The Future of Onco-Cardiology: We Are Not Just "Side Effect Hunters".

The clinical correlations linking diabetes mellitus with accelerated atherosclerosis, cardiomyopathy, and increased post-myocardial infarction fatality rates are increasingly understood in mechanistic terms. The multiple mechanisms discussed in this review seem to share a common element: prolonged increases in reactive oxygen species (ROS) production in diabetic cardiovascular cells. Intracellular hyperglycemia causes excessive ROS production. This activates nuclear poly(ADP-ribose) polymerase, which inhibits GAPDH, shunting early glycolytic intermediates into pathogenic signaling pathways. ROS and poly(ADP-ribose) polymerase also reduce sirtuin, PGC-1α, and AMP-activated protein kinase activity. These changes cause decreased mitochondrial biogenesis, increased ROS production, and disturbed circadian clock synchronization of glucose and lipid metabolism. Excessive ROS production also facilitates nuclear transport of proatherogenic transcription factors, increases transcription of the neutrophil enzyme initiating NETosis, peptidylarginine deiminase 4, and activates the NOD-like receptor family, pyrin domain-containing 3 inflammasome. Insulin resistance causes excessive cardiomyocyte ROS production by increasing fatty acid flux and oxidation. This stimulates overexpression of the nuclear receptor PPARα and nuclear translocation of forkhead box O 1, which cause cardiomyopathy. ROS also shift the balance between mitochondrial fusion and fission in favor of increased fission, reducing the metabolic capacity and efficiency of the mitochondrial electron transport chain and ATP synthesis. Mitochondrial oxidative stress also plays a central role in angiotensin II-induced gap junction remodeling and arrhythmogenesis. ROS contribute to sudden death in diabetics after myocardial infarction by increasing post-translational protein modifications, which cause increased ryanodine receptor phosphorylation and downregulation of sarco-endoplasmic reticulum Ca(++)-ATPase transcription. Increased ROS also depress autonomic ganglion synaptic transmission by oxidizing the nAch receptor α3 subunit, potentially contributing to the increased risk of fatal cardiac arrhythmias associated with diabetic cardiac autonomic neuropathy.

Telomere Dysfunction and DNA Damage Checkpoints in Diseases and Cancer of the Gastrointestinal Tract

Mitochondria are dynamic organelles that continuously change their shape through fission and fusion. Disruption of mitochondrial dynamics is involved in disease pathology through excessive reactive oxygen species (ROS) production. Accelerated cellular senescence resulting from cigarette smoke exposure with excessive ROS production has been implicated in the pathogenesis of chronic obstructive pulmonary disease (COPD). Hence, we investigated the involvement of mitochondrial dynamics and ROS production in terms of cigarette smoke extract (CSE)-induced cellular senescence in human bronchial epithelial cells (HBEC). Mitochondrial morphology was examined by electron microscopy and fluorescence microscopy. Senescence-associated β-galactosidase staining and p21 Western blotting of primary HBEC were performed to evaluate cellular senescence. Mitochondrial-specific superoxide production was measured by MitoSOX staining. Mitochondrial fragmentation was induced by knockdown of mitochondrial fusion proteins (OPA1 or Mitofusins) by small-interfering RNA transfection. N-acetylcysteine and Mito-TEMPO were used as antioxidants. Mitochondria in bronchial epithelial cells were prone to be more fragmented in COPD lung tissues. CSE induced mitochondrial fragmentation and mitochondrial ROS production, which were responsible for acceleration of cellular senescence in HBEC. Mitochondrial fragmentation induced by knockdown of fusion proteins also increased mitochondrial ROS production and percentages of senescent cells. HBEC senescence and mitochondria fragmentation in response to CSE treatment were inhibited in the presence of antioxidants. CSE-induced mitochondrial fragmentation is involved in cellular senescence through the mechanism of mitochondrial ROS production. Hence, disruption of mitochondrial dynamics may be a part of the pathogenic sequence of COPD development.

