Contributing roles of mitochondrial dysfunction and hepatocyte apoptosis in liver diseases through oxidative stress, post-translational modifications, inflammation, and intestinal barrier dysfunction.

This review provides an update on recent findings from basic, translational, and clinical studies on the molecular mechanisms of mitochondrial dysfunction and apoptosis of hepatocytes in multiple liver diseases, including but not limited to alcohol-associated liver disease (ALD), metabolic dysfunction-associated steatotic liver disease (MASLD), and drug-induced liver injury (DILI). While the ethanol-inducible cytochrome P450-2E1 (CYP2E1) is mainly responsible for oxidizing binge alcohol via the microsomal ethanol oxidizing system, it is also responsible for metabolizing many xenobiotics, including pollutants, chemicals, drugs, and specific diets abundant in n-6 fatty acids, into toxic metabolites in many organs, including the liver, causing pathological insults through organelles such as mitochondria and endoplasmic reticula. Oxidative imbalances (oxidative stress) in mitochondria promote the covalent modifications of lipids, proteins, and nucleic acids through enzymatic and non-enzymatic mechanisms. Excessive changes stimulate various post-translational modifications (PTMs) of mitochondrial proteins, transcription factors, and histones. Increased PTMs of mitochondrial proteins inactivate many enzymes involved in the reduction of oxidative species, fatty acid metabolism, and mitophagy pathways, leading to mitochondrial dysfunction, energy depletion, and apoptosis. Unique from other organelles, mitochondria control many signaling cascades involved in bioenergetics (fat metabolism), inflammation, and apoptosis/necrosis of hepatocytes. When mitochondrial homeostasis is shifted, these pathways become altered or shut down, likely contributing to the death of hepatocytes with activation of inflammation and hepatic stellate cells, causing liver fibrosis and cirrhosis. This review will encapsulate how mitochondrial dysfunction contributes to hepatocyte apoptosis in several types of liver diseases in order to provide recommendations for targeted therapeutics.

Cholinergic signaling via the α7 nicotinic acetylcholine receptor regulates the migration of monocyte-derived macrophages during acute inflammation.

BACKGROUND: The involvement of the autonomic nervous system in the regulation of inflammation is an emerging concept with significant potential for clinical applications. Recent studies demonstrate that stimulating the vagus nerve activates the cholinergic anti-inflammatory pathway that inhibits pro-inflammatory cytokines and controls inflammation. The α7 nicotinic acetylcholine receptor (α7nAChR) on macrophages plays a key role in mediating cholinergic anti-inflammatory effects through a downstream intracellular mechanism involving inhibition of NF-κB signaling, which results in suppression of pro-inflammatory cytokine production. However, the role of the α7nAChR in the regulation of other aspects of the immune response, including the recruitment of monocytes/macrophages to the site of inflammation remained poorly understood. RESULTS: We observed an increased mortality in α7nAChR-deficient mice (compared with wild-type controls) in mice with endotoxemia, which was paralleled with a significant reduction in the number of monocyte-derived macrophages in the lungs. Corroborating these results, fluorescently labeled α7nAChR-deficient monocytes adoptively transferred to WT mice showed significantly diminished recruitment to the inflamed tissue. α7nAChR deficiency did not affect monocyte 2D transmigration across an endothelial monolayer, but it significantly decreased the migration of macrophages in a 3D fibrin matrix. In vitro analysis of major adhesive receptors (L-selectin, ß1 and ß2 integrins) and chemokine receptors (CCR2 and CCR5) revealed reduced expression of integrin αM and αX on α7nAChR-deficient macrophages. Decreased expression of αMß2 was confirmed on fluorescently labeled, adoptively transferred α7nAChR-deficient macrophages in the lungs of endotoxemic mice, indicating a potential mechanism for α7nAChR-mediated migration. CONCLUSIONS: We demonstrate a novel role for the α7nAChR in mediating macrophage recruitment to inflamed tissue, which indicates an important new aspect of the cholinergic regulation of immune responses and inflammation.

Pneumococcal pneumonia and endotoxemia: An experimental and clinical reappraisal.

BACKGROUND: Circulating endotoxins could result from bacterial digestive translocation during sepsis, thus contributing to uncontrolled systemic inflammation, leading in turn to organ dysfunction. We addressed this issue in the setting of severe pneumococcal pneumonia. METHODS: Endotoxemia was measured in a clinically relevant rabbit model of ventilated pneumococcal pneumonia and in 110 patients with bacteraemic pneumonia, using a patented mass spectrometry (LC-MS/MS) method for detection of 3-OH fatty acids (C10, C12, C14, C16 and C18), which are molecules bound to the lipid A motif of LPS. RESULTS: Whereas higher levels of systemic inflammation and organ dysfunctions were found, there was no significant difference in lipopolysaccharide concentrations when infected rabbits were compared to non-infected ones, or when patients were compared to healthy volunteers. CONCLUSIONS: Seemingly, endotoxins do not drive the overwhelming inflammation associated with severe forms of pneumococcal pneumonia.

