Metabolomics and proteomics insights into subacute ruminal acidosis etiology and inhibition of proliferation of yak rumen epithelial cells in vitro.

BACKGROUND: Untargeted metabolomics and proteomics were employed to investigate the intracellular response of yak rumen epithelial cells (YRECs) to conditions mimicking subacute rumen acidosis (SARA) etiology, including exposure to short-chain fatty acids (SCFA), low pH5.5 (Acid), and lipopolysaccharide (LPS) exposure for 24 h. RESULTS: These treatments significantly altered the cellular morphology of YRECs. Metabolomic analysis identified significant perturbations with SCFA, Acid and LPS treatment affecting 259, 245 and 196 metabolites (VIP > 1, P < 0.05, and fold change (FC) ≥ 1.5 or FC ≤ 0.667). Proteomic analysis revealed that treatment with SCFA, Acid, and LPS resulted in differential expression of 1251, 1396, and 242 proteins, respectively (FC ≥ 1.2 or ≤ 0.83, P < 0.05, FDR < 1%). Treatment with SCFA induced elevated levels of metabolites involved in purine metabolism, glutathione metabolism, and arginine biosynthesis, and dysregulated proteins associated with actin cytoskeleton organization and ribosome pathways. Furthermore, SCFA reduced the number, morphology, and functionality of mitochondria, leading to oxidative damage and inhibition of cell survival. Gene expression analysis revealed a decrease the genes expression of the cytoskeleton and cell cycle, while the genes expression associated with inflammation and autophagy increased (P < 0.05). Acid exposure altered metabolites related to purine metabolism, and affected proteins associated with complement and coagulation cascades and RNA degradation. Acid also leads to mitochondrial dysfunction, alterations in mitochondrial integrity, and reduced ATP generation. It also causes actin filaments to change from filamentous to punctate, affecting cellular cytoskeletal function, and increases inflammation-related molecules, indicating the promotion of inflammatory responses and cellular damage (P < 0.05). LPS treatment induced differential expression of proteins involved in the TNF signaling pathway and cytokine-cytokine receptor interaction, accompanied by alterations in metabolites associated with arachidonic acid metabolism and MAPK signaling (P < 0.05). The inflammatory response and activation of signaling pathways induced by LPS treatment were also confirmed through protein interaction network analysis. The integrated analysis reveals co-enrichment of proteins and metabolites in cellular signaling and metabolic pathways. CONCLUSIONS: In summary, this study contributes to a comprehensive understanding of the detrimental effects of SARA-associated factors on YRECs, elucidating their molecular mechanisms and providing potential therapeutic targets for mitigating SARA.

A radiomics model to predict γδ T-cell abundance and overall survival in head and neck squamous cell carcinoma.

γδ T cells are becoming increasingly popular because of their attractive potential for antitumor immunotherapy. However, the role and assessment of γδ T cells in head and neck squamous cell carcinoma (HNSCC) are not well understood. We aimed to explore the prognostic value of γδ T cell and predict its abundance using a radiomics model. Computer tomography images with corresponding gene expression data and clinicopathological data were obtained from online databases. After outlining the volumes of interest manually, the radiomic features were screened using maximum melevance minimum redundancy and recursive feature elimination algorithms. A radiomics model was developed to predict γδ T-cell abundance using gradient boosting machine. Kaplan-Meier survival curves and univariate and multivariate Cox regression analyses were used for the survival analysis. In this study, we confirmed that γδ T-cell abundance was an independent predictor of favorable overall survival (OS) in patients with HNSCC. Moreover, a radiomics model was built to predict the γδ T-cell abundance level (the areas under the operating characteristic curves of 0.847 and 0.798 in the training and validation sets, respectively). The calibration and decision curves analysis demonstrated the fitness of the model. The high radiomic score was an independent protective factor for OS. Our results indicated that γδ T-cell abundance was a promising prognostic predictor in HNSCC, and the radiomics model could discriminate its abundance levels and predict OS. The noninvasive radiomics model provided a potentially powerful prediction tool to aid clinical judgment and antitumor immunotherapy.

Establishment of SV40 Large T-Antigen-Immortalized Yak Rumen Fibroblast Cell Line and the Fibroblast Responses to Lipopolysaccharide.