Journal of Cardiac SurgeryVolume 32, Issue 3 p. 233-234 IMAGES IN CARDIAC SURGERY Perforation of a HeartMate II outflow graft Masashi Kawabori MD, Corresponding Author Masashi Kawabori MD kawabori.masashi@gmail.com Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, Texas Correspondence Masashi Kawabori MD, Texas Heart Institute, 6770 Bertner Avenue, Houston, TX 77030. Email: kawabori.masashi@gmail.comSearch for more papers by this authorChitaru Kurihara MD, Chitaru Kurihara MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this authorWilliam E. Cohn MD, William E. Cohn MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this authorAndrew B. Civitello MD, Andrew B. Civitello MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this authorO. H. Frazier MD, O. H. Frazier MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this authorJeffrey A. Morgan MD, Jeffrey A. Morgan MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this author Masashi Kawabori MD, Corresponding Author Masashi Kawabori MD kawabori.masashi@gmail.com Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, Texas Correspondence Masashi Kawabori MD, Texas Heart Institute, 6770 Bertner Avenue, Houston, TX 77030. Email: kawabori.masashi@gmail.comSearch for more papers by this authorChitaru Kurihara MD, Chitaru Kurihara MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this authorWilliam E. Cohn MD, William E. Cohn MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this authorAndrew B. Civitello MD, Andrew B. Civitello MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this authorO. H. Frazier MD, O. H. Frazier MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this authorJeffrey A. Morgan MD, Jeffrey A. Morgan MD Division of Cardiothoracic Transplantation and Circulatory Support, Texas Heart Institute, Baylor College of Medicine, Houston, TexasSearch for more papers by this author First published: 01 March 2017 https://doi.org/10.1111/jocs.13115Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. Volume32, Issue3March 2017Pages 233-234 RelatedInformation

During reperfusion, the interplay between excess reactive oxygen species (ROS) production, mitochondrial Ca(2+) overload, and mitochondrial permeability transition pore (mPTP) opening, as the crucial mechanism of cardiomyocyte injury, remains intriguing. Here, we investigated whether an induction of a partial decrease in mitochondrial membrane potential (DeltaPsi(m)) is an underlying mechanism of protection by anesthetic-induced preconditioning (APC) with isoflurane, specifically addressing the interplay between ROS, Ca(2+), and mPTP opening. The magnitude of APC-induced decrease in DeltaPsi(m) was mimicked with the protonophore 2,4-dinitrophenol (DNP), and the addition of pyruvate was used to reverse APC- and DNP-induced decrease in DeltaPsi(m). In cardiomyocytes, DeltaPsi(m), ROS, mPTP opening, and cytosolic and mitochondrial Ca(2+) were measured using confocal microscope, and cardiomyocyte survival was assessed by Trypan blue exclusion. In isolated cardiac mitochondria, antimycin A-induced ROS production and Ca(2+) uptake were determined spectrofluorometrically. In cells exposed to oxidative stress, APC and DNP increased cell survival, delayed mPTP opening, and attenuated ROS production, which was reversed by mitochondrial repolarization with pyruvate. In isolated mitochondria, depolarization by APC and DNP attenuated ROS production, but not Ca(2+) uptake. However, in stressed cardiomyocytes, a similar decrease in DeltaPsi(m) attenuated both cytosolic and mitochondrial Ca(2+) accumulation. In conclusion, a partial decrease in DeltaPsi(m) underlies cardioprotective effects of APC by attenuating excess ROS production, resulting in a delay in mPTP opening and an increase in cell survival. Such decrease in DeltaPsi(m) primarily attenuates mitochondrial ROS production, with consequential decrease in mitochondrial Ca(2+) uptake.