Thrombomodulin Improves Cognitive Deficits in Heat-Stressed Mice.

BACKGROUND: Thrombomodulin (TM) exerts anticoagulant and anti-inflammatory effects to improve the survival of patients with septic shock. Heat stroke resembles septic shock in many aspects. We tested whether TM would improve cognitive deficits and related causative factors in heat-stressed (HS) mice. METHODS: Adult male mice were exposed to HS (33°C for 2 hours daily for 7 consecutive days) to induce cognitive deficits. Recombinant human soluble TM (1 mg/kg, i.p.) was administered immediately after the first HS trial and then once daily for 7 consecutive days. We performed the Y-maze, novel objective recognition, and passive avoidance tests to evaluate cognitive function. Plasma levels of lipopolysaccharide (LPS), high-mobility group box 1 (HMGB1), coagulation parameters, and both plasma and tissue levels of inflammatory and oxidative stress markers were biochemically measured. The duodenum and hippocampus sections were immunohistochemically stained. The intestinal and blood-brain barrier permeability were determined. RESULTS: Compared with controls, HS mice treated with TM had lesser extents of cognitive deficits, exacerbated stress reactions, gut barrier disruption, endotoxemia, blood-brain barrier disruption, and inflammatory, oxidative, and coagulatory injury to heart, duodenum, and hippocampal tissues, and increased plasma HMGB1. In addition to reducing cognitive deficits, TM therapy alleviated all the abovementioned complications in heat-stressed mice. CONCLUSIONS: The findings suggest that HS can lead to exacerbated stress reactions, endotoxemia, gut barrier disruption, blood-brain barrier disruption, hippocampal inflammation, coagulopathy, and oxidative stress, which may act as causative factors for cognitive deficits. TM, an anti-inflammatory, antioxidant, and anti-coagulatory agent, inhibited heat stress-induced cognitive deficits in mice.

Dietary Anthocyanins Mitigate High-Fat Diet-Induced Hippocampal Inflammation in Mice.

BACKGROUND: Obesity and consumption of high-fat diets (HFD) are associated with intestinal permeabilization and increased paracellular transport of endotoxins, which can promote neuroinflammation. Inflammation can affect the hypothalamic pituitary adrenal (HPA) axis, which controls responses to stress and downregulates the brain-derived neurotrophic factor (BDNF), which can promote anxiety and depression, conditions frequently found in obesity. We previously showed that consumption of anthocyanins (AC) mitigate HFD-induced insulin resistance, intestinal permeability, and inflammation. OBJECTIVES: This study investigated if a dietary supplementation with a cyanidin- and delphinidin-rich extract (CDRE) could counteract HFD/obesity-induced hippocampal inflammation in mice. METHODS: C57BL/6J male mice were fed for 14 wk on one of the following diets: 1) a control diet containing 10% total calories from fat (C), 2) a control diet supplemented with 40 mg AC/kg body weight (BW) (CAC), 3) a HFD containing 60% total calories from fat (lard) (HF), or 4) the HFD supplemented with 2, 20, or 40 mg AC/kg BW (HFA2, HFA20, and HFA40, respectively). In plasma and in the hippocampus, parameters of neuroinflammation and the underlying cause (endotoxemia) and consequences (alterations to the HPA and BDNF downregulation) were measured. RESULTS: Consumption of the HFD caused endotoxemia. Accordingly, hippocampal Tlr4 mRNA levels were 110% higher in the HF group, which were both prevented by CDRE supplementation. Consumption of the HFD also caused: 1) microgliosis and increased expression of genes involved in neuroinflammation, that is, Iba-1, Nox4, Tnfα, and Il-1ß, 2) alterations of HPA axis regulation, that is, with low expression of mineralocorticoid (MR) and glucocorticoid (GR) receptors; and 3) decreased Bdnf expression. Supplementation of HFD-fed mice with CDRE mitigated neuroinflammation, microgliosis, and MR and BDNF decreases. CONCLUSIONS: CDRE supplementation mitigates the negative effects associated with HFD consumption and obesity in mouse hippocampus, in part by decreasing inflammation, improving glucocorticoid metabolism, and upregulating BDNF.

Perception of unfamiliar caregivers during sickness - Using the new Caregiver Perception Task (CgPT) during experimental endotoxemia.

