidative stress by the mitochondrial-targeted antioxidant peptide SS-31 in hypertensive cardiomyopathy.
Oxidative stress has been implicated in hypertensive cardiovascular diseases. Mitochondria and NADPH oxidase have been proposed as primary sites of reactive oxygen species (ROS) generation.
The mitochondrial targeted antioxidant peptide SS-31 was used to determine the role of mitochondrial oxidative stress in Angiotensin II (Ang)-induced cardiomyopathy, as well as in Gαq overexpressing mice with heart failure.
Angiotensin II induces mitochondrial ROS in neonatal cardiomyocytes, which is prevented by SS-31, but not the non-targeted antioxidant N-acetyl cysteine (NAC). Continuous administration of Ang for 4 weeks in mice significantly increased both systolic and diastolic blood pressure, and this was not affected by SS-31 treatment. Ang was associated with upregulation of NADPH oxidase 4 (NOX4) expression, increased cardiac mitochondrial protein oxidative damage and induced the signaling for mitochondrial biogenesis. Reducing mitochondrial ROS by SS-31 substantially attenuated Ang-induced NOX4 upregulation, mitochondrial oxidative damage, upregulation of mitochondrial biogenesis, phosphorylation of p38 MAP kinase, and prevented apoptosis, concomitant with amelioration of Ang induced cardiac hypertrophy, diastolic dysfunction, and fibrosis, despite the absence of blood pressure lowering effect. NAC did not show any beneficial effect. SS-31 administration for 4 weeks also partially rescued the heart failure phenotype of Gαq overexpressing mice.
Mitochondrial targeted peptide SS-31 ameliorates cardiomyopathy resulting from prolonged Ang stimulation as well as Gαq overexpression, suggesting its potential clinical application for target organ protection in hypertensive cardiovascular diseases.
PMCID: PMC3742010  PMID: 21620606
mitochondria; hypertension; cardiomyopathy
5.  Pan-colonic field defects are detected by CGH in the colons of UC patients with dysplasia/cancer 
Cancer Letters  2012;320(2):180-188.
BAC arrays were used to evaluate genomic instability along the colon of patients with ulcerative colitis (UC). Genomic instability increases with disease progression and biopsies more proximal to dysplasia showed increased instability. Pan-colonic field copy number gain or loss involving small (< 1 Mb) regions were detected in most patients and were particularly apparent in the UC progressor patients who had dysplasia or cancer. Chromosomal copy gains or losses affecting large regions were mainly restricted to dysplastic biopsies. Areas of significant chromosomal losses were detected in the UC progressors on chromosomes 2q36, 3q25, 3p21, 4q34, 4p16.2, 15q22, and 16p13 (p-value ≤ 0.04). These results extend our understanding of the dynamic nature of pan-colonic genomic instability in this disease.
PMCID: PMC3406733  PMID: 22387989
ulcerative colitis; genomic instability; BAC array
6.  mTOR is a key modulator of ageing and age-related disease 
Nature  2013;493(7432):338-345.
Many experts in the biology of ageing believe that pharmacological interventions to slow ageing are a matter of ‘when’ rather than ‘if’. A leading target for such interventions is the nutrient response pathway defined by the mechanistic target of rapamycin (mTOR). Inhibition of this pathway extends lifespan in model organisms and confers protection against a growing list of age-related pathologies. Characterized inhibitors of this pathway are already clinically approved, and others are under development. Although adverse side effects currently preclude use in otherwise healthy individuals, drugs that target the mTOR pathway could one day become widely used to slow ageing and reduce age-related pathologies in humans.
PMCID: PMC3687363  PMID: 23325216
7.  Mitochondria-targeted catalase reduces abnormal APP processing, amyloid β production and BACE1 in a mouse model of Alzheimer's disease: implications for neuroprotection and lifespan extension 
Human Molecular Genetics  2012;21(13):2973-2990.
