Potential of lysophosphatidylinositol as a prognostic indicator of cardiac arrest using a rat model

Aims: The potential of a lysophosphatidylinositol species, LPI(18:0), as a biomarker of ischaemia was tested using a rat model of cardiac arrest (CA).

Methods: Male Sprague-Dawley rats were subjected to asphyxia-induced CA or CA followed by cardiopulmonary bypass (CPB) resuscitation. The brain, heart, kidney and liver tissues were harvested from rats after 0, 5, 10, 20, 30 and 60?min CA and 30?min CA followed by 60?min CPB resuscitation. Blood samples were collected from inferior vena cava and hepatic veins following 30?min CA. Phospholipids were extracted from the four tissues and blood and analysed by HPLC-MS.

Results: The relative content of LPI(18:0) compared to a phosphatidylinositol species, PI(18:0,22:4), was significantly increased in the brain, heart, liver and kidney following 30?min CA and decreased following CPB resuscitation. In addition, the increase of the LPI(18:0)/PI(18:0,22:4) ratio in the four tissues was proportional to the duration of ischaemia for CA lasting up to 60?min. The ratio was also found to be increased in plasma from the hepatic vein following 30?min CA.

Conclusion: LPI(18:0) is a good indicator of CA downtime and has a potential to be used for early prognostication of outcome in CA.     

Recover of peripheral nerve function after prolong hypothermic cardiac arrest in a porcine model with extra corporeal life support

ObjectivesSurviving long lasting cardiac arrest following accidental hypothermia has been reported after treatment with extra corporeal life support (ECLS), but there is a risk of neurologic injury. Most surviving hypothermia patients have a prolonged stay in the intensive care unit, where most patients experience polyneuropathy. Theoretically, accidental hypothermic cardiac arrest may in itself contribute to polyneuropathy. This study was designed to examine the impact of three hours of cardiac arrest at a core temperature of 20 °C followed by reanimation of peripheral nerve function.MethodsSeven pigs were cannulated for ECLS and cooled to a core temperature of 20 °C followed by three hours of circulatory arrest where the extremities were packed with ice. After three hours, ECLS was started and rewarming was performed. During the process, neural testing of the ulnar nerve (a somatic nerve) and of the vagus nerve (an autonomic nerve) were performed and blood was drawn for analysis of p-potassium, serum-neuron-specific enolase, and S100b protein.ResultsThe ulnar nerve was cooled from 34.9±1.6 °C to 12.8±3.8 °C and the vagus nerve from 36.2±1.2 °C to 15.4±1.4 °C. Physiologic function of both somatic and autonomic nerves were strongly affected by cooling, but recovered to almost normal levels during rewarming, even after three hours of hypothermic cardiac arrest. P-potassium rose from 3.9 (3.6–4.6) mmol/l to 8.1 (7.2–9.1) mmol/l after three hours of cardiac arrest, but normalized after recirculation. There was no rise in serum-neuron-specific enolase, but a slight rise in S100b protein during three hours of hypothermic cardiac arrest was observed. All pigs obtained return of spontaneous circulation (ROSC).ConclusionsReanimation after three hours of hypothermic cardiac arrest using ECLS was possible with no or, if present, minor damage to the two nerves tested.     

Monitoring microbiological and biochemical changes in fermented yam

The microbiological and biochemical changes occurring in yam (Dioscorea rotundata L.) tissues fermented in 2% brine under anaerobic conditions at ambient temperature were studied. The ash. fibre and fat contents of the yam tissues were reduced at an average rate of 1.98, 1.72 and 0.42 mg g⁻¹ d⁻¹, respectively.Lactobacillus andLeuconostoc spp., as well as other fermentative microorganisms, showed a tendency to increased growth. The decrease in the total aerobic count after a 4-d fermentation was due to depletion of available nutrients, increased acidity of the medium and complete anaerobiosis of the environment. The fermenting microbes showed a higher amylolytic activity, releasing between 52 and 73 mg glucose per mL, than proteolytic activity, where between 120 and 250 μg amino acids per mL was released.     

