Safety evaluation of high-risk myocardial micro-biopsy in a swine model.

The objective of the study was to investigate the safety profile of high-risk micro-endomyocardial biopsy (micro-EMB) compared to conventional EMB in a large animal model. Twenty pigs were subjected to a maximum of 30 consecutive biopsies, including sampling from the free ventricular wall, with either micro-EMB (n = 10) or conventional EMB (n = 10). There were no major complications in the micro-EMB group (0/10), compared to six major complications in the EMB group (6/10; p = 0.003). Survival analysis further highlighted these differences (p = 0.004). There were significantly higher volumes of pericardial effusion in the EMB group (p = 0.01). The study shows a safety advantage of micro-EMB compared to standard EMB in the experimental high-risk circumstances investigated in this animal study. These results indicate enhanced possibilities to collect samples from sensitive areas by using the micro-EMB technique instead of standard EMB.

Micro-biopsy for detection of gene expression changes in ischemic swine myocardium: A pilot study.

Micro-endomyocardial biopsy (micro-EMB) is a novel catheter-based biopsy technique, aiming to increase flexibility and safety compared to conventional EMB. The technique was developed and evaluated in healthy swine. Therefore, the ability to detect disease related tissue changes could not be evaluated. The aim of the present pilot study was to investigate the ability to detect disease related gene expression changes using micro-EMB. Myocardial infarction was induced in three swine by coronary artery balloon occlusion. Micro-EMB samples (n = 164) were collected before, during, and after occlusion. RNA-sequencing was performed on 85 samples, and 53 of these were selected for bioinformatic analysis. A large number of responding genes was detected from the infarcted area (n = 1911). The early responding genes (n = 1268) were mostly related to apoptosis and inflammation. There were fewer responding genes two days after infarction (n = 6), which were related to extra-cellular matrix changes, and none after 14 days. In contrast to the infarcted area, samples harvested from a non-infarcted myocardial region showed considerably fewer regulated genes (n = 33). Deconvolution analysis, to estimate the proportion of different cell types, revealed a higher proportion of fibroblasts and a reduced proportion of cardiomyocytes two days after occlusion compared to baseline (p < 0.02 and p < 0.01, respectively. S5 File). In conclusion, this pilot study demonstrates the capabilities of micro-EMB to detect local gene expression responses at an early stage after ischemia, but not at later timepoints.

The Role of Carbon Dioxide in the Rat Acute Stroke Penumbra.

PURPOSE: The vasodilatory response to inhaled CO2 occurs in the acute stroke ischemic penumbra and may be a potential therapeutic modality. METHODS: Twenty-two Sprague-Dawley rats were subjected to 90-min occlusion of the M2 segment of the middle cerebral artery (M2CAO) by endovascular technique. The animals were administered different C02 concentrations and scanned serially with 9.4 T MRI. Infarct tissue was determined by diffusion-weighted imaging (DWI) and hypoperfused tissue was determined by arterial spin labeling (PWI). RESULTS: 4 animals were administered room air (RA)+ 6% CO2 (group 1), 6 animals RA+12% CO2 (Group 2) and 4 animals only RA (group 3). In the rats with CO2 administered (groups 1 and 2), the DWI lesion to cerebral hypoperfusion volume ratio (SD) at pre-CO2 administration, was 0.145(0.168), which increased to 0.708(0.731) during CO2 administration and reduced to 0.533(0.527) post-CO2 administration. In 9 of 10 rats the hypoperfused volume decreased when CO2 was administered. When CO2 was stopped the hypoperfused volume became larger again. Administration of RA+12% CO2 (Group 2) decreased the volume of CBF hypoperfusion significantly compared to the control group (95%CI: 0.084 ± 0.0213, p = 0.004). CONCLUSION: Inhaled CO2 appears to reduce the size of the hypoperfused tissue volume during acute stroke and may be a potential modality for treatment of acute ischemic stroke. These findings will nonetheless need to be validated in a larger cohort in other centers.

Myocardial micro-biopsy procedure for molecular characterization with increased precision and reduced trauma.