The purpose of this study was to investigate the protective effects of the mitochondria-targeted antioxidant catalase (MCAT) and lifespan extension in mice that express amyloid beta (Aβ). Using immunoblotting and immunostaining analyses, we measured the production of full-length amyloid precursor protein (APP), soluble APPα, C-terminal fragments CTF99 and CTF83, monomeric and oligomeric Aβ, Aβ deposits and beta site amyloid precursor protein cleaving enzyme 1 (BACE1), in different stages of disease progression in MCAT/AβPP and AβPP mice. Using quantitative reverse transcriptase polymerase chain reaction and immunostaining analyses, we studied the expression of catalase, BACE1, the Alzheimer's disease (AD) markers, synaptophysin, APP, neprilysin, insulin-degrading enzyme and transthyretin in MCAT, AβPP, MCAT/AβPP and wild-type (WT) mice. Using the high pressure liquid chromatography analysis of 8-hydroxy-2-deoxyguanosine, we measured oxidative DNA damage in the cerebral cortical tissues from MCAT, AβPP, MCAT/AβPP and WT mice. We found that the AβPP transgenic mice that carried the human MCAT gene lived 5 months longer than did the AβPP mice. We also found that the overexpression of MCAT in the brain sections from the MCAT/AβPP transgenic mice significantly correlated with a reduction in the levels of full-length APP, CTF99, BACE1, Aβ levels (40 and 42), Aβ deposits and oxidative DNA damage relative to the brain sections from the AβPP mice. Interestingly, we found significantly increased levels of soluble APPα and CTF83 in the MCAT/AβPP mice, relative to the AβPP mice. These data provide direct evidence that oxidative stress plays a primary role in AD etiopathology and that in MCAT mice express Aβ, MCAT prevents abnormal APP processing, reduces Aβ levels and enhances Aβ-degrading enzymes in mice at different ages, corresponding to different stages of disease progression. These findings indicate that mitochondria-targeted molecules may be an effective therapeutic approach to treat patients with AD.
PMCID: PMC3373244  PMID: 22492996
8.  Ex-vivo imaging of excised tissue using vital dyes and confocal microscopy 
Current Protocols in Cytometry  2012;CHAPTER:Unit9.39.
Vital dyes routinely used for staining cultured cells can also be used to stain and image live tissue slices ex-vivo. Staining tissue with vital dyes allows researchers to collect structural and functional data simultaneously and can be used for qualitative or quantitative fluorescent image collection. The protocols presented here are useful for structural and functional analysis of viable properties of cells in intact tissue slices, allowing for the collection of data in a structurally relevant environment. With these protocols, vital dyes can be applied as a research tool to disease processes and properties of tissue not amenable to cell culture based studies.
PMCID: PMC3401092  PMID: 22752953
9.  Cardiac Aging: From Molecular Mechanisms to Significance in Human Health and Disease 
Antioxidants & Redox Signaling  2012;16(12):1492-1526.
Cardiovascular diseases (CVDs) are the major causes of death in the western world. The incidence of cardiovascular disease as well as the rate of cardiovascular mortality and morbidity increase exponentially in the elderly population, suggesting that age per se is a major risk factor of CVDs. The physiologic changes of human cardiac aging mainly include left ventricular hypertrophy, diastolic dysfunction, valvular degeneration, increased cardiac fibrosis, increased prevalence of atrial fibrillation, and decreased maximal exercise capacity. Many of these changes are closely recapitulated in animal models commonly used in an aging study, including rodents, flies, and monkeys. The application of genetically modified aged mice has provided direct evidence of several critical molecular mechanisms involved in cardiac aging, such as mitochondrial oxidative stress, insulin/insulin-like growth factor/PI3K pathway, adrenergic and renin angiotensin II signaling, and nutrient signaling pathways. This article also reviews the central role of mitochondrial oxidative stress in CVDs and the plausible mechanisms underlying the progression toward heart failure in the susceptible aging hearts. Finally, the understanding of the molecular mechanisms of cardiac aging may support the potential clinical application of several “anti-aging” strategies that treat CVDs and improve healthy cardiac aging.