The yak lives in harsh alpine environments and the rumen plays a crucial role in the digestive system. Rumen-associated cells have unique adaptations and functions. The yak rumen fibroblast cell line (SV40T-YFB) was immortalized by introducing simian virus 40 large T antigen (SV40T) by lentivirus-mediated transfection. Further, we have reported the effects of lipopolysaccharide (LPS) of different concentrations on cell proliferation, extracellular matrix (ECM), and proinflammatory mediators in SV40T-YFB. The results showed that the immortalized yak rumen fibroblast cell lines were identified as fibroblasts that presented oval nuclei, a fusiform shape, and positive vimentin and SV40T staining after stable passage. Chromosome karyotype analysis showed diploid characteristics of yak (n = 60). LPS at different concentrations inhibited cell viability in a dose-dependent manner. SV40T-YFB treated with LPS increased mRNA expression levels of matrix metalloproteinases (MMP-2 and MMP-9), inflammatory cytokines (TNF-α, IL-1ß, IL-6), and urokinase-type plasminogen activator system components (uPA, uPAR). LPS inhibits the expression of tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2), plasminogen activator inhibitor-2 (PAI-2), fibronectin (FN), anti-inflammatory factor IL-10, and collagen I (COL I) in SV40T-YFB. Overall, these results suggest that LPS inhibits cell proliferation and induces ECM degradation and inflammatory response in SV40T-YFB.

Glucose Regulates Glucose Transport and Metabolism via mTOR Signaling Pathway in Bovine Placental Trophoblast Cells.

It has been confirmed that improving the energy level of the diet contributed to the greater reproductive performance and birth weight of calves in periparturient dairy cows. To investigate the effect of glucose on nutrient transport during fetal development, the bovine placental trophoblast cells (BPTCs) were cultured in media with different glucose concentrations (1, 2, 4, 8, or 16 mg/mL). Subsequently, the BPTCs were cultured in media with 1, 8 mg/mL glucose and 8 mg/mL glucose plus 100 nmol/L rapamycin (the inhibitor of mTOR pathway). Compared with the 1 mg/mL glucose, the addition of 8 mg/mL glucose stimulated cell proliferation, upregulated the mRNA abundance of the glucose transporter GLUT1 and GLUT4, and increased the activity of glucose metabolism-related enzyme glucose-6-phosphate dehydrogenease (G6PD), lactate dehydrogenase (LDHA) and phosphoglycerate kinase 1 (PGK1), as well as adenosine-triphosphate (ATP) content (p < 0.05).Furthermore, compared with the treatment of 1 mg/mL glucose, adding 8 mg/mL of glucose-upregulated gene expression in the mTOR signaling pathway, including phosphatidylinositol3-kinase (PI3K), protein kinase B (Akt), mammalian target of rapamycin (mTOR) and 70 kDa ribosomal protein S6 kinase 2 (P70S6K) (p < 0.05).The supplementation of rapamycin downregulated the gene and protein expression of the mTOR signaling pathway, including mTOR, P70S6K, EIF4E-binding protein 1 (4EBP1), hypoxia-inducible factor 1-alpha (HIF-1α) and gene expression of glucose transporter upregulated by 8 mg/mL glucose (p < 0.05). Thus, these results indicated that the addition of 8 mg/mL glucose regulated the glucose transport and metabolism in BPTCs through the mTOR signaling pathway, thereby promoting the supply of nutrients to fetus.

Establishment of Immortalized Yak Ruminal Epithelial Cell Lines by Lentivirus-Mediated SV40T and hTERT Gene Transduction.

Yak is a unique species of cattle that is adapted to the harsh natural environment of the Qinghai-Tibet Plateau. Research on the function of the yak rumen is limited to animal experiments, and the cell molecular mechanism is very limited. The high cost of isolation and culture of adult yak rumen epithelial cells (YRECs), low success rate, and limited cell life limit the scope of long-term physiological functions and nutrient absorption mechanisms of yak rumen epithelium in vitro studies. This study aimed to explore the isolation and immortal culture methods of primary YRECs and establish a new cell line model for studying cell molecular mechanisms. The human telomerase reverse transcriptase gene (hTERT) and simian virus 40 large T antigen (SV40T) were transferred into primary YDECs using mammalian gene expression lentiviral vectors. The immortalized cell line (SV40T-YREC-hTERT) retains the morphological and functional characteristics of primary cells. The epithelial cell marker protein cytokeratin 18 of the immortalized cell lines was positive, and the cell proliferation and karyotype were normal. The SV40T and hTERT genes were successfully transferred into immortalized cell lines and maintained high expression. Simultaneously, the immortalized cell lines had normal function of short-chain fatty acid (SCFA) transport and absorption, and the immortalized yak rumen epithelial cell lines were successfully established. In addition, the transepithelial electrical resistance value gradually increased with culture time, and the permeability of epithelial cells decreased by culturing epithelial cells in Transwell culture chambers. Transmission electron microscopy demonstrated the submicroscopic structure of cells in the integrity barrier model and established the YREC barrier model in vitro.

microRNA-20b-5p overexpression combing Pembrolizumab potentiates cancer cells to radiation therapy via repressing programmed death-ligand 1.

