ROLE OF CASPASE-1/CASPASE-11-HMGB1-RAGE/TLR4 SIGNALING IN THE EXACERBATION OF EXTRAPULMONARY SEPSIS INDUCED LUNG INJURY BY MECHANICAL VENTILATION.

BACKGROUND: Mechanical ventilation (MV) is a clinically important measure for respiratory support in critically ill patients. Although moderate tidal volume MV does not cause lung injury, it can further exacerbate lung injury in pathological state such as sepsis. This pathological process is known as the 'two-hit' theory, whereby an initial lung injury (e.g., infection, trauma, or sepsis) triggers an inflammatory response that activates immune cells, presenting the lung tissue in a fragile state and rendering it more susceptible to subsequent injury. The second hit occurs when mechanical ventilation is applied to lung tissue in a fragile state, and it is noteworthy that this mechanical ventilation is harmless to healthy lung tissue, further aggravating pre-existing lung injury through unknown mechanisms. This interaction between initial injury and subsequent mechanical ventilation develops a malignant cycle significantly exacerbating lung injury and severely hampering patient prognosis. The two-hit theory is critical to understanding the complicated mechanisms of ventilator-associated lung injury and facilitates the subsequent development of targeted therapeutic strategies. METHODS AND RESULTS: CLP mice model was used to mimic clinical sepsis patients. After 12 hours the mice were mechanical ventilated for 2-6 hours. MV by itself didn't lead to HMGB1 release, but significantly strengthened HMGB1 in plasma and cytoplasm of lung tissue in septic mice. Plasma and lung tissue activation of cytokines and chemokines, MAPK signaling pathway, neutrophil recruitment, and ALI were progressively decreased in LysM HMGB1-/- (Hmgb1 deletion in myeloid cells) and iHMGB1-/- mice (inducible HMGB1-/- mouse strain where the Hmgb1 gene was globally deleted after tamoxifen treatment). Compared with C57BL/6 mice, although EC-HMGB1-/- (Hmgb1 deletion in endothelial cells) mice didn't have lower levels of inflammation, neutrophil recruitment and lung injury were reduced. Compared with LysM HMGB1-/- mice, EC-HMGB1-/- mice had higher levels of inflammation but significantly lower neutrophil recruitment and lung injury. Overall, iHMGB1-/- mice had the lowest levels of all the above indicators. The level of inflammation, neutrophil recruitment and the degree of lung injury were decreased in RAGE-/- mice, and even the above indices were further decreased in TLR4/RAGE-/- mice. Levels of inflammation and neutrophil recruitment were decreased in Caspase-11-/- and Caspase-1/11-/- mice, but no statistical difference between these two gene knockout mice. CONCLUSIONS: These data show for the first time that the Caspase-1/Caspase-11-HMGB1-TLR4/RAGE signaling pathway plays a key role in mice model of sepsis induced lung injury exacerbated by MV. Different species of HMGB1 knockout mice have different lung protective mechanisms in the 'two hits' model, and location is the key to function. Specifically, LysM HMGB1-/- mice due to the deletion of HMGB1 in myeloid cells resulted in a pulmonary protective mechanism that was associated with a downregulation of the inflammatory response. EC HMGB1-/- mice are deficient in HMGB1 owing to endothelial cells, resulting in a distinct pulmonary protective mechanism independent of the inflammatory response and more relevant to the improvement of alveolar-capillary permeability. iHMGB1-/- mice, which are systemically HMGB1-deficient, share both of these lung-protective mechanisms.

In vitro and in vivo inhibitory effects of the Sanghuang mushroom extracts against Candida albicans.

Aim: To explore the antifungal potential of Sanghuang mushroom, a traditional Chinese medicine. Materials & methods: The antifungal properties and the potential mechanism of Sanghuang mushroom extracts against Candida albicans were studied in vitro and in vivo. Results: Sanghuang mushroom extracts inhibited the biofilm formation, increased the cell membrane permeability and promoted cell apoptosis of C. albicans in vitro. In a murine model of vulvovaginal candidiasis, Sanghuang mushroom extracts reduced the vaginal fungal load, improved inflammatory cell infiltration and downregulated the expression of TNF-α, IL-1ß and IL-6. Untargeted metabolomic analysis suggested the presence of ten antifungal components in Sanghuang mushroom extracts. Conclusion: Sanghuang mushroom extracts showed promise as antifungal agent against candidiasis, with potential therapeutic implications.

