Programmed cell death, from liver Ischemia-Reperfusion injury perspective: An overview.

Liver ischemia-reperfusion injury (LIRI) commonly occurs in liver resection, liver transplantation, shock, and other hemorrhagic conditions, resulting in profound local and systemic effects via associated inflammatory responses and hepatic cell death. Hepatocyte death is a significant component of LIRI and its mechanism was previously thought to be limited to apoptosis and necrosis. With the discovery of novel types of programmed cell death (PCD), necroptosis, ferroptosis, pyroptosis, autophagy, NETosis, and parthanatos have been shown to be involved in LIRI. Understanding the mechanisms underlying cell death following LIRI is indispensable to mitigating the widespread effects of LIRI. Here, we review the roles of different PCD and discuss potential therapy in LIRI.

Diagnosis and Molecular Characterization of Potential RNA Binding Protein Involved in the Pathogenesis of Liver Ischemia Reperfusion Injury.

Background: Liver ischemia-reperfusion is one of the common complications after liver surgery. Uncontrolled liver ischemia-reperfusion will lead to many serious consequences such as surgical failure. It is an urgent clinical problem to search for diagnostic markers and explore its potential pathogenesis. Methods: In this study, we focus on 1411 candidate RNA binding protein. Through several GEO (Gene Expression Omnibus) online datasets, we construct a diagnostic model and perform interactive validation. We evaluate the efficacy of the prognostic model. Using bioinformatics methods, we predicted the relevant signaling pathways of liver ischemia-reperfusion and key genes. We also evaluated the association of RNA binding protein with immune cell infiltration. Single cell sequencing datasets were used to explore the expression profiles of key genes at the single cell level. Machine learning algorithm is used to predict key gene RNA binding domains. Results: ROC (Receiver Operating Characteristic) and DCA (Decision Curve Analysis) curves showed that the above diagnostic model had good and stable diagnostic efficacy and clinical practicability. We identified three key genes (BTG2, CCNL1 and DNAJB1) in liver ischemia-reperfusion. DNAJB1, BTG2 and CCNL1 are mainly expressed in immune cells such as macrophages and T cells, and are closely related to inflammatory pathways such as TNF-α, highlighting their importance in hepatic ischemia reperfusion. We identified RNA-binding domains of the above three genes. We found that the expression of DNAJB1, CCNL1 and BTG2 in the ischemia-reperfusion group were significantly higher than those in the sham operation group. Conclusion: Our study revealed the importance of the candidate RNA binding protein in liver ischemia reperfusion injury and provided new insights into the therapeutic of hepatic ischemia-reperfusion injury.

A comprehensive analysis of m6A/m7G/m5C/m1A-related gene expression and immune infiltration in liver ischemia-reperfusion injury by integrating bioinformatics and machine learning algorithms.

BACKGROUND: Liver ischemia-reperfusion injury (LIRI) is closely associated with immune infiltration, which commonly occurs after liver surgery, especially liver transplantation. Therefore, it is crucial to identify the genes responsible for LIRI and develop effective therapeutic strategies that target immune response. Methylation modifications in mRNA play various crucial roles in different diseases. This study aimed to identify potential methylation-related markers in patients with LIRI and evaluate the corresponding immune infiltration. METHODS: Two Gene Expression Omnibus datasets containing human liver transplantation data (GSE12720 and GSE151648) were downloaded for integrated analysis. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were conducted to investigate the functional enrichment of differentially expressed genes (DEGs). Differentially expressed methylation-related genes (DEMRGs) were identified by overlapping DEG sets and 65 genes related to N6-methyladenosine (m6A), 7-methylguanine (m7G), 5-methylcytosine (m5C), and N1-methyladenosine (m1A). To evaluate the relationship between DEMRGs, a protein-protein interaction (PPI) network was utilized. The core DEMRGs were screened using three machine learning algorithms: least absolute shrinkage and selection operator, random forest, and support vector machine-recursive feature elimination. After verifying the diagnostic efficacy using the receiver operating characteristic curve, we validated the expression of the core DEMRGs in clinical samples and performed relative cell biology experiments. Additionally, the immune status of LIRI was comprehensively assessed using the single sample gene set enrichment analysis algorithm. The upstream microRNA and transcription factors of the core DEMRGs were also predicted. RESULTS: In total, 2165 upregulated and 3191 downregulated DEGs were identified, mainly enriched in LIRI-related pathways. The intersection of DEGs and methylation-related genes yielded 28 DEMRGs, showing high interaction in the PPI network. Additionally, the core DEMRGs YTHDC1, METTL3, WTAP, and NUDT3 demonstrated satisfactory diagnostic efficacy and significant differential expression and corresponding function based on cell biology experiments. Furthermore, immune infiltration analyses indicated that several immune cells correlated with all core DEMRGs in the LIRI process to varying extents. CONCLUSIONS: We identified core DEMRGs (YTHDC1, METTL3, WTAP, and NUDT3) associated with immune infiltration in LIRI through bioinformatics and validated them experimentally. This study may provide potential methylation-related gene targets for LIRI immunotherapy.

