Proprotein convertase subtilisin/kexin type 9 (PCSK9) deficiency leads to lower cholesterol and is associated with reduced vascular complications in the general population. Cholesterol lowering may also have beneficial effects in sickle cell disease (SCD). The objective of this study was to determine effects of PCSK9 deficiency in a mouse model of SCD. Bone marrow transplantation (BMT) was performed from donor SCD mice to wild-type, PCSK9-deficient, and LDLR-deficient recipients to generate SCD controls (Pcsk9+/+, SCDbmt) with preserved PCSK9 status, SCD mice with deficiency of PCSK9 (Pcsk9−/−, SCDbmt), and SCD mice with deficiency of LDLR (Ldlr−/−, SCDbmt). Although cholesterol levels were lower in Pcsk9−/−, SCDbmt mice compared to Pcsk9+/+, SCDbmt mice, anemia was more severe in Pcsk9−/−, SCDbmt mice. Increased reticulocytosis, enhanced ex vivo erythrocyte sickling, and increased erythrocyte phosphatidylserine exposure was also observed. Livers, spleens, and kidneys contained increased iron in Pcsk9−/−, SCDbmt mice compared to Pcsk9+/+, SCDbmt mice consistent with greater hemolysis. SCD mice with deficiency of LDLR (Ldlr−/−, SCDbmt mice) had similar anemia as Ldlr+/+, SCDbmt mice despite higher serum cholesterol. In conclusion, deficiency of PCSK9 is associated with worsened anemia in SCD mice due to increased hemolysis. These findings may have implications for lipid-lowering strategies in patients with SCD, as well as for potential novel modifiers of anemia severity.

Objective Implantation of donor hearts with prolonged ischemic times is associated with worse survival. We sought to identify risk factors that modulate the effects of prolonged preservation. Methods Retrospective review of the United Network for Organ Sharing database (2000–2018) to identify transplants with >5 (n = 1526) or ≤5 h (n = 35,733) of donor heart preservation. In transplanted hearts preserved for >5 h, Cox‐proportional hazards identify modifiers for survival. Results Compared to ≤5 h, transplanted patients with >5 h of preservation spent less time in status 1B (76 ± 160 vs. 85 ± 173 days, p = .027), more commonly had ischemic cardiomyopathy (42.3% vs. 38.3%, p = .002), and less commonly received a blood type O heart (45.4% vs. 50.8%, p < .001). Longer heart preservation time was associated with a higher incidence of postoperative stroke (4.5% vs. 2.5%, p < .001), and dialysis (16.4% vs. 10.6%, p < .001). Prolonged preservation was associated with a greater likelihood of death from primary graft dysfunction (2.8% vs. 1.5%, p < .001) but there was no difference in death from acute (2.0% vs. 1.7%, p = .402) or chronic rejection (2.0% vs. 1.9%, p = .618). In transplanted patients with >5 h of heart preservation, multivariable analysis identified greater mortality with ischemic cardiomyopathy etiology (hazard ratio [HR] = 1.36, p < 0.01), pre‐transplant dialysis (HR = 1.84, p < .01), pre‐transplant extracorporeal membrane oxygenation (ECMO, HR = 2.36, p = .09), and O blood type donor hearts (HR = 1.35, p < .01). Conclusion Preservation time >5 h is associated with worse survival. This mortality risk is further amplified by preoperative dialysis and ECMO, ischemic cardiomyopathy etiology, and use of O blood type donor hearts.

The blood-brain barrier (BBB) maintains a stable brain microenvironment, and is a primary target of ischemic stroke. Breakdown of BBB integrity has been shown to initiate a devastating cascade of events and eventual neuronal loss in cerebral ischemia. Thus, it is important to identify mechanisms by which BBB integrity can be maintained or loss of BBB integrity can be reduced under ischemic stroke conditions. MicroRNAs (miRs) function as a novel class of small noncoding RNAs that negatively modulate protein expression. We and others have recently shown the involvement of miRs in the pathogenesis of ischemic brain injury. However, the functional significance and molecular mechanisms of miRs in regulating cerebrovascular pathology are poorly understood in ischemic stroke. In this study, we investigate the essential role of the miR-15a/16-1 cluster in the regulation of post-ischemic BBB dysfunction. We demonstrated that expression of the miR-15a/16-1 cluster is significantly increased in the cerebral vasculature and cultured BMECs after in vivo and in vitro ischemic stimuli. EC-selective miR-15a/16-1 genetic deficiency leads to reduced BBB leakage and smaller brain infarct in mice after middle cerebral artery occlusion (MCAO). In contrast, EC-selective miR-15a/16-1 transgenic overexpression aggravates BBB damage and brain infarct. Mechanistically, we found that the miR-15a/16-1 cluster can bind to the 3’-UTRs of major BBB tight junctions, Claudins, and inhibit their translation. Genetic manipulation of miR-15a/16-1 level by lentivirus significantly alters the expression of Claudin 2, 5 and 12 in BMECs under the Oxygen Glucose Deprivation (OGD) conditions. Accordingly, EC-selective miR-15a/16-1 transgenic overexpression remarkably inhibits cerebral expression of Claudins in mice following MCAO. These findings suggest that the miR-15a/16-1 cluster functions as a novel master regulator in BBB pathologies and increases brain injury after ischemic stroke. Elucidating the molecular mechanisms of miR-15a/16-1-mediated BBB dysfunction may lead us to discover novel pharmaceutical targets for the development of effective therapies against ischemic stroke.