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Gamma-oryzanol alleviates osteoarthritis development by targeting Keap1-Nrf2 binding to interfere with chondrocyte ferroptosis.

Osteoarthritis (OA) is a prevalent joint disorder pathologically correlated to chondrocyte ferroptosis. Gamma-oryzanol (γ-Ory), as a first-line drug for autonomic disorders, aroused our interest because of its antioxidant, lipid-lowering, and hypoglycemic potential. The purpose of this study was to investigate the potential impact and mechanism of γ-Ory in treating OA. And the inhibition of γ-Ory in extracellular matrix molecule (ECM) degradation, ferroptosis, and Keap1-Nrf2 binding in IL-1ß-exposed chondrocytes was detected via immunoblotting, immunofluorescence, and co-immunoprecipitation. Micro-CT, SO staining, and immunofluorescence have been conducted to assess the impact of γ-Ory treatment on ACLT-mediated OA in rats at both imaging and histological stages. We found that γ-Ory dose-dependently suppressed IL-1ß-induced ECM deterioration and chondrocyte ferroptosis. Our animal experiments revealed that γ-Ory delayed ACLT-mediated OA development. Mechanistically, γ-Ory interfered with the binding of Keap1 to Nrf2 to promote the latter's nuclear import, thereby increasing the expression of detoxification enzymes. Summarily, our works support γ-Ory's potential as a candidate drug for the treatment of OA.

Poliumoside protects against type 2 diabetes-related osteoporosis by suppressing ferroptosis via activation of the Nrf2/GPX4 pathway.

BACKGROUND: Type 2 diabetes is often linked with osteoporosis (T2DOP), a condition that accelerates bone degeneration and increases the risk of fractures. Unlike conventional menopausal osteoporosis, the diabetic milieu exacerbates the likelihood of fractures and osteonecrosis. In particular poliumoside (Pol), derived from Callicarpa kwangtungensis Chun, has shown promising anti-oxidant and anti-inflammatory effects. Yet, its influence on T2DOP remains to be elucidated. PURPOSE: The focus of this study was to elucidate the influence of Pol in HGHF-associated ferroptosis and its implications in T2DOP. STUDY DESIGN: A murine model of T2DOP was established using a minimal dosage of streptozotocin (STZ) through intraperitoneal infusion combined with a diet high in fat and sugar. Concurrently, to mimic the diabetic condition in a lab environment, bone mesenchymal stem cells (BMSCs) were maintained in a high-glucose and high-fat (HGHF) setting. METHODS: The impact of Pol on BMSCs in an HGHF setting was determined using methods, such as BODIPY-C11, FerroOrange staining, mitochondrial functionality evaluations, and Western blot methodologies, coupled with immunoblotting and immunofluorescence techniques. To understand the role of Pol in a murine T2DOP model, techniques including micro-CT, hematoxylin and eosin (H&E) staining, dual-labeling with calcein-alizarin red, and immunohistochemistry were employed for detailed imaging and histological insights. RESULTS: Our findings suggest that Pol acts against HGHF-induced bone degradation and ferroptosis, as evidenced by an elevation in glutathione (GSH) and a decline in malondialdehyde (MDA) levels, lipid peroxidation, and mitochondrial reactive oxygen species (ROS). Furthermore, Pol treatment led to increased bone density, enhanced GPX4 markers, and reduced ROS in the distal femur region. On investigating the underlying mechanism of action, it was observed that Pol triggers the Nrf2/GPX4 pathway, and the introduction of lentivirus-Nrf2 negates the beneficial effects of Pol in HGHF-treated BMSCs. CONCLUSION: Pol is effective in treating T2DOP by activating the Nrf2/GPX4 signaling pathway to inhibit ferroptosis.

Hesperetin promotes diabetic wound healing by inhibiting ferroptosis through the activation of SIRT3.

