Liver protects neuron viability and electrocortical activity in post-cardiac arrest brain injury.

Brain injury is the leading cause of mortality among patients who survive cardiac arrest (CA). Clinical studies have shown that the presence of post-CA hypoxic hepatitis or pre-CA liver disease is associated with increased mortality and inferior neurological recovery. In our in vivo global cerebral ischemia model, we observed a larger infarct area, elevated tissue injury scores, and increased intravascular CD45+ cell adhesion in reperfused brains with simultaneous hepatic ischemia than in those without it. In the ex vivo brain normothermic machine perfusion (NMP) model, we demonstrated that addition of a functioning liver to the brain NMP circuit significantly reduced post-CA brain injury, increased neuronal viability, and improved electrocortical activity. Furthermore, significant alterations were observed in both the transcriptome and metabolome in the presence or absence of hepatic ischemia. Our study highlights the crucial role of the liver in the pathogenesis of post-CA brain injury.

Metabolomics Differences of the Donor Livers Between In Situ and Ex Situ Conditions During Ischemia-free Liver Transplantation.

BACKGROUND: Ischemia-free liver transplantation (IFLT) has been innovated to avoid graft ischemia during organ procurement, preservation, and implantation. However, the metabolism activity of the donor livers between in the in situ and ex situ normothermic machine perfusion (NMP) conditions, and between standard criteria donor and extend criteria donor remains unknown. METHODS: During IFLT, plasma samples were collected both at the portal vein and hepatic vein of the donor livers in situ during procurement and ex situ during NMP. An ultra-high performance liquid chromatography-mass spectrometry was conducted to investigate the common and distinct intraliver metabolite exchange. RESULTS: Profound cysteine and methionine metabolism, and aminoacyl-tRNA biosynthesis were found in both in situ and ex situ conditions. However, obvious D-arginine and D-ornithine metabolism, arginine and proline metabolism were only found in the in situ condition. The suppressed activities of the urea cycle pathway during ex situ condition were confirmed in an RNA expression level. In addition, compared with extend criteria donor group, standard criteria donor group had more active intraliver metabolite exchange in metabonomics level. Furthermore, we found that the relative concentration of p-cresol, allocystathionine, L-prolyl-L-proline in the ex situ group was strongly correlated with peak alanine aminotransferase and aspartate aminotransferase at postoperative days 1-7. CONCLUSIONS: In the current study, we show the common and distinct metabolism activities during IFLT. These findings might provide insights on how to modify the design of NMP device, improve the perfusate components, and redefine the criteria of graft viability.

Abrogation of graft ischemia-reperfusion injury in ischemia-free liver transplantation.

BACKGROUND: Ischemia-reperfusion injury (IRI) is considered an inherent component of organ transplantation that compromises transplant outcomes and organ availability. The ischemia-free liver transplantation (IFLT) procedure has been developed to avoid interruption of blood supply to liver grafts. It is unknown how IFLT might change the characteristics of graft IRI. METHODS: Serum and liver biopsy samples were collected from IFLT and conventional liver transplantation (CLT) recipients. Pathological, metabolomics, transcriptomics, and proteomics analyses were performed to identify the characteristic changes in graft IRI in IFLT. RESULTS: Peak aspartate aminotransferase (539.59 ± 661.76 U/L versus 2622.28 ± 3291.57 U/L) and alanine aminotransferase (297.64 ± 549.50 U/L versus 1184.16 ± 1502.76 U/L) levels within the first 7 days and total bilirubin levels by day 7 (3.27 ± 2.82 mg/dl versus 8.33 ± 8.76 mg/dl) were lower in the IFLT versus CLT group (all p values < 0.001). The pathological characteristics of IRI were more obvious in CLT grafts. The antioxidant pentose phosphate pathway remained active throughout the procedure in IFLT grafts and was suppressed during preservation and overactivated postrevascularization in CLT grafts. Gene transcriptional reprogramming was almost absent during IFLT but was profound during CLT. Proteomics analysis showed that "metabolism of RNA" was the major differentially expressed process between the two groups. Several proinflammatory pathways were not activated post-IFLT as they were post-CLT. The activities of natural killer cells, macrophages, and neutrophils were lower in IFLT grafts than in CLT grafts. The serum levels of 14 cytokines were increased in CLT versus IFLT recipients. CONCLUSIONS: IFLT can largely avoid the biological consequences of graft IRI, thus has the potential to improve transplant outcome while increasing organ utilization.

Transplantation of a beating heart: A first in man.