Radiation therapy (RT) is widely applied in cancer treatment. The sensitivity of tumor cells to RT is the key to the treatment. This study probes the role and mechanism of miR-20b-5p in Pembrolizumab's affecting the radiosensitivity of tumor cells. After Pembrolizumab treatment or cell transfection (miR-20b-5p mimics and miR-20b-5p inhibitors), tumor cells (NCI-H460 and ZR-75-30) were exposed to RT. The sensitivity of NCI-H460 and ZR-75-30 to RT was evaluated by monitoring cell proliferation and apoptosis. The dual-luciferase reporter assay and RNA immunoprecipitation (RIP) were adopted to evaluate the binding relationship between miR-20b-5p and CD274 (PD-L1). The xenograft model was established in nude mice to examine the mechanism of action of Pembrolizumab in vivo. Our outcomes exhibited that either Pembrolizumab treatment or miR-20b-5p overexpression potentiated radiosensitivity of tumor cells. Overexpressing miR-20b-5p enhanced radiosensitization of Pembrolizumab in vivo and in vitro by targeting PD-L1 and inactivating PD-L1/PD1. Overall, miR-20b-5p overexpression combined with Pembrolizumab potentiated cancer cells' sensitivity to RT by repressing PD-L1/PD1.Abbreviations Akt: serine/threonine kinase 1; cDNA: complementary DNA; CO2: carbon dioxide; EDTA: Ethylene Diamine Tetraacetic Acid; ENCORI: The Encyclopedia of RNA Interactomes; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; IGF2BP2: insulin like growth factor 2 mRNA binding protein 2; IHC: Immunohistochemistry; LncRNA MALAT1: Long non-coding RNA metastasis-associated lung adenocarcinoma transcript 1; miRNAs: MicroRNAs; Mt: Mutant type; MTT: 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide; NC: negative control; NR2F2: nuclear receptor subfamily 2 group F member 2; NSCLC: non-small cell lung cancer; OD: optical density; PBS: phosphate-buffered saline; PD-L1: Programmed death-ligand 1; PD-1: programmed death 1; PI3K: phosphatidylinositol 3-kinase; qRT-PCR: Quantitative reverse transcription-polymerase chain reaction; RIP: RNA immunoprecipitation; RIPA: Radio Immunoprecipitation Assay; RRM2: ribonucleotide reductase regulatory subunit M2; RT: Radiation therapy; U6: U6 small nuclear RNA; V: volume; WB: Western blot; Wt: wild type; x ± sd: mean ± standard deviation.

A novel role for apatinib in enhancing radiosensitivity in non-small cell lung cancer cells by suppressing the AKT and ERK pathways.

BACKGROUND: Radioresistance is still the major cause of radiotherapy failure and poor prognosis in patients with non-small cell lung cancer (NSCLC). Apatinib (AP) is a highly selective inhibitor of vascular endothelial growth factor receptor 2 (VEGFR2). Whether and how AP affects radiosensitivity in NSCLC remains unknown. The present study aimed to explore the radiosensitization effect of AP in NSCLC and its underlying mechanism as a radiosensitizer. METHODS: The NSCLC cell lines A549 and LK2 were treated with AP, ionizing radiation (IR), or both AP and IR. Expression of VEGFR2 was analyzed by western blot and RT-PCR. Cell proliferation was measured using CCK-8 and colony formation assays. Apoptosis and cell cycle distribution in NSCLC cells were analyzed by flow cytometry. Nuclear phosphorylated histone H2AX foci immunofluorescence staining was performed to evaluate the efficacy of the combination treatment. Western blot was used to explore the potential mechanisms of action. RESULTS: AP inhibited cell proliferation in a dose- and time-dependent manner. Flow cytometry analysis indicated that AP significantly increased radiation-induced apoptosis. Colony formation assays revealed that AP enhanced the radiosensitivity of NSCLC cells. AP strongly restored radiosensitivity by increasing IR-induced G2/M phase arrest. AP effectively inhibited repair of radiation-induced DNA double-strand breaks. Western blot analysis showed that AP enhanced radiosensitivity by downregulating AKT and extracellular signal-regulated kinase (ERK) signaling. CONCLUSION: Our findings suggest that AP may enhance radiosensitivity in NSCLC cells by blocking AKT and ERK signaling. Therefore, AP may be a potential clinical radiotherapy synergist and a novel small-molecule radiosensitizer in NSCLC. Our study fills a gap in the field of anti-angiogenic drugs and radiosensitivity.

