Rational design of a near-infrared fluorescent probe for monitoring butyrylcholinesterase activity and its application in development of inhibitors.

Butyrylcholinesterase (BChE) is widely expressed in multiple tissues and has a vital role in several key human disorders, such as Alzheimer's disease and tumorigenesis. However, the role of BChE in human disorders has not been investigated. Thus, to quantitatively detect and visualize dynamical variations in BChE activity is essential for exploring the biological roles of BChE in the progression of a number of human disorders. Herein, based on the substrate characteristics of BChE, we customized and synthesized three near-infrared (NIR) fluorescent probe substrates with cyanine-skeleton, and finally selected a NIR fluorescence probe substrate named CYBA. The CYBA demonstrated a significant increase in fluorescence when interacting with BChE, but mainly avoided AChE. Upon the addition of BChE, CYBA could be specifically hydrolyzed to TBO, resulting in a significant NIR fluorescence signal enhancement at 710 nm. Systematic evaluation revealed that CYBA exhibited exceptional chemical stability in complex biosamples and possessed remarkable selectivity and sensitivity towards BChE. Moreover, CYBA was successfully applied for real-time imaging of endogenous BChE activity in two types of nerve-related living cells. Additionally, CYBA demonstrated exceptional stability in the detection of complex biological samples in plasma recovery studies (97.51%-104.01%). Furthermore, CYBA was used to construct a high-throughput screening (HTS) method for BChE inhibitors using human plasma as the enzyme source. We evaluated inhibitory effects of a series of natural products and four flavonoids were identified as potent inhibitors of BChE. Collectively, CYBA can serve as a practical tool to track the changes of BChE activity in complicated biological environments due to its excellent capabilities.

Microwave ablation enhances the systemic immune response in patients with lung cancer.

Microwave ablation (MWA) is a key alternative therapy to conventional surgery for the treatment of lung cancer. In addition to eliminating local tumors, MWA may promote antitumor immunological responses, such as abscopal effects in distant lesions. However, the intensity of MWA is limited and the underlying mechanisms are not well-defined. The present study assessed the impact of MWA on immune cell subsets and cytokines in patients with lung cancer. A total of 45 patients with lung cancer who underwent percutaneous lung tumor MWA were enrolled. Peripheral blood samples were collected before and 24 h after MWA and changes in immune cell subsets [lymphocytes, CD3+, CD4+ and CD8+ T cells, B cells and natural killer (NK) cells] and serum cytokine levels (IL-1ß, IL-2, IL-4-6, IL-8, IL-10, IL-12p70, IL-17A and F, IL-22, TNF-α, TNF-ß and IFN-γ) were assessed by flow cytometry and ELISA. The number of total lymphocytes, CD4+ T and NK cells in the peripheral blood significantly decreased 24 h after MWA, while number of CD8+ T cells remained stable, leading to a higher proportion of CD8+ T cells. In addition, the serum levels of IL-2, IL-1ß, IL-6, IL-12p70, IL-22, TNF-α and IFN-γ were significantly increased 24 h after MWA, indicating a T helper 1 type immune response. The immune response in patients with advanced stage disease was comparable with patients in the early stage group; however, the number of total lymphocytes and CD3+ T cells significantly decreased and the ratio of CD4/CD8 and IL-2 levels significantly increased. The early immune response after MWA may contribute to systemic antitumor immunity in patients with both early and advanced disease. Thus, MWA may exhibit potential as a local therapy and trigger abscopal effects in distant lesions in patients with lung cancer.

The mechanism of Sanzi Yangqin decoction for asthma treatment based on network pharmacology and experimental verification.