Introduction Excessive reactive oxygen species (ROS) production is a prominent feature of obesity‐related conditions including impairments in endothelial function. NADPH oxidases (Nox) and mitochondria are major sources of ROS production within the vasculature. However, studies involving direct measurement of in vivo ROS production are limited in humans. Thus, the relative contributions of Nox‐ and mitochondrial‐derived ROS in human obesity remain to be fully characterized. The objective of this study was to determine the predominant sources of in vivo ROS production in the skeletal muscle microvasculature of individuals with obesity. We hypothesize that both Nox‐ and mitochondria‐dependent ROS production are upregulated in human obesity. Methods Two sedentary men with Class II and above obesity (body mass index ≥ 35 kg/m 2 ) and the Metabolic Syndrome were studied. Microdialysis was used for measurements of in vivo skeletal muscle ROS production. Microdialysis probes were perfused with saline containing Amplex Ultrared, horseradish peroxidase and superoxide dismutase to measure local hydrogen peroxide and superoxide concentrations. To assess ROS production from distinct sources, microdialysis probes were perfused with the following: apocynin (non‐specific Nox inhibitor), MitoTEMPO (mitochondrial‐ROS inhibitor), and GKT 137831 (specific Nox 4 inhibitor). Results The concentrations of hydrogen peroxide and superoxide decreased upon addition of apocynin to microdialysis probes (mean ± SEM; hydrogen peroxide: 1.18 ± 0.58 to 0.76 ± 0.01 μM; superoxide: 1.47 ± 0.46 to 1.10 ± 0.62 μM). MitoTEMPO perfusion elicited an increase in hydrogen peroxide (1.12 ± 0.76 to 1.51 ± 1.08 μM) and decrease in superoxide (1.47 ± 0.46 to 0.78 ± 0.10 μM). The combination of apocynin plus MitoTEMPO resulted in concentrations of hydrogen peroxide (0.22 ± 0.10 μM) and superoxide (0.23 ± 0.06 μM) that were 69.7% and 79.1% lower than apocynin alone. In addition, the hydrogen peroxide (0.56 ± 0.25 μM) and superoxide (0.43 ± 0.18 μM) concentrations with co‐perfusion of apocynin and GKT were 26.3% and 60.9% lower than with apocynin alone. Conclusions Our preliminary data suggest that mitochondria and Nox together are significant sources of ROS production within the skeletal muscle microvasculature of obese individuals. The local inhibition of Nox‐ and mitochondrial‐derived ROS resulted in marked decreases in extracellular ROS levels within the skeletal muscle microvasculature. Although the small sample size currently limits our interpretation of the results, this study is anticipated to become the first to report on distinct sources of in vivo ROS production in human obesity.

<b>Introduction:</b> Early coronary reperfusion has been established as the best therapeutic strategy to limit infarct size and improve prognosis. Therefore, elucidating the mechanisms underlying cardiomyocyte death may yield novel therapeutic targets to limit ischemia reperfusion (I/R) injury. I/R is accompanied and influenced by perturbations of the β-adrenergic receptor pathway which acts through cAMP dependent signaling cascade to modulate cardiac function and remodeling. <b>Purpose:</b> Although the involvement of the cAMP-binding protein Epac1 in cardiac hypertrophy and arrhythmia has been recently described, its role in I/R induced-cardiomyocyte death has not yet been investigated. <b>Methods:</b> Isolated adult cardiomyocytes from Epac1 knock-out (Epac1-/-) mice or wild-type (WT) littermates were exposed to hypoxia (HX) for 4h and 2h of reoxygenation period (HX+R). Cell death was determined by Trypan blue staining and LDH release. The mitochondrial permeability transition pore (mPTP) opening was monitored by the calcein loading CoCl(2)-quenching technique. The area at risk was examined by Evans blue and infarct size was evaluated by TTC staining. <b>Results:</b> Our data showed that HX+R-induced cardiomyocyte death were significantly prevented in adult cardiomyocytes isolated from Epac1-/- mice. In addition, we found that the increased expression of apoptotic markers (Bax, cleaved Caspase-9,Caspase-3) during HX+R conditions were also inhibited in Epac1-/- cardiomyocytes compared to WT cells. Interestingly, HX+R induced a decrease in calcein fluorescence corresponding to mPTP opening (56%±1.1 vs control cells set at 100%) in WT cardiomyocytes while genetic deletion of Epac1 prevented mPTP opening in HX+R conditions. Concomitantly, we found that the infarct size was significantly reduced in the Epac1-/- mice compared to the WT animals (53±4 % vs 33±4%, p<0.01) despite the same area at risk. <b>Conclusion:</b> Epac1 deletion confers resistance to I/R injury via the inhibition of a mitochondrial death signaling. Our study shed light the therapeutic potential of the inhibition of Epac1 and, the development of Epac1 inhibitors as new drugs to treat I/R injury.

