Sustained Ischemia Research Articles

Synthesis and Pharmacological Evaluation of Novel Adenine–Hydrogen Sulfide Slow Release Hybrids Designed as Multitarget Cardioprotective Agents

This work deals with the design, synthesis, and evaluation of the cardioprotective properties of a number of novel hybrid compounds combining the adenine nucleus with a suitable H2S slow-releasing moiety, coupled via a stable ether bond. The H2S release rate of the hybrids and their ability to increase cGMP were estimated in vitro. The most promising derivatives 4 and 11, both containing 4-hydroxythiobenzamide moiety as H2S donor, were selected for further in vivo evaluation. Their ability to release H2S in vivo was recorded using a new fully validated UPLC-DAD method. Both compounds reduced significantly the infarct size when administered at the end of sustained ischemia. Mechanistic studies showed that they conferred enhanced cardioprotection compared to adenine or 4-hydroxythiobenzamide. They activate the PKG/PLN pathway in the ischemic myocardium, suggesting that the combination of both pharmacophores results in synergistic cardioprotective activity through the combination of both molecular pathways that trigger cardioprotection.

Effects of wortmannin on cardioprotection exerted by ischemic preconditioning in rat hearts subjected to ischemia-reperfusion.

Ischemic preconditioning (IPC) is one of the most powerful interventions to reduce ischemia-reperfusion injury. The aim of the present study was to investigate the involvement of the phosphatidylinositol-3-kinases (PI3Ks) family in cardioprotection exerted by IPC and the relationship between preservation of mitochondrial morphology and ATP synthesis capacity. In this regard, macroautophagy (autophagy) is considered a dynamic process involved in the replacement of aged or defective organelles under physiological conditions. IPC consisted of four 5-min cycles of ischemia-reperfusion followed by sustained ischemia. Wortmannin (W), a PI3K family inhibitor, was added to the perfusion medium to study the involvement of autophagy in the beneficial effects of IPC. In the present study, LC3-II/I expression was significantly increased in the IPC group when compared with the control group. The hearts subjected to IPC showed greater degradation of p62 than control groups, establishing the existence of an autophagic flow. Electron microscopy showed that IPC preserves the structural integrity of mitochondria after ischemia and at the end of reperfusion. Moreover, hearts subjected to IPC exhibited increased mitochondrial ATP synthesis. The beneficial effects of IPC were abolished by W in all trials of this study, abolishing the differences between the IPC and control groups. These results suggest that IPC could partly reduce injury by ischemia-reperfusion (I/R) by decreasing mitochondrial damage and promoting autophagy. Since W is a nonspecific inhibitor of the PI3Ks family, further research is required to confirm participation of PI3K in the response to IPC.

Beyond Preconditioning: Postconditioning as an Alternative Technique in the Prevention of Liver Ischemia‐Reperfusion Injury

Liver ischemia/reperfusion injury may significantly compromise hepatic postoperative function. Various hepatoprotective methods have been improvised, aiming at attenuating IR injury. With ischemic preconditioning (IPC), the liver is conditioned with a brief ischemic period followed by reperfusion, prior to sustained ischemia. Ischemic postconditioning (IPostC), consisting of intermittent sequential interruptions of blood flow in the early phase of reperfusion, seems to be a more feasible alternative than IPC, since the onset of reperfusion is more predictable. Regarding the potential mechanisms involved, it has been postulated that the slow intermittent oxygenation through controlled reperfusion decreases the burst production of oxygen free radicals, increases antioxidant activity, suppresses neutrophil accumulation, and modulates the apoptotic cascade. Additionally, favorable effects on mitochondrial ultrastructure and function, and upregulation of the cytoprotective properties of nitric oxide, leading to preservation of sinusoidal structure and maintenance of blood flow through the hepatic circulation could also underlie the protection afforded by postconditioning. Clinical studies are required to show whether biochemical and histological improvements afforded by the reperfusion/reocclusion cycles of postconditioning during early reperfusion can be translated to a substantial clinical benefit in liver resection and transplantation settings or to highlight more aspects of its molecular mechanisms.

Ischemic preconditioning modifies mortality and inflammatory response.