BACKGROUND: Asthma is a chronic airway inflammatory disease characterized by airway inflammation, mucus hypersecretion, airway hyper-reactivity. Sanzi Yangqin Decoction (SZYQD) is widely prescribed for asthma treatment. Its anti-asthma activities have been reported in animal model, but the exact mechanism and targets of SZYQD in asthma treatment have not been fully elucidated. METHODS: A network pharmacological approach was used to predict the active components, targets, and signalling pathways of SZYQD in asthma, including potential target prediction, protein‒protein interaction (PPI) network construction and analysis, and Gene Ont (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. The active ingredients were identified from the SZYQD, and were molecular docked according to the results of network pharmacology. A mouse model of asthma induced by ovalbumin (OVA) and lipopolysaccharide (LPS) was constructed to evaluate the therapeutic effect of SZYQD. Furthermore, the effects of SZYQD and its active ingredients were tested in vitro for regulating inflammation and MUC5AC expression (two main pathophysiologic abnormalities of asthma) in macrophages and airway epithelial cells by using Real-time PCR and western blotting. RESULTS: A total of 28 active ingredients and 111 HUB genes were screened in the relevant databases, including three key ingredients (luteolin, ß-carotene, and Sinapine) and nine core target genes (JUN, CTNNB1, IL10, TP53, AKT1, STAT3, TNF, IL6 and EGFR). KEGG and GO analysis indicated that the potential anti-asthmatic mechanisms of SZYQD were related to PI3K-Akt signalling pathway and response to lipopolysaccharide, etc. In the in vivo asthmatic model, our findings demonstrated that SZYQD exerted a protective effect against asthmatic mice induced by OVA and LPS through the inhibition of inflammation and mucus overproduction. Consistently, cell experiments showed that the SZYQD extract or the key active ingredients luteolin significantly decreased lipopolysaccharide (LPS)-induced IL-6 expression and activation of the NF-κB pathway in macrophages. In addition, SZYQD extract or luteolin inhibited activation of the AKT pathway and expression of MUC5AC induced by EGF in airway epithelial cells. CONCLUSION: The anti-asthmatic mechanism of SZYQD might be associated with inhibiting inflammation and airway mucus hypersecretion by regulating the NF-κB and AKT signalling pathways as predicted by network pharmacology, which provides more evidence for the application of SZYQD in asthma treatment.

Wogonin alleviates BaP-induced DNA damage and oxidative stress in human airway epithelial cells by dual inhibiting CYP1A1 activity and expression.

Benzo[a]pyrene (BaP) is a common air pollutant that has been reported to cause oxidative stress and carcinogenesis. Wogonin, a flavonoid compound extracted from the roots of Scutellaria baicalensis, has been found to possess a variety of pharmacological activities, including anti-inflammatory and anti-cancer effects. The purpose of this study was to examine the ability of wogonin to alleviate the cytotoxicity induced by BaP in human airway epithelial cells and explore the corresponding mechanism. Our study found that wogonin treatment inhibited DNA damage and reactive oxygen species overproduction induced by BaP in human airway epithelial cells. In vitro enzyme assays showed that wogonin significantly inhibited the enzymatic activity of CYP1A1. In addition, wogonin decreased the basal level of CYP1A1 and inhibited the CYP1A1 overexpression induced by BaP, whereas overexpression of CYP1A1 partially reversed the effect of wogonin on BaP-induced DNA damage. Meanwhile, a CYP1A1 inhibitor and CYP1A1 knockdown also showed these same effects. Further studies showed that wogonin regulates CYP1A1 expression by inhibiting CDK7 and CDK9 activity. The use of CDK7 or CDK9 inhibitors decreased BaP-induced cytotoxicity and CYP1A1 expression. Finally, we found that the methoxy group of wogonin was crucial for its inhibitory activity. In conclusion, our data indicated that wogonin could effectively relieve BaP induced cytotoxicity, and its mechanism was related to the dual inhibition of CYP1A1 activity and expression.

Therapeutic effects of natural polyphenols on colorectal adenomas: Focus on preclinical studies (Review).

Colorectal adenoma (CRA) is a premalignant lesion of colorectal cancer. The current treatment is surgical resection, but CRA is prone to recurrence, and there is no safe and effective drug to prevent adenoma recurrence and canceration. Recent studies have shown that natural compounds in plants have favorable antitumor effects. According to preclinical studies, natural polyphenols can regulate different signal pathways and targets to play a role in the treatment of CRA, which is closely related to its inhibition of proliferation, induction of apoptosis, inhibition of inflammation and oxidative stress, and regulation of intestinal flora. Natural polyphenols are potential candidates for CRA therapy due to their remarkable efficacy and safety. In the present review, attention was paid to the experimental research progress of natural polyphenols extracted from numerous plants in the treatment of CRA in the last 10 years. The present review provided new guidance for the study of CRA, clarified the therapeutic role of polyphenols in CRA, and evaluated for the first time, to the best of our knowledge, the therapeutic potential of natural polyphenols to treat CRA by targeting multiple genes and signal pathways and epigenetic modification.

Autocrine TGF-alpha is associated with Benzo(a)pyrene-induced mucus production and MUC5AC expression during allergic asthma.