I. Introduction
II. Aging and Epidemiology of CVDs
III. Physiology of Cardiac Aging
A. Ventricular changes
B. Valvular changes
IV. Animal Models of Cardiac Aging
A. Rodents
B. Drosophila
C. Canines
D. Nonhuman primates
V. Mitochondria and the Free Radical Theory of Aging
A. ROS and aging
B. Pleiotropy of ROS
C. Mitochondrial hormesis in aging
D. Mitochondrial turnover in aging
VI. Molecular Mechanisms of Cardiac Aging
A. Mitochondrial oxidative stress in cardiac aging
B. Neurohormonal regulation of cardiac aging
1. Renin-angiotensin system in cardiac aging
2. B-adrenergic signaling
3. Insulin/insulin-like growth factor 1/PI3K signaling
4. Natriuretic peptides signaling
C. Nutrient signaling in cardiac aging
D. Cardiac stem cell aging and telomeres
VII. Aging, Oxidative Stress, and CVDs
A. Oxidative stress and mitochondria in CVDs
1. The central role of mitochondrial oxidative stress and redox status in hypertension and heart failure
2. The role of mitochondria and oxidative stress in IR injury
B. Mechanisms of progression to heart failure in the aged hypertrophic heart
1. Increased cardiomyocyte death
2. ECM remodeling
3. Alteration of calcium handling proteins
4. Hypoxic response and angiogenesis
5. Mitochondrial dysfunction and abnormalities in energetics
VIII. Exercise, Cardiovascular Risks, and Cardiac Aging
IX. Emerging “Anti-Aging” Interventional Strategies for Cardiac Aging and CVDs
A. Dietary restriction
B. Antioxidant interventions
1. Nontargeted antioxidants
2. Mitochondrial-targeted antioxidants
a. TPP+conjugated antioxidants
b. Szeto-schiller peptides
C. Resveratrol and SIRTs activators
X. Conclusion and Future Directions
PMCID: PMC3329953  PMID: 22229339
10.  Rapamycin Reverses Elevated mTORC1 Signaling in Lamin A/C–Deficient Mice, Rescues Cardiac and Skeletal Muscle Function, and Extends Survival 
Science translational medicine  2012;4(144):144ra103.
Mutations in LMNA, the gene that encodes A-type lamins, cause multiple diseases including dystrophies of the skeletal muscle and fat, dilated cardiomyopathy, and progeria-like syndromes (collectively termed laminopathies). Reduced A-type lamin function, however, is most commonly associated with skeletal muscle dystrophy and dilated cardiomyopathy rather than lipodystrophy or progeria. The mechanisms underlying these diseases are only beginning to be unraveled. We report that mice deficient in Lmna, which corresponds to the human gene LMNA, have enhanced mTORC1 (mammalian target of rapamycin complex 1) signaling specifically in tissues linked to pathology, namely, cardiac and skeletal muscle. Pharmacologic reversal of elevated mTORC1 signaling by rapamycin improves cardiac and skeletal muscle function and enhances survival in mice lacking A-type lamins. At the cellular level, rapamycin decreases the number of myocytes with abnormal desmin accumulation and decreases the amount of desmin in both muscle and cardiac tissue of Lmna–/– mice. In addition, inhibition of mTORC1 signaling with rapamycin improves defective autophagic-mediated degradation in Lmna–/– mice. Together, these findings point to aberrant mTORC1 signaling as a mechanistic component of laminopathies associated with reduced A-type lamin function and offer a potential therapeutic approach, namely, the use of rapamycin-related mTORC1 inhibitors.
PMCID: PMC3613228  PMID: 22837538
11.  Mitochondrial proteome remodelling in pressure overload-induced heart failure: the role of mitochondrial oxidative stress 
Cardiovascular Research  2011;93(1):79-88.
We investigate the role of mitochondrial oxidative stress in mitochondrial proteome remodelling using mouse models of heart failure induced by pressure overload.
Methods and results
We demonstrate that mice overexpressing catalase targeted to mitochondria (mCAT) attenuate pressure overload-induced heart failure. An improved method of label-free unbiased analysis of the mitochondrial proteome was applied to the mouse model of heart failure induced by transverse aortic constriction (TAC). A total of 425 mitochondrial proteins were compared between wild-type and mCAT mice receiving TAC or sham surgery. The changes in the mitochondrial proteome in heart failure included decreased abundance of proteins involved in fatty acid metabolism, an increased abundance of proteins in glycolysis, apoptosis, mitochondrial unfolded protein response and proteolysis, transcription and translational control, and developmental processes as well as responses to stimuli. Overexpression of mCAT better preserved proteins involved in fatty acid metabolism and attenuated the increases in apoptotic and proteolytic enzymes. Interestingly, gene ontology analysis also showed that monosaccharide metabolic processes and protein folding/proteolysis were only overrepresented in mCAT but not in wild-type mice in response to TAC.