To evaluate the effect of ischemic preconditioning on mortality, inflammatory mediators and oxidative stress after intestinal ischemia and reperfusion. Male Wistar rats were allocated according to the period of ischemia with or without ischemic preconditioning which consist on clamping the superior mesenteric artery for 10 minutes followed by reperfusion for 10 minutes before the sustained ischemia period. Mortality was assessed in Phase 1 study, and the CINC-1, CINC-2 and MDA levels in the lungs were analyzed in Phase 2. Mortality was lower in the ischemic preconditioning group subjected to 90 minutes of ischemia compared to the group without ischemic preconditioning (I-90: 50% and IPC-90: 15%, p=0.018), and it was lower in the ischemic preconditioning group as a whole compared to the groups without ischemic preconditioning (IPC-14% and I=30%, p=0.006). Lower levels of MDA, CINC-1, and CINC-2 were observed in the animals that were subjected to ischemic preconditioning compared to the animals that were not (MDA: I-45=1.23 nmol/mg protein, and IPC-45=0.62 nmol/mg protein, p=0.0333; CINC-1: I-45=0.82 ng/mL and IPC-45=0.67 ng/mL, p=0.041; CINC-2: I-45=0.52 ng/mL and IPC-45=0.35 ng/mL, p=0.032). Ischemic preconditioning reduces mortality, inflammatory process and oxidative stress in rats subjected to intestinal ischemia and reperfusion.

Mechanical cytoprotection: A review of cytoskeleton-protection approaches for cells

We review a class of cutting-edge approaches for cytoprotection of cells exposed to assaults such as sustained deformations, chemotherapy, radiotherapy, or ischemia. These approaches will enhance cell survival by mechanically protecting the structure and dynamics of the actin cytoskeleton (CSK). Cortical actin provides structural support to the plasma membrane (PM), protecting its integrity. Consequently, assaults can fragment the actin cortex leading to local, mechanical failure of the PM and poration of the cell. This disrupts normal trafficking of biomolecules across the PM, leading to loss of homeostasis and eventually, to cell death and tissue necrosis. Two different approaches to cytoskeletal protection are covered in this review paper. The first is to supply energy-related molecules to maintain and enhance the energy-consuming dynamics of the actin CSK. The second is to stabilize newly formed actin CSK directly¸ for example through cross-linking or reinforcement at PM anchoring sites. Research in this area is clearly still in its infancy. Very few studies have gone beyond characterizing the effects of induced damage to the actin CSK (and subsequent PM collapse). Recent work, focusing instead on sustaining the actin under non-physiological or pathophysiological conditions, has shown great promise. Such cytoskeletal-protection may find medical applications in preventing or minimizing tissue damage when tissues are unhealthy or at risk, or in enhancing cell performance under stress. Here, we condense the relevant cell biology and biomechanics background, assess candidate cytoskeletal protective agents, and review published works that have shown potential for medical benefit in experimental model systems.

Value of serum tenascin-C in patients with acute myocardial infarction

BackgroundMyocardial infarction (MI) is defined as myocardial cell necrosis due to significant and sustained ischemia. TN-C is an extracellular matrix glycoprotein that is expressed in several important steps during the very early stage of cardiogenesis. TN-C is not normally expressed in the adult heart, but transiently appears during pathological conditions and plays important roles in tissue remodeling. AimTo study the role of TN-C in myocardial infarction patients and to evaluate its role as a predictor of HF in these patients. MethodsThis study was conducted on 45 cases uniformly divided into 3 closely matched (in age and sex) groups as follows: Group (I) includes 15 patients who were suffering from AMI; Group (II) includes 15 patients who were suffering from HF on top of MI; and Group (III) includes 15 healthy volunteers coming for regular annual checkup. 3–6ml venous blood was collected on the day of admission under complete aseptic conditions and stored at −70°C until assayed by ELISA. ResultsTN-C levels in the sera of patients with AMI Group (I) were significantly higher than those of healthy volunteers. Moreover, in Group I of AMI, a positive correlation between TN-C level on one side and CK, CK-MB and troponin T level on the other side was found. TN-C levels in the sera of patients with congestive heart failure on top of acute MI Group (II) were significantly higher than those of healthy volunteers. Pro-BNP levels in patients with heart failure Group (II) were significantly higher than those with AMI not complicated with heart failure Group (I). Levels of pro-BNP were also positively correlated with those of TN-C in patients with heart failure on top of AMI Group (II). ConclusionsSerum TN-C might be a novel marker reflecting active structural remodeling in the myocardium following infarction, with high TN-C levels at acute stages possibly predicting progression of LV remodeling. Also, the incorporation of a combination of serum TN-C and plasma BNP levels may improve risk stratification for congestive heart failure after AMI. Further studies on large scale are needed for more evaluation of TN-C role in HF.

