Cardiac transplantation with hearts from donors after circulatory declaration of death: haemodynamic and biochemical parameters at procurement predict recovery following cardioplegic storage in a rat model

OBJECTIVES: Donation after circulatory declaration of death (DCDD) could significantly improve the number of cardiac grafts for transplantation. Graft evaluation is particularly important in the setting of DCDD given that conditions of cardio-circulatory arrest and warm ischaemia differ, leading to variable tissue injury. The aim of this study was to identify, at the time of heart procurement, means to predict contractile recovery following cardioplegic storage and reperfusion using an isolated rat heart model. Identification of reliable approaches to evaluate cardiac grafts is key in the development of protocols for heart transplantation with DCDD. METHODS: Hearts isolated from anaesthetized male Wistar rats (n = 34) were exposed to various perfusion protocols. To simulate DCDD conditions, rats were exsanguinated and maintained at 37°C for 15-25 min (warm ischaemia). Isolated hearts were perfused with modified Krebs-Henseleit buffer for 10 min (unloaded), arrested with cardioplegia, stored for 3 h at 4°C and then reperfused for 120 min (unloaded for 60 min, then loaded for 60 min). Left ventricular (LV) function was assessed using an intraventricular micro-tip pressure catheter. Statistical significance was determined using the non-parametric Spearman rho correlation analysis. RESULTS: After 120 min of reperfusion, recovery of LV work measured as developed pressure (DP)-heart rate (HR) product ranged from 0 to 15 ± 6.1 mmHg beats min(-1) 10(-3) following warm ischaemia of 15-25 min. Several haemodynamic parameters measured during early, unloaded perfusion at the time of heart procurement, including HR and the peak systolic pressure-HR product, correlated significantly with contractile recovery after cardioplegic storage and 120 min of reperfusion (P < 0.001). Coronary flow, oxygen consumption and lactate dehydrogenase release also correlated significantly with contractile recovery following cardioplegic storage and 120 min of reperfusion (P < 0.05). CONCLUSIONS: Haemodynamic and biochemical parameters measured at the time of organ procurement could serve as predictive indicators of contractile recovery. We believe that evaluation of graft suitability is feasible prior to transplantation with DCDD, and may, consequently, increase donor heart availability.

Pregnancy outcome following exposure to angiotensin-converting enzyme inhibitors or angiotensin receptor antagonists: a systematic review

The objective was to analyze the outcome following prenatal exposure to angiotensin-converting enzyme inhibitors (ACE-Is) or angiotensin receptor antagonists (ARBs). For this purpose, a systematic review of published case reports and case series dealing with intrauterine exposure to ACE-Is or to ARBs using Medline as the source of data was performed. The publications retained for analysis included patients who were described individually, revealing, at minimum, the gestational age, substance used, period of medication intake, and the outcome. In total, 72 reports were included; 37 articles (118 well-documented cases) described the prenatal exposure to ACE-Is; and 35 articles (68 cases) described the prenatal exposure to ARBs. Overall, 52% of the newborns exposed to ACE-Is and 13% of the newborns exposed to ARBs did not exhibit any complications (P<0.0001). Neonatal complications were more frequent following exposure to ARBs and included renal failure, oligohydramnios, death, arterial hypotension, intrauterine growth retardation, respiratory distress syndrome, pulmonary hypoplasia, hypocalvaria, limb defects, persistent patent ductus arteriosus, or cerebral complications. The long-term outcome is described as positive in only 50% of the exposed children. Fetopathy caused by exposure to ACE-Is or ARBs has relevant neonatal and long-term complications. The outcome is poorer following exposure to ARBs. We propose the term "fetal renin-angiotensin system blockade syndrome" to describe the related clinical findings. Thirty years after the first description of ACE-I fetopathy, relevant complications are, at present, regularly described, indicating that the awareness of the deleterious effect of prenatal exposure to drugs inhibiting the renin-angiotensin system should be improved.