Social withdrawal is a well-established part of sickness behavior, but in some contexts sick animals might gain from keeping close instead of keeping away. For instance, sick individuals are more willing to be near known individuals who can provide care and safety (close others) compared to when healthy. Yet, interactions with some strangers might also be beneficial (i.e., healthcare professionals), but it is not known how sickness interplay with social behavior towards such individuals. Here, we assessed if sickness affects perception of caregivers, and developed a new task, the Caregiver Perception Task (CgPT). Twenty-six participants performed the CgPT, once after an injection of lipopolysaccharide (LPS, 0.8 ng/kg body weight, n = 24), and once after an injection of saline (n = 25), one hour and forty-five minutes post-injection. During the task, participants watched short video clips of three types of caregivers: a healthcare professional taking care of a sick individual, a healthcare professional not taking care of a sick individual, and a non-healthcare professional taking care of their sick adult child or partner. After each video clip, the likability, trustworthiness, professionalism, and willingness to interact with and receive care from the caregiver were rated on visual analogue scales. Results showed that participants injected with saline rated healthcare professionals who did not take care of a sick individual less positively on all aspects compared to healthcare professionals who took care of a sick individual. Moreover, compared to saline, LPS increased the participants' willingness to receive care from healthcare professionals and non-healthcare professionals providing care, but not from healthcare professionals not providing care. Thus, our results indicate that sick individuals may approach unknown individuals with potential to provide care and support.

Ursodeoxycholic and chenodeoxycholic bile acids attenuate systemic and liver inflammation induced by lipopolysaccharide in rats.

Bacterial lipopolysaccharide (LPS) induces general inflammation, by activating pathways involving cytokine production, blood coagulation, complement system activation, and acute phase protein release. The key cellular players are leukocytes and endothelial cells, that lead to tissue injury and organ failure. The aim of this study was to explore the anti-inflammatory, antioxidant, and cytoprotective properties of two bile acids, ursodeoxycholic acid (UDCA) and chenodeoxycholic acid (CDCA) in LPS-induced endotoxemia in rats. The experiment involved six distinct groups of Wistar rats, each subjected to different pretreatment conditions: control and LPS groups were pretreated with propylene glycol, as a bile acid solvent, while the other groups were pretreated with UDCA or CDCA for 10 days followed by an LPS injection on day 10. The results showed that both UDCA and CDCA reduced the production of pro-inflammatory cytokines: TNF-α, GM-CSF, IL-2, IFNγ, IL-6, and IL-1ß and expression of nuclear factor-κB (NF-κB) induced by LPS. In addition, pretreatment with these bile acids showed a positive impact on lipid profiles, a decrease in ICAM levels, an increase in antioxidant activity (SOD, |CAT, GSH), and a decrease in prooxidant markers (H2O2 and O2-). Furthermore, both bile acids alleviated LPS-induced liver injury. While UDCA and CDCA pretreatment attenuated homocysteine levels in LPS-treated rats, only UDCA pretreatment showed reductions in other serum biochemical markers, including creatine kinase, lactate dehydrogenase, and high-sensitivity troponin I. It can be concluded that both, UDCA and CDCA, although exerted slightly different effects, can prevent the inflammatory responses induced by LPS, improve oxidative stress status, and attenuate LPS-induced liver injury.

The role of gut barrier dysfunction in postoperative complications in liver transplantation: pathophysiological and therapeutic considerations.

PURPOSE: Gut barrier dysfunction is a pivotal pathophysiological alteration in cirrhosis and end-stage liver disease, which is further aggravated during and after the operational procedures for liver transplantation (LT). In this review, we analyze the multifactorial disruption of all major levels of defense of the gut barrier (biological, mechanical, and immunological) and correlate with clinical implications. METHODS: A narrative review of the literature was performed using PubMed, PubMed Central and Google from inception until November 29th, 2023. RESULTS: Systemic translocation of indigenous bacteria through this dysfunctional barrier contributes to the early post-LT infectious complications, while endotoxin translocation, through activation of the systemic inflammatory response, is implicated in non-infectious complications including renal dysfunction and graft rejection. Bacterial infections are the main cause of early in-hospital mortality of LT patients and unraveling the pathophysiology of gut barrier failure is of outmost importance. CONCLUSION: A pathophysiology-based approach to prophylactic or therapeutic interventions may lead to enhancement of gut barrier function eliminating its detrimental consequences and leading to better outcomes for LT patients.

Neo-epitope detection identifies extracellular matrix turnover in systemic inflammation and sepsis: an exploratory study.

