Synbindin restrains proinflammatory macrophage activation against microbiota and mucosal inflammation during colitis.

OBJECTIVE: As a canonical membrane tethering factor, the function of synbindin has been expanding and indicated in immune response. Here, we investigated the role of synbindin in the regulation of toll-like receptor 4 (TLR4) signalling and macrophage response to microbiota during colitis. DESIGN: Three distinct mouse models allowing global, myeloid-specific or intestinal epithelial cell-specific synbindin heterozygous deletion were constructed and applied to reveal the function of synbindin during dextran sodium sulfate (DSS) colitis. Effects of synbindin on TLR4 signalling and macrophage activation in response to bacterial lipopolysaccharide (LPS) or Fusobacterium nucleatum were evaluated. The colocalisation and interaction between synbindin and Rab7b were determined by immunofluorescence and coimmunoprecipitation. Synbindin expression in circulating monocytes and intestinal mucosal macrophages of patients with active IBD was detected. RESULTS: Global synbindin haploinsufficiency greatly exacerbated DSS-induced intestinal inflammation. The increased susceptibility to DSS was abolished by gut microbiota depletion, while phenocopied by specific synbindin heterozygous deletion in myeloid cells rather than intestinal epithelial cells. Profoundly aberrant proinflammatory gene signatures and excessive TLR4 signalling were observed in macrophages with synbindin interference in response to bacterial LPS or Fusobacterium nucleatum. Synbindin was significantly increased in intestinal mucosal macrophages and circulating monocytes from both mice with DSS colitis and patients with active IBD. Interleukin 23 and granulocyte-macrophage colony-stimulating factor were identified to induce synbindin expression. Mechanistic characterisation indicated that synbindin colocalised and directly interacted with Rab7b, which coordinated the endosomal degradation pathway of TLR4 for signalling termination. CONCLUSION: Synbindin was a key regulator of TLR4 signalling and restrained the proinflammatory macrophage activation against microbiota during colitis.

Roles of PD-1/PD-L1 Pathway: Signaling, Cancer, and Beyond.

Immunotherapies that target PD-1/PD-L1 axis have shown unprecedented success in a wide variety of human cancers. PD-1 is one of the key coinhibitory receptors expressed on T cells upon T cell activation. After engagement with its ligands, mainly PD-L1, PD-1 is activated and recruits the phosphatase SHP-2 in proximity to T cell receptor (TCR) and CD28 signaling. This event results in dephosphorylation and attenuation of key molecules in TCR and CD28 pathway, leading to inhibition of T cell proliferation, activation, cytokine production, altered metabolism and cytotoxic T lymphocytes (CTLs) killer functions, and eventual death of activated T cells. Bodies evolve coinhibitory pathways controlling T cell response magnitude and duration to limit tissue damage and maintain self-tolerance. However, tumor cells hijack these inhibitory pathways to escape host immune surveillance by overexpression of PD-L1. This provides the scientific rationale for clinical application of immune checkpoint inhibitors in oncology. The aberrantly high expression of PD-L1 in tumor microenvironment (TME) can be attributable to the "primary" activation of multiple oncogenic signaling and the "secondary" induction by inflammatory factors such as IFN-γ. Clinically, antibodies targeting PD-1/PD-L1 reinvigorate the "exhausted" T cells in TME and show remarkable objective response and durable remission with acceptable toxicity profile in large numbers of tumors such as melanoma, lymphoma, and mismatch-repair deficient tumors. Nevertheless, most patients are still refractory to anti-PD-1/PD-L1 therapy. Identifying the predictive biomarkers and design rational PD-1-based combination therapy become the priorities in cancer immunotherapy. PD-L1 expression, cytotoxic T lymphocytes infiltration, and tumor mutation burden (TMB) are generally considered as the most important factors affecting the effectiveness of PD-1/PD-L1 blockade. The revolution in cancer immunotherapy achieved by PD-1/PD-L1 blockade offers the paradigm for scientific translation from bench to bedside. The next decades will without doubt witness the renaissance of immunotherapy.

Synbindin deficiency inhibits colon carcinogenesis by attenuating Wnt cascade and balancing gut microbiome.

