Machine learning for identifying tumor stemness genes and developing prognostic model in gastric cancer.

Gastric cancer presents a formidable challenge, marked by its debilitating nature and often dire prognosis. Emerging evidence underscores the pivotal role of tumor stem cells in exacerbating treatment resistance and fueling disease recurrence in gastric cancer. Thus, the identification of genes contributing to tumor stemness assumes paramount importance. Employing a comprehensive approach encompassing ssGSEA, WGCNA, and various machine learning algorithms, this study endeavors to delineate tumor stemness key genes (TSKGs). Subsequently, these genes were harnessed to construct a prognostic model, termed the Tumor Stemness Risk Genes Prognostic Model (TSRGPM). Through PCA, Cox regression analysis and ROC curve analysis, the efficacy of Tumor Stemness Risk Scores (TSRS) in stratifying patient risk profiles was underscored, affirming its ability as an independent prognostic indicator. Notably, the TSRS exhibited a significant correlation with lymph node metastasis in gastric cancer. Furthermore, leveraging algorithms such as CIBERSORT to dissect immune infiltration patterns revealed a notable association between TSRS and monocytes and other cell. Subsequent scrutiny of tumor stemness risk genes (TSRGs) culminated in the identification of CDC25A for detailed investigation. Bioinformatics analyses unveil CDC25A's implication in driving the malignant phenotype of tumors, with a discernible impact on cell proliferation and DNA replication in gastric cancer. Noteworthy validation through in vitro experiments corroborated the bioinformatics findings, elucidating the pivotal role of CDC25A expression in modulating tumor stemness in gastric cancer. In summation, the established and validated TSRGPM holds promise in prognostication and delineation of potential therapeutic targets, thus heralding a pivotal stride towards personalized management of this malignancy.

CELSR1 variants are associated with partial epilepsy of childhood.

CELSR1 gene, encoding cadherin EGF LAG seven-pass G-type receptor 1, is mainly expressed in neural stem cells during the embryonic period. It plays an important role in neurodevelopment. However, the relationship between CELSR1 and disease of the central nervous system has not been defined. In this study, we performed trios-based whole-exome sequencing in a cohort of 356 unrelated cases with partial epilepsy without acquired causes and identified CELSR1 variants in six unrelated cases. The variants included one de novo heterozygous nonsense variant, one de novo heterozygous missense variant, and four compound heterozygous missense variants that had one variant was located in the extracellular region and the other in the cytoplasm. The patients with biallelic variants presented severe epileptic phenotypes, whereas those with heterozygous variants were associated with a mild epileptic phenotype of benign epilepsy with centrotemporal spikes (BECTS). These variants had no or low allele frequency in the gnomAD database. The frequencies of the CELSR1 variants in this cohort were significantly higher than those in the control populations. The evidence from ClinGen Clinical-Validity Framework suggested a strong association between CELSR1 variants and epilepsy. These findings provide evidence that CELSR1 is potentially a candidate pathogenic gene of partial epilepsy of childhood.

Machine learning models for predicting one-year survival in patients with metastatic gastric cancer who experienced upfront radical gastrectomy.

Tumor metastasis is a common event in patients with gastric cancer (GC) who previously underwent curative gastrectomy. It is meaningful to employ high-volume clinical data for predicting the survival of metastatic GC patients. We aim to establish an improved machine learning (ML) classifier for predicting if a patient with metastatic GC would die within 12 months. Eligible patients were enrolled from a Chinese GC cohort, and the complete detailed information from medical records was extracted to generate a high-dimensional dataset. Appropriate feature engineering and feature filter were conducted before modeling with eight algorithms. A 10-fold cross validation (CV) nested in a holdout CV (8:2) was employed for hyperparameter tuning and model evaluation. Model selection was based on the area under the receiver operating characteristic (AUROC) curve, recall, and precision. The selected model was globally explained using interpretable surrogate models. Of the total 399 cases (median survival of 8.2 months), 242 patients survived less than 12 months. The linear discriminant analysis (LDA), support vector machine (SVM), and random forest (RF) model had the highest AUROC (0.78 ± 0.021), recall (0.93 ± 0.031), and precision (0.80 ± 0.026), respectively. The LDA model created a new function that generally separated the two classes. The predicted probability of the SVM model was interpreted using a linear regression model visualized by a nomogram. The predicted class of the RF model was explained using a decision tree model. In summary, analyzing high-volume medical data by ML is helpful to produce an improved model for predicting the survival in patients with metastatic GC. The algorithm should be carefully selected in different practical scenarios.