Hesperetin (HST) is a flavonoid compound naturally occurring in citrus fruits and is widespread in various traditional medicinal herbs such as grapefruit peel, orange peel, and tangerine peel. These plant materials are commonly used in traditional Chinese medicine to prepare herbal remedies. The study aimed to investigate the potential molecular mechanisms through which HST reduces ferroptosis in human umbilical vein endothelial cells (HUVECs) and promotes angiogenesis and wound healing. We employed network pharmacology to predict the downstream targets affected by HST. The expression of markers related to ferroptosis was assessed through Western blot (WB) and polymerase chain reaction. Intracellular levels of ferroptosis-related metabolism were examined using glutathione/oxidized glutathione (GSH/GSSG) and malondialdehyde (MDA) assay kits. Mitochondrial status and iron levels within the cells were investigated through staining with Mitosox, FerroOrange, and JC1 staining. Potential downstream direct targets of HST were identified using molecular docking. Additionally, wound healing and neovascularization within the wound site were analyzed using various methods including HE staining, Masson's staining, immunohistochemistry, and Doppler hemodynamics assessment. HST effectively inhibits the elevated levels of intracellular ferroptosis stimulated by ERASTIN. Furthermore, we observed that HST achieves this inhibition of ferroptosis by activating SIRT3. In a diabetic rat wound model, HST significantly promotes wound healing, reducing levels of tissue ferroptosis, consistent with our in vitro findings. This study demonstrates that HST can inhibit the progression of ferroptosis and protect the physiological function of HUVECs by activating SIRT3. HST holds promise as a natural compound for promoting diabetic wound healing.

Identification of ZIP8-induced ferroptosis as a major type of cell death in monocytes under sepsis conditions.

Sepsis is a heterogenous syndrome with concurrent hyperinflammation and immune suppression. A prominent feature of immunosuppression during sepsis is the dysfunction and loss of monocytes; however, the major type of cell death contributing to this depletion, as well as its underlying molecular mechanisms, are yet to be identified. In this study, we confirmed the monocyte loss in septic patients based on a pooled gene expression data of periphery leukocytes. Using the collected reference gene sets from databases and published studies, we identified ferroptosis with a greater capacity to distinguish between sepsis and control samples than other cell death types. Further investigation on the molecular drivers, by a genetic algorithm-based feature selection and a weighted gene co-expression network analysis, revealed that zrt-/irt-like protein 8 (ZIP8), encoded by SLC39A8, was closely associated with ferroptosis of monocytes during sepsis. We validated the increase of ZIP8 of monocytes with in vivo and in vitro experiments. The in vitro studies also showed that downregulation of ZIP8 alleviated the lipopolysaccharide-induced lipid peroxidation, as well as restoring the reduction of GPX4, FTH1 and xCT. These findings suggest that ferroptosis might be a key factor in the loss of monocytes during sepsis, and that the heightened expression of ZIP8 may facilitate this progression.

Prevention of new-onset atrial fibrillation in elderly patients undergoing anatomic pulmonary resection by infusion of magnesium sulfate: protocol for a randomized controlled trial.

Atrial fibrillation (AF) is the most commonly sustained arrhythmia after pulmonary resection, which has been shown to predict higher hospital morbidity and mortality. The lack of strong evidence-based medical evidence makes doctors have very few options for medications to prevent new-onset AF following thoracic surgery. Magnesium can prevent perioperative AF in patients undergoing cardiac surgery. However, this has not yet been fully studied in patients undergoing non-cardiac thoracic surgery, which is the aim of this study. This is a single-center, prospective, double-blind, randomized controlled trial. In total, 838 eligible patients were randomly assigned to one of two study groups, namely, the control group or the magnesium group. The patients in the magnesium group preoperatively received 80 mg magnesium sulfate/kg ideal weight in 100 ml normal saline 30 min. The control group received the same volumes of normal saline simultaneously. The primary outcome is the incidence of new-onset AF intra-operative and on the first, second, and third postoperative days. The secondary outcomes are bradycardia, hypertension, hypotension, and flushing. The occurrence of stroke or any other type of arrhythmia is also recorded. Postoperative respiratory suppression and gastrointestinal discomfort, intensive care unit stays and total duration of hospital stays, in-hospital mortality, and 3-month all-cause mortality are also recorded as important outcomes. This study aims to prospectively evaluate the prophylactic effects of magnesium sulfate against AF compared with a placebo control group during and following anatomic pulmonary resection. The results may provide reliable evidence for the prophylactic value of magnesium against AF in patients with lung cancer. The trial was approved by the Clinical Research Ethics Committee of Shanghai Pulmonary Hospital and has been registered at Chinese Clinical Trial Registry: www.chictr.org.cn, identifier: ChiCTR2300068046.

Structure and ecological function of the soil microbiome associated with 'Sanghuang' mushrooms suffering from fungal diseases.