兔心脏骤停模型方法的建立

犬心脏骤停模型报道较多,而兔心脏骤停模型建立方法的研究尚未见报道.为了配合"心脏骤停再灌注期内皮素、氧自由基、降钙素的影响及对抗作用研究"的实施,我们对兔心脏骤停模型建立的方法进行了研究,并测定了缺血再灌注后兔血中自由基含量的变化,现报告如下.     

Immobilization for carbon ion beam ablation of cardiac structures in a porcine model

IntroductionWhereas hadron therapy of static targets is clinically established, treatment of moving organs remains a challenge. One strategy is to minimize motion of surrounding tissue mechanically and to mitigate residual motion with an appropriate irradiation technique. In this technical note, we present and characterize such an immobilization technique for a novel noncancerous application: the irradiation of small targets in hearts with scanned carbon ion beams in a porcine model for elimination of arrhythmias.Material and methodsA device for immobilization was custom-built. Both for the treatment planning 4D-CT scan and for irradiation, breath-hold at end-exhale was enforced using a remotely-controlled respirator. Target motion was thus reduced to heartbeat only. Positioning was verified by orthogonal X-rays followed by couch shift if necessary. Reproducibility of bony anatomy, diaphragm, and heart position after repositioning and between repeated breath-hold maneuvers was evaluated on X-rays and cardiac-gated 4D-CTs. Treatment was post hoc simulated on sequential 4D-CTs for a subset of animals, after immediate repositioning and after a delay of one week, similar to the delay between imaging and irradiation.ResultsBreath-hold without repositioning was highly reproducible with an RMS deviation of at most one millimeter. 4D-CTs showed larger deformations in soft tissue, but treatment simulation on sequential images resulted in full target coverage (V95 >95%).ConclusionThe method of immobilization permitted reproducible positioning of mobile, thoracic targets for range-sensitive particle therapy. The presented immobilization strategy could be a reasonable approach for future animal investigations with the ultimate goal of translation to therapy in men.     

Diabetes and arterial extracellular matrix changes in a porcine model of atherosclerosis.

Patients with diabetes are at substantially increased risk for atherosclerosis and clinical cardiovascular events. Because arterial extracellular matrix contains several molecules, including biglycan, versican, hyaluronan, and elastin, that may affect plaque lipid retention and stability, we determined whether diabetes affects plaque content of these molecules in a porcine model of hyperlipidemia and diabetes. Coronary artery sections were studied from non-diabetic normolipidemic (n=11, N-NL), diabetic normolipidemic (n=10, DM-NL), non-diabetic hyperlipidemic (n=16, N-HL), and diabetic hyperlipidemic (n=15, DM-HL) animals. Hyaluronan, biglycan, versican, and apolipoprotein B (apoB) were detected with monospecific peptides or antisera, and elastin with Movat's pentachrome stain, and contents of each were quantified by computer-assisted morphometry. In the hyperlipidemic groups, diabetes was associated with a 4-fold increase in intimal area, with strong correlations between intimal area and immunostained areas for hyaluronan (R(2) = 0.83, p<0.0001), biglycan (R(2) = 0.72, p<0.0001), and apoB (R(2) = 0.23, p=0.0069). In contrast, median (interquartile range) intimal elastin content was significantly lower with diabetes [N-HL: 5.2% (2.4-8.2%) vs DM-HL: 1.5% (0.5-4.2%), p=0.01], and there was a strong negative correlation between intimal total and elastin areas (Spearman r = -0.62, p=0.001). In this porcine model, diabetes was associated with multiple extracellular matrix changes that have been associated with increased lesion instability, greater atherogenic lipoprotein retention, and accelerated atherogenesis.     