Effects of targeted-edited oncogenic insulin-like growth factor-1 receptor with specific-sgRNA on biological behaviors of HepG2 cells.

BACKGROUND: Insulin-like growth factor-1 receptor (IGF-1R) is over-expressed in hepatocellular carcinoma (HCC). However, the relationship between IGF-1R activation and HCC progression remains unidentified. AIM: To investigate the effects of editing IGF-1R on the biological features of HCC cells. METHODS: Immunohistochemistry analyzed the expressions of IGF-1R and P-glyco protein (P-gp) in HCC tissues and their distal non-cancerous tissues (non-Ca). IGF-1R was edited with Crispr/Cas9 system, screened specific sgRNAs, and then transfected into HepG2 cells. CCK-8, scratch wound test detected cell proliferation, migration, invasion and transwell assays, respectively. Alterations of IGF-1R and P-gp were confirmed by Western blotting. Alterations of anti-cancer drug IC50 values were analyzed at the cell level. RESULTS: The positive rates of IGF-1R (93.6%, χ 2 = 63.947) or P-gp (88.2%, χ2 = 58.448) were significantly higher (P < 0.001) in the HCC group than those (36.6% in IGF-1R or 26.9% in P-gp) in the non-Ca group. They were positively correlated between high IGF-1R and P-gp expression, and they were associated with hepatitis B virus infection and vascular invasion of HCC. Abnormal expressions of circulating IGF-1R and P-gp were confirmed and associated with HCC progression. Biological feature alterations of HCC cells transfected with specific sgRNA showed IGF-1R expression down-regulation, cell proliferation inhibition, cell invasion or migration potential decreasing, and enhancing susceptibility of HepG2 cells to anti-cancer drugs. CONCLUSION: Edited oncogenic IGF-1R was useful to inhibit biological behaviors of HepG2 cells.

[Characteristics of PM2.5 Pollution in Beijing During the Historical Period of the 2022 Olympic Winter Games].

Based on the dataset derived from January to March between 2015 and 2021 in Beijing, the PM2.5 pollution characteristics and its potential source regions during the historical period of the Beijing 2022 Olympic Winter Games and Paralympic Winter Games were investigated. From 2015 to 2018, both the number of severely polluted days (daily average ρ(PM2.5)>75 µg·m-3) and the average PM2.5 concentrations during severe pollution episodes decreased significantly in the period of January to March. While, neither variable has changed obviously since 2018. On average, severely polluted days occurred 23 times in each year between 2018 and 2021 during the period of January to March, and the average of ρ(PM2.5) was approximately 120.0 µg·m-3 during such polluted days. From January to March in 2015-2021, the severely polluted event with more than 5 consecutive polluted days occurred 2-3 times in each year, and the severest one lasted 8 d. During the historical period of the Beijing 2022 Olympic Winter Games, severely polluted days took place 2-9 d every year. The large quantities of fireworks during the Spring Festival maybe one of important primary sources of the PM2.5. The number of severely polluted days during the historical period of the Paralympic Winter Games ranged from 1 to 5 d, except for 2021 with 9 d owing to the frequent stagnant weather condition. The PM2.5 chemical composition was dominated by secondary species on severely polluted days during the historical period of the Beijing 2022 Olympic Winter Games and Paralympic Winter Games. Nitrate accounted for 46% of the measurable chemical components of PM2.5 during severe pollution events in 2020, which was remarkably higher than that during clean days in the same year (11%). The mass fraction of SO42- ranged from 12% to 19% in 2018-2020, indicating that the contribution of sulfate was much less, but cannot be ignored. The main potential source regions of PM2.5 in Beijing during the period concerned in this study were central and western Inner Mongolia, Hebei Province, Tianjin City, Shanxi Province, Shaanxi Province, central and western Shandong Province, and northern Henan Province.