BACKGROUND: The most serious challenges in medicinal 'Sanghuang' mushroom production are the fungal diseases caused by various molds. Application of biological agents has been regarded as a potential crop disease management strategy. Here, the soil microbiome associated with 'Sanghuang' mushroom affected by fungal diseases grown under field cultivation (FC) and hanging cultivation (HC) was characterized using culture-dependent and culture-independent methods. RESULTS: A total of 12,525 operational taxonomic units (OTUs) and 168 pure cultures were obtained using high-throughput sequencing and a culture-dependent method, respectively. From high-throughput sequencing, we found that HC samples had more OTUs, higher α-diversity, and greater microbial community complexity than FC samples. Analysis of ß-diversity divided the soil microbes into two groups according to cultivation mode. Basidiomycota (48.6%) and Ascomycota (46.5%) were the two dominant fungal phyla in FC samples, with the representative genera Trichoderma (56.3%), Coprinellus (29.4%) and Discosia (4.8%), while only the phylum Ascomycota (84.5%) was predominant in HC samples, with the representative genera Discosia (34.0%), Trichoderma (30.2%), Penicillium (14.9%), and Aspergillus (7.8%). Notably, Trichoderma was predominant in both the culture-independent and culture-dependent analyses, with Trichoderma sp. FZ0005 showing high host pathogenicity. Among the 87 culturable bacteria, 15 exhibited varying extents of antifungal activity against Trichoderma sp. FZ0005, with three strains of Bacillus spp. (HX0037, HX0016, and HX0039) showing outstanding antifungal capacity. CONCLUSIONS: Overall, our results suggest that Trichoderma is the major causal agent of 'Sanghuang' fungal diseases and that Bacillus strains may be used as biocontrol agents in 'Sanghuang' cultivation.

Ginsenoside Rb1 inhibits ferroptosis to ameliorate hypoxic-ischemic brain damage in neonatal rats.

Hypoxic ischemic encephalopathy (HIE) is among the leading causes of neonatal mortality, and currently there is no effective treatment. Ginsenoside Rb1 (GsRb1) is one of the principal active components of ginseng, and has protective benefits against oxidative stress, inflammation, hypoxic injury, and so on. However, the role and underlying mechanism of GsRb1 on HIE are unclear. Here, we established the neonatal rat hypoxic-ischemic brain damage (HIBD) model in vivo and the PC12 cell oxygen-glucose deprivation (OGD) model in vitro to investigate the neuroprotective effects of GsRb1 on HIE, and illuminate the potential mechanism. Our results showed that GsRb1 and the ferroptosis inhibitor liproxstatin-1 (Lip-1) could significantly restore System Xc activity and antioxidant levels as well as inhibit lipid oxidation levels and inflammatory index levels of HIBD and OGD models. Taken together, GsRb1 might inhibit ferroptosis to exert neuroprotective effects on HIE through alleviating oxidative stress and inflammation, which will set the foundation for future research on ferroptosis by reducing hypoxic-ischemic brain injury and suggest that GsRb1 might be a promising therapeutic agent for HIE.

Vertical dispersal of Aedes albopictus within multi-story buildings in downtown Shanghai, China.

BACKGROUND: Shanghai has numerous high-rise apartment and office buildings, but the effects of these high-rise spaces on the vertical dispersal, oviposition and blood feeding behavior of Aedes albopictus are unknown. METHODS: In six multi-story building blocks in downtown Shanghai, 174 mosq-ovitraps (MOT) were placed both indoors and outdoors for Ae. albopictus collection at different vertical heights from the 1st to 6th floors and a terrace on the 8th floor. Collections were made for 4 months. The human landing catch (HLC) method for Ae. albopictus monitoring was also conducted on 6 consecutive days on six floors of two of the six buildings to study the feeding behavior of Ae. albopictus at different heights. RESULTS: Both MOTs and HLCs collected Ae. albopictus at all monitored heights. The vertical distribution, oviposition pattern and biting behavior varied significantly among the seven heights (1st-6th floors and 8th floor) (mosq-ovitrap index (MOI): X2 = 140.616, df = 6, P < 0.001; HLC: F (5, 138) = 15.111, P < 0.001). The MOI at low heights (1st + 2nd floors) was significantly higher than that at medium (3rd + 4th floor, P < 0.001) and high heights (5th + 6th floors, P < 0.001), and there was no significant difference in the MOI for the 3rd-6th floors. The outdoor MOIs were significantly higher than indoor MOIs at all heights (outdoor 23.09% vs. indoor 9.58%, X2 = 74.121, df = 1, P < 0.001). Aedes albopictus HLC density on the ground floor was significantly higher than that on all other heights (5.04 vs. 0.13, 0.29, 0.58, 0.79 and 1.50 per half hour, P < 0.05), while no difference was detected among the heights above the ground floor (P > 0.05). CONCLUSIONS: Aedes albopictus is more common near the ground level, but it can easily disperse to higher floors in the multi-story buildings of urban Shanghai. No significant differences in Ae. albopictus density were detected within the 3rd-6th floors using MOT or HLC. This suggests that Ae. albopictus might also disperse to areas above the 6th floor and seek hosts there. Aedes albopictus prefers to oviposit outdoors; however, Ae. albopictus was also able to inhabit, oviposit and engage in blood-feeding behavior indoors on different floors. The three-dimensional dispersal pattern of Ae. albopictus in urban areas could facilitate arbovirus transmission and increase the difficulty of dengue control.