OBJECTS: Benzo(a)pyrene (BaP), an environmental pollutant, is present in high concentrations in urban smog and cigarette smoke and has been reported to promote high mucin 5AC (MUC5AC) expression. Epithelium-derived inflammatory cytokines are considered an important modulator of mucus oversecretion and MUC5AC overexpression. Here, we investigated whether the effect of BaP on MUC5AC overexpression was associated with cytokine autocrine activity in vivo and in vitro. METHODS: In vivo, BALB/c mice were treated with ovalbumin (OVA) in the presence or absence of BaP. Allergy-induced mucus production was assessed by Alcian Blue Periodic acid Schiff (AB-PAS) staining. The human airway epithelial cell line NCI-H292 was used in vitro. MUC5AC and transforming growth factor (TGF)-α mRNA levels were assessed with real-time quantitative PCR. The concentration of cytokines was measured by ELISA. The MUC5AC, p-ERK, ERK, p-EGFR and EGFR proteins were detected by Western blotting in cells or by immunohistochemistry in mouse lungs. Small-interfering RNAs were used for gene silencing. RESULTS: TGF-α was overproduced in the supernatant of NCI-H292 cells treated with BaP. Knockdown of TGF-α expression inhibited the BaP-induced increase in MUC5AC expression and subsequent activation of the EGFR-ERK signalling pathway. Knocking down aryl hydrocarbon receptor (AhR) expression or treatment with an ROS inhibitor (N-acetyl-L-cysteine) could relieve the TGF-α secretion induced by BaP in epithelial cells. In an animal study, coexposure to BaP with OVA increased mucus production, MUC5AC expression and ROS-EGFR-ERK activation in the lung as well as TGF-α levels in bronchoalveolar lavage fluid (BALF). Furthermore, the concentration of TGF-α in BALF was correlated with MUC5AC mRNA levels. Additionally, TGF-α expression was found to be positively correlated with MUC5AC expression in the airway epithelial cells of smokers. Compared with non-smoker asthma patients, TGF-α serum levels were also elevated in smoker asthma patients. CONCLUSION: Autocrine TGF-α was associated with BaP-induced MUC5AC expression in vitro and in vivo. BaP induced TGF-α secretion by activating AhR and producing ROS, which led to activation of the EGFR-ERK pathway.

Resveratrol inhibits MUC5AC expression by regulating SPDEF in lung cancer cells.

BACKGROUND: MUC5AC was recently identified to play important roles in the proliferation and metastasis of malignant mucinous lung tumor cells. Resveratrol (Res), a natural compound with anticancer effects in lung cancer cells, has been reported to inhibit mucin production in airway epithelial cells. This study aimed to investigate the inhibitory effect of Res on MUC5AC expression in lung mucinous adenocarcinoma cells and the potential mechanisms. METHODS: Mucus-producing A549 human lung carcinoma cells were used to test the effects of Res on SPDEF and MUC5AC expression. Gene and protein expression was assessed by real-time quantitative PCR (qPCR), immunofluorescence and western blotting assays. SPDEF lentivirus was used to upregulate SPDEF expression levels in mucus-producing A549 human lung carcinoma cells. Cell proliferation was assessed by Cell Counting Kit-8 (CCK-8) assay. RESULTS: Res decreased MUC5AC expression in an SPDEF-dependent manner in mucus-producing A549 human lung carcinoma cells, and this change was accompanied by decreased ERK expression and AKT pathway activation. Moreover, SPDEF was found to be overexpressed in lung adenocarcinoma (LUAD), especially in mucinous adenocarcinoma. In-vitro functional assays showed that overexpression of SPDEF reduced the chemosensitivity of A549 cells to cisplatin (DDP). In addition, Res treatment increased A549 cell chemosensitivity to DDP by inhibiting the SPDEF-MUC5AC axis. CONCLUSION: Our results indicate that the SPDEF-MUC5AC axis is associated with DDP sensitivity, and that Res decreases SPDEF and MUC5AC expression by inhibiting ERK and AKT signaling in A549 cells, which provides a potential pharmacotherapy for the prevention and therapeutic management of mucinous adenocarcinoma.

Investigation of the Active Ingredients and Mechanism of Polygonum cuspidatum in Asthma Based on Network Pharmacology and Experimental Verification.

