Therapeutic Efficacy of Cryopreserved, Allogeneic Extracellular Vesicles for Treatment of Acute Myocardial Infarction.

Extracellular vesicles (EV) that are derived from endothelial progenitor cells (EPC) have been determined to be a novel therapy for acute myocardial infarction, with a promise for immediate "off-the-shelf" delivery. Early experience suggests delivery of EVs from allogeneic sources is safe. Yet, clinical translation of this therapy requires assurances of both EV stability following cryopreservation and absence of an adverse immunologic response to EVs from allogeneic donors. Thus, more bioactivity studies on allogeneic EVs after cold storage are necessary to establish quality standards for its widespread clinical use. Thus, in this study, we aimed to demonstrate the safety and efficacy in delivering cryopreserved EVs in allogeneic recipients as a therapy for acute myocardial infarction.In this present study, we have analyzed the cardioprotective effects of allogeneic EPC-derived EVs after storage at -80°C for 2 months, using a shear-thinning gel (STG) as an in vivo delivery vehicle. EV size, proteome, and nucleic acid cargo were observed to remain steady through extended cryopreservation via nanoparticle tracking analysis, mass spectrometry, and nanodrop analysis, respectively. Fresh and previously frozen EVs in STG were delivered intramyocardially in a rat model of myocardial infarction (MI), with both showing improvements in contractility, angiogenesis, and scar thickness in comparison to phosphate-buffered saline (PBS) and STG controls at 4 weeks post-MI. Pathologic analyses and flow cytometry revealed minimal inflammatory and immune upregulation upon exposure of tissue to EVs pooled from allogeneic donor cells.Allogeneic EPC-EVs have been known to elicit minimal immune activity and retain therapeutic efficacy after at least 2 months of cryopreservation in a post-MI model.

Brain proteomic atlas of alcohol use disorder in adult males.

Alcohol use disorder (AUD) affects transcriptomic, epigenetic and proteomic expression in several organs, including the brain. There has not been a comprehensive analysis of altered protein abundance focusing on the multiple brain regions that undergo neuroadaptations occurring in AUD. We performed a quantitative proteomic analysis using a liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis of human postmortem tissue from brain regions that play key roles in the development and maintenance of AUD, the amygdala (AMG), hippocampus (HIPP), hypothalamus (HYP), nucleus accumbens (NAc), prefrontal cortex (PFC) and ventral tegmental area (VTA). Brain tissues were from adult males with AUD (n = 11) and matched controls (n = 16). Across the two groups, there were >6000 proteins quantified with differential protein abundance in AUD compared to controls in each of the six brain regions. The region with the greatest number of differentially expressed proteins was the AMG, followed by the HYP. Pathways associated with differentially expressed proteins between groups (fold change > 1.5 and LIMMA p < 0.01) were analyzed by Ingenuity Pathway Analysis (IPA). In the AMG, adrenergic, opioid, oxytocin, GABA receptor and cytokine pathways were among the most enriched. In the HYP, dopaminergic signaling pathways were the most enriched. Proteins with differential abundance in AUD highlight potential therapeutic targets such as oxytocin, CSNK1D (PF-670462), GABAB receptor and opioid receptors and may lead to the identification of other potential targets. These results improve our understanding of the molecular alterations of AUD across brain regions that are associated with the development and maintenance of AUD. Proteomic data from this study is publicly available at www.lmdomics.org/AUDBrainProteomeAtlas/ .

Delayed delivery of endothelial progenitor cell-derived extracellular vesicles via shear thinning gel improves postinfarct hemodynamics.

BACKGROUND: Extracellular vesicles (EVs) are promising therapeutics for cardiovascular disease, but poorly-timed delivery might hinder efficacy. We characterized the time-dependent response to endothelial progenitor cell (EPC)-EVs within an injectable shear-thinning hydrogel (STG+EV) post-myocardial infarction (MI) to identify when an optimal response is achieved. METHODS: The angiogenic effects of prolonged hypoxia on cell response to EPC-EV therapy and EV uptake affinity were tested in vitro. A rat model of acute MI via left anterior descending artery ligation was created and STG+EV was delivered via intramyocardial injections into the infarct border zone at time points corresponding to phases of post-MI inflammation: 0 hours (immediate), 3 hours (acute inflammation), 4 days (proliferative), and 2 weeks (fibrosis). Hemodynamics 4 weeks post-treatment were compared across treatment and control groups (phosphate buffered saline [PBS], shear-thinning gel). Scar thickness and ventricular diameter were assessed histologically. The primary hemodynamic end point was end systolic elastance. The secondary end point was scar thickness. RESULTS: EPC-EVs incubated with chronically versus acutely hypoxic human umbilical vein endothelial cells resulted in a 2.56 ± 0.53 versus 1.65 ± 0.15-fold increase (P = .05) in a number of vascular meshes and higher uptake of EVs over 14 hours. End systolic elastance improved with STG+EV therapy at 4 days (0.54 ± 0.08) versus PBS or shear-thinning gel (0.26 ± 0.03 [P = .02]; 0.23 ± 0.02 [P = .01]). Preservation of ventricular diameter (6.20 ± 0.73 mm vs 8.58 ± 0.38 mm [P = .04]; 9.13 ± 0.25 mm [P = .01]) and scar thickness (0.89 ± 0.05 mm vs 0.62 ± 0.03 mm [P < .0001] and 0.58 ± 0.05 mm [P < .0001]) was significantly greater at 4 days, compared wit PBS and shear-thinning gel controls. CONCLUSIONS: Delivery of STG+EV 4 days post-MI improved left ventricular contractility and preserved global ventricular geometry, compared with controls and immediate therapy post-MI. These findings suggest other cell-derived therapies can be optimized by strategic timing of therapeutic intervention.