Exosomal microRNA-15a from mesenchymal stem cells impedes hepatocellular carcinoma progression via downregulation of SALL4.

Hepatocellular carcinoma (HCC) is a heterogeneous tumor with an increased incidence worldwide accompanied by high mortality and dismal prognosis. Emerging evidence indicates that mesenchymal stem cells (MSCs)-derived exosomes possess protective effects against various human diseases by transporting microRNAs (miRNAs or miRs). We aimed to explore the role of exosomal miR-15a derived from MSCs and its related mechanisms in HCC. Exosomes were isolated from transduced MSCs and co-incubated with Hep3B and Huh7 cells. miR-15a expression was examined by RT-qPCR in HCC cells, MSCs, and secreted exosomes. CCK-8, transwell, and flow cytometry were used to detect the effects of miR-15a or spalt-like transcription factor 4 (SALL4) on cell proliferative, migrating, invasive, and apoptotic properties. A dual-luciferase reporter gene assay was performed to validate the predicted targeting relationship of miR-15a with SALL4. Finally, in vivo experiments in nude mice were implemented to assess the impact of exosome-delivered miR-15a on HCC. The exosomes from MSCs restrained HCC cell proliferative, migrating, and invasive potentials, and accelerated their apoptosis. miR-15a was expressed at low levels in HCC cells and could bind to SALL4, thus curtailing the proliferative, migrating, and invasive abilities of HCC cells. Exosomes successfully delivered miR-15a to HCC cells. Exosomal miR-15a depressed tumorigenicity and metastasis of HCC tumors in vivo. Overall, exosomal miR-15a from MSCs can downregulate SALL4 expression and thereby retard HCC development.

MicroRNA-499 serves as a sensitizer for lung cancer cells to radiotherapy by inhibition of CK2α-mediated phosphorylation of p65.

The present study aimed to define the tumor-suppressive role of microRNA-499 (miR-499) in lung cancer cells and its underlying mechanism. First, qRT-PCR analysis revealed poor expression of miR-499 in clinical samples and cell lines of lung cancer. Next, we performed loss- and gain-of-function experiments for the expression of miR-499 in lung cancer cells exposed to irradiation (IR) to determine the effect of miR-499 expression on cell viability and apoptosis as well as tumor growth. Results showed that overexpression of miR-499 inhibited cell viability, enhanced the radiosensitivity of lung cancer cells, and promoted cell apoptosis under IR. Furthermore, CK2α was verified to be a target of miR-499, and miR-499 was identified to repress p65 phosphorylation by downregulating CK2α expression, which ultimately diminished the survival rate of lung cancer cells under IR. Collectively, the key findings of the study illustrate the tumor-inhibiting function of miR-499 and confirmed that miR-499-mediated CK2α inhibition and altered p65 phosphorylation enhances the sensitivity of lung cancer cells to IR.

M2 macrophage-derived exosomal microRNA-155-5p promotes the immune escape of colon cancer by downregulating ZC3H12B.

