Objective—: Ischemia–reperfusion (I/R) injury significantly contributes to organ dysfunction and failure after myocardial infarction, stroke, and transplantation. In addition to its established role in the fibrinolytic system, plasminogen activator inhibitor-1 has recently been implicated in the pathogenesis of I/R injury. The underlying mechanisms remain largely obscure. Approach and Results—: Using different in vivo microscopy techniques as well as ex vivo analyses and in vitro assays, we identified that plasminogen activator inhibitor-1 rapidly accumulates on microvascular endothelial cells on I/R enabling this protease inhibitor to exhibit previously unrecognized functional properties by inducing an increase in the affinity of β2 integrins in intravascularly rolling neutrophils. These events are mediated through low-density lipoprotein receptor–related protein-1 and mitogen-activated protein kinase–dependent signaling pathways that initiate intravascular adherence of these immune cells to the microvascular endothelium. Subsequent to this process, extravasating neutrophils disrupt endothelial junctions and promote the postischemic microvascular leakage. Conversely, deficiency of plasminogen activator inhibitor-1 effectively reversed leukocyte infiltration, microvascular dysfunction, and tissue injury on experimental I/R without exhibiting side effects on microvascular hemostasis. Conclusions—: Our experimental data provide novel insights into the nonfibrinolytic properties of the fibrinolytic system and emphasize plasminogen activator inhibitor-1 as a promising target for the prevention and treatment of I/R injury.

Programmed necrosis (necroptosis), a newly discovered form of cell death, is mediated by receptor-interacting protein 1 (RIP1) and plays a pivotal role after myocardial, renal, and cerebral ischemia-reperfusion (I/R). The relevance of necroptosis in the postischemic liver remains, however, unclear. The aim of this study was to analyze the role of programed necrosis during hepatic I/R. C57BL6 mice were subjected to warm hepatic I/R (90 min/240 min). The animals were pretreated with either the RIP1 inhibitor necrostatin-1 (Nec-1, 3.5 μg kg) or vehicle (Nec-1inactive, 3.5 μg kg) administered systemically before ischemia. Sham-operated animals served as controls (n = 6 each group). The inflammatory response was evaluated by intravital microscopy. The hepatic transaminases alanine aminotransferase/aspartate aminotransferase in plasma as well as the activity of caspase-3 in tissue were determined as markers of hepatocellular injury. Leukocyte recruitment to the liver, sinusoidal perfusion failure, as well as the transaminase activities were strongly increased on I/R as compared with the sham-operated mice. Inhibition of the RIP1-dependent pathway with Nec-1, however, did not attenuate I/R-induced leukocyte migration, perfusion failure, and hepatocellular injury. Western blot analysis showed a baseline RIP1 expression in livers from sham-operated mice, whereas RIP1 expression was not detectable in both Nec-1-treated and vehicle-treated I/R group. Caspase-3 activity was significantly elevated after I/R in both postischemic groups. Our in vivo data show that RIP1-mediated necroptosis is not present in the postischemic liver and that I/R-induced caspase activation is associated with loss of RIP1 expression. Because caspases are able to cleave RIP1, we hypothesize that I/R-triggered caspase activation negatively regulates necroptosis and, thereby, determines apoptosis as a preferred route of cell death after hepatic I/R.

T-cells have been demonstrated to modulate ischemia–reperfusion injury (IRI) in the kidney, lung, liver, and intestine. Whereas most T-cell subpopulations contribute primarily to the antigen-specific effector and memory phases of immunity, γδ-T-cells combine adaptive features with rapid, innate-like responses that can place them in the initiation phase of immune reactions. Therefore, we aimed to clarify the role of γδ-T-cells in intestinal IRI. Adult wild-type (WT) and γδ-T-cell-deficient mice were subjected to acute intestinal IRI. Gene expression of pro-inflammatory cytokines and influx of leukocyte subpopulations in the gut were assessed by qPCR and flow cytometry. Serum transaminases were measured as an indicator of distant organ IRI. Intestinal IRI led to increased influx of neutrophils, pro-inflammatory cytokine expression and LDH/ALT/AST elevation. Selective deficiency of γδ-T-cells significantly decreased pro-inflammatory cytokine levels and neutrophil infiltration in the gut following IRI compared to controls. Furthermore, γδ-T-cell deficiency resulted in decreased LDH and transaminases levels in sera, indicating amelioration of distant organ injury. Increasing evidence demonstrates a key role of T-cell subpopulations in IRI. We demonstrate that γδ-T-cell deficiency ameliorated pro-inflammatory cytokine production, neutrophil recruitment and distant organ injury. Thus, γδ-T-cells may be considered as mediators contributing to the inflammatory response in the acute phase of intestinal IRI.