Outcome at two years of age in a Swiss national cohort of extremely preterm infants born between 2000 and 2008

Background While survival rates of extremely preterm infants have improved over the last decades, the incidence of neurodevelopmental disability (ND) in survivors remains high. Representative current data on the severity of disability and of risk factors associated with poor outcome in this growing population are necessary for clinical guidance and parent counselling. Methods Prospective longitudinal multicentre cohort study of preterm infants born in Switzerland between 240/7 and 276/7 weeks gestational age during 2000–2008. Mortality, adverse outcome (death or severe ND) at two years, and predictors for poor outcome were analysed using multilevel multivariate logistic regression. Neurodevelopment was assessed using Bayley Scales of Infant Development II. Cerebral palsy was graded after the Gross Motor Function Classification System. Results Of 1266 live born infants, 422 (33%) died. Follow-up information was available for 684 (81%) survivors: 440 (64%) showed favourable outcome, 166 (24%) moderate ND, and 78 (11%) severe ND. At birth, lower gestational age, intrauterine growth restriction and absence of antenatal corticosteroids were associated with mortality and adverse outcome (p < 0.001). At 360/7 weeks postmenstrual age, bronchopulmonary dysplasia, major brain injury and retinopathy of prematurity were the main predictors for adverse outcome (p < 0.05). Survival without moderate or severe ND increased from 27% to 39% during the observation period (p = 0.02). Conclusions In this recent Swiss national cohort study of extremely preterm infants, neonatal mortality was determined by gestational age, birth weight, and antenatal corticosteroids while neurodevelopmental outcome was determined by the major neonatal morbidities. We observed an increase of survival without moderate or severe disability.

Maternal and fetal cord blood lipids in intrauterine growth restriction

Small for gestational age neonates (SGA) could be subdivided into two groups according to the underlying causes leading to low birth weight. Intrauterine growth restriction (IUGR) is a pathologic condition with diminished growth velocity and fetal compromised well-being, while non-growth restricted SGA neonates are constitutionally (genetically determined) small. Antenatal sonographic measurements are used to differentiate these two subgroups. Maternal metabolic changes contribute to the pathogenesis of IUGR. A disturbed lipid metabolism and cholesterol supply might affect the fetus, with consequences for fetal programming of cardiovascular diseases. We evaluated fetal serum lipids and hypothesized a more atherogenic lipoprotein profile in IUGR fetuses.

The evaluation of the oxidative state of low-density lipoproteins in intrauterine growth restriction and preeclampsia

To evaluate the oxidative state of lipoproteins in pregnancies complicated by intrauterine growth restriction (IUGR) in comparison to preeclampsia (PE) and healthy pregnant control subjects (CN).

Detection and distribution of apoptotic cell death in normal and diseased canine cranial cruciate ligaments

One of the possible initiating factors in canine cranial cruciate ligament (CCL) rupture could be an abnormal pattern of ligament cell death. This study compared apoptotic cell death in sections of ruptured CCLs and normal controls, and examined nitric oxide (NO) production in joint tissues and correlated this to apoptosis. CCLs and cartilage from the lateral femoral condyle were harvested from 10 healthy dogs and 15 dogs with CCL rupture and ligaments were further processed to detect cleaved caspase-3 and to determine supernatant NO production in explant cultures. Apoptotic activity was greater in ruptured ligaments compared to controls. NO in ligaments showed a moderate but significant positive correlation with caspase-positive cells. The results suggest that increased apoptosis has a role in CCL rupture and that apoptosis may be influenced by local NO production.

Cardiac glycosides initiate Apo2L/TRAIL-induced apoptosis in non-small cell lung cancer cells by up-regulation of death receptors 4 and 5