Nepetoidin B Alleviates Liver Ischemia/Reperfusion Injury via Regulating MKP5 and JNK/P38 Pathway.

Background: Nepetoidin B (NB) has been reported to possess anti-inflammatory, antibacterial, and antioxidant properties. However, its effects on liver ischemia/reperfusion (I/R) injury remain unclear. Methods: In this study, a mouse liver I/R injury model and a mouse AML12 cell hypoxia reoxygenation (H/R) injury model were used to investigate the potential role of NB. Serum transaminase levels, liver necrotic area, cell viability, oxidative stress, inflammatory response, and apoptosis were evaluated to assess the effects of NB on liver I/R and cell H/R injury. Quantitative polymerase chain reaction (qPCR) and Western blotting were used to measure mRNA and protein expression levels, respectively. Molecular docking was used to predict the binding capacity of NB and mitogen-activated protein kinase phosphatase 5 (MKP5). Results: The results showed that NB significantly reduced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, liver necrosis, oxidative stress, reactive oxygen species (ROS) content, inflammatory cytokine content and expression, inflammatory cell infiltration, and apoptosis after liver I/R and AML12 cells H/R injury. Additionally, NB inhibited the JUN protein amino-terminal kinase (JNK)/P38 pathway. Molecular docking results showed good binding between NB and MKP5 proteins, and Western blotting results showed that NB increased the protein expression of MKP5. MKP5 knockout (KO) significantly diminished the protective effects of NB against liver injury and its inhibitory effects on the JNK/P38 pathway. Conclusion: NB exerts hepatoprotective effects against liver I/R injury by regulating the MKP5-mediated P38/JNK signaling pathway.

tBHQ mitigates fatty liver ischemia-reperfusion injury by activating Nrf2 to attenuate hepatocyte mitochondrial damage and macrophage STING activation.

BACKGROUND: Liver ischemia-reperfusion (IR) injury is an inevitable pathophysiological process in various liver surgeries. Previous studies have found that IR injury is exacerbated in fatty liver due to significant hepatocellular damage and macrophage inflammatory activation, though the underlying mechanisms are not fully understood. In this study, we aim to explore the role and mechanism of Nrf2 (Nuclear factor erythroid 2-related factor 2) signaling in regulating hepatocellular damage and macrophage immune response in fatty liver IR injury. METHODS: The study used high-fat diet-induced fatty liver mice to establish an IR model, alongside an in vitro co-culture system of primary hepatocytes and macrophages. This approach was used to examine mitochondrial dysfunction, oxidative stress, mitochondrial DNA (mtDNA) release, and activation of macrophage STING (Stimulator of interferon genes) signaling. We also conducted recovery verification using H-151 (a STING inhibitor) and tBHQ (an Nrf2 activator). RESULTS: Compared to the control group, mice on a high-fat diet demonstrated more severe liver IR injury, as evidenced by increased histological damage, elevated liver enzyme levels, and heightened inflammatory markers. The HFD group showed significant oxidative stress and mitochondrial dysfunction and damage post-IR, as indicated by elevated levels of ROS and lipid peroxidation markers, and decreased antioxidant enzyme activity. Elevated mtDNA release from hepatocytes post-IR activated macrophage STING signaling, worsening inflammation and liver damage. However, STING signaling inhibition with H-151 in vivo or employing STING knockout macrophages significantly reduced these injuries. In-depth mechanism studies have found that the transfer of Nrf2 protein into the nucleus of liver cells after IR in fatty liver is reduced. Pre-treatment with tBHQ ameliorated liver oxidative stress, mitochondrial damage and suppressed the macrophage STING signaling activation. CONCLUSIONS: Our study reveals a novel mechanism where the interaction between hepatocellular damage and macrophage inflammation intensifies liver IR injury in fatty liver. Enhancing Nrf2 activation to protect mitochondrial from oxidative stress damage and inhibiting macrophage STING signaling activation emerge as promising strategies for clinical intervention in fatty liver IR injury.

