Cadmium aggravates liver injury by activating ferroptosis and neutrophil extracellular traps formation in Nile tilapia (Oreochromis niloticus).

Cadmium (Cd) is a pervasive environmental contaminant and a significant risk factor for liver injury. The present study was undertaken to evaluate the involvement of ferroptosis and neutrophil extracellular traps (NETs) in Cd-induced liver injury in Nile tilapia (Oreochromis niloticus), and to explore its underlying mechanism. Cd-induced liver injury was associated with increased total iron, malondialdehyde (MDA), and Acyl-CoA synthetase long-chain family member 4 (ACSL4), together with reduced levels of glutathione, glutathione peroxidase-4a (Gpx4a), and solute carrier family 7 member 11 (SLC7A11), which are all hallmarks of ferroptosis. Moreover, liver hyperemia, neutrophil infiltration, increased inflammatory factors and myeloperoxidase, as well as elevated serum DNA content in Cd-stimulated Nile tilapia suggested that a considerable number of neutrophils were recruited to the liver. Furtherly, in vitro experiments demonstrated that Cd induced the formation of NETs, and the possible mechanism was related to the generation of reactive oxygen species and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, along with the P38 and extracellular regulated protein kinase (ERK) signaling pathways. We concluded that ferroptosis and NETs are the critical mechanisms contributing to Cd-induced liver injury in Nile tilapia. These findings will contribute to Cd toxicological studies in aquatic animals.

Single-cell transcriptomic sequencing identifies subcutaneous patient-derived xenograft recapitulated medulloblastoma.

BACKGROUND: Medulloblastoma (MB) is one of the most common malignant brain tumors that mainly affect children. Various approaches have been used to model MB to facilitate investigating tumorigenesis. This study aims to compare the recapitulation of MB between subcutaneous patient-derived xenograft (sPDX), intracranial patient-derived xenograft (iPDX), and genetically engineered mouse models (GEMM) at the single-cell level. METHODS: We obtained primary human sonic hedgehog (SHH) and group 3 (G3) MB samples from six patients. For each patient specimen, we developed two sPDX and iPDX models, respectively. Three Patch+/- GEMM models were also included for sequencing. Single-cell RNA sequencing was performed to compare gene expression profiles, cellular composition, and functional pathway enrichment. Bulk RNA-seq deconvolution was performed to compare cellular composition across models and human samples. RESULTS: Our results showed that the sPDX tumor model demonstrated the highest correlation to the overall transcriptomic profiles of primary human tumors at the single-cell level within the SHH and G3 subgroups, followed by the GEMM model and iPDX. The GEMM tumor model was able to recapitulate all subpopulations of tumor microenvironment (TME) cells that can be clustered in human SHH tumors, including a higher proportion of tumor-associated astrocytes and immune cells, and an additional cluster of vascular endothelia when compared to human SHH tumors. CONCLUSIONS: This study was the first to compare experimental models for MB at the single-cell level, providing value insights into model selection for different research purposes. sPDX and iPDX are suitable for drug testing and personalized therapy screenings, whereas GEMM models are valuable for investigating the interaction between tumor and TME cells.

ß-Carotene Protects Mice against Lipopolysaccharide and D-Galactosamine Induced Acute Liver Injury via Regulation of NF-κB, MAPK, and Nrf2 Signaling.

Acute liver injury (ALI), posing a serious threaten to our life, has emerged as a public health issue around the world. ß-carotene has plenty of pharmacologic effects, such as anti-inflammatory, antioxidant, and antitumor activities. In this study, we focused on studying the protective role and potential molecular mechanisms of ß-carotene against D-galactosamine (D-GalN) and lipopolysaccharide (LPS) induced ALI. Our results indicated that ß-carotene pretreatment effectively hindered abnormal changes induced by LPS/D-GalN in liver histopathology. Meanwhile, serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were downgraded with ß-carotene pretreatment. ß-carotene pretreatment also decreased malondialdehyde content and myeloperoxidase activity, increased glutathione peroxidase and superoxide dismutase levels, and reduced the levels of tumor necrosis factor-a (TNF-α) and interleukin 6 (IL-6) in liver tissues. Further investigations found that ß-carotene mediated multiple signaling pathways in LPS/D-GalN-induced ALI, inhibiting NF-κB and MAPK signaling and upregulating the expression of Nrf2 and HO-1 proteins. All findings indicate that ß-carotene appears to protect mice against LPS/D-GalN induced ALI by reducing oxidative stress and inflammation, possibly via regulating NF-κB, MAPK, and Nrf2 signaling.

