Hepatic nicotinamide adenine dinucleotide (NAD+) reduction is a pathological hallmark and pathogenic basis for alcohol-associated liver disease (ALD). This study investigated the therapeutic role of nicotinamide mononucleotide (NMN), a NAD+ precursor naturally presenting in various dietary sources, against ALD. Here, we show that NMN supplementation attenuates NAD+ decline, which in turn attenuates hepatic lipid deposition, dysregulated lipid metabolism genes, oxidative stress, and inflammation in ALD mice, and ultimately ameliorated liver injury. Transcriptomics-based mechanistic analysis demonstrates that alcohol-reduced hepcidin antimicrobial peptide (Hamp) is rescued by NMN, alongside the recovery of circulatory and hepatic iron levels. The hepatic Hamp expression is positively associated with NAD+ content in the liver. Hamp knockdown significantly abolishes NMN-improved lipid accumulation and the dysregulation of lipid metabolism-related genes in ethanol-treated hepatocytes. Further analysis identifies a C/EBPα-involved transcriptional regulation mechanism in NMN-protected Hamp in ALD. These findings identify NMN as a promising dietary therapeutic for ALD, acting via the C/EBPα/Hamp signaling axis.

Alcohol-associated liver disease (ALD) is a major cause of liver-related morbidity and mortality worldwide. Ursolic acid (UA), a plant-derived pentacyclic triterpenoid, has been reported to exert protective effects against ALD; however, its molecular targets remain unclear. This study aimed to identify the potential targets and underlying mechanisms of UA in ALD. C57BL/6N mice were fed a Lieber-DeCarli liquid alcohol diet to establish an ALD model and were administered UA by gavage. We found that UA treatment alleviated hepatic steatosis in both ALD mice and ethanol-exposed AML-12 hepatocytes, accompanied by normalization of lipid metabolism-related gene expression. UA also exerted hepatoprotective effects by attenuating alcohol-induced liver injury and inflammation, while rebalancing ethanol metabolism and antioxidant defense, particularly through increasing the activity of glutathione S-transferases. Mechanistically, UA selectively upregulated glutathione S transferase mu 1 (GSTM1) at both the transcriptional and protein levels compared with other GST isoforms. In vitro, UA increased GSTM1 expression, whereas GSTM1 silencing abolished the protective effects of UA against intracellular lipid accumulation, dysregulation of lipid metabolism-related genes, and inflammatory responses in ethanol-exposed hepatocytes, indicating that GSTM1 is a key mediator of UA action. In addition, UA reduced alcohol-induced phosphorylation of apoptosis signal-regulating kinase 1 (ASK1) and its downstream effectors c-Jun N-terminal kinase (JNK) and p38. Pharmacological inhibition of ASK1 reversed the exacerbated intracellular lipid accumulation caused by GSTM1 knockdown during ethanol exposure. Collectively, these findings demonstrate that UA alleviates ALD by upregulating GSTM1 and inhibiting the ASK1-JNK/p38 pathway, thereby identifying GSTM1 as a promising therapeutic target in ALD.

BACKGROUND:Exertional heat stroke (EHS) requires immediate cooling to prevent organ injury and death. Although cold-water immersion (CWI) is the recommended first-line cooling modality, it is often difficult to implement safely in the emergency department (ED), particularly in patients requiring airway management or ongoing resuscitation. This study aimed to describe an ice-water-soaked sheet coverage protocol and preliminarily evaluate its cooling efficacy and feasibility in patients with EHS. METHODS:We conducted a retrospective study of patients with EHS admitted to the ED between May 2022 and July 2024. Patients treated with either CWI or ice-water sheet coverage were included. The ice-water sheet protocol used 2.0 × 2.5 m sheets soaked in 3-8 °C ice water, applied to the trunk and extremities, and replaced every 4-5 min. Core temperature was continuously monitored via rectal probe. Cooling performance, procedural feasibility, and adverse events were assessed descriptively. RESULTS:A total of 25 male patients were included, 10 in the CWI group and 15 in the ice-water sheet coverage group. Mean age was 32.5 ± 7.4 years. The mean time from arrival in the resuscitation unit to cooling initiation was 5.2 ± 1.8 min in the ice-water sheet coverage group and 8.5 ± 1.3 min in the CWI group. All patients achieved a core temperature < 39.0 °C within 30 min of cooling initiation. The mean cooling rate was 0.09 ± 0.02 °C/min in the ice-water sheet coverage group and 0.12 ± 0.01 °C/min in the CWI group. All patients reached <38.5 °C within 45.3 ± 10.2 min. Ice-water sheet coverage did not interfere with intravenous access, endotracheal intubation, or continuous monitoring. Mild shivering occurred in 20% of patients and was controlled with low-dose sedation. No frostbite, severe arrhythmia, or hemodynamic instability was observed. All patients survived following comprehensive treatment. CONCLUSIONS:Whole-body ice-water sheet coverage achieved rapid and clinically effective cooling in patients with EHS while allowing concurrent resuscitative care. This protocol may represent a practical alternative to CWI in ED settings where immersion cooling is difficult to implement.