Active dry yeast supplementation improves the growth performance, rumen fermentation, and immune response of weaned beef calves.

The objective of this experiment was to investigate the potential benefits of active dry yeast (ADY) on the growth performance, rumen fermentation, nutrient digestibility, and serum parameters of weaned beef calves. Thirty Simmental crossbred male calves (body weight = 86.47 ± 4.41 kg and 70 ± 4 d of age) were randomly divided into 2 groups: control (CON) (fed basal ration) and ADY (fed basal ration and 5 g/d ADY per calf). The dietary concentrate-to-roughage ratio was 35:65. All the calves were regularly provided rations 3 times a day at 07:00, 13:00, and 19:00 and had free access to water. The experiment lasted for 60 d. The average daily gain of ADY group was higher (P = 0.007) than that of the CON group, and the ratio of feed intake to average daily gain in the ADY group was reduced (P = 0.022) as compared to the CON group. The concentration of ruminal ammonia-N was higher (P = 0.023) in the CON group than that in the ADY group, but an opposite trend of microbial protein was found between the 2 groups. Also, the ruminal concentrations of propionate and butyrate were higher (P < 0.05) in the ADY group than those in the CON group. Calves fed ADY exhibited higher (P < 0.05) crude protein and neutral detergent fiber digestibility. Supplementation of ADY increased (P < 0.05) the contents of glucose, glutathione peroxidase, superoxide dismutase, immunoglobulin A, immunoglobulin M, and interleukin 10 in the serum of calves, but an opposite trend was observed in malondialdehyde, interleukin 1 beta, and tumor necrosis factor alpha contents between the 2 groups. In conclusion, dietary supplementation with ADY could improve the growth performance, rumen fermentation, nutrient digestibility, antioxidant ability, and immune response of weaned beef calves.

Clinical implication of xenotropic and polytropic retrovirus receptor 1 in papillary thyroid carcinoma.

To investigate the expression of xenotropic and polytropic retrovirus receptor 1 （） in papillary thyroid cancer （PTC） and its clinical implication. The HPA and UALCAN databases were used to explore the expression of XPR1 in PTC and normal tissues. The cBioPortal database was used to obtain the clinical data of PTC patients and gene expression profile. The correlation of expression with gender，age，sub-types，T stage，N stage，M stage and clinical stage of patients were analyzed. Cox regression was conducted to analysis the factors affecting the prognosis of PTC patients. The mutation of was assessed through cBioPortal database. GO and KEGG analyses were used to explore the related biological pathway of involved in PTC. HPA database analysis showed that XPR1 was highly expressed in PTC tissue compared with normal tissues. UALCAN analysis displayed that expression was significantly higher in PTC tissue compared with normal tissues （<0.01），and the highest and lowest expressions of were observed in tall cell and follicular sub-type of PTC，respectively. The expression of was correlated with age，sub-types，T stage，N stage and disease stage of PTC patients （<0.05 or <0.01），but was not correlated with gender and M stage （all >0.05）. Cox regression analysis showed that was an independent prognostic factor of PTC patients （=2.894，<0.05）. The cBioPortal database indicated that the mutation appeared in 6% PTC patients; the mutation type mainly was missense and the mutation point was located at the E615K. Enrichment analysis indicated that might affect the PTC progression through involvement in metabolic pathway. is highly expressed in PTC tissues，which is associated with the prognosis of patients. Metabolic pathway associated with might play an important role in PTC progression，indicating that might be a novel biomarker for diagnosis and treatment of PTC.

ITGB1 enhances the Radioresistance of human Non-small Cell Lung Cancer Cells by modulating the DNA damage response and YAP1-induced Epithelial-mesenchymal Transition.

