Maternal n-3 enriched diet reprograms the offspring neurovascular transcriptome and blunts inflammation induced by endotoxin in the neonate.

Infection during the perinatal period can adversely affect brain development, predispose infants to ischemic stroke and have lifelong consequences. We previously demonstrated that diet enriched in n-3 polyunsaturated fatty acids (n-3 PUFA) transforms brain lipid composition in the offspring and protects the neonatal brain from stroke, in part by blunting injurious immune responses. Critical to the interface between the brain and systemic circulation is the vasculature, endothelial cells in particular, that support brain homeostasis and provide a barrier to systemic infection. Here, we examined whether maternal PUFA-enriched diets exert reprograming of endothelial cell signalling in postnatal day 9 mice after modeling aspects of infection using LPS. Transcriptome analysis was performed on microvessels isolated from brains of pups from dams maintained on 3 different maternal diets from gestation day 1: standard, n-3 enriched or n-6 enriched diets. Depending on the diet, in endothelial cells LPS produced distinct regulation of pathways related to immune response, cell cycle, extracellular matrix, and angiogenesis. N-3 PUFA diet enabled higher immune reactivity in brain vasculature, while preventing imbalance of cell cycle regulation and extracellular matrix cascades that accompanied inflammatory response in standard diet. Cytokine analysis revealed a blunted LPS response in blood and brain of offspring from dams on n-3 enriched diet. Analysis of cerebral vasculature in offspring in vivo revealed no differences in vessel density. However, vessel complexity was decreased in response to LPS at 72 h in standard and n-6 diets. Thus, LPS modulates specific transcriptomic changes in brain vessels of offspring rather than major structural vessel characteristics during early life. N-3 PUFA-enriched maternal diet in part prevents an imbalance in homeostatic processes, alters inflammation and ultimately mitigates changes to the complexity of surface vessel networks that result from infection. Importantly, maternal diet may presage offspring neurovascular outcomes later in life.

Donepezil ameliorates gut barrier disruption in doxorubicin-treated rats.

An impact of donepezil against doxorubicin-induced gut barrier disruption and gut dysbiosis has never been investigated. Twenty-four male Wistar rats were divided into three groups. Each group was treated with either vehicle as a control, doxorubicin, or doxorubicin-cotreated with donepezil. Heart rate variability was assessed to reflect the impact of doxorubicin and donepezil. Then, animals were euthanized, and the ileum and its contents were collected in each case to investigate the gut barrier and gut microbiota, respectively. The microbiota-derived endotoxin, trimethylamine N-oxide (TMAO), and short-chain fatty acids (SCFAs) in the serum were determined. An increase in the sympathetic tone, endotoxins, and TMAO levels with disruption of the gut barrier and a decrease in SCFAs levels were observed in doxorubicin-treated rats. Gut microbiota of doxorubicin-treated rats was significantly different from that of the control group. Donepezil treatment significantly decreased the sympathetic tone, restored the gut barrier, and reduced endotoxin and TMAO levels in doxorubicin-treated rats. Nonetheless, donepezil administration did not alter the gut microbiota profile and levels of SCFAs in doxorubicin-treated rats. Doxorubicin impaired the autonomic balance and the gut barrier, and induced gut dysbiosis, resulting in gut toxicity. Donepezil partially improved the doxorubicin-induced gut toxicity through balancing the autonomic disturbance.

[Nuclear factor E2-related factor 2 attenuates endotoxin-induced acute lung injury by up-regulating cellular tight junction protein Claudin-18 expression].