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (Apo2L/TRAIL) belongs to the TNF family known to transduce their death signals via cell membrane receptors. Because it has been shown that Apo2L/TRAIL induces apoptosis in tumor cells without or little toxicity to normal cells, this cytokine became of special interest for cancer research. Unfortunately, cancer cells are often resistant to Apo2L/TRAIL-induced apoptosis; however, this can be at least partially negotiated by parallel treatment with other substances, such as chemotherapeutic agents. Here, we report that cardiac glycosides, which have been used for the treatment of cardiac failure for many years, sensitize lung cancer cells but not normal human peripheral blood mononuclear cells to Apo2L/TRAIL-induced apoptosis. Sensitization to Apo2L/TRAIL mediated by cardiac glycosides was accompanied by up-regulation of death receptors 4 (DR4) and 5 (DR5) on both RNA and protein levels. The use of small interfering RNA revealed that up-regulation of death receptors is essential for the demonstrated augmentation of apoptosis. Blocking of up-regulation of DR4 and DR5 alone significantly reduced cell death after combined treatment with cardiac glycosides and Apo2L/TRAIL. Combined silencing of DR4 and DR5 abrogated the ability of cardiac glycosides and Apo2L/TRAIL to induce apoptosis in an additive manner. To our knowledge, this is the first demonstration that glycosides up-regulate DR4 and DR5, thereby reverting the resistance of lung cancer cells to Apo2/TRAIL-induced apoptosis. Our data suggest that the combination of Apo2L/TRAIL and cardiac glycosides may be a new interesting anticancer treatment strategy.

CD40 ligation protects bronchial epithelium against oxidant-induced caspase-independent cell death

CD40 and its ligand regulate pleiotropic biological responses, including cell proliferation, differentiation, and apoptosis. In many inflammatory lung diseases, tissue damage by environmental or endogenous oxidants plays a major role in disease pathogenesis. As the epithelial barrier is a major target for these oxidants, we postulated that CD40, the expression of which is increased in asthma, plays a role in the regulation of apoptosis of bronchial epithelial cells exposed to oxidants. Using 16HBE 14o- cells exposed to oxidant stress, we found that ligation of CD40 (induced by G28-5 monoclonal antibodies) enhanced cell survival and increased the number of cells in G2/M (interphase between DNA synthesis and mitosis) of the cell cycle. This was associated with NF-kappaB and activator protein-1 activation and increased expression of the inhibitor of apoptosis, c-IAP1. However, oxidant stress-induced apoptosis was found to be caspase- and calpain-independent implicating CD40 ligation as a regulator of caspase-independent cell death. This was confirmed by the demonstration that CD40 ligation prevented mitochondrial release and nuclear translocation of apoptosis inducing factor. In conclusion, we demonstrate a novel role for CD40 as a regulator of epithelial cell survival against oxidant stress. Furthermore, we have identified, for the first time, an endogenous inhibitory pathway of caspase-independent cell death.

Causes of death and determinants of outcome in critically ill patients

INTRODUCTION: Whereas most studies focus on laboratory and clinical research, little is known about the causes of death and risk factors for death in critically ill patients. METHODS: Three thousand seven hundred patients admitted to an adult intensive care unit (ICU) were prospectively evaluated. Study endpoints were to evaluate causes of death and risk factors for death in the ICU, in the hospital after discharge from ICU, and within one year after ICU admission. Causes of death in the ICU were defined according to standard ICU practice, whereas deaths in the hospital and at one year were defined and grouped according to the ICD-10 (International Statistical Classification of Diseases and Related Health Problems) score. Stepwise logistic regression analyses were separately calculated to identify independent risk factors for death during the given time periods. RESULTS: Acute, refractory multiple organ dysfunction syndrome was the most frequent cause of death in the ICU (47%), and central nervous system failure (relative risk [RR] 16.07, 95% confidence interval [CI] 8.3 to 31.4, p < 0.001) and cardiovascular failure (RR 11.83, 95% CI 5.2 to 27.1, p < 0.001) were the two most important risk factors for death in the ICU. Malignant tumour disease and exacerbation of chronic cardiovascular disease were the most frequent causes of death in the hospital (31.3% and 19.4%, respectively) and at one year (33.2% and 16.1%, respectively). CONCLUSION: In this primarily surgical critically ill patient population, acute or chronic multiple organ dysfunction syndrome prevailed over single-organ failure or unexpected cardiac arrest as a cause of death in the ICU. Malignant tumour disease and chronic cardiovascular disease were the most important causes of death after ICU discharge.