Previous evidence has highlighted M2 macrophage regulation of cancer cells via exosome shuttling of microRNAs (miRNAs or miRs). The current study set out to explore the possible role of M2 macrophage-derived exosomal miR-155-5p in regard to immune escape of colon cancer cells. Experimental data from quantitative reverse-transcriptase PCR (qRT-PCR) and western blot analysis revealed highly expressed miR-155-5p and interleukin (IL)-6 and poorly expressed ZC3H12B in M2 macrophage-derived exosomes. Additionally, miR-155-5p could be transferred by M2 macrophage-isolated exosomes to colon cancer cells, which targeted ZC3H12B by binding to the 3¢ UTR, as identified by dual luciferase reporter gene. Meanwhile, gain- and loss-of function experimentation on miR-155-5p and ZC3H12B in SW48 and HT29 cells cocultured with M2 macrophage-secreted exosomes demonstrated that miR-155-5p overexpression or ZC3H12B silencing promoted the proliferation and antiapoptosis ability of SW48 and HT29 cells, as well as augmenting the CD3+ T cell proliferation and the proportion of interferon (IFN)-γ+ T cells. Xenograft models confirmed that M2 macrophage-derived exosomal miR-155-5p reduced the ZC3H12B expression to upregulate IL-6, which consequently induced immune escape and tumor formation. Collectively, our findings indicated that M2 macrophage-derived exosomal miR-155-5p can potentially promote the immune escape of colon cancer by impairing ZC3H12B-mediated IL-6 stability reduction, thereby promoting the occurrence and development of colon cancer.

Cancer-associated fibroblast-derived exosomal microRNA-24-3p enhances colon cancer cell resistance to MTX by down-regulating CDX2/HEPH axis.

MicroRNA-24-3p (miR-24-3p) has been implicated as a key promoter of chemotherapy resistance in numerous cancers. Meanwhile, cancer-associated fibroblasts (CAFs) can secret exosomes to transfer miRNAs, which mediate tumour development. However, little is known regarding the molecular mechanism of CAF-derived exosomal miR-24-3p in colon cancer (CC). Hence, this study intended to characterize the functional relevance of CAF-derived exosomal miR-24-3p in CC cell resistance to methotrexate (MTX). We identified differentially expressed HEPH, CDX2 and miR-24-3p in CC through bioinformatics analyses, and validated their expression in CC tissues and cells. The relationship among HEPH, CDX2 and miR-24-3p was verified using ChIP and dual-luciferase reporter gene assays. Exosomes were isolated from miR-24-3p inhibitor-treated CAFs (CAFs-exo/miR-24-3p inhibitor), which were used in combination with gain-of-function and loss-of-function experiments and MTX treatment. CCK-8, flow cytometry and colony formation assays were conducted to determine cell viability, apoptosis and colony formation, respectively. Based on the findings, CC tissues and cells presented with high expression of miR-24-3p and low expression of HEPH and CDX2. CDX2 was a target gene of miR-24-3p and could up-regulate HEPH. Under MTX treatment, overexpressed CDX2 or HEPH and down-regulated miR-24-3p reduced cell viability and colony formation and elevated cell apoptosis. Furthermore, miR-24-3p was transferred into CC cells via CAF-derived exosomes. CAF-derived exosomal miR-24-3p inhibitor diminished cell viability and colony formation and increased cell apoptosis in vitro and inhibited tumour growth in vivo under MTX treatment. Altogether, CAF-derived exosomal miR-24-3p accelerated resistance of CC cells to MTX by down-regulating CDX2/HEPH axis.

KDM5A silencing transcriptionally suppresses the FXYD3-PI3K/AKT axis to inhibit angiogenesis in hepatocellular cancer via miR-433 up-regulation.