OBJECTIVE: To investigate the effect of nuclear factor E2-related factor 2 (Nrf2) on the cellular tight junction protein Claudin-18 in endotoxin-induced acute lung injury (ALI). METHODS: Eighteen healthy male C57BL/6 mice were divided into control group, endotoxin-induced ALI model group (ALI group) and Nrf2 activator tert-butylhydroquinone (tBHQ) pretreatment group (tBHQ+ALI group) according to random number table method, with 6 mice in each group. Mice endotoxin-induced ALI model was reproduced by intraperitoneal injection of lipopolysaccharide (LPS, 15 mg/kg), and the mice in the control group was injected with an equal amount of phosphate buffer solution (PBS). The mice in the tBHQ+ALI group received three intraperitoneal injections of tBHQ (a total of 50 mg/kg) at an interval of 1 hour before molding. The last injection of tBHQ was accompanied by LPS of 15 mg/kg. The mice in the control group and model group were given equal amounts of PBS, and PBS or LPS was given at the last injection. The mice were sacrificed at 12 hours after LPS injection to take lung tissues. After the lung tissue was stained with hematoxylin-eosin (HE) staining, the pathological changes were observed under light microscopy, and the lung injury score was calculated. The lung wet/dry ratio (W/D) was determined. Nrf2 protein expression in the lung tissue was detected by Western blotting. Positive expression of Claudin-18 in the lung tissue was determined by immunohistochemistry. RESULTS: The lung tissue showed normal structure, without significant pathological change in the control group. Compared with the control group, the alveolar septum widened accompanied by inflammatory cell infiltration, capillary hyperemia and tissue edema in the ALI group, the lung injury score and lung W/D ratio were significantly increased (lung injury score: 6.50±1.05 vs. 1.83±0.75, lung W/D ratio: 3.79±0.22 vs. 3.20±0.14, both P < 0.01), and the Nrf2 protein expression and Claudin-18 positive expression in the lung tissue were significantly lowered [Nrf2 protein (Nrf2/ß-actin): 0.41±0.33 vs. 1.22±0.33, Claudin-18 (A value): 0.28±0.07 vs. 0.44±0.10, both P < 0.05]. After tBHQ pretreatment, the degree of lung histopathological injury was significantly reduced compared with the ALI group, the alveolar space slightly abnormal, inflammatory cell infiltration and tissue edema reduced, the lung injury score and lung W/D ratio were significantly decreased (lung injury score: 3.00±0.89 vs. 6.50±1.05, lung W/D ratio: 3.28±0.19 vs. 3.79±0.22, both P < 0.01), and Nrf2 protein expression and Claudin-18 positive expression in the lung tissue were significantly increased [Nrf2 protein (Nrf2/ß-actin): 1.26±0.09 vs. 0.41±0.33, Claudin-18 (A valure): 0.45±0.04 vs. 0.28±0.07, both P < 0.05]. CONCLUSIONS: Nrf2 alleviated pulmonary edema and improved endotoxin-induced ALI by up-regulating Claudin-18 expression.

Toxicity of Cry- and Vip3Aa-Class Proteins and Their Interactions against Spodoptera frugiperda (Lepidoptera: Noctuidae).

The fall armyworm (FAW), Spodoptera frugiperda (J.E. Smith), is one of the most important insect pests affecting corn crops worldwide. Although planting transgenic corn expressing Bacillus thuringiensis (Bt) toxins has been approved as being effective against FAW, its populations' resistance to Bt crops has emerged in different locations around the world. Therefore, it is important to understand the interaction between different Bt proteins, thereby delaying the development of resistance. In this study, we performed diet-overlay bioassays to evaluate the toxicity of Cry1Ab, Cry1Ac, Cry1B, Cry1Ca, Cry1F, Cry2Aa, Cry2Ab, Vip3Aa11, Vip3Aa19, and Vip3Aa20, as well as the interaction between Cry1Ab-, Cry1F-, Cry2Ab-, and Vip3Aa-class proteins against FAW. According to our results, the LC50 values of Bt proteins varied from 12.62 ng/cm2 to >9000 ng/cm2 (protein/diet), among which the Vip3Aa class had the best insecticidal effect. The combination of Cry1Ab and Vip3Aa11 exhibited additive effects at a 5:1 ratio. Cry1F and Vip3Aa11 combinations exhibited additive effects at 1:1, 1:2, and 5:1 ratios. The combination of Cry1F and Vip3Aa19 showed an antagonistic effect when the ratio was 1:1 and an additive effect when the ratio was 1:2, 2:1, 1:5, and 5:1. Additionally, the combinations of Cry1F and Vip3Aa20 showed antagonistic effects at 1:2 and 5:1 ratios and additive effects at 1:1 and 2:1 ratios. In addition to the above combinations, which had additive or antagonistic effects, other combinations exhibited synergistic effects, with variations in synergistic factors (SFs). These results can be applied to the establishment of new pyramided transgenic crops with suitable candidates, providing a basis for FAW control and resistance management strategies.