The rate of mitosis of cancer cells is significantly higher than normal primary cells with increased metabolic needs, which in turn enhances the generation of reactive oxygen species (ROS) production. Higher ROS production is known to increase cancer cell dependence on ROS scavenging systems to counteract the increased ROS. Therapeutic options which selectively modulate the levels of intracellular ROS in cancers are likely candidates for drug discovery. Docetaxel (DTX) has demonstrated antitumor activity in preclinical and clinical studies. It is thought that DTX induces cell death through excessive ROS production and increased Ca2+ entry. The Ca2+ permeable TRPM2 channel is activated by ROS. Selenium (Se) has been previously used to stimulate apoptosis for the treatment of glioblastoma cells resistant to DTX. However, the potential mechanism(s) of the additive effect of DTX on TRPM2 channels in cancer cells remains unclear. The aim of this study was to evaluate the effect of combination therapy of DTX and Se on activation of TRPM2 in DBTRG glioblastoma cells. DBTRG cells were divided into four treatment groups: control, DTX (10nM for 10h), Se (1μM for 10h), and DTX+Se. Our study showed that apoptosis (Annexin V and propidium iodide), mitochondrial membrane depolarization (JC1), and ROS production levels were increased in DBTRG cells following treatment with Se and DTX respectively. Cell number and viability, and the levels of apoptosis, JC1, ROS, and [Ca2+]i, induced by DTX, were further increased following addition of Se. We also observed an additive increase in the activation of the NAD-dependent DNA repair enzyme poly (ADP-ribose) polymerase-1 (PARP-1) activity, which was accompanied by a decline in its essential substrate NAD+. As well, the Se- and DTX-induced increases in intracellular Ca2+ florescence intensity were decreased following treatment with the TRPM2 antagonist N-(p-amylcinnamoyl) anthranilic acid (ACA). Therefore, combination therapy with Se and DTX may represent an effective strategy for the treatment of glioblastoma cells and may be associated with TRPM2-mediated increases in oxidative stress and [Ca2+]i.

Glioma is a common primary malignant tumor that has a poor prognosis and often develops drug resistance. The coumarin derivative osthole has previously been reported to induce cancer cell apoptosis. Recently, we found that it could also trigger glioma cell necroptosis, a type of cell death that is usually accompanied with reactive oxygen species (ROS) production. However, the relationship between ROS production and necroptosis induced by osthole has not been fully elucidated. In this study, we found that osthole could induce necroptosis of glioma cell lines U87 and C6; such cell death was distinct from apoptosis induced by MG‐132. Expression of necroptosis inhibitor caspase‐8 was decreased, and levels of necroptosis proteins receptor‐interacting protein 1 (RIP1), RIP3 and mixed lineage kinase domain‐like protein were increased in U87 and C6 cells after treatment with osthole, whereas levels of apoptosis‐related proteins caspase‐3, caspase‐7, and caspase‐9 were not increased. Lactate dehydrogenase release and flow cytometry assays confirmed that cell death induced by osthole was primarily necrosis. In addition, necroptosis induced by osthole was accompanied by excessive production of ROS, as observed for other necroptosis‐inducing reagents. Pretreatment with the RIP1 inhibitor necrostatin‐1 attenuated both osthole‐induced necroptosis and the production of ROS in U87 cells. Furthermore, the ROS inhibitor N‐acetylcysteine decreased osthole‐induced necroptosis and growth inhibition. Overall, these findings suggest that osthole induces necroptosis of glioma cells via ROS production and thus may have potential for development into a therapeutic drug for glioma therapy.

Oxidative stress, resulting from increased production of reactive oxygen species (ROS) and/or reduced antioxidant defences, has been implicated in cardiovascular disease pathophysiology for over 2 decades. Based on the concept that this drives both the genesis and progression of conditions such as heart failure, numerous clinical trials of antioxidant therapies were undertaken but were unsuccessful. Nevertheless, experimental data linking oxidative stress and heart disease remain compelling and support continued efforts to develop more effective therapies than antioxidant vitamins.1 In the current issue of Circulation , Zhao et al .2 report that cardiomyocyte-specific high-level overexpression of the ROS-generating enzyme NADPH oxidase-4 (Nox4) aggravated angiotensin II-induced cardiac remodeling and was mitigated by a small molecule Nox inhibitor. The authors propose that Nox4 inhibition may have therapeutic potential to treat cardiac remodeling. Is this proposal reasonable and how should such studies be interpreted within a pathophysiological framework for the roles of ROS in heart failure?