BACKGROUND: Sepsis is associated with high morbidity and mortality, primarily due to systemic inflammation-induced tissue damage, resulting organ failure, and impaired recovery. Regulated extracellular matrix (ECM) turnover is crucial for maintaining tissue homeostasis in health and in response to disease-related changes in the tissue microenvironment. Conversely, uncontrolled turnover can contribute to tissue damage. Systemic Inflammation is implicated to play a role in the regulation of ECM turnover, but the relationship between the two is largely unclear. METHODS: We performed an exploratory study in 10 healthy male volunteers who were intravenously challenged with 2 ng/kg lipopolysaccharide (LPS, derived from Escherichia coli) to induce systemic inflammation. Plasma samples were collected before (T0) and after (T 1 h, 3 h, 6 h and 24 h) the LPS challenge. Furthermore, plasma was collected from 43 patients with septic shock on day 1 of ICU admission. Circulating neo-epitopes of extracellular matrix turnover, including ECM degradation neo-epitopes of collagen type I (C1M), type III (C3M), type IV (C4Ma3), and type VI (C6M), elastin (ELP-3) and fibrin (X-FIB), as well as the ECM synthesis neo-epitopes of collagen type III (PRO-C3), collagen type IV (PRO-C4) and collagen type VI (PRO-C6) were measured by ELISA. Patient outcome data were obtained from electronic patient records. RESULTS: Twenty-four hours after LPS administration, all measured ECM turnover neo-epitopes, except ELP-3, were increased compared to baseline levels. In septic shock patients, concentrations of all measured ECM neo-epitopes were higher compared to healthy controls. In addition, concentrations of C6M, ELP-3 and X-FIB were higher in patients with septic shock who ultimately did not survive (N = 7) compared to those who recovered (N = 36). CONCLUSION: ECM turnover is induced in a model of systemic inflammation in healthy volunteers and was observed in patients with septic shock. Understanding interactions between systemic inflammation and ECM turnover may provide further insight into mechanisms underlying acute and persistent organ failure in sepsis.

Lactobacillus strains reduce the toxic effects of a subchronic exposure to arsenite through drinking water.

The aim of the present study was to determine the efficacy of LAB strains in reducing the intestinal toxicity of arsenite [As(III)] and its tissue accumulation. For this purpose, Balb/c mice were randomly separated in four groups. One group received no treatment (control), one group received only As(III) (30 mg/L) via drinking water and the remaining two groups received As(III) via water and a daily dose of two LAB strains (Lactobacillus intestinalis LE1 and Lacticaseibacillus paracasei BL23) by gavage during 2 months. The results show that both strains reduce the pro-inflammatory and pro-oxidant response observed at the colonic level, partially restore the expression of the intercellular junction proteins (CLDN3 and OCLN) responsible for the maintenance of epithelial integrity, and increase the synthesis of the major mucin of the colonic mucus layer (MUC2), compared to animals treated with As(III) alone. Microbial metabolism of short-chain fatty acids also undergoes a recovery and the levels of fatty acids in the lumen reach values similar to those of untreated animals. All these positive effects imply the restoration of mucosal permeability, and a reduction of the marker of endotoxemia LPS binding protein (LBP). Treatment with the bacteria also has a direct impact on intestinal absorption, reducing the accumulation of As in the internal organs. The data suggest that the protective effect may be due to a reduced internalization of As(III) in intestinal tissues and to a possible antioxidant and anti-inflammatory activity of the bacteria through activation of pathways such as Nrf2 and IL-10. In vitro tests show that the protection may be the result of the combined action of structural and metabolic components of the LAB strains.

Inhibiting the CB1 receptor in CIH-induced animal model alleviates colon injury.

Obstructive sleep apnea (OSA) can lead to intestinal injury, endotoxemia, and disturbance of intestinal flora. Additionally, as a crucial component of the endocannabinoid system, some studies have demonstrated that cannabinoid 1 (CB1) receptors are closely linked to the multiple organ dysfunction triggered by OSA. However, the role of the CB1 receptor in alleviating OSA-induced colon injury remains unclear. Here, through the construction of the OSA classic model, we found that the colon tissue of chronic intermittent hypoxia (CIH)-induced mice exhibited an overexpression of the CB1 receptor. The results of hematoxylin-eosin staining and transmission electron microscopy revealed that inhibition of the CB1 receptor could decrease the gap between the mucosa and muscularis mucosae, alleviate mitochondrial swelling, reduce microvilli shedding, and promote the recovery of tight junctions of CIH-induced mice. Furthermore, CB1 receptor inhibition reduced the levels of metabolic endotoxemia and inflammatory responses, exhibiting significant protective effects on the colon injury caused by CIH. At the molecular level, through western blotting and real-time polymerase chain reaction techniques, we found that inhibiting the CB1 receptor can significantly increase the expression of ZO-1 and Occludin proteins, which are closely related to the maintenance of intestinal mucosal barrier function. Through 16S rRNA high-throughput sequencing and short-chain fatty acid (SCFA) determination, we found that inhibition of the CB1 receptor increased the diversity of the microbial flora and controlled the makeup of intestinal flora. Moreover, butyric acid concentration and the amount of SCFA-producing bacteria, such as Ruminococcaceae and Lachnospiraceae, were both markedly elevated by CB1 receptor inhibition. The results of the spearman correlation study indicated that Lachnospiraceae showed a positive association with both ZO-1 and Occludin but was negatively correlated with the colon CB1 receptor, IL-1ß, and TNF-α. According to this study, we found that inhibiting CB1 receptor can improve CIH-induced colon injury by regulating gut microbiota, reducing mucosal damage and promoting tight junction recovery. KEY POINTS: •CIH leads to overexpression of CB1 receptor in colon tissue. •CIH causes intestinal flora disorder, intestinal mucosal damage, and disruption of tight junctions. •Inhibition of CB1 receptor can alleviate the colon injury caused by CIH through regulating the gut microbiota, reducing mucosal injury, and promoting tight junction recovery.

