Study of the mechanism by which Xiaoyan decoction combined with E7449 regulates tumorigenesis in lung adenocarcinoma.

TNKS is a new target for the treatment of lung adenocarcinoma, the synergistic effects of the TCM compound Xiaoyan decoction and the TNKS inhibitor E7449 in the intervention on TNKS were investigated, and the possible underlying mechanisms involved were clarified. Immunohistochemistry was used to analyse TNKS expression in tumour tissues. The impact of targeting TNKS on cell growth, invasion, apoptosis, key genes and signalling pathways was investigated in tumour cells by Western blotting, rescue experiments, colony formation assays, flow cytometry and label-free experiments. Tumour xenografts with A549 cells were then transplanted for in vivo study. We found that TNKS high expression was closely related to the advanced tumour stage and tumour size in lung adenocarcinom. After TNKS was knocked down in vitro, the growth, proliferation, migration and invasion were markedly reduced in A549 and H1975 cells. We subsequently applied the Xiaoyan decoction and TNKS inhibitors to intervene in lung adenocarcinoma. Xiaoyan decoction and E7449 suppressed TNKS expression and inhibited adenocarcinoma cell proliferation, migration, invasion and apoptosis in vitro. Proteomic analysis revealed that E7449 treatment may be most closely associated with the classic Wnt/ß-catenin pathway, whereas Xiaoyan decoction treatment may be related to the WNT/PLAN pathway. Xenograft studies confirmed that E7449 or Xiaoyan decoction inhibited lung tumour growth in vivo and attenuated the Wnt signalling pathway in adenocarcinoma. These findings suggest that TNKS is a novel therapeutic target. TCM preparations and small molecule inhibitors are expected to constitute an effective combination strategy.

Roles of post-translational modifications of UHRF1 in cancer.

UHRF1 as a member of RING-finger type E3 ubiquitin ligases family, is an epigenetic regulator with five structural domains. It has been involved in the regulation of a series of biological functions, such as DNA replication, DNA methylation, and DNA damage repair. Additionally, aberrant overexpression of UHRF1 has been observed in over ten cancer types, indicating that UHRF1 is a typical oncogene. The overexpression of UHRF1 repressed the transcription of such tumor-suppressor genes as CDKN2A, BRCA1, and CDH1 through DNMT1-mediated DNA methylation. In addition to the upstream transcription factors regulating gene transcription, post-translational modifications (PTMs) also contribute to abnormal overexpression of UHRF1 in cancerous tissues. The types of PTM include phosphorylation, acetylation, methylationand ubiquitination, which regulate protein stability, histone methyltransferase activity, intracellular localization and the interaction with binding partners. Recently, several novel PTM types of UHRF1 have been reported, but the detailed mechanisms remain unclear. This comprehensive review summarized the types of UHRF1 PTMs, as well as their biological functions. A deep understanding of these crucial mechanisms of UHRF1 is pivotal for the development of novel UHRF1-targeted anti-cancer therapeutic strategies in the future.

Ethanol extract of Andrographis paniculata alleviates aluminum-induced neurotoxicity and cognitive impairment through regulating the p62-keap1-Nrf2 pathway.

