Cirbp suppression compromises DHODH-mediated ferroptosis defense and attenuates hypothermic cardioprotection in an aged donor transplantation model.

Hypothermia is commonly used to protect donor hearts during transplantation. However, patients transplanted with aged donor hearts still have severe myocardial injury and decreased survival rates, but the underlying mechanism remains unknown. Because aged hearts are not considered suitable for donation, the number of patients awaiting heart transplants is increasing. In this study, we examined whether hypothermic cardioprotection was attenuated in aged donor hearts during transplantation and evaluated potential therapeutic targets. Using a rat heart transplantation model, we found that hypothermic cardioprotection was impaired in aged donor hearts but preserved in young donor hearts. RNA-Seq showed that cold-inducible RNA-binding protein (Cirbp) expression was decreased in aged donor hearts, and these hearts showed severe ferroptosis after transplantation. The young donor hearts from Cirbp-KO rats exhibited attenuated hypothermic cardioprotection, but Cirbp overexpression in aged donor hearts ameliorated hypothermic cardioprotection. Cardiac proteomes revealed that dihydroorotate dehydrogenase (DHODH) expression was significantly decreased in Cirbp-KO donor hearts during transplantation. Consequently, DHODH-mediated ubiquinone reduction was compromised, thereby exacerbating cardiac lipid peroxidation and triggering ferroptosis after transplantation. A cardioplegic solution supplemented with CIRBP agonists improved hypothermic cardioprotection in aged donor hearts, indicating that this method has the potential to broaden the indications for using aged donor hearts in transplantation.

Rescue of cardiac dysfunction during chemotherapy in acute myeloid leukaemia by blocking IL-1α.

BACKGROUND AND AIMS: Patients with acute myeloid leukaemia (AML) suffer from severe myocardial injury during daunorubicin (DNR)-based chemotherapy and are at high risk of cardiac mortality. The crosstalk between tumour cells and cardiomyocytes might play an important role in chemotherapy-related cardiotoxicity, but this has yet to be demonstrated. This study aimed to identify its underlying mechanism and explore potential therapeutic targets. METHODS: Cardiac tissues were harvested from an AML patient after DNR-based chemotherapy and were subjected to single-nucleus RNA sequencing. Cardiac metabolism and function were evaluated in AML mice after DNR treatment by using positron emission tomography, magnetic resonance imaging, and stable-isotope tracing metabolomics. Plasma cytokines were screened in AML mice after DNR treatment. Genetically modified mice and cell lines were used to validate the central role of the identified cytokine and explore its downstream effectors. RESULTS: In the AML patient, disruption of cardiac metabolic homeostasis was associated with heart dysfunction after DNR-based chemotherapy. In AML mice, cardiac fatty acid utilization was attenuated, resulting in cardiac dysfunction after DNR treatment, but these phenotypes were not observed in similarly treated tumour-free mice. Furthermore, tumour cell-derived interleukin (IL)-1α was identified as a primary factor leading to DNR-induced cardiac dysfunction and administration of an anti-IL-1α neutralizing antibody could improve cardiac functions in AML mice after DNR treatment. CONCLUSIONS: This study revealed that crosstalk between tumour cells and cardiomyocytes during chemotherapy could disturb cardiac energy metabolism and impair heart function. IL-1α neutralizing antibody treatment is a promising strategy for alleviating chemotherapy-induced cardiotoxicity in AML patients.

Corrigendum: Identification of ACKR4 as an immune checkpoint in pulmonary arterial hypertension.

[This corrects the article DOI: 10.3389/fimmu.2023.1153573.].

Application of fecal collection device in Intensive Care Unit Patients with fecal incontinence Receiving Extracorporeal Membrane Oxygenation: A Comparison Cohort Study.