This is the first study to demonstrate that scavenging mitochondrial reactive oxygen species (ROS) by mCAT not only attenuates most of the mitochondrial proteome changes in heart failure, but also induces a subset of unique alterations. These changes represent processes that are adaptive to the increased work and metabolic requirements of pressure overload, but which are normally inhibited by overproduction of mitochondrial ROS.
PMCID: PMC3243039  PMID: 22012956
Mitochondria; Oxidative stress; Proteome; Pressure overload; Cardiomyopathy
12.  A novel radial water tread maze tracks age-related cognitive decline in mice 
Pathobiology of Aging & Age Related Diseases  2013;3:10.3402/pba.v3i0.20679.
There is currently no treatment and cure for age-related dementia and cognitive impairment in humans. Mice suffer from age-related cognitive decline just as people do, but assessment is challenging because of cumbersome and at times stressful performance tasks. We developed a novel radial water tread (RWT) maze and tested male C57BL/6 (B6) and C57BL/6 x Balb/c F1 (CB6F1) mice at ages 4, 12, 20, and 28 months. B6 mice showed a consistent learning experience and memory retention that gradually decreased with age. CB6F1 mice showed a moderate learning experience in the 4 and 12 month groups, which was not evident in the 20 and 28 month groups. In conclusion, CB6F1 mice showed more severe age-related cognitive impairment compared to B6 mice and might be a suitable model for intervention studies. In addition, the RWT maze has a number of operational advantages compared to currently accepted tasks and can be used to assess age-related cognition impairment in B6 and CB6F1 mice as early as 12 months of age.
PMCID: PMC3791354  PMID: 24106580
memory impairment; aging; water tread radial maze; mouse cognition
13.  Correction: Cardiomyocyte-Specific Expression of Lamin A Improves Cardiac Function in Lmna−/− Mice 
PLoS ONE  2012;7(9):10.1371/annotation/92be6b32-d8e7-44c2-80a9-21097ad27965.
PMCID: PMC3465171
14.  Cardiomyocyte-Specific Expression of Lamin A Improves Cardiac Function in Lmna−/− Mice 
PLoS ONE  2012;7(8):e42918.
Lmna−/− mice display multiple tissue defects and die by 6–8 weeks of age reportedly from dilated cardiomyopathy with associated conduction defects. We sought to determine whether restoration of lamin A in cardiomyocytes improves cardiac function and extends the survival of Lmna−/− mice. We observed increased total desmin protein levels and disorganization of the cytoplasmic desmin network in ∼20% of Lmna−/− ventricular myocytes, rescued in a cell-autonomous manner in Lmna−/− mice expressing a cardiac-specific lamin A transgene (Lmna−/−; Tg). Lmna−/−; Tg mice displayed significantly increased contractility and preservation of myocardial performance compared to Lmna−/− mice. Lmna−/−; Tg mice attenuated ERK1/2 phosphorylation relative to Lmna−/− mice, potentially underlying the improved localization of connexin43 to the intercalated disc. Electrocardiographic recordings from Lmna−/− mice revealed arrhythmic events and increased frequency of PR interval prolongation, which is partially rescued in Lmna−/−; Tg mice. These findings support our observation that Lmna−/−; Tg mice have a 12% median extension in lifespan compared to Lmna−/− mice. While significant, Lmna−/−; Tg mice only have modest improvement in cardiac function and survival likely stemming from the observation that only 40% of Lmna−/−; Tg cardiomyocytes have detectable lamin A expression. Cardiomyocyte-specific restoration of lamin A in Lmna−/− mice improves heart-specific pathology and extends lifespan, demonstrating that the cardiac pathology of Lmna−/− mice limits survival. The expression of lamin A is sufficient to rescue certain cellular defects associated with loss of A-type lamins in cardiomyocytes in a cell-autonomous fashion.