Heat stroke (HS) is a life-threatening condition exacerbated by rising global temperatures, with children identified as a particularly vulnerable population. Despite this, basic research on age-related differences in thermotolerance remains limited. In this study, we established a high-temperature and high-humidity exposure model with real-time core body temperature (CBT) monitoring to investigate thermotolerance in young versus adult rats. The results showed that young rats exhibited prolonged CBT plateau phases and delayed HS onset, indicating enhanced thermotolerance compared to adult rats. This was accompanied by significantly milder multi-organ injury and reduced intestinal barrier damage. Young rats displayed lower serum levels of D-lactate and intestinal fatty acid-binding protein, better-preserved intestinal epithelial ultrastructure, and higher expression of tight junction proteins such as ZO-1, Occludin, and E-cadherin. Moreover, young rats showed elevated expression of heat shock proteins (HSP40 and HSP70) in intestinal tissues, which likely contributed to improved barrier integrity and cellular protection. These findings suggest that enhanced intestinal barrier stability and robust HSP responses underlie the superior thermotolerance observed in young rats. However, despite their physiological advantages, infants and young children often suffer poor HS outcomes due to behavioral limitations and caregiver negligence, especially in enclosed environments such as parked vehicles. This highlights the critical need for enhanced caregiver awareness, improved pediatric emergency response training, and preventive strategies to mitigate pediatric HS risk.

Atractylenolide I (AO-I), a major pharmacological ingredient derived from Atractylodes macrocephala, has been reported to possess multiple bioactivities. This study aimed to investigate the potential protective effects of AO-I against lipotoxicity in both cultured hepatocytes and mice with non-alcoholic fatty liver disease (NAFLD). The NAFLD model was established by feeding C57BL/6 mice a high-fat diet (HFD) for 12 weeks. Oral administration of AO-I at doses of 20 or 50 mg/kg body weight per day was explored. Lipotoxicity in AML12 hepatocytes was induced by incubation with palmitate for in vitro investigations on the roles of AO-I. The results demonstrated that supplementation with AO-I improved alanine transaminase (ALT), aspartate transaminase (AST), and liver histology in NAFLD mice. Furthermore, AO-I directly alleviated palmitate-induced lipotoxicity by reducing hepatocyte death. Mechanistic studies revealed that AO-I significantly attenuated lipotoxicity through its potential resistance to endoplasmic reticulum (ER) stress and IRE1α-XBP1 splicing both in vivo and in vitro. We identified that AO-I substantially rescued HFD- and palmitate-induced SIRT1 decline in mouse livers and hepatocytes, respectively, thereby improving liver injury and hepatocyte cell death. Mechanistic studies demonstrated that the amelioration of ER stress and lipotoxicity by AO-I supplementation depended on SIRT1 activity. Moreover, AO-I also mitigated IRE1α activation-triggered JNK/p38 MAPK hyperphosphorylation. In conclusion, our findings suggest that AO-I exhibits significant therapeutic effects for improving lipotoxicity in the liver and may be considered as a candidate for NAFLD therapy.

