OBJECTIVES: To investigate YEATS2 expression in gastric cancer (GC), its prognostic value, and its regulatory role in epithelial-mesenchymal transition (EMT) of GC cells. METHODS: YEATS2 expression in GC was analyzed using publicly available databases. Paired GC and adjacent tissues were collected from 100 patients undergoing radical surgery for immunohistochemical detection of YEATS2 expression, and its correlations with the patients' clinicopathological parameters and Ki67 expression were analyzed. The prognostic value of YEATS2 was assessed using Kaplan-Meier analysis, Cox regression and ROC curves, and its regulatory mechanisms were analyzed using KEGG enrichment analysis. In cultured GC cell lines (HGC-27 and AGS), the effect of YEATS2 knockdown and overexpression on migration, invasion and EMT of the cells were examined with scratching assay, Transwell assay and Western blotting. RESULTS: YEATS2 was significantly overexpressed in GC tissues with a positive correlation with Ki67 (P<0.05). High YEATS2 expression was associated with elevated CEA (≥5 μg/L), CA19-9 (≥37 kU/L), T3-4 stage, and N2-3 stage (all P<0.05). Patients with high YEATS2 expression had significantly reduced 5-year survival (P<0.001); ROC analysis showed that YEATS2 expression levels had a sensitivity of 80.00% and a specificity of 66.67% for predicting patient survival (P<0.05). Cox regression identified high YEATS2 as an independent risk factor for poor postoperative 5-year survival outcome of GC patients (HR: 1.675, 95%CI: 1.013-2.771; P=0.045). KEGG enrichment analysis suggested involvement of YEATS2 in EMT in GC and Wnt/β-catenin signaling. In cultured GC cells, YEATS2 overexpression significantly promoted cell migration and invasion, upregulated the expressions of vimentin, N-cadherin, Wnt and active β-catenin, and downregulated E-cadherin expression, and these changes were obviously suppressed by treatment with XAV-939 (a Wnt/β-catenin inhibitor). CONCLUSIONS High YEATS2 expression activates Wnt/β-catenin signaling to promote EMT in GC and is correlated with poor prognosis of GC patients.
Humans ; Stomach Neoplasms/pathology* ; Epithelial-Mesenchymal Transition ; Wnt Signaling Pathway ; Cell Line, Tumor ; Prognosis ; Cell Movement ; Male ; Female ; beta Catenin/metabolism*

OBJECTIVES: To investigate the effect of hypaphorine (HYP) on Crohn's disease (CD)‑like colitis in mice and its molecular mechanism. METHODS: Thirty male C57BL/6J mice were equally randomized into WT, TNBS, and HYP groups, and in the latter two groups, mouse models of CD-like colitis were established using TNBS with daily gavage of 15 mg/kg HYP or an equivalent volume of saline. The treatment efficacy was evaluated by assessing the disease activity index (DAI), body weight changes, colon length and histopathology. The effect of HYP was also tested in a LPS-stimulated Caco-2 cell model mimicking intestinal inflammation by evaluating inflammatory responses and barrier function of the cells using qRT-PCR and immunofluorescence staining. GO and KEGG analyses were conducted to explore the therapeutic mechanism of HYP, which was validated in both the cell and mouse models using Western blotting. RESULTS: In the mouse models of CD-like colitis, HYP intervention obviously alleviated colitis as shown by significantly reduced body weight loss, colon shortening, DAI and inflammation scores, and expressions of pro-inflammatory factors in the colon tissues. HYP treatment also significantly increased the TEER values, reduced bacterial translocation to the mesenteric lymph nodes, liver, and spleen, lowered serum levels of I-FABP and FITC-dextran, increased the number of colonic tissue cup cells, and upregulated colonic expressions of MUC2 and tight junction proteins (claudin-1 and ZO-1) in the mouse models. In LPS-stimulated Caco-2 cells, HYP treatment significantly inhibited the expressions of pro-inflammatory factors and increased the expressions of tight junction proteins. Western blotting showed that HYP downregulated the expressions of the key proteins in the TLR4/MyD88 signaling pathway in both the in vitro and in vivo models. CONCLUSIONS HYP alleviates CD-like colitis in mice possibly by suppressing intestinal epithelial inflammation and improving gut barrier function.
Animals ; Male ; Mice, Inbred C57BL ; Crohn Disease/drug therapy* ; Mice ; Humans ; Caco-2 Cells ; Intestinal Mucosa/metabolism* ; Colitis/drug therapy* ; Disease Models, Animal ; Inflammation ; Toll-Like Receptor 4/metabolism* ; Myeloid Differentiation Factor 88/metabolism* ; Intestinal Barrier Function