The molecular mechanisms that control the development of colorectal cancer (CRC) remain poorly defined. Here we show Synbindin promoted CRC oncogenesis by activating Wnt signaling and altering gut microbiome. Synbindin upregulation in human CRCs was associated with poor patient prognosis. Intestine-specific disruption of Synbindin balanced the disturbed gut microbiota and protected mice against tumor formation in the colitis-associated cancer (CAC) model. The protective role was compromised after gut microbiota depletion. In host, increased goblet cells and mucin2 expression, together with increased intestinal epithelial cells (IECs) apoptosis and decreased epithelial proliferation were observed. Further transcriptomic sequencing identified Wnt signaling a major regulatory node downstream of Synbindin. Combined molecular and cellular characterizations revealed that Synbindin confers Disheveled-3 (DVL3)-based signalosome assembly and acts as a modular scaffold for DVL3 and Axin2 complex, orchestrating the intensity of Wnt signaling. These findings identify a critical role of Synbindin in gut microbiome composition and Wnt signaling activation in colorectal carcinogenesis, and highlight Synbindin as an adaptor protein with multifaceted roles.

Upregulation of ASAP3 contributes to colorectal carcinogenesis and indicates poor survival outcome.

The function and clinical implication of ArfGAP with SH3 domain, ankyrin repeat, and PH domain 3 (ASAP3) in colorectal cancer (CRC) remains undefined. In the present study, we showed that the expression level of ASAP3 was dramatically increased in CRC and its upregulation was associated with American Joint Committee on Cancer stage (P < 0.001) and poor prognosis (P = 0.0022). The combination of stage and ASAP3 expression improved the prediction of survival in CRC patients. Suppression of ASAP3 inhibited cell proliferation by inducing G1 phase arrest without influencing apoptosis. ASAP3 promoted growth of colon tumors in mice with colitis, and accelerated cell invasion and migration in vitro. Increased ASAP3 was associated with activation of the nuclear factor-κB (NF-κB) canonical pathway in CRC. Upregulation of ASAP3 increased the phosphorylation and nuclear translocation of the p65 NF-κB subunit. Mechanistically, ASAP3 interacts with NF-κB essential modulator (NEMO) and could reduce the polyubiquitinylation of NEMO. Overall, ASAP3 might regulate NF-κB via binding to NEMO. ASAP3 acts as an oncogene in colonic cancer and could be a potential biomarker of colon carcinogenesis.

A20 reduces lipid storage and inflammation in hypertrophic adipocytes via p38 and Akt signaling.

Adipose tissue plays a vital role in the development of obesity and related disorders. Our previous study showed that A20, an ubiquitin-editing enzyme with anti-inflammation function, attenuated free fatty acids (FFAs)-induced lipid accumulation in nonalcoholic steatohepatitis. Here, we investigated A20 expression in adipose tissue of obese individuals and its effects on 3T3-L1 lipogenesis as well as the likely mechanisms underlying this process. By re-annotation of raw microarray data downloaded from Gene Expression Omnibus, we found that obese individuals showed significantly higher A20 mRNA levels in adipocytes. In vitro, A20 inhibited MCP-1 and IL-6 secretion in adipocytes. Forced expression of A20 resulted in decreased expression of key markers of lipogenesis and adipogenesis, such as sterol regulatory element binding protein 1c (SREBP-1c) and adipogenesis (aP2), leading to less lipids accumulation in differentiated 3T3-L1 cells. This process was concomitant with attenuated activation of p38 and Akt signaling. Our results suggest that A20 may have therapeutic potential for obesity and related diseases. The mechanisms involved the suppression of lipid storage and inflammation in adipocytes.

Tislelizumab plus cetuximab and irinotecan in refractory microsatellite stable and RAS wild-type metastatic colorectal cancer: a single-arm phase 2 study.

