Diabetes increases apoptosis and necrosis in the ischemic and non-ischemic human myocardium: role of caspases and poly (ADP-ribose) polymerase

Background: Diabetes is an important predictor of morbidity and mortality after cardiac surgery but the reason is unclear. Therefore the aims of these studies were to elucidate whether cell death is greater in the ischemic and non- ischemic diabetic human myocardium than in the non-diabetic and to investigate the underlying mechanism. Methods: The right atrial appendages (n=8/group) of non-diabetics, non-insulin dependent diabetics (NIDDM) and insulin dependent diabetics (IDDM) were subjected to 90 minutes simulated ischemia/120 minutes reoxygenation (SI/R). Tissue injury was measured by the release of CK into the media, and cellular apoptosis and necrosis were assessed in tissue by the TUNEL assay and propidium iodide. Initiator and effector caspases activation was also measured. Results: Apoptosis and necrosis were greater in the NIDDM and the IDDM groups than in the non-diabetic group both in fresh tissue and after SI/R. Activation of effector caspases was also higher in the NIDDM and IDDM groups than in the non-diabetic group after SI/R. Caspase-3 inhibition reduced apoptosis in all study groups without influencing necrosis; however, PARP inhibition resulted in a similar reduction in apoptosis and in necrosis in all groups and caspase-2 inhibition did not. Conclusions: Diabetes increases both apoptosis and necrosis in the human myocardium in fresh and after being subjected to ischemia/reoxygenation, an effect that is mediated, at least in part, by caspase-3 and PARP activation. These results may explain the increased cardiac-related morbidity and mortality during cardiac surgery in patients with diabetes

Characterization of an in vitro model for the study of the short and prolonged effects of myocardial ischaemia and reperfusion in man

The mechanisms underlying myocardial ischaemia and reperfusion-induced injury have been investigated, mainly by using animal experimental preparations in vitro and in vivo, but little is known of the process in human myocardium. The present studies characterize an in vitro model using human myocardium for the study of early and delayed effects of ischaemia and reperfusion. The right atrial appendage was manually sliced and incubated in buffer through which was bubbled O2/CO2 (19:1, v/v) for various time periods. Lactate dehydrogenase (LDH) leakage, 3- [4,5-dimethylthiazol-2-yl]-2,5 diphenyl-2H-tetrazolium bromide (MTT) reduction, oxygen consumption, nucleotide levels and tissue morphology were all investigated as markers of myocardial injury. The specimens remained stable and viable up to 24 h, but had significantly deteriorated by 48 h. The preparation responded to ischaemia in a time-related manner. Tissue viability was reduced by 25% after 30 min ischaemia, declined to 60% after 60 min ischaemia and to 75% after 120 min ischaemia. Interestingly, the tissue was more susceptible when ischaemia was induced after 24 h of aerobic incubation. The effects of the duration of reperfusion were investigated after a fixed 60 min ischaemic insult. The results of LDH leakage suggest that reperfusion injury is mainly sustained within the first 2 h of reperfusion. However, the results of MTT reduction show that there is a progressive decrease in tissue viability over the 24 h reperfusion period, possibly reflecting the occurrence of tissue necrosis and apoptosis at different reperfusion times. In conclusion, the data provide evidence that the incubation of human atrial tissue in vitro is stable, and slices are viable for at least 24 h, which permits the study of early and delayed consequences of ischaemia and reperfusion in the human myocardium

Replacement of the Ascending Aorta, Aortic Root and Valve with a Novel Stentless Valved-Conduit

Purpose:- Biological valved-conduit grafts avoid the need for anticoagulation, and can exploit the excellent hemodynamic performance of stentless valves. Incorporation of sinuses of Valsalva into the neoaortic root can improve the function of the stentless valves. Description:- Here we present a novel prefabricated stentless valved-conduit incorporating sinuses of Valsalva and describe the technique of inplantation. The BioValsalva™ valved-conduit incorporates a stentless porcine aortic valve (Vascutek Elan™) suspended within a triple-layered vascular conduit (Triplex™, Vascutek, Renfrewshire, Scotland) constructed with sinuses of Valsalva. Evaluation:- The BioValsalva™ valve-conduit was used in twelve patients with aortic regurgitation due to annuloaortic ectasia unsuitable for aortic valve repair and concomitant ascending aorta aneurysm with no mortality and excellent functioned result. Conclusion:- The prefabricated, composite stentless-valved-conduit, which material is hemostatic and reduces bleeding, is easy to implant with short ischemic time, and lends itself well to a variety of insertion techniques

Lineage tracing of cardiac explant derived cells.

