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Journal of Hepatology Oct 2019Hepatic ischemia-reperfusion injury (IRI) is a major complication of hemorrhagic shock, liver resection and transplantation. YAP, a key downstream effector of the Hippo...
BACKGROUND & AIMS
Hepatic ischemia-reperfusion injury (IRI) is a major complication of hemorrhagic shock, liver resection and transplantation. YAP, a key downstream effector of the Hippo pathway, is essential for determining cell fate and maintaining homeostasis in the liver. We aimed to elucidate its role in IRI.
METHODS
The role of YAP/Hippo signaling was systematically studied in biopsy specimens from 60 patients after orthotopic liver transplantation (OLT), and in a mouse model of liver warm IRI. Human biopsy specimens were collected after 2-10 h of cold storage and 3 h post-reperfusion, before being screened by western blot. In the mouse model, the role of YAP was probed by activating or inhibiting YAP prior to ischemia-reperfusion.
RESULTS
In human biopsies, high post-OLT YAP expression was correlated with well-preserved histology and improved hepatocellular function at postoperative day 1-7. In mice, the ischemia insult (90 min) triggered intrinsic hepatic YAP expression, which peaked at 1-6 h of reperfusion. Activation of YAP protected the liver against IR-stress, by promoting regenerative and anti-oxidative gene induction, while diminishing oxidative stress, necrosis/apoptosis and the innate inflammatory response. Inhibition of YAP aggravated hepatic IRI and suppressed repair/anti-oxidative genes. In mouse hepatocyte cultures, activating YAP prevented hypoxia-reoxygenation induced stress. Interestingly, YAP activation suppressed extracellular matrix synthesis and diminished hepatic stellate cell (HSC) activation, whereas YAP inhibition significantly delayed hepatic repair, potentiated HSC activation, and enhanced liver fibrosis at 7 days post-IRI. Notably, YAP activation failed to protect Nrf2-deficient livers against IR-mediated damage, leading to extensive fibrosis.
CONCLUSION
Our novel findings document the crucial role of YAP in IR-mediated hepatocellular damage and liver fibrogenesis, providing evidence of a potential therapeutic target for the management of sterile liver inflammation in transplant recipients.
LAY SUMMARY
In the clinical arm, graft YAP expression negatively correlated with liver function and tissue damage after human liver transplantation. YAP activation attenuated hepatocellular oxidative stress and diminished the innate immune response in mouse livers following ischemia-reperfusion injury. In the mouse model, YAP inhibited hepatic stellate cell activation, and abolished injury-mediated fibrogenesis up to 7 days after the ischemic insult.
Topics: Animals; Apoptosis; Cell Cycle Proteins; Cells, Cultured; Disease Models, Animal; Hippo Signaling Pathway; Humans; Inflammation; Liver; Liver Diseases; Liver Transplantation; Oxidative Stress; Protein Serine-Threonine Kinases; Reperfusion Injury; Shock, Hemorrhagic; Signal Transduction; Transcription Factors; Warm Ischemia
PubMed: 31201834
DOI: 10.1016/j.jhep.2019.05.029 -
Nature Aug 2022After cessation of blood flow or similar ischaemic exposures, deleterious molecular cascades commence in mammalian cells, eventually leading to their death. Yet with...
After cessation of blood flow or similar ischaemic exposures, deleterious molecular cascades commence in mammalian cells, eventually leading to their death. Yet with targeted interventions, these processes can be mitigated or reversed, even minutes or hours post mortem, as also reported in the isolated porcine brain using BrainEx technology. To date, translating single-organ interventions to intact, whole-body applications remains hampered by circulatory and multisystem physiological challenges. Here we describe OrganEx, an adaptation of the BrainEx extracorporeal pulsatile-perfusion system and cytoprotective perfusate for porcine whole-body settings. After 1 h of warm ischaemia, OrganEx application preserved tissue integrity, decreased cell death and restored selected molecular and cellular processes across multiple vital organs. Commensurately, single-nucleus transcriptomic analysis revealed organ- and cell-type-specific gene expression patterns that are reflective of specific molecular and cellular repair processes. Our analysis comprises a comprehensive resource of cell-type-specific changes during defined ischaemic intervals and perfusion interventions spanning multiple organs, and it reveals an underappreciated potential for cellular recovery after prolonged whole-body warm ischaemia in a large mammal.
Topics: Animals; Cell Death; Cell Survival; Cytoprotection; Gene Expression Profiling; Ischemia; Organ Specificity; Perfusion; Swine; Warm Ischemia
PubMed: 35922506
DOI: 10.1038/s41586-022-05016-1 -
Sulforaphane Is Protective against Warm Ischemia/Reperfusion Injury and Partial Hepatectomy in Rats.International Journal of Molecular... Jan 2024Sulforaphane (SFN) has various beneficial effects on organ metabolism. However, whether SFN affects inflammatory mediators induced by warm hepatic ischemia/reperfusion...