PMCID: PMC3419749  PMID: 22905185
15.  Mitochondrial oxidative stress mediates induction of autophagy and hypertrophy in angiotensin-II treated mouse hearts 
Autophagy  2011;7(8):917-918.
Autophagy is characterized by recycling of cellular organelles and can be induced by several stimuli, including nutrient deprivation and oxidative stress. As a major site of free radical production during oxidative phosphorylation, mitochondria are believed to be primary targets of oxidative damage during stress. Our recent study demonstrated that angiotensin II increases cardiac mitochondrial reactive oxygen species (ROS) production, causes a decline of mitochondrial membrane potential in cardiomyocytes and increases cardiac mitochondrial protein oxidative damage and mitochondrial DNA deletions. The deleterious effects of angiotensin II on mitochondria are associated with an increase in autophagosomes and increased signaling of mitochondrial biogenesis, interpreted as an attempt to replenish the damaged mitochondria and restore energy production. Direct evidence for the central role of mitochondrial ROS was investigated by comparing the effect on mice overexpressing catalase targeted to mitochondria (mCAT) and mice overexpressing peroxisomal targeted catalase (pCAT, the natural site of catalase) challenged by angiotensin II or Gαq overexpression. The mCAT, but not pCAT, mice are resistant to cardiac hypertrophy, fibrosis and mitochondrial damage, biogenesis and autophagy induced by angiotensin II, as well as heart failure induced by overexpression of Gαq.
PMCID: PMC3359471  PMID: 21505274
mitochondria; autophagy; reactive oxygen species; catalase; angiotensin II; heart failure
16.  Ulcerative colitis-associated colorectal cancer arises in a field of short telomeres, senescence, and inflammation 
Cancer research  2011;71(5):1669-1679.
Inflammation plays a role in the progression to cancer and it is linked to the presence of senescent cells. Ulcerative colitis (UC) is a chronic inflammatory disease that predisposes to colorectal cancer. Tumorigenesis in this setting is associated with telomere shortening that can be observed in the non-dysplastic epithelium of UC patients with high-grade dysplasia (HGD) or cancer (UC Progressors). We hypothesized that a pre-neoplastic field of inflammation, telomere shortening, and senescence underlies tumor progression in UC Progressors. Multiple biopsies of varying histological grade were collected along the colon of 9 UC Progressors and analyzed for telomere length, DNA damage, senescence, p53, p16, and chronic and acute inflammation. Twenty biopsies from 4 UC non- Progressors and 21 biopsies from control individuals without UC were also analyzed. Short telomeres and increased DNA damage, senescence, and infiltrating leukocytes were observed in biopsies located less than 10 cm from HGD or cancer. Low-grade dysplasia had the shortest telomeres along with the highest levels of senescence and infiltrating leukocytes, whereas HGD biopsies showed the opposite pattern. p16 and p53 expression was low in non-dysplastic biopsies, but progressively increased in LGD and HGD. Additionally, high levels of infiltrating leukocytes were associated with telomere shortening, senescence, and reduced p53 expression. These results suggest that dysplasia arises in a pre-neoplastic field of chronic inflammation which leads to telomere shortening, DNA damage, and senescence. Our findings argue that senescence acts as a tumor suppressor mechanism that is abrogated during the transition from LGD to HGD in ulcerative colitis.
PMCID: PMC3077943  PMID: 21363920
ulcerative colitis; telomere length; inflammation; senescence; colon
17.  Age-Dependent Cardiomyopathy in Mitochondrial Mutator Mice is Attenuated by Overexpression of Catalase Targeted to Mitochondria 
Aging Cell  2010;9(4):536-544.
Mitochondrial defects have been found in aging and several age-related diseases. Mice with a homozygous mutation in the exonuclease encoding domain of mitochondrial DNA polymerase gamma (Polgm/m) are prone to age-dependent accumulation of mitochondrial DNA mutations and have shown a broad spectrum of aging-like phenotypes. However, the mechanism of cardiac phenotypes in relation to the role of mitochondrial DNA mutations and oxidative stress in this mouse model has not been fully addressed. We demonstrate age-dependent cardiomyopathy in Polgm/m mice, which by 13-14 months of age displays marked cardiac hypertrophy and dilatation, impairment of systolic and diastolic function and increased cardiac fibrosis. This age-dependent cardiomyopathy is associated with increases in mitochondrial DNA (mtDNA) deletions and protein oxidative damage, increased expression of apoptotic and senescence markers, as well as a decline in signaling for mitochondrial biogenesis. The relationship of these changes to mitochondrial reactive oxygen species (ROS) was tested by crossing Polgm/m mice with mice that overexpress mitochondrial targeted catalase (mCAT). All of the above phenotypes were partially rescued in Polgm/m/mCAT mice. These data indicate that accumulation of mitochondrial DNA damage with age can lead to cardiomyopathy, and that this phenotype is partly mediated by mitochondrial oxidative stress.
