Fredrik Yannopoulos

Remote Ischemic Preconditioning Protects the Brain Against Injury After Hypothermic Circulatory Arrest

Background— Ischemic preconditioning (IPC) is a mechanism protecting tissues from injury during ischemia and reperfusion. Remote IPC (RIPC) can be elicited by applying brief periods of ischemia to tissues with ischemic tolerance, thus protecting vital organs more susceptible to ischemic damage. Using a porcine model, we determined whether RIPC of the limb is protective against brain injury caused by hypothermic circulatory arrest (HCA). Methods and Results— Twelve piglets were randomized to control and RIPC groups. RIPC was induced in advance of cardiopulmonary bypass by 4 cycles of 5 minutes of ischemia of the hind limb. All animals underwent cardiopulmonary bypass followed by 60 minutes of HCA at 18°C. Brain metabolism and electroencephalographic activity were monitored for 8 hours after HCA. Assessment of neurological status was performed for a week postoperatively. Finally, brain tissue was harvested for histopathological analysis. Study groups were balanced for baseline and intraoperative parameters. Brain lactate concentration was significantly lower (P<0.0001, ANOVA) and recovery of electroencephalographic activity faster (P<0.05, ANOVA) in the RIPC group. RIPC had a beneficial effect on neurological function during the 7-day follow-up (behavioral score; P<0.0001 versus control, ANOVA). Histopathological analysis demonstrated a significant reduction in cerebral injury in RIPC animals (injury score; mean [interquartile range]: control 5.8 [3.8 to 7.5] versus RIPC 1.5 [0.5 to 2.5], P<0.001, t test). Conclusions— These data demonstrate that RIPC protects the brain against HCA-induced injury, resulting in accelerated recovery of neurological function. RIPC might be neuroprotective in patients undergoing surgery with HCA and improve long-term outcomes. Clinical trials to test this hypothesis are warranted.

Safety and biodistribution study of bone marrow–derived mesenchymal stromal cells and mononuclear cells and the impact of the administration route in an intact porcine model

BACKGROUND AIMS Bone marrow mononuclear cells (BM-MNCs) and bone marrow-derived mesenchymal stem stromal cells (BM-MSCs) could have therapeutic potential for numerous conditions, including ischemia-related injury. Cells transplanted intravascularly may become entrapped in the lungs, which potentially decreases their therapeutic effect and increases the risk for embolism. METHODS Twelve pigs were divided into groups of 3 and received (99m)Tc- hydroxymethyl-propylene-amine-oxime-labeled autologous BM-MNCs or allogeneic BM-MSCs by either intravenous (IV) or intra-arterial (IA) transplantation. A whole body scan and single photon emission computed tomography/computed tomography (SPECT/CT) were performed 8 h later, and tissue biopsies were collected for gamma counting. A helical CT scan was also performed on 4 pigs to detect possible pulmonary embolism, 2 after IV BM-MSC injection and 2 after saline injection. RESULTS The transplantation route had a greater impact on the biodistribution of the BM-MSCs than the BM-MNCs. The BM-MNCs accumulated in the spleen and bones, irrespective of the administration route. The BM-MSCs had relatively higher uptake in the kidneys. The IA transplantation decreased the deposition of BM-MSCs in the lungs and increased uptake in other organs, especially in the liver. Lung atelectases were frequent due to mechanical ventilation and attracted transplanted cells. CT did not reveal any pulmonary embolism. CONCLUSIONS Both administration routes were found to be safe, but iatrogenic atelectasis might be an issue when cells accumulate in the lungs. The IA administration is effective in avoiding pulmonary entrapment of BM-MSCs. The cell type and administration method both have a major impact on the acute homing.