BACKGROUND: Alzheimer's disease (AD) is the most prevalent neurodegenerative and remains incurable. Aluminum is a potent neurotoxin associated with AD. The main pathological features of AD are extracellular amyloid-ß protein deposition and intracellular hyperphosphorylated Tau protein. A body of evidence suggest that oxidative stress and autophagy are involved in the pathogenesis of AD. Andrographis paniculata (AP) is a native plant with anti-inflammatory, anti-oxidative stress, and regulation of autophagy properties. AP significantly alleviated cognitive impairments, reduced Aß deposition and has neuroprotective effect. However, its effects on aluminum-induced AD model have not been studied much. In this study, we investigated whether AP protect against aluminum-induced neurotoxicity through regulation of p62-Kelch-like ECH-associated protein 1(Keap1)-Nuclear factor E2 related factor 2 (Nrf2) pathway and activation autophagy in vivo and in vitro. METHODS: UPLC-ESI-qTOF-MS/MS was used to identify the chemical constituents of AP ethanol extract. The mice with cognitive deficit were established by injecting aluminum chloride and D-galactose, and treated with either AP extract (200, 400, or 600 mg/kg/d) or andrographolide (2 mg/kg/2d).The spatial memory ability was detected by Morris water maze, HE staining were used to detect in brain tissue,Oxidative stress indexs and SOD activity in both serum and brain tissue were detected by kit.The expression of p62-Nrf2 pathway proteins were measured via western blotting. Furthermore, the neurotoxicity model was induced by aluminum maltolate (700 µM) in PC12 cells. Following AP and andrographolide treatment, the cell viability was detected. The relevant mRNA and protein expressions were detected in cells transfected with the p62 siRNA. RESULTS: The main active components of AP included andrographolide, neoandrographolide and deoxyandrographolide as identified. AP and andrographolide significantly improved the spatial memory ability of mice, attenuated pathological changes of hippocampal cells, reduced the level of malondialdehyde, and increased superoxide dismutase activity in serum or brain tissue as compared to model control. In addition, the Nrf2, p62 and LC3B-II proteins expression were increased, and p-Tau and Keap1 proteins were decreased in the hippocampus after AP and andrographolide treatment.Furthermore, AP increased aluminum maltolate-induced cell viability in PC12 cells. Silencing p62 could reverse the upregulation expression of Nrf2 and downregulation of Keap1 and Tau proteins induced by AP in aluminum maltolate-treated cells. CONCLUSIONS: AP had neuroprotective effects against aluminum -induced cognitive dysfunction or cytotoxicity, which was involved in the activation of the p62-keap1-Nrf2 pathway and may develop as therapeutic drugs for the treatment of AD. However, this study has certain limitations, further optimize the protocol or model and study the molecular mechanism of AP improving AD.

Nomogram for predicting the unfavourable outcomes of percutaneous endoscopic transforaminal discectomy for lumbar disc herniation: a retrospective study.

Objective: To investigate and integrate multiple independent risk factors to establish a nomogram for predicting the unfavourable outcomes of percutaneous endoscopic transforaminal discectomy (PETD) for lumbar disc herniation (LDH). Methods: From January 2018 to December 2019, a total of 425 patients with LDH undergoing PETD were included in this retrospective study. All patients were divided into the development and validation cohort at a ratio of 4:1. Univariate and multivariate logistic regression analyses were used to investigate the independent risk factors associated with the clinical outcomes of PETD for LDH in the development cohort, and a prediction model (nomogram) was established to predict the unfavourable outcomes of PETD for LDH. In the validation cohort, the nomogram was validated by the concordance index (C-index), calibration curve, and decision curve analysis (DCA). Results: 29 of 340 patients showed unfavourable outcomes in the development cohort, and 7 of 85 patients showed unfavourable outcomes in the validation cohort. Body mass index (BMI), course of disease (COD), protrusion calcification (PC), and preoperative lumbar epidural steroid injection (LI) were independent risk factors associated with the unfavourable outcomes of PETD for LDH and were identified as predictors for the nomogram. The nomogram was validated by the validation cohort and showed high consistency (C-index = 0.674), good calibration and high clinical value. Conclusions: The nomogram based on patients' preoperative clinical characteristics, including BMI, COD, LI and PC, can be used to accurately predict the unfavourable outcomes of PETD for LDH.

PLCB1 Enhances Cell Migration and Invasion in Gastric Cancer Via Regulating Actin Cytoskeletal Remodeling and Epithelial-Mesenchymal Transition.

Phospholipase C Beta 1 (PLCB1) regulates the abundance of PI(4,5)P2 in the plasma membrane and is implicated in various kinds of cancers. This study aimed to investigate the role and underlying mechanisms of PLCB1 in gastric cancer. Herein, it was found that PLCB1 mRNA and protein were highly expressed in gastric cancer, and high levels of PLCB1 were correlated with poor outcomes of patients with gastric cancer via the GEPIA database. Moreover, our results revealed that PLCB1 depletion inhibited gastric cancer cell proliferation, migration, and invasion. Meanwhile, PLCB1 overexpression resulted in an inverse result. Furthermore, PLCB1 mediated actin cytoskeleton rearrangement and activated the RhoA/LIMK/Cofilin pathway. Besides, PLCB1 promoted the Epithelial-Mesenchymal transition process via activating ATK signaling. In conclusion, PLCB1 promoted gastric cancer cell migratory and invasive abilities via regulating actin cytoskeleton rearrangement and Epithelial-Mesenchymal transition process. These findings imply that targeting PLCB1 may be a potential strategy to improve the prognosis of gastric cancer patients.