PMCID: PMC3265170  PMID: 20456298
mitochondria; mutation; aging; cardiomyopathy; oxidative stress
18.  Deletion at Fragile Sites is a Common and Early Event in Barrett’s Esophagus 
Molecular cancer research : MCR  2010;8(8):1084-1094.
Barrett’s esophagus is a premalignant intermediate to esophageal adenocarcinoma, which develops in the context of chronic inflammation and exposure to bile and acid. We asked whether there might be common genomic alterations that could be identified as potential clinical biomarker(s) for Barrett’s esophagus by whole genome profiling. We detected copy number alterations and/or loss of heterozygosity (LOH) at fifty-six fragile sites in 20 patients with premalignant Barrett’s esophagus (BE). Chromosomal fragile sites are particularly sensitive to DNA breaks and have been shown to be frequent sites of rearrangement or loss in many human cancers. 78% of all genomic alterations detected by array-CGH were associated with fragile sites. Copy number losses in early BE were observed at particularly high frequency at FRA3B (81%), FRA9A/C (71.4%), FRA5E (52.4%) and FRA 4D (52.4%), and at lower frequencies in other fragile sites, including FRA1K (42.9%), FRAXC (42.9%), FRA 12B (33.3%) and FRA16D (33.3%). Due to the consistency of the region of copy number loss, we were able to verify these results by quantitative PCR which detected loss of FRA3B and FRA16D, in 83% and 40% of early molecular stage BE patients respectively. LOH in these cases was confirmed via pyrosequencing at FRA3B and FRA16D (75% and 70% respectively). Deletion and genomic instability at FRA3B and other fragile sites could thus be a biomarker of genetic damage in BE patients and a potential biomarker of cancer risk.
PMCID: PMC3100793  PMID: 20647332
19.  Leukocyte Telomere Length Is Associated With Disability In Older U.S. Population 
To determine whether the mean leukocyte telomere length (LTL) serves as a biomarker of disability assessed by Activities of Daily Living (ADL) and what factors may modify this relationship.
Retrospective cross-sectional study.
A subset of the National Long Term Care Survey (NTLCS), a Medicare-based U.S. population longitudinal study focused on trends of overall health and functional status in the elderly.
Six hundred and twenty four individuals from the 1999 wave of the NTLCS cohort.
Relative LTL determined by quantitative PCR. LTL has previously been shown to correlate with common age-related disorders and mortality, as well as with socioeconomical status.
We observed gender difference of LTL, but not age-dependent shortening or association with socio-economical status. Importantly, LTL was associated with disability and functional status assessed by ADL. The association between ADL and LTL was more significant among non-diabetic subjects, while associations were not seen when diabetic subjects only were analyzed. Associations of LTL with cardiovascular diseases and cancer were also present in the non-diabetic group, but not in the diabetic group.
Our findings support the concept that LTL is a biomarker of overall well-being that is predictive of disability of older individuals in the US population. Diabetes plays an important role as a modifier of the association of LTL with disability, cardiovascular diseases, and cancer. These associations have obvious clinical implications due to the potential predictive value of LTL and deserve further investigation.
PMCID: PMC2918372  PMID: 20579170
disability; telomere; aging; disease; human
20.  Mitochondrial oxidative stress and mammalian healthspan 
Mechanisms of ageing and development  2010;131(7-8):527-535.