Quercetin Alleviates Deoxynivalenol-Induced Intestinal Damage by Suppressing Inflammation and Ferroptosis in Mice.

Deoxynivalenol (DON), one of the most prevalent mycotoxins found in food and feed, can cause gastrointestinal inflammation and systemic immunosuppression, presenting a serious hazard to human and animal health. Quercetin (QUE) is a plant polyphenol with anti-inflammatory and antioxidant properties. In this research, we investigated the potential function of QUE as a treatment for DON-induced intestinal damage. Thirty male specific-pathogen-free BALB/c mice were randomly allocated to treatment with QUE (50 mg/kg) and/or DON (0, 0.5, 1, and 2 mg/kg). We found that QUE attenuated DON-induced intestinal damage in mice by improving jejunal structural injury and changing tight junction proteins (claudin-1, claudin-3, ZO-1, and occludin) levels. QUE also suppressed DON-triggered intestinal inflammation by inhibiting the TLR4/NF-κB signaling pathway. Meanwhile, QUE decreased the oxidative stress caused by DON by enhancing the concentrations of SOD and GSH, while diminishing the contents of MDA. In particular, QUE reduced DON-induced intestinal ferroptosis. DON-induced intestinal damage elevated TfR and 4HNE levels, along with transcription levels of ferroptosis-related genes (PTGS2, ACSL4, and HAMP1) while diminishing mRNA levels of FTH1, SLC7A11, GPX4, FPN1, and FSP1, all of which were reversed by QUE treatment. Our findings imply that QUE alleviates DON-induced intestinal injury in mice by inhibiting the TLR4/NF-κB signaling pathway and ferroptosis. In this study, we elucidate the toxicological mechanism of DON, provide a basic foundation or theory for future DON prevention and treatment, and explore strategies to prevent and alleviate DON's hazardous effects.

De Novo Mutations Contributes Approximately 7% of Pathogenicity in Inherited Eye Diseases.

Purpose: The purpose of this study was to describe genotype-phenotype associations and novel insights into genetic characteristics in a trio-based cohort of inherited eye diseases (IEDs). Methods: To determine the etiological role of de novo mutations (DNMs) and genetic profile in IEDs, we retrospectively reviewed a large cohort of proband-parent trios of Chinese origin. The patients underwent a detailed examination and was clinically diagnosed by an ophthalmologist. Panel-based targeted exome sequencing was performed on DNA extracted from blood samples, containing coding regions of 792 IED-causative genes and their flanking exons. All participants underwent genetic testing. Results: All proband-parent trios were divided into 22 subgroups, the overall diagnostic yield was 48.67% (605/1243), ranging from 4% to 94.44% for each of the subgroups. A total of 108 IED-causative genes were identified, with the top 24 genes explaining 67% of the 605 genetically solved trios. The genetic etiology of 6.76% (84/1243) of the trio was attributed to disease-causative DNMs, and the top 3 subgroups with the highest incidence of DNM were aniridia (n = 40%), Marfan syndrome/ectopia lentis (n = 38.78%), and retinoblastoma (n = 37.04%). The top 10 genes have a diagnostic yield of DNM greater than 3.5% in their subgroups, including PAX6 (40.00%), FBN1 (38.78%), RB1 (37.04%), CRX (10.34%), CHM (9.09%), WFS1 (8.00%), RP1L1 (5.88%), RS1 (5.26%), PCDH15 (4.00%), and ABCA4 (3.51%). Additionally, the incidence of DNM in offspring showed a trend of correlation with paternal age at reproduction, but not statistically significant with paternal (P = 0.154) and maternal (P = 0.959) age at reproduction. Conclusions: Trios-based genetic analysis has high accuracy and validity. Our study helps to quantify the burden of the full spectrum IED caused by each gene, offers novel potential for elucidating etiology, and plays a crucial role in genetic counseling and patient management.

Forsythin inhibits ß-hydroxybutyrate-induced oxidative stress in bovine macrophages by regulating p38/ERK, PI3K/Akt, and Nrf2/HO-1 signaling pathways.