Endotoxin tolerance ameliorates lipopolysaccharide/D-galactosamine-induced acute liver failure by negative regulation of the NF-κB/NLRP3 and activation of Nrf2/HO-1 via Sitr1.

Acute liver failure (ALF) is a potentially fatal disorder characterized by extensive hepatocyte necrosis and rapid decline in liver function. Numerous factors, including oxidative stress, cell death, and inflammatory responses, are associated with its pathogenesis. Endotoxin tolerance (ET) refers to the phenomenon in which the body or cells exhibit low or no response to high-dose lipopolysaccharide (LPS) stimulation after pre-stimulation with low-dose LPS. However, the specific mechanism through which ET regulates LPS/D-galactosamine (D-GalN)-induced ALF remains unclear. An ALF mouse model was established by intraperitoneal injection of D-GalN (400 mg/kg) and LPS (10 mg/kg). A low dose of LPS (0.1 mg/kg/d) was continuously administered to mice for 5 d before modeling to assess the protective effect of ET. The data from this study showed that ET alleviated the inflammatory response in mice with LPS/D-GalN-induced ALF. ET inhibited LPS-induced oxidative damage and pyroptosis in macrophages in vitro. RNA sequencing analysis showed that the NF-κB/NLRP3 pathway was linked to the anti-inflammatory and antioxidative effects of ET. Furthermore, using western blot, RT-qPCR, and immunofluorescence, we verified that ET inhibited the NF-κB/NLRP3 pathway and triggered the Nrf2/HO-1 signaling pathway to attenuate oxidative stress and cell pyroptosis. Sirt1 knockdown reversed this protective effect. In summary, our research elucidates that ET prevents ALF advancement by upregulating Sirt1 levels, triggering the Nrf2/HO-1 signaling axis, and suppressing the NF-κB/NLRP3 signaling cascade to inhibit oxidative stress and cell pyroptosis. Our results provide a mechanistic explanation for the protective effect of ET against ALF.

The endotoxin hypothesis of Alzheimer's disease.

Lipopolysaccharide (LPS) constitutes much of the surface of Gram-negative bacteria, and if LPS enters the human body or brain can induce inflammation and act as an endotoxin. We outline the hypothesis here that LPS may contribute to the pathophysiology of Alzheimer's disease (AD) via peripheral infections or gut dysfunction elevating LPS levels in blood and brain, which promotes: amyloid pathology, tau pathology and microglial activation, contributing to the neurodegeneration of AD. The evidence supporting this hypothesis includes: i) blood and brain levels of LPS are elevated in AD patients, ii) AD risk factors increase LPS levels or response, iii) LPS induces Aß expression, aggregation, inflammation and neurotoxicity, iv) LPS induces TAU phosphorylation, aggregation and spreading, v) LPS induces microglial priming, activation and neurotoxicity, and vi) blood LPS induces loss of synapses, neurons and memory in AD mouse models, and cognitive dysfunction in humans. However, to test the hypothesis, it is necessary to test whether reducing blood LPS reduces AD risk or progression. If the LPS endotoxin hypothesis is correct, then treatments might include: reducing infections, changing gut microbiome, reducing leaky gut, decreasing blood LPS, or blocking LPS response.

The role of ATP-binding cassette transporters in arthropod pesticide toxicity and resistance.