Mogroside Alleviates Diabetes Mellitus and Modulates Intestinal Microflora in Type 2 Diabetic Mice.

Mogroside, the main component of Siraitia grosvenorii (Swingle) C. Jeffrey (Cucurbitaceae) is a natural product with hypoglycemic and intestinal microbiota regulating properties. However, whether the alteration of intestinal microbiota is associated with the antidiabetic effect of mogroside remains poorly understood. This study investigated the mechanism underlying the hypoglycemic effect of mogroside in regulating intestinal flora and attenuating metabolic endotoxemia. Kunming mice with type 2 diabetes mellitus (T2DM) induced by high-fat diet and intraperitoneal injection of streptozotocin were randomly divided into model, pioglitazone (2.57 mg/kg) and mogroside (200, 100, and 50 mg/kg) groups. After 28 d of administration, molecular changes related to glucose metabolism and metabolic endotoxemia in mice were evaluated. The levels of insulin receptor substrate-1 (IRS-1), cluster of differentiation 14 (CD14) and toll-like receptor 4 (TLR4) mRNAs were measured, and the composition of intestinal microflora was determined by 16s ribosomal DNA (rDNA) sequencing. The results showed that mogroside treatment significantly improved hepatic glucose metabolism in T2DM mice. More importantly, mogroside treatment considerably reduced plasma endotoxin (inhibition rate 65.93%, high-dose group) and inflammatory factor levels, with a concomitant decrease in CD14 and TLR4 mRNA levels. Moreover, mogroside treatment reduced the relative abundance of Firmicutes and Proteobacteria (the inhibition rate of Proteobacteria was 85.17% in the low-dose group) and increased the relative abundance of Bacteroidetes (growth rate up to 40.57%, high-dose group) in the intestines of diabetic mice. This study reveals that mogroside can relieve T2DM, regulating intestinal flora and improving intestinal mucosal barrier, indicating that mogroside can be a potential therapeutic agent or intestinal microbiota regulator in the treatment of T2DM.

Inactivation of Mst/Nrf2/Keap1 signaling flexibly mitigates MAPK/NQO-HO1 activation in the reproductive axis of experimental fluorosis.

Fluoride induced reprotoxicity through oxidative stress-mediated reproductive cell death. Hence, the current study evaluated the importance of the MST/Nrf2/MAPK/NQO-HO1 signaling pathway in fluorosis-induced reproductive toxicity. For this purpose, the reproductive toxicity of sodium fluoride (NaF) at physiological, biochemical, and intracellular levels was evaluated. In-vivo, NaF at 100 mg/L instigated physiological dysfunction, morphological, stereological, and structural injuries in the gut-gonadal axis of fluorosis mice through weakening the antioxidant signaling, Nrf2/HO-1/NQO1signaling pathway, causing the gut-gonadal barrier disintegrated via oxidative stress-induced inflammation, mitochondrial damage, apoptosis, and autophagy. Similar trends were also observed in-vitro in the isolated Leydig cells (LCs) challenging with 20 mg/L NaF. Henceforth, activating the cellular antioxidant signaling pathway, Nrf2/HO-1/NQO1, inactivating autophagy and apoptosis, or attenuating lipopolysaccharide (LPS) can be the theoretical basis and valuable therapeutic targets for coping with NaF-induced reproductive toxicity.

The natural sesquiterpene lactone inulicin suppresses the production of pro-inflammatory mediators via inhibiting NF-κB and AP-1 pathways in LPS-activated macrophages.