Feed-forward activation in a theoretical first-order biochemical pathway which contains an anticipatory model

This paper explores the consequences of the theoretical forward activation enzymatic pathway A ₀ A ₁ A ₂ A ₃ where E ₁ convents A ₀ to A ₁, E ₂ converts A ₁ to A ₂ and E ₃ converts A ₂ to A ₃. A ₀, which is environmentally determined, also serves to activate (or modulate) the activity of E ₃ in such a way as to keep the concentration of A ₂ ([A ₂]) constant at a particular set-point value. For mathematical simplicity, first order rate kinetics are used where k ₁, k ₂ and k ₃ are the rate constants for E ₁, E ₂, and E ₃ respectively. It is shown that if k ₃ is modulated appropriately so as keep [A ₂] at the setpoint value with a changing upstream [A ₀], then the modulation of k ₃ must be anticipatory of the dynamics of the biochemical pathway. In other words, the rate of change of k ₃, will be a function of [A ₀], k ₁, k ₂, k ₃, and the set-point value of [A ₂]. If the modulation of k ₃ does not perfectly model (anticipate) the reaction pathway of which it is a part, then the actual [A ₂] will deviate from the set-point value. This type of anticipatory feed-forward activation may represent an important aspect of biological organization.     

Approximate model of cooperative activation and crossbridge cycling in cardiac muscle using ordinary differential equations

We develop a point model of the cardiac myofilament (MF) to simulate a wide variety of experimental muscle characterizations including Force-Ca relations and twitches under isometric, isosarcometric, isotonic, and auxotonic conditions. Complex MF behaviors are difficult to model because spatial interactions cannot be directly implemented as ordinary differential equations. We therefore allow phenomenological approximations with careful consideration to the relationships with the underlying biophysical mechanisms. We describe new formulations that avoid mean-field approximations found in most existing MF models. To increase the scope and applicability of the model, we include length- and temperature-dependent effects that play important roles in MF responses. We have also included a representation of passive restoring forces to simulate isolated cell shortening protocols. Possessing both computational efficiency and the ability to simulate a wide variety of muscle responses, the MF representation is well suited for coupling to existing cardiac cell models of electrophysiology and Ca-handling mechanisms. To illustrate this suitability, the MF model is coupled to the Chicago rabbit cardiomyocyte model. The combined model generates realistic appearing action potentials, intracellular Ca transients, and cell shortening signals. The combined model also demonstrates that the feedback effects of force on Ca binding to troponin can modify the cytosolic Ca transient.     

Percentile growth charts for biomedical studies using a porcine model

Increasing rates of obesity and heart disease are compromising quality of life for a growing number of people. There is much research linking adult disease with the growth and development both in utero and during the first year of life. The pig is an ideal model for studying the origins of developmental programming. The objective of this paper was to construct percentile growth curves for the pig for use in biomedical studies. The body weight (BW) of pigs was recorded from birth to 150 days of age and their crown-to-rump length was measured over the neonatal period to enable the ponderal index (PI; kg/m3) to be calculated. Data were normalised and percentile curves were constructed using Cole's lambda-mu-sigma (LMS) method for BW and PI. The construction of these percentile charts for use in biomedical research will allow a more detailed and precise tracking of growth and development of individual pigs under experimental conditions.     

Asphyxial cardiac arrest, resuscitation and neurological outcome in a Landrace/Large-White swine model

The vast majority of laboratory studies on animals have focused on ventricular fibrillation (VF) and not on cardiac arrest (CA) resulting from asphyxia. The aim of this study was to develop a clinically relevant animal model in Landrace/Large-White swine of asphyxial CA resuscitated using the European Resuscitation Council guidelines. Survival and 24 h neurological outcome in terms of functional deficit were also evaluated. Asphyxial arrest was induced by clamping the endotracheal tube (ETT) in 10 Landrace/Large-White piglets. After 4 min of untreated arrest, resuscitation was initiated by unclamping the ETT, 100% oxygen mechanical ventilation, 2 min chest compressions and epinephrine administration. Advanced Life Support algorithm was followed. In case of restoration of spontaneous circulation, the animals were supported for one hour and then observed for 23 h. Coronary perfusion pressure was significantly higher in surviving animals (P < 0.001) during cardiopulmonary resuscitation. End-tidal CO(2) was significantly higher in the animals that survived than in non-surviving animals (P = 0.001). All of the animals were severely neurologically impaired 24 h after CA. This refined model of asphyxia CA is easily reproducible and may be used for pharmacological studies in CA.     