Andrographolide exerts a neuroprotective effect by regulating the LRP1-mediated PPARγ/NF-κB pathway.

Low-density lipoprotein receptor-associated protein 1 (LRP1) is widely expressed in neurons, microglia and astrocytes. Studies have revealed that the suppression of LRP1 expression in the brain significantly exacerbates Alzheimer's disease (AD)-related neuropathology. Andrographolide (Andro) has been demonstrated to possess neuroprotective properties, although its underlying mechanisms remain largely unknown. This study aims to investigate whether Andro can inhibit neuroinflammation in AD by modulating the LRP1-mediated PPARγ/NF-κB pathway. In Aß-induced BV-2 cells, Andro was found to increase cell viability and enhance the expression of LRP1, while decreasing the expression of p-NF-κB (p65) and NF-κB(p65), as well as IL-1ß, IL-6 and TNF-α levels. In addition, when Aß was cotreatment with Andro to BV2 cells with either LRP1 or PPARγ knockdown, increased mRNA and protein expression of p-NF-κB(p65) and NF-κB(p65), NF-κB DNA binding activity as well as IL-1ß, IL-6 and TNF-α levels were observed. These findings suggested that Andro could attenuate Aß induced cytotoxicity by reducing neuroinflammation which may be partly attributed to its effects on this LRP1 mediated PPARγ/NF-κB pathway.

Reduced Serological Response to COVID-19 Booster Vaccine is Associated with Reduced B Cell Memory in Patients With Inflammatory Bowel Disease; VARIATION [VAriability in Response in IBD AgainsT SARS-COV-2 ImmunisatiON].

BACKGROUND AND AIMS: Patients with inflammatory bowel disease [IBD] have an attenuated response to initial COVID-19 vaccination. We sought to characterize the impact of IBD and its treatment on responses after the third vaccine against SARS-CoV-2. METHODS: This was a prospective multicentre observational study of patients with IBD [n = 202] and healthy controls [HC, n = 92]. Serological response to vaccination was assessed by quantification of anti-spike protein [SP] immunoglobulin [Ig]G levels [anti-SPIgG] and in vitro neutralization of binding to angiotensin-converting enzyme 2 [ACE2]. Peripheral blood B-cell phenotype populations were assessed by flow cytometry. SARS-CoV-2 antigen-specific B-cell responses were assessed in ex vivo culture. RESULTS: Median anti-SP IgG post-third vaccination in our IBD cohort was significantly lower than HCs [7862 vs 19 622 AU/mL, p < 0.001] as was ACE2 binding inhibition [p < 0.001]. IBD patients previously infected with COVID-19 [30%] had similar quantitative antibody response as HCs previously infected with COVID-19 [p = 0.12]. Lowest anti-SP IgG titres and neutralization were seen in IBD patients on anti-tumour necrosis factor [anti-TNF] agents, without prior COVID-19 infection, but all IBD patients show an attenuated vaccine response compared to HCs. Patients with IBD have reduced memory B-cell populations and attenuated B-cell responses to SARS-CoV-2 antigens if not previously infected with COVID-19 [p = 0.01]. Higher anti-TNF drug levels and zinc levels <65 ng/ml were associated with significantly lower serological responses. CONCLUSIONS: Patients with IBD have an attenuated response to three doses of SARS-CoV-2 vaccine. Physicians should consider patients with higher anti-TNF drug levels and/or zinc deficiency as potentially at higher risk of attenuated response to vaccination.

Interleukin-6 promotes skeletal muscle catabolism by activating tryptophan-indoleamine 2,3-dioxygenase 1-kynurenine pathway during intra-abdominal sepsis.