Hepatocellular cancer (HCC) has been reported to belong to one of the highly vascularized solid tumours accompanied with angiogenesis of human umbilical vein endothelial cells (HUVECs). KDM5A, an attractive drug target, plays a critical role in diverse physiological processes. Thus, this study aims to investigate its role in angiogenesis and underlying mechanisms in HCC. ChIP-qPCR was utilized to validate enrichment of H3K4me3 and KDM5A on the promotor region of miR-433, while dual luciferase assay was carried out to confirm the targeting relationship between miR-433 and FXYD3. Scratch assay, transwell assay, Edu assay, pseudo-tube formation assay and mice with xenografted tumours were conducted to investigate the physiological function of KDM5A-miR-433-FXYD3-PI3K-AKT axis in the progression of HCC after loss- and gain-function assays. KDM5A p-p85 and p-AKT were highly expressed but miR-433 was down-regulated in HCC tissues and cell lines. Depletion of KDM5A led to reduced migrative, invasive and proliferative capacities in HCC cells, including growth and a lowered HUVEC angiogenic capacity in vitro. Furthermore, KDM5A suppressed the expression of miR-433 by demethylating H3K4me3 on its promoterregion. miR-433 negatively targeted FXYD3. Depleting miR-433 or re-expressing FXYD3 restores the reduced migrative, invasive and proliferative capacities, and lowers the HUVEC angiogenic capacity caused by silencing KDM5A. Therefore, KDM5A silencing significantly suppresses HCC tumorigenesis in vivo, accompanied with down-regulated miR-433 and up-regulated FXYD3-PI3K-AKT axis in tumour tissues. Lastly, KDM5A activates the FXYD3-PI3K-AKT axis to enhance angiogenesis in HCC by suppressing miR-433.

Whole-Transcriptome Sequencing Identifies Key Differentially Expressed mRNAs, miRNAs, lncRNAs, and circRNAs Associated with CHOL.

To systematically evaluate the whole-transcriptome sequencing data of cholangiocarcinoma (CHOL) to gain more insights into the transcriptomic landscape and molecular mechanism of this cancer, we performed whole-transcriptome sequencing based on the tumorous (C) and their corresponding non-tumorous adjacent to the tumors (CP) from eight CHOL patients. Subsequently, differential expression analysis was performed on the C and CP groups, followed by functional interaction prediction analysis to investigate gene-regulatory circuits in CHOL. In addition, The Cancer Genome Atlas (TCGA) for CHOL data was used to validate the results. In total, 2,895 differentially expressed messenger RNAs (dif-mRNAs), 56 differentially expressed microRNAs (dif-miRNAs), 151 differentially expressed long non-coding RNAs (dif-lncRNAs), and 110 differentially expressed circular RNAs (dif-circRNAs) were found in CHOL samples compared with controls. Enrichment analysis on those differentially expressed genes (DEGs) related to miRNA, lncRNA, and circRNA also identified the function of spliceosome. The downregulated hsa-miR-144-3p were significantly enriched in the competing endogenous RNA (ceRNA) complex network, which also included 7 upregulated and 13 downregulated circRNAs, 7 upregulated lncRNAs, and 90 upregulated and 40 downregulated mRNAs. Moreover, most of the DEGs and a few of the miRNAs (such as hsa-miR-144-3p) were successfully validated by TCGA data. The genes involved in RNA splicing and protein degradation processes and miR-144-3p may play fundamental roles in the pathogenesis of CHOL.

Prognosis for intrahepatic cholangiocarcinoma patients treated with postoperative adjuvant transcatheter hepatic artery chemoembolization.

OBJECTIVE: We used meta-analysis to evaluate the efficacy of transcatheter hepatic arterial chemoembolization (TACE) for the treatment of intrahepatic cholangiocarcinoma (ICC). METHODS: We performed the meta-analysis using the R 3.12 software and the quality evaluation of data using the Newcastle-Ottawa Scale. The main outcomes were recorded as 1-year overall survival (OS), 3-year OS, 5-year OS, and hazard ratio (HR) of TACE treatment or non-TACE treatment. The heterogeneity test was performed using the Q-test based on chi-square and I2 statistics. Egger's test was used to test the publication bias. The odds ratio or HR and 95% confidence interval (CI) were used to represent the effect index. RESULTS: Nine controlled clinical trials involving 1724 participants were included in this study; patients came mainly from China, Italy, South Korea, and Germany. In the OS meta-analysis, the 1-year and 3-year OS showed significant heterogeneity, but not the 5-year OS. TACE increased the 1-year OS (odds ratio = 2.66, 95% CI: 1.10-6.46) of the patients with ICC, but the 3- and 5-year OS rates were not significantly increased. The results had no publication bias, but the stability was weak. The HR had significant heterogeneity (I2 = 0%, P= 0.54). TACE significantly decreased the HR of ICC patients (HR = 0.59, 95% CI: 0.48-0.73). The results had no publication bias, and the stability was good. CONCLUSIONS: Treatment with TACE is effective for patients with ICC. Regular updating and further research and analysis still need to be carried out.