Immunotherapy confers little to no benefit in the treatment of microsatellite stable (MSS) metastatic colorectal cancer (mCRC). Mechanistic insights suggested that epidermal growth factor receptor (EGFR) antibody plus irinotecan might augment the tumor immune response in mCRC. Therefore, we conducted a proof-of-concept, single-arm, phase 2 study (ChiCTR identifier: ChiCTR2000035642) of a combination treatment regimen including tislelizumab (anti-PD-1), cetuximab (anti-EGFR) and irinotecan in 33 patients with MSS and RAS wild-type (WT) mCRC who were previously treated with ≥2 lines of therapy. The primary endpoint was met, with a confirmed objective response rate of 33%. As secondary endpoints, the disease control rate was 79%, and the median progression-free survival and overall survival were 7.3 and 17.4 months respectively. Among the 33 patients, 32 (97.0%) had treatment-related adverse events (AEs). Three (9.1%) reported grade ≥ 3 AEs, including rash (n = 1), neutropenia (n = 2). The post-hoc evaluation of dynamic circulating tumor DNA using next generation sequencing and the analysis of peripheral immune proteomics landscape using Olink revealed that lower variant allele frequency (VAF) at baseline, greater reduction in VAF on treatment, and a hot peripheral macroenvironment were associated with the treatment response independently. Our study showed the antitumor activity of tislelizumab, cetuximab, and irinotecan combination with a tolerable safety profile in previously treated MSS and RAS WT mCRC.

Cardiovascular Outcomes in the Patients With Colorectal Cancer: A Multi-Registry-Based Cohort Study of 197,699 Cases in the Real World.

Purpose: We aimed to investigate the mortality patterns and quantitatively assess the risks of cardiovascular death (CVD) in patients with colorectal cancer (CRC). We also established a competing-risk model to predict the probability of CVD for patients with CRC. Patients and Methods: Patients with CRC who diagnosed between 2007 and 2015 in the Surveillance, Epidemiology, and End Results (SEER) database were included in the present study. The cumulative incidence function (CIF) was used for CVD and other causes of death, and Gray's test was used to determine the subgroup difference in CIF. The Fine-Gray proportional subdistribution hazards model was used for identifying independent risk factors for CVD. A novel competing-risk model was established to evaluate the probability of CVD for patients with CRC. The performance of the nomogram was measured by concordance index (C-index), calibration curve, decision curve analysis (DCA), and risk stratification. Results: After a median follow-up of 37.00 months, 79,455 deaths occurred, of whom 56,185 (70.71%) succumbed to CRC and 23,270 (29.29%) patients died due to non-CRC, among which CVD accounted for 9,702 (41.69%), being the major cause of non-cancer deaths. The 1-, 3-, and 5-year cumulative rates for CVD were 12.20, 24.25, and 30.51%, respectively. In multivariate analysis, age, race, marital status, tumor size, tumor stage, advanced stage, surgery, and chemotherapy were independent risk factors of CVD among patients with CRC. The nomogram was well calibrated and had good discriminative ability, with a c-index of 0.719 (95% CI, 0.738-0.742) in the training cohort and 0.719 (95% CI, 0.622-0.668) in the validation cohort. DCA demonstrated that nomogram produced more benefit within wide ranges of threshold probabilities for 1-, 3-, and 5-year CVD, respectively. Conclusion: This study was the first to analyze the CIF and risk factors for CVD among CRC based on a competing-risk model. We have also built the first 1-, 3-, and 5-year competing nomogram for predicting CVD. This nomogram had excellent performance and could help clinicians to provide individualized management in clinical practice.

Heparanase modulates the prognosis and development of BRAF V600E-mutant colorectal cancer by regulating AKT/p27Kip1/Cyclin E2 pathway.

BRAF V600E-mutant colorectal cancer (CRC) is a rare subtype of colorectal cancer with poor prognosis. Compelling evidence indicates that the heparanase (HPSE) gene has multiple functions in cancer, however, its role in BRAF V600E-mutant CRC remains elusive. Differentially expressed genes between BRAF V600E-mutant and wild-type patients were explored by analyzing public data from The Cancer Genome Atlas and the Gene Expression Omnibus. Clinical samples of 172 patients with BRAF V600E-mutant CRC diagnosed at Zhongshan Hospital Fudan University were collected. Overall survival was analyzed using Kaplan-Meier curves and Cox regression models. Cell models and xenografts were utilized to investigate the effect of HPSE on tumor proliferation. HPSE was significantly highly expressed in the BRAF V600E-mutant group. High HPSE expression level was independently associated with inferior survival in the BRAF V600E-mutant cohort. HPSE knockdown impeded tumor proliferation of BRAF V600E-mutant CRC cells in vitro and in vivo. Mechanistically, HPSE silencing arrested cell cycle in G0/G1 phase by downregulating Cyclin E2 expression via the AKT/p27Kip1 pathway. These findings support a role for HPSE in promoting BRAF V600E-mutant CRC progression, which suggests it holds great promise as a prognostic biomarker and a potential therapeutic target for the aggressive CRC subtype.