BACKGROUND: Polygonum cuspidatum is a Chinese medicine commonly used to treat phlegm-heat asthma. However, its anti-asthmatic active ingredients and mechanism are still unknown. The aim of this study was to predict the active ingredients and pathways of Polygonum cuspidatum and to further explore the potential molecular mechanism in asthma by using network pharmacology. METHODS: The active ingredients and their targets related to Polygonum cuspidatum were seeked out with the TCM systematic pharmacology analysis platform (TCMSP), and the ingredient-target network was constructed. The GeneCards, DrugBank and OMIM databases were used to collect and screen asthma targets, and then the drug-target-disease interaction network was constructed with Cytoscape software. A target protein-protein interaction (PPI) network was constructed using the STRING database to screen key targets. Finally, GO and KEGG analyses were used to identify biological processes and signaling pathways. The anti-asthmatic effects of Polygonum cuspidatum and its active ingredients were tested in vitro for regulating airway smooth muscle (ASM) cells proliferation and MUC5AC expression, two main symptoms of asthma, by using Real-time PCR, Western blotting, CCK-8 assays and annexin V-FITC staining. RESULTS: Twelve active ingredients in Polygonum cuspidatum and 479 related target proteins were screened in the relevant databases. Among these target proteins, 191 genes had been found to be differentially expressed in asthma. PPI network analysis and KEGG pathway enrichment analysis predicted that the Polygonum cuspidatum could regulate the AKT, MAPK and apoptosis signaling pathways. Consistently, further in vitro experiments demonstrated that Polygonum cuspidatum and resveratrol (one active ingredient of Polygonum cuspidatum) were shown to inhibit ASM cells proliferation and promoted apoptosis of ASM cells. Furthermore, Polygonum cuspidatum and resveratrol inhibited PDGF-induced AKT/mTOR activation in ASM cells. In addition, Polygonum cuspidatum decreased H2O2 induced MUC5AC overexpression in airway epithelial NCI-H292 cells. CONCLUSION: Polygonum cuspidatum could alleviate the symptoms of asthma including ASM cells proliferation and MUC5AC expression through the mechanisms predicted by network pharmacology, which provides a basis for further understanding of Polygonum cuspidatum in the treatment of asthma.

Benzo(a)pyrene induces MUC5AC expression through the AhR/mitochondrial ROS/ERK pathway in airway epithelial cells.

OBJECTIVES: Benzo(a)pyrene (BaP) is a ubiquitous air pollutants, and BaP exposure leads to a risk of respiratory diseases. The oversecretion of airway mucus and high expression of mucin 5AC (MUC5AC) are associated with common respiratory disorders caused by air pollution. We aimed to investigate the effect of BaP on MUC5AC expression, especially the mechanisms by which BaP induces MUC5AC gene expression. METHODS: The human airway epithelial cell NCI-H292 was used to test the effects of BaP on the expression of MUC5AC in vitro. MUC5AC mRNA and protein expression were assessed with real-time quantitative PCR, immunochemistry, and western blotting. A luciferase assay was conducted to detect the activity of the promoter. The total cellular ROS and mitochondrial ROS were measured by corresponding probes. Small-interfering RNAs were used for gene silencing. AhR-overexpressing cell lines were constructed by transfection with AhR overexpression lentivirus. RESULTS: We found that BaP stimulation upregulated the MUC5AC mRNA and protein levels and activated the ERK pathway. Suppressing ERK with U0126 (an ERK inhibitor) or knocking down ERK with siRNA decreased BaP-induced MUC5AC expression. The luciferase activity transfected with the MUC5AC promoter and cAMP-response element (CRE) was increased after BaP treatment, whereas CREB siRNA suppressed the BaP-induced overexpression of MUC5AC. In addition, BaP increased mitochondrial ROS production, and Mito-TEMP, a mitochondrial ROS inhibitor, inhibited BaP-induced MUC5AC expression and ERK activation. BaP increased the mRNA levels of CYP1A1 and CYP1B1, while Alizarin, a CYP1s inhibitor, suppressed the effects of BaP, including the MUC5AC overexpression, ERK activation and mitochondrial ROS generation. BaP induced the translocation of aryl hydrocarbon receptor (AhR) from the cytoplasm to the nucleus. SiRNA-mediated knockdown or chemical inhibition of AhR decreased the BaP-induced expression of MUC5AC, while the overexpression of AhR significantly enhanced the BaP-induced expression of MUC5AC. ITE, an endogenous AhR ligand, also upregulated the mRNA and protein expression of MUC5AC. Furthermore, resveratrol treatment inhibited the BaP-induced MUC5AC overexpression, AhR translocation, mitochondrial ROS production and ERK pathway activation. CONCLUSION: Here, we highlighted the crucial role of AhR/mitochondrial ROS/ERK pathway activation in BaP-induced MUC5AC overexpression and identified resveratrol as a promising drug to reduce BaP-induced MUC5AC overexpression.