Safranal inhibits estrogen-deficiency osteoporosis by targeting Sirt1 to interfere with NF-κB acetylation.

BACKGROUND: Osteoporosis is a prevalent bone metabolic disease in menopause, and long-term medication is accompanied by serious side effects. Estrogen deficiency-mediated hyperactivated osteoclasts is the initiating factor for bone loss, which is regulated by nuclear factor-κB (NF-κB) signaling. Safranal (Saf) is a monoterpene aldehyde produced from Saffron (Crocus sativus L.) and possesses multiple biological properties, particularly the anti-inflammatory property. However, Saf's role in osteoporosis remains unknown. PURPOSE: This study aims to validate the role of Saf in osteoporosis and explore the potential mechanism. STUDY DESIGN: The RANKL-exposed mouse BMM (bone marrow monocytes) and the castration-mediated osteoporosis model were applied to explore the effect and mechanism of Saf in vitro and in vivo. METHOD: The effect of Saf on osteoclast formation and function were assessed by TRAcP staining, bone-resorptive experiment, qPCR, immunoblotting and immunofluorescence, etc. Micro-CT, HE, TRAcP and immunohistochemical staining were performed to estimate the effects of Saf administration on OVX-mediated osteoporosis in mice at imaging and histological levels. RESULTS: Saf concentration-dependently inhibited RANKL-mediated osteoclast differentiation without affecting cellular viability. Meanwhile, Saf-mediated anti-osteolytic capacity and Sirt1 upregulation were also found in ovariectomized mice. Mechanistically, Saf interfered with NF-κB signaling by activating Sirt1 to increase p65 deacetylation and inactivating IKK to decrease IκBα degradation. CONCLUSION: Our results support the potential application of Saf as a therapeutic agent for osteoporosis.

VK2 Promotes Osteogenic Differentiation of BMSCs against High Glucose Exposure via Modulation of Intracellular Oxidative Stress.

INTRODUCTION: Diabetic osteoporosis (DOP) has gradually gained public attention. The clinical manifestations of DOP include bone mass loss, bone microstructural damage, and increased bone fragility.

Cedrol, a Ginger-derived sesquiterpineol, suppresses estrogen-deficient osteoporosis by intervening NFATc1 and reactive oxygen species.

Osteoporosis is a prevalent bone metabolic disease in menopause, and long-term medication is accompanied by serious side effects. Ginger, a food spice and traditional medicine with ancient history, exhibits the potential to alleviate osteoporosis in preclinical experiments, whereas its complex composition leads to ambiguous pharmacological mechanisms. The purpose of this study was to investigate the effect and mechanism of Ced in estrogen-deficient osteoporosis, a sesquiterpene alcohol recently discovered from Ginger with multiple pharmacological properties. RANKL was stimulated BMM (bone marrow macrophages) differentiation into osteoclasts in vitro. And the osteoclast activity and number were assessed by TRAcP and SEM. We found that Ced mitigated RANKL-induced osteoclastogenesis by descending the ROS content and obstructing NFATc1, NF-κB, and MAPK signaling. Also, Ced-mediated anti-osteolytic property was found in ovariectomized mice by Micro-CT scanning and histological staining. Summarily, our works demonstrated the anti-osteoporotic potential of Cedrol in Ginger for the first time, which also offered more pharmacological evidence for Ginger as food or medicine used for bone metabolic disease.