Improved Cerebral Recovery From Hypothermic Circulatory Arrest After Remote Ischemic Preconditioning

BACKGROUND Remote ischemic preconditioning is a novel method of reducing ischemia-reperfusion injury in which a transient ischemic period of the limb provides systemic protection against a prolonged ischemic insult. This method of preconditioning has shown some potential in ameliorating ischemia-related injury in various organs and experimental settings. We hypothesized that remote ischemic preconditioning might also improve the recovery from hypothermic circulatory arrest (HCA). METHODS Twenty-four juvenile pigs underwent 60 minutes of HCA at 18 degrees C with either transient right hind leg ischemic preconditioning or no ischemic preconditioning. Preconditioning was induced by four cycles of 5-minute ischemia periods with three 5-minute reperfusion periods in between. Microdialysis and electroencephalography (EEG) data were recorded to detect any possible changes during the recovery phase. RESULTS The EEG data showed that the remote ischemic preconditioning group had significantly better EEG recovery time and a lower burst suppression ratio throughout the follow-up period. Cerebral extracellular glucose and glycerol content rose significantly immediately after HCA in the control group compared with the remote ischemic preconditioning group, and significantly higher lactate concentrations were measured in the control group at 5 and 6 hours after reperfusion, indicating a difference in cerebral metabolism. CONCLUSIONS Our data imply that remote ischemic preconditioning improves the recovery from HCA. It provides a faster recovery of cortical neuronal activity and protection against potential oxygen radical-mediated ischemia damage during and after HCA. In addition, it seems to protect from a late phase lactate and pyruvate burst, mitigating possible damage from an anaerobic metabolism phase.

Review of remote ischemic preconditioning: from laboratory studies to clinical trials

Abstract In remote ischemic preconditioning (RIPC) short periods of non-lethal ischemia followed by reperfusion of tissue or organ prepare remote tissue or organ to resist a subsequent more severe ischemia-reperfusion injury. The signaling mechanism of RIPC can be humoral communication, neuronal stimulation, systemic modification of circulating immune cells, and activation of hypoxia inducible genes. Despite promising evidence from experimental studies, the clinical effects of RIPC have been controversial. Heterogeneity of inclusion and exclusion criteria and confounding factors such as comedication, anesthesia, comorbidities, and other risk factors may have influenced the efficacy of RIPC. Although the cardioprotective pathways of RIPC are more widely studied, there is also evidence of benefits in CNS, kidney and liver protection. Future research should explore the potential of RIPC, not only in cardiac protection, but also in patients with threatening ischemia of the brain, organ transplantation of the heart, liver and kidney and extensive cardiovascular surgery. RIPC is generally well-tolerated, safe, effective, and easily feasible. It has a great prospect for ischemic protection of the heart and other organs.

High number of transplanted stem cells improves myocardial recovery after AMI in a porcine model

Abstract Objective. The clinical data considering the bone marrow mononuclear cell (BMMNC) therapy in treatment for acute myocardial infarction (AMI) are controversial and the mechanisms remain unknown. Our objective was to study the cardiac function and changes in cytokine levels after administration of BMMNC in experimental AMI model. Design. Unlabeled or Super-Paramagnetic-Iron-Oxide-labeled BMMNCs or saline was injected into myocardium of 31 pigs after circumflex artery occlusion. Ejection fraction (EF) was measured preoperatively, postoperatively and at 21 days by echocardiography. Cardiac MRI was performed postoperatively and after 21 days in 7 BMMNC animals. Serum cytokine levels were measured at baseline, 24 h and 21 days. Cellular homing was evaluated comparing MRI and histology. Results. From baseline to 21 days EF decreased less in BMMNC group (EF mean control -19 SD 12 vs. BMMNC -4 SD 15 percentage points p = 0.02). Cytokine concentrations showed high variability between the animals. MRI correlated with histology in cell detection and revealed BMMNCs in the infarction area. By MRI, EF improved 11 percentage points. The improvement in EF was associated with the number of transplanted BMMNCs detected in the myocardium. Conclusion. BMMNC injection after AMI improved cardiac function. Quantity of transplanted BMMNCs correlated with the improvement in cardiac function after AMI.