Novel anti-inflammatory peptide alleviates liver ischemia-reperfusion injury.

Ischemia-reperfusion injury (IRI) remains inevitable in liver surgeries, macrophages play a critical role in the development of IRI, but little is known about the macrophages regulate pathogenesis of IRI. Based on target-guided screening, we identified a small 3 kDa peptide (SjDX5-271) from various schistosome egg-derived peptides that induced M2 macrophage polarization. SjDX5-271 treatment protected the mice against liver IRI through promoting M2 macrophage polarization, the protective effect was abrogated when the macrophages were depleted. Transcriptomic sequencing showed that the TLR signaling pathway was significantly inhibited in macrophages derived from the SjDX5-271 treatment group. We further identified that SjDX5-271 promotes M2 macrophage polarization by inhibiting the TLR4/MyD88/NF-κB signaling pathway and further alleviates hepatic inflammation in liver IRI. Collectively, SjDX5-271 exhibits promising therapeutic effects in IRI and represents a novel therapeutic approach for IRI, even in immune-related diseases. This study revealed the development of a new biologic from the parasite and enhanced our understanding of host-parasite interplay, providing a blueprint for future therapies for immune-related diseases.

Oridonin attenuates liver ischemia-reperfusion injury by suppressing PKM2/NLRP3-mediated macrophage pyroptosis.

BACKGROUND: The NOD-like receptor protein 3 (NLRP3) mediated pyroptosis of macrophages is closely associated with liver ischemia reperfusion injury (IRI). As a covalent inhibitor of NLRP3, Oridonin (Ori), has strong anti-inflammasome effect, but its effect and mechanisms for liver IRI are still unknown. METHODS: Mice and liver macrophages were treated with Ori, respectively. Co-IP and LC-MS/MS analysis of the interaction between PKM2 and NLRP3 in macrophages. Liver damage was detected using H&E staining. Pyroptosis was detected by WB, TEM, and ELISA. RESULTS: Ori ameliorated liver macrophage pyroptosis and liver IRI. Mechanistically, Ori inhibited the interaction between pyruvate kinase M2 isoform (PKM2) and NLRP3 in hypoxia/reoxygenation（H/R）-induced macrophages, while the inhibition of PKM2/NLRP3 reduced liver macrophage pyroptosis and liver IRI. CONCLUSION: Ori exerted protective effects on liver IRI via suppressing PKM2/NLRP3-mediated liver macrophage pyroptosis, which might become a potential therapeutic target in the clinic.

Acute Liver Ischemia Caused by Vasculitis Due to the Use of Synthetic Cannabinoids.

Teaching Point: Synthetic cannabinoids are drugs whose use has increased significantly in recent years and whose toxicological effects cannot be ignored. Chronic inflammatory processes such as vasculitis that may be caused by these substances pose serious health problems at all ages.

Impact of prolonged liver ischemia during intermittent Pringle maneuver on postoperative outcomes following liver resection.