Ketosis is a metabolic disease of dairy cows in the perinatal period, ß-hydroxybutyrate (ß-HB) is the main component of ketosis. High levels of ß-HB can trigger oxidative stress and inflammatory response in dairy cows, leading to decreased milk yield and multiple postpartum diseases. Forsythin (FOR), the major constituent of the herbal medicine Forsythia, has anti-inflammatory, anti-oxidant, and antiviral effects. FOR was demonstrated to have an antioxidant effect on PC12 cells. However, the effects of FOR on ß-HB-stimulated bovine macrophages (BMs) has not been reported. Thus, the aim of the present study was to investigate the effects of FOR on ß-HB-stimulated BMs. Firstly, the CCK8 test confirmed that FOR (50, 100, 200 µg/mL) has no effect on BMs activity, and we selected these concentrations for subsequent experiments. Secondly, through detecting the oxidation indexes ROS, MDA and antioxidant indexes CAT and SOD, we confirmed the antioxidant effect of FOR on BMs. Next, qRT-PCR confirmed that FOR dramatically reduced the mRNA levels of IL-1ß and IL-6. Furthermore, the western blotting confirmed that FOR observably down-regulated ß-HB-stimulated phosphorylation of p38, ERK and Akt and up-regulated expression of Nrf2, and HO-1. Above results suggested that FOR plays antioxidant effects on ß-HB-induced BMs through p38, ERK and PI3K/Akt, Nrf2 and HO-1 signaling pathways. Therefore, we speculated that FOR may be a potential medicine to alleviate ß-HB-induced inflammatory response and provide a preliminary reference for the research and development of FOR.

Lysine specific demethylase 1 inhibitor alleviated lipopolysaccharide/D-galactosamine-induced acute liver injury.

Acute liver injury is a severe clinical syndrome with markedly high mortality and poor prognosis. An accumulating body of evidence has demonstrated that epigenetic mechanisms have essential roles in the pathogenesis of acute liver injury. Lysine-specific demethylase 1 (LSD1) belongs to the amine oxidase superfamily of flavin adenine dinucleotide (FAD)-dependent enzymes, specifically demethylates H3 lysine 4. In the study, we investigated the effects and mechanisms of LSD1 in lipopolysaccharide (LPS)/D-Galactosamine (D-Gal)-induced acute liver injury in mice. Western blot analysis showed that LSD1 phosphorylation and di-methylated histone H3 on lysine 4 (H3K4me2) protein expression were significantly increased after LPS/D-Gal treatment (2.3 and 2.4 times higher than control respectively). GSK-LSD1 2HCl is an irreversible and selective LSD1 inhibitor. Pre-treatment with LSD1 inhibitor alleviated LPS/D-Gal-induced liver damage, decreased serum levels of alanine transaminase and aspartate aminotransferase in mice. Moreover, the LSD1 phosphorylation level in low, medium, and high LSD1 inhibitor groups was lower by a factor of 1.6, 1.9, and 2.0 from the LPS/D-Gal group, respectively. Mechanistically, LSD1 inhibitor further inhibited NF-κB signaling cascades and subsequently inhibited the production of pro-inflammatory cytokine TNF-α, IL-6, and IL-1ß induced by LPS/D-Gal in liver tissues. Furthermore, LSD1 inhibitor upregulated the protein expression of Nrf2/HO-1 signaling pathways, and the activities of related antioxidant enzymes were enhanced. Collectively, our data demonstrated that LSD1 inhibitor protected against the LPS/D-Gal-induced acute liver injury via inhibiting inflammation and oxidative stress, and targeting the epigenetic marker may be a potent therapeutic strategy for acute liver injury.

CuO-NPs-triggered heterophil extracellular traps exacerbate liver injury in chicks by promoting oxidative stress and inflammatory responses.

With the widespread use of copper oxide nanoparticles (CuO-NPs), their potential toxicity to the environment and biological health has attracted close attention. Heterophil extracellular traps (HETs) are an innate immune mechanism of chicken heterophils against adverse stimuli, but excessive HETs cause damage. Here, we explored the effect and mechanism of CuO-NPs on HETs formation in vitro and further evaluated the potential role of HETs in chicken liver and kidney injury. Heterophils were exposed to 5, 10, and 20 µg/mL of CuO-NPs for 2 h. The results showed that CuO-NPs induced typical HETs formation, which was dependent on NADPH oxidase, P38 and extracellular regulated protein kinases (ERK1/2) pathways, and glycolysis. In in vivo experiments, fluorescence microplate and morphological analysis showed that CuO-NPs elevated the level of HETs in chicken serum and caused liver and kidney damage. Meanwhile, CuO-NPs caused hepatic oxidative stress (MDA, SOD, CAT, and GSH-PX imbalance), and also induced an increase in mRNA expression of their inflammatory and apoptosis-related factors (IL-1ß, IL-6, TNF-α, COX-2, iNOS, NLRP3, and Caspase-1, 3, 11). However, these results were significantly altered by DNase I (HETs degradation reagent). In conclusion, the present study demonstrates for the first time that CuO-NPs induce the formation of HETs and that HETs exacerbate pathological damage in chicken liver and kidney by promoting oxidative stress and inflammation, providing insights into immunotoxicity and potential prevention and treatment targets caused by CuO-NPs overexposure.