Pesticide resistance in arthropods threatens agricultural productivity and the control of vector-borne diseases. The ATP-binding cassette (ABC) transporters have emerged as important factors in the toxicity of synthetic pesticides, as well as for Bacillus thuringiensis insecticidal Cry protein binding. Depending on the localization of expression, both higher and lower expression of ABCs have been linked with pesticide resistance. The recent development of genetic-based approaches such as RNAi and CRISPR/Cas9 gene editing in nonmodel species, has greatly contributed to unveil their functional importance in pesticide toxicity and resistance. Using these tools, we are now poised to further unravel the molecular genetic mechanisms of gene regulation uncovering more elusive regulatory resistance genes.

Regional pulmonary perfusion, blood volume, and their relationship change in experimental early ARDS.

Regional pulmonary perfusion (Q) has been investigated using blood volume (Fb) imaging as an easier-to-measure surrogate. However, it is unclear if changing pulmonary conditions could affect their relationship. We hypothesized that vascular changes in early acute respiratory distress syndrome (ARDS) affect Q and Fb differently. Five sheep were anesthetized and received lung protective mechanical ventilation for 20 h while endotoxin was continuously infused. Using dynamic 18F-FDG and 13NN Positron Emission Tomography (PET), regional Fb and Q were analysed in 30 regions of interest (ROIs) and normalized by tissue content (Fbn and Qn, respectively). After 20 h, the lung injury showed characteristics of early ARDS, including gas exchange and lung mechanics. PET images of Fbn and Qn showed substantial differences between baseline and lung injury. Lung injury caused a significant change in the Fbn-Qn relationship compared to baseline (p < 0.001). The best models at baseline and lung injury were Fbn = 0.32 + 0.690Qn and Fbn = 1.684Qn-0.538Qn2, respectively. Endotoxine-associated early ARDS changed the relationship between Fb and Q, shifting from linear to curvilinear. Effects of endotoxin exposure on the vasoactive blood flow regulation were most likely the key factor for this change limiting the quantitative accuracy of Fb imaging as a surrogate for regional Q.

Inhaled endotoxin induces a systemic neutrophil response without affecting cardiovascular measures in a randomized cross-over exposure study.

OBJECTIVE: The gram-negative bacterial cell wall component endotoxin (lipopolysaccharide, LPS) is a key component of particulate matter (PM). PM exposure is associated with cardiovascular morbidity and mortality. However, the contribution of individual components of PM to acute and chronic cardiovascular measures is not clear. This study examines whether systemic inflammation induced by LPS inhalation causes acute changes in cardiovascular physiology measures. MATERIALS AND METHODS: In this double blinded, placebo-controlled crossover study, fifteen adult volunteers underwent inhalation exposure to 20,000 EU Clinical Center Reference Endotoxin (CCRE). Peripheral blood and induced sputum neutrophils were obtained at baseline and six hours post-exposure. Blood pressure, measures of left ventricular function (ejection fraction (LVEF) and global longitudinal strain (LVGLS)), and indices of endothelial function (flow mediated dilation (FMD) and velocity time integral during hyperemia (VTIhyp)) were measured before and after treatment. Wilcoxon sign-rank tests and linear mixed models were used for statistical analysis. RESULTS: In comparison with normal saline, LPS inhalation resulted in significant increases in peripheral blood and sputum neutrophils but was not associated with significant alterations in blood pressure, LVGLS, LVEF, FMD, or VTIhyp. DISCUSSION AND CONCLUSIONS: In healthy adults, systemic inflammation after LPS inhalation was not associated with acute changes in cardiovascular physiology. Larger studies are needed to investigate the effects of other PM components on inflammation induced cardiovascular dysfunction.

Occupational exposure to endotoxins and small cell lung cancer: a systematic review with meta-analysis.