BACKGROUND: The aim of this study was to compare postoperative outcomes following liver resection between patients with prolonged cumulative ischemia time (CIT) which exceeded 60 min and patients with CIT less than 60 min. METHODS: Between March 2020 and October 2022, 164 consecutive patients underwent liver resection at the Unit for hepato-bilio-pancreatic surgery, University Clinic for Digestive Surgery in Belgrade, Serbia. Intermittent Pringle maneuver was routinely applied. Depending on CIT during transection, patients were divided into two groups: group 1 (CIT <60 min) included 101 patients, and group 2 (CIT ≥60 min) included 63 patients. RESULTS: Median operative time (210 vs. 400 min) and CIT (30 vs. 76 min) were longer in the Group 2 (p < 0.001). Intraoperative blood loss was higher in the Group 2 (150 vs 500 ml), p < 0.001. The perioperative transfusion rate was similar between the groups (p = 0.107). There was no difference in postoperative overall morbidity (50.5% vs. 58.7%, p = 0.337) and major morbidity (18.8 vs. 19%, p = 0.401). In-hospital mortality, 30-day mortality, and 90-day mortality were similar between the groups (p = 0.408; p = 0.408; p = 0.252, respectively). Non-anatomical liver resection was the only predictive factor of 90-day mortality identified by multivariate analysis (p = 0.047; Relative Risk = 0.179; 95% Confidence Interval 0.033-0.981). CONCLUSION: Intermittent Pringle maneuver with CIT exceeding 60 min is a safe method for bleeding control during liver resection with no impact on postoperative morbidity and mortality.

Sirtuin 5 Alleviates Liver Ischemia/Reperfusion Injury by Regulating Mitochondrial Succinylation and Oxidative Stress.

Aims: Mitochondrial dysfunction is the primary mechanism of liver ischemia/reperfusion (I/R) injury. The lysine desuccinylase sirtuin 5 (SIRT5) is a global regulator of the mitochondrial succinylome and has pivotal roles in mitochondrial metabolism and function; however, its hepatoprotective capacity in liver I/R remains unclear. In this study, we established liver I/R model in SIRT5-silenced and SIRT5-overexpressed mice to examine the role and precise mechanisms of SIRT5 in liver I/R injury. Results: Succinylation was strongly enriched in liver mitochondria during I/R, and inhibiting mitochondrial succinylation significantly attenuated liver I/R injury. Importantly, the levels of the desuccinylase SIRT5 were notably decreased in liver transplant patients, as well as in mice subjected to I/R and in AML12 cells exposed to hypoxia/reoxygenation. Furthermore, SIRT5 significantly ameliorated liver I/R-induced oxidative injury, apoptosis, and inflammation by regulating mitochondrial oxidative stress and function. Intriguingly, the hepatoprotective effect of SIRT5 was mediated by PRDX3. Mechanistically, SIRT5 specifically desuccinylated PRDX3 at the K84 site, which enabled PRDX3 to alleviate mitochondrial oxidative stress during liver I/R. Innovation: This study denoted the new effect and mechanism of SIRT5 in regulating mitochondrial oxidative stress through lysine desuccinylation, thus preventing liver I/R injury. Conclusions: Our findings demonstrate for the first time that SIRT5 is a key mediator of liver I/R that regulates mitochondrial oxidative stress through the desuccinylation of PRDX3, which provides a novel strategy to prevent liver I/R injury. Antioxid. Redox Signal. 40, 616-631.

Adelmidrol ameliorates liver ischemia-reperfusion injury through activating Nrf2 signaling pathway.

Liver ischemia/reperfusion (I/R) injury commonly occurs after various liver surgeries. Adelmidrol, an N- palmitoylethanolamide analog, has anti-inflammatory, anti-oxidant, and anti-injury properties. To investigate whether adelmidrol could reduce liver I/R injury, we established a mouse of liver I/R injury and an AML12 cell hypoxia-reoxygenation model to perform experiments using multiple indicators. Serum ALT and AST levels, and H&E staining were used to measure liver damage; MDA content, superoxide dismutase and glutathione activities, and dihydroethidium staining were used to measure oxidative stress; mRNA expression levels of tumor necrosis factor-α, interleukin (IL)-1ß, IL-6, MCP-1, and Ly6G staining were used to measure inflammatory response; and protein expression of Bax, Bcl-2, C-caspase3, and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining were used to measure apoptosis. The experimental results showed that adelmidrol reduced liver I/R injury. In addition, adelmidrol pretreatment elevated AML12 cell activity and reduced I/R-and H/R-induced apoptosis, inflammatory injury, and oxidative stress. ML385, an inhibitor of nuclear factor erythroid2-related factor 2 (Nrf2), reverses liver I/R injury attenuated by adelmidrol. These results suggest that adelmidrol ameliorates liver I/R injury by activating the Nrf2 signaling pathway.