The JMJD3 histone demethylase inhibitor GSK-J1 ameliorates lipopolysaccharide-induced inflammation in a mastitis model.

Jumonji domain-containing 3 (JMJD3/KDM6B) is a histone demethylase that plays an important role in regulating development, differentiation, immunity, and tumorigenesis. However, the mechanisms responsible for the epigenetic regulation of inflammation during mastitis remain incompletely understood. Here, we aimed to investigate the role of JMJD3 in the lipopolysaccharide (LPS)-induced mastitis model. GSK-J1, a small molecule inhibitor of JMJD3, was applied to treat LPS-induced mastitis in mice and in mouse mammary epithelial cells in vivo and in vitro. Breast tissues were then collected for histopathology and protein/gene expression examination, and mouse mammary epithelial cells were used to investigate the mechanism of regulation of the inflammatory response. We found that the JMJD3 gene and protein expression were upregulated in injured mammary glands during mastitis. Unexpectedly, we also found JMJD3 inhibition by GSK-J1 significantly alleviated the severity of inflammation in LPS-induced mastitis. These results are in agreement with the finding that GSK-J1 treatment led to the recruitment of histone 3 lysine 27 trimethylation (H3K27me3), an inhibitory chromatin mark, in vitro. Furthermore, mechanistic investigation suggested that GSK-J1 treatment directly interfered with the transcription of inflammatory-related genes by H3K27me3 modification of their promoters. Meanwhile, we also demonstrated that JMJD3 depletion or inhibition by GSK-J1 decreased the expression of toll-like receptor 4 and negated downstream NF-κB proinflammatory signaling and subsequently reduced LPS-stimulated upregulation of Tnfa, Il1b, and Il6. Together, we propose that targeting JMJD3 has therapeutic potential for the treatment of inflammatory diseases.

Alumina nanoparticles-induced heterophil extracellular traps exacerbate liver injury by regulating oxidative stress and inflammation in chickens.

Widely used alumina nanoparticles (Al2O3 NPs) exposed to the environment pose a serious threat to human and animal health. The formation of heterophil extracellular traps (HETs) is a mechanism of innate immune defense against infection, but excessive HETs cause pathological damage. Here, we aim to explore the influence and mechanism of Al2O3 NPs on the formation of HETs in vitro, and further investigate the role of HETs release in histopathological damage after Al2O3 NPs treatment. Immunofluorescence analysis showed that Al2O3 NPs induced the formation of HETs, which was characterized by modified histones and elastase in the DNA backbone. Fluorescence microplate analysis showed that HETs formation was dependent on NADPH oxidase, P38, extracellular regulated protein kinases (ERK1/2) pathways and glycolysis. In vivo investigation showed that Al2O3 NPs significantly caused HETs release and liver damage. Biochemical analysis showed that Al2O3 NPs inhibited the activity of antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-PX). Real-time fluorescence quantification results showed that Al2O3 NPs caused the overexpression of inflammation-related molecules interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), caspase-1 and caspase-11. All these changes were significantly changed by DNase I (Degradation reagent for HETs). Together, these suggest that Al2O3 NPs-induced HETs exacerbate liver injury by regulating oxidative stress and inflammatory responses, which provide a new perspective and potential prophylaxis and treatment targets for Al2O3 NPs toxicological research.

Morin alleviates aflatoxin B1-induced liver and kidney injury by inhibiting heterophil extracellular traps release, oxidative stress and inflammatory responses in chicks.