The relationship of occupational exposure to endotoxins with different histologic subtypes of lung cancer has not been established. Our objective was to conduct a systematic review with meta-analysis to assess the effect of exposure to endotoxins on the development of small cell lung cancer (SCLC). A bibliographic search was conducted using MEDLINE, Embase, CENTRAL, and Web of Science databases until December 2022, including all cohort and/or case-control studies that examined occupational exposure to endotoxins and SCLC. Risk of bias was assessed using the U.S. Office of Health Assessment and Translation tool. A random effects model was applied, publication bias were assessed, and a sensitivity analysis was conducted. Four papers were selected for meta-analysis purposes. A total of 144 incident cases of SCLC and 897 population or hospital controls were included. Occupational exposure to endotoxins was considered for textile/leather industry and agricultural sector workers exposed to endotoxins originating from wool, cotton, or leather dust. Except for one study, all investigations were classified as having a low probability of risk of biases. The results of the meta-analysis were not statistically significant (pooled OR: 0.86; 95% CI:0.69-1.08). In addition, neither between-study heterogeneity (I2=0%;p=0.92) nor publication bias was observed (p=0.49). The results of the sensitivity analysis, after including five studies that assessed the risk of SCLC among textile industry and crop/livestock farm workers (not specifically exposed to endotoxins), showed a negative statistically non-significant association and low between-study heterogeneity (pooled OR: 0.90; 95% CI:0.79-1.02; I2=22%;p=0.23). Subjects exposed to occupational exposure to endotoxins seem to exhibit a negative association with the development of SCLC, although the results are not conclusive.

Mutational analysis of the transmembrane α4-helix of Bacillus thuringiensis mosquito-larvicidal Cry4Aa toxin.

Cry4Aa, produced by Bacillus thuringiensis subsp. israelensis, exhibits specific toxicity to larvae of medically important mosquito genera. Cry4Aa functions as a pore-forming toxin, and a helical hairpin (α4-loop-α5) of domain I is believed to be the transmembrane domain that forms toxin pores. Pore formation is considered to be a central mode of Cry4Aa action, but the relationship between pore formation and toxicity is poorly understood. In the present study, we constructed Cry4Aa mutants in which each polar amino acid residues within the transmembrane α4 helix was replaced with glutamic acid. Bioassays using Culex pipiens mosquito larvae and subsequent ion permeability measurements using symmetric KCl solution revealed an apparent correlation between toxicity and toxin pore conductance for most of the Cry4Aa mutants. In contrast, the Cry4Aa mutant H178E was a clear exception, almost losing its toxicity but still exhibiting a moderately high conductivity of about 60% of the wild-type. Furthermore, the conductance of the pore formed by the N190E mutant (about 50% of the wild-type) was close to that of H178E, but the toxicity was significantly higher than that of H178E. Ion selectivity measurements using asymmetric KCl solution revealed a significant decrease in cation selectivity of toxin pores formed by H178E compared to N190E. Our data suggest that the toxicity of Cry4Aa is primarily pore related. The formation of toxin pores that are highly ion-permeable and also highly cation-selective may enhance the influx of cations and water into the target cell, thereby facilitating the eventual death of mosquito larvae.

Behavior, synaptic mitochondria, and microglia are differentially impacted by chronic adolescent stress and repeated endotoxin exposure in male and female rats.

Early life adversity and chronic inflammation have both been associated with cognitive impairment and neural compromise. In this study, we investigated the interactions between a history of chronic adolescent stress (CAS) and repeated endotoxin exposure on behavior, synaptic mitochondria, and microglia in adult male and female Wistar rats. Adult rats from chronic stress and control conditions were exposed to either repeated endotoxin (lipopolysaccharide; LPS) or saline injections every 3 days for 9 weeks. In both sexes, repeated LPS, regardless of stress history, impaired working memory in the Y maze. Regarding spatial memory, LPS impaired function for females; whereas, CAS altered function in males. Although males had an increase in anxiety-like behavior shortly after CAS, there were no long-term effects on anxiety-like behavior or social interaction observed in males or females. Stress did not alter synaptic mitochondrial function in either sex. Repeated LPS altered synaptic mitochondrial function such that ATP production was increased in females only. There were no observed increases in IBA-1 positive cells within the hippocampus for either sex. However, LPS and CAS altered microglia morphology in females. Impact of repeated LPS was evident at the terminal endpoint with increased spleen weight in both sexes and decreased adrenal weight in males only. Circulating cytokines were not impacted by repeated LPS at the terminal endpoint, but evidence of CAS effects on cytokines in females were evident. These data suggest a long-term impact of chronic stress and an impact of repeated endotoxin challenge in adulthood; however, not all physiological and behavioral metrics examined were impacted by the paradigm employed in this study and the two environmental challenges rarely interacted.