Protein liquid-liquid phase separation (LLPS), a pivotal phenomenon intricately linked to cellular processes, is regulated by various other proteins. However, there is still a lack of high-throughput methods for screening protein regulators of LLPS in target proteins. Here, we developed a CRISPR/Cas9-based screening method to identify protein phase separation regulators by integrating bimolecular fluorescence complementation (BiFC) and fluorescence-activated cell sorting (FACS). Using this newly developed method, we screened the RNA-binding proteins that regulate PABPN1 phase separation and identified the tumor suppressor QKI as a promoter of PABPN1 phase separation. Furthermore, QKI exhibits decreased expression levels and diminished nuclear localization in colorectal cancer cells, resulting in reduced PABPN1 phase separation, which, in turn, promotes alternative polyadenylation (APA), cell proliferation, and migration in colorectal cancer.
Humans ; Colorectal Neoplasms/genetics* ; RNA-Binding Proteins/genetics* ; Poly(A)-Binding Protein I/genetics* ; CRISPR-Cas Systems ; Flow Cytometry ; Cell Proliferation ; Cell Line, Tumor ; Cell Movement

ObjectiveBased on ultra performance liquid chromatography-quadrupole-time-of-flight tandem mass spectrometry（UPLC-Q-TOF-MS）， to evaluate the establishment of a mouse model of liver Yin deficiency by thyroid tablet suspension combined with 10% carbon tetrachloride（CCl₄） from the perspective of non-targeted metabolomics， in order to lay the foundation for the establishment of a traditional Chinese medicine（TCM） syndrome model. MethodA total of 24 mice were randomly divided into blank group and model group. The model group was given thyroid tablet suspension（0.003 2 g·kg^-1） by gavage for 14 consecutive days， and 10% CCl₄（5 mL·kg^-1） was intraperitoneally injected once a week to establish a liver Yin deficiency model， while the blank group was injected with an equal amount of olive oil intraperitoneally and gavaged with an equal amount of distilled water， and was fed with normal feed. After the modeling was completed， 6 mice in each group were randomly selected， the levels of alanine aminotransferase（ALT）， aspartate aminotransferase（AST）， cyclic adenosine monophosphate（cAMP）， cyclic guanosine monophosphate（cGMP）， interleukin（IL）-6， IL-10， tumor necrosis factor-α（TNF-α）were measured in the mice serum， and malondialdehyde（MDA）， superoxide dismutase（SOD）， total protein（TP）， hydroxyproline（HYP） and other indicators were measured in the mice liver. Liver tissue sections were taken for hematoxylin-eosin（HE） staining and observing pathological changes. The remaining 6 mice in each group were subjected to UPLC-Q-TOF-MS combined with principal component analysis（PCA） and orthogonal partial least squares-discriminant analysis（OPLS-DA） were used to screen differential metabolites in the liver Yin deficiency mouse model， Kyoto Encyclopedia of Genes and Genomes（KEGG） database was used to analyze the corresponding metabolic pathways of differential metabolites. ResultCompared with the blank group， mice in the model group showed liver Yin deficiency manifestations such as reduced body weight， fatigue and sleepiness， disheveled and lusterless hair， irritability. The levels of ALT， cAMP/cGMP， IL-6， AST， MDA， cAMP， TNF-α significantly increased（P<0.05， P<0.01）， while the levels of SOD， IL-10 and cGMP significantly decreased（P<0.05， P<0.01）， and the changes of HYP and TP were not statistically significant. Hepatic steatosis and distortion of the radial arrangement of the liver plate cells were seen in the section images of the model group， endogenous substances were clearly separated， and 252 differential metabolites were identified in the serum samples， which were mainly involved in the metabolic pathways of purine metabolism， steroid hormone biosynthesis and pyrimidine metabolism. A total of 229 differential metabolites were identified in the liver samples， mainly involving nucleotide metabolism， purine metabolism， steroid hormone biosynthesis， pyrimidine metabolism， antifolate resistance， insulin resistance， primary bile acid biosynthesis， prostate cancer， sulfur relay system， arachidonic acid metabolism and other metabolic pathways. ConclusionThe successful establishment of liver Yin deficiency model in mice by CCl₄ combined with thyroid hormone is evaluated through the investigation of serum and liver metabolomics， combined with biochemical indicators， which provides a biological basis and experimental foundation for the Yin deficiency syndrome model of TCM.