Remimazolam relieved the injury of hypoxia/reperfusion treated human embryo liver cell line through the targeting METTL3 mediated m6A modification of P53.

Background: This study was performed to explore the role of Re in liver IRI progression. Hypoxia and reperfusion (H/R) treated human embryo liver cell line (L-02) was used to establish a liver IRI model. Materials and methods: Cell behaviors were detected using CCK-8, flow cytometry and TUNEL staining assays. The m6A content was detected using m6A dot blot assay. RT-qPCR and western blot assays were used to assessed the relative mRNA and protein levels. MeRIP assay was conducted to determine the m6A levels of P53. The relationship between METTL3 and P53 was demonstrated using RIP and dual-luciferase reporter assays. Results: The results showed that Re treatment significantly decreased the cell apoptosis and promoted the cell viability in the H/R treated L-02 cells. Besides, H/R treatment increased the METTL3 and m6A levels in the L-02 cells, and Re treatment decreased them. Additionally, METTL3 overexpression reversed the role of Re in the H/R treated L-02 cells. Mechanistically, METTL3 overexpression enhanced the m6A levels and mRNA stability and expressions of P53. The combination of METTL3 and P53 was further confirmed. Conclusion: In conclusion, this study demonstrated that Re treatment relieved the H/R induced injury in the L-02 cells through decreasing the METTL3 levels. METTL3 enhanced the mRNA stability and expressions of P53 through m6A modification. Re-METTL3-P53 axis might a new direction for the treatment of liver IRI in the future.

Multi-Method Quantification of Acetyl-Coenzyme A and Further Acyl-Coenzyme A Species in Normal and Ischemic Rat Liver.

Thioesters of coenzyme A (CoA) carrying different acyl chains (acyl-CoAs) are central intermediates of many metabolic pathways and donor molecules for protein lysine acylation. Acyl-CoA species largely differ in terms of cellular concentrations and physico-chemical properties, rendering their analysis challenging. Here, we compare several approaches to quantify cellular acyl-CoA concentrations in normal and ischemic rat liver, using HPLC and LC-MS/MS for multi-acyl-CoA analysis, as well as NMR, fluorimetric and spectrophotometric techniques for the quantification of acetyl-CoAs. In particular, we describe a simple LC-MS/MS protocol that is suitable for the relative quantification of short and medium-chain acyl-CoA species. We show that ischemia induces specific changes in the short-chain acyl-CoA relative concentrations, while mild ischemia (1-2 min), although reducing succinyl-CoA, has little effects on acetyl-CoA, and even increases some acyl-CoA species upstream of the tricarboxylic acid cycle. In contrast, advanced ischemia (5-6 min) also reduces acetyl-CoA levels. Our approach provides the keys to accessing the acyl-CoA metabolome for a more in-depth analysis of metabolism, protein acylation and epigenetics.

Caspase 6 promotes innate immune activation by functional crosstalk between RIPK1-IκBα axis in liver inflammation.