Cumulative Score Based on Preoperative Fibrinogen and Pre-albumin Could Predict Long-term Survival for Patients with Resectable Gastric Cancer.

Background: To investigate the prognostic significance of the cumulative score based on preoperative fibrinogen and pre-albumin (FP score) in patients with gastric cancer after radical gastrectomy. Methods: Baseline characteristics, preoperative fibrinogen and pre-albumin levels were retrospectively reviewed in patients who underwent radical gastrectomy. The optimal cut-off values for fibrinogen and pre-albumin were defined as 4.0 g/L and 230.0 mg/L, respectively. Patients with elevated fibrinogen (≥ 4.0 g/L) and decreased pre-albumin (< 230.0 mg/L) levels were allocated an FP score of 2, those with only one of these two abnormalities were assigned a score of 1, and those with neither of the two abnormalities were allocated a score of 0. The prognostic value was examined by univariate and multivariate regression analyses. Results: The preoperative FP score was significantly correlated with age, tumor size, fibrinogen level, pre-albumin level and white blood cell count. No significant differences based on sex, tumor location, degree of differentiation, depth of invasion, lymph node status, tumor-node-metastasis (TNM) stage or adjuvant chemotherapy were identified between the groups. In addition, univariate survival analysis revealed that a high preoperative FP score was significantly associated with unfavorable disease-free survival (DFS) [hazard ratio (HR), 1.482; 95% confidence interval (CI), 1.222-1.796; P < 0.001] and overall survival (OS) (HR, 1.623; 95% CI, 1.315-2.002; P < 0.001). Moreover, after adjusting for other factors, a high preoperative FP score remained an independent predictor for impaired DFS (HR, 1.434; 95% CI, 1.177-1.747; P < 0.001) and OS (HR, 1.413; 95% CI, 1.136-1.758; P = 0.002) in multivariate Cox regression analysis. Conclusions: The preoperative FP score significantly predicts long-term survival for gastric cancer patients who have undergone radical gastrectomy.

Nomograms for predicting survival in patients with metastatic gastric adenocarcinoma who undergo palliative gastrectomy.

BACKGROUND: Recently, evidence has emerged that palliative gastrectomy in patients with stage IV gastric cancer may offer some survival benefits. However, the decision whether to perform primary tumor surgery remains challenging for surgeons, and investigations into models that are predictive of prognosis are scarce. Current study aimed to develop and validate prognostic nomograms for patients with metastatic gastric adenocarcinoma treated with palliative gastrectomy. METHODS: The development dataset comprised 1186 patients from the Surveillance, Epidemiology, and End Results Program who were diagnosed with metastatic gastric adenocarcinoma in 2004-2011, while the validation dataset included 407 patients diagnosed in 2012-2015. Variables were incorporated into a Cox proportional hazards model to identify independent risk factors for survival. Both pre- and postoperative nomograms for predicting 1- or 2-year survival probabilities were constructed using the development dataset. The concordance index (c-index) and calibration curves were plotted to determine the accuracy of the nomogram models. Finally, the cut-off value of the calculated total scores based on preoperative nomograms was set and validated by comparing survival with contemporary cases without primary tumor surgery. RESULTS: Age, tumor size, location, grade, T stage, N stage, metastatic site, scope of gastrectomy, number of examined lymph node(s), chemotherapy and radiotherapy were risk factors of survival and were included as variables in the postoperative nomogram; the c-indices of the development and validation datasets were 0.701 (95% confidence interval [CI]: 0.693-0.710) and 0.699 (95% CI: 0.682-0.716), respectively. The preoperative nomogram incorporated age, tumor size, location, grade, depth of invasion, regional lymph node(s) status, and metastatic site. The c-indices for the internal (bootstrap) and external validation sets were 0.629 (95% CI: 0.620-0.639) and 0.607 (95% CI: 0.588-0.626), respectively. Based on the preoperative nomogram, patients with preoperative total score > 28 showed no survival benefit with gastrectomy compared to no primary tumor surgery. CONCLUSIONS: Our survival nomograms for patients with metastatic gastric adenocarcinoma undergoing palliative gastrectomy can assist surgeons in treatment decision-making and prognostication.