Degradation efficiencies of 2,4,6-TCP by Fe0-based advanced oxidation processes (AOPs) with common peroxides.

Degradation of 2,4,6-trichlorophenol (2,4,6-TCP) by zero-valent iron (ZVI) activating three common peroxides (peroxymonosulfate (PMS), hydrogen peroxide (H2O2), and peroxydisulfate (PS)) was investigated. The effects of ZVI dosage, peroxides concentration, initial pH, and Cl- concentration were examined. The 2,4,6-TCP degradation efficiencies by Fe0/peroxides (PMS, H2O2, PS) were compared. Results showed that the order for degradation efficiency was H2O2≥PMS>PS. The degradation efficiency of 2,4,6-TCP in ZVI/peroxides systems were optimal at c(Ox) = 1 mmol•L-1; c(Fe0) = 0.1 g/L; initial pH = 3.2. Additionally, pH had a vital effect on 2,4,6-TCP degradation. At pH<3.2, ferrous play a vital role in all reaction, and accelerate the reaction rate rapidly. The existence of NaCl showed different results in the four systems. Chloride had little effect on 2,4,6-TCP degradation when chloride concentration at 5 mM, whereas the presence of 300 mM chloride significantly accelerated the degradation of 2,4,6-TCP from 72.7% to 95.2% in ZVI-PMS system. Notably, the other three systems showed opposite results. In contrast, the AOX (Absorbable Organic Halogen) values were highest in ZVI-PMS-Cl- system, due to the formation of lots of refractory chlorinated phenols as identified by GC-MS. These findings are good for choosing the most suitable technology for chlorophenol wastewater treatment.

TRAPPC4 regulates the intracellular trafficking of PD-L1 and antitumor immunity.

Tumor cells evade T cell-mediated immunosurveillance via the interaction between programmed death-1 (PD-1) ligand 1 (PD-L1) on tumor cells and PD-1 on T cells. Strategies disrupting PD-1/PD-L1 have shown clinical benefits in various cancers. However, the limited response rate prompts us to investigate the molecular regulation of PD-L1. Here, we identify trafficking protein particle complex subunit 4 (TRAPPC4), a major player in vesicular trafficking, as a crucial PD-L1 regulator. TRAPPC4 interacts with PD-L1 in recycling endosomes, acting as a scaffold between PD-L1 and RAB11, and promoting RAB11-mediated recycling of PD-L1, thus replenishing its distribution on the tumor cell surface. TRAPPC4 depletion leads to a significant reduction of PD-L1 expression in vivo and in vitro. This reduction in PD-L1 facilitates T cell-mediated cytotoxicity. Overexpression of Trappc4 sensitizes tumor cells to checkpoint therapy in murine tumor models, suggesting TRAPPC4 as a therapeutic target to enhance anti-tumor immunity.

Treatment strategy for myocarditis in patients using immune checkpoint inhibitors or combined anti-vascular endothelial growth factor therapy by clinical severity.

BACKGROUND: Combination of immune checkpoint inhibitor (ICI) and anti-vascular endothelial growth factor (VEGF) therapy has increasingly become a promising strategy in various tumors. However, the combination might be associated with increased cardiotoxicities. Myocarditis is a potentially fatal complication in cancer patients treated with ICI. Currently, there are no clear guidelines for the management of this disease nor data characterizing the clinical course of myocarditis patients due to the combination of ICI and anti-VEGF therapy. PATIENTS AND METHODS: This study included all patients consecutively admitted to Shanghai Zhongshan Hospital, Fudan University for the diagnosis of ICI-associated myocarditis according to Bonaca's criteria. The clinical presentation and outcome of myocarditis patients were collected receiving ICI and anti-VEGF combined therapy. RESULTS: A total of 48 patients were included (23 received combined treatment of ICI and anti-VEGF while 25 received ICI only). No differences in baseline characteristics, clinical course, and outcomes were observed among patients receiving ICI with or without anti-VEGF treatment. The patients were subdivided into 3 groups including 8 fulminant cases, 25 clinically significant cases, and 15 subclinical cases based on clinical severity. The fulminant group was associated with a higher rate of cardiovascular deaths (CVDs) compared with clinically significant and subclinical groups (87.5% versus 4.0% versus 0.0%, p < 0.01). When stratified by the dose of corticosteroids used, cases with high-dose usage were more likely to have a CVD when compared to low dose or no use and intermediate dose groups (0.0% versus 4.0% versus 57.1%, p < 0.01). CONCLUSIONS: No significant differences between myocarditis patients receiving ICI or combined anti-VEGF therapy in terms of clinical presentation and outcome were observed. Treatment strategy for myocarditis in patients using ICI or combined anti-VEGF therapy should be based on clinical severity. Specifically, immunosuppressive therapy besides high-dose corticosteroids is needed for fulminant cases.