BACKGROUND: Inflammatory cytokine interleukin-6 (IL-6) plays a pivotal role in skeletal muscle degradation after intra-abdominal sepsis (IAS), with mechanism remained to be elucidated. Indoleamine 2,3-dioxygenase 1 (IDO-1), a key enzyme in converting tryptophan into kynurenine, could be activated by IL-6, and kynurenine has been shown to be involved in muscle degradation. We hypothesized that IL-6 could promote muscle degradation via tryptophan-IDO-1-kynurenine pathway in IAS patients. METHODS: Serum and rectus abdominis (RA) were obtained from IAS or non-IAS patients. Mouse model of IAS-induced muscle wasting was generated by caecal ligation and puncture (CLP) and lipopolysaccharide (LPS) injection. IL-6 signalling was blocked by anti-mouse IL-6 antibody (IL-6-AB), and the IDO-1 pathway was blocked by navoximod. To elucidate the role of kynurenine in muscle mass and physiology, kynurenine was administered to IAS mice treated with IL-6-AB. RESULTS: Compared to non-IAS patients, kynurenine levels in serum (+2.30-fold vs. non-IAS, P < 0.001) and RA (+3.11-fold vs. non-IAS, P < 0.001) were elevated, whereas tryptophan levels in serum (-53.65% vs. non-IAS, P < 0.01) and RA (-61.39% vs. non-IAS, P < 0.01) were decreased. Serum IL-6 level of the IAS group was significantly higher compared to non-IAS patients (+5.82-fold vs. non-IAS, P = 0.01), and muscle cross-sectional area (MCSA) was markedly reduced compared to non-IAS patients (-27.73% vs. non-IAS, P < 0.01). In animal experiments, IDO-1 expression was up-regulated in the small intestine, colon and blood for CLP or LPS-treated mice, and there was correlation (R2  = 0.66, P < 0.01) between serum and muscle kynurenine concentrations. Navoximod significantly mitigated IAS-induced skeletal muscle loss according to MCSA analysis (+22.94% vs. CLP, P < 0.05; +23.71% vs. LPS, P < 0.01) and increased the phosphorylated AKT (+2.15-fold vs. CLP, P < 0.01; +3.44-fold vs. LPS, P < 0.01) and myosin heavy chain (+3.64-fold vs. CLP, P < 0.01; +2.13-fold vs. LPS, P < 0.01) protein expression in myocytes. In the presence of anti-IL-6 antibody, a significantly decreased IDO-1 expression was observed in the small intestine, colon and blood in CLP or LPS mice (all P < 0.01), whereas the decrease of MCSA was alleviated (+37.43% vs. CLP + IgG, P < 0.001; +30.72% vs. LPS + IgG, P < 0.001). In contrast, additional supplementation of kynurenine decreased the MCSA in septic mice treated with IL-6-AB (both P < 0.01). CONCLUSIONS: This study provided novel insights into the tryptophan-IDO-1-kynurenine-dependent mechanisms that underlie inflammatory cytokine-induced skeletal muscle catabolism during intra-abdominal sepsis.

Anti-mesothelin CAR-T immunotherapy in patients with ovarian cancer.

Recently, chimeric antigen receptor T cell (CAR-T) therapy has received increasing attention as an adoptive cellular immunotherapy that targets tumors. However, numerous challenges remain for the effective use of CAR-T to treat solid tumors, including ovarian cancer, which is an aggressive and metastatic cancer with a poor therapeutic response. We screened for an effective anti-MSLN single-chain Fv antibody with comparable binding activity and non-off-target properties using human phage display library. A second-generation of anti-MSLN CAR was designed and generated. We demonstrated the efficacy of our anti-MSLN CAR-T cells for ovarian cancer treatment in an in vitro experiment to kill ovarian tumor cell lines. The anti-MSLN CAR-T cells impeded MSLN-positive tumor growth concomitant with a significant increase in cytokine levels compared with the control. Then, we demonstrated the efficacy of anti-MSLN CAR-T cells in an in vivo experiment against ovarian cancer cell-derived xenografts. Furthermore, we herein report three cases with ovarian cancer who were treated with autologous anti-MSLN CAR-T cells and evaluate the safety and effectiveness of adoptive cell therapy. In this investigator-initiated clinical trials, no patients experienced cytokine release syndrome or neurological symptoms over 2 grads. Disease stabilized in two patients, with progression-free survival times of 5.8 and 4.6 months. Transient CAR expression was detected in patient blood after infusion each time. The tumor partially subsided, and the patient's condition was relieved. In conclusion, this work proves the efficacy of the anti-MSLN CAR-T treatment strategy in ovarian cancer and provides preliminary data for the development of further clinical trials.