Exertional heat stroke (EHS) is a severe and potentially life-threatening condition that primarily affects healthy individuals engaged in strenuous physical activity under hot and humid environments, representing an escalating public health challenge in the context of global climate change. This study evaluated the preventive potential of a synbiotic formulation containing Lactiplantibacillus plantarum S58 and β-glucan (S58/βG) in a rat model of EHS. Rats received S58/βG pretreatment for 14 or 28 days before EHS induction, and a range of indicators including core temperature, intestinal histopathology, tight junction protein expression, MAPK signaling pathway activation, epithelial apoptosis, and gut microbiota composition were systematically assessed. The results demonstrated that S58/βG effectively enhanced thermotolerance, maintained intestinal integrity, and mitigated systemic organ injury. Mechanistically, it inhibited the phosphorylation of p38 and ERK, attenuated epithelial apoptosis, and maintained the expression of ZO-1, occludin, and E-cadherin. Moreover, S58/βG modulated the gut microbiota by progressively increasing beneficial bacteria and reducing potentially pathogenic bacteria over time. These findings demonstrate, for the first time, that S58/βG confers protection against EHS by concurrently modulating the gut microbiota and MAPK pathway, highlighting its potential as a dietary intervention to prevent EHS in vulnerable populations.

Alcohol-associated bowel disease (ABD), a classic consequence of alcohol abuse, is a significant underlying pathology closely linked to alcohol-related diseases and injuries. The dysfunction of intestinal epithelial cells (IECs) is a primary driver and fundamental pathological basis in ABD; however, the potential mechanisms have not yet been fully elucidated. In this study, ABD model was established by feeding C57BL/6N mice with Lieber DeCarli alcohol diet. Transcriptomics of IECs isolated from ileum was performed to systematically delineate the profiles of genes expression differences in ABD. Gene editing was used to verify the mechanisms underlying alcohol-induced IECs damage. Antioxidant (MitoQ) was administrated to ABD mice to elucidate the concept that oxidative stress was involved in alcohol-stimulated IECs dysfunction. Our results showed that alcohol-fed significantly induced ileal structure and barrier impairment, tight junction proteins loss, intestinal permeability enhancement, and proinflammatory factors expression increase. Transcriptomics analysis revealed that amino acid (AA) transporters and redox-related pathways were the top down- and up-regulated pathways, respectively. Further verification confirmed that alcohol feeding inhibited the uptake capacity of fluorescently labeled-AA by IECs. While decreased AA transporters were positively associated with ileal injury indices. Genetically knocking-down AA transporters, including Slc15a1, Slc6a19, and Slc3a1, aggravated ethanol exposure stimulated tight junction proteins transcriptional repression or cell damage in cultured IECs. While MitoQ intervention reversed alcohol-suppressed expression and activity of AA transporters and further IECs damage. In summary, AA transporters impairment contributes to chronic-plus-binge alcohol intake-induced IECs dysfunction in ABD. Antioxidant treatment might be a promising choice for ABD management.

Heat stroke (HS) is the most severe heat-related emergency, and its pathophysiology remains largely unknown, especially for exertional HS (EHS), which affects younger populations, athletes, and manual workers. Herein, we performed single-cell-transcriptomics, T cell receptor sequencing, and flow cytometry of PBMCs from 9 healthy control participants, 9 patients with heat exhaustion, and 9 patients with EHS to explore complex immunological responses associated with HS pathobiology. We showcased that granzyme-positive T cells and CD56 dim NK cells with high cytotoxicity features and IL-1B + NLRP3 + monocytes with high inflammation and pyroptosis scores were enriched in HS, while the CD161 + T cells with innate immune-like, low cytotoxicity, and clonal expansion features were reduced in HS. Importantly, elevated granzyme-positive T and NK cells might interact with monocytes to induce pyroptosis of hepatic and renal cells and target organ injuries, and blocking the NLRP3 inflammasome pathway prior to the induction could alleviate organ injury in HS. This study offers deeper insights into the pathogenesis of HS, supporting the development of optimal treatment strategies.