Nivolumab-associated DRESS in a genetic susceptible individual.

The use of immune checkpoint inhibitors (ICIs) is rising exponentially in numerous cancers, but immune-related adverse events can occur. We report a rare case of high-grade drug reaction with eosinophilia and systemic symptoms (DRESS) syndrome developed stepwise in a patient with gastric cancer after nivolumab treatment. Subclinical myocarditis was sensitively detected by cardiovascular magnetic resonance 3 weeks after initiating nivolumab. Eruption, eosinophilia, and interstitial pneumonitis occurred 1 week later. Corticosteroids were started and his condition improved. Four months later, when he was still on steroids tapering off, acute kidney injury and sequential herpes zoster virus activation developed. Severe acute tubulointerstitial nephritis (ATN) with an intense infiltration of lymphocytes was observed on renal biopsy. In blood, a substantial shift to Th2 response, an increase of Th17 cells, and strikingly enriched granzyme B+ and perforin+ CD8+ T cells were detected at ATN onset. Serum interleukin (IL)-5, IL-17, interferon gamma, and IL-6 levels were consistently elevated. Further molecular profiling identified a DRESS risk allele human leukocyte antigen (HLA)-A*31:01 in this patient. His ATN responded favorably to a high dose of corticosteroids. In parallel, complete antitumor response was observed during the clinical course of DRESS. This is the first ever case report of nivolumab-associated DRESS syndrome with exploration of the mechanisms from the histopathological, cellular and molecular aspects. Nivolumab-induced DRESS may result from type IV hypersensitivity-related 'off-target effect' and PD-1 block-mediated 'on-target effect'. HLA risk alleles may constitute the genetic susceptible basis. HLA typing assay has the potential to screen susceptible individuals to avoid ICI-induced DRESS.

Association between non-alcoholic fatty liver disease and silent carotid plaque in Chinese aged population: a cross-sectional study.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is a risk factor for carotid plaque in the general population; however, whether NAFLD is associated with carotid plaque in aged population remains unknown. This cross-sectional study was conducted to evaluate the association between NAFLD and carotid plaque in a Chinese aged population. METHODS: A total number of 12,990 Chinese aged adults (7,685 men and 5,305 women) were included. NAFLD was diagnosed based on the recommendation of Asia-Pacific Working Party on NAFLD and Chinese Association for the Study of Liver Disease and excessive alcohol consumption (weekly alcohol consumption ≤210 g in men and ≤140 g in women) was excluded. Carotid plaque was confirmed by ultrasonography. Potential confounders and biochemical findings were collected at baseline. Logistic regression model was employed to evaluate the association between NAFLD and carotid plaque. RESULTS: The prevalence of carotid plaque was significantly higher in aged participants with NAFLD than in those without NAFLD (22.4% vs. 16.3%, P<0.001). NAFLD is associated with carotid plaque [odd ratio (OR) =1.89, 95% confidence interval (CI): 1.59-2.24], after adjusting for age, gender, BMI, liver and kidney function, glucose level, lipid profiles, and white blood cell (WBC) count. The association between NAFLD and carotid plaque was attenuated when participants with elevated ALT (≥75 IU/L), a history of cardiovascular diseases (CVDs) and obesity were censored although the significant association remained. CONCLUSIONS: NAFLD is associated with carotid plaque in Chinese aged population.

On peroxymonosulfate-based treatment of saline wastewater: when phosphate and chloride co-exist.