Objective: The objective of this study was to assess the effectiveness of fecal collection devices in preventing incontinence-associated dermatitis (IAD) and reducing skin care time in ICU patients with fecal incontinence undergoing Extracorporeal Membrane Oxygenation (ECMO). Methods: A nonrandomized comparison cohort (quasi-experimental) study with pre-post comparison was carried out in a general intensive care unit. 85 bedridden patients receiving ECMO with fecal incontinence (FI) in a general intensive care unit between June 2017 and May 2022 participated in the study and separated into two groups according to the fecal collection device they received. 40 were assigned to the Control group (structured IAD preventive care protocol alone) and 45 to the Intervention group (structured IAD preventive care protocol plus application of fecal collection device). The status of IAD was assessed using the Incontinence Associated Dermatitis Intervention Tool (IAD-IT). Fecal consistency was evaluated via the Bristol Stool Scale. Outcome measures included the nursing time for skin care and the incidence of IAD, and bleeding complications between the two groups during the period. Results: Participants in the Intervention group had fewer IAD occurrences than participants in the Control group (13.33% vs. 52.50%, P < .05). The patients in the Intervention group significantly reduced skincare time (63.30±14.09 min in the Control group versus 28.44±2.04 min in the Intervention group, P < .01). There was 3 turning complications for bleeding in the Intervention group and 11 in the Control group and had a significant reduction in urning complications(3 vs.11, P = .022). Conclusions: Applying a fecal collection device may reduce skincare time and reduce occurrences of IAD and bleeding related to turning position for skin care in ICU patients with FI associated with diarrhea receiving ECMO Support.This study offers a more efficient way to use the fecal collection device in ECMO patients.Future research needs to focus on the perianal skin in ECMO patients regarding fecal collection devices connected to continuous low-negative-pressure suction devices.

Effects of trimetazidine on cardiac function in adult cyanotic congenital heart disease patients: Protocol for a 3-month multicenter, randomized, double-blind controlled trial.

BACKGROUND: Nearly 20% Patients with cyanotic congenital heart disease (CCHD) are not able to receive surgery. These patients experience a decline in cardiac function as they age, which has been demonstrated to be associated with changes in energy metabolism in cardiomyocytes. Trimetazidine (TMZ), a metabolic regulator, is supposed to alleviate such maladaptation and reserve cardiac function in CCHD patients. METHODS: This is a randomized, double-blind, placebo-controlled clinical trial. Eighty adult CCHD patients will be recruited and randomized to the TMZ (20 mg TMZ 3 times a day for 3 months) or placebo group (placebo 3 times a day for 3 months). The primary outcome is the difference in cardiac ejection fractions (EF) measured by cardiac magnetic resonance (MRI) between baseline and after 3 months of TMZ treatment. The secondary outcomes include TMZ serum concentration, rate of cardiac events, NYHA grading, fingertip SpO2, NT-proBNP levels, 6-minute walking test (6MWT), KCCQ-CSS questionnaire score, echocardiography, ECG, routine blood examination, liver and kidney function test, blood pressure and heart rate. DISCUSSION: This trial is designed to explore whether the application of TMZ in adult CCHD patients can improve cardiac function, reduce cardiac events, and improve exercise performance and quality of life. The results will provide targeted drug therapy for CCHD patients with hypoxia and support the application of TMZ in children with CCHD.

Signaling via a CD28/CD40 chimeric costimulatory antigen receptor (CoStAR™), targeting folate receptor alpha, enhances T cell activity and augments tumor reactivity of tumor infiltrating lymphocytes.

Transfer of autologous tumor infiltrating lymphocytes (TIL) to patients with refractory melanoma has shown clinical efficacy in a number of trials. However, extending the clinical benefit to patients with other cancers poses a challenge. Inefficient costimulation in the tumor microenvironment can lead to T cell anergy and exhaustion resulting in poor anti-tumor activity. Here, we describe a chimeric costimulatory antigen receptor (CoStAR) comprised of FRα-specific scFv linked to CD28 and CD40 intracellular signaling domains. CoStAR signaling alone does not activate T cells, while the combination of TCR and CoStAR signaling enhances T cell activity resulting in less differentiated T cells, and augmentation of T cell effector functions, including cytokine secretion and cytotoxicity. CoStAR activity resulted in superior T cell proliferation, even in the absence of exogenous IL-2. Using an in vivo transplantable tumor model, CoStAR was shown to improve T cell survival after transfer, enhanced control of tumor growth, and improved host survival. CoStAR could be reliably engineered into TIL from multiple tumor indications and augmented TIL activity against autologous tumor targets both in vitro and in vivo. CoStAR thus represents a general approach to improving TIL therapy with synthetic costimulation.

Identification of ACKR4 as an immune checkpoint in pulmonary arterial hypertension.

Objective: Inflammation is recognized as a contributor in the development of pulmonary arterial hypertension (PAH), and the recruitment and functional capacity of immune cells are well-orchestrated by chemokines and their receptors. This study is aimed at identification of critical chemokines in the progression of PAH via transcriptomic analysis. Methods: Differentially expressed genes (DEGs) from lungs of PAH patients were achieved compared to controls based on Gene Expression Omnibus (GEO) database. Gene set enrichment analysis (GSEA) was applied for functional annotation and pathway enrichement. The abundance of immune cells was estimated by the xCell algorithm. Weighted correlation network analysis (WGCNA) was used to construct a gene expression network, based on which a diagnostic model was generated to determine its accuracy to distinguish PAH from control subjects. Target genes were then validated in lung of hypoxia-induce pulmonary hypertension (PH) mouse model. Results: ACKR4 (atypical chemokine receptor 4) was downregulated in PAH lung tissues in multiple datasets. PAH relevant biological functions and pathways were enriched in patients with low-ACKR4 level according to GSEA enrichment analysis. Immuno-infiltration analysis revealed a negative correlation of activated dendritic cells, Th1 and macrophage infiltration with ACKR4 expression. Three gene modules were associated with PAH via WGCNA analysis, and a model for PAH diagnosis was generated using CXCL12, COL18A1 and TSHZ2, all of which correlated with ACKR4. The ACKR4 expression was also downregulated in lung tissues of our experimental PH mice compared to that of controls. Conclusions: The reduction of ACKR4 in lung tissues of human PAH based on transcriptomic data is consistent with the alteration observed in our rodent PH. The correlation with immune cell infiltration and functional annotation indicated that ACKR4 might serve as a protective immune checkpoint for PAH.

Safe and Effective Delivery of mRNA Using Modified PEI-Based Lipopolymers.

Chemically modified mRNA (modRNA) has proven to be a versatile tool for the treatment of various cancers and infectious diseases due to recent technological advancements. However, a safe and effective delivery system to overcome the complex extracellular and intracellular barriers is required in order to achieve higher therapeutic efficacy and broaden clinical applications. Here, we explored All-Fect and Leu-Fect C as novel transfection reagents derived from lipopolymers, which demonstrated excellent biocompatibility, efficient delivery capabilities, and a robust ability to escape the lysosomes. These properties directly increase mRNA stability by preventing mRNA degradation by nucleases and simultaneously promote efficient gene translation in vitro and in vivo. The modRNA delivered with lipopolymer vectors sustained effective transfection in mouse hearts following direct intramyocardial injection, as well as in major organs (liver and spleen) after systemic administration. No observable immune reactions or systemic toxicity were detected following the systemic administration of lipopolymer-mRNA complexes to additional solid organs. This study identified commercial reagents for the effective delivery of modRNA and may help facilitate the advancement of gene-based interventions involving the safe and effective delivery of nucleic acid drug substances.

Impact of Double-Machine Replacement Protocol at Start of Continuous Renal Replacement Therapy in Vasopressor-Dependent Patients: A Retrospective Cohort Study.

INTRODUCTION: When starting continuous renal replacement therapy (CRRT), vasopressor-dependent patients are at risk of hemodynamic instability. Thus far, only a few studies have analyzed the impact of CRRT circuit replacement for vasopressor-dependent patients. Hence, we compared the effect of double-machine replacement protocol (DMRP) with single-machine replacement protocol (SMRP) for CRRT circuit replacement in vasopressor-dependent patients. METHODS: The medical records of 96 vasopressor-dependent patients treated with CRRT in the general intensive care unit of the Shunde Hospital, Southern Medical University, between January 2017 and April 2018 were retrospectively analyzed. The major measures of the SMRP included returning the blood to the patient and sealing access catheter with heparin and starting a new CRRT machine with a slow blood pump, while DMRP involved simultaneous drawing and return of blood with two machines using a slow blood pump for circuit replacement. The primary outcome measures were changes in vasopressor dose and hemodynamic parameters, and the secondary outcome measure was the pause time difference between the two groups during the period. RESULTS: A total of 53 patients were treated with SMRP and 43 patients with DMRP. Heart rate was higher in the SMRP group as compared to the DMRP group (p < 0.05). There were no significant changes in central venous pressure, mean arterial pressure, and vasopressor dose in either group (p > 0.05). The patients in the DMRP group had a significant reduction in CRRT pause time (5.62 ± 0.69 min in DMRP group vs. 37.01 ± 8.72 min in SMRP group, p < 0.01). The DMRP group needed a lower volume of circuit purging and priming fluid related to CRRT circuit replacement (0 mL in DMRP group vs. 463 mL in SMRP group). CONCLUSIONS: Implementation of the DMRP for CRRT circuit replacement had a slight hemodynamic effect on vasopressor-dependent patients. It also reduced the pause time and volume of circuit purging and priming fluid related to CRRT circuit replacement compared with SMRP.

Gelatin/Polycaprolactone Electrospun Nanofibrous Membranes: The Effect of Composition and Physicochemical Properties on Postoperative Cardiac Adhesion.

Cardiovascular diseases have become a major threat to human health. The adhesion formation is an inevitable pathophysiological event after cardiac surgery. We have previously shown that gelatin/polycaprolactone (GT/PCL, mass ratio 50:50) electrospun nanofibrous membranes have high potential in preventing postoperative cardiac adhesion, but the effect of GT:PCL composition on anti-adhesion efficacy was not investigated. Herein, nanofibrous membranes with different GT:PCL mass ratios of 0:100, 30:70, 50:50, and 70:30 were prepared via electrospinning. The 70:30 membrane failed to prevent postoperative cardiac adhesion, overly high GT contents significantly deteriorated the mechanical properties, which complicated the suturing during surgery and hardly maintained the structural integrity after implantation. Unexpectedly, the 0:100 membrane (no gelatin contained) could not effectively prevent either, since its large pore size allowed the penetration of numerous inflammatory cells to elicit a severe inflammatory response. Only the GT:PCL 50:50 membrane exhibited excellent mechanical properties, good biocompatibility and effective anti-cell penetration ability, which could serve as a physical barrier to prevent postoperative cardiac adhesion and might be suitable for other biomedical applications such as wound healing, guided tissue or bone regeneration.

Resveratrol Reduced Liver Damage After Liver Resection in a Rat Model by Upregulating Sirtuin 1 (SIRT1) and Inhibiting the Acetylation of High Mobility Group Box 1 (HMGB1).

BACKGROUND Liver failure after resection for liver cancer is associated with increased patient mortality. This study aimed to investigate the mechanism of the protective effects of resveratrol, a natural plant-derived compound, on liver injury in a rat model of partial hepatectomy. MATERIAL AND METHODS Adult male Sprague-Dawley (SD) rats (n=60) were divided into the sham group (n=20), the liver resection group (n=20), and the liver resection plus resveratrol-treated group (n=20). Liver resection removed 2/3 of the liver resection; resveratrol was given at a dose of 30 mg/kg/day from one week before surgery until death. Liver injury was assessed by serum liver function tests, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), γ-glutamyl-transferase (γ-GT) and total bilirubin, histological examination of the rat liver, and liver cell apoptosis using the TUNEL assay. High mobility group box 1 (HMGB1) expression was measured by enzyme-linked immunoassay (ELISA). Sirtuin 1 (SIRT1) and acetylated HMGB1 (Ac-HMGB1) expression were detected by Western blot. Normal human liver cells and HepG2 liver cancer cells were incubated with acetylated HMGB1, and albumin production and ammonia elimination assays were performed. RESULTS Resveratrol reduced postoperative liver injury as shown by reduced ALT, AST, γ-GT, and total bilirubin levels, maintained liver structure, and reduced cell apoptosis. Resveratrol treatment reduced the expression and acetylation levels of HMGB1 via the SIRT1 signaling pathway. Resveratrol reversed Ac-HMGB1 induced dysfunction in liver cells cultured in vitro. CONCLUSIONS Resveratrol reduced liver damage after liver resection in a rat model by upregulating SIRT1 and reducing the acetylation of HMGB1.

Long-term self-renewal of naïve neural stem cells in a defined condition.

During embryonic development, neural stem cells (NSCs) emerge as early as the neural plate stage and give rise to the nervous system. Early-stage NSCs express Sry-related-HMG box-1 (Sox1) and are biased towards neuronal differentiation. However, long-term maintenance of early-stage NSCs in vitro remains a challenge. Here, we report development of a defined culture condition for the long-term maintenance of Sox1-positive early-stage mouse NSCs. The proliferative ability of these Sox1-positive NSCs was confirmed by clonal propagation. Compared to the NSCs cultured using the traditional culture condition, the long-term self-renewing Sox1-positive NSCs efficiently differentiate into neurons and exhibit an identity representative of the anterior and midbrain regions. These early-stage Sox1-positive NSCs could also be switched to late-stage NSCs by being cultured with bFGF/EGF, which can then differentiate into astrocytes and oligodendrocytes. The long-term self-renewing Sox1-positive NSCs were defined as naïve NSCs, based on their high neuronal differentiation capacity and anterior regional identity. This culture condition provides a robust platform for further dissection of the NSC self-renewal mechanism and promotes potential applications of NSCs for cell-based therapy on nervous system disorders.

Suppression of stress induction of the 78-kilodalton glucose regulated protein (GRP78) in cancer by IT-139, an anti-tumor ruthenium small molecule inhibitor.

In many cancers, combination therapy regimens are successfully improving response and survival rates, but the challenges of toxicity remain. GRP78, the master regulator of the unfolded protein response, is emerging as a target that is upregulated in tumors, specifically following treatment, and one that impacts tumor cell survival and disease recurrence. Here, we show IT-139, an antitumor small molecule inhibitor, suppresses induction of GRP78 from different types of endoplasmic reticulum (ER) stress in a variety of cancer cell lines, including those that have acquired therapeutic resistance, but not in the non-cancer cells being tested. We further determined that IT-139 treatment exacerbates ER stress while at the same time suppresses GRP78 induction at the transcriptional level. Our studies revealed a differential effect of IT-139 on chaperone protein family expression at multiple levels in different cancer cell lines. In xenograft studies, IT-139 decreased BRAF inhibitor upregulation of GRP78 expression in the tumor, while having minimal effect on GRP78 expression in the adjacent normal cells. The preferential decrease in GRP78 levels in tumor cells over normal cells, supported by the manageable safety profile seen in the Phase 1 clinical trial, reinforce the value IT-139 brings to combination therapies as it continues its clinical development.

Modulation of GSK-3ß/ß-Catenin Signaling Contributes to Learning and Memory Impairment in a Rat Model of Depression.

Background: It is widely accepted that cognitive processes, such as learning and memory, are affected in depression, but the molecular mechanisms underlying the interactions of these 2 disorders are not clearly understood. Recently, glycogen synthase kinase-3 beta (GSK-3ß)/ß-catenin signaling was shown to play an important role in the regulation of learning and memory. Methods: The present study used a rat model of depression, chronic unpredictable stress, to determine whether hippocampal GSK-3ß/ß-catenin signaling was involved in learning and memory alterations. Results: Our results demonstrated that chronic unpredictable stress had a dramatic influence on spatial cognitive performance in the Morris water maze task and reduced the phosphorylation of Ser9 of GSK-3ß as well as the total and nuclear levels of ß-catenin in the hippocampus. Inhibition of GSK3ß by SB216763 significantly ameliorated the cognitive deficits induced by chronic unpredictable stress, while overexpression of GSK3ß by AAV-mediated gene transfer significantly decreased cognitive performance in adult rats. In addition, chronic unpredictable stress exposure increased the expression of the canonical Wnt antagonist Dkk-1. Furthermore, chronic administration of corticosterone significantly increased Dkk-1 expression, decreased the phosphorylation of Ser9 of GSK-3ß, and resulted in the impairment of hippocampal learning and memory. Conclusions: Our results indicate that impairment of learning and memory in response to chronic unpredictable stress may be attributed to the dysfunction of GSK-3ß/ß-catenin signaling mediated by increased glucocorticoid signaling via Dkk-1.