Objectives: Radiotherapy has played a limited role in the treatment of non-small cell lung cancer (NSCLC) due to the risk of tumour radioresistance. We previously established the radioresistant non-small cell lung cancer (NSCLC) cell line H460R. In this study, we identified differentially expressed genes between these radioresistant H460R cells and their radiosensitive parent line. We further evaluated the role of a differentially expressed gene, ITGB1, in NSCLC cell radioresistance and as a potential target for improving radiosensitivity. Materials and Methods: The radiosensitivity of NSCLC cells was evaluated by flow cytometry, colony formation assays, immunofluorescence, and Western blotting. Bioinformatics assay was used to identify the effect of ITGB1 and YAP1 expression in NSCLC tissues. Results: ITGB1 mRNA and protein expression levels were higher in H460R than in the parental H460 cells. We observed lower clonogenic survival and cell viability and a higher rate of apoptosis of ITGB1-knockdown A549 and H460R cells than of wild type cells post-irradiation. Transfection with an ITGB1 short hairpin (sh) RNA enhanced radiation-induced DNA damage and G2/M phase arrest. Moreover, ITGB1 induced epithelial-mesenchymal transition (EMT) of NSCLC cells. Silencing ITGB1 suppressed the expression and intracellular translocation of Yes-associated protein 1 (YAP1), a downstream effector of ITGB1. Conclusions: ITGB1 may induce radioresistance via affecting DNA repair and YAP1-induced EMT. Taken together, our data suggest that ITGB1 is an attractive therapeutic target to overcome NSCLC cell radioresistance.

Comparing the gastrointestinal barrier function between growth-retarded and normal yaks on the Qinghai-Tibetan Plateau.

BACKGROUND: Yak (Bos grunniens) is an ancient bovine species on the Qinghai-Tibetan Plateau. Due to extremely harsh condition in the plateau, the growth retardation of yaks commonly exist, which can reduce the incomes of herdsman. The gastrointestinal barrier function plays a vital role in the absorption of nutrients and healthy growth. Functional deficiencies of the gastrointestinal barrier may be one of the contributors for yaks with growth retardation. METHODS: To this end, we compared the growth performance and gastrointestinal barrier function of growth-retarded (GRY) and normal yaks (GNY) based on average daily gain (ADG), serum parameters, tissue slice, real-time PCR, and western blotting, with eight yaks in each group. RESULTS: GRY exhibited lower (P < 0.05) average daily gain as compared to GNY. The diamine oxidase, D-lactic acid, and lipopolysaccharide concentrations in the serum of GRY were significantly higher (P < 0.05) than those of GNY. Compared to GNY, the papillae height in the rumen of GRY exhibited lower (P = 0.004). In jejunum, with the exception of higher villus height, width, and surface area in GNY, numerical difference (P = 0.61) was detected between two groups for crypt depth. Both in rumen and jejunum, the mRNA expression of interleukin-1beta in GRY was markedly higher (P < 0.05) than that in GNY, but an opposite trend was found in interleukin-10 expression. Moreover, GRY showed a higher (P < 0.05) tumor necrosis factor-alpha mRNA expression in the rumen. The claudin-1 (CLDN1), occludin (OCLN), and zonula occludens-1 (ZO1) expressions of GRY in rumen and jejunum were significantly down-regulated (P < 0.05) as compared to GNY. The correlation analysis identified that in rumen and jejunum, there was a positive correlation between interleukin-10 and CLDN1, OCLN, and ZO1 mRNA expressions, but the tumor necrosis factor-alpha was negatively correlated with CLDN1, OCLN, and ZO1. In the rumen, the ADG was positively correlated with papillae surface area, and a same relationship between ADG and CLDN1, OCLN, and ZO1 expressions was found. CONCLUSION: The results indicated that the ruminal and jejunal barrier functions of GRY are disrupted as compared to GNY. In addition, our study provides a potential solution for promoting the growth of GRY by enhancing the gastrointestinal barrier function.

Identification of Aberrantly Expressed Genes in Murine Glioblastoma During Radiotherapy via Bioinformatic Data Mining.

OBJECTIVE: Glioblastoma (GBM) is an aggressive tumor with a fast growth rate. Radioresistance of GBM can lead to high recurrence. In general, due to the protection of the blood-brain barrier, the immune environment of the central nervous system is unique. The immune response induced by radiotherapy is weak in GBM. In the present study, aberrantly expressed genes during radiotherapy were assessed in murine models based on microarray RNA data. METHODS: The microarray data were extracted from the Intergovernmental Group on Earth Observations and differentially expressed genes (DEGs) screened out. Gene expression profiles of 115 samples in GSE56113 were analyzed and 104 genes were identified as aberrantly expressed based on GEO2R 8 d after radiotherapy. Then, the Database for Annotation, Visualization, and Integrated Discovery was used to analyze Genome Kyoto Encyclopedia of Gene pathways and Gene Ontology (GO) terms. The 20 core candidate genes were identified using protein-protein interaction network analysis and Cytoscape software with Molecular Complex Detection plