Biomass fuels related-PM2.5 promotes lung fibroblast-myofibroblast transition through PI3K/AKT/TRPC1 pathway.

Emerging evidence has suggested that exposure to PM2.5 is a significant contributing factor to the development of chronic obstructive pulmonary disease (COPD). However, the underlying biological effects and mechanisms of PM2.5 in COPD pathology remain elusive. In this study, we aimed to investigate the implication and regulatory effect of biomass fuels related-PM2.5 (BRPM2.5) concerning the pathological process of fibroblast-to-myofibroblast transition (FMT) in the context of COPD. In vivo experimentation revealed that exposure to biofuel smoke was associated with airway inflammation in rats. After 4 weeks of exposure, there was inflammation in the small airways, but no significant structural changes in the airway walls. However, after 24 weeks, airway remodeling occurred due to increased collagen deposition, myofibroblast proliferation, and tracheal wall thickness. In vitro, cellular immunofluorescence results showed that with stimulation of BRPM2.5 for 72 h, the cell morphology of fibroblasts changed significantly, most of the cells changed from spindle-shaped to star-shaped irregular, α-SMA stress fibers appeared in the cytoplasm and the synthesis of type I collagen increased. The collagen gel contraction experiment showed that the contractility of fibroblasts was enhanced. The expression level of TRPC1 in fibroblasts was increased. Specific siRNA-TRPC1 blocked BRPM2.5-induced FMT and reduced cell contractility. Additionally, specific siRNA-TRPC1 resulted in a decrease in the augment of intracellular Ca2+ concentration ([Ca2+]i) induced by BRPM2.5. Notably, it was found that the PI3K inhibitor, LY294002, inhibited enhancement of AKT phosphorylation level, FMT occurrence, and elevation of TRPC1 protein expression induced by BRPM2.5. The findings indicated that BRPM2.5 is capable of inducing the FMT, with the possibility of mediation by PI3K/AKT/TRPC1. These results hold potential implications for the understanding of the molecular mechanisms involved in BRPM2.5-induced COPD and may aid in the development of novel therapeutic strategies for pathological conditions characterized by fibrosis.

Kai-xin-san improves cognitive impairment in D-gal and Aß25-35 induced ad rats by regulating gut microbiota and reducing neuronal damage.

ETHNOPHARMACOLOGICAL RELEVANCE: Kai-Xin-San (KXS) is a classic herbal formula for the treatment and prevention of AD (Alzheimer's disease) with definite curative effect, but its mechanism, which involves multiple components, pathways, and targets, is not yet fully understood. AIM OF THE STUDY: To verify the effect of KXS on gut microbiota and explore its anti-AD mechanism related with gut microbiota. MATERIALS AND METHODS: AD rat model was established and evaluated by intraperitoneal injection of D-gal and bilateral hippocampal CA1 injections of Aß25-35. The pharmacodynamics of KXS in vivo includes general behavior, Morris water maze test, ELISA, Nissl & HE staining and immunofluorescence. Systematic analysis of gut microbiota was conducted using 16S rRNA gene sequencing technology. The potential role of gut microbiota in the anti-AD effect of KXS was validated with fecal microbiota transplantation (FMT) experiments. RESULTS: KXS could significantly improve cognitive impairment, reduce neuronal damage and attenuate neuroinflammation and colonic inflammation in vivo in AD model rats. Nine differential intestinal bacteria associated with AD were screened, in which four bacteria (Lactobacillus murinus, Ligilactobacillus, Alloprevotella, Prevotellaceae_NK3B31_group) were very significant. CONCLUSION: KXS can maintain the ecological balance of intestinal microbiota and exert its anti-AD effect by regulating the composition and proportion of gut microbiota in AD rats through the microbiota-gut-brain axis.

Active microbial metabolites study on antitussive and expectorant effects and metabolic mechanisms of platycosides fraction of Platycodonis Radix.