Diagnostic accuracy and prognostic significance of osteopontin in liver cirrhosis and hepatocellular carcinoma: a Meta-analysis.

OBJECTIVE: At present, there is no definite suggestion about effective tumour biomarkers for the diagnostic accuracy and prognostic significance of hepatocellular carcinoma (HCC) and liver cirrhosis (LC). The aim of our research was to determine the value of the tumour biomarker osteopontin (OPN), which is encoded by the Spp1 gene, in the diagnosis, prognosis and development of HCC and LC through meta-analysis. METHODS: A systematic literature search was performed in the PubMed, Embase, Cochrane Library and China National Knowledge Infrastructure electronic databases up to March 2021. Studies evaluating the diagnostic and/or prognostic value of OPN in HCC and/or LC were included. RESULTS: From the systematic search, 35 studies including 9150 participants were eligible, 25 of which provided data on the diagnostic value of OPN overexpression, while 15 studies provided data on the prognostic value. OPN had high diagnostic accuracy in both HCC and LC patients compared with healthy controls, and the diagnostic efficiency was increased by the biomarker combination OPN + AFP. CONCLUSIONS: OPN may be adopted as a promising predictive tumour biomarker for the diagnosis and prognosis of HCC and LC and may be a potential therapeutic target.

Mechanical Ventilation Exacerbates Poly (I:C) Induced Acute Lung Injury: Central Role for Caspase-11 and Gut-Lung Axis.

Background: The mechanisms by which moderate tidal volume ventilation (MTV) exacerbates preexisting lung injury are unclear. We hypothesized that systemic endotoxemia via the gut-lung axis would lead to non-canonical and canonical inflammasome activation and pyroptosis in a two-hit model involving polyinosinic-polycytidylic acid (Poly(I:C)), a synthetic analog of dsRNA and MTV and that this would associate with acute lung injury (ALI). Methods: Anesthetized mice were administered Poly(I:C) intratracheally and then 6 h later, they were mechanically ventilated for 4 h with otherwise non-injurious MTV (10ml/kg). Changes in intestinal and alveolar capillary permeability were measured. Further documentation of ALI was assessed by evans blue albumin permeability, protein and IL-1 family concentration in bronchoalveolar lavage fluid (BALF) or plasma, and histopathology in cohorts of wildtype (WT), whole body genetically ablated caspase-11 (caspase-11-/-), caspase-1/caspase-11 double knockout (caspase-1/11-/-), gasdermin D (GSDMD)-/-, nucleotide-binding domain leucine-rich repeat-containing protein 3 (NLRP3)-/- and advanced glycosylation end product-specific receptor (RAGE) -/- mice. Results: Non-injurious MTV exacerbated the mild lung injury associated with Poly(I:C) administration. This included the disruption of alveolar-capillary barrier and increased levels of interleukin (IL)-6, high mobility group proteins 1 (HMGB-1), IL-1ß in BALF and IL-18 in plasma. Combined (Poly(I:C)-MTV) injury was associated with increase in gastrointestinal permeability and endotoxin in plasma and BALF. Poly(I:C)-MTV injury was sensitive to caspase-11 deletion with no further contribution of caspase-1 except for maturation and release of IL-18 (that itself was sensitive to deletion of NLRP3). Combined injury led to large increases in caspase-1 and caspase-11. Genetic ablation of GSDMD attenuated alveolar-capillary disruption and release of cytokines in combined injury model. Conclusions: The previously noted exacerbation of mild Poly(I:C)-induced ALI by otherwise non-injurious MTV is associated with an increase in gut permeability resulting in systemic endotoxemia. The gut-lung axis resulted in activation of pulmonary non-canonical (cytosolic mediated caspase-11 activation) and canonical (caspase-1) inflammasome (NLRP3) mediated ALI in this two-hit model resulting in GSDMD sensitive alveolar capillary barrier disruption, pyroptosis (alveolar macrophages) and cytokine maturation and release (IL-1ß; IL-18). Pharmacologic strategies aimed at disrupting communication between gut and lung, inhibition of inflammasomes or GSDMD in pyroptosis may be useful in ALI.

Genotyping Helicobacter pylori antibiotic resistance and virulence-associated genes in patients with gastric cancer in Wenzhou, China.