BACKGROUND: Caspase 6 is an essential regulator in innate immunity, inflammasome activation and host defense. We aimed to characterize the causal mechanism of Caspase 6 in liver sterile inflammatory injury. METHODS: Human liver tissues were harvested from patients undergoing ischemia-related hepatectomy to evaluate Caspase 6 expression. Subsequently, we created Caspase 6-knockout (Caspase 6KO) mice to analyze roles and molecular mechanisms of macrophage Caspase 6 in murine models of liver ischemia/reperfusion (IR) injury. RESULTS: In human liver biopsies, Caspase 6 expression was positively correlated with more severe histopathological injury and higher serum ALT/AST level at one day postoperatively. Moreover, Caspase 6 was mainly elevated in macrophages but not hepatocytes in ischemic livers. Unlike in controls, the Caspase 6-deficient livers were protected against IR injury, as evidenced by inhibition of inflammation, oxidative stress and iron overload. Disruption of macrophage NF-κB essential modulator (NEMO) in Caspase 6-deficient livers deteriorated liver inflammation and ferroptosis. Mechanistically, Caspase 6 deficiency spurred NEMO-mediated IκBα phosphorylation in macrophage. Then phosphorylated-inhibitor of NF-κBα (p-IκBα) co-localized with receptor-interacting serine/ threonine-protein kinase 1 (RIPK1) in the cytoplasm to degradate RIPK1 under inflammatory conditions. The disruption of RIPK1-IκBα interaction preserved RIPK1 degradation, triggering downstream apoptosis signal-regulating kinase 1 (ASK1) phosphorylation and inciting NIMA-related kinase 7/NOD-like receptor family pyrin domain containing 3 (NEK7/NLRP3) activation in macrophages. Moreover, ablation of macrophage RIPK1 or ASK1 diminished NEK7/NLRP3-driven inflammatory response and dampened hepatocyte ferroptosis by reducing HMGB1 release from macrophages. CONCLUSIONS: Our findings underscore a novel mechanism of Caspase 6 mediated RIPK1-IκBα interaction in regulating macrophage NEK7/NLRP3 function and hepatocytes ferroptosis, which provides therapeutic targets for clinical liver IR injury. Video Abstract.

Proanthocyanidin Alleviates Liver Ischemia/Reperfusion Injury by Suppressing Autophagy and Apoptosis via the PPARα/PGC1α Signaling Pathway.

Background and Aims: Hepatic ischemia-reperfusion injury (IRI) is a common pathophysiological phenomenon in clinical practice, which usually occurs in liver transplantation, liver resection, severe trauma, and hemorrhagic shock. Proanthocyanidin (PC), exerted from various plants with antioxidant, antitumor, and antiaging activity, were administrated in our study to investigate the underlying mechanism of its protective function on IRI. Methods: Two doses of PC (50 mg/kg, 100 mg/kg) were given to BALB/c mice by intragastric administration for 7 days before partial (70%) warm IR surgery. Serum and liver tissues were collected 2, 8, and 24 h after reperfusion for relevant experiments. Results: The results of transaminase and hematoxylin and eosin staining indicated that PC pretreatment significantly alleviated IRI in mice. Serum total superoxide dismutase increased and malondialdehyde decreased in PC pretreatment groups. Enzyme-linked immunosorbent assays, western blotting, quantitative real-time polymerase chain reaction, and immunohistochemistry showed that inflammation, apoptosis, and autophagy in PC preprocessing groups were significantly inhibited and were dose-dependent. The protein, mRNA expression, and immunohistochemical staining results of peroxisome proliferator-activated receptor alpha (PPARα) and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) in the PC pretreatment groups were significantly upregulated compared with the IR group in a dose-dependent manner. Conclusions: PC pretreatment suppressed inflammation, apoptosis, and autophagy via the PPAR-α signaling pathway to protect against IRI of the liver in mice.

Trans-anethole pretreatment ameliorates hepatic ischemia-reperfusion injury via regulation of soluble epoxide hydrolase.

Hepatic ischemia reperfusion injury (IRI) is a risk factor for early graft nonfunction and graft rejection after liver transplantation (LT). The process of liver IRI involves inflammatory response, oxidative stress, apoptosis and other pathophysiological processes. So far, there is still a lack of effective drugs to ameliorate liver IRI. Trans-anethole (TA) is an aromatic compound. Many medications as well as natural foods contain TA. TA has multiple effects such as anti-inflammation, anti-oxidative stress and anti-apoptosis. However, the mechanism of TA pretreatment in liver IRI is unclear. The mice hepatic IRI model was constructed after gavage pretreatment with TA (10 mg/kg, 20 mg/kg, 40 mg/kg) for 7 consecutive days. Our study confirmed that TA pretreatment significantly improve liver function and reduce serum AST, ALT in hepatic IRI. HE staining showed that TA pretreatment alleviated liver injury. Meanwhile, TA (20 mg/kg) pretreatment attenuated hepatocyte apoptosis in hepatic IRI. In addition, TA (20 mg/kg) pretreatment reduced the inflammatory factors TNF-α, IL-6 and infiltration of CD11b positive cells in liver tissues during hepatic IRI in mice. TA pretreatment also alleviated oxidative stress in mice hepatic IRI. Our study further indicated that TA pretreatment attenuated mice hepatic IRI through inhibiting NLRP3 inflammasome activation via regulation of soluble epoxide hydrolase (sEH). This study provides a novel and effective potential drug with few side effects for easing liver IRI.