Medicinal fungus Phellinus igniarius alleviates gout in vitro by modulating TLR4/NF-kB/NLRP3 signaling.

Background: Phellinus igniarius (P. igniarius) is a valuable medicinal and edible fungus with various biological activities such as anti-inflammation, antioxidation, and immune regulation. In this study, we explored the effects of P. igniarius on a gout model in vitro. Methods: The DPPH, ABTS, and FRAP methods were combined to determine and compare the antioxidant activities of wild P. igniarius total polyphenols (WPP) and cultivated P. igniarius total polyphenols (CPP) in vitro. Spectrophotometry was used to compare the inhibitory effect of WPP and CPP on xanthine oxidase (XO) activity to evaluate anti-hyperuricemia activity in vitro. HUVECs were stimulated with monosodium urate (MSU) crystals for 24 h to establish an acute gouty inflammation model in vitro. The protective effects were compared by measuring cell viability; the contents of ICAM-1, IL-1ß, IL-6 and VCAM-1; the protein expressions of TLR4 and NLRP3; reactive oxygen species production; and the nuclear translocation of NF-κB p65. UHPLC-QE-MS technology was used to explore the potential metabolic mechanism of P. igniarius against gout. Results: WPP and CPP had strong antioxidant capacity, and the antioxidant capacity of CPP was similar to that of WPP. In a comparative experiment of xanthine oxidase activity inhibition by WPP and CPP, the IC50 values were 88.19 µg/ml and 108.0 µg/ml, respectively. At a dose of 40 µg/ml, WPP and CPP significantly improved the decrease in cell viability induced by monosodium urate (150 µg/ml) and inhibited the increase in inflammatory factors such as ICAM-1, IL-1ß, IL-6, and VCAM-1. The increase in TLR4 and NLRP3 protein expression induced by MSU crystals in HUVECs was also significantly inhibited by total polyphenols from wild and cultivated P. igniarius. In addition, both significantly improved MSU-induced ROS overproduction and NF-κB p65 nuclear translocation. WPP and CPP may primarily be involved in phenylalanine metabolism and lysophosphatidylcholine metabolism in their role in the treatment of gout. Conclusion: CPP and WPP both showed good antioxidant activity and xanthine oxidase inhibitory activity and had good therapeutic effects on the gout model in vitro. Furthermore, this study indicated that cultivated P. igniarius had a protective effect similar to that of wild P. igniarius, which would be expected to improve the shortage of wild P. igniarius and promote the development of the cultivated P. igniarius industry and product development.

Role of Wnt/ß-catenin signaling pathway in ameloblast differentiation in relevance to dental fluorosis.

Excess consumption of fluoride during the development of tooth enamel will cause dental fluorosis, but the exact molecular mechanisms remain to be elucidated. Circadian rhythm is implicated in many physiological processes and various diseases. There is increasing evidence indicates that ameloblast differentiation is under the control of clock genes. However, it has not been reported whether fluoride regulates ameloblast differentiation through clock genes and the downstream PPARγ. To explore the effect of fluoride on ameloblast differentiation and the underlying regulatory mechanism, we used both rat dental fluorosis model and an ameloblast cell line LS8 to conduct a series of experiments. Our results showed that fluoride significantly reduced the expression of PCNA, RUNX2 and MMP9 in rat ameloblasts and LS8 cells (P < 0.05). Fluoride increased nuclear translocation of ß-catenin in vivo and in vitro, and 0.1 µg/ml Dkk1 pretreatment ameliorated the decreased expression of CXXC5, RUNX2 and MMP9 induced by fluoride. Furthermore, we found fluoride significantly inhibited the expression of Clock, Bmal1, Per2 and PPARγ in rat mandibular ameloblasts and LS8 cells by immunostaining, qPCR and Western blot (P < 0.05). Flow cytometry analysis showed that fluoride promoted ROS generation. Remarkably, 50 µM resveratrol significantly ameliorated the inhibitory effect of fluoride on ameloblast differentiation markers, clock genes and PPARγ, and inhibited the Wnt/ß-catenin signaling (P < 0.05). Taken together, these findings suggested that excessive fluoride promoted ROS generation, leading to the inhibition of clock genes, which resulted in reduced PPARγ and activated Wnt/ß-catenin signaling pathway, thus inhibiting ameloblast differentiation and matrix degradation. This study provides a better understanding of the molecular mechanism of enamel defects in dental fluorosis.