Role of Endogenous Opioid System in Ischemic-Induced Late Preconditioning.

BackgroundOpioid receptors (OR) are involved in myocardial late preconditioning (LPC) induced by morphine and δ1-opioid receptor (δ1-OR) agonists. The role of OR in ischemic-induced LPC is unknown. We investigated whether 1) OR are involved in the trigger and/or mediation phase of LPC and 2) a time course effect on the expression of different opioid receptors and their endogenous ligands exists.MethodsMale Wistar rats were randomly allocated to four groups (each group n = 8). Awake animals were ischemic preconditioned by a 5 minutes coronary occlusion. 24 hours later, anesthetized animals underwent 25 minutes coronary occlusion followed by 2 hours of reperfusion. The role of OR was investigated by treatment with intraperitoneal naloxone (Nal) 10 minutes prior to LPC (Nal-LPC; trigger phase) or 10 min prior to sustained ischemia (LPC-Nal; mediation phase).ResultsLPC reduced infarct size from 61±10% in controls to 25±9% (P<0.001). Naloxone during trigger or mediation phase completely abolished LPC-induced cardioprotection (59±9% and 62±9%; P<0.001 vs. LPC). 8, 12 and 24 hours after the ischemic stimulus, expression of δ-OR in the heart was increased, whereas μ-opioid receptor (μ-OR) and κ-opioid receptor (κ-OR) were not. Plasma concentrations of β-endorphin and leu-enkephalin but not dynorphin were increased by LPC.ConclusionIschemic LPC is triggererd and mediated by OR. Expression of δ-OR and plasma levels of endogenous opioid peptides are increased after ischemic LPC.

The calcium-ROS-pH triangle and mitochondrial permeability transition: challenges to mimic cardiac ischemia-reperfusion.

The calcium-ROS-pH triangle and mitochondrial permeability transition: challenges to mimic cardiac ischemia-reperfusion.

Molecular basis of cardioprotection: signal transduction in ischemic pre-, post-, and remote conditioning.

Reperfusion is mandatory to salvage ischemic myocardium from infarction, but reperfusion per se contributes to injury and ultimate infarct size. Therefore, cardioprotection beyond that by timely reperfusion is needed to reduce infarct size and improve the prognosis of patients with acute myocardial infarction. The conditioning phenomena provide such cardioprotection, insofar as brief episodes of coronary occlusion/reperfusion preceding (ischemic preconditioning) or following (ischemic postconditioning) sustained myocardial ischemia with reperfusion reduce infarct size. Even ischemia/reperfusion in organs remote from the heart provides cardioprotection (remote ischemic conditioning). The present review characterizes the signal transduction underlying the conditioning phenomena, including their physical and chemical triggers, intracellular signal transduction, and effector mechanisms, notably in the mitochondria. Cardioprotective signal transduction appears as a highly concerted spatiotemporal program. Although the translation of ischemic postconditioning and remote ischemic conditioning protocols to patients with acute myocardial infarction has been fairly successful, the pharmacological recruitment of cardioprotective signaling has been largely disappointing to date.

Remifentanil and opioid-induced cardioprotection.

CARDIAC SURGERY frequently involves ischemia and reperfusion of the myocardium, which can lead to perioperative damage, arrhythmia, and dysfunction. In noncardiac surgery, cardiac complications are the major cause of morbidity and mortality. This is a consequence of the high underlying prevalence of cardiovascular disease and the metabolic responses to surgical stress and pain, which alter physiology and may trigger a cascade of prothrombotic biochemical events. Myocardial oxygen consumption also is increased during the perioperative period, which affects the oxygen supply/demand balance, and intraluminal shear forces in the coronary artery may be increased, producing a “perfect storm” for atheromatous plaque rupture and subsequent coronary artery thrombosis. Intense research has been directed toward preventing or ameliorating myocardial ischemiareperfusion injury, stemming from the groundbreaking work of Murry et al. They “conditioned” the heart to tolerate the effects of acute ischemia-reperfusion injury by inducing brief nonlethal episodes of ischemia and reperfusion to the heart before sustained lethal myocardial ischemia. The subsequent dramatic reduction in myocardial injury is now referred to as ischemic preconditioning, and it has become apparent that similar levels of cardioprotection can be achieved by applying the stimulus during or after the lethal ischemia (preconditioning or postconditioning, respectively).

Role of the MPTP in conditioning the heart - translatability and mechanism.