Dual Regulatory Mechanisms of Expression and Mutation Involving Metabolism-Related Genes FDFT1 and UQCR5 during CLM.

Colorectal cancer (CRC) is the third most common cancer worldwide, and liver metastasis presents a major cause of CRC-associated death. Extensive genomic analysis has provided valuable insight into the pathogenesis and progression of CRC; however, a comprehensive proteogenomic characterization of CRC liver metastasis (CLM) has yet to be reported. Here, we analyzed the proteomes of 44 paired normal colorectal tissues and CRC tissues with or without liver metastasis, as well as analyzed genomics of CRC characterized previously by The Cancer Genome Atlas (TCGA) to conduct integrated proteogenomic analyses. We identified a total of 2,170 significantly deregulated proteins associated with CLM, 14.88% of which were involved in metabolic pathways. The mutated peptide number was found to have potential prognosis value, and somatic variants revealed two metabolism-related genes UQCR5 and FDFT1 that frequently mutated only in the liver metastatic cohort and displayed dysregulated protein abundance with biological function and clinical significance in CLM. Proteogenomic characterization and integrative and comparative genomic analysis provides functional context and prognostic value to annotate genomic abnormalities and affords a new paradigm for understanding human colon and rectal cancer liver metastasis.

Differentially Expressed Protein Are Involved in Dynamic Changes of Catechins Contents in Postharvest Tea Leaves under Different Temperatures.

In this study, isobaric tags for relative and absolute quantitation (iTRAQ) technology were used to investigate three samples from postharvest tea leaves that were treated at room temperature (25 °C, control group), high temperature (38 °C), and low temperature (4 °C) for 4 h. In heat and cold treatments, a total of 635 and 566 differentially expressed proteins (DEPs) were determined, respectively. DEPs were annotated to GO and KEGG databases, which revealed that DEPs involved in various aspects of biological process. Three catechins-related DEPs, CsCHI, CsF3H, and CsANR, were identified. Both catechins contents and the expression profiles of catechins biosynthesis-related genes changed significantly under different temperature treatments. The correlations between catechins contents, gene expression profiles, and DEPs were analyzed. This study provides potential new insights into the molecular basis for tea production of postharvest leaves and catechins content changes at diverse temperature conditions and will guide the improvement of tea-processing technology.

Three new sesquiterpenes from Ainsliaea glabra.

Three new sesquiterpenes: 4-acrylic-6-methyl-α-tetralone (1), ainsliaea acid A (2) and ainsliaea acid B (3), together with 8 known compounds (4-11) were isolated from the whole herb of Ainsliaea glabra and their structures were established by means of 1D and 2D NMR spectroscopy and HR-ESIMS. Compounds 1-6 were tested for the inhibition of nuclear factor kappa B (NF-κB) in the 293-NF-κB-luciferase reporter cell line induced by lipopolysaccharide (LPS), and compound 2 was further tested for the production of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), IL-6 and IL-10 in RAW264.7 macrophages induced by LPS. The isolated compound 2 exhibited significant anti-inflammatory activity.

Differentially expressed protein and gene analysis revealed the effects of temperature on changes in ascorbic acid metabolism in harvested tea leaves.