BACKGROUND AND STUDY AIMS: Helicobacter pylori infection affects approximately 50% of the global population and has become a serious health concern related to gastric cancer, gastritis, and peptic ulcers. This organism acquires drug resistance through gene mutations, and its increasing resistance to antibiotics has severely influenced the effectiveness of eradication efforts. Therefore, we designed this study to determine the prevalence of H. pylori virulence- (cagA and vacA) and antibiotic resistance - associated genotypes in patients with gastric cancer infected with H. pylori in Whenzhou, China. PATIENTS AND METHODS: We used polymerase chain reaction (PCR) to confirm H. pylori in cancerous and paracancerous tissue specimens from 225 patients. Then we tested the prevalence of virulence- and antibiotic resistance - associated genotypes in H. pylori using a PCR-based DNA-sequencing assay. RESULTS: We observed H. pylori DNA in 222 of the 225 patients and found the most prevalent virulence-associated genotypes in cagA+ (97.75%) and vacAs1m1 (93.25%). Metronidazole resistance - associated gene mutation was G616A in rdxA; levofloxacin resistance - associated gene mutations were N87K, N87I, and D91G in gyrA; clarithromycin resistance - associated gene mutations were A2143G and A2142G in 23SrRNA; and amoxicillin resistance - associated gene mutation was T556S in pbp1. The most prevalent mutation related to antibiotic resistance was present in rdxA (97.30%), followed by gyrA (41.44%) and 23SrRNA (16.67%); the least prevalent was in pbp1 (2.25%). We observed single-gene mutations in 102 patients (45.95%) and found mutations in multiple genes (≥2 genes) in 116 patients (52.25%). CONCLUSION: Patients with gastric cancer in Wenzhou, China, have high incidence infection caused by H. pylori with high-toxicity virulence genotypes. The frequency of gene mutations associated with metronidazole, levofloxacin, and clarithromycin resistances was high and that associated with amoxicillin resistance was relatively low. The mutation patterns were diverse, and the rates of multiple gene mutations were high.

Patterns of immune infiltration in gastric cancer and their clinical significance.

OBJECTIVE: The malignant phenotypes of cancer are defined not only by its intrinsic tumor cells but also by the tumor-infiltrating immune cells activated and recruited to the cancer microenvironment. However, a comprehensive introduction of gastric cancer immune cell infiltration has not been identified so far. METHODS: In this study, we comprehensively analyzed the tumor-infiltrating immune cells abundance in gastric cancer for the first time by CIBERSORT. The meta-analysis, single-sample gene set enrichment analysis and hierarchical agglomerative clustering were used to measure and evaluate the respective proportions of 22 cell types of immune infiltration using normalized gene expression data. The fraction of tumor-infiltrating immune cells subpopulations was also evaluated to determine the associations with clinical features and molecular subtypes. RESULTS: Tumor-infiltrating immune cells are extensively involved in the pathogenesis and development of the gastric cancer. We discovered Tfh and activated CD4+ memory T cells were associated with poorer overall survival and Progression-free survival (PFS), but that naïve B cells were opposite for PFS. Unsupervised clustering analysis revealed there existed three tumor-infiltrating immune cells subgroups with distinct survival patterns. Specially, cluster 1 showed significantly better clinical outcome than other two clusters. CONCLUSIONS: Collectively, our data explored the differences of tumor-infiltrating immune cells in gastric cancer, and these variations were likely to be important clues for prognosis and management of its future clinical implementation.

LPS Induces Active HMGB1 Release From Hepatocytes Into Exosomes Through the Coordinated Activities of TLR4 and Caspase-11/GSDMD Signaling.

High-mobility group box-1 (HMGB1), a ubiquitous nuclear protein, acts as a late mediator of lethality when released extracellularly during sepsis. The major source of circulating HMGB1 in sepsis is hepatocytes. However, the mechanism of HMGB1 release of hepatocytes during sepsis is not very clear. We have previously shown that bacterial endotoxin [lipopolysaccharide (LPS)] sensing pathways, including Toll-like receptor (TLR)4 and caspase-11, regulate hepatocyte HMGB1 release in response to LPS. Here, we report the novel function of caspase-11 and gasdermin D (GsdmD) in LPS-induced active HMGB1 released from hepatocytes. HMGB1 release during endotoxemia was caspase-11/GsdmD dependent via an active way in vivo and in vitro. Caspase-11/GsdmD was responsible for HMGB1 translocation from nucleus to the cytoplasm via calcium changing-induced phosphorylation of calcium-calmodulin kinase kinase (camkk)ß during endotoxemia. Cleaved GsdmD accumulated on the endoplasmic reticulum, suggesting this may lead to calcium leak and intracellular calcium increase. Furthermore, we investigated that exosome was an important pathway for HMGB1 release from hepatocytes; this process was dependent on TLR4, independent of caspase-11 and GsdmD in vivo and in vitro. These findings provide a novel mechanism that TLR4 signaling results in an increase in caspase-11 expression, as well as increased exosome release, while caspase-11/GsdmD activation/cleavage leads to accumulation of HMGB1 in the cytoplasm through a process associated with the release of calcium from the endoplasmic reticulum and camkkß activation.