Aflatoxin B1 (AFB1) is a secondary metabolite produced by Aspergillus flavus and parasitic aspergillus, mainly existing in cereals, peanuts, corn, and other crops, which seriously endanger poultry, human health, and environment. Morin, a flavonoid compound extracted from moraceae plants, possess antioxidant, antibacterial, and anti-inflammatory effects. However, whether morin has a protective effect on AFB1-induced liver and kidney damage in chicks has not been specifically reported. In this study, we mainly confirmed the protective effect of morin on AFB1-induced liver and kidney damage in chicks and clarified its mechanism. It was found that morin can significantly reduce the liver biochemical indicators of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and kidney indicators of creatinine (CRE) and urea nitrogen (BUN) levels. Meanwhile, histopathological examination showed that morin effectively relieved AFB1-caused liver damage, including hepatocyte disruption, swelling, and inflammatory cell infiltration, and effectively relieved kidney damage, including renal cell necrosis, exfoliation, and vacuolization. Further investigation of its mechanism demonstrated that morin significantly inhibited AFB1-induced heterophil extracellular traps (HETs) release, and decreased the level of malondialdehyde (MDA) but increased the levels of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) in vivo. Moreover, quantitative real-time PCR (qRT-PCR) analysis showed that morin also significantly decreased AFB1-induced mRNA expressions of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1ß (IL-1ß), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), caspase-1, caspase-3, and caspase-11. In conclusion, all results confirmed that morin could protect AFB1-caused liver and kidney damage by inhibiting HETs release, regulating oxidative stress, and inhibiting inflammatory response, suggesting that morin can be utilized as a potential drug for prevention and treatment of aflatoxicosis in poultry breeding industry.

Baicalin protects against zearalenone-induced chicks liver and kidney injury by inhibiting expression of oxidative stress, inflammatory cytokines and caspase signaling pathway.

Zearalenone (ZEA) is a secondary metabolite produced by fungi such as Fusarium and Fusarium flavum, which is classified as a mycotoxin. Crops and feed in a humid surrounding are widely polluted by ZEA, which further endangering the healthful aquaculture of poultry and even human health. Up to now, prevention and cure of mycotoxicosis is still a crucial subject of poultry husbandry. Baicalin (BAI) is a flavonoid refined from dried roots of Scutellaria baicalensis possessing the function of hepatoprotective, anti-inflammatory, anti-oxidant, and anti-atherosclerotic efficacies.etc. But whether Baicalin also has a protective effect against ZEA intoxication is unclear. Therefore, the aim of this study was to establish a model of ZEA-induced toxic injury in chicks, and then to investigate the way in which Baicalin plays a protective role in the mechanism of ZEA-induced liver and kidney injury in chicks. The results exhibit that Baicalin could not only significantly decrease aspartate aminotransferase (AST) , alanine aminotransferase (ALT) and creatinine (Cre) levels in serum, but also ameliorate ZEA-induced pathologic changes of liver and kidney. Baicalin could also significantly regulate ZEA-induced the changes of catalase (CAT) , malondialdehyde (MDA) , total sulfhydryl group , except for glutathione peroxidase (GSH-px) , and inhibit the mRNA levels of inflammatory cytokines tumor necrosis factor-α (TNF-α) , interleukin-1ß (IL-1ß) and cyclooxygenase-2 (COX-2) with caspase-3 and caspase-11 in the caspase signaling pathway , meanwhile inhibit the cell apoptosis in immunohistochemistry. In summary, we successfully established a model of ZEA-induced liver injury in chicks, and confirm that Baicalin can reduce ZEA-induced liver and kidney injury in chicks. The mechanism of these effects is via inhibiting inflammation, oxidative stress and apoptosis, which also indicates the potential applicability of Baicalin for the prevention and treatment of ZEA-induced toxicity in chicks.

A new nairo-like virus associated with human febrile illness in China.

ABSTRACTSeveral nairo-like viruses have been discovered in ticks in recent years, but their relevance to public health remains unknown. Here, we found a patient who had a history of tick bite and suffered from a febrile illness was infected with a previously discovered RNA virus, Beiji nairovirus (BJNV), in the nairo-like virus group of the order Bunyavirales. We isolated the virus by cell culture assay. BJNV could induce cytopathic effects in the baby hamster kidney and human hepatocellular carcinoma cells. Negative-stain electron microscopy revealed enveloped and spherical viral particles, morphologically similar to those of nairoviruses. We identified 67 patients as BJNV infection in 2017-2018. The median age of patients was 48 years (interquartile range 41-53 years); the median incubation period was 7 days (interquartile range 3-12 days). Most patients were men (70%), and a few (10%) had underlying diseases. Common symptoms of infected patients included fever (100%), headache (99%), depression (63%), coma (63%), and fatigue (54%), myalgia or arthralgia (45%); two (3%) patients became critically ill and one died. BJNV could cause growth retardation, viremia and histopathological changes in infected suckling mice. BJNV was also detected in sheep, cattle, and multiple tick species. These findings demonstrated that the newly discovered nairo-like virus may be associated with a febrile illness, with the potential vectors of ticks and reservoirs of sheep and cattle, highlighting its public health significance and necessity of further investigation in the tick-endemic areas worldwide.