Mitochondria have long been known to be the gatekeepers of cell fate. This is particularly so in the response to acute ischaemia‐reperfusion injury (IRI). Following an acute episode of sustained myocardial ischaemia, the opening of the mitochondrial permeability transition pore (MPTP) in the first few minutes of reperfusion, mediates cell death. Preventing MPTP opening at the onset of reperfusion using either pharmacological inhibitors [such as cyclosporin A (CsA) ] or genetic ablation has been reported to reduce myocardial infarct (MI) size in animal models of acute IRI. Interestingly, the endogenous cardioprotective intervention of ischaemic conditioning, in which the heart is protected against MI by applying cycles of brief ischaemia and reperfusion to either the heart itself or a remote organ or tissue, appears to be mediated through the inhibition of MPTP opening at reperfusion. Small proof‐of‐concept clinical studies have demonstrated the translatability of this therapeutic approach to target MPTP opening using CsA in clinical settings of acute myocardial IRI. However, given that CsA is a not a specific MPTP inhibitor, more novel and specific inhibitors of the MPTP need to be discovered – the molecular identification of the MPTP should facilitate this. In this paper, we review the role of the MPTP as a target for cardioprotection, the potential mechanisms underlying MPTP inhibition in the setting of ischaemic conditioning, and the translatability of MPTP inhibition as a therapeutic approach in the clinical setting.Linked ArticlesThis article is part of a themed section on Conditioning the Heart – Pathways to Translation. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue‐8

Remote ischemic preconditioning for myocardial protection: update on mechanisms and clinical relevance.

Ischemic heart disease is the leading cause of death for both men and women worldwide, accruing 7.4 million deaths in 2012. There has been a continued search for better cardioprotective modalities that would reduce myocardial ischemia-reperfusion injury. Among these attempts, a more convenient model of ischemic preconditioning, known as remote ischemic preconditioning (RIPC) was first introduced in 1993 by Przyklenk and colleagues who reported that brief regional occlusion-reperfusion episodes in one vascular bed of the heart render protection to remote myocardial tissue. Subsequently, major advances in myocardial RIPC came with the use of skeletal muscle as the ischemic stimulus. To date, numerous studies have revealed that RIPC applied to the kidney, liver, mesentery, and skeletal muscle, have all exhibited cardioprotective effects. The main purpose of this review article is to summarize the new advances in understanding the molecular mechanisms of RIPC during the past 5years, including those related to capsaicin-activated C sensory fibers, hypoxia-inducible factor 1α, connexin 43, extracellular vesicles, microRNA-144, microRNA-1, and nitrite. In addition, we have discussed results from several recent human clinical trials with RIPC. Taken together, the emerging clinical evidence supports the concept that the effectiveness of RIPC paired with its low-cost and non-invasive features makes it an ideal treatment before reperfusion after sustained ischemia. More carefully designed studies are warranted to fully exploit the clinical benefits of RIPC and its potential implications in patients with cardiovascular disease.

Extracellular signalling molecules in the ischaemic/reperfused heart - druggable and translatable for cardioprotection?

In patients with acute myocardial infarction, timely reperfusion is essential to limit infarct size. However, reperfusion also adds to myocardial injury. Brief episodes of ischaemia/reperfusion in the myocardium or on organ remote from the heart, before or shortly after sustained myocardial ischaemia effectively reduce infarct size, provided there is eventual reperfusion. Such conditioning phenomena have been established in many experimental studies and also translated to humans. The underlying signal transduction, that is the molecular identity of triggers, mediators and effectors, is not clear yet in detail, but several extracellular signalling molecules, such as adenosine, bradykinin and opioids, have been identified to contribute to cardioprotection by conditioning manoeuvres. Several trials have attempted the translation of cardioprotection by such autacoids into a clinical scenario of myocardial ischaemia and reperfusion. Adenosine and its selective agonists reduced infarct size in a few studies, but this benefit was not translated into improved clinical outcome. All studies with bradykinin or drugs which increase bradykinin's bioavailability reported reduced infarct size and some of them also improved clinical outcome. Synthetic opioid agonists did not result in a robust infarct size reduction, but this failure of translation may relate to the cardioprotective properties of the underlying anaesthesia per se or of the comparator drugs. The translation of findings in healthy, young animals with acute coronary occlusion/reperfusion to patients of older age, with a variety of co-morbidities and co-medications, suffering from different scenarios of myocardial ischaemia/reperfusion remains a challenge.