Tea is an important non-alcoholic beverage worldwide. Tea quality is determined by numerous secondary metabolites in harvested tea leaves, including tea polyphenols, theanine, caffeine, and ascorbic acid (AsA). AsA metabolism in harvested tea leaves is affected by storage and transportation temperature. However, the molecular mechanisms underlying AsA metabolism in harvested tea leaves exposed to different storage and transportation temperature conditions remain unclear. Here we performed RP-HPLC to detect dynamic changes in AsA content in tea leaves subjected to high- (38 °C), low- (4 °C), or room-temperature (25 °C) treatments. The AsA distribution and levels in the treated tea leaves were analyzed using cytological-anatomical characterization methods. The differentially expressed CsAPX1 and CsDHAR2 proteins, which are involved in the AsA recycling pathway, were identified from the corresponding proteomic data using iTRAQ. We also analyzed the expression profiles of 18 genes involved in AsA metabolism, including CsAPX1 and CsDHAR2. AsA was mainly distributed in tea leaf mesophyll cells. High- and low-temperature treatments upregulated the CsAPX1 and CsDHAR2 proteins and induced CsAPX and CsDHAR2 gene expression. These results indicated that the CsAPX1 and CsDHAR2 proteins might have critical roles in AsA recycling in tea leaves. Our results provide a foundation for the in-depth investigation of AsA metabolism in tea leaves during storage and transportation, and they will promote better tea flavor in tea production.

Tumor suppressive microRNA-124a inhibits stemness and enhances gefitinib sensitivity of non-small cell lung cancer cells by targeting ubiquitin-specific protease 14.

Increasing evidence has shown that microRNAs (miRNAs) play a significant functional role by directly regulating respective targets in cancer stem cell (CSC)-induced non-small cell lung cancer (NSCLC) progression and resistance to therapy. In this study, we found that hsa-miR-124a was downregulated during spheroid formation of the NSCLC cell lines SPC-A1 and NCI-H1650 and NSCLC tissues compared with normal lung cells and tissues. Patients with lower hsa-miR-124a expression had shorter overall survival (OS) and progression free survival (PFS). Moreover, ubiquitin-specific protease 14 (USP14) was confirmed to be a direct target of hsa-miR-124a. Furthermore, concomitant low hsa-miR-124a expression and high USP14 expression were correlated with a shorter median OS and PFS in NSCLC patients. Cellular functional analysis verified that the tumor suppressor hsa-miR-124a negatively regulated cell growth and self-renewal, and promoted apoptosis and gefitinib sensitivity of lung cancer stem cells by suppressing its target gene USP14. Our results provide the first evidence that USP14 is a direct target of hsa-miR-124a, and that hsa-miR-124a inhibits stemness and enhances the gefitinib sensitivity of NSCLC cells by targeting USP14. Thus, hsa-miR-124a and USP14 may be useful as tumor biomarkers for the diagnosis and treatment of NSCLC.

DRR1 promotes glioblastoma cell invasion and epithelial-mesenchymal transition via regulating AKT activation.

Metastatic invasion is the primary cause of treatment failure for GBM. EMT is one of the most important events in the invasion of GBM; therefore, understanding the molecular mechanisms of EMT is crucial for the treatment of GBM. In this study, high expression of DRR1 was identified to correlate with a shorter median overall and relapse-free survival. Loss-of-function assays using shDRR1 weakened the invasive potential of the GBM cell lines through regulation of EMT-markers. The expressions of p-AKT were significantly decreased after DRR-depletion in SHG44 and U373 cells. Moreover, the invasion was inhibited by the AKT inhibitor, MK-2206. The expression of Vimentin, N-cadherin, MMP-7, snail and slug was significantly inhibited by MK-2206, while the expression of E-cadherin was upregulated. Our results provide the first evidence that DRR1 is involved in GBM invasion and progression possibly through the induction of EMT activation by phosphorylation of AKT.