Aging of the American society is leading to a growing need for disease-modifying interventions to treat age-related diseases and enhance healthspan. Mitochondria and mitochondrially-generated reactive oxygen species appear to play a central role in these processes and are a likely target for interventions. Conventional, untargeted antioxidants have not demonstrated a clear benefit in human studies. As a result, approaches have been developed to target antioxidants specifically to mitochondria. Studies have employed a wide array of targeted molecules including antioxidant enzymes such as catalase, peroxiredoxin, superoxide dismutases and small molecular compounds which recapitulate the antioxidant activities of these enzymes. Lifespan and healthspan effects differ between interventions suggesting varied roles for specific mitochondrial reactive oxygen species and their impact on usual aging. Consistent findings in myocardial protection across various interventions support a focus on the impact of cardiac aging on healthspan. The advancement of mitochondrially-targeted small molecule antioxidants suggests the prospect of swift translation to human use.
PMCID: PMC2933331  PMID: 20566356
Mitochondria; antioxidants; drug targeting; healthspan
21.  Array Comparative Genomic Hybridization in Ulcerative Colitis Neoplasia: Single Non-Dysplastic Biopsies Distinguish Progressors from Non-Progressors 
Approximately 10% of ulcerative colitis patients develop colorectal neoplasia. At present, identification of this subset is markedly limited and necessitates lifelong colonoscopic surveillance for the entire ulcerative colitis population. Better risk markers are needed to focus surveillance onto the patients most likely to benefit. Using array-based comparative genomic hybridization, we analyzed single, non-dysplastic biopsies from three patient groups: ulcerative colitis progressors (n=9) with cancer or high-grade dysplasia at a mean distance of 18 cm from the analyzed site; ulcerative colitis nonprogressors (n=8) without dysplasia during long-term surveillance; and non-ulcerative colitis normal controls (n=2). Genomic DNA from fresh colonic epithelium purified from stroma was hybridized to 287 (low-density) and 4,342 (higher-density) feature bacterial artificial chromosome arrays. Sample-to-reference fluorescence ratios were calculated for individual chromosomal targets and globally across the genome. The low-density arrays yielded pronounced genomic gains and losses in 3 of 9 (33%) ulcerative colitis progressors but in none of the 10 control patients. Identical DNA samples analyzed on the higher density arrays, using a combination of global and individual high variance assessments, distinguished all 9 progressors from all 10 controls. These data confirm that genomic alterations in ulcerative colitis progressors are widespread, even involving single non-dysplastic biopsies far distant from neoplasia. They therefore show promise toward eliminating full colonoscopic surveillance with extensive biopsy sampling in the majority of ulcerative colitis patients.
PMCID: PMC2932629  PMID: 20802465
Ulcerative Colitis; Array Comparative Genomic Hybridization; Colonic Neoplasia
22.  Targeted Expression of Catalase to Mitochondria Prevents Age-Associated Reductions in Mitochondrial Function and Insulin Resistance 
Cell metabolism  2010;12(6):668-674.
Aging-associated muscle insulin resistance has been hypothesized to be due to decreased mitochondrial function, secondary to cumulative free radical damage, leading to increased intramyocellular lipid content. To directly test this hypothesis we examined both in vivo and in vitro mitochondrial function, intramyocellular lipid content and insulin action in lean healthy mice with targeted overexpression of the human catalase gene to mitochondria (MCAT mice). Here we show that MCAT mice are protected from age-induced decrease in muscle mitochondrial function (~30%), energy metabolism (~7%) and lipid-induced muscle insulin resistance. This protection from age-induced reduction in mitochondrial function was associated with reduced mitochondrial oxidative damage, preserved mitochondrial respiration and muscle ATP synthesis and AMP-activated protein kinase-induced mitochondrial biogenesis. Taken together these data suggest that the preserved mitochondrial function maintained by reducing mitochondrial oxidative damage may prevent age-associated whole body energy imbalance and muscle insulin resistance.
PMCID: PMC3013349  PMID: 21109199
23.  Genetic variation in prostaglandin E2 synthesis and signaling, prostaglandin dehydrogenase, and risk of colorectal adenoma 
Prostaglandins are important inflammatory mediators; PGE2 is the predominant prostaglandin in colorectal neoplasia and affects colorectal carcinogenesis. Prostaglandins are metabolites of omega-6 and omega-3 polyunsaturated fatty acids; their biosynthesis is the primary target of nonsteroidal anti-inflammatory drugs (NSAIDs), which reduce colorectal neoplasia risk.