Diagnosis and Decision Making for Patients With Disorders of Consciousness: A Survey Among Family Members

ObjectivesTo examine the perceptions of family members of patients with disorders of consciousness (DOC) in regard to the patients' level of consciousness, communicative status, and prognosis as compared with the objective medical categories, and to elicit the family members' self-reported practice of treatment decision-making. DesignCross-sectional semiquantitative survey. SettingFive specialized neurologic rehabilitation facilities. ParticipantsConsecutive sample of primary family members (N=44) of patients with DOC as determined by the Coma Recovery Scale-Revised, surveyed 6 months after the patient's brain injury. InterventionNot applicable. Main Outcome MeasuresPerception of level of consciousness as compared with the medical diagnosis; assessment of communicative status and prognosis; and practice of treatment decision-making. ResultsThe study included 44 family members of patients, most of whom had sustained global cerebral ischemia. Six months after brain injury, 36% were in a vegetative state (VS), 20% were in a minimally conscious state (MCS), and 39% had emerged from an MCS. In 76% of cases, the relatives assumed the same level of consciousness that diagnostic tests showed. In the other cases, consciousness was mostly underestimated. While relatives of patients in a VS, and to a lesser extent of those in an MCS, were more skeptical about the patients' chances to advance to an independent life, all had high hopes that the patients would regain the ability to communicate. Yet, 59% of family members had thought about limiting life-sustaining treatment. Most of them base treatment decisions on the patient's well-being; very few relied on previously expressed patient wishes. ConclusionsAccording to our sample, family members of patients with DOC largely assess the level of consciousness correctly and express high hopes to reestablish communication with the patient.

Pharmacological postconditioning with tanshinone IIA attenuates myocardial ischemia-reperfusion injury in rats by activating the phosphatidylinositol 3-kinase pathway.

Tanshinone IIA, one of the active ingredients in the Chinese medicine Danshen, is cardioprotective when applied prior to sustained myocardial ischemia. The present study aimed to investigate whether pharmacological postconditioning with tanshinone IIA attenuates myocardial ischemia-reperfusion injury when applied prior to prolonged reperfusion following a sustained ischemia. A total of 88 Sprague-Dawley rats received 30 min myocardial ischemia followed by 5 or 120 min reperfusion. Compared with the ischemia-reperfusion model group, the group that received an intravenous injection of 10 mg/kg tanshinone IIA prior to reperfusion had a reduced myocardial infarct size, higher levels of phospho-Akt and phospho-endothelial nitric oxide synthase and less reduction in the optical density of the mitochondria at 540 nm, indicating that the mitochondrial permeability transition (MPT) was attenuated. The cardioprotective effect conferred by tanshinone IIA was abolished by LY294002, a specific inhibitor of phosphatidylinositol 3-kinase (PI3K). These results demonstrate that tanshinone IIA postconditioning protects the myocardium from ischemia-reperfusion injury through the PI3K/Akt pathway, and the MPT may be also involved in this process.

Reply to “Letter to the editor: ‘Cyclosporin A in left ventricular remodeling after myocardial infarction’”

Reply to “Letter to the editor: ‘Cyclosporin A in left ventricular remodeling after myocardial infarction’”

Cardioprotection by remote ischemic preconditioning of the rat heart is mediated by extracellular vesicles

Remote ischemic preconditioning (RIPC) of the heart is exerted by brief ischemic insults affected on a remote organ or a remote area of the heart before a sustained cardiac ischemia. To date, little is known about the inter-organ transfer mechanisms of cardioprotection by RIPC. Exosomes and microvesicles/microparticles are vesicles of 30–100nm and 100–1000nm in diameter, respectively (collectively termed extracellular vesicles [EVs]). Their content of proteins, mRNAs and microRNAs, renders EV ideal conveyors of inter-organ communication. However, whether EVs are involved in RIPC, is unknown. Therefore, here we investigated whether (1) IPC induces release of EVs from the heart, and (2) EVs are necessary for cardioprotection by RIPC. Hearts of male Wistar rats were isolated and perfused in Langendorff mode. A group of donor hearts was exposed to 3×5-5min global ischemia and reperfusion (IPC) or 30min aerobic perfusion, while coronary perfusates were collected. Coronary perfusates of these hearts were given to another set of recipient isolated hearts. A group of recipient hearts received IPC effluent depleted of EVs by differential ultracentrifugation. Infarct size was determined after 30min global ischemia and 120min reperfusion. The presence or absence of EVs in perfusates was confirmed by dynamic light scattering, the EV marker HSP60 Western blot, and electron microscopy. IPC markedly increased EV release from the heart as assessed by HSP60. Administration of coronary perfusate from IPC donor hearts attenuated infarct size in non-preconditioned recipient hearts (12.9±1.6% vs. 25.0±2.7%), similarly to cardioprotection afforded by IPC (7.3±2.7% vs. 22.1±2.9%) on the donor hearts. Perfusates of IPC hearts depleted of EVs failed to exert cardioprotection in recipient hearts (22.0±2.3%). This is the first demonstration that EVs released from the heart after IPC are necessary for cardioprotection by RIPC, evidencing the importance of vesicular transfer mechanisms in remote cardioprotection.