Klebsiella pneumoniae hijacks the Toll-IL-1R protein SARM1 in a type I IFN-dependent manner to antagonize host immunity.

Many bacterial pathogens antagonize host defense responses by translocating effector proteins into cells. It remains an open question how those pathogens not encoding effectors counteract anti-bacterial immunity. Here, we show that Klebsiella pneumoniae exploits the evolutionary conserved innate protein SARM1 to regulate negatively MyD88- and TRIF-governed inflammation, and the activation of the MAP kinases ERK and JNK. SARM1 is required for Klebsiella induction of interleukin-10 (IL-10) by fine-tuning the p38-type I interferon (IFN) axis. SARM1 inhibits the activation of Klebsiella-induced absent in melanoma 2 inflammasome to limit IL-1ß production, suppressing further inflammation. Klebsiella exploits type I IFNs to induce SARM1 in a capsule and lipopolysaccharide O-polysaccharide-dependent manner via the TLR4-TRAM-TRIF-IRF3-IFNAR1 pathway. Absence of SARM1 reduces the intracellular survival of K. pneumoniae in macrophages, whereas sarm1-deficient mice control the infection. Altogether, our results illustrate an anti-immunology strategy deployed by a human pathogen. SARM1 inhibition will show a beneficial effect to treat Klebsiella infections.

A novel mechanism for macrophage pyroptosis in rheumatoid arthritis induced by Pol ß deficiency.

Rheumatoid arthritis (RA) is a chronic and inflammatory autoimmune disease. Macrophage pyroptosis, a proinflammatory form of cell death, is critically important in RA; however, the detailed mechanism underlying pyroptosis induction is not yet well understood. Here, we report that DNA polymerase ß (Pol ß), a key enzyme in base excision repair, plays a pivotal role in RA pathogenesis. Our data shows that Pol ß expression is significantly decreased in peripheral blood mononuclear cells (PBMCs) from active RA patients and collagen-induced arthritis (CIA) mice, and Pol ß deficiency increases the incidence of RA, macrophage infiltration, and bone destruction in CIA mouse models. In vitro, experiments showed that Pol ß deficiency exacerbated macrophage pyroptosis induced by LPS plus ATP, while overexpression of Pol ß inhibited macrophage pyroptosis. Further characterization revealed that Pol ß knockout resulted in DNA damage accumulation and cytosolic dsDNA leakage, which activated the cGAS-STING-NF-κB signaling pathway and upregulated the expression of NLRP3, IL-1 ß, and IL-18. In conclusion, our findings clarify the influence of Pol ß on the development of RA and provide a detailed explanation for the STING-NF-κB pathway to induce macrophage pyroptosis.

Evaluation of the Efficiency of MRI-Based Radiomics Classifiers in the Diagnosis of Prostate Lesions.