Vitamin D receptor mediates liver ischemia and reperfusion injury by autophagy-regulated M2 macrophage polarization.

Liver ischemia and reperfusion (IR) injury is the major complication of liver-related operations. Macrophage polarization has an essential effect on the mechanism of liver IR injury. Vitamin D receptor (VDR) has been found to regulate macrophage polarization and alleviate IR injury. Nevertheless, the correlation between VDR and macrophage polarization in liver IR injury has not been clearly elucidated. VDR knockout mice and wild-type littermates underwent partial liver ischemia for 90 min and reperfusion for 6 h. RAW264.7 cells were also used to verify the influence of VDR on macrophage polarization in vitro. VDR activation could promote M2 macrophage polarization and then reduce liver injury. In contrast, VDR deficiency aggravated the liver injury by disturbing M2 macrophage polarization. Moreover, autophagy participated in the effect of VDR on M2 macrophage polarization through mediating suppressor of cytokine signaling. Therefore, VDR plays a vital influence in liver IR injury. The protective role of VDR activation in liver IR injury is related to regulate M2 macrophage polarization by autophagy.

Hepatocyte HSPA12A inhibits macrophage chemotaxis and activation to attenuate liver ischemia/reperfusion injury via suppressing glycolysis-mediated HMGB1 lactylation and secretion of hepatocytes.

Rationale: Liver ischemia-reperfusion (LI/R) injury is characterized by two interconnected phases: local ischemia that causes hepatic cell damage to release damage-associated molecular pattern (DAMPs), and DAMPs that recruit immune cells to elicit inflammatory cascade for further injury of hepatocytes. High-mobility group box 1 (HMGB1) is a representative DAMP. Studies in macrophages demonstrated that HMGB1 is secreted after lactylation during sepsis. However, whether lactylation mediates HMGB1 secretion from hepatocytes after LI/R is known. Heat shock protein A12A (HSPA12A) is an atypical member of HSP70 family. Methods: Gene expression was examined by microarray analysis and immunoblotting. The hepatic injury was analyzed using released ALT and AST activities assays. Hepatic macrophage chemotaxis was evaluated by Transwell chemotaxis assays. Inflammatory mediators were evaluated by immunoblotting. HMGB1 secretion was examined in exosomes or serum. HMGB1 lactylation was determined using immunoprecipitation and immunoblotting. Results: Here, we report that LI/R decreased HSPA12A expression in hepatocytes, while hepatocyte-specific HSPA12A overexpression attenuated LI/R-induced hepatic dysfunction and mortality of mice. We also noticed that hepatocyte HSPA12A overexpression suppressed macrophage chemotaxis to LI/R-exposed livers in vivo and to hypoxia/reoxygenation (H/R)-exposed hepatocytes in vitro. The LI/R-increased serum HMGB1 levels of mice and the H/R-increased HMGB1 lactylation and secretion levels of hepatocytes were also inhibited by hepatocyte HSPA12A overexpression. By contrast, HSPA12A knockout in hepatocytes promoted not only H/R-induced HMGB1 lactylation and secretion of hepatocytes but also the effects of H/R-hepatocytes on macrophage chemotaxis and inflammatory activation, while all these deleterious effects of HSPA12A knockout were reversed following hepatocyte HMGB1 knockdown. Further molecular analyses showed that HSPA12A overexpression reduced glycolysis-generated lactate, thus decreasing HMGB1 lactylation and secretion from hepatocytes, thereby inhibiting not only macrophage chemotaxis but also the subsequent inflammatory cascade, which ultimately protecting against LI/R injury. Conclusion: Taken together, these findings suggest that hepatocyte HSPA12A is a novel regulator that protects livers from LI/R injury by suppressing glycolysis-mediated HMGB1 lactylation and secretion from hepatocytes to inhibit macrophage chemotaxis and inflammatory activation. Therefore, targeting hepatocyte HSPA12A may have therapeutic potential in the management of LI/R injury in patients.