A new method involving gut microbiota biotransformation, spectrum-effect relationship analysis and metabolomics analysis was developed to study the antitussive and expectorant microbial metabolites of platycosides fraction (MPFs) of Platycodonis Radix. Furthermore, their possible metabolic mechanisms were studied for the first time. The findings showed that the antitussive and expectorant effects of the platycosides fraction (PF) were significantly enhanced by the gut microbiota biotransformation. 11 active antitussive microbial metabolites and 12 active expectorant microbial metabolites, which shared 8 components, were successfully screened out via spectrum-effect relationship analysis. The prototypes of the active microbial metabolites could be reversely traced according to the gut microbiota biotransformation pathways. It was found out that one platycoside could produce several active microbial metabolites and several different platycosides could produce the same active microbial metabolite. In addition, the metabolomics analysis showed that both the PF and its active microbial metabolites could regulate the same metabolomic pathways of Linoleic acid metabolism, Arachidonic acid metabolism and Glycerophospholipid metabolism to exert antitussive activity, and regulate the same metabolomic pathway of Arachidonic acid metabolism to exert expectorant activity. These findings suggested the microbial metabolites may be the active forms of the platycosides. Overall, the proposed approach was useful in screening the active microbial metabolites; this work explained the in vivo antitussive and expectorant metabolic mechanisms of multi-constituents, multi-targets and synergistic effects of PF of Platycodonis Radix.

A Comparative Study of Serum Pharmacochemistry of Kai-Xin-San in Normal and AD Rats Using UPLC-LTQ-Orbitrap-MS.

Kai-Xin-San (KXS) is a classic formula for the treatment of Alzheimer's disease (AD). KXS has been widely used to treat emotional diseases; however, its active components remain unknown. There have been some reports about the efficacy and metabolic analysis of KXS, which are mainly based on studying normal animals. The current work first established an AD rat model by injecting D-galactose into the abdominal cavity and injecting Aß25-35 into the hippocampus on both sides, followed by intragastric administration of KXS for a consecutive week; then, the analytical method for ethanol extraction from the serum of normal and model rats was developed using UPLC-LTQ-Orbitrap-MS; finally, the transitional components in the blood were systematically compared and analyzed by multivariate statistical analysis. A total of 36 components of KXS were identified in the rat serum of the normal group, including 24 prototype components (including ginsenosides, triterpenoid acids of Poria cocos, polygala saponins, polygala xanthones and polygala ester) and 13 metabolites (including desugar, hydration and oxidation products of ginsenosides, triterpenoid acid hydroxylation, deoxygenation, demethylation, desaturation, and glycine-conjugated products of Poria cocos). Twenty KXS-relevant components were detected in the rat serum of the model group, including 11 prototypes and 9 metabolites. The normal group and the model group shared 12 common components, including 9 prototypes and 3 metabolites. The intestinal microecological balance of the model rats probably was destroyed, affecting the absorption/metabolism of saponins by the body, which resulted in fewer transitional components in the model group. This study reflected the drug-body interaction from an objective and accurate perspective, offering references and insights for elucidating the basis of active components and mechanism of action of KXS for treating AD.

Hepatotoxicity Comparison of Crude and Licorice-Processed Euodiae Fructus in Rats With Stomach Excess-Cold Syndrome.

In recent years, drug-induced liver injury (DILI) has become an important issue of public health. Euodiae Fructus (EF) is a commonly used herb with mild toxicity in clinic, and large doses of EF can cause significant liver damage. Licorice processing might reduce the hepatotoxicity of CEF (crude EF), but up to now, studies on the hepatotoxicity of EF have been hardly reported, let alone its material basis and mechanism of detoxification by licorice processing. This work firstly established a stomach excess-cold syndrome animal model induced by intragastric administration of cold Zhimu (Anemarrhena asphodeloides Bge). Secondly, multiple approaches and indexes were used to evaluate the hepatotoxicity of the drugs in the rats including general behavior, biochemical analysis, protein expressions, and histopathological examination. Thirdly, the hepatotoxicity of three doses of three CEF and LPEF (licorice-processed EF) extracts was systematically investigated, and the hepatotoxicity differences were analyzed and compared comprehensively among the three extracts, three doses, and CEF and LPEF. Finally, the connotation of detoxification of EF by licorice processing was preliminarily discussed according to the changes in toxic components after processing, toxicological characteristics, and TCM (traditional Chinese medicine) theory. All extracts of EF were found to have dose-dependent hepatotoxicity, and the toxicity was in the descending order of water extract, ethanol extract, and volatile oil. The hepatotoxic mechanism of EF may be related to peroxidation damage, inflammatory factor, and mitochondrial injury. The CEF hepatotoxicity can be significantly reduced by licorice processing. EF should be safe for short-term use at pharmacopeial dose under the guidance of the TCM theory. The detoxification mechanism is probably related to the reduction of toxic components and antagonistic action of licorice.

Bioactive constituents of Mosla chinensis-cv. Jiangxiangru ameliorate inflammation through MAPK signaling pathways and modify intestinal microbiota in DSS-induced colitis mice.

BACKGROUND: Mosla chinensis Maxim. cv. Jiangxiangru (JXR), a traditional Chinese medicine, commonly used for the therapy of cold, fever, diarrhea, digestive disorders, and other diseases. Inflammatory bowel disease (IBD) is a chronic disorder of the human gastrointestinal tract. Research about the effect of JXR on IBD and the active ingredient composition of JXR remains deficiency. PURPOSE: This study aims to determine the phytochemical composition and the anti-inflammatory property of JXR, as well as the possible anti-inflammatory mechanisms. METHODS: The bioactive profile of JXR extracts was determined by UPLC-LTQ-Orbitrap-MS. A DSS induced colitis mouse model was applied to explore the anti-inflammatory activity of JXR. The body weight, colon length and histopathological status of colon tissue were evaluated. The content of inflammatory mediators (nitric oxide (NO), prostaglandin E2 (PGE2)) and cytokines (tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1ß (IL-1ß)), corresponding mRNA and protein expression levels were analyzed. Oxidation pressure and gut microbial composition were also explored. RESULTS: Totally 63 constitutes were identified from JXR, among them, phenolic acids and flavonoids comprised a large part, and rosmarinic acid (RA) was the main compound. The results of DSS-induced colitis mice model indicated that JXR effectively ameliorated inflammation, restore the redox balance in the gut. JXR treatment significantly reduced the production of reactive oxygen species (ROS), increased the activity of antioxidative enzyme, suppressed the secretion of inflammatory mediators (NO, PGE2) and cytokines (TNF-α, IL-6, IL-1ß). JXR also restrained the activation of mitogen-activated protein kinases (MAPKs) signaling pathway. Furthermore, JXR could restore the microbial diversity by suppressing Bacteroidaceae, increasing Bifidobacteriales and Melainabacteria in DSS colitis mouse model. CONCLUSIONS: This study demonstrated that JXR composed with various bioactive compounds, effectively ameliorated colitis, restored the redox balance and regulated gut microbiota. Results from the present study provide an insight of therapeutic potential of JXR in IBD based on its anti-inflammatory and antioxidant properties, also provide a scientific basis for using JXR as a functional ingredient to promote colon health.

Spectrum-effect relationship between UHPLC-Q-TOF/MS fingerprint and promoting gastrointestinal motility activity of Fructus aurantii based on multivariate statistical analysis.

ETHNOPHARMACOLOGICAL RELEVANCE: Nowadays, gastrointestinal motility disorders (GMD) have reduced the quality of people's daily life worldwide, but there is still a lack of effective western medicine treatment. Fructus aurantii (FA), a representative regulating-qi herbal medicine, has been widely used to treat GMD in China for thousands of years, but it is not clear that which specific components contribute to the efficacy. AIM OF THE STUDY: The efficacy differences of various fractions of FA on normal mice and GMD rats were compared, so as to find out the main effective fraction of FA, and to screen the main regulating-qi components based on spectrum-effect relationship and multivariate statistical analysis. MATERIALS AND METHODS: The fingerprints of different fractions of FA were established and main compounds were identified with UHPLC-Q-TOF/MS technique. The promoting gastrointestinal motility activities of FA were evaluated by defecation test, gastric emptying and intestinal propulsion test in mice, and further investigated according to the biochemical analysis of 5-HT, SP, MLT, GAS and VIP in GMD rats' plasma. One-way ANOVA was used to find out the difference of efficacy. The active components were screened through spectrum-effect relationship with PCA-X, Pearson bivariate correlation analysis and OPLS analysis. CONCLUSIONS: Ethyl acetate fraction is the main active fraction, and nine compounds are the major regulating-qi components. The developed spectrum-effect analysis can be used for the screening of bioactive components in natural products with high accuracy and reliability.

Components study on antitussive effect and holistic mechanism of Platycodonis Radix based on spectrum-effect relationship and metabonomics analysis.

The antitussive effect of Platycodonis Radix is closely related to the components in saponins fraction of Platycodonis Radix extract (SFPRE); however, these active components and their holistic mechanism remain unknown. Hence, a new method by integrating spectrum-effect relationship analysis with metabolomics analysis was applied to study the active components and their holistic mechanism simultaneously. For spectrum-effect relationship analysis, chemical fingerprints of ten batches of SFPRE were developed using UHPLC-LTQ-Orbitrap MSn; antitussive effect were evaluated using a classic mice-cough model induced by ammonia liquor. Spectrum-effect relationship was analyzed by partial least squares regression (PLSR) analysis. For metabolomics analysis, the altered metabolites related to cough in serum were identified by UHPLC-Q-TOF/MS and orthogonal partial least squares-discriminant analysis (OPLS-DA); metabolic pathway analysis was depended on MetaboAnalyst 4.0, KEGG database, METLIN database and HMDB database. Our findings showed that 10 identified components of Polygalacin D (peak 26), Deapio-platycodin D (peak 21), Platycodin D (peak 23), ß-Gentiotriosyl platycodigenin (peak 37), Platycoside G3 (peak 17), Platycoside C (peak 25), Platycodin D3 (peak 16), 3-O-ß-D-glucopyranosyl platycodigenin (peak 33), Platycoside F (peak 19) and 3″-O-acetyl platycodin D3 (peak 15), and 2 unidentified components (peak 45 and 44) possessed antitussive effects. The metabolomics analysis result showed that 19 metabolites were potential biomarkers related to the cough, 16 of which could be restored to normal levels by SFPRE. These biomarkers were involved in arachidonic acid metabolism, linoleic acid metabolism and glycerophospholipid metabolism. The current study may facilitate the development of antitussive medicines with fewer side-effects based on Platycodonis Radix.

Open tubular capillary column immobilized with sulfobutylether-ß-cyclodextrin for chiral separation in capillary electrochromatography.

A novel chiral open tubular capillary column was fabricated with sulfobutylether-ß-cyclodextrin and glycidyl methacrylate for enantioseparation in capillary electrochromatography. First, the pretreated silica-fused capillary was treated with 3-trimethoxysilyl propyl methacrylate to attach double bond ligand onto the surface. A copolymer layer was formed on the surface of capillary using glycidyl methacrylate and ethylene dimethacrylate by in situ one-pot polymerization. Sulfobutylether-ß-cyclodextrin was encapsulated inside the copolymerized layer. The morphology of the developed column was characterized by field emission scanning electron microscopy. The effect of organic percentage and pH value of the mobile phase on electroosmotic flow and resolution was also investigated. The performance of the fabricated column was validated by separation of amlodipine besilate, 2,3-diphenylpropionic acid, tropic acid, and pantoprazole enantiomers with good resolutions of 3.67, 4.82, 3.34, and 2.61, respectively. The repeatabilities of column-to-column and day-to-day through relative standard deviation were found better than 4%, exhibiting satisfactory repeatability of the developed column. The results reveal that open tubular capillary columns modified with ß-cyclodextrin show a great prospect for enantioseparation of chiral drugs in capillary electrochromatography.

A phase 2, randomized, double-blind, placebo-controlled trial of AMG 301, a pituitary adenylate cyclase-activating polypeptide PAC1 receptor monoclonal antibody for migraine prevention.

OBJECTIVE: To assess the safety and efficacy of AMG 301, an inhibitor of the pituitary adenylate cyclase-activating polypeptide (PACAP)-1 (PAC1) receptor, for prevention of migraine. METHODS: In a double-blind trial, patients were randomized 4:3:3 to placebo, AMG 301 210 mg every 4 weeks, or AMG 301 420 mg every 2 weeks for 12 weeks. Effect on monthly migraine days and other secondary measures were assessed over weeks 9-12. Safety and tolerability were assessed. RESULTS: Of 343 randomized patients (mean age, 41.8-42.5 years), the majority were women (85.4-90.4%), white (94.1-96.2%), and had episodic migraine (62.5-67.9%). A total of 305 patients completed treatment (placebo, n = 124; AMG 301 210 mg, n = 94; AMG 301 420 mg, n = 87). Least squares mean reduction at week 12 in monthly migraine days from baseline was -2.5 (0.4) days for placebo and -2.2 (0.5) days for both AMG 301 treatment groups. No difference between AMG 301 and placebo on any measure of efficacy was observed; mean (95% confidence interval) treatment difference versus placebo for monthly migraine days for AMG 301 210 mg, 0.3 (-0.9 to 1.4); AMG 301 420 mg, 0.3 (-0.9 to 1.4). The incidence of adverse events was similar across groups. CONCLUSION: AMG 301 offered no benefit over placebo for migraine prevention; further studies may be necessary to fully understand the role of PACAP isoforms and its receptors in migraine pathophysiology. STUDY REGISTRATION: ClinicalTrials.gov: NCT03238781.

Evaluation of hepatic steatosis before liver transplantation in ex vivo by volumetric quantitative PDFF-MRI.

PURPOSE: Over the last two decades, extended criteria have promoted an increased number of donor livers available for liver transplantation. But posttransplant graft loss is still a major concern. Macrovesicular hepatic steatosis (MHS) is recognized as the most significant prognostic histologic parameter in predicting posttransplant graft loss. We aimed to evaluate the utility of ex vivo volumetric quantitative MRI for quantifying MHS before liver transplantation using proton density fat-fraction (PDFF-MRI) histogram analysis. METHODS: PDFF-MRI was performed at 3.0T in 40 livers. We obtained histogram parameters of whole-liver volume of interest, including the mean, median, 5th, 10th, 25th, 75th, 90th, and 95th percentile PDFF; skewness; kurtosis; entropy; and volume. RESULTS: Livers from 40 cadaveric donors were included, and histologic ex vivo fat quantification was available for 33 livers. Ten livers had MHS and 23 had normal fat content. The MHS group had higher mean, median, 5th, 10th, 25th, 75th, 90th, and 95th percentile PDFF, and entropy than the group with normal fat content (P < .05). Median PDFF had greater area under the curve value than other parameters. Mean PDFF showed an excellent correlation with entropy and a moderate correlation with MHS quantification on histology. CONCLUSIONS: Ex vivo volumetric quantitative PDFF-MRI histogram analysis is a very useful and noninvasive method to detect MHS before liver transplantation. Median PDFF was the best predictor of the presence of MHS. Entropy is a very promising parameter.

Correction: TRPC6-dependent Ca2+ signaling mediates airway inflammation in response to oxidative stress via ERK pathway.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Gegen Qinlian Decoction abates nonalcoholic steatohepatitis associated liver injuries via anti-oxidative stress and anti-inflammatory response involved inhibition of toll-like receptor 4 signaling pathways.

Gegen Qilian Decoction (GGQLD) is a well-established classic Chinese medicine prescription in treating nonalcoholic steatohepatitis (NASH). However, the molecular mechanism of GGQLD action on NASH is still not clear. This study aimed to assess the anti-NASH effect of GGQLD, and to explore its molecular mechanisms in vivo and in vitro. In HFD-fed rats, GGQLD decreased significantly serum triglyceride (TG), cholesterol (CHO), total bile acid (TBA), low-density lipoprotein (LDL), free fatty acid (FFA) and lipopolysaccharide (LPS) levels, increased levels of differentially expressed proteins (DEPs) Ahcy, Gpx1, Mat1a, GNMT, and reduced the expression of ALDOB. In RAW264.7 macrophages, GGQLD reduced the expression levels of inflammatory factors TNF-α and IL-6 mRNA, and diminished NASH by increasing differentially expressed genes (DEGs) CBS, Mat1a, Hnf4α and Pparα to reduce oxidative stress or lipid metabolism. The results of DEGs verification also showed that GGQLD up-regulated expressions of Hnf4α, Pparα and Cbs genes. In HepG2 cells, GGQLD decreased IL-6 levels and intracellular TG content, and inhibited FFA-induced expression of toll-like receptor 4 (TLR4). In summary, GGQLD abates NASH associated liver injuries via anti-oxidative stress and anti-inflammatory response involved inhibition of TLR4 signal pathways. These findings provide new insights into the anti-NASH therapy by GGQLD.

TRPC6-dependent Ca2+ signaling mediates airway inflammation in response to oxidative stress via ERK pathway.

Ozone (O3) plays an extremely important role in airway inflammation by generating reactive oxygen species (ROS) including hydrogen peroxide, then promoting redox actions and causing oxidative stress. Evidences indicate that TRPC6 (canonical transient receptor potential channel 6) is a redox-regulated Ca2+ permeable nonselective cation channel, but its role in the setting of oxidative stress-related airway inflammation remains unknown. Here, we found that both TRPC6-/- mice and mice pretreated with SAR7334, a potent TRPC6 inhibitor, were protected from O3-induced airway inflammatory responses. In vitro, both knockdown of TRPC6 expression with shRNA and TRPC6 blockage markedly attenuated the release of cytokines IL-6 and IL-8 induced by O3 or H2O2 in 16HBE cells (human bronchial epithelial cell line). Treatment with O3 or H2O2 enhanced TRPC6 protein expression in vivo and vitro. We also observed that TRPC6-dependent increase of intracellular Ca2+ concentration ([Ca2+]i) was triggered by H2O2, which consisted of the release from intracellular calcium store and the influx of extracellular Ca2+ and could be further strengthened by 6-h O3 exposure in both 16HBE cells and HBEpiCs (primary human bronchial epithelial cells). Moreover, we confirmed that the activation of MAPK signals (ERK1/2, p38, JNK) was required for the inflammatory response induced by O3 or H2O2 while only the phosphorylation of ERK pathway was diminished in the TRPC6-knockdown situation. These results demonstrate that oxidative stress regulates TRPC6-mediated Ca2+ cascade, which leads to the activation of ERK pathway and inflammation and could become a potential target to treat oxidative stress-associated airway inflammatory diseases.

CT features and quantitative analysis of subsolid nodule lung adenocarcinoma for pathological classification prediction.

BACKGROUND: The value of the CT features and quantitative analysis of lung subsolid nodules (SSNs) in the prediction of the pathological grading of lung adenocarcinoma is discussed. METHODS: Clinical data and CT images of 207 cases (216 lesions) with CT manifestations of an SSNs lung adenocarcinoma confirmed by surgery pathology were retrospectively analysed. The pathological results were divided into three groups, including atypical adenomatous hyperplasia (AAH)/adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA) and invasive adenocarcinoma (IAC). Then, the quantitative and qualitative data of these nodules were compared and analysed. RESULTS: The mean size, maximum diameter, mean CT value and maximum CT value of the nodules were significantly different among the three groups of AAH/AIS, MIA and IAC and were different between the paired groups (AAH/AIS and MIA or MIA and IAC) (P < 0.05). The critical values of the above indicators between AAH/AIS and MIA were 10.05 mm, 11.16 mm, - 548.00 HU and - 419.74 HU. The critical values of the above indicators between MIA and IAC were 14.42 mm, 16.48 mm, - 364.59 HU and - 16.98 HU. The binary logistic regression analysis of the features with the statistical significance showed that the regression model between AAH/AIS and MIA is logit(p) = - 0.93 + 0.216X1 + 0.004X4. The regression model between MIA and IAC is logit(p) = - 1.242-1.428X5(1) - 1.458X6(1) + 1.146X7(1) + 0.272X2 + 0.005X3. The areas under the curve (AUC) obtained by plotting the receiver operating characteristic curve (ROC) using the regression probabilities of regression models I and II were 0.815 and 0.931. CONCLUSIONS: Preoperative prediction of pathological classification of CT image features has important guiding value for clinical management. Correct diagnosis results can effectively improve the patient survival rate. Through comprehensive analysis of the CT features and qualitative data of SSNs, the diagnostic accuracy of SSNs can be effectively improved. The logistic regression model established in this study can better predict the pathological classification of SSNs lung adenocarcinoma on CT, and the predictive value is significantly higher than the independent use of each quantitative factor.

TRPC6 modulates adhesion of neutrophils to airway epithelial cells via NF-κB activation and ICAM-1 expression with ozone exposure.