Objective: To compare the performance of different imaging classifiers in the prospective diagnosis of prostate diseases based on multiparameter MRI. Methods: A total of 238 patients with pathological outcomes were enrolled from September 2019 to July 2021, including 142 in the training set and 96 in the test set. After the regions of interest were manually segmented, decision tree (DT), Gaussian naive Bayes (GNB), XGBoost, logistic regression, random forest (RF) and support vector machine classifier (SVC) models were established on the training set and tested on the independent test set. The prospective diagnostic performance of each classifier was compared by using the AUC, F1-score and Brier score. Results: In the patient-based data set, the top three classifiers of combined sequences in terms of the AUC were logistic regression (0.865), RF (0.862), and DT (0.852); RF "was significantly different from the other two classifiers (P =0.022, P =0.005), while logistic regression and DT had no statistical significance (P =0.802). In the lesions-based data set, the top three classifiers of combined sequences in terms of the AUC were RF (0.931), logistic regression (0.922) and GNB (0.922). These three classifiers were significantly different from. Conclusion: The results of this experiment show that radiomics has a high diagnostic efficiency for prostate lesions. The RF classifier generally performed better overall than the other classifiers in the experiment. The XGBoost and logistic regression models also had high classification value in the lesions-based data set.

Polß modulates the expression of type I interferon via STING pathway.

DNA Polymerase ß (Polß) is a key enzyme in base excision repair (BER), which is very important in maintaining the stability and integrity of the genome. Mutant Polß is closely associated with carcinogenesis. However, Polß is highly expressed in most cancers, but the underlying mechanism is not well understood. Here, we found that breast cancer cells MCF-7 with Polß knockdown exhibited high levels of type I interferon and were easily eliminated by natural killer (NK) cells.Similarly, Polß-mutant (R137Q) mice exhibited chronic inflammation symptoms in multiple organs and upregulated type I interferon levels. Further results showed that Polß deficiency caused more DNA damage accumulation in cells and triggered the leakage of damaged DNA into the cytoplasm, which activated the STING/IRF3 pathway, promoted phosphorylated IRF3 translocating into the nucleus and enhanced the expression of type I interferon and proinflammatory cytokines. In addition, this effect could be eliminated by Polß overexpression, STING inhibitor or STING knockdown. Taken together, our findings provide mechanistic insight into the role of Polß in cancers by linking DNA repair and the inflammatory STING pathway.

TLR7 Mediates Acute Respiratory Distress Syndrome in Sepsis by Sensing Extracellular miR-146a.

TLR7 (Toll-like receptor 7), the sensor for single-stranded RNA, contributes to systemic inflammation and mortality in murine polymicrobial sepsis. Recent studies show that extracellular miR-146a-5p serves as a TLR7 ligand and plays an important role in regulating host innate immunity. However, the role of miR-146a-5p and TLR7 signaling in pulmonary inflammation, endothelial activation, and sepsis-associated acute respiratory distress syndrome remains unclear. Here, we show that intratracheal administration of exogenous miR-146a-5p in mice evokes lung inflammation, activates endothelium, and increases endothelial permeability via TLR7-dependent mechanisms. TLR7 deficiency attenuates pulmonary barrier dysfunction and reduces lung inflammatory response in a murine sepsis model. Moreover, the impact of miR-146a-5p-TLR7 signaling on endothelial activation appears to be a secondary effect because TLR7 is undetectable in the human pulmonary artery and microvascular endothelial cells (ECs), which show no response to direct miR-146a-5p treatment in vitro. Both conditioned media of miR-146a-5p-treated macrophages (Mϕ) and septic sera of wild-type mice induce a marked EC barrier disruption in vitro, whereas Mϕ conditioned media or septic sera of TLR7-/- mice do not exhibit such effect. Cytokine array and pathway enrichment analysis of the Mϕ conditioned media and septic sera identify TNFα (tumor necrosis factor α) as the main downstream effector of miR-146a-5p-TLR7 signaling responsible for the EC barrier dysfunction, which is further supported by neutralizing anti-TNFα antibody intervention. Together, these data demonstrate that TLR7 activation elicits pulmonary inflammation and endothelial barrier disruption by sensing extracellular miR-146a-5p and contributes to sepsis-associated acute respiratory distress syndrome.

L-theanine prevents progression of nonalcoholic hepatic steatosis by regulating hepatocyte lipid metabolic pathways via the CaMKKß-AMPK signaling pathway.