Objective To explore the application value of chest dual-energy subtraction (DES) technology in the diagnosis of occupational pneumoconiosis. Methods A total of 86 patients with suspected pneumoconiosis and 21 dust-exposure workers were selected as the research subjects using random sampling method. The posterior and anterior chest radiographs were taken by digital radiography (DR) and DES technology, and the difference of chest radiographs of DR, DES and combined groups were compared. Results The positive rate of superior chest radiographs in DR group was higher than that in DES group (72.9% vs 56.1%, P<0.05). The determination of shadow shape and size, total density and stage of pneumoconiosis on chest radiographs in DES group and combined group were consistent with those in DR group (all Kappa values >0.75, all P<0.01). However, the judgment of small shadow intensity, small shadow aggregation and large shadow distribution in the two groups were not superior to those in the DR group (Kappa value was 0.67, 0.74, both P<0.01). There was no significant difference between DES group and DR group in the determination of small shadow intensity, small shadow aggregation and large shadow distribution (P>0.05). However, there were statistically significant difference in this index between the combined group and DES group and DR group (all P<0.01). There was no statistically significant difference in shadow shape and size, total density and stage of pneumoconiosis in chest radiographs among these three groups (all P>0.05). Conclusion There was no difference between DES alone and DES combined with DR for the diagnosis of pneumoconiosis in terms of shadow shape and size, total density and stage of pneumoconiosis when compared with the gold standard DR. The value of DES in the diagnosis of pneumoconiosis needs further study.

Objective To analyze clinicopathological characteristics and the potential risk-related factors of female breast hyperplasia in different age groups.Method From Jan 2015 to Dec 2016,patients diagnosed with breast hyperplasia in 12 hospitals were evaluated.All patients completed the self-designed questionnaires on women'health,including basic demographic information,clinic examination information,radiologic information and pathologic results.The patients were divided into a young group (＜ 45 years old) and an elderly group (from 45 to 75 years old).Results There were 3 684 cases of breast hyperplasia,including 2 291 cases in young group and 1 393 cases in elder group,respectively Clinically breast pain type were most commonly observed in both young and older groups (50.3％ vs.42.7％,P ＜ 0.001).While pathological research based on biopsy showed that breast adenopathy were the most common changes in both groups (67.9％ vs.61.7％,P ＜0.001).More breast cancer cases were identified in elder group than that in young group,especially in clinically lump type patients (9.4％ vs.4.2％,P ＜ 0.001).Compared with elder group,patients in young group have different distribution characteristics regarding to fertility factors,lifestyle factors and psychology scale including anxiety and depression.Conclusion Distributions of clinicopathological characteristics and risk factors of female breast hyperplasia differ across different age groups.

Objective To investigate the short?term functional connectivity (FC) changes of default mode network (DMN) after liver transplantation (LT) by using seed?based functional connectivity analysis of resting?state functional MRI (rs?fMRI). Methods Eighteen cirrhotic patients as transplant candidates and 20 healthy controls were included in this study. All the patients underwent rs?fMRI examination before and one month after LT. The data were analyzed using DPARSF and REST software. Seed?based functional connectivity analysis was used to isolate the DMN. The posterior cingulate cortex (PCC) was chosen as seed region for the DMN map. Maps of the DMN were compared among the groups. Values of Z reflecting the functional connectivity of 3 groups were obtained. Two?sample t?test was performed to explore the DMN difference between cirrhotic patients and controls, and we used paired t tests to examine for any differences in functional connectivity before and after LT. Pearson correlation analysis was performed to explore the relationship between the changes of functional connectivity with that of clinical indexes and neuropsychological test scores pre?and post?LT. Results Typically spatial distributions of the DMN were found in all the groups. According to paired t test, post?LT patients showed increased FC in left medial prefrontal cortex (t=3.31, P<0.05), while decreased FC in left precuneus and left lateral temporal cortex (t=-3.37,-4.53,respectively, both P<0.05). Compared to healthy controls, pre?LT patients showed decreased FC in the right precuneus, bilateral medial prefrontal cortex, bilateral inferior parietal cortex, and bilateral lateral temporal cortex(t=-4.32—3.00, all P<0.05), and post?LT patients showed decreased FC in right precuneus, left posterior cingulate cortex, bilateral inferior parietal cortex, and bilateral lateral temporal cortex(t=-4.33—2.99,all P<0.05). Pearson correlation analysis revealed positively correlation between the changes of functional connectivity in left medial prefrontal cortex with that of DST (r=0.543, P=0.02). Conclusions This study found that the DMN FC of post?LT patients still lower than that of healthy controls one month after LT. The left medial prefrontal cortex was the first brain region that showed increased FC, while the FC of some regions continued to decrease, suggesting that brain function reorganization can continue after LT. Rs?fMRI can be used to observe the DMN changes in post?LT patients.