Ozone (O3) is a major component of air pollution, which has been associated with airway inflammation characterized by the influx of neutrophils in asthmatic subjects. Canonical transient receptor potential 6 (TRPC6) channel is recently identified as a target of oxidative stress which is involved in airway inflammation. However, the regulatory role of TRPC6 in airway epithelial cells and neutrophils has not yet been illuminated in detail. In this study, we investigated the role of TRPC6 in neutrophil adhesion to airway epithelial cells exposed to O3 in vivo and in vitro approaches. Using transgenic mice, the results showed that TRPC6-deficiency attenuated O3-induced neutrophil recruitment to airway epithelial cells and intercellular adhesion molecule-1 (ICAM-1) expression. In vitro, O3 induced ICAM-1 expression and neutrophil adhesion to 16HBE cells (human airway epithelial cell line) and which were reduced by both TRPC6 silencing short hairpin RNA (shRNA) and TRPC6 inhibitor Larixyl Acetate (LA). We also confirmed that TRPC6-dependent Ca2+ entry and NF-κB activation in 16HBE cells were required for ICAM-1-mediated neutrophil adhesion exposed to O3. In conclusion, this study demonstrated the contribution of TRPC6 to O3-induced neutrophil adhesion to airway epithelial cells via NF-κB activation and ICAM-1 expression, which may provide new potential concepts for preventing and treating air pollutant-related inflammatory lung diseases.

The anti-diabetic activities, gut microbiota composition, the anti-inflammatory effects of Scutellaria-coptis herb couple against insulin resistance-model of diabetes involving the toll-like receptor 4 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Scutellaria-coptis herb couple (SC) is one of the well-known herb couples in many traditional Chinese compound formulas used for the treatment of diabetes mellitus (DM), which has been used to treat DM for thousands of years in China. AIM OF THE STUDY: Few studies have confirmed in detail the anti-diabetic activities of SC in vivo and in vitro. The present investigations aimed to evaluate the anti-diabetic activity of SC in type 2 diabetic KK-Ay mice and in RAW264.7 macrophages to understand its possible mechanism. MATERIALS AND METHODS: High-performance liquid chromatography with ultraviolet detection (HPLC-UV) and LC-LTQ-Orbitrap Pro mass spectrometry were used to analyze the active ingredients of SC extracts and control the quality. A type 2 diabetic KK-Ay mice model was established by high-fat diet. Body weight, fasting blood glucose levels, fasting blood insulin levels, glycosylated hemoglobin and glycosylated serum protein were measured. The effects of SC on total cholesterol (TC), high-density lipoprotein (HDL) and triglyceride (TG) levels were examined. The lipopolysaccharide (LPS), interleukin-6 (IL-6) and tumour necrosis factor alpha (TNF-α) levels were measured. Gut microbial communities were assayed by polymerase chain reaction (PCR) and PCR-denaturing gradient gel electrophoresis (PCR-DGGE) methods. The expressions of Toll-like receptor 4 (TLR4) and MyD88 protein in the colons were measured by western blot. In RAW264.7 macrophages, IL-6, TNF-α, TLR4 and MyD88 protein levels were measured by enzyme-linked immunosorbent assay (ELISA) kits or western blot, and the mRNA expression of IL-6, TNF-α and TLR4 was examined by the real time PCR. RESULTS: The present results showed that the SC significantly increased blood HDL and significantly reduced fasting blood glucose, fasting blood insulin, glycosylated hemoglobin, glycosylated serum protein, TC, TG, LPS, IL-6 and TNF-α levels (P < 0.05 or P < 0.01) in type-2 diabetic KK-Ay mice. Furthermore, SC could regulate the structure of intestinal flora. Additionally, the expressions of TLR4 and MyD88 protein in the colons were significantly decreased in the model group (P < 0.05 or P < 0.01). However, SC had no significant effect on weight gain. In RAW264.7 macrophages, SC containing serum (SC-CS) (5%, 10% and 20%) significantly decreased IL-6, TNF-α, TLR4 and MyD88 protein levels and the mRNA expression of IL-6, TNF-α and TLR4 (P < 0.05 or P < 0.01). CONCLUSIONS: The anti-diabetic effects of SC were attributed to its regulation of intestinal flora and anti-inflammation involving the TLR4 signaling pathway. These findings provide a new insight into the anti-diabetic application for SC in clinical settings and display the potential of SC in the treatment of DM.

Nicotinamide nucleotide transhydrogenase-mediated redox homeostasis promotes tumor growth and metastasis in gastric cancer.

Overcoming oxidative stress is a critical step for tumor growth and metastasis, however the underlying mechanisms in gastric cancer remain unclear. In this study, we found that overexpression of nicotinamide nucleotide transhydrogenase (NNT) was associated with shorter overall and disease free survival in gastric cancer. The NNT is considered a key antioxidative enzyme based on its ability to regenerate NADPH from NADH. Knockdown of NNT caused significantly NADPH reduction, induced high levels of ROS and significant cell apoptosis under oxidative stress conditions such as glucose deprival and anoikis. In vivo experiments showed that NNT promoted tumor growth, lung metastasis and peritoneal dissemination of gastric cancer. Moreover, intratumoral injection of NNT siRNA significantly suppressed gastric tumor growth in patient-derived xenograft (PDX) models. Overall, our study highlights the crucial functional roles of NNT in redox regulation and tumor progression and thus raises an important therapeutic hypothesis in gastric cancer.