Both chloride and phosphate are common inorganic anions in industrial wastewater, however, their effects on peroxymonosulfate (PMS)-based oxidation systems are largely unknown. The present results show that addition of chloride (>1 mM) apparently enhanced the degradation of Acid Orange 7 (AO7) independent of the presence of phosphate (PBS) buffer. Both PBS and chloride favored the degradation of AO7, while PBS played a more important role when they co-existed. The degradation efficiency of AO7 was enhanced by increasing the concentration of PBS and chloride. A maximum of absorbable organic halides (AOX) accumulation was observed; indicating some chlorinated byproducts could be initially generated and further oxidized by increasing the reaction time. It is demonstrated that the PBS/PMS system, with a lower AOX formation at the same chloride concentration, is superior to the Co/PMS system, a typical sulfate radical-based system. The active chlorine species (HClO/Cl2) were found to be the dominant oxidants in the presence of higher chloride concentration (>50 mM) under neutral conditions. The findings of this work may promote the further application of PMS-based oxidation processes in saline effluents treatment.

Peroxymonosulfate/base process in saline wastewater treatment: The fight between alkalinity and chloride ions.

Both Cl- and base can affect PMS activation to produce reactive chlorine or oxygen species, but the overall effects of chloride on this emerging PMS/base technology in saline wastewater treatment are unknown. Here effectiveness of PMS/base, PMS/Cl- and PMS/base/Cl- is compared with a gradient concentration of chloride and alkalinity, by probing the degradation of methylene blue (MB). Both PMS/base and PMS/Cl- systems can rapidly degrade MB due to the generation of singlet oxygen and reactive chlorine, respectively. Interestingly, dye degradation and adsorbable organic halides (AOX) formation are inhibited in the PMS/base/Cl- system as high concentrations of Cl- and base co-exist. Reaction of PMS with chloride diminishes the effective concentration of PMS by base activation, whereas in return high alkalinity decreases the oxidation capacity of reactive species. Therefore, this finding may have significant technical implications for evaluating the applicability of the emerging PMS/base technology and optimizing the conditions for AOX abatement in PMS-based processes.

Systematic evaluation of supervised classifiers for fecal microbiota-based prediction of colorectal cancer.

Predicting colorectal cancer (CRC) based on fecal microbiota presents a promising method for non-invasive screening of CRC, but the optimization of classification models remains an unaddressed question. The purpose of this study was to systematically evaluate the effectiveness of different supervised machine-learning models in predicting CRC in two independent eastern and western populations. The structures of intestinal microflora in feces in Chinese population (N = 141) were determined by 454 FLX pyrosequencing, and different supervised classifiers were employed to predict CRC based on fecal microbiota operational taxonomic unit (OTUs). As a result, Bayes Net and Random Forest displayed higher accuracies than other algorithms in both populations, although Bayes Net was found with a lower false negative rate than that of Random Forest. Gut microbiota-based prediction was more accurate than the standard fecal occult blood test (FOBT), and the combination of both approaches further improved the prediction accuracy. Moreover, when unclassified OTUs were used as input, the BayesDMNB text algorithm achieved higher accuracy in the Chinese population (AUC=0.994). Taken together, our results suggest that Bayes Net classification model combined with unclassified OTUs may present an accurate method for predicting CRC based on the compositions of gut microbiota.

On the kinetics of organic pollutant degradation with Co2+/peroxymonosulfate process: When ammonium meets chloride.

A large amount of chloride and ammonium ions were produced and released from industrial processes with non-biodegradable organic pollutants to affect efficiencies of advanced oxidation processes (AOPs). Here, the influences of chloride and ammonium ions on Co/peroxymonosulfate (Co/PMS) reaction system, a widely used AOPs to produce sulfate radicals, were investigated by examining the degradation efficiency of an azo dye (Acid Orange 7, AO7). The experimental results showed that a significant decrease in the degradation rate of AO7 was observed in the presence of NH4+, while a dual effect of chloride on AO7 bleaching appeared. The presence of NH4Cl was unfavorable for AO7 degradation at low concentration (<20 mM), whereas further addition of NH4Cl (>20 mM) apparently accelerated AO7 discoloration rate. The apparent effects of the two co-existing inorganic ions were determined by roles of the dominating ions at varied molar ratio of [NH4+]/[Cl-]. The present study may have technical implications for the treatment of industrial wastewater containing diverse ions in practice.

A20 Attenuates Liver Fibrosis in NAFLD and Inhibits Inflammation Responses.