Qi-Xian Decoction Upregulated E-cadherin Expression in Human Lung Epithelial Cells and Ovalbumin-Challenged Mice by Inhibiting Reactive Oxygen Species-Mediated Extracellular-Signal-Regulated Kinase (ERK) Activation.

BACKGROUND Loss of the epithelial barrier is characterized by a reduction in E-cadherin expression and is a hallmark of asthma. Qi-xian decoction (QXT) is a Chinese medicinal formula that has been used to effectively treat asthma. This study aimed to investigate the effect of QXT on E-cadherin expression in human lung epithelial 16HBE cells and ovalbumin-challenged mice and to explore the underlying molecular mechanism. MATERIAL AND METHODS Ovalbumin (OVA)-induced mice were used as a model of asthma. Real-time PCR and Western blotting were utilized to examine mRNA and protein levels. Lung tissue reactive oxygen species (ROS) levels were evaluated using dichloro-dihydro-fluorescein diacetate (DCFH-DA). Serum superoxide dismutase (SOD) and the total antioxidant capacity (TAOC) were measured via enzyme-linked immunosorbent assay (ELISA)-based analyses. 16HBE cells were utilized to explore the effect of QXT or hydrogen peroxide (H2O2) on the expression of E-cadherin in vitro. RESULTS We found that QXT treatment increased E-cadherin expression and decreased extracellular-signal-regulated kinase (ERK) phosphorylation levels in the lung tissues of OVA-challenged mice. QXT also downregulated ROS levels and increased serum SOD and TAOC levels in OVA-challenged mice. In vitro studies demonstrated that increased ROS generation induced by H2O2 resulted in decreased E-cadherin expression levels in 16HBE cells, which was attenuated by inhibition of ERK signaling. Moreover, the H2O2-induced downregulation of E-cadherin expression, increased ROS generation, and ERK activation in 16HBE cells were restored by treatment with QXT water or ethanol extract. CONCLUSIONS These data demonstrate that one mechanism by which QXT protects against asthma is to restore E-cadherin expression in vivo and in vitro by inhibiting ROS-mediated ERK activation.

Decreasing Arl4c expression by inhibition of AKT signal in human lung adenocarcinoma cells.

PURPOSE: Arl4c is overexpressed in several cancer tissues and is involved in cancer development. Nevertheless, the exact mechanism that regulates Arl4c expression in lung cancer has not been fully elucidated. The aim of this study was to investigate the regulatory mechanism of Arl4c and to explore potential chemotherapeutic drugs targeting Arl4c. METHODS: Immunohistochemistry was used to examine Arl4c expression levels in human lung adenocarcinoma cancer specimens. Protein expression was detected by western blot. Overexpression of Arl4c-Flag protein was used to detect the ubiquitination of Arl4c. A short interfering RNA against Arl4c was used for gene silencing. RESULTS: Arl4c was overexpressed in lung cancer tissues, and knockdown of Arl4c expression by siRNA decreased lung cancer A549 and 95-D cell proliferation. In addition, Arl4c expression was downregulated via inhibition of the AKT pathway in A549 and 95-D cells, whereas exposure to benzo (a) pyrene (a carcinogen in smoke) increased Arl4c expression in 16HBE cells via AKT activation. Finally, we found that chemotherapy drug hydroxycamptothecin (HCPT) could decrease Arl4c expression levels by inhibiting the activation of the AKT pathway in A549 and 95-D cells. Moreover, accumulation of ubiquitinated Arl4c protein was increased by HCPT and LY294002 (an AKT inhibitor) treatment whereas the proteasome inhibitor MG-132 attenuated the inhibitory effect of HCPT and LY294002 on Arl4c expression. CONCLUSION: Here, we highlighted the AKT pathway as an important regulatory pathway for Arl4c expression in lung cancer cells and identified HCPT as a promising drug for lung adenocarcinoma treatment that functioned by targeting Arl4c expression.

Ursolic Acid Reverses the Chemoresistance of Breast Cancer Cells to Paclitaxel by Targeting MiRNA-149-5p/MyD88.