Exertional heat stroke (EHS) is a life-threatening condition characterized by hyperthermia and multi-organ dysfunction, often associated with intestinal barrier disruption. This study evaluated the protective effects of Huoxiang Zhengqi Dropping Pills (HXZQD) against EHS in a rat model. HXZQD was administered via oral gavage at low, medium, and high doses, followed by EHS induction through exercise under high-temperature and high-humidity conditions. The findings revealed that high-dose HXZQD significantly delayed the onset of EHS, reduced core body temperature elevations, and mitigated multi-organ injury, as evidenced by biochemical markers and histopathological examination. This study showed that HXZQD alleviated EHS-induced intestinal damage by preserving barrier proteins (ZO-1, Occludin, and Ecadherin) and maintaining intestinal barrier integrity. Transmission electron microscopy confirmed the preservation of tight junction structures. Further analysis indicated that HXZQD modulated the MAPK/NF-κB signaling pathways, inhibiting heat stress-induced activation and reducing inflammation. Additionally, HXZQD positively regulated gut microbiota, increasing the proportion of beneficial Lactococcus and decreasing harmful Streptococcus. These findings suggest that HXZQD maintains intestinal homeostasis during EHS by preserving barrier function and modulating gut microbiota, offering a promising preventive approach for EHS management.

Hepatic lipotoxicity, resulting from excessive lipid accumulation in hepatocytes, plays a central role in the pathogenesis of various metabolic liver diseases. Despite recent progress, the precise mechanisms remain incompletely understood. Using excessive exposure to palmitate in hepatocytes as our primary experimental model and mice studies, we aimed to uncover the mechanisms behind hepatic lipotoxicity, thereby developing potential treatments. Our data reveal for the first time that exposure to palmitate leads to downregulated expression of poly (ADP-ribose) polymerase 1 (PARP-1) in hepatocytes, inhibiting its enzymatic activity. Whereas inhibiting PARP-1 worsens palmitate-induced hepatotoxicity, preventing PARP-1 suppression, using nicotinamide adenine dinucleotide (NAD+) precursors, nicotinamide N-methyltransferase (NNMT) inhibitors, or a poly(ADP-ribose) glycohydrolase (PARG) inhibitor, prevents it. Moreover, we uncover that PARP-1 suppression contributes to palmitate-triggered mechanistic target of rapamycin complex 1 (mTORC1) activation, which has been previously reported by us to contribute to palmitate-induced hepatocyte cell death. Furthermore, our results identify p300 as a downstream target of mTORC1 activation upon palmitate exposure. Importantly, p300 inhibition via either pharmacological or genetic approaches protects against palmitate hepatotoxicity. In addition, we provide evidence that the toll-like receptor 4 (TLR4)-nuclear factor κB (NF-κB) pathway activation in response to palmitate plays a mechanistic role in mediating palmitate-induced PARP-1 downregulation in that both TLR4 antagonist and NF-κB inhibitors prevent palmitate-induced PARP-1 reduction and protect against hepatocyte cell death. In conclusion, our study presents new evidence that the PARP-1-mTORC1-p300 pathway serves as a novel molecular mechanism underlying palmitate-induced hepatic lipotoxicity. Targeting the PARP-1 pathway by increasing cellular NAD+ availability either through its precursor supplementation or by inhibiting its degradation represents a promising therapeutic approach for treating hepatic lipotoxicity.NEW & NOTEWORTHY This study explores the mechanisms of palmitate-induced hepatotoxicity, highlighting the role of PARP-1 downregulation in triggering the mTORC1-p300 pathway and resultant hepatocyte cell death. It further reveals that enhancing cellular NAD+ levels through either precursor supplementation or NNMT inhibitors prevents lipotoxicity by restoring PARP-1 activity. Finally, the study identifies that the TLR4-NF-κB activation mediates palmitate-induced PARP-1 suppression and offers potential therapeutic insights for metabolic liver diseases caused by lipotoxicity.