Monitoring of cell therapy and assessment of cardiac function using magnetic resonance imaging in a mouse model of myocardial infarction

We have developed a mouse severe combined immunodeficient (SCID) model of myocardial infarction based on permanent coronary artery occlusion that allows long-term functional analysis of engrafted human embryonic stem cell-derived cardiomyocytes, genetically marked with green fluorescent protein (GFP), in the mouse heart. We describe methods for delivery of dissociated cardiomyocytes to the left ventricle that minimize scar formation and visualization and validation of the identity of the engrafted cells using the GFP emission spectrum, and histological techniques compatible with GFP epifluorescence, for monitoring phenotypic changes in the grafts in vivo. In addition, we describe how magnetic resonance imaging can be adapted for use in mice to monitor cardiac function non-invasively and repeatedly. The model can be adapted to include multiple control or other cell populations. The procedure for a cohort of six mice can be completed in a maximum of 13 weeks, depending on follow-up, with 30 h of hands-on time.     

A biochemical model of matrix metalloproteinase 9 activation and inhibition

Matrix metalloproteinases (MMPs) are a class of extracellular and membrane-bound proteases involved in an array of physiological processes, including angiogenesis. We present a detailed computational model of MMP9 activation and inhibition. Our model is validated to existing biochemical experimental data. We determine kinetic rate constants for the processes of MMP9 activation by MMP3, MMP10, MMP13, and trypsin; inhibition by the tissue inhibitors of metalloproteinases (TIMPs) 1 and 2; and MMP9 deactivation. This computational approach allows us to investigate discrepancies in our understanding of the interaction of MMP9 with TIMP1. Specifically, we find that inhibition due to a single binding event cannot describe MMP9 inhibition by TIMP1. Temporally accurate biphasic inhibition requires either an additional isomerization step or a second lower affinity isoform of MMP9. We also theoretically characterize the MMP3/TIMP2/pro-MMP9 and MMP3/TIMP1/pro-MMP9 systems. We speculate that these systems differ significantly in their time scales of activation and inhibition such that MMP9 is able to temporarily overshoot its final equilibrium value in the latter. Our numerical simulations suggest that the ability of pro-MMP9 to complex TIMP1 increases this overshoot. In all, our analysis serves as a summary of existing kinetic data for MMP9 and a foundation for future models utilizing MMP9 or other MMPs under physiologically well defined microenvironments.     

Ischemic injury of the liver in a porcine model of cardiac death assessed by in vivo microdialysis

This study aims to evaluate the ischemic injury of the liver in a porcine model of cardiac death assessed by in vivo microdialysis. A porcine model of cardiac death was established by the suffocation method. Metabolic indicators were monitored using the microdialysis technique during warm ischemia time (WIT) and cold ischemia time (CIT). Pathological changes in ischemic-injured livers were observed by haematoxylin–eosin staining. The predictive values of biochemical parameters regarding the liver donor were evaluated by receiver operating characteristic curve analysis. All statistical analyses were conducted using the SPSS 18.0 software (SPSS Inc, Chicago, Illinois, USA). The degree of warm ischemic injury of the livers increased with prolonged WIT. Serum glucose, glycerol, pyruvate, lactic acid levels and lactate-to-pyruvate (L/P) ratio increased gradually during WIT. Results from Pearson correlation analyses indicated that serum lactate level and L/P ratio were positively associated with the degree of warm ischemic injury of the livers. The degree of cold ischemic injury of the livers gradually increased after 12 h CIT. Serum glucose, lactic acid and L/P ratio achieved a peak after 6–8 h of CIT, but gradually decreased with prolonged CIT. The peak of glycerol occurred after 8 h of CIT, while no changes were found with prolonged CIT. Serum pyruvate level exhibited an increasing trend after 12 h CIT. Our results confirmed that serum glucose and lactate levels were negatively correlated with cold ischemic injury of the liver. However, serum glycerol and pyruvate levels showed positive correlations with cold ischemic injury of the liver. The liver donor was unavailable after 30 min WIT and 24 h CIT. The cut-off value of serum lactate level for warm ischemic injury of the livers was 2.374 with a sensitivity (Sen) of 90 % and specificity (Spe) of 95 %; while the L/P radio was 0.026 (Sen = 80 %, Spe = 83 %). In addition, the cut-off values of serum glucose, lactate, glycerol and pyruvate levels for cold ischemic injury of the livers were 0.339 (Sen = 100 %, Spe = 77 %), 1.172 (Sen = 100 %, Spe = 61 %), 56.359 (Sen = 100 %, Spe = 65 %) and 0.020 (Sen = 100 %, Spe = 67 %), respectively. Our findings provide empirical evidences that serum glucose, lactate levels and L/P ratio may be good indicators for the degree of warm ischemic injury of the livers after cardiac death; while serum glucose, lactate, glycerol and pyruvate levels may be important in predicting cold ischemic injury.     