BACKGROUND: L-theanine, a non-protein amino acid was found principally in the green tea, has been previously shown to exhibit potent anti-obesity property and hepatoprotective effect. Herein, we investigated the effects of L-theanine on alleviating nonalcoholic hepatic steatosis in vitro and in vivo, and explored the underlying molecular mechanism. METHODS: In vitro, HepG2 and AML12 cells were treated with 500 µM oleic acid (OA) or treated with OA accompanied by L-theanine. In vivo, C57BL/6J mice were fed with normal control diet (NCD), high-fat diet (HFD), or HFD along with L-theanine for 16 weeks. The levels of triglycerides (TG), accumulation of lipid droplets and the expression of genes related to hepatocyte lipid metabolic pathways were detected in vitro and in vivo. RESULTS: Our data indicated that, in vivo, L-theanine significantly reduced body weight, hepatic steatosis, serum levels of alanine transaminase (ALT), aspartate transaminase (AST), TG and LDL cholesterol (LDL-C) in HFD-induced nonalcoholic fatty liver disease (NAFLD) mice. In vitro, L-theanine also significantly alleviated OA induced hepatocytes steatosis. Mechanic studies showed that L-theanine significantly inhibited the nucleus translocation of sterol regulatory element binding protein 1c (SREBP-1c) through AMPK-mTOR signaling pathway, thereby contributing to the reduction of fatty acid synthesis. We also identified that L-theanine enhanced fatty acid ß-oxidation by increasing the expression of peroxisome proliferator-activated receptor α (PPARα) and carnitine palmitoyltransferase-1 A (CPT1A) through AMP-activated protein kinase (AMPK). Furthermore, our study indicated that L-theanine can active AMPK through its upstream kinase Calmodulin-dependent protein kinase kinase-ß (CaMKKß). CONCLUSIONS: Taken together, our findings suggested that L-theanine alleviates nonalcoholic hepatic steatosis by regulating hepatocyte lipid metabolic pathways via the CaMKKß-AMPK signaling pathway.

DDX3X functionally and physically interacts with Estrogen Receptor-alpha.

DEAD-box protein 3X (DDX3X) is a human DEAD-box protein with conventional roles in RNA metabolism and unconventional functions in signalling pathways that do not require its enzymatic activity. For example, DDX3X acts as a multifunctional adaptor molecule in anti-viral innate immune signalling pathways, where it interacts with and regulates the kinase IKB-kinase-epsilon (IIKKε). Interestingly, both DDX3X and IKKɛ have also independently been shown to act as breast cancer oncogenes. IKKɛ's oncogenic functions are likely multifactorial, but it was suggested to phosphorylate the transcription factor Estrogen receptor alpha (ERα) at Serine 167, which drives expression of Erα target genes in an estrogen-independent manner. In this study, we identified a novel physical interaction between DDX3X and ERα that positively regulates ERα activation. DDX3X knockdown in ER+ breast cancer cell lines resulted in reduced ERα phosphorylation, reduced Estrogen Response Element (ERE)-controlled reporter gene expression, decreased expression of ERα target genes, and decreased cell proliferation. Vice versa, overexpression of DDX3X resulted in enhanced ERα phosphorylation and activity. Furthermore, we provide evidence that DDX3X physically binds to ERα from co-immunoprecipitation and pulldown experiments. Based on our data, we propose that DDX3X acts as an adaptor to facilitate IKKε-mediated ERα activation, akin to the mechanism we previously elucidated for IKKε-mediated Interferon Regulatory factor 3 (IRF3) activation in innate immune signalling. In conclusion, our research provides a novel molecular mechanism that might contribute to the oncogenic effect of DDX3X in breast cancer, potentially linking it to the development of resistance against endocrine therapy.

FEN1 inhibitor synergizes with low-dose camptothecin to induce increased cell killing via the mitochondria mediated apoptotic pathway.

Camptothecin has been used in tumor therapy for a long time but its antitumor effect is rather limited due to the side effect and the drug resistance. FEN1, a major component of DNA repair systems, plays important roles in maintaining genomic stability via DNA replication and repair. Here we found that FEN1 inhibitor greatly sensitizes cancer cells to low-dose camptothecin. The combinative treatment of FEN1 inhibitor and 1 nM camptothecin induced a synthetic lethal effect, which synergistically suppressed cancer cell proliferation and significantly mediated apoptosis both in vitro and in vivo. Our study suggested that targeting FEN1 could be a potent strategy for tumor-targeting cancer therapy.