Paclitaxel (PTX) is widely used as a front-line chemotherapy for breast cancer treatment. However, its clinical applications are limited by the development of chemoresistance. The objective of this study was to investigate the reversal effects of ursolic acid (UA) on PTX resistance and the possible mechanisms in breast cancer. The role of miRNA-149-5p/MyD88 in the regulation of PTX resistance was investigated by the transfection of breast cancer cells with MDA-MB-231 (231) and MDA-MB-231/PTX-resistance (231/PTX) with lentiviruses carrying the MyD88 gene, shRNA specific for MyD88, the miR-149-5p gene, and shRNA specific for miR-149-5p. The PTX sensitivity was assessed by a CCK-8 assay. qRT-PCR and Western blot analyses were used to detect changes in the mRNA and protein levels. Flow cytometry was used to measure the rate of cell apoptosis. A luciferase activity assay was used to detect the binding site of miR-149-5p on the 3'UTR of MyD88. 231/PTX cells were injected into the flanks of female athymic nude mice, and the mice were randomly divided into the five following groups: PBS, PTX (low), PTX (high), UA, and PTX+UA. Our data show that UA reversed the resistance of breast cancer 231/PTX cells to PTX in vitro and in vivo. UA treatment significantly increased the expression of miR-149-5p, which was lower in 231/PTX cells than in 231 cells. Furthermore, the overexpression of miR-149-5p increased the sensitivity of 231/PTX cells to PTX treatment, whereas the knockdown of the miR-149-5p gene attenuated the effects of UA on the regulation of PTX sensitivity. A luciferase assay demonstrated that miR-149-5p could directly regulate the transcriptional activity of MyD88, a known PTX-resistance gene, by targeting the 3'UTR of MyD88. Meanwhile, the downregulation of MyD88 through the overexpression of miR-149-5p or UA treatment inhibited the activation of the Akt signaling pathway in 231/PTX cells. Thus, our data indicate that UA can reverse PTX resistance by targeting the miRNA-149-5p/MyD88 axis in breast cancer cells.

The Zuo Jin Wan Formula increases chemosensitivity of human primary gastric cancer cells by AKT mediated mitochondrial translocation of cofilin-1.

Resistance to cisplatin (DDP)-based chemotherapy is a major cause of treatment failure in human gastric cancer (GC). It is necessary to identify the drugs to re-sensitize GC cells to DDP. In our previous research, Zuo Jin Wan Formula (ZJW) has been proved could increase the mitochondrial apoptosis via cofilin-1 in a immortalized cell line, SGC-7901/DDP. Due to the immortalized cells may still difficult highly recapitulate the important molecular events in vivo, primary GC cells model derived from clinical patient was constructed in the present study to further evaluate the effect of ZJW and the underlying molecular mechanism. Immunofluorescent staining was used to indentify primary cultured human GC cells. Western blotting was carried out to detect the protein expression. Cell Counting Kit-8 (CCK-8) was used to evaluate cell proliferation. Flow cytometry analysis was performed to assess cell apoptosis. ZJW inhibited proliferation and induced apoptosis in primary DDP-resistant GC cells. Notably, the apoptosis in GC cells was mediated by inducing cofilin-1 mitochondrial translocation, down-regulating Bcl-2 and up-regulating Bax expression. Surprisingly, the level of p-AKT protein was higher in DDP-resistant GC cells than that of the DDP-sensitive GC cells, and the activation of AKT could attenuate ZJW-induced sensitivity to DDP. These data revealed that ZJW can increase the chemosensitivity in DDP-resistant primary GC cells by inducing mitochondrial apoptosis and AKT inactivation. The combining chemotherapy with ZJW may be an effective therapeutic strategy for GC chemoresistance patients.

Expression and biological function of rhotekin in gastric cancer through regulating p53 pathway.

BACKGROUND/AIM: Gastric cancer (GC) is one of a most threatening cancer globally. Rhotekin (RTKN), a Rho effector, has been reported to be upregulated in GC tissues. This study aimed to investigate the underlying regulatory roles of RTKN in the biological behavior of GC. METHODS: Real-time PCR and Western blotting were carried out to detect the mRNA and protein expression, respectively. Cell Counting Kit-8 and xenograft nude mice model were used to evaluate cell proliferation. Flow cytometry analysis was performed to assess cell cycle distribution and cell apoptosis. RESULTS: RTKN had high expression level in GC compared with normal tissues. RTKN expression strongly associated with tumor size, TNM stage, lymphnode metastasis and the poor prognosis of patients with GC. Downregulation of RTKN significantly repressed GC cell proliferation, but increased cell population in G1/S phase and induced cell apoptosis. Moreover, the RTKN expression level was related to the p53 signaling pathway and histone deacetylase (HDAC) Class I pathway. RTKN knockdown caused a notable increment in the acetylation level of p53 (Lys382), and the expression of p53-target genes (p21, Bax, and PUMA), as well as a reduction in the expression of a potential deacetylase for p53, HDAC1. Notably, downregulation of HDAC1 had similar effects as RTKN knockdown, and RTKN overexpression could hardly abrogate the effects of HDAC1 knockdown on GC cells. CONCLUSION: RTKN could work as an oncogene via regulating HDAC1/p53 and may become a promising treatment strategy for GC.