Bacillus thuringiensis Cry9Aa Insecticidal Protein Domain I Helices α3 and α4 Are Two Core Regions Involved in Oligomerization and Toxicity.

Bacillus thuringiensis Cry9 proteins show high insecticidal activity against different lepidopteran pests. Cry9 could be a valuable alternative to Cry1 proteins because it showed a synergistic effect with no cross-resistance. However, the pore-formation region of the Cry9 proteins is still unclear. In this study, nine mutations of certain Cry9Aa helices α3 and α4 residues resulted in a complete loss of insecticidal activity against the rice pest Chilo suppressalis; however, the protein stability and receptor binding ability of these mutants were not affected. Among these mutants, Cry9Aa-D121R, Cry9Aa-D125R, Cry9Aa-D163R, Cry9Aa-E165R, and Cry9Aa-D167R are unable to form oligomers in vitro, while the oligomers formed by Cry9Aa-R156D, Cry9Aa-R158D, and Cry9Aa-R160D are unstable and failed to insert into the membrane. These data confirmed that helices α3 and α4 of Cry9Aa are involved in oligomerization, membrane insertion, and toxicity. The knowledge of Cry9 pore-forming action may promote its application as an alternative to Cry1 insecticidal proteins.

V-ATPase E mediates Cry2Ab binding and toxicity in Helicoverpa armigera.

Cry2Ab is one of the important alternative Bt proteins that can be used to manage insect pests resistant to Cry1A toxins and to expand the insecticidal spectrum of pyramided Bt crops. Previous studies have showed that vacuolar H+-ATPase subunits A and B (V-ATPase A and B) may be involved in Bt insecticidal activities. The present study investigated the role of V-ATPases subunit E in the toxicity of Cry2Ab in Helicoverpa amigera. RT-PCR analysis revealed that oral exposure of H. amigera larvae to Cry2Ab led to a significant reduction in the expression of H. armigera V-ATPase E (HaV-ATPase E). Ligand blot, homologous and heterologous competition experiments confirmed that HaV-ATPases E physically and specifically bound to activated Cry2Ab toxin. Heterologous expressing of HaV-ATPase E in Sf9 cells made the cell line more susceptible to Cry2Ab, whereas knockdown of the endogenous V-ATPase E in H. zea midgut cells decreased Cry2Ab's cytotoxicity against this cell line. Further in vivo bioassay showed that H. armigera larvae fed a diet overlaid with both Cry2Ab and E. coli-expressed HaV-ATPase E protein suffered significantly higher mortality than those fed Cry2Ab alone. These results support that V-ATPases E is a putative receptor of Cry2Ab and can be used to improve Cry2Ab toxicity and manage Cry2Ab resistance at least in H. armigera.

Cytokine Adsorption Effects of a Novel Hemofiltration Column for the Treatment of Experimental Endotoxemia.

INTRODUCTION: The TKM-101 is a new hemofiltration column packed with a polymer alloy membrane consisting of polyethersulfone, polyvinylpyrrolidone, and sulfonated poly (arylene ether) copolymers. We examined the ability of the TKM-101 column to remove cytokines and humoral mediators from blood in vitro and the effects of extracorporeal treatment with the TKM-101 column on the mortality rate and inflammatory responses to endotoxic shock in vivo. METHODS: In vitro and in vivo laboratory investigations were conducted. In the in vitro experiment, the adsorption abilities of TKM-101, AN69-ST, and control columns for cytokine-related sepsis in blood were compared using human serum samples. In the in vivo experiment, male Sprague-Dawley rats were anesthetized and injected with Escherichia coli endotoxin (15 mg/kg, intravenously). Afterward, the rats were assigned (in a double-blind manner) to one of three groups (n = 17 per group): apheresis with a control column (control group), apheresis with an AN69-ST column (AN69-ST group), or apheresis with a TKM-101 column (TKM-101 group). Outcomes were compared among the groups. RESULTS: In vitro, the concentrations of all evaluated cytokines significantly decreased with the TKM-101 column compared to those with the control column; however, there were no significant differences between the TKM-101 and AN69-ST columns. In vivo, the mortality rates 8 h after endotoxin injection were 65%, 29%, and 29% for the control, AN69-ST, and TKM-101 groups, respectively. Hypotension and elevated plasma cytokine concentrations were less prominent in the TKM-101 and AN69-ST groups compared to those in the control group. CONCLUSIONS: TKM-101 effectively removed proteins of varying sizes, from small-sized proteins such as interleukin (IL)-8 to mid-sized protein such as IL-10 in vitro. Moreover, TKM-101 treatment reduced mortality and had inhibitory effects on inflammatory responses in endotoxemic rats. These findings suggest that TKM-101 treatment may be available for use in patients with sepsis and/or endotoxemia.