OBJECTIVE: Inulicin is a sesquiterpene lactone in Inulae Flos which is clinically used for the treatment of inflammatory diseases, such as cough, sputum production, and vomiting. This study aimed to demonstrate the anti-inflammatory activity and the underlying mechanism of inulicin by using lipopolysaccharide (LPS)-induced in vitro and in vivo models. METHODS: LPS-stimulated RAW264.7 macrophages and mouse peritoneal macrophages (MPMs) were used for evaluating the in vitro anti-inflammatory activity of inulicin, while endotoxemia mice were used for evaluating its in vivo action. Cytokines' levels were determined by ELISA. RT-qPCR and western blot were used for assaying the mRNA and protein levels of target genes. RAW264.7 macrophages transfected with reporter plasmid pNFκB-TA-luc or pAP1-TA-luc were used for assaying the activation of NF-κB or AP-1 signaling. RESULTS: Inulicin significantly inhibited LPS-induced production of NO, IL-6, c-c motif chemokine ligand 2 (CCL2), and IL-1ß in both RAW264.7 cells and MPMs. Mechanism study indicated that it could suppress inducible nitric oxide synthase, IL-6, CCL2, and IL-1ß mRNA levels in LPS-stimulated RAW264.7 cells. Moreover, inulicin inhibited IκBα phosphorylation and prevented the nuclear translocation of p65, thereby inactivating NF-κB signaling. Concurrently, it also inhibited AP-1 signaling by reducing the phosphorylation of C-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK). In endotoxemia mice, a single intraperitoneal administration of inulicin could decrease the production of pro-inflammatory cytokines in serum and peritoneal lavage fluid. CONCLUSIONS: The present study demonstrates that inulicin possesses anti-inflammatory effects in vitro and in vivo, which suggests that inulicin might be a promising candidate for the treatment of inflammatory diseases.

Metabolic Endotoxemia: From the Gut to Neurodegeneration.

Metabolic endotoxemia is a severe health problem for residents in developed countries who follow a Western diet, disrupting intestinal microbiota and the whole organism's homeostasis. Although the effect of endotoxin on the human immune system is well known, its long-term impact on the human body, lasting many months or even years, is unknown. This is due to the difficulty of conducting in vitro and in vivo studies on the prolonged effect of endotoxin on the central nervous system. In this article, based on the available literature, we traced the path of endotoxin from the intestines to the blood through the intestinal epithelium and factors promoting the development of metabolic endotoxemia. The presence of endotoxin in the bloodstream and the inflammation it induces may contribute to lowering the blood-brain barrier, potentially allowing its penetration into the central nervous system; although, the theory is still controversial. Microglia, guarding the central nervous system, are the first line of defense and respond to endotoxin with activation, which may contribute to the development of neurodegenerative diseases. We traced the pro-inflammatory role of endotoxin in neurodegenerative diseases and its impact on the epigenetic regulation of microglial phenotypes.

Central activation of the fatty acid sensor GPR120 suppresses microglia reactivity and alleviates sickness- and anxiety-like behaviors.

G protein-coupled receptor 120 (GPR120, Ffar4) is a sensor for long-chain fatty acids including omega-3 polyunsaturated fatty acids (n-3 PUFAs) known for beneficial effects on inflammation, metabolism, and mood. GPR120 mediates the anti-inflammatory and insulin-sensitizing effects of n-3 PUFAs in peripheral tissues. The aim of this study was to determine the impact of GPR120 stimulation on microglial reactivity, neuroinflammation and sickness- and anxiety-like behaviors by acute proinflammatory insults. We found GPR120 mRNA to be enriched in  both murine and human microglia, and in situ hybridization revealed GPR120 expression in microglia of the nucleus accumbens (NAc) in mice. In a manner similar to or exceeding n-3 PUFAs, GPR120 agonism (Compound A, CpdA) strongly attenuated lipopolysaccharide (LPS)-induced proinflammatory marker expression in primary mouse microglia, including tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß), and inhibited nuclear factor-ĸB translocation to the nucleus. Central administration of CpdA to adult mice blunted LPS-induced hypolocomotion and anxiety-like behavior and reduced TNF-α, IL-1ß and IBA-1 (microglia marker) mRNA in the NAc, a brain region modulating anxiety and motivation and implicated in neuroinflammation-induced mood deficits. GPR120 agonist pre-treatment attenuated NAc microglia reactivity and alleviated sickness-like behaviors elicited by central injection TNF-α and IL-1ß. These findings suggest that microglial GPR120 contributes to neuroimmune regulation and behavioral changes in response to acute infection and elevated brain cytokines. GPR120 may participate in the protective action of n-3 PUFAs at the neural and behavioral level and offers potential as treatment target for neuroinflammatory conditions.

PI3K/AKT and STAT3 pathways mediate the neuroprotective effect of dasatinib from acute cerebral injury in endotoxemic mice.