Sitagliptin reduces insulin resistance and improves rat liver steatosis via the SIRT1/AMPKα pathway.

Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease. It is asymptomatic at presentation and is frequently identified among individuals with metabolic dysfunction, including obesity and diabetes. NAFLD is primarily characterized by the accumulation of triacylglycerol in the liver. Since insulin resistance and fat metabolism dysregulation are major causes of type 2 diabetes and NAFLD, anti-diabetes agents are widely considered as potential therapy strategies for NAFLD. Sitagliptin, an inhibitor of dipeptidyl peptidase-4, has been developed as an oral anti-hyperglycemic agent. In the present study, the effect of sitagliptin on the progression of NAFLD was evaluated in a rat model fed with a high fat diet (HFD). It was identified that sitagliptin significantly suppressed lipid accumulation in rat blood and liver and improved insulin resistance. Furthermore, it was revealed that sitagliptin reactivated the HFD-suppressed SIRT1/AMPK axis pathway and upregulated its downstream target genes, modulating fatty acid metabolism. These findings demonstrate a preventive effect of sitagliptin on hepatic lipid dysregulation and suggest that sitagliptin has potential as a clinical therapeutic strategy for NAFLD.

Curcumin suppresses MUC5AC production via interfering with the EGFR signaling pathway.

Excessive mucin production in the airway may contribute to airway inflammatory diseases. Curcumin has been reported to prevent mucin 5AC (MUC5AC) production in human airway epithelial cells; however, the molecular targets of curcumin involved in regulating MUC5AC expression have remained elusive. The present study aimed to elucidate the molecular mechanisms by which curcumin regulates MUC5AC production, utilizing the NCI­H292 human airway epithelial cell line featuring MUC5AC hypersecretion. Curcumin was able to counteract the endothelial growth factor (EGF)­stimulated mRNA and protein expression of MUC5AC. In addition, curcumin treatment prevented EGF­induced phosphorylation of EGF receptor (EGFR) as well as the downstream AKT and signal transducer and activator of transcription 3 (STAT3), while inhibition of PI3K and STAT3 signaling significantly attenuated the expression of MUC5AC that was induced by EGF. Furthermore, EGF­induced increases in the levels of phosphorylated STAT3 in the nuclear fraction were inhibited by curcumin and PI3K inhibitors. In addition, treatment with curcumin significantly decreased MUC5AC and EGFR expression in a time­dependent manner under basal conditions. These results demonstrated that curcumin inhibited MUC5AC protein expression in NCI­H292 cells under basal conditions as well under EGF stimulation. This inhibition was accompanied by decreased activation of the EGFR/AKT/STAT3 pathway and reduced EGFR expression, which indicated that curcumin may have a dual role in interfering with the EGFR signaling pathway and inhibiting mucin expression in human airway epithelial cells.

The outcome and the influencing factors of the age of onset in post-mortem of chronic bronchitis patients: a retrospective study.

Purpose: Chronic bronchitis is thought to occur in elderly patients, and smoking seems to be an important risk factor. The outcomes related to the age of onset in patients with chronic bronchitis are still unclear. Patients and methods: A retrospective study was conducted on deceased patients whose diagnosis included bronchitis from 2010 to 2016. Patients were separated into two groups according to the age of onset (Group I, age ≤50 years old; Group II, age >50 years old). Information regarding disease course, smoking history, death age, number of admissions per year, Hugh Jones Index, and self-reported comorbidities of the patients was recorded. Results: The courses of chronic cough and sputum were 33.38±7.73 years and 14.44±8.60 years in Group I and Group II, respectively (p<0.05). The death ages of Group I and Group II were 77.65±7.87 years and 84.69±6.67 years, respectively (p<0.05). There was a significant negative correlation between the number of hospital admissions per year and the age of onset. The age of onset was negatively associated with daily smoking count (r=-0.210) and total smoking count (r=-0.146). In Group I, there were fewer cases of coronary heart disease (OR =0.41 [0.24-0.71]), neurological diseases (OR =0.48 [0.24-0.97]), and total comorbidities (OR =0.67 [0.54-0.85]) than in Group II. Conclusion: Patients with early onset chronic bronchitis had a longer history, younger death age, poorer health status, and lower incidence of comorbidities.