Alpinetin prevents inflammatory responses in OVA-induced allergic asthma through modulating PI3K/AKT/NF-κB and HO-1 signaling pathways in mice.

Allergic asthma is the most common type of asthma which characterized by inflammatory responses of the airways. Alpinetin, a flavonoid compound derived from the ginger family of medicinal herbs, possesses various biological properties including anti-inflammatory, anti-oxidant and other medical effects. In this study, we aimed to evaluate the effects of alpinetin on OVA-induced allergic asthma, and further to examine its molecular mechanisms underlying these processes in vivo and in vitro. Mice were sensitized and challenged with OVA to build allergic asthma model in vivo. Bronchoalveolar lavage fluid (BALF) was collected for inflammatory cells analysis and lung tissues were examined for histopathological examination. The levels of IL-5, IL-13, IL-4, IgE, TNF-α, IL-6 and IL-1ß were determined by the respective ELISA kits. The PI3K/AKT/NF-κB and HO-1 signaling pathways were examined by western blot analysis. The results showed that alpinetin significantly ameliorated OVA-induced pathologic changes of lungs, such as decreasing massive inflammatory cell infiltration and mucus hypersecretion, and reduced the number of inflammatory cells in BALF. Alpinetin also decreased the OVA-induced levels of IL-4, IL-5, IL-13 and IgE. Furthermore, alpinetin inhibited OVA-induced phosphorylation of p65, IκB, PI3K and AKT, and the activity of HO-1 in vivo. More importantly, these anti-inflammatory effects and molecular mechanisms of alpinetin has also been confirmed in LPS-stimulated RAW 264.7 macrophages in vitro. In conclusion, above results indicate that alpinetin exhibites a potent anti-inflammatory activity in allergic asthma through modulating PI3K/AKT/NF-κB and HO-1 signaling pathways, which would be used as a promising therapy agent for allergic asthma.

Protective Effects of Pterostilbene on Lipopolysaccharide-Induced Acute Lung Injury in Mice by Inhibiting NF-κB and Activating Nrf2/HO-1 Signaling Pathways.

Pterostilbene (PTER) is a kind of stilbene compound with biological activity isolated from plants such as red sandalwood, blueberry and grape. It has anti-tumor, anti-bacterial, anti-oxidation and other pharmacological activities. However, the underlying mechanism of the protective effect of PTER on lipopolysaccharide (LPS)-induced acute lung injury (ALI) remained not clarified. In this study, LPS was used to establish a mouse model of ALI. Bronchoalveolar lavage fluid (BALF) was collected for inflammatory cells, and the wet-to-dry weight ratio of the lungs was measured. The activities of myeloperoxidase (MPO), antioxidant indexes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px) and oxidation index such as malondialdehyde (MDA) in lung tissues of mice were measured by the corresponding kits. The levels of Cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), TNF-α, IL-6 and IL-1ß in lung tissues of mice were detected by quantitative real-time polymerase chain reaction (qRT-PCR). The activities of Nrf2, HO-1, p-p65 and p-IκB were determined by western blotting. The results showed that the model of LPS-induced ALI was successfully replicated, and it was found that PTER could significantly improve the pathological degree of ALI such as sustained the integrity of the lung tissue structure, alleviated pulmonary interstitial edema and alveolar wall thickening, reduced infiltrated inflammatory cells. PTER could decrease the number of inflammatory cells and obviously inhibit the increase of W/D ratio caused by LPS. PTER could also significantly reduce LPS-induced MPO and MDA, and increase LPS-decreased SOD, CAT and GSH-Px in the lungs. In addition, it was also found that PTER has the ability to decrease LPS-induced production of COX-2, iNOS, TNF-α, IL-6 and IL-1ß. The underlying mechanism involved in the protective effect of PTER on ALI were via activating Nrf2 and HO-1, and inhibiting the phosphorylation of p65 and IκB. These results suggested that PTER can protect LPS-induced ALI in mice by inhibiting inflammatory response and oxidative stress, which provided evidence that PTER may be a potential therapeutic candidate for LPS-induced ALI intervention.