Invasive surgery reduces infarct size and preserves cardiac function in a porcine model of myocardial infarction

Reperfusion injury following myocardial infarction (MI) increases infarct size (IS) and deteriorates cardiac function. Cardioprotective strategies in large animal MI models often failed in clinical trials, suggesting translational failure. Experimentally, MI is induced artificially and the effect of the experimental procedures may influence outcome and thus clinical applicability. The aim of this study was to investigate if invasive surgery, as in the common open chest MI model affects IS and cardiac function. Twenty female landrace pigs were subjected to MI by transluminal balloon occlusion. In 10 of 20 pigs, balloon occlusion was preceded by invasive surgery (medial sternotomy). After 72 hrs, pigs were subjected to echocardiography and Evans blue/triphenyl tetrazoliumchloride double staining to determine IS and area at risk. Quantification of IS showed a significant IS reduction in the open chest group compared to the closed chest group (IS versus area at risk: 50.9 ± 5.4% versus 69.9 ± 3.4%, P = 0.007). End systolic LV volume and LV ejection fraction measured by echocardiography at follow‐up differed significantly between both groups (51 ± 5 ml versus 65 ± 3 ml, P = 0.033; 47.5 ± 2.6% versus 38.8 ± 1.2%, P = 0.005). The inflammatory response in the damaged myocardium did not differ between groups. This study indicates that invasive surgery reduces IS and preserves cardiac function in a porcine MI model. Future studies need to elucidate the effect of infarct induction technique on the efficacy of pharmacological therapies in large animal cardioprotection studies.     

Endovascular hypothermia improves post-resuscitation myocardial dysfunction by increasing mitochondrial biogenesis in a pig model of cardiac arrest

The aim of the study was to investigate the effects of endovascular hypothermia on mitochondrial biogenesis in a pig model of prolonged cardiac arrest (CA). Ventricular fibrillation was electrically induced, and animals were left untreated for 10 min; then after 6min of cardiopulmonary resuscitation (CPR), defibrillation was attempted. 25 animals that were successfully resuscitated were randomized into three groups: Sham group (SG, 5, no CA), normal temperature group (NTG, 5 for 12 h observation and 5 for 24 h observation), and endovascular hypothermia group (EHG, 5 for 12 h observation and 5 for 24 h observation). The core temperatures (T_c) in the EHG were maintained at 34 ± 0.5 °C for 6 h by an endovascular hypothermia device (Coolgard 3000), then actively increased at the speed of 0.5 °C per hour during the next 6 h to achieve a normal body temperature, while T_c were maintained at 37.5 ± 0.5 °C in the NTG. Cardiac and mitochondrial functions, the quantification of myocardial mitochondrial DNA (mtDNA), peroxisome proliferator-activated receptor coactivator-1α (PGC-1α), nuclear respiratory factor (NRF)-1, and NRF-2 were examined. Results showed that myocardial and mitochondrial injury and dysfunction increased significantly at 12 h and 24 h after CA. Endovascular hypothermia offered a method to rapidly achieve the target temperature and provide stable target temperature management (TTM). Cardiac outcomes were improved and myocardial injuries were alleviated with endovascular hypothermia. Compared with NTG, endovascular hypothermia significantly increased mitochondrial activity and biogenesis by amplifying mitochondrial biogenesis factors’ expressions, including PGC-1α, NRF-1, and NRF-2. In conclusions, endovascular hypothermia after CA alleviated myocardial and mitochondrial dysfunction, and was associated with increasing mitochondrial biogenesis.     