Curcumin Inhibits Growth of Human NCI-H292 Lung Squamous Cell Carcinoma Cells by Increasing FOXA2 Expression.

Lung squamous cell carcinoma (LSCC) is a common histological lung cancer subtype, but unlike lung adenocarcinoma, limited therapeutic options are available for treatment. Curcumin, a natural compound, may have anticancer effects in various cancer cells, but how it may be used to treat LSCC has not been well studied. Here, we applied curcumin to a human NCI-H292 LSCC cell line to test anticancer effects and explored underlying potential mechanisms of action. Curcumin treatment inhibited NCI-H292 cell growth and increased FOXA2 expression in a time-dependent manner. FOXA2 expression was decreased in LSCC tissues compared with adjacent normal tissues and knockdown of FOXA2 increased NCI-H292 cells proliferation. Inhibition of cell proliferation by curcumin was attenuated by FOXA2 knockdown. Moreover inhibition of STAT3 pathways by curcumin increased FOXA2 expression in NCI-H292 cells whereas a STAT3 activator (IL-6) significantly inhibited curcumin-induced FOXA2 expression. Also, SOCS1 and SOCS3, negative regulators of STAT3 activity, were upregulated by curcumin treatment. Thus, curcumin inhibited human NCI-H292 cells growth by increasing FOXA2 expression via regulation of STAT3 signaling pathways.

Zuo Jin Wan Reverses DDP Resistance in Gastric Cancer through ROCK/PTEN/PI3K Signaling Pathway.

Gastric cancer (GC) is the third leading cause of cancer-related death. Chemotherapy resistance remains the major reason for GC treatment failure and poor overall survival of patients. Our previous studies have proved that Zuo Jin Wan (ZJW), a traditional Chinese medicine (TCM) formula, could significantly enhance the sensitivity of cisplatin (DDP)-resistant gastric cancer cells to DDP by inducing apoptosis via mitochondrial translocation of cofilin-1. However, the underlying mechanism remains poorly understood. This study aimed to evaluate the effects of ROCK/PTEN/PI3K on ZJW-induced apoptosis in vitro and in vivo. We found that ZJW could significantly activate the ROCK/PTEN pathway, inhibit PI3K/Akt, and promote the apoptosis of SGC-7901/DDP cells. Inhibition of ROCK obviously attenuated ZJW-induced apoptosis as well as cofilin-1 mitochondrial translocation, while inhibition of PI3K had the opposite effects. In vivo, combination treatment of DDP and ZJW (2000 mg/kg) significantly reduced tumor growth compared with DDP alone. Moreover, the combined administration of ZJW and DDP increased the expression of cleaved ROCK and p-PTEN while it decreased p-PI3K and p-cofilin-1, which was consistent with our in vitro results. These findings indicated that ZJW could effectively inhibit DDP resistance in GC by regulating ROCK/PTEN/PI3K signaling and provide a promising treatment strategy for gastric cancer.

Association of 758 G/A polymorphism of 3'untranslated region of prohibitin with risk of gastric cancer.

BACKGROUND: A polymorphic variant allele (T-allele) in the 3'-UTR of prohibitin (C-to-T at nucleotide 729) was reported to be associated with an increased risk of breast cancer. However, the association between the 3'-UTR polymorphism of prohibitin and the susceptibility to gastric cancer remains unknown. Thus, we investigated the distribution of prohibitin genotypes in Chinese patients with gastric cancer and subsequently analyzed the association between the 3'-UTR polymorphism of prohibitin and the risk of gastric cancer in that population. METHODS: The distribution of 3'-UTR polymorphism of prohibitin in 82 gastric cancer patients was determined by sequencing and compared with that of 171 healthy controls. Luciferase reporter assay was used to investigate the effect of 3'-UTRs variant on PHB expression. RESULTS: Our study discovered two major polymorphic sites in the 3'-UTR of prohibitin (C-to-T at nucleotide 729 and G to A at nucleotide 758). The C/T polymorphism at 729 site was not associated with the increased risk of gastric cancer (P=.961, OR=1.044, 95%CI: 0.187-5.818); however, G/A polymorphism at nucleotide 758 increased the risk of gastric cancer (P=.017, OR=1.923, 95%CI: 1.119-3.305). Luciferase reporter constructs containing the 758A allele showed higher luciferase activity compared with the wild-type allele, which indicated that 758 G>A in 3'-UTR increased PHB expression. CONCLUSIONS: The G to A transition but not the C-to-T transition in the 3'-UTR of prohibitin was associated with an increased risk of gastric cancer in Chinese population.