Morin alleviates LPS-induced mastitis by inhibiting the PI3K/AKT, MAPK, NF-κB and NLRP3 signaling pathway and protecting the integrity of blood-milk barrier.

Mastitis is a common veterinary clinical disease that restricts the development of dairy farming around the world. Morin, extracted from Mulberry Tree and other herbs, has been reported to possess the function of anti-bacteria, anti-oxidant, and anti-inflammatory. However, whether morin could protect lipopolysaccharide (LPS)-induced mouse mastitis in vivo has not well known. This study firstly aims to evaluate the effects of morin on LPS-induced mouse mastitis in vivo, and then try to illustrate the mechanism involved in the process. Before injected with LPS, mice were intraperitoneally pre-injected with different concentrations of morin, and mice of the control and LPS group were injected with the same amount of saline. Pathologic changes of mammary gland were determined by histopathological examination. Myeloperoxidase (MPO) activities of mammary gland were determined by the MPO kits. The mRNA expressions of inflammatory cytokines including TNF-α, IL-1ß and IL-6, and those of chemokine factors CCL2 and CXCL2, and those of tight junctions occludin claudin-3 were examined by qRT-PCR analysis. The activities of IκB, p65, ERK, P38, AKT, PI3K, NLPR3, claudin-1, claudin-3 and occludin were determined by western blotting. The results showed that morin alleviated LPS-induced edema, destructed structures and infiltrated inflammatory cells of mammary gland. Morin administration significantly decreased LPS-induced TNF-α, IL-1ß, IL-6, CCL2 and CXCL2 mRNA expressions. Furthermore, western blot analysis also showed that morin significantly reduced LPS-induced phosphorylation of p65, IκB, p38 and ERK, and enhanced LPS-induced phosphorylation of AKT and PI3K. It was also found that LPS-decreased claudin-3 and occludin expressions were also inhibited by morin treatment. In summary, above results suggest that morin indeed protect LPS-induced mouse mastitis in vivo, and the mechanism was through inhibiting the PI3K/AKT, MAPK, NF-κB and NLRP3 signaling pathways and protecting the integrity of blood-milk barrier by regulating the tight junction proteins expressions.

Neutrophil extracellular traps promote cadmium chloride-induced lung injury in mice.

Cadmium (Cd) is a ubiquitous toxic heavy metal derived mainly from industrial processes. In industrialized societies, individuals are exposed to a plethora of sources of Cd pollution. Cd can trigger serious diseases such as rheumatoid arthritis (RA) and chronic obstructive pulmonary disease (COPD) by the over-activating immune system. As an effector mechanism in innate immunity, neutrophil extracellular traps (NETs) not only play an important role in defending against infection but also lead to tissue damage. However, the role of NETs in Cd-induced lung damage process has not been previously studied. In this study, we aimed to investigate the potential effects of Cd-induced NETs on lung injury in vivo and further to clarify the molecular mechanisms of Cd-induced NETs formation. In vivo, Cd treatment destroyed the structural integrity of lung tissue and significantly increased the levels of NETs in the bronchoalveolar lavage fluid (BALF). The known NETs inhibitor DNase I ameliorated pathologic changes and significantly decreased levels of NETs in BALF, which suggesting the curial role of NETs in Cd-induced lung injury. Further investigation showed that Cd could significantly trigger NETs formation, which is composed of DNA backbone decorated with histones (H3) and neutrophils elastase (NE). The inhibitors of NADPH oxidase, ERK1/2 and p38 MAPK-signaling pathways significantly reduced the formation of NETs, and western blotting analysis also showed that Cd significantly increased the phosphorylation of p38 and ERK1/2 signaling pathways. Above results confirmed that NADPH oxidase, ERK1/2 and p38 MAPK-signaling pathways were related to Cd-induced NETs formation. In conclusion, NETs was involved in Cd-induced lung injury, and the mechanisms of Cd-induced NETs formation was via activating NADPH oxidase, ERK1/2 and p38 MAPK-signaling pathways, which might provide a new perspective in Cd-induced lung injury.

Mixed adenoendocrine carcinoma in the extrahepatic biliary tract: A case report and literature review.