Oxidative damage is thought to be a major causal factor for replicative senescence and human aging (Harman, 1994). Leakage of superoxide from the mitochondrial respiratory chain is an important source of oxidative stress (Raha & Robinson, 2000). Targeting antioxidants to mitochondria is an efficient way to attenuate oxidative damage in mitochondria due to the production of reactive oxygen species (ROS) in isolated mitochondria (Kelso et al., 2001; Echtay et al., 2002) and in mitochondria within cells (Kelso et al., 2001; Hwang et al., 2001). Therefore, by determining the effect of these antioxidants it should be possible to establish whether oxidative damage has a role in telomere shortening. It has been shown that selective targeting of a potent antioxidant to mitochondria is achieved by attaching the redox active moiety of ubiquinol to the decyltriphenylphosphonium (DPPT) cation, resulting in the mitochondria-specific antioxidant mitoQ [10-(6′-ubiquinonyl) decyltriphenylphosphonium bromide] (Kelso et al., 2001). Ubiquinol acts as an antioxidant by donating a hydrogen atom from one of its hydroxyl groups to a lipid peroxyl radical, which decreases lipid peroxidation within the mitochondrial inner membrane (Ingold et al., 1993). MitoQ reduces oxidative damage and decreases ROS-induced apoptosis in short-term experiments (Kelso et al., 2001; Echtay et al., 2002). As mitoQ is predominantly located within mitochondria in cells due to its accumulation by the mitochondrial membrane potential, its effects in cells are thought to be largely due to the prevention of mitochondrial oxidative damage (Kelso et al., 2001) and there is also evidence that mitoQ decreases the release of ROS from mitochondria (Hwang et al., 2001). The possibility of such effects being due to non-specific interactions with mitochondria within cells can be discounted by the use of control compounds such as DPPT, which are also accumulated within mitochondria driven by the membrane potential but which do not act as antioxidants. Therefore, the blocking of a process by mitoQ but not by DPPT indicates a role for ROS production in the process and is consistent with the increased ROS production being primarily mitochondrial. Telomeres act as 'mitotic clocks' in human fibroblasts because they shorten with each round of replication due to both the inability of DNA polymerases to replicate the very ends of chromosomes (Olovnikow, 1973) and the specific accumulation of stress-induced DNA damage (von Zglinicki, 2002). Eventually, telomere dysfunction triggers replicative senescence (Bodnar et al., 1998). Although intense stress can cause a senescence-like arrest without involvement of telomeres (Chen et al., 2001; Gorbunova et al., 2002), one possibility is that ROS production accelerates replicative senescence via its contribution to telomere shortening under conditions of mild stress. Therefore, we wanted to find out whether mitoQ could prolong the replicative lifespan of human fibroblasts under mild stress conditions, and whether this would correlate with a reduction in the rate of telomere shortening. MitoQ in micromolar concentrations selectively blocks mitochondrial oxidative damage and prevents apoptosis induced by acute treatments with hydrogen peroxide (Kelso et al., 2001). When mitoQ was incubated with MRC-5 fibroblasts, concentrations above 50–100 nm were cytostatic in long-term culture, and even for concentrations of 10–20 nm an adaptation period of at least one week under normoxic conditions was necessary before beneficial effects on growth could be seen (data not shown). Such an adaptation period seems to be a characteristic effect of powerful antioxidants and may reflect the involvement of ROS in a multitude of cellular signal transduction chains (Forman et al., 2002). Neither DPPT nor mitoQ had a significant effect on the intracellular peroxide content under normoxic culture conditions as measured by 2′,7′-dichlorofluorescein fluorescence. However, mitoQ, but not DPPT, abolished nearly half of the rise in peroxides induced by hyperoxic culture in untreated cultures (Fig. 1a). Chronically increased oxidative stress exerted by culture under mild hyperoxia (40% oxygen partial pressure) shortens the replicative lifespan of MRC-5 fibroblasts down to few population doublings (von Zglinicki et al., 1995; von Zglinicki, 2002). This premature aging phenotype is indistinguishable from replicative senescence under standard culture conditions (von Zglinicki et al., 1995; Saretzki et al., 1998; Toussaint et al., 2000). Treatment of MRC-5 cells under these conditions with mitoQ significantly elongated the replicative lifespan by an average of 40% (ranging