This study aims to evaluate the long-term efficacy and reintervention rate after primary percutaneous portal vein stent angioplasty for portal vein stenosis (PVS) in pediatric liver transplantation (LT) recipients. From 2004 to 2020, a total of 470 pediatric LTs were performed in our center. All cases were screened for interventional PVS treatment and analyzed retrospectively. We identified 44 patients with 46 percutaneous angioplasties for posttransplantation PVS. The median interval from LT to percutaneous catheter intervention was 5 months (16 days-104 months) with a median follow-up (f/u) period after catheter intervention of 5.7 years (2-156 months). In 40 patients, an endovascular stent was placed as primary (n = 38) or secondary (n = 2) intervention. The median age at stent placement was 23 (6-179) months with a median weight of 10 kg (6-46 kg). Technical success and relief of PVS were achieved in all patients irrespective of age or weight. Adverse events occurred peri-interventionally in two patients and were resolved with standard care. All primary portal vein (PV) stents remained patent until the end of f/u. Reinterventions have been successfully performed in 10 patients for suspected or proven restenosis, resulting in a primary patency rate of 75% and an assisted patency rate of 25%. The median time to reintervention was 6.2 years (range 1-10 years). The need for reintervention was independent of age or weight at both transplantation and initial angioplasty as well as of additional risk factors due to portal hypertension. Percutaneous transhepatic PV stent angioplasty in children is safe and effective in all age groups, with excellent long-term patency. Primary stent angioplasty should be considered as first-line treatment for PVS after pediatric LT.

CD4 ⁺ T cells recruited to the liver play a key role in the pathogenesis of ischemia/reperfusion (I/R) injury. The mechanism of their activation during alloantigen‐independent I/R is not completely understood. We hypothesized that liver‐resident dendritic cells (DCs) interact with CD4 ⁺ T cells in the postischemic liver and that modulation of DCs or T‐cell–DC interactions attenuates liver inflammation. In mice, warm hepatic I/R (90/120–240 min) was induced. Tolerogenic DCs were generated in situ by pretreatment of animals with the vitamin D analog paricalcitol. A mAb‐CD44 was used for blockade of CD4 ⁺ T‐cell–DC interactions. As shown by 2‐photon in vivo microscopy as well as confocal microscopy, CD4 ⁺ T cells were closely colocalized with DCs in the postischemic liver. Pretreatment with paricalcitol attenuated I/R‐induced maturation of DCs (flow cytometry), CD4 ⁺ T‐cell recruitment into the liver (intravital microscopy), and hepatocellular/microvascular damage (intravital microscopy, alanine aminotransferase/aspartate aminotransferase, histology). However, interruption of T‐cell–DC interaction increased proinflammatory DC maturation and even enhanced tissue damage. Simultaneous treatment with an anti‐CD44mAb completely abolished the beneficial effect of paricalcitol on T‐cell migration and tissue injury. Our study demonstrates for the first time that hepatic DCs interact with CD4 ⁺ T cells in the postischemic liver in vivo ; modulation of DCs and/or generation of tolerogenic DCs attenuates intrahepatic CD4 ⁺ T‐cell recruitment and reduces I/R injury; and interruption of CD44‐dependent CD4 ⁺ T‐cell–DC interactions enhances tissue injury by preventing the modulatory effect of hepatic DCs on T cells, especially type 1 T helper effector cells. Thus, hepatic DCs are strongly involved in the promotion of CD4 ⁺ T‐cell–dependent postischemic liver inflammation.—Funken, D., Ishikawa‐Ankerhold, H., Uhl, B., Lerchenberger, M., Rentsch, M., Mayr, D., Massberg, S., Werner, J., Khandoga, A. In situ targeting of dendritic cells sets tolerogenic environment and ameliorates CD4 ⁺ T‐cell response in the postischemic liver. FASEB J. 31, 4796–4808 (2017). www.fasebj.org

Background: The inflammasome NLRP3 (NOD-like receptor family pyrin domain containing 3) is critical for epithelial barrier integrity. Allergic asthma is characterized by airway inflammation, airway hyperresponsiveness (AHR), and mucus hypersecretion. To date, the key players and underlying mechanisms in the interaction between NLRP3 and epithelial barrier integrity in type 2-mediated allergic asthma are poorly understood. Objective: Our study aims to evaluate the protective mechanisms of NLRP3 on airway epithelium and its structural and functional components during type 2-mediated allergic asthma inflammation. Methods: Using an experimental model of allergic airway disease, NLRP3-deficient (NLRP3 ^-/- ) and wild-type (WT) mice were analyzed for AHR, mucus hyperplasia, airway inflammation and the alterations in the airway epithelium transcriptome. Results: In comparison to WT mice, NLRP3 ^-/- mice exhibited significantly enhanced AHR and mucus production, while eosinophilic airway inflammation was comparable. Analysis of epithelial cell markers revealed decreased gene expression of the tight junction proteins Cld-18 and Tjp-1 , and decreased expression of the epithelial transmembrane protein E-cadherin in the lungs of naïve NLRP3 ^-/- mice compared to WT mice. Moreover, intranasal treatment with FITC-labelled OVA resulted in significantly higher allergen uptake by lung conventional dendritic cells (cDCs) in NLRP3 ^-/- compared to WT mice indicating increased epithelial leakiness. In vitro , inhibition of NLRP3 in the human bronchial epithelial cell line 16HBE14o- with MCC950 resulted in the downregulation of Tjp-1 and CDH1 (E-cadherin). Conclusion: NLRP3 is essential for epithelial barrier integrity in the lung and protects from the development of allergic asthma in a murine model.