Neuroendocrine neoplasm (NEN) comprises a group of tumors that exhibit neuroendocrine phenotypes. NEN is subclassified into neuroendocrine tumor (NET), neuroendocrine carcinoma (NEC) and mixed adenoendocrine carcinoma (MANEC), based on histopathological parameters. NEN in the extrahepatic biliary tract (EHBT) is uncommon. Little is known about its clinicopathological features and prognostic indicators. The present study presented a case of MANEC in the distal common bile duct (CBD) and reviewed previous cases of NENs in the EHBT to characterize the clinical settings of this disease entity and to identify influencing factors of survival outcomes. A 64-year-old Chinese woman presented with abdominal pain and jaundice. Imaging studies demonstrated malignant stenosis in the distal CBD. Bile duct brush cytology revealed small clusters of atypical cells. Following an initial diagnosis of distal cholangiocarcinoma (CCA), the patient underwent pancreaticoduodenectomy. Histological analysis combined with immunohistochemical investigation of the resected specimen revealed a collision tumor that was composed of poorly differentiated adenocarcinoma and NEC. Each histological component accounted for >30% of the tumor. The definitive diagnosis was a MANEC in the distal CBD. Multiple intrahepatic and pulmonary metastases were observed postoperatively over 8 months. The patient succumbed to the disease 12 months after surgery. In conclusion, NEN occurs infrequently in the EHBT, with NET being the predominant type. NEN in the EHBT is extremely challenging to diagnose preoperatively due to its tendency to mimic CCA. Patients with NEN in the EHBT exhibited extremely distinct oncology outcomes according to pathological types. Additionally, old age (>60 years) and the presence of tumor recurrence were associated with decreased survival of patients with NEN.

Effects of Neutrophil Extracellular Traps on Bovine Mammary Epithelial Cells in vitro.

Bovine mastitis is a common infectious disease which causes huge economic losses in dairy cattle. Bovine mammary epithelial cell (BMEC) damage usually directly causes the decrease of milk production, which is one of the most important causes of economic loss. NETs, novel effector mechanisms, are reported to exacerbate the pathogenesis of several inflammatory diseases. NETs formation has also been observed in the milk and mammary glands of sheep. However, the effects and detailed mechanisms of NETs on BMEC damage remain unclear. Thus, we aim to examine the effects of NETs on BMECs in vitro, and further to investigate the detail mechanism. In this study, the cytotoxicity of NETs on BMECs was determined using lactic dehydrogenase (LDH) levels in culture supernatants. Histone-induced BMEC damage was examined by flow cytometry and immunofluorescence analysis. The activities of caspase 1, caspase 3, caspase 11, and NLRP3 was detected using western blotting and immunohistochemical analysis. The results showed that NETs and their component histone significantly increased cytotoxicity to BMECs, suggesting the critical role of NETs, and their component histone in BMEC damage. In addition, histone could also induce necrosis, pyroptosis, and apoptosis of BMECs, and the mechanisms by which histone leads to BMEC damage occurred via activating caspase 1, caspase 3, and NLRP3. Altogether, NETs formation regulates inflammation and BMEC damage in mastitis. Inhibiting excess NETs formation may be useful to ameliorate mammary gland damage associated with mastitis.

Zearalenone Induces Estrogen-Receptor-Independent Neutrophil Extracellular Trap Release in Vitro.

Zearalenone (ZEA) is a non-steroidal estrogenic mycotoxin synthesized in Fusarium species, mainly Fusarium graminearum and Fusarium culmorum, and it has strong estrogenic activity and causes genotoxic effects, reproductive disorders, and immunosuppressive effects. Neutrophil extracellular trap (NET) has been studied for many years. Initially, NET was considered a form of the innate response that combats invading microorganisms. However, NET is involved in a series of pathophysiological mechanisms, including thrombosis, tissue necrosis, autoinflammation, and even autoimmunity. We recently found that polymorphonuclear neutrophils response to ZEA exposure by undergoing NET formation. However, the molecular mechanisms involved in this process remain poorly characterized. Here, we analyze whether estrogen receptors (ERs) can affect NET formation after ZEA stimulation. The involvement of ERs is investigated with the selective ER antagonists. Moreover, we investigate the mechanisms of NET formation using immunofluorescence staining, fluorescence microplate, and western blot analysis. Our results show that ERs (ERα and ERß) are not involved in ZEA-induced NET formation, but reactive oxygen species (ROS), extracellular signal-regulated kinase (ERK), and p38 are postulated to be involved. Specifically, we provide data demonstrating that ZEA-induced ROS may promote activation of ERK and p38 as well as subsequent NET release. We are the first to demonstrate this new mechanism of ZEA-induced NET formation, which may help in understanding the role of ZEA in overexposure diseases and provide a relevant basis for therapeutic applications.