Background and objective All the advanced NSCLC patients that received EGFR-TKI therapy will eventually relapse after a period of efficacy. The aim of this study is to investigate the serum biomarkers as potential predictive factors for the efficacy of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) targeted therapy in advanced non-small cell lung cancer. Methods Twenty serf-paired serum samples were collected from 9 advanced NSCLC patients that evaluated as disease control (SD or PR) after gefinitib therapy, at the time points of before and after gefinitib treatment but 2 weeks before being evaluated as disease progress. AII samples were pre-separated by WCX microbeads, and then detected on the MALDI-TOF-MS platform of Bruker AutoflexTM. ClinProTools (Version: 2.1) was used to analyze the differentially expressed proteins. Results There were 7 protein peaks (m/z), 3 242.09, 8 690.36, 2 952.64, 3 224.04, 1 450.51, 1 887.8 and 3 935.73 found statistically differentially expressed between the self-paired samples. Three proteins (3 242.09, 2 952.64 and 3 224.04) were down-regulated and four proteins (8 690.36, 1 450.51, 1 887.8 and 3 935.73) up-regulated in gefinitib treated sera. Conclusion The data here suggest that several specific protein peaks might indicate gefinitib resistance, yet the identities of these proteins and the mechanisms underlying the responsiveness to gefinitib treatment need further investigation.

BACKGROUNDDendritic cells (DCs) are the unique antigen-presenting cells that can activate naive T lymphocytes. This function is critical for inducing specific immune response. DCs-based vaccines have been used broadly in immunotherapy for many carcinomas. Constructing vaccines by transfecting total tumor RNA into DCs can be done with a few tumor tissues and need not to identify tumor antigens, so it is especially suitable for lung cancer which lacks tumor-specific antigens but has great heterogenicity and weak immunogenicity. Currently, the best transfection stage and method are still indefinite. So, the objective of this study is to explore the best condition of transfecting total RNA extracted from lung cancer tissues into DCs.

METHODSTen patients with lung cancer were enrolled whose tumor tissues were CEA and MUC1 positive in immunohistochemical staining. Total tumor RNA were extracted by one-step method. Then DCs and T cells were separated and cultured from peripheral blood monocytes and the RNA was transfected into the DCs in different stages with different methods. CEA and MUC1 expression in the transfected DCs were measured by flow cytometry analysis and T cells' proliferation was examined by mixed lymphocyte reaction (MLR).

RESULTSThe expression of CEA and MUC1 protein in immature DCs (11.33±2.64, 39.68±7.25) was remarkably higher than that in mature DCs (5.46±1.63, 27.17±4.16) after transfection with total RNA of lung cancer tissues (P < 0.01), and the DCs presented more powerful effects on T cell proliferation. The CEA and MUC1 expression on DCs were significantly higher in electroporation transfection group (20.53±3.64, 65.39± 9.33) than that in lipofection group (11.33±2.64, 39.68±7.25) and passive pulsing transfection group ( 0.91±0.27,18.53±3.26)(P < 0.01), and the DCs in electroporation transfection group presented more powerful effects on stimulating T cell proliferation than the other two groups did.

CONCLUSIONSTransfecting total tumor RNA into immature DCs by using electroporation is a good way to construct DCs-based vaccines for lung cancer and to achieve a higher activity to stimulate T cell proliferation.

BACKGROUNDTo explore the experience of gefitinib molecular target therapy for Chinese patients with non-small cell lung cancer (NSCLC).

METHODSThe unpublished data of gefitinib for advanced NSCLC in 7 hospitals were collected. The detailed data from Guangdong Provincial People's Hospital were analyzed.

RESULTSA total of 282 patients with advanced NSCLC was treated with gefitinib from July 2001 to December 2003. Response rate was 22.2%-47.7%, disease control rate 62.6%-81.8%. No severe side effects were surveyed.

CONCLUSIONSGefitinib can be used safely and effectively in Chinese patients with advanced NSCLC.