Effects of lamotrigine alone and in combination with MK-801, phenobarbital, or phenytoin on cell death in the neonatal rat brain

The neonatal rat brain is vulnerable to neuronal apoptosis induced by antiepileptic drugs (AEDs), especially when given in combination. This study evaluated lamotrigine alone or in combination with phenobarbital, phenytoin, or the glutamate antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK-801) for a proapoptotic action in the developing rat brain. Cell death was assessed in brain regions (striatum, thalamus, and cortical areas) of rat pups (postnatal day 8) by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay, 24 h after acute drug treatment. Lamotrigine alone did not increase neuronal apoptosis when given in doses up to 50 mg/kg; a significant increase in cell death occurred after 100 mg/kg. Combination of 20 mg/kg lamotrigine with 0.5 mg/kg MK-801 or 75 mg/kg phenobarbital resulted in a significant increase in TUNEL-positive cells, compared with MK-801 or phenobarbital treatment alone. A similar enhancement of phenytoin-induced cell death occurred after 30 mg/kg lamotrigine. In contrast, 20 mg/kg lamotrigine significantly attenuated phenytoin-induced cell death. Lamotrigine at 10 mg/kg was without effect on apoptosis induced by phenytoin. Although the functional and clinical implications of AED-induced developmental neuronal apoptosis remain to be elucidated, our finding that lamotrigine alone is devoid of this effect makes this drug attractive as monotherapy for the treatment of women during pregnancy, and for preterm or neonatal infants. However, because AEDs are often introduced as add-on medication, careful selection of drug combinations and doses may be required to avoid developmental neurotoxicity when lamotrigine is used in polytherapy.

The death-associated protein kinase 2 is up-regulated during normal myeloid differentiation and enhances neutrophil maturation in myeloid leukemic cells

The death-associated protein kinase 2 (DAPK2) belongs to a family of Ca(2+)/calmodulin-regulated serine/threonine kinases involved in apoptosis. During investigation of candidate genes operative in granulopoiesis, we identified DAPK2 as highly expressed. Subsequent investigations demonstrated particularly high DAPK2 expression in normal granulocytes compared with monocytes/macrophages and CD34(+) progenitor cells. Moreover, significantly increased DAPK2 mRNA levels were seen when cord blood CD34(+) cells were induced to differentiate toward neutrophils in tissue culture. In addition, all-trans retinoic acid (ATRA)-induced neutrophil differentiation of two leukemic cell lines, NB4 and U937, revealed significantly higher DAPK2 mRNA expression paralleled by protein induction. In contrast, during differentiation of CD34(+) and U937 cells toward monocytes/macrophages, DAPK2 mRNA levels remained low. In primary leukemia, low expression of DAPK2 was seen in acute myeloid leukemia samples, whereas chronic myeloid leukemia samples in chronic phase showed intermediate expression levels. Lentiviral vector-mediated expression of DAPK2 in NB4 cells enhanced, whereas small interfering RNA-mediated DAPK2 knockdown reduced ATRA-induced granulocytic differentiation, as evidenced by morphology and neutrophil stage-specific maturation genes, such as CD11b, G-CSF receptor, C/EBPepsilon, and lactoferrin. In summary, our findings implicate a role for DAPK2 in granulocyte maturation.

Active caspase 3 and DNA fragmentation as markers for apoptotic cell death in primary and metastatic liver tumours

AIMS: The induction of tumour cell death by apoptosis is a major goal of cancer therapy and the in situ detection of apoptosis in tumour tissue has become an important diagnostic parameter. Different apoptosis detection methods assess distinct biochemical processes in the dying cell. Thus, their direct comparison is mandatory to evaluate their diagnostic value. The aim of this study was to compare the immunohistochemical detection of active caspase 3 and single-stranded DNA in primary and metastatic liver tumours as markers of apoptotic cell death. METHODS: We studied detection of active caspase 3 and single-stranded DNA in 20 primary hepatocellular carcinomas (HCC) and 20 liver metastases from colorectal carcinomas (CRC) using immunohistochemistry on paraffin sections. RESULTS: Our results reveal that both methods are suitable and sensitive techniques for the in situ detection of apoptosis, however, they also demonstrate that immunohistochemistry for active caspase 3 and single-stranded DNA have differential sensitivities in HCC and CRC. CONCLUSION: The sensitivity of apoptosis detection using immunohistochemistry for active caspase 3 and single-stranded DNA may be tumour cell type dependent.