MSCs Ameliorate Hepatic IR Injury by Modulating Phenotypic Transformation of Kupffer Cells Through Drp-1 Dependent Mitochondrial Dynamics.

BACKGROUND: Hepatic ischemia and reperfusion (IR) injury, characterized by reactive oxygen species (ROS) production and immune disorders, leads to exogenous antigen-independent local inflammation and hepatocellular death. Mesenchymal stem cells (MSCs) have been shown to be immunomodulatory, antioxidative and contribute to liver regeneration in fulminant hepatic failure. We aimed to investigate the underlying mechanisms by which MSCs protect against liver IR injury in a mouse model. METHODS: MSCs suspension was injected 30 min prior to hepatic warm IR. Primary kupffer cells (KCs) were isolated. Hepatic injury, inflammatory responses, innate immunity, KCs phenotypic polarization and mitochondrial dynamics were evaluated with or without KCs Drp-1 overexpression RESULTS: MSCs markedly ameliorated liver injury and attenuated inflammatory responses and innate immunity after liver IR injury. MSCs significantly restrained M1 phenotypic polarization but boosted M2 polarization of KCs extracted from ischemic liver, as demonstrated by lowered transcript levels of iNOS and IL-1ß but raised transcript levels of Mrc-1 and Arg-1 combined with p-STAT6 up-regulation and p-STAT1 down-regulation. Moreover, MSCs inhibited KCs mitochondrial fission, as evidenced by decreased Drp1 and Dnm2 levels. We overexpressed Drp-1 in KCs which promote mitochondrial fission during IR injury. the regulation of MSCs towards KCs M1/M2 polarization was abrogated by Drp-1 overexpression after IR injury. Ultimately, in vivo Drp-1 overexpression in KCs hampered the therapeutic effects of MSCs against hepatic IR injury CONCLUSIONS: We revealed that MSCs facilitated M1-M2 phenotypic polarization through inhibiting Drp-1 dependent mitochondrial fission and further attenuated liver IR injury. These results add a new insight into regulating mechanisms of mitochondrial dynamics during hepatic IR injury and may offer novel opportunities for developing therapeutic targets to combat hepatic IR injury.

TRIM37 exacerbates hepatic ischemia/reperfusion injury by facilitating IKKγ translocation.

BACKGROUND: Hepatic ischemia/reperfusion (I/R) injury is one of the major pathological processes associated with various liver surgeries. However, there is still a lack of strategies to protect against hepatic I/R injury because of the unknown underlying mechanism. The present study aimed to identify a potential strategy and provide a fundamental experimental basis for treating hepatic I/R injury. METHOD: A classic 70% ischemia/reperfusion injury was established. Immunoprecipitation was used to identify direct interactions between proteins. The expression of proteins from different subcellular localizations was detected by Western blotting. Cell translocation was directly observed by immunofluorescence. HE, TUNEL and ELISA were performed for function tests. RESULT: We report that tripartite motif containing 37 (TRIM37) aggravates hepatic I/R injury through the reinforcement of IKK-induced inflammation following dual patterns. Mechanistically, TRIM37 directly interacts with tumor necrosis factor receptor-associated factor 6 (TRAF6), inducing K63 ubiquitination and eventually leading to the phosphorylation of IKKß. TRIM37 enhances the translocation of IKKγ, a regulatory subunit of the IKK complex, from the nucleus to the cytoplasm, thereby stabilizing the cytoplasmic IKK complex and prolonging the duration of inflammation. Inhibition of IKK rescued the function of TRIM37 in vivo and in vitro. CONCLUSION: Collectively, the present study discloses some potential function of TRIM37 in hepatic I/R injury. Targeting TRIM37 might be potential for treatment against hepatic I/R injury.Targeting TRIM37 might be a potential treatment strategy against hepatic I/R injury.