Effect of a 1-Piece Drainable Pouch on Incontinence-Associated Dermatitis in Intensive Care Unit Patients With Fecal Incontinence: A Comparison Cohort Study.

PURPOSE: The purpose of this study was to compare the effect of a 1-piece drainable pouch to standard care on occurrences of incontinence-associated dermatitis (IAD) in intensive care unit (ICU) patients with fecal incontinence (FI). DESIGN: Nonrandomized comparison cohort (quasi-experimental) study. METHODS: Sixty-two bedridden patients with FI and indwelling urinary catheters in the ICU of the Shunde Hospital, Southern Medical University, Foshan, China, participated in the study. Thirty-one were assigned to the control group (standard IAD preventive care alone) and 31 to the intervention group (standard IAD preventive care plus application of a 1-piece drainable pouch). Stool consistency was evaluated via the Bristol Stool Scale. Trained nurses assessed the status of IAD using the Incontinence-Associated Dermatitis Intervention Tool. The incidence of IAD and the perianal skin status were investigated over a 7-day period. RESULTS: Participants in the experimental group had fewer IAD occurrences than participants in the control group (12.9% vs 41.9%, P < .05). Occurrences of IAD in the perianal were also significantly lower in the experimental group than in the control group (χ = 7.884, P < .05). CONCLUSION: Applying a 1-piece drainable pouch may reduce occurrences of IAD in ICU patients with FI compared with the patients receiving standard IAD preventive care.

Cytoplasmic and Nuclear TAZ Exert Distinct Functions in Regulating Primed Pluripotency.

Mouse epiblast stem cells (mEpiSCs) and human embryonic stem cells (hESCs) are primed pluripotent stem cells whose self-renewal can be maintained through cytoplasmic stabilization and retention of ß-catenin. The underlying mechanism, however, remains largely unknown. Here, we show that cytoplasmic ß-catenin interacts with and retains TAZ, a Hippo pathway effector, in the cytoplasm. Cytoplasmic retention of TAZ promotes mEpiSC self-renewal in the absence of nuclear ß-catenin, whereas nuclear translocation of TAZ induces mEpiSC differentiation. TAZ is dispensable for naive mouse embryonic stem cell (mESC) self-renewal but required for the proper conversion of mESCs to mEpiSCs. The self-renewal of hESCs, like that of mEpiSCs, can also be maintained through the cytoplasmic retention of ß-catenin and TAZ. Our study indicates that how TAZ regulates cell fate depends on not only the cell type but also its subcellular localization.

A new approach combining venoarterial extracorporeal membrane oxygenation and CRRT for adults: a retrospective study.

PURPOSE: This study aimed to assess a new approach combining venoarterial (VA) extracorporeal membrane oxygenation (ECMO) and continuous renal replacement therapy (CRRT) in adults, while monitoring CRRT circuit pressures. METHODS: The inlet and outlet of the CRRT circuit were connected to preoxygenator port and postoxygenator port, respectively. Then, complications and CRRT circuit pressures were evaluated. RESULTS: 7 patients underwent combined VA-ECMO and CRRT; 16 filters were used. CRRT blood flow ranged from 150 to 200 mL/min; the CRRT to ECMO blood flow ratio was <0.1. The CRRT pressures at treatment initiation were normal. No complications were reported. CONCLUSIONS: This approach combining VA-ECMO and CRRT in adults did not compromise the accuracy of pressure monitoring systems for CRRT circuit function, and caused no complications. Hence, it may be a feasible method for performing combined VA-ECMO and CRRT in adults.

Depletion of Tcf3 and Lef1 maintains mouse embryonic stem cell self-renewal.