Sulforaphane (SFN) has various beneficial effects on organ metabolism. However, whether SFN affects inflammatory mediators induced by warm hepatic ischemia/reperfusion injury (HIRI) is unclear. To investigate the hepatoprotective effects of SFN using an in vivo model of HIRI and partial hepatectomy (HIRI + PH), rats were subjected to 15 min of hepatic ischemia with blood inflow occlusion, followed by 70% hepatectomy and release of the inflow occlusion. SFN (5 mg/kg) or saline was randomly injected intraperitoneally 1 and 24 h before ischemia. Alternatively, ischemia was prolonged for 30 min to evaluate the effect on mortality. The influence of SFN on the associated signaling pathways was analyzed using the interleukin 1β (IL-1β)-treated primary cultured rat hepatocytes. In the HIRI + PH-treated rats, SFN reduced serum liver enzyme activities and the frequency of pathological liver injury, such as apoptosis and neutrophil infiltration. SFN suppressed tumor necrosis factor-alpha (TNF-α) mRNA expression and inhibited nuclear factor-kappa B (NF-κB) activation by HIRI + PH. Mortality was significantly reduced by SFN. In IL-1β-treated hepatocytes, SFN suppressed the expression of inflammatory cytokines and NF-κB activation. Taken together, SFN may have hepatoprotective effects in HIRI + PH in part by inhibiting the induction of inflammatory mediators, such as TNF-α, via the suppression of NF-κB in hepatocytes.
Topics: Animals; Rats; Hepatectomy; NF-kappa B; Tumor Necrosis Factor-alpha; Warm Ischemia; Reperfusion Injury; Inflammation Mediators; Interleukin-1beta; Ischemia; Sulfoxides; Isothiocyanates
PubMed: 38203749
DOI: 10.3390/ijms25010579 -
Transplantation Reviews (Orlando, Fla.) Oct 2019Ischemia has been a persistent and largely unavoidable element in solid organ transplantation, contributing to graft deterioration and adverse post-transplant outcomes.... (Review)
Review
Ischemia has been a persistent and largely unavoidable element in solid organ transplantation, contributing to graft deterioration and adverse post-transplant outcomes. In liver transplantation, where available organs arise with greater frequency from marginal donors (i.e., ones that are older, obese, and/or declared dead following cardiac arrest through the donation after circulatory death process), there is increasing interest using dynamic perfusion strategies to limit, assess, and even reverse the adverse effects of ischemia in these grafts. Normothermic perfusion, in particular, is used to restore the flow of oxygen and other metabolic substrates at physiological temperatures. It may be used in liver transplantation both in situ following cardiac arrest in donation after circulatory death donors or during part or all of the ex situ preservation phase. This review article addresses issues relevant to use of normothermic perfusion strategies in liver transplantation, including technical and logistical aspects associated with establishing and maintaining normothermic perfusion in its different forms and clinical outcomes that have been reported to date.
Topics: Female; Graft Rejection; Graft Survival; Humans; Liver Transplantation; Male; Organ Preservation; Perfusion; Sensitivity and Specificity; Tissue and Organ Procurement; Warm Ischemia
PubMed: 31239189
DOI: 10.1016/j.trre.2019.06.001 -
Journal of Visualized Experiments : JoVE Jun 2023Porcine models of liver ex situ normothermic machine perfusion (NMP) are increasingly being used in transplant research. Contrary to rodents, porcine livers are...
Porcine models of liver ex situ normothermic machine perfusion (NMP) are increasingly being used in transplant research. Contrary to rodents, porcine livers are anatomically and physiologically close to humans, with similar organ size and bile composition. NMP preserves the liver graft at near-to-physiological conditions by recirculating a warm, oxygenated, and nutrient-enriched red blood cell-based perfusate through the liver vasculature. NMP can be used to study ischemia-reperfusion injury, preserve a liver ex situ before transplantation, assess the liver's function prior to implantation, and provide a platform for organ repair and regeneration. Alternatively, NMP with a whole blood-based perfusate can be used to mimic transplantation. Nevertheless, this model is labor-intensive, technically challenging, and carries a high financial cost. In this porcine NMP model, we use warm ischemic damaged livers (corresponding to donation after circulatory death). First, general anesthesia with mechanical ventilation is initiated, followed by the induction of warm ischemia by clamping the thoracic aorta for 60 min. Cannulas inserted in the abdominal aorta and portal vein allow flush-out of the liver with cold preservation solution. The flushed-out blood is washed with a cell saver to obtain concentrated red blood cells. Following hepatectomy, cannulas are inserted in the portal vein, hepatic artery, and infra-hepatic vena cava and connected to a closed perfusion circuit primed with a plasma expander and red blood cells. A hollow fiber oxygenator is included in the circuit and coupled to a heat exchanger to maintain a pO2 of 70-100 mmHg at 38 °C. NMP is achieved by a continuous flow directly through the artery and via a venous reservoir through the portal vein. Flows, pressures, and blood gas values are continuously monitored. To evaluate the liver injury, perfusate and tissue are sampled at predefined time points; bile is collected via a cannula in the common bile duct.