Background and purpose: Sepsis induces brain dysfunction and there is still a requirement for an unemployed viable restorative approach. This study aimed to investigate the role of dasatinib in the modulation of proinflammatory mediators, attenuating neuroinflammatory response, and it's signaling pathway during endotoxemia. Experimental approach: Twenty-four adult male Swiss-albino mice were randomized into four groups: sham (undergo laparotomy without cecal ligation and puncture, sepsis (laparotomy with cecal ligation and puncture), vehicle-dimethyl sulfoxide, dasatinib (20 mg/kg/day) intraperitoneally. Brain tissue used for assessment of interleukin (IL)-6, IL-1ß, tumor necrosis factor-alpha (TNF-α), IL-10, Toll-like receptor 4 (TLR4), protein kinase B (AKT), phosphoinositide 3-kinases (PI3K), signal transducer and activator of transcription 3 (STAT3), and histopathological examination. Findings/Results: Brain tissue levels of TNF-α, IL-6, and IL1 ß were higher in the sepsis group than in the sham and vehicle groups. The dasatinib group had considerably lower tissue levels of these markers and significantly higher tissue values of IL-10 than the sepsis and vehicle groups. The sham group had much lower tissue values of TLR4, AKT, STAT3, and PI3k than in sepsis and vehicle groups. Furthermore, tissue levels of these markers in the dasatinib group were considerably lower than those in the sepsis and vehicle groups. Histopathology demonstrated that dasatinib might considerably reduce brain damage and the intensity of neuroinflammation when compared to sepsis and vehicle groups that showed extensive brain inflammation and damage. Conclusion and implication: Dasatinib attenuated endotoxemia-induced acute brain damage in mice via modulating effects on TLR4, PI3K, AKT, and STAT3 downstream signaling pathways.

Propofol and Dexmedetomidine Ameliorate Endotoxemia-Associated Encephalopathy via Inhibiting Ferroptosis.

Background: Sepsis is recognized as a multiorgan and systemic damage caused by dysregulated host response to infection. Its acute systemic inflammatory response highly resembles that of lipopolysaccharide (LPS)-induced endotoxemia. Propofol and dexmedetomidine are two commonly used sedatives for mechanical ventilation in critically ill patients and have been reported to alleviate cognitive impairment in many diseases. In this study, we aimed to explore and compare the effects of propofol and dexmedetomidine on the encephalopathy induced by endotoxemia and to investigate whether ferroptosis is involved, finally providing experimental evidence for multi-drug combination in septic sedation. Methods: A total of 218 C57BL/6J male mice (20-25 g, 6-8 weeks) were used. Morris water maze (MWM) tests were performed to evaluate whether propofol and dexmedetomidine attenuated LPS-induced cognitive deficits. Brain injury was evaluated using Nissl and Fluoro-Jade C (FJC) staining. Neuroinflammation was assessed by dihydroethidium (DHE) and DCFH-DA staining and by measuring the levels of three cytokines. The number of Iba1+ and GFAP+ cells was used to detect the activation of microglia and astrocytes. To explore the involvement of ferroptosis, the levels of ptgs2 and chac1; the content of iron, malondialdehyde (MDA), and glutathione (GSH); and the expression of ferroptosis-related proteins were investigated. Conclusion: The single use of propofol and dexmedetomidine mitigated LPS-induced cognitive impairment, while the combination showed poor performance. In alleviating endotoxemic neural loss and degeneration, the united sedative group exhibited the most potent capability. Both propofol and dexmedetomidine inhibited neuroinflammation, while propofol's effect was slightly weaker. All sedative groups reduced the neural apoptosis, inhibited the activation of microglia and astrocytes, and relieved neurologic ferroptosis. The combined group was most prominent in combating genetic and biochemical alterations of ferroptosis. Fpn1 may be at the core of endotoxemia-related ferroptosis activation.

Protective Effect of Silibinin on Lipopolysaccharide-Induced Endotoxemia by Inhibiting Caspase-11-Dependent Cell Pyroptosis.

