Endovascular transplantation of mRNA-enhanced mesenchymal stromal cells results in superior therapeutic protein expression in swine heart.

Heart failure has a poor prognosis and no curative treatment exists. Clinical trials are investigating gene- and cell-based therapies to improve cardiac function. The safe and efficient delivery of these therapies to solid organs is challenging. Herein, we demonstrate the feasibility of using an endovascular intramyocardial delivery approach to safely administer mRNA drug products and perform cell transplantation procedures in swine. Using a trans-vessel wall (TW) device, we delivered chemically modified mRNAs (modRNA) and mRNA-enhanced mesenchymal stromal cells expressing vascular endothelial growth factor A (VEGF-A) directly to the heart. We monitored and mapped the cellular distribution, protein expression, and safety tolerability of such an approach. The delivery of modRNA-enhanced cells via the TW device with different flow rates and cell concentrations marginally affect cell viability and protein expression in situ. Implanted cells were found within the myocardium for at least 3 days following administration, without the use of immunomodulation and minimal impact on tissue integrity. Finally, we could increase the protein expression of VEGF-A over 500-fold in the heart using a cell-mediated modRNA delivery system compared with modRNA delivered in saline solution. Ultimately, this method paves the way for future research to pioneer new treatments for cardiac disease.

A novel noble metal stent coating reduces in vitro platelet activation and acute in vivo thrombosis formation: a blinded study.

Inherent to any stenting procedure is the prescription of dual antiplatelet therapy (DAPT) to reduce the platelet response. Clinical guidelines recommend 6-12 months of DAPT, depending on stent type, clinical picture and patient factors. Our hypothesis is that a nanostructured noble metal coating has the potential to reduce protein deposition and platelet activation. These effects would reduce subsequent thrombo-inflammatory reactions, potentially mitigating the need for an extensive DAPT in the acute phase. Here, a noble metal nanostructure coating on stents is investigated. Twelve pigs underwent endovascular implantation of coated and non-coated stents for paired comparisons in a blinded study design. The non-coated control stent was placed at the contralateral corresponding artery. Volumetric analysis of angiographic data, performed by a treatment blinded assessor, demonstrated a significant thrombus reduction for one of the coatings compared to control. This effect was already seen one hour after implantation. This finding was supported by in vitro data showing a significant reduction of coagulation activation in the coated group. This novel coating shows promise as an implant material addition and could potentially decrease the need for DAPT in the early phases of stent implementation.

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.

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.

Standards for opioid use disorder care: An assessment of Nordic approaches.

AIMS: Outcomes in opioid use disorder (OUD) in Nordic countries have improved with integrated treatment and harm-reduction programmes. Approaches and the standard of care are different across the region. Evidence of treatment needs and current approaches are defined from evidence to inform development of a common standard. METHOD: Evidence of population sizes and treatment approach collected. Common standards for care (harm reduction, pharmacotherapy, psychology/social therapy) defined for each country. RESULTS: Evidence defines number in treatment; potential population needing treatment not defined for all countries. Populations sizes, treatment access (ratio in treatment programme compared to total country population) defined: Sweden 4,000 in OUD care (access ratio 40); Finland 3,000 (55); Norway 8,000 (154); Denmark 7,500 (132). Approach to treatment similar: integrated treatment programmes standard. Care provided by specialists in outpatient clinics/primary care; secondary care/inpatient services are available. Harm reduction is limited in Sweden but available and more accessible elsewhere. Treatment entry criteria: access relatively unlimited in Norway and Denmark, more limited in Finland and Sweden. Standards of care defined: easy access to high-quality services, individual planning, care not limited by time, management of relapse, education for patients, continuous engagement, holistic approach including management of comorbidities, needle equipment programmes without limit, treatment in prisons as community. CONCLUSION: There are opportunities to improve OUD care in the Nordics. Policy makers and clinicians can advance OUD care and share common success factors. Collaborative work across the Nordic countries is valuable. Further research in clinical practice development can yield important results for the benefit of patients with OUD.

Principles for managing OUD related to chronic pain in the Nordic countries based on a structured assessment of current practice.

BACKGROUND: Long-term use of opioid analgesics (OA) for chronic pain may result in opioid use disorder (OUD). This is associated with adverse outcomes for individuals, families and society. Treatment needs of people with OUD related to chronic pain are different compared to dependence related to use, and also injection, of illicit opioids. In Nordic countries, day-to-day practical advice to assist clinical decision-making is insufficient. AIM: To develop principles based on expert clinical insights for treatment of OUD related to the long-term use of OA in the context of chronic pain. METHODS: Current status including an assessment of barriers to effective treatment in Finland, Denmark, Iceland, Norway, Sweden was defined using a patient pathway model. Evidence to describe best practice was identified from published literature, clinical guidelines and expert recommendations from practice experience. RESULTS: Availability of national treatment guidelines for OUD related to chronic pain is limited across the Nordics. Important barriers to effective care identified: patients unlikely to present for help, healthcare system set up limits success, diagnosis tools not used, referral pathways unclear and treatment choices not elucidated. Principles include the development of a specific treatment pathway, awareness/ education programs for teams in primary care, guidance on use of diagnostic tools and a flexible treatment plan to encourage best practice in referral, treatment assessment, choice and ongoing management via an integrated care pathway. Healthcare systems and registries in Nordic countries offer an opportunity to further research and identify population risks and solutions. CONCLUSIONS: There is an opportunity to improve outcomes for patients with OUD related to chronic pain by developing and introducing care pathways tailored to specific needs of the population.

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.