Heat stroke (HS) is a severe condition associated with prolonged exposure to high temperatures, leading to systemic inflammation and multi-organ damage. Disruption of the intestinal barrier plays a crucial role in HS progression by allowing endotoxins to enter circulation and trigger widespread inflammation. Lactoferrin (LF), known for its immune-modulating and barrier-protective properties, shows promise in mitigating HS-related damage. This study investigated the protective effects of LF on the intestinal barrier under HS conditions using in vitro and in vivo models. In Caco-2 cell monolayers exposed to heat stress, LF improved barrier integrity by increasing transepithelial electrical resistance (TEER) and maintaining tight junctions. In mice, LF supplementation enhanced heat tolerance, delayed HS onset, and reduced multi-organ damage, whereas LF knockout (KO) mice exhibited exacerbated damage. Further analysis revealed that heat stress induced ferroptosis by suppressing GPX4 and SLC7A11, contributing to intestinal injury. LF alleviated ferroptosis by upregulating these proteins and reducing Fe2+ accumulation, reactive oxygen species (ROS) generation, and lipid peroxidation. Additionally, LF inhibited heat stress-induced activation of the MAPK pathway by suppressing the phosphorylation of p38, ERK, and JNK, suggesting that MAPK signaling mediates its protective effects. These findings suggest that LF supplementation may enhance heat tolerance and prevent HS-induced intestinal barrier damage through ferroptosis inhibition and MAPK pathway regulation.

Importance:Image-guided thermal ablation has been administered for patients with T1N0M0 papillary thyroid carcinoma (PTC) who elect to not undergo surgery or receive active surveillance. Considering the indolent nature of PTC, long-term outcomes of ablation are needed. Objective:To investigate l0-year outcomes of thermal ablation in treating T1N0M0 PTC. Design, Setting, and Participants:This multicenter study was conducted at 4 university-affiliated hospitals in China and included 179 consecutive patients with T1N0M0 PTC (median [IQR] volume, 88.0 [163.2] mm3) who underwent thermal ablation between June 2010 and March 2014. Patients who were ineligible to undergo surgery or elected not to were included, and patients had PTC tumors that were smaller than 20 mm as confirmed by biopsy; no clinical or imaging evidence of extrathyroidal extension, lymph node metastasis (LNM), or distant metastasis; and no history of neck irradiation. Main Outcomes and Measures:The primary outcomes were disease progression (LNM, newly developed tumors, persistent tumors, and distant metastasis) and disease-free survival (DFS). Secondary outcomes were technical success, volume reduction rate, tumor disappearance, complications, and delayed surgery. DFS was calculated using a Kaplan-Meier analysis. Results:Among the 179 patients, the mean (SD) age was 45.8 (12.7) years, and 118 (65.9%) were female. During a mean (SD) follow-up period of 120.8 (10.8) months, disease progression was found in 11 of 179 patients (6.1%), including LNM in 4 patients (2.2%), newly developed tumors in 6 patients (3.3%), and persistent tumor in 1 patient (0.6%). The 10-year DFS was 93.9%. The technical success, median volume reduction rate, and tumor disappearance rate was 100%, 100%, and 97.2%, respectively. The magnitude of the disease progression (6.1% vs 7.1%; difference, 1.0%; 95% CI, -6.5% to 25.6%) and DFS (93.9% vs 92.9%; difference, 1.0%, 95% CI, -6.5% to 25.6%) between patients with T1a and T1b tumors was small. The difference in the rate of tumor disappearance between T1a and T1b tumors was large (99.4% vs 71.4%; difference, 28.0%; 95% CI, 10.9%-54.0%). One patient experienced transient voice hoarseness (0.6%). Because of anxiety, 1 patient underwent delayed surgery (0.6%). Conclusions and Relevance:The results of this 10-year multicenter cohort study suggest that thermal ablation is an effective and safe alternative for patients with T1N0M0 PTC who do not undergo surgery or receive active surveillance. For safe and effective treatment, accurate radiologic evaluation, an understanding of ablation techniques, and experienced physicians are recommended.