The effect of ischemic preconditioning on nitric oxide synthase activity during myocardial ischemia and reperfusion in anesthetized dogs

obJeCtives: To examine whether changes in nitric oxide (NO) bioavailability occurring during a 25 min occlusion of the left anterior descending coronary artery and following ischemic preconditioning (PC) in anesthetized dogs (chloralose and urethane, intravenous) are associated with changes in the activation of NO synthase (NOS) enzymes. Methods: In 12 control and 16 preconditioned dogs, total NOS activity (measured using radioimmunoassay), superoxide (measured using confocal microscopy) and nitrotyrosine (NT; measured using Western blot) production were assessed in tissue samples obtained from the left ventricular wall both after the PC (two 5 min occlusions) procedure and at two time points (5 min and 25 min) during prolonged ischemia. Plasma nitrate/nitrite concentrations in the blood of the coronary sinus were also determined. ResuLts: Compared with the sham control group that was not subjected to ischemia, occlusion of the left anterior descending coronary artery resulted in an initial increase, followed by a significant decrease in NOS activity and nitrate/nitrite levels. There were parallel increases in superoxide and NT production by the end of the ischemic period. PC itself enhanced NOS activity and the concentration of NO metabolites, which were maintained during the prolonged occlusion. Furthermore, PC prevented prolonged ischemia-induced superoxide and NT formation. ConCLusions: The present study demonstrated that changes in NO bioavailability during ischemia are closely associated with changes in NOS activation. PC stimulates and maintains the activation of NOS, thereby providing better NO bioavailability and suppressed free radical formation during sustained ischemia. All of these effects may have importance in the previously observed marked antiarrhythmic effect of PC.

Reference Gene Identification for Reverse Transcription-Quantitative Polymerase Chain Reaction Analysis in an Ischemic Wound-Healing Model

Reference genes are often used in RT-quantitative PCR (qPCR) analysis to normalize gene expression levels to a gene that is expressed stably across study groups. They ultimately serve as a control in RT-qPCR analysis, producing more accurate interpretation of results. Whereas many reference genes have been used in various wound-healing studies, the most stable reference gene for ischemic wound-healing analysis has yet to be identified. The goal of this study was to determine systematically the most stable reference gene for studying gene expression in a rat ischemic wound-healing model using RT-qPCR. Twelve commonly used reference genes were analyzed using RT-qPCR and geNorm data analysis to determine stability across normal and ischemic skin tissue. It was ultimately determined that Ubiquitin C (UBC) and β-2 Microglobulin (B2M) are the most stably conserved reference genes across normal and ischemic skin tissue. UBC and B2M represent reliable reference genes for RT-qPCR studies in the rat ischemic wound model and are unaffected by sustained tissue ischemia. The geometric mean of these two stable genes provides an accurate normalization factor. These results provide insight on dependence of reference-gene stability on experimental parameters and the importance of such reference-gene investigations.

Protection tissulaire: une nouvelle piste

Visceral infarctions remain the leading cause of mortality in the world, in spite of the development of early reperfusion strategies. Infarct size is determined not only by the severity of ischemia but also by pathological processes initiated at reperfusion. Experimental evidences suggest that brief ischemia-reperfusion periods, called ischemic preconditioning or postconditioning, and performed either before or just after a sustained ischemia, respectively, limit reperfusion injuries. Ischemic conditioning activates several primary pathways that inhibit mitochondrial permeability transition pore opening. Thus, pharmacological interventions like cyclosporine A, a potent permeability transition pore inhibitor, are also able to reduce cell death, when administered at reperfusion (pharmacological postconditioning), in numerous ischemiareperfusion models including in humans. This review will focus on the physiological preclinical data on both ischemic and pharmacological conditioning that are relevant to their translation to clinical therapeutics in intensive care medicine.