Aims Cultured cardiac explants produce a heterogeneous population of cells including a distinctive population of refractile cells described here as small round cardiac explant derived cells (EDCs). The aim of this study was to explore the source, morphology and cardiogenic potential of EDCs. Methods Transgenic MLC2v-Cre/ZEG, and actin-eGFP mice were used for lineage-tracing of EDCs in vitro and in vivo. C57B16 mice were used as cell transplant recipients of EDCs from transgenic hearts, as well as for the general characterisation of EDCs. The activation of cardiac-specific markers were analysed by: immunohistochemistry with bright field and immunofluorescent microscopy, electron microscopy, PCR and RT-PCR. Functional engraftment of transplanted cells was further investigated with calcium transient studies. Results Production of EDCs was highly dependent on the retention of blood-derived cells or factors in the cultured explants. These cells shared some characteristics of cardiac myocytes in vitro and survived engraftment in the adult heart in vivo. However, EDCs failed to differentiate into functional cardiac myocytes in vivo as demonstrated by the absence of stimulation-evoked intracellular calcium transients following transplantation into the peri-infarct zone. Conclusions This study highlights that positive identification based upon one parameter alone such as morphology or immunofluorescene is not adequate to identify the source, fate and function of adult cardiac explant derived cells

Lactate dehydrogenase (LDH) leakage (A1) and 3-(4,5-dimethyl thiazol-2-yl)-2,5diphenyl tetrazolium bromide (MTT) reduction (B1) and the correlation between IPreC and I/R alone for LDH release (A2) and MTT reduction (B2) of human myocardium muscles (n = 300 per group) subjected to 250 min of aerobic condictions (AC), 90 min of ischemia followed by 120 min of reoxygenation (I/R alone), or ischemic preconditioning (IPreC) induced by 5 min of ischemia followed by 5 min of reoxygenation prior to the 90 min of ischemia.

<p>The mean values are shown. *<i>p</i> < 0.05 vs AC group and †<i>p</i> < 0.05 vs I/R alone group.</p

Conditions affecting the capacity for cardioprotection by preconditioning (IPreC) obtained by substracting the LDH values of the I/R alone group from the IPreC group for each individual patients and then compared with the mean value of the remainder study population.

<p>Conditions affecting the capacity for cardioprotection by preconditioning (IPreC) obtained by substracting the LDH values of the I/R alone group from the IPreC group for each individual patients and then compared with the mean value of the remainder study population.</p

Randomized Controlled Trial of Intramuscular or Intracoronary Injection of Autologous Bone Marrow Cells into Scarred Myocardium

Introduction Previous studies on the transplantation of autologous bone marrow cells (BMC) in patients with chronic ischemic heart disease have focused on their effects on viable, peri-infarct tissue. We conducted a blinded, randomized controlled study to investigate whether intramuscular (IM) or intracoronary (IC) administration of BMC into non-viable scarred myocardium during coronary artery bypass grafting (CABG) improves contractile function of scar segments compared with CABG alone. Methods 63 elective CABG patients, with established myocardial scars diagnosed by dobutamine stress echocardiography (DSE) and confirmed at surgery, were randomized into control, IM or IC treatment groups. The BMC obtained at the time of surgery were injected into the middepth of the scar in the IM group or via the graft conduit supplying the scar in the IC group. Contractile function of the scar segments was assessed by DSE before and 6 months after treatment. Cardiac magnetic resonance imaging was also performed in the last 33 patients at the same time points. Results 12.5-29.4% of patients showed improved wall motion in at least one scar segment after BMC treatment but this effect was similar to that in the control group. Quantitatively, %systolic fractional thickening in scar segments did not improve with BMC administration. Scar transmurality, %infarct volume, left ventricular volumes and ejection fractions were also not improved by BMC. Discussion Autologous BMC, injected directly into the scar or the artery supplying the scar, do not improve contractility of non-viable scarred myocardium. Furthermore, BMC do not reduce scar size nor improve left ventricular function

Conditions affecting the susceptibility to ischemia/reoxygenation (I/R)-induced injury obtained by substracting the LDH values of the AC group from the I/R alone group for each individual patients and then compared with the mean value of the remainder study population.

<p>Conditions affecting the susceptibility to ischemia/reoxygenation (I/R)-induced injury obtained by substracting the LDH values of the AC group from the I/R alone group for each individual patients and then compared with the mean value of the remainder study population.</p

Indexes of basal redox status in human right atrial tissue (n = 90) from patients with conditions that affect myocardial susceptibilility to I/R-induced injury and the capacity for IPreC-mediated protection.

<p>Indexes of basal redox status in human right atrial tissue (n = 90) from patients with conditions that affect myocardial susceptibilility to I/R-induced injury and the capacity for IPreC-mediated protection.</p

Experimental protocol.

<p>Tissues in all groups were equilibrated for 30–40 min at 37°C in aerobic conditions. Muscle tissues were either maintained under aerobic conditions (AC) for the entire experimental period or subjected to 90 min of ischemia followed by 120 min of ischemia/reoxygenation (I/R alone) or preconditioned (IPreC) with 5 min of ischemia and 5 min of reoxygenation.</p