BACKGROUND:Exertional heat stroke (EHS) is acknowledged as a leading cause of sudden death among athletes and manual laborers. Rapid cooling treatment serves as a primary strategy for preventing and managing exercise-induced hyperthermia, underscoring the growing demand for swift and effective cooling devices. OBJECTIVES:This study aimed to evaluate the cooling performance and effectiveness of a lightweight water circulation cooling blanket. METHODS:This randomized crossover design study recruited 12 male volunteers engaged in professional endurance exercise training outdoors. Participants completed a 3-km exercise session in a hot chamber (ambient temperature ≈ 40 °C; relative humidity [RH] ≈ 40 %), which was immediately followed by a 15-min cooling test (ambient temperature ≈ 31 °C; RH ≈ 65 %). During this cooling phase, participants were assigned to either lay on a lightweight water circulation cooling blanket (LWC condition) or to receive natural cooling (CON condition). Subsequently, all participants were required to complete another 3-km endurance exercise and a subsequent cooling phase under the same environmental conditions as those in the first round. The analysis included measurements of core temperature (Tcore), heart rate (HR), skin temperature (Tskin), thermal sensation, and rating of perceived exertion (RPE). RESULTS:After two cooling sessions, Tcore measurements were significantly reduced in the LWC condition during the cooling phase. The peak Tcore recorded after the second exercise session was lower in the LWC condition compared to the CON condition. Additionally, HR, Tskin, thermal sensation, and RPE were lower in the LWC condition than those observed in the CON condition post-cooling. CONCLUSIONS:The lightweight water circulation cooling blanket demonstrated effective cooling capabilities and enhanced recovery of both HR and RPE. This device may serve as a practical solution for emergency cooling following intense exercise or during recovery intervals between training sessions in extreme heat, benefiting both athletes and manual laborers.

BACKGROUND:Exertional heat stroke (EHS) is a life-threatening condition induced by high-temperature environments, which poses significant health risks. The Qingshu Yiqi Decoction (QSYQD) is a traditional Chinese medicine (TCM) formula that is known to clear summer heat, replenish Qi, and protect intestinal function. However, experimental evidence and mechanistic insights into its role in EHS prevention are limited. PURPOSE:The aim of this study is to evaluate the protective effects of QSYQD against EHS-induced hyperthermia and multi-organ injury and investigate the underlying molecular mechanisms. METHODS:An EHS rat model was established using treadmill exercise in a high-temperature environment. The rats were pretreated with low, medium, or high doses of the QSYQD. The intestinal barrier function was assessed using biomarkers, an ultrastructural analysis, and key protein expressions. Network pharmacology was used to identify potential targets, and molecular mechanisms were validated using western blot. The gut microbiota composition was analyzed using 16S rRNA sequencing. RESULTS:The QSYQD significantly alleviated hyperthermia and multi-organ injury in the EHS rats, and the medium dose had the most pronounced effects. It preserved intestinal barrier integrity by maintaining the tight junction protein levels and reducing biomarkers of epithelial damage. Network pharmacology and experimental validation revealed that the QSYQD inhibited the NF-κB and MLC signaling pathways, key regulators of intestinal barrier function. Additionally, the QSYQD altered the gut microbiota composition and notably increased the abundance of Lactobacillus, that has known heat stress protective effects. CONCLUSIONS:QSYQD protects against EHS-induced damage by preserving intestinal barrier integrity, modulating gut microbiota, and inhibiting the NF-κB and MLC signaling pathways. Future studies will focus on identifying active compounds within QSYQD that enhance heat tolerance and provide EHS protection.

With the intensifying trend of global warming, heat stroke (HS) has emerged as a growing public health threat. HS is a life-threatening condition triggered by excessive core body temperature (CBT), often leading to multi-organ failure and high mortality. Sleep deprivation (SD), a common physiological state among individuals frequently exposed to hot environments—such as military personnel and manual laborers—has been clinically associated with increased HS susceptibility. However, its underlying mechanisms remain unclear. In this study, we demonstrate that acute SD significantly reduces thermotolerance and exacerbates HS-induced multi-organ injury in rats. Notably, SD intensifies intestinal barrier disruption by activating the MAPK signaling pathway and promoting epithelial apoptosis. These findings suggest that SD may heighten HS vulnerability through intestinal barrier dysfunction. Our results align with clinical observations and identify intestinal integrity as a potential intervention target. Ensuring sufficient sleep and maintaining intestinal barrier function may represent key strategies to prevent HS, particularly in heat-exposed populations.