Leg ischaemia before circulatory arrest alters brain leucocyte count and respiratory chain redox state

OBJECTIVES Remote ischaemic preconditioning and its neuroprotective abilities are currently under investigation and the method has shown significant effects in several small and large animal studies. In our previous studies, leucocyte filtration during cardiopulmonary bypass reduced cerebrocortical adherent leucocyte count and mitigated cerebral damage after hypothermic circulatory arrest (HCA) in piglets. This study aimed to obtain and assess direct visual data of leucocyte behaviour in cerebral vessels after hypothermic circulatory arrest following remote ischaemic preconditioning. METHODS Twelve native stock piglets were randomized into a remote ischaemic preconditioning group (n = 6) and a control group (n = 6). The intervention group underwent hind-leg ischaemia, whereas the control group received a sham-treatment before a 60-min period of hypothermic circulatory arrest. An intravital microscope was used to obtain measurements from the cerebrocortical vessel in vivo. It included three sets of filters: a violet filter to visualize microvascular perfusion and vessel diameter, a green filter for visualization of rhodamine-labelled leucocytes and an ultraviolet filter for reduced nicotinamide adenine dinucleotide (NADH) analysis. The final magnification on the microscope was 400. After the experiment, cerebral and cerebellar biopsies were collected and analysed with transmission electron microscope by a blinded analyst. RESULTS In the transmission electron microscope analysis, the entire intervention group had normal, unaffected rough endoplasmic reticulums in their cerebellar tissue, whereas the control group had a mean score of 1.06 (standard deviation 0.41) (P = 0.026). The measured amount of adherent leucocytes was lower in the remote ischaemic preconditioning group. The difference was statistically significant at 5, 15 and 45 min after circulatory arrest. Statistically significant differences were seen also in the recovery phase at 90 and 120 min after reperfusion. Nicotinamide adenine dinucleotide autofluorescence had statistically significant differences at 10 min after cooling and at 120 and 180 min after hypothermic circulatory arrest. CONCLUSIONS Remote ischaemic preconditioning seems to provide better mitochondrial respiratory chain function as indicated by the higher NADH content. It simultaneously provides a reduction of adherent leucocytes in cerebral vessels after hypothermic circulatory arrest. Additionally, it might provide some degree of cellular organ preservation as implied by the electron microscopy results.

Intra-arterial bone marrow mononuclear cell distribution in experimental global brain ischaemia

Abstract Objectives. Bone marrow mononuclear cells (BM-MNCs) can ameliorate focal ischaemic brain injury. A global ischaemic brain injury, which can occur after cardiac or thoracic surgery, could be an essential target for BM-MNCs. No studies using BM-MNCs for this indication have been conducted. Design. Ten porcine underwent a global normothermic ischaemic insult, followed by an intra-arterial injection of Technetium99m-HMPAO-labelled BM-MNCs after 2, 4, 6, 12 or 24 hours. A whole-body scan and a SPECT/CT were performed 2 hours after the injection. Severity of the injury was assessed with EEG and tissue biopsies were analysed by scintigraphy. Results. The majority of the cells appeared in the lungs and the liver. Only a minimal number of cells were located in the brain. Median distribution of cells between organs in all animals was as follows: lungs 32.7% (30.6–38.2), liver 14.2% (12.0–17.2), spleen 7.3% (3.3–11.3) and kidneys 2.5% (2.0–3.3). The transplanted cells could not be detected within the brain tissue by radionuclide imaging. Conclusions. Intra-arterially transplanted BM-MNCs did not migrate to the damaged brain tissue in significant quantity when transplanted during the first 24 hours after the global ischaemic insult, contrary to results with models of focal brain injury.

Remote ischemic precondition preserves cerebral oxygen tension during hypothermic circulatory arrest

Abstract Objectives. Remote ischemic preconditioning (RIPC) is a novel and promising method of mitigating neurological injury. In previous animal studies, RIPC has provided substantial neuroprotective effects. We hypothesized that the promising neuroprotective properties were a consequence of a better oxygen consumption profile during hypothermic circulatory arrest (HCA). Design. Six 7-week-old female pigs were randomly assigned to undergo the 60 minutes of HCA with the right hind leg receiving transient RIPC preoperatively and six animals were assigned to a control group that underwent 60 minutes of HCA without any preconditioning. A combined temperature/oxygen-tension probe was inserted into the parietal cortex of each animal to monitor cerebral oxygen tension during experiments. Results. The RIPC group had significantly higher cerebral oxygen tension readings throughout the HCA. Statistically significant differences were measured from the 20 minute time point onwards in every time point up to the 60 minute time point. Conclusions. This study shows that RIPC performed before HCA conserves the cerebral oxygen tension during a circulatory arrest. RIPC could possibly prolong the safe operating time during HCA as cerebral oxygen content is preserved throughout circulatory arrest.