Short-term residential exposure to endotoxin emitted from livestock farms in relation to lung function in non-farming residents.

BACKGROUND: Evidence on the public health relevance of exposure to livestock farm emissions is increasing. Research mostly focused on chemical air pollution, less on microbial exposure, while endotoxins are suggested relevant bacterial components in farm emissions. Acute respiratory health effects of short-term exposure to livestock-related air pollution has been shown for NH3 and PM10, but has not yet been studied for endotoxin. We aimed to assess associations between lung function and short-term exposure to livestock farming emitted endotoxin in co-pollutant models with NH3 and PM10. METHODS: In 2014/2015, spirometry was conducted in 2308 non-farming residents living in a rural area in the Netherlands. Residential exposure to livestock farming emitted endotoxin during the week prior to spirometry was estimated by dispersion modelling. The model was applied to geo-located individual barns within 10 km of each home address using provincial farm data and local hourly meteorological conditions. Regional week-average measured concentrations of NH3 and PM10 were obtained through monitoring stations. Lung function parameters (FEV1, FVC, FEV1/FVC, MMEF) were expressed in %-predicted value based on GLI-2012. Exposure-response analyses were performed by linear regression modelling. RESULTS: Week-average endotoxin exposure was negatively associated with FVC, independently from regional NH3 and PM10 exposure. A 1.1% decline in FVC was estimated for an increase of endotoxin exposure from 10th to 90th percentile. Stratified analyses showed a larger decline (3.2%) for participants with current asthma and/or COPD. FEV1 was negatively associated with week-average endotoxin exposure, but less consistent after co-pollutant adjustment. FEV1/FVC and MMEF were not associated with week-average endotoxin exposure. CONCLUSIONS: Lower lung function in non-farming residents was observed in relation to short-term residential exposure to livestock farming emitted endotoxin. This study indicates the probable relevance of exposure to microbial emissions from livestock farms considering public health besides chemical air pollution, necessitating future research incorporating both.

Nicotinamide mononucleotide alleviates endotoxin-induced acute lung injury by modulating macrophage polarization via the SIRT1/NF-κB pathway.