ObjectiveTo evaluate the impact of Nafamostat mesylate (NM) in improving survival outcomes among rats subjected to exertional heat stroke.MethodsThis study involved a cohort of 45 specific pathogen-free (SPF) male Sprague Dawley (SD) rats. After successfully inducing exertional heat stroke, the rats were randomly divided into three groups: the Control group (Con, n = 15), the Exertional Heat Stroke group (EHS, n = 15), and the Nafamostat Mesylate group (NM, n = 15). A subset of ten rats from each group was selected for a 72-h survival analysis. Three hours following the successful establishment of the model, blood samples were collected under anesthesia for comprehensive analysis. This included routine hematological tests, coagulation assessments, and quantitative proteomics analysis, which were later validated using Parallel Reaction Monitoring (PRM). Additionally, tissue samples were harvested from the brain, heart, lung, kidney, liver, and duodenum of rats in each group for subsequent pathological examination.ResultsThe 72-h survival rate in the NM group was markedly higher than that observed in the EHS group. Pathological assessments indicated a notable reduction in thrombus formation within the brain, lungs, and liver in the NM group when compared to the EHS group. Furthermore, the NM group exhibited an elevated platelet count and a significant reduction in prothrombin time (PT) and activated partial thromboplastin time (APTT) relative to the EHS group. Proteomic profiling identified a total of 1,971 differentially expressed proteins, with 160 proteins being downregulated and 52 upregulated in the NM group as compared to the EHS group. PRM validation confirmed that the NM group significantly dampened the expression levels of key differential proteins, including ribosomal protein P2 (rpLP2), Histone 4c16 (H4c16), neutrophilic granule protein (NGP), and inositol monophosphatase 1 (Impa1), which are implicated in anti-inflammatory responses, suppression of immune-mediated thrombosis, and enhancement of cellular metabolism.ConclusionNM mitigates coagulopathy, alleviates thrombus burden, and improves the 72-h survival rate in EHS rats through the modulation of differentially expressed proteins, specifically rpLP2, H4c16, NGP, and Impa1.

Acetaminophen (APAP) overdose is a leading cause of drug-induced liver injury (DILI), and glutathione S-transferase mu 1 (GSTM1), a phase II enzyme involved in drug metabolism and detoxification, is associated with DILI incidence. However, the role of hepatic GSTM1 in APAP-induced liver injury is not fully understood. This study aimed to explore the role of hepatic GSTM1 in APAP-induced hepatotoxicity. A mouse model of APAP-induced liver injury was employed, and liver-specific GSTM1 and/or miR-743a-3p silencing was achieved via adeno-associated virus-8 (AAV8)-mediated RNA delivery. Our data showed that GSTM1 expression was significantly downregulated in APAP-treated mice. APAP induced miR-743a-3p upregulation, which directly suppressed GSTM1 expression, exacerbating liver injury, oxidative damage, and inflammation. Liver-specific miR-743a-3p knockdown rescued GSTM1 knockdown and mitigated liver injury. Mechanistically, GSTM1 interacted with Sp1 and promoted its S-glutathionylation. Loss of GSTM1 decreased Sp1 S-glutathionylation, impairing its nuclear translocation and transcriptional activity, which interfered with sirtuin 1 (SIRT1) expression. Activation of SIRT1 significantly alleviated GSTM1 knockdown-induced liver injury. Our findings suggest that miR-743a-3p-mediated GSTM1 silencing exacerbates APAP-induced liver injury through disruption of the GSTM1-Sp1-SIRT1 axis.