Background: In the current practice, graft ischaemia and reperfusion injury (IRI) is considered an inevitable component in organ transplantation, contributes to compromised organ quality, inferior graft survival and limitations in organ availability. Among all the donor organs, the heart is most vulnerable to IRI and the tolerated ischaemic time is the shortest. Methods: By combining adapted surgical techniques and normothermic machine perfusion (NMP), we performed the first case of ischaemia-free beating heart transplantation (IFBHT) in man. The donor heart was procured after an in situ NMP circuit was established, then underwent ex situ NMP and implanted under NMP support. The post-transplant graft function was monitored. Findings: The donor heart was procured, preserved, and implanted under a continuously perfused, normothermic, oxygenated, beating state. During ex situ NMP, the donor heart beat with sinus rhythm and adequate ventricular contraction, consumed oxygen and lactate, suggesting a good cardiac function. The dynamic electrocardiogram demonstrated an absence of ischaemic injury of the donor heart during the entire procedure. The echocardiogram showed an immediate graft function with a left ventricle ejection fraction (LVEF) of 70%. The patient was discharged on post-transplantation day 20 and was followed up for 8 months with normal cardiac function and life. Interpretation: This study shows the feasibility of IFBHT procedure, which might be able to completely avoid graft IRI, has thus the potential to improve transplant outcome while increasing organ utilization. Funding: This study was funded by National Natural Science Foundation of China, Guangdong Provincial Key Laboratory Construction Projection on Organ Donation and Transplant Immunology, and Guangdong Provincial International Cooperation Base of Science and Technology.

Resolving the graft ischemia-reperfusion injury during liver transplantation at the single cell resolution.

Ischemia-reperfusion injury (IRI) remains the major reason for impaired donor graft function and increased mortality post-liver transplantation. The mechanism of IRI involves multiple pathophysiological processes and numerous types of cells. However, a systematic and comprehensive single-cell transcriptional profile of intrahepatic cells during liver transplantation is still unclear. We performed a single-cell transcriptome analysis of 14,313 cells from liver tissues collected from pre-procurement, at the end of preservation and 2 h post-reperfusion. We made detailed annotations of mononuclear phagocyte, endothelial cell, NK/T, B and plasma cell clusters, and we described the dynamic changes of the transcriptome of these clusters during IRI and the interaction between mononuclear phagocyte clusters and other cell clusters. In addition, we found that TNFAIP3 interacting protein 3 (TNIP3), specifically and highly expressed in Kupffer cell clusters post-reperfusion, may have a protective effect on IRI. In summary, our study provides the first dynamic transcriptome map of intrahepatic cell clusters during liver transplantation at single-cell resolution.

Mild hypothermia improves neurological outcome in mice after cardiopulmonary resuscitation through Silent Information Regulator 1-actviated autophagy.

Mild hypothermia treatment (MHT) improves the neurological function of cardiac arrest (CA) patients, but the exact mechanisms of recovery remain unclear. Herein, we generated a CA and cardiopulmonary resuscitation (CPR) mouse model to elucidate such function. Naïve mice were randomly divided into two groups, a normothemia (NT) group, in which animals had normal body temperature, and a MHT group, in which animals had a body temperature of 33 °C (range: 32-34 °C), after the return of spontaneous circulation (ROSC), followed by CA/CPR. MHT significantly improved the survival rate of CA/CPR mice compared with NT. Mechanistically, MHT increased the expression of Silent Information Regulator 1 (Sirt1) and decreased P53 phosphorylation (p-P53) in the cortex of CA/CPR mice, which coincided with the elevated autophagic flux. However, Sirt1 deletion compromised the neuroprotection offered by MHT, indicating that Sirt1 plays an important role. Consistent with the observations obtained from in vivo work, our in vitro study utilizing cultured neurons subjected to oxygen/glucose deprivation and reperfusion (OGD/R) also indicated that Sirt1 knockdown increased OGD/R-induced neuron necrosis and apoptosis, which was accompanied by decreased autophagic flux and increased p-P53. However, the depletion of P53 did not suppress neuron death, suggesting that P53 was not critically involved in MHT-induced neuroprotection. In contrast, the application of autophagic inhibitor 3-methyladenine attenuated MHT-improved neuron survival after OGD/R, further demonstrating that increased autophagic flux significantly contributes to MHT-linked neuroprotection of CA/CRP mice. Our findings indicate that MHT improves neurological outcome of mice after CA/CPR through Sirt1-mediated activation of autophagic flux.

Expression profiles of long noncoding RNAs and mRNAs in post-cardiac arrest rat brains.

To investigate long noncoding (lnc)-RNA and mRNA expression profiles in post­cardiac arrest (CA) brains, an external transthoracic electrical current was applied for 8 min to induce CA (the CA group). A total of 4 rats received sham-operations and served as the blank control (BC) group. Upon return of spontaneous circulation (ROSC), lncRNA and mRNA expression in the rat cerebral cortex was assayed with high­throughput Agilent lncRNA and mRNA microarrays. In total, 37 lncRNAs were upregulated and 21 lncRNAs were downregulated in the CA group, and 258 mRNA transcripts were differentially expressed with 177 mRNAs upregulated and 81 mRNAs downregulated in the CA group. The differentially expressed lncRNAs in the CA group were co­expressed with thousands of mRNAs. The differentially expressed lncRNAs could be clustered into >100 signaling pathways and processes according to Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes analyses. The most common predicted functions involved metabolic pathways, protein synthesis, transport and degradation during CA­ROSC. CA­ROSC led to significant alterations in cerebral lncRNA and mRNA expression profiles. Thus, lncRNA­mRNA network interactions have the potential to regulate vital metabolic pathways and processes involved in CA-ROSC.