Topics: Humans; Swine; Animals; Organ Preservation; Liver; Liver Transplantation; Perfusion; Warm Ischemia
PubMed: 37358290
DOI: 10.3791/65336 -
Experimental and Clinical... Feb 2020
Topics: Anastomosis, Surgical; Delayed Graft Function; Graft Survival; Humans; Kidney Transplantation; Treatment Outcome; Warm Ischemia
PubMed: 32170860
DOI: 10.6002/ect.2019.0317 -
Experimental and Clinical... Mar 2021Chronic kidney disease is the most common type of organ failure worldwide, with a prevalence of 13.4% for all stages. Organ transplant is the only curative option for... (Review)
Review
Chronic kidney disease is the most common type of organ failure worldwide, with a prevalence of 13.4% for all stages. Organ transplant is the only curative option for end-stage kidney failure. However, the shortage of organ donors remains a major obstacle in organ transplant, with donation after circulatory death being the most viable path to increasing the donor pool. The circumstances that surround this type of donation are different from donation after brain death, namely concerning warm ischemia times, which are longer and may preclude a successful transplant. This article describes the pathophysiology of warm ischemia and summarizes recent developments in technological and methodological practices that mitigate the mechanisms of warm ischemia. Anoxia, mitochondrial dysfunction, calcium overload, oxidative and nitrosative stress, immune response, and no reflow are the main mechanisms by which ischemia leads to cell death and organ dysfunction. In situ oxygenated recirculation, abdominal normothermic organ recirculation, abdominal hypothermic organ recirculation, and ex vivo machine perfusion ensure continued organ perfusion and prevent prolonged warm ischemia in organ donation. These practices, coupled with optimizations in the identification and assessment of potential donors after circulatory death, may lead to a significant increase in the number and success rates of organ transplant worldwide.
Topics: Death; Humans; Tissue Donors; Warm Ischemia
PubMed: 32799784
DOI: 10.6002/ect.2020.0081 -
The Urologic Clinics of North America May 2017Renal function after renal cancer surgery is a critical component of survivorship. Quantity and quality of preserved parenchyma are the most important determinants of... (Review)
Review
Renal function after renal cancer surgery is a critical component of survivorship. Quantity and quality of preserved parenchyma are the most important determinants of functional recovery; type and duration of ischemia play secondary roles. Several studies evaluated surgical techniques to minimize ischemia; however, long-term outcomes and potential benefits over clamped partial nephrectomy (PN) have not been consistently demonstrated. Analysis of acute kidney injury (AKI) after PN suggest that most kidneys recover strongly even if AKI is experienced after surgery. Ongoing study is required to evaluate long-term implications of AKI after PN and further assess impact of ischemia on functional outcomes.
Topics: Acute Kidney Injury; Cold Ischemia; Humans; Kidney Neoplasms; Nephrectomy; Postoperative Complications; Treatment Outcome; Warm Ischemia
PubMed: 28411916
DOI: 10.1016/j.ucl.2016.12.010 -
Urology Aug 2015
Topics: Female; Humans; Kidney; Kidney Neoplasms; Male; Nephrectomy; Warm Ischemia
PubMed: 26199169
DOI: 10.1016/j.urology.2015.04.045 -
Biopreservation and Biobanking Aug 2021High-quality RNA extraction from tissue samples is of key importance for scientific research and translational medicine. Tissue collection and preparation may affect...
High-quality RNA extraction from tissue samples is of key importance for scientific research and translational medicine. Tissue collection and preparation may affect RNA quality. In this study, we investigated effects of warm ischemia time, cryopreservation, and grinding methods on RNA quality. Total RNA was extracted from mouse kidney tissues with warm ischemia times of 0, 30, 60, 90, and 120 minutes. Half of the tissues were used to extract RNA immediately, while the others were cryopreserved in the vapor phase of liquid nitrogen for 6 months before RNA extraction. A mortar, homogenizer, and tissue lyser were used to grind tissues. RNA was extracted by TRIzol, and RNA integrity was assessed by the RNA integrity number (RIN) value. For fresh tissues and frozen tissues with warm ischemia time within 60 minutes, RIN values were above 7.0 and remained above 6.0 with warm ischemia time within 120 minutes. For the same warm ischemia time, RIN values of frozen tissues were slightly lower than those of fresh tissues. No significant RIN value alterations were observed among grinding methods, but for RNA extraction efficiency, a mortar was much less efficient than the homogenizer or tissue lyser. For frozen tissues, RNA tended to degrade within 8 minutes at room temperature. Mouse kidney tissues with a warm ischemia time within 120 minutes are suitable for general RNA-related research. For tissues with a warm ischemia time within 60 minutes, cryopreservation may not affect RNA quality. The duration of frozen tissues held at room temperature before grinding affects the integrity of RNA, while grinding methods do not affect RNA integrity.
Topics: Animals; Cryopreservation; Kidney; Mice; RNA; Tissue Banks; Warm Ischemia
PubMed: 33577406
DOI: 10.1089/bio.2020.0129