from 15 to 70%) in four independent experiments whereas the lifespan of DPPT-treated control cells remained unchanged (Fig. 1b). This is in agreement with effects of other potent antioxidants on the replicative lifespan of human cells, e.g. the spin trap α-phenyl-N-t-butyl nitrone (Chen et al., 1995; von Zglinicki et al., 2000), its derivative N-t-butyl hydroxylamine (Atamna et al., 2000) or an oxidation-resistant vitamin C derivative (Asp-2-O-phosphate (Furumoto et al., 1998). MitoQ decreases the cellular peroxide content and prolongs replicative lifespan of MRC-5 cells under hyperoxia. Cells (PD between 20 and 28) were left untreated (no) or were pretreated for one week with 10–20 nm (data pooled) of either mitoQ or DPPT and then subjected to 40% hyperoxia until cessation of proliferation (4–6 weeks). MitoQ or DPPT, respectively, were prepared as stock solutions in 100% alcohol and added to the medium every other day. (a) Cell peroxide content under normoxia (filled bars) and after one week of hyperoxia (open bars) was measured by 2′,7′-dichlorofluorescein using flow cytometry as described (Lorenz et al., 2001). Data are mean ± SEM from 3 to 5 independent experiments. (B) Replicative lifespan under hyperoxia (in population doublings, PD) was measured in nine (no treatment), three (DPPT) or four (MitoQ) independent experiments. Box plots indicate the median, upper and lower quartiles and percentiles. Asterisks indicate a significant difference (P < 0.05) between mitoQ and either DPPT or no treatment. To examine the involvement of telomeres, we measured telomere length at the start point of the treatment and at up to four different time points under hyperoxic culture by non-radioactive Southern blotting as described (Petersen et al., 1998; Serra et al., 2002). MitoQ treatment minimized telomere shortening under hyperoxia (Fig. 2a). Quantitative evaluation of data from four independent experiments (Fig. 2b) indicated that hyperoxia increased the rate of telomere shortening per population doubling (PD), similar to that seen in DPPT-treated and untreated control cells. This is in close agreement with earlier data (von Zglinicki, 2002). However, mitoQ treatment completely prevented this rise in telomere shortening rate due to hyperoxia and instead gave a negligible rate of telomere shortening. Again, this is in agreement with the reported protective effect of PBN (von Zglinicki et al., 2000), Asp-2-O-phosphate (Furumoto et al., 1998) or superoxide dismutase overexpression (Serra et al., 2002) on telomere maintenance. MitoQ counteracts telomere shortening under hyperoxia. Cells were grown under normoxia (NO) and, after one week pretreatment with either DPPT or mitoQ, under 40% hyperoxia (HO) and DNA was sampled at the times and PD as indicated. (A) Telomere Southern blot including λHindIII marker (M). Blots were scanned with a FUJI LAS-100 luminescence imager. Average telomere length was calculated using the AIDA software and is indicated by white lines. (B) Telomere shortening rates per PD (mean ± SEM) under normoxia (filled bar) and hyperoxia (open bars). Cells were untreated (no) or treated with either DPPT or mitoQ. Data were calculated by linear regression of triplicate measurements from three or four independent experiments. Telomere shortening rate in mitoQ-treated cells under hyperoxia is not significantly different from zero, but is significantly (P < 0.05, anova) smaller than in both untreated or DPPT-treated cells. Together, these data indicate that minimizing oxidative stress significantly slows down telomere shortening and prolongs replicative lifespan. Moreover, they suggest that it is accelerated telomere shortening in response to increased mitochondrial ROS production that induces premature senescence-like arrest under conditions of mild stress such as chronic hyperoxia. Intense, acute stress, which leads to an immediate arrest in the vast majority of cells, is of course telomere length-independent (Chen et al., 2001; Gorbunova et al., 2002). Such stress will damage DNA all over the genome and thus trigger arrest before replication and concomitant telomere shortening occurs. On the other hand, low levels of interstitial DNA damage are compatible with cell proliferation, but damage will contribute to telomere shortening because of the low telomere-specific efficiency of single-strand break repair (Petersen et al., 1998). Thus, telomeres appear to act as cellular sentinels for oxidative damage under near-physiological conditions by limiting cell proliferation if and when stress and thus greater mutational risks accumulate (von Zglinicki, 2002). This work was supported by MRC grant G0100140 57030 to T.v.Z.