We investigated candidate and tagSNPs in PGE2 synthase (PGES), PGE2 receptors (EP2 and EP4), and prostaglandin dehydrogenase (PGDH) in a case-control study of adenomas (n=483) vs. polyp-free controls (n=582) and examined interactions with NSAID use or fish intake, a source of omega-3 fatty acids.
A 30% adenoma risk reduction was observed for EP2 4950G>A (intron 1; ORGA/AA vs. GG: 0.71; 95% CI: 0.52-0.99). For the candidate polymorphism EP4 Val294Ile, increasing fish intake was associated with increased adenoma risk among those with variant genotypes, but not among those with the Val/Val genotype, (p-interaction=0.02). An interaction with fish intake was also observed for PGES -664A>T (5’UTR; p-interaction=0.01). Decreased risk with increasing fish intake was only seen among those with the AT or TT genotypes (OR>2 t/wk vs. <1 t/wk: 0.56; 95% CI: 0.28-1.13). We also detected interactions between NSAIDs and EP2 9814C>A (intron 1) and PGDH 343C>A (intron 1). However, none of the observed associations was statistically significant after adjustment for multiple testing. We investigated potential gene-gene interactions using the Chatterjee 1df Tukey test and logic regression; neither method detected significant interactions.
These data provide little support for associations between adenoma risk and genetic variability related to PGE2, yet suggest gene-environment interactions with anti-inflammatory exposures.
PMCID: PMC2849740  PMID: 20086108
prostaglandins; colorectal cancer; colorectal polyps; NSAIDs; aspirin; fish; fat intake
24.  Divergent cellular phenotypes of human and mouse cells lacking the Werner syndrome RecQ helicase 
DNA repair  2009;9(1):11.
Werner syndrome (WS) is a human autosomal recessive genetic instability and cancer predisposition syndrome with features of premature aging. Several genetically determined mouse models of WS have been generated, however none develops features of premature aging or an elevated risk of neoplasia unless additional genetic perturbations are introduced. In order to determine whether differences in cellular phenotype could explain the discrepant phenotypes of Wrn−/− mice and WRN-deficient humans, we compared the cellular phenotype of newly derived Wrn−/− mouse primary fibroblasts with previous analyses of primary and transformed fibroblasts from WS patients and with newly derived, WRN-depleted human primary fibroblasts. These analyses confirmed previously reported cellular phenotypes of WRN-mutant and WRN-deficient human fibroblasts, and demonstrated that the human WRN-deficient cellular phenotype can be detected in cells grown in 5% or in 20% oxygen. In contrast, we did not identify prominent cellular phenotypes present in WRN-deficient human cells in Wrn−/− mouse fibroblasts. Our results indicate that human and mouse fibroblasts have different functional requirements for WRN protein, and that the absence of a strong cellular phenotype may in part explain the failure of Wrn−/− mice to develop an organismal phenotype resembling Werner syndrome.
PMCID: PMC2818259  PMID: 19896421
25.  Cardiac Aging in Mice and Humans: the Role of Mitochondrial Oxidative Stress 
Trends in cardiovascular medicine  2009;19(7):213-220.
Age is a major risk factor for cardiovascular diseases, not only because it prolongs exposure to several other cardiovascular risks, but also owing to intrinsic cardiac aging, which reduces cardiac functional reserve, predisposes the heart to stress and contributes to increased cardiovascular mortality in the elderly. Intrinsic cardiac aging in the murine model closely recapitulates age-related cardiac changes in humans, including left ventricular hypertrophy, fibrosis and diastolic dysfunction. Cardiac aging in mice is accompanied by accumulation of mitochondrial protein oxidation, increased mitochondrial DNA mutations, increased mitochondrial biogenesis, as well as decreased cardiac SERCA2 protein. All of these age-related changes are significantly attenuated in mice overexpressing catalase targeted to mitochondria (mCAT). These findings demonstrate the critical role of mitochondrial reactive oxygen species (ROS) in cardiac aging and support the potential application of mitochondrial antioxidants to cardiac aging and age-related cardiovascular diseases.
PMCID: PMC2858758  PMID: 20382344

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