Carfilzomib in relapsed or refractory multiple myeloma patients with early or late relapse following prior therapy: A subgroup analysis of the randomized phase 3 ASPIRE and ENDEAVOR trials.

We performed analyses of the randomized phase 3 ASPIRE and ENDEAVOR trials to investigate the efficacy of carfilzomib among subgroups of relapsed or refractory multiple myeloma patients who had early or late disease relapse following initiation of the immediately prior therapy. In ASPIRE and ENDEAVOR, patients had received 1 to 3 prior lines of therapy. Patients in ASPIRE received carfilzomib, lenalidomide, and dexamethasone (KRd) or lenalidomide and dexamethasone (Rd), and patients in ENDEAVOR received carfilzomib and dexamethasone (Kd) or bortezomib and dexamethasone (Vd). Patients with relapse ≤1 year after initiating the most recent prior line of therapy were categorized as early relapsers, and patients with relapse after >1 year were categorized as late relapsers. The median progression-free survival (PFS) in ASPIRE for early relapsers was 21.4 months for KRd vs 10.7 months for Rd (hazard ratio [HR]: 0.714; 95% confidence interval [CI]: 0.508-1.004; P = 0.0257), and for late relapsers was 29.7 months for KRd vs 18.2 months for Rd (HR: 0.675; 95% CI: 0.533-0.854; P = 0.0005). The overall response rate (ORR) for early relapsers was 83.2% for KRd vs 54.8% for Rd, and for late relapsers was 89.0% for KRd vs 69.7% for Rd. The median PFS in ENDEAVOR (Kd vs Vd) for early relapsers was 13.9 months vs 5.7 months (HR: 0.598; 95% CI: 0.423-0.846; P = 0.0017), and for late relapsers was 22.2 months vs 10.2 months (HR: 0.486; 95% CI: 0.382-0.620; P < 0.0001). The ORR (Kd vs Vd) for early relapsers was 63.4% vs 49.1% and for late relapsers was 81.8% vs 66.8%. In conclusion, patients with relapsed or refractory multiple myeloma who received carfilzomib-containing regimens had improved PFS and ORR compared with control groups, regardless of whether they had an early or late relapse following the most recent prior therapy.

TRPC6 contributes to LPS-induced inflammation through ERK1/2 and p38 pathways in bronchial epithelial cells.

receptor potential canonical (TRPC) channels are presently an emerging target for airway disorders. Recent evidence has indicated that TRPC6 as a member of the TRPC family plays an important role in airway inflammation, but its precise function in bronchial epithelial cells remains unclear. The aim of this study was to investigate the role of TRPC6 in Toll-like receptor 4 (TLR4)-mediated inflammation in human bronchial epithelial cells stimulated by endotoxin [lipopolysaccharide (LPS)]. Hyp9 is a simplified phloroglucinol derivative of hyperforin that highly selectively activates TRPC6 channels. The results show that the activation of TRPC6 by Hyp9 induced the production of interleukin (IL)-8 and IL-6. LPS was also able to induce the release of IL-8 and IL-6, which was significantly aggravated by Hyp9 and reduced by knockdown of TRPC6. Treatment with LPS not only chronically induced the expression of TRPC6 mRNA and protein in a TLR4-dependent manner but also acutely increased Ca2+ influx through TRPC6 channels. In addition, LPS-induced overexpression of TRPC6 and Ca2+ influx were associated with the phosphorylation of phosphatidylinositol 3-kinase (PI3K) and Akt. Importantly, TRPC6 was required for the activation of ERK1/2, p38, and NF-κB. In conclusion, these data reveal that LPS induced the overexpression of TRPC6 and TRPC6-dependent Ca2+ influx via the TLR4/PI3K/Akt pathway resulting in Ca2+ mobilization, which subsequently promoted the activation of ERK1/2, p38, and NF-κB and the inflammatory response in bronchial epithelial cells.