Excessive ROS production and enhanced autophagy contribute to myocardial injury induced by branched-chain amino acids: Roles for the AMPK-ULK1 signaling pathway and α7nAChR

Ulcerative colitis (UC) is a multifactorial inflammatory disease, and increasing evidence has demonstrated that the mechanism of UC pathogenesis is associated with excessive cellular apoptosis and reactive oxygen species (ROS) production. However, their function and molecular mechanisms related to UC remain unknown. In this study, Rab27A mRNA and protein were proven to be overexpressed in intestinal epithelial cells of UC patients and DSS‐induced colitis mice, compared with control (P < 0.05). And Rab27A silencing inhibits inflammatory process in DSS‐induced colitis mice (P < 0.05). Then, it was shown that knockdown of Rab27A suppressed apoptosis and ROS production through modulation of miR‐124‐3p, whereas overexpression of Rab27A promoted apoptosis and ROS production in LPS‑induced colonic cells. In addition, enhanced expression of miR‐124‐3p attenuated apoptosis and ROS production by targeting regulation of STAT3 in LPS‑induced colonic cells. Mechanistically, we found Rab27A reduced the expression and activity of miR‐124‐3p to activate STAT3/RelA signalling pathway and promote apoptosis and ROS production in LPS‑induced colonic cells, whereas overexpression of miR‐124‐3p abrogated these effects of Rab27A. More importantly, animal experiments illustrated that ectopic expression of Rab27A promoted the inflammatory process, whereas overexpression of miR‐124‐3p might interfere with the inflammatory effect in DSS‐induced colitis mice. In summary, Rab27A might modulate the miR‐124‐3p/STAT3/RelA axis to promote apoptosis and ROS production in inflammatory colonic cells, suggesting that Rab27A as a novel therapeutic target for the prevention and treatment of UC patients.

Circulating levels of soluble lectin-like oxidized low-density lipoprotein receptor-1 (sLOX-1) play an important role in the development and progression of atherosclerosis. We hypothesized that the inflammatory marker C-reactive protein (CRP) might stimulate sLOX-1 release by activating tumor necrosis factor-α converting enzyme (TACE). Macrophages differentiated from THP-1 cells were stimulated with TNF-α and further treated with CRP in the absence or presence of specific inhibitors or small interfering RNA (siRNA). Our results showed that CRP increased sLOX-1 release from activated macrophages in a dose-dependent manner and that these effects were regulated by Fc γ receptor II (FcγRII)-mediated p47(phox) phosphorylation, reactive oxygen species (ROS) production, and TACE activation. CRP also enhanced sLOX-1 release from macrophages derived from peripheral blood mononuclear cells (PBMC) of patients with acute coronary syndrome (ACS). Pretreatment with antibody against FcγRII or with CD32 siRNA, p47(phox) siRNA, apocynin, N-acetylcysteine, tumor necrosis factor-α protease inhibitor 1 (TAPI-1) or TACE siRNA attenuated sLOX-1 release induced by CRP. CRP also elevated serum sLOX-1 levels in a rabbit model of atherosclerosis. Thus, CRP might stimulate sLOX-1 release, and the underlying mechanisms possibly involved FcγRII-mediated p47(phox) phosphorylation, ROS production, and TACE activation.

Hypertension is associated with excessive reactive oxygen species (ROS) production in vascular cells. Mitochondria undergo fusion and fission, a process playing a role in mitochondrial function. OPA1 is essential for mitochondrial fusion. Loss of OPA1 is associated with ROS production and cell dysfunction. We hypothesized that mitochondria fusion could reduce oxidative stress that defect in fusion would exacerbate hypertension. Using (a) Opa1 haploinsufficiency in isolated resistance arteries from Opa1+/- mice, (b) primary vascular cells from Opa1+/- mice, and (c) RNA interference experiments with siRNA against Opa1 in vascular cells, we investigated the role of mitochondria fusion in hypertension. In hypertension, Opa1 haploinsufficiency induced altered mitochondrial cristae structure both in vascular smooth muscle and endothelial cells but did not modify protein level of long and short forms of OPA1. In addition, we demonstrated an increase of mitochondrial ROS production, associated with a decrease of superoxide dismutase 1 protein expression. We also observed an increase of apoptosis in vascular cells and a decreased VSMCs proliferation. Blood pressure, vascular contractility, as well as endothelium-dependent and -independent relaxation were similar in Opa1+/- , WT, L-NAME-treated Opa1+/- and WT mice. Nevertheless, chronic NO-synthase inhibition with L-NAME induced a greater hypertension in Opa1+/- than in WT mice without compensatory arterial wall hypertrophy. This was associated with a stronger reduction in endothelium-dependent relaxation due to excessive ROS production. Our results highlight the protective role of mitochondria fusion in the vasculature during hypertension by limiting mitochondria ROS production.