Endomyocardial biopsy is a valuable tool in cardiac diagnostics but is limited by low diagnostic yield and significant complication risks. Meanwhile, recent developments in transcriptomic and proteomic technologies promise a wealth of biological data from minimal tissue samples. To take advantage of the minimal tissue amount needed for molecular analyses, we have developed a sub-millimeter endovascular biopsy device, considerably smaller than current clinical equipment, and devised a low-input RNA-sequencing protocol for analyzing small tissue samples. In in vivo evaluation in swine, 81% of biopsy attempts (n = 157) were successful. High quality RNA-sequencing data was generated from 91% of the sequenced cardiac micro-biopsy samples (n = 32). Gene expression signatures of samples taken with the novel device were comparable with a conventional device. No major complications were detected either during procedures or during 7 days' follow-up, despite acquiring a relatively large number of biopsies (median 30) in each animal. In conclusion, the novel device coupled with RNA-sequencing provides a feasible method to obtain molecular data from the myocardium. The method is less traumatic and has a higher flexibility compared to conventional methods, enabling safer and more targeted sampling from different parts of the myocardium.

Oxygen metabolism MRI - A comparison with perfusion imaging in a rat model of MCA branch occlusion and reperfusion.

The penumbra is sustained by an increased extraction of oxygen (OEF) from blood to brain tissue. Metabolic imaging may improve penumbra specificity when examining stroke patients with wake-up stroke and a long time between admission and symptom onset. We used MRI to examine OEF, and compared the volume of regions with elevated OEF to the volume of regions with perfusion deficit in a M2 occlusion model (M2CAO) with preserved collateral blood flow. OEF was calculated from BOLD MRI examining tissue R2', with ASL perfusion imaging employed to determine cerebral blood flows (CBF) and volumes. Diffusion imaging was used to identify the ischemic core (IC). Examinations were performed during and after transient M2CAO in rats. The IC-OEF mismatch was significantly smaller than the IC-CBF mismatch during M2CAO. The penumbra OEF was significantly increased during M2CAO, and decreased significantly after reperfusion. The IC-OEF mismatch may provide increased penumbra specificity compared to IC-CBF mismatch regimens. Results strongly indicate the potential of metabolic MRI for thrombectomy patient selection in cases with a long time from symptom onset to admission. Results demonstrate the effectiveness of reperfusion in alleviating metabolic disturbances in ischemic regions, emphasizing fast treatment to achieve significant neurological recovery in stroke patients.

Transcriptomic analysis of the harvested endothelial cells in a swine model of mechanical thrombectomy.

PURPOSE: In mechanical thrombectomy (MT) for ischemic stroke, endothelial cells (ECs) from intracranial blood vessels adhere to the stent retriever device and can be harvested. However, understanding the molecular biology and the role of the endothelium in different pathological conditions remains insufficient. The purpose of the study was to characterize and analyze the molecular aspect of harvested ECs using cell culture and transcriptomic techniques in an MT swine model relevant to clinical ischemic stroke. METHODS: In swine, preformed thrombi were injected into the external carotid and subclavian arteries to occlude their branches. MT was performed according to clinical routine. The stent retriever device and thrombus were treated with cell dissociation buffer. The resulting cell suspension was analyzed by immunohistochemistry and was cultured. Cultured cells were analyzed using single-cell RNA sequencing (scRNA-seq) after fluorescence-activated cell sorting (FACS). RESULTS: A total number of 37 samples were obtained containing CD31-positive cells. Cell culture was successful in 90% of samples, and the cells expressed multiple typical EC protein markers. Eighty-nine percent of the sorted cells yielded high-quality transcriptomes, and single-cell transcriptomes from cultured cells showed that they expressed typical endothelial gene patterns. Gene expression analysis of ECs from an occluded artery did not show distinctive clustering into subtypes. CONCLUSION: ECs harvested during MT can be cultured and analyzed using single-cell transcriptomic techniques. This analysis can be implemented in clinical practice to study the EC gene expression of comorbidities, such as hypertension, diabetes mellitus, and metabolic syndrome, in patients suffering from acute ischemic stroke.