Nicotinamide mononucleotide adenylyltransferase 1 (NMNAT1), a nicotinamide adenine dinucleotide (NAD+) synthetase in Preiss-Handler and salvage pathways, governs nuclear NAD+ homeostasis. This study investigated the role of NMNAT1 in alcohol-associated liver disease (ALD). Decreased NMNAT1 expression and activity were observed in the liver of patients with alcohol-associated hepatitis and either liver or primary hepatocytes from ALD mice. F-box and WD repeat domain containing 7 (FBXW7)-regulated interferon regulatory factor 1 (IRF1) ubiquitination degradation contributed to the alcohol-inhibited NMNAT1 transcriptional level. Hepatic NMNAT1 knockout aggravated alcohol-induced hepatic NAD+ decline and further hepatic steatosis and liver injury. Metabolomics and transcriptomics interaction revealed that the cysteine sulfinic acid decarboxylase (CSAD)-regulated taurine pathway was involved in NMNAT1-disrupted hepatic lipid metabolism in ALD. Hepatic CSAD overexpression or taurine supply attenuated hepatic NMNAT1 knockout-aggravated ALD. Hepatic NMNAT1 loss inhibited NMN-protected ALD. Replenishing hepatic NMNAT1 reversed liver lipid accumulation in ALD mice. These findings identified NMNAT1 as a promising therapeutic target for ALD.

ABSTRACT This study aims to identify the risk factors associated with clinical outcomes and the proteomic changes in organs related to fatal SARS‐CoV‐2 infection within the super‐elderly population. This retrospective analysis included all elderly individuals with COVID‐19 admitted to the Second Medical Center of PLA General Hospital from December 2022 to January 2023. The follow‐up period ended on March 30, 2023. During this time, epidemiological, demographic, laboratory, and outcome data were analyzed descriptively. Proteomic sequencing was performed on super‐elderly patients who died from COVID‐19 at different stages of the disease. A total of 352 elderly COVID‐19 patients, with a mean age of 89.84 ± 8.54 years, were included in this study. During a median follow‐up period of 98 days, 79 patients died. Deceased patients were older and more likely to have cardiovascular and cerebrovascular diseases, with a lower prevalence of lipid‐lowering therapy. The number of deaths in the acute and post‐acute phases were 34 and 45, respectively. Proteomics data suggest that the immune systems of patients who died in the acute phase underwent a more rapid and severe onslaught. Patients in the post‐acute phase showed higher levels of viral genome replication and a more robust immune response. However, the over‐activation of the immune system led to systemic organ dysfunction. Effective management of comorbidities may improve the prognosis of COVID‐19 in super‐elderly patients. The continuous replication of the SARS‐CoV‐2 virus and its subsequent impact on the immune system are critical determinants of survival time in this demographic.

Alcohol-associated liver disease (ALD) is characterized by the reduction of hepatic nicotinamide adenine dinucleotide (NAD+), which exacerbates hepatic steatosis. The present study was conducted to investigate the protective role of nicotinamide (NAM), a foodborne precursor of NAD+ biosynthesis, in ALD. C57BL/6N mice were employed to establish the ALD model and were administered NAM by gavage. Our results showed that NAM supplementation significantly ameliorated alcohol-induced NAD+ reduction and lipid accumulation in both mice liver and cultured AML-12 hepatocytes and improved lipid metabolism-associated gene disorders. Alcohol-induced liver injury and oxidative stress were also blocked by NAM administration. Further transcriptomics analysis and validation revealed that alcohol-stimulated sphingomyelin phosphodiesterase 3 (SMPD3) was significantly reversed by NAM, along with the reduction of hepatic ceramide levels. Importantly, SMPD3 was upregulated in the livers of ALD patients. Genetically silencing SMPD3 alleviated alcohol-induced lipid accumulation in hepatocytes. ChIP assay identified SMPD3 as a direct downstream target of hypoxia-inducible factor 1 alpha (HIF-1α). Liver-specific Hif1α knockdown reduced the level of hepatic SMPD3 expression in mice. Activation of HIF-1α abolished the prevention of intrahepatic liver lipid deposition by NAM, while SMPD3 knockdown reversed HIF-1α activation-stimulated lipid accumulation, indicating that a HIF-1α-regulated SMPD3 pathway was involved in the beneficial role of NAM. NAM improved liver oxidative stress, while antioxidant MitoQ administration rescued HIF-1α/SMPD3 activation in ALD mice, implying that the antioxidant effect of NAM contributed to its inhibitory role on the HIF-1α/SMPD3 pathway. In conclusion, NAM ameliorates chronic alcohol intake-induced hepatic steatosis by inhibiting SMPD3. This study provides new insights into the mechanistic understanding of ALD and highlights NAM as a therapeutic choice for ALD treatment.