CONTEXT: Sepsis-induced acute lung injury (ALI) is a severe condition with limited effective therapeutics; nicotinamide mononucleotide (NMN) has been reported to exert anti-inflammatory activities. OBJECTIVE: This study explores the potential mechanisms by which NMN ameliorates sepsis-induced ALI in vivo and in vitro. MATERIALS AND METHODS: Cultured MH-S cells and a murine model were used to evaluate the effect of NMN on sepsis-induced ALI. MH-S cells were stimulated with LPS (1 µg/mL) and NMN (500 µM) for 12 h grouping as control, LPS, and LPS + NMN. Cell viability, apoptotic status, and M1/2 macrophage-related markers were detected. The mice were pretreated intraperitoneally with NMN (500 mg/kg) and/or EX-527 (5 mg/kg) 1 h before LPS injection and randomized into 7 groups (n = 8): control, LPS, LPS + NMN, NMN, LPS + NMN + EX-527 (a SIRT1 inhibitor), LPS + EX-527, and EX-527. After 12 h, lung histopathology, W/D ratio, MPO activity, NAD+ and ATP levels, M1/2 macrophage-related markers, and expression of the SIRT1/NF-κB pathway were detected. RESULTS: In MH-S cells, NMN significantly decreased the apoptotic rate from 12.25% to 5.74%. In septic mice, NMN improved the typical pathologic findings in lungs and reduced W/D ratio and MPO activity, but increased NAD+ and ATP levels. Additionally, NMN suppressed M1 but promoted M2 polarization, and upregulated the expression of SIRT1, with inhibition of NF-κB-p65 acetylation and phosphorylation. Furthermore, inhibition of SIRT1 reversed the effects of NMN-induced M2 macrophage polarization. CONCLUSIONS: NMN protects against sepsis-induced ALI by promoting M2 macrophage polarization via the SIRT1/NF-κB pathway, it might be an effective strategy for preventing or treating sepsis-induced ALI.

Serum amyloid A aggravates endotoxin-induced ocular inflammation through the regulation of retinal microglial activation.

Serum amyloid A (SAA) are major acute-phase response proteins which actively participate in many inflammatory diseases. This study was designed to explore the function of SAA in acute ocular inflammation and the underlying mechanism. We found that SAA3 was upregulated in endotoxin-induced uveitis (EIU) mouse model, and it was primarily expressed in microglia. Recombinant SAA protein augmented intraocular inflammation in EIU, while the inhibition of Saa3 by siRNA effectively alleviated the inflammatory responses and rescued the retina from EIU-induced structural and functional damage. Further study showed that the recombinant SAA protein activated microglia, causing characteristic morphological changes and driving them further to pro-inflammatory status. The downregulation of Saa3 halted the amoeboid change of microglia, reduced the secretion of pro-inflammatory factors, and increased the expression of tissue-reparative genes. SAA3 also regulated the autophagic activity of microglial cells. Finally, we showed that the above effect of SAA on microglial cells was at least partially mediated through the expression and signaling of Toll-like receptor 4 (TLR4). Collectively, our study suggested that microglial cell-expressed SAA could be a potential target in treating acute ocular inflammation.

Channel Formation in Cry Toxins: An Alphafold-2 Perspective.

Bacillus thuringiensis (Bt) strains produce pore-forming toxins (PFTs) that attack insect pests. Information for pre-pore and pore structures of some of these Bt toxins is available. However, for the three-domain (I-III) crystal (Cry) toxins, the most used Bt toxins in pest control, this crucial information is still missing. In these Cry toxins, biochemical data have shown that 7-helix domain I is involved in insertion in membranes, oligomerization and formation of a channel lined mainly by helix α4, whereas helices α1 to α3 seem to have a dynamic role during insertion. In the case of Cry1Aa, toxic against Manduca sexta larvae, a tetrameric oligomer seems to precede membrane insertion. Given the experimental difficulty in the elucidation of the membrane insertion steps, we used Alphafold-2 (AF2) to shed light on possible oligomeric structural intermediates in the membrane insertion of this toxin. AF2 very accurately (<1 Å RMSD) predicted the crystal monomeric and trimeric structures of Cry1Aa and Cry4Ba. The prediction of a tetramer of Cry1Aa, but not Cry4Ba, produced an 'extended model' where domain I helices α3 and α2b form a continuous helix and where hydrophobic helices α1 and α2 cluster at the tip of the bundle. We hypothesize that this represents an intermediate that binds the membrane and precedes α4/α5 hairpin insertion, together with helices α6 and α7. Another Cry1Aa tetrameric model was predicted after deleting helices α1 to α3, where domain I produced a central cavity consistent with an ion channel, lined by polar and charged residues in helix α4. We propose that this second model corresponds to the 'membrane-inserted' structure. AF2 also predicted larger α4/α5 hairpin n-mers (14 ≤n ≤ 17) with high confidence, which formed even larger (~5 nm) pores. The plausibility of these models is discussed in the context of available experimental data and current paradigms.