We previously reported A20 was able to inhibit lipid accumulation in nonalcoholic steatohepatitis. We want to investigate whether A20 influences liver fibrosis in this study. Liver tissues from patients with hepatic fibrosis (n = 9) and healthy individuals (n = 7) were studied for A20 protein level by immunohistochemistry. A20 messenger RNA (mRNA) and protein level were also analyzed in two murine hepatic fibrosis models: methionine- and choline-deficient (MCD) diet and extrahepatic bile duct ligation (BDL) operation by real-time PCR and western blot. In vitro, the LX-2 human hepatic stellate cell line was treated by LPS at 0, 0.001, 0.01, 0.1, and 1 µg/mL for 6 h or at the concentration of 0.1 µg/mL for 0, 6, 12, and 24 h, then A20 expression levels were detected by western blot and PCR. The mRNA level of α-SMA, collagen I, collagen III, TGF-ß, IL-6, MCP-1, and TLR4 was also examined by PCR. We then overexpressed A20 in LX-2 cells using adenovirus technique. Levels of α-SMA, collagen I, collagen III, TGF-ß, IL-6, MCP-1, and TLR4 were examined in A20-overexpression LX-2 cells. Patients with hepatic fibrosis showed significantly higher A20 protein level compared with healthy controls. A20 mRNA and protein levels were also increased in livers from MCD feeding or BDL operation mice in comparison to normal controls. In LX-2 cells, LPS induced A20 protein in a concentration-dependent manner. The mRNA levels of α-SMA, collagen I, collagen III, TGF-ß, IL-6, MCP-1, and TLR4 were increased after LPS treatment. Overexpression of A20 in LX-2 cells inhibited α-SMA deposition and collagen I, collagen III secretion. TGF-ß, IL-6, MCP-1, and TLR4 mRNA levels were also reduced in A20-overexpression LX-2 cells in response to LPS stimulation. A20 overexpression inhibits hepatic stellate cell activation, which could be the mechanism for high A20 expression protected livers from fibrosis. Enhancement of A20 expression seems to be rational therapeutic strategies for liver fibrosis.

ASAP3 regulates microvilli structure in parietal cells and presents intervention target for gastric acidity.

Gastric acidity-associated disorders such as peptic ulcer and reflux diseases are widespread, and the reported resistance and side effects of currently used medicines suggest an urgent requirement for alternative therapeutic approaches. Here we demonstrate a critical role of ASAP3 in regulating the microvilli structure of parietal cells in vivo, and reveal the feasibility of controlling gastric acidity by targeting ASAP3. Conditional knockout of ASAP3 in mice caused elongation and stacking of microvilli in parietal cells, and substantially decreased gastric acid secretion. These were associated with active assembly of F-actin caused by a higher level of GTP-bound Arf6 GTPase. Consistently, a small molecular compound QS11 inhibited ASAP3 function and significantly reduced gastric acidity in vivo. Of note, the expression of ASAP3 was positively correlated with gastric acid secretion in 90 human cases, and high expression of ASAP3 was associated with reflux disease and peptic ulcer. These results reveal for the first time that ASAP3 regulates the microvilli structures in parietal cells. Our data also suggest ASAP3 as a feasible and drugable therapeutic target for gastric acidity-associated diseases.

Long noncoding RNA expression profiles in gut tissues constitute molecular signatures that reflect the types of microbes.

The gut microbiota is commonly referred to as a hidden organ due to its pivotal effects on host physiology, metabolism, nutrition and immunity. The gut microbes may be shaped by environmental and host genetic factors, and previous studies have focused on the roles of protein-coding genes. Here we show a link between long non-coding RNA (lncRNA) expression and gut microbes. By repurposing exon microarrays and comparing the lncRNA expression profiles between germ-free, conventional and different gnotobiotic mice, we revealed subgroups of lncRNAs that were specifically enriched in each condition. A nearest shrunken centroid methodology was applied to obtain lncRNA-based signatures to identify mice in different conditions. The lncRNA-based prediction model successfully identified different gnotobiotic mice from conventional and germ-free mice, and also discriminated mice harboring transplanted microbes from fecal samples of mice or zebra fishes. To achieve optimal prediction accuracy, fewer lncRNAs were required in the prediction model than protein-coding genes. Taken together, our study demonstrated the effecacy of lncRNA expression profiles in discriminating the types of microbes in the gut. These results also provide a resource of gut microbe-associated lncRNAs for the development of lncRNA biomarkers and the identification of functional lncRNAs in host-microbes interactions.