Developmental origins of pregnancy-induced cardiac changes: establishment of a novel model using the atrial natriuretic peptide gene-disrupted mice

Molecular and Cellular Biochemistry - Pregnancy evokes many challenges on the maternal cardiovascular system that may unmask predispositions for future disease. This is particularly evident for...     

Modulation of collateral artery growth in a porcine hindlimb ligation model using MCP-1

For an appropriate extrapolation to patients with peripheral arterial obstructive disease, we tested the efficacy of monocyte chemoattractant protein 1 (MCP-1) treatment in a porcine hindlimb ligation model. In 40 minipigs, a femoral artery ligation was performed. Control animals were examined immediately after ligation (n = 4) or after 2 wk of intra-arterial infusion of PBS (n = 11). A second group of animals was evaluated after intra-arterial infusion of 2.0 microg/h of MCP-1 for 48 h (followed by 12 days of PBS; n = 13) or 2 wk continuously (n = 12). In the terminal experiment after 2 wk, resting flow to the leg and peripheral arterial pressures were assessed without vasodilatation. Subsequently, vascular conductance was determined by using a pump-driven extracorporal circulation during maximal vasodilatation. The results showed that resting blood flow to the hindlimb was 53% of the normal after 2 wk of infusion of PBS, compared with 81% in both MCP-1 treatment groups (P < 0.05). Collateral conductance was 645 +/- 346 ml x min(-1) x mmHg(-1) after 2 wk of infusion with PBS, compared with 1,070 +/- 530 and 1,158 +/- 535 ml x min(-1) x mmHg(-1) after 48 h and 2 wk treatment with MCP-1, respectively (P < 0.05). Modulation of the process of arteriogenesis is feasible in this large animal model via intra-arterial infusion of the Cys-Cys-chemokine MCP-1.     

Evaluation of a porcine model for pulmonary gene transfer using a novel synthetic vector

Background

The pig lung, given its gross anatomical, histological and physiological similarities to the human lung, may be useful as a large animal model, in addition to rodents, in which to assess the potential of vectors for pulmonary airway gene transfer. The aim of this study was to assess the utility of the pig lung as a model of gene transfer to the human lung with a synthetic vector system.

Methods

The LID vector system consists of a complex of lipofectin (L), integrin‐binding peptide (I) and plasmid DNA (D). LID complexes containing a β‐galactosidase reporter gene under a CMV promoter or a control plasmid at1 mg/3 ml PBS, or 3 ml buffer, was administered to the right lower lobe ofthe pig lung through a bronchoscope. Pigs were culled at 48 h and lung sections prepared for immunohistochemical and histological analysis. Bronchoalveolar lavage fluid was collected and analysed for TNF‐α by ELISA.

Results

Immunohistochemical staining for the β‐galactosidase reporter gene indicated high efficiency of gene transfer by the LID vector to pig bronchial epithelium with 46% of large bronchi staining positively. There was no evidence for vector‐specific inflammation assessed by leukocytosis and cytokine production.

Conclusions

This study demonstrates the use of the pig for studies of gene transfer in the lung and confirms in a second species the potential of the LID vector for gene therapy of pulmonary diseases such as cystic fibrosis. Copyright © 2002 John Wiley & Sons, Ltd.