OBJECTIVE: To explore the protective effect and the underlying mechanism of silibinin (SIB), one of the active compounds from Silybum marianum (L.) Gaertn in endotoxemia. METHODS: Mouse peritoneal macrophage were isolated via intraperitoneally injection of BALB/c mice with thioglycolate medium. Cell viability was assessed using the cell counting kit-8, while cytotoxicity was determined through lactate dehydrogenase cytotoxicity assay. The protein expressions of interleukin (IL)-1 α, IL-1 ß, and IL-18 were determined by enzyme-linked immunosorbent assay. Intracellular lipopolysaccharide (LPS) levels were measured by employing both the limulus amoebocyte lysate assay and flow cytometry. Additionally, proximity ligation assay was employed for the LPS and caspase-11 interaction. Mice were divided into 4 groups: the control, LPS, high-dose-SIB (100 mg/kg), and low-dose-SIB (100 mg/kg) groups (n=8). Zebrafish were divided into 4 groups: the control, LPS, high-dose-SIB (200 εmol/L), and low-dose-SIB (100 εmol/L) groups (n=30 for survival experiment and n=10 for gene expression analysis). The expression of caspase-11, gasdermin D (GSDMD), and N-GSDMD was determined by Western blot and the expressions of caspy2, gsdmeb, and IL-1 ß were detected using quantitative real-time PCR. Histopathological observation was performed through hematoxylineosin staining, and protein levels in bronchoalveolar lavage fluid were quantified using the bicinchoninicacid protein assay. RESULTS: SIB noticeably decreased caspase-11 and GSDMD-mediated pyroptosis and suppressed the secretion of IL-1 α, IL-1 ß, and IL-18 induced by LPS (P<0.05). Moreover, SIB inhibited the translocation of LPS into the cytoplasm and the binding of caspase-11 and intracellular LPS (P<0.05). SIB also attenuated the expression of caspase-11 and N-terminal fragments of GSDMD, inhibited the relative cytokines, prolonged the survival time, and up-regulated the survival rate in the endotoxemia models (P<0.05). CONCLUSIONS: SIB can inhibit pyroptosis in the LPS-mediated endotoxemia model, at least in part, by inhibiting the caspase-11-mediated cleavage of GSDMD. Additionally, SIB inhibits the interaction of LPS and caspase-11 and inhibits the LPS-mediated up-regulation of caspase-11 expression, which relieves caspase-11-dependent cell pyroptosis and consequently attenuates LPS-mediated lethality.

Calcium trafficking and gastrointestinal physiology following an acute lipopolysaccharide challenge in pigs.

The influence of systemic immune activation on whole-body calcium (Ca) trafficking and gastrointestinal tract (GIT) physiology is not clear. Thus, the study objectives were to characterize the effects of lipopolysaccharide (LPS) on Ca pools and GIT dynamics to increase understanding of immune-induced hypocalcemia, ileus, and stomach hemorrhaging. Twelve crossbred pigs [44 ±â€…3 kg body weight (BW)] were randomly assigned to 1 of 2 intramuscular treatments: (1) control (CON; 2 mL saline; n = 6) or (2) LPS (40 µg LPS/kg BW; n = 6). Pigs were housed in metabolism stalls to collect total urine and feces for 6 h after treatment administration, at which point they were euthanized, and various tissues, organs, fluids, and digesta were weighed, and analyzed for Ca content. Data were analyzed with the MIXED procedure in SAS 9.4. Rectal temperature and respiration rate increased in LPS relative to CON pigs (1.4 °C and 32%, respectively; P ≤ 0.05). Inflammatory biomarkers such as circulating alkaline phosphatase, aspartate aminotransferase, and total bilirubin increased in LPS compared with CON pigs whereas albumin decreased (P ≤ 0.02). Plasma glucose and urea nitrogen decreased and increased, respectively, after LPS (43% and 80%, respectively; P < 0.01). Pigs administered LPS had reduced circulating ionized calcium (iCa) compared to CON (15%; P < 0.01). Considering estimations of total blood volume, LPS caused an iCa deficit of 23 mg relative to CON (P < 0.01). Adipose tissue and urine from LPS pigs had reduced Ca compared to CON (39% and 77%, respectively; P ≤ 0.05). There did not appear to be increased Ca efflux into GIT contents and no detectable increases in other organ or tissue Ca concentrations were identified. Thus, while LPS caused hypocalcemia, we were unable to determine where circulating Ca was trafficked. LPS administration markedly altered GIT dynamics including stomach hemorrhaging, diarrhea (increased fecal output and moisture), and reduced small intestine and fecal pH (P ≤ 0.06). Taken together, changes in GIT physiology suggested dyshomeostasis and alimentary pathology. Future research is required to fully elucidate the etiology of immune activation-induced hypocalcemia and GIT pathophysiology.

Lipopolysaccharide (LPS) activates the immune system and this is accompanied with hypocalcemia and altered gastrointestinal tract (GIT) physiology. The study objectives were to characterize whole-body calcium (Ca) trafficking and evaluate GIT dynamics during LPS-induced immune activation. Ca concentrations were analyzed after intramuscular LPS injection. Administering LPS caused marked alterations in metabolic and inflammatory biomarkers and GIT dynamics, characterized by increased lower GIT motility and stomach hemorrhaging. Circulating Ca and adipose tissue and urine Ca output were decreased after LPS. Ca concentrations in other tissues and GIT contents were not detectably different. Thus, we were unable to account for about 110 mg Ca following LPS. Where and how circulating Ca is partitioned during immune activation remains unclear.