Mouse and rat embryonic stem cell (ESC) self-renewal can be maintained by dual inhibition of glycogen synthase kinase 3 (GSK3) and mitogen-activated protein kinase kinase (MEK). Inhibition of GSK3 promotes ESC self-renewal by abrogating T-cell factor 3 (TCF3)-mediated repression of the pluripotency network. How inhibition of MEK mediates ESC self-renewal, however, remains largely unknown. Here, we show that inhibition of MEK can significantly suppress lymphoid enhancer factor 1 (LEF1) expression in mouse ESCs. Knockdown or knockout of Lef1 partially mimics the self-renewal-promoting effect of MEK inhibitors. Moreover, depletion of both Tcf3 and Lef1 enables maintenance of undifferentiated mouse ESCs without exogenous factors, cytokines or inhibitors. Transcriptome resequencing analysis reveals that LEF1 is closely associated with endoderm specification in ESCs. Thus, our study adds support to the notion that the key to maintaining the ESC ground state is to shield ESCs from differentiative cues.

The myeloid heat shock transcription factor 1/ß-catenin axis regulates NLR family, pyrin domain-containing 3 inflammasome activation in mouse liver ischemia/reperfusion injury.

Heat shock transcription factor 1 (HSF1) has been implicated in the differential regulation of cell stress and disease states. ß-catenin activation is essential for immune homeostasis. However, little is known about the role of macrophage HSF1-ß-catenin signaling in the regulation of NLRP3 inflammasome activation during ischemia/reperfusion (I/R) injury (IRI) in the liver. This study investigated the functions and molecular mechanisms by which HSF1-ß-catenin signaling influenced NLRP3-mediated innate immune response in vivo and in vitro. Using a mouse model of IR-induced liver inflammatory injury, we found that mice with a myeloid-specific HSF1 knockout (HSF1M-KO ) displayed exacerbated liver damage based on their increased serum alanine aminotransferase levels, intrahepatic macrophage/neutrophil trafficking, and proinflammatory interleukin (IL)-1ß levels compared to the HSF1-proficient (HSF1FL/FL ) controls. Disruption of myeloid HSF1 markedly increased transcription factor X-box-binding protein (XBP1), NLR family, pyrin domain-containing 3 (NLRP3), and cleaved caspase-1 expression, which was accompanied by reduced ß-catenin activity. Knockdown of XBP1 in HSF1-deficient livers using a XBP1 small interfering RNA ameliorated hepatocellular functions and reduced NLRP3/cleaved caspase-1 and IL-1ß protein levels. In parallel in vitro studies, HSF1 overexpression increased ß-catenin (Ser552) phosphorylation and decreased reactive oxygen species (ROS) production in bone-marrow-derived macrophages. However, myeloid HSF1 ablation inhibited ß-catenin, but promoted XBP1. Furthermore, myeloid ß-catenin deletion increased XBP1 messenger RNA splicing, whereas a CRISPR/CRISPR-associated protein 9-mediated XBP1 knockout diminished NLRP3/caspase-1. CONCLUSION: The myeloid HSF1-ß-catenin axis controlled NLRP3 activation by modulating the XBP1 signaling pathway. HSF1 activation promoted ß-catenin, which, in turn, inhibited XBP1, leading to NLRP3 inactivation and reduced I/R-induced liver injury. These findings demonstrated that HSF1/ß-catenin signaling is a novel regulator of innate immunity in liver inflammatory injury and implied the therapeutic potential for management of sterile liver inflammation in transplant recipients. (Hepatology 2016;64:1683-1698).

Induction of site-specific chromosomal translocations in embryonic stem cells by CRISPR/Cas9.

Chromosomal translocation is the most common form of chromosomal abnormality and is often associated with congenital genetic disorders, infertility, and cancers. The lack of cellular and animal models for chromosomal translocations, however, has hampered our ability to understand the underlying disease mechanisms and to develop new therapies. Here, we show that site-specific chromosomal translocations can be generated in mouse embryonic stem cells (mESCs) via CRISPR/Cas9. Mouse ESCs carrying translocated chromosomes can be isolated and expanded to establish stable cell lines. Furthermore, chimeric mice can be generated by injecting these mESCs into host blastocysts. The establishment of ESC-based cellular and animal models of chromosomal translocation by CRISPR/Cas9 provides a powerful platform for understanding the effect of chromosomal translocation and for the development of new therapeutic strategies.