Field evaluation of Mosq-ovitrap, Ovitrap and a CO2-light trap for Aedes albopictus sampling in Shanghai, China.

BACKGROUND: The Mosq-ovitrap (MOT) is currently used for routine surveillance of container-breeding Aedes in China. However, the effectiveness of monitoring Aedes albopictus using the MOT and other mosquito monitoring methods, such as the Ovitrap (OT) and the CO2-light trap (CLT), have not been extensively compared. Moreover, little is known about the spatial-temporal correlations of eggs with adult Ae. albopictus abundance among these three types of traps. METHODS: Comparative field evaluation of MOT, OT and CLT for Ae. albopictus monitoring was conducted simultaneously at two city parks and three residential neighborhoods in downtown Shanghai for 8 months from April 21 to December 21, 2017. RESULTS: Significantly more Ae. albopictus eggs were collected from both MOTs and OTs when traps remained in the field for 10 d or 7 d compared with 3 d (MOT: 50.16, 34.15 vs. 12.38 per trap, P < 0.001; OT: 3.98, 2.92 vs. 0.63 per trap, P < 0.001). Egg collections of MOTs were significantly greater than OTs for all three exposure durations (Percent positive: X 2 = 72.251, 52.420 and 51.429, P value all < 0.001; egg collections: t = 8.068, 8.517 and 10.021, P value all <0.001). Significant temporal correlations were observed between yields of MOT and CLT in all sampling locations and 3 different MOT exposure durations (correlation coefficient r ranged from 0.439 to 0.850, P values all < 0.05). However, great variation was found in the spatial distributions of Ae. albopictus density between MOT and CLT. MOT considerably underestimated Ae. albopictus abundances in areas with high Ae. albopictus density (>25.56 per day ⋅ trap by CLT). CONCLUSION: The MOT was more efficient than the OT in percent positive scores and egg collections of Ae. albopictus. The minimum length of time that MOTs are deployed in the field should not be less than 7 d, as Ae. albopictus collections during this period were much greater than for 3 d of monitoring. MOT considerably underestimated Ae. albopictus abundance in areas with high Aedes albopictus density compared to CLT. In areas with moderate Aedes albopictus densities, MOT results were significantly correlated with CLT catches.

Human Adipose Tissue-Derived Stromal Cells Attenuate the Multiple Organ Injuries Induced by Sepsis and Mechanical Ventilation in Mice.

Mechanical ventilation (MV) can augment sepsis-induced organ injury. Previous studies indicate that human mesenchymal stem cells (hMSCs) have immune-modulatory effect. We hypothesize that human adipose tissue-derived stromal cells (hADSCs) could attenuate MV and sepsis-induced organ injury. Male C57BL/6 mice were randomized to five groups: Sham group; MV group; cecal ligation and puncture (CLP) group; CLP + MV group; and CLP + MV + hADSC group. Anesthetized mice were subjected to cecal ligation and puncture surgery. The mice then received mechanical ventilation (12 ml/kg), with or without the intervention of hADSCs. The survival rate, organ injury of the liver and kidney, total protein and cells in bronchoalveolar lavage fluid (BALF), and histological changes of the lung and liver were examined. The level of IL-6 in BALF was measured by ELISA. Real-time quantitative PCR was used to analyze mRNA of IL-6 and tumor necrosis factor-α (TNF-α). hADSC treatment increased survival rate of septic mice with MV. hADSCs attenuated dysfunction of the liver and kidney and decreased lung inflammation and tissue injury of the liver and lung. IL-6 level in BALF and TNF-α and IL-6 mRNA expression in the tissue of the lung, liver, and kidney were significantly reduced by hADSC treatment. MV with conventional tidal volume aggravates CLP-induced multiple organ injuries. hADSCs inhibited the compound injuries possibly through modulation of immune responses.