Granulation tissue is altered after intramyocardial and intracoronary bone marrow-derived cell transfer for experimental acute myocardial infarction

BACKGROUND Bone marrow-derived mononuclear cell (BMMC) treatment in acute myocardial infarction (AMI) has been shown to have a beneficial effect. Our objective was to study in detail the histopathological process after the cell therapy after intramyocardial (IM) or intracoronary (IC) administration of BMMCs following experimental AMI. METHODS Twenty-fours pigs were randomized to the IM group (n=8), the IC group (n=8), and the control group (n=8).After 90 min of transient occlusion of the circumflex coronary artery, BMMCs were injected either intramyocardially or by a transfemoral catheter into the circumflex coronary artery. Echocardiography was performed preoperatively, postoperatively, and after a 21-day recovery period. The heart biopsies were examined histopathologically. Volumetric ex vivo CT scan was performed to evaluate calcification of the infarcted myocardium. RESULTS The ejection fraction (EF) showed significant recovery in the IM group compared to the control group at Day 21 (P=.05). Despite beneficial histological changes in the infarction site in the IC group, compared to the control group, EF failed to recover. Reduction of collagen density that depicts scar formation was seen in both cell therapy groups compared to the control (P<.001). The number of mitotic cells was higher in the control group compared to the cell therapy groups (P<.001). The IC and IM groups differed significantly from each other in muscle-specific actin staining (P<.001) and smooth muscle actin staining (P<.004). The IM therapy group showed higher density for both stainings. Additionally, macrophage density was higher in the IC group compared to the IM and control groups (P<.002). Both cell therapy regimens substantially diminished tissue calcification; due to the large variation, the effect was not statistically significant. CONCLUSION BMMC therapy launches cellular changes that affect mostly the repair process in the granulation tissue. The cell transplantation method might have some effect on the magnitude of the effect.

Performance of CHA2DS2-VASc score for stroke prediction after surgical aortic valve replacement

Objective: Stroke is a frequent complication occurring early and late after surgical aortic valve replacement. There is an unmet clinical need for simple tools to assess postoperative stroke risk. We sought to assess the predictive performance of Congestive heart failure; Hypertension; Age ≥75 (doubled); Diabetes mellitus; prior Stroke, transient ischemic attack or thromboembolism (doubled); Vascular disease; Age 65 to 74; Sex category (female) (CHA2DS2‐VASc) score in patients undergoing surgical aortic valve replacement with a bioprosthesis. Methods: Seven hundred fourteen patients undergoing isolated surgical aortic valve replacement with a bioprosthesis at 4 university hospitals were included. Data were collected retrospectively from patient records and monitored by an independent party. Results: Median follow‐up time was 4.8 years. Mean CHA2DS2‐VASc score was 4.1 ± 1.6. Low (scores, 0‐1), high (scores, 2‐4), and very high (scores, 5‐9) CHA2DS2‐VASc scores were observed in 39 (5.5%), 400 (56.0%), and 262 (38.5%) patients, respectively. Incidences of stroke or transient ischemic attack at 1 year were 2.6%, 4.8%, and 10.7%; at 5 years incidences were 5.2%, 14.0%, and 21.9%; and at 10 years incidence were 5.2%, 20.7%, and 37.9% for patients in low, high, and very high scores, respectively. Incidences of major bleeds at 1 year were 0%, 1.8%, and 2.7%; at 5 years incidences were 0%, 5.4%, and 8.7%; and at 10 years incidences were 0%, 9.0%, and 27.1%, respectively. Competing risk analysis showed that patients with CHA2DS2‐VASc score of 5 through 9 had a significantly increased risk of stroke or transient ischemic attack (hazard ratio, 4.75; 95% confidence interval, 1.09‐20.6; P = .037) irrespective of preoperative or new‐onset in‐hospital atrial fibrillation compared with low‐risk patients. Conclusions: CHA2DS2‐VASc is a valuable tool to identify patients with increased risk of stroke and major bleeding, and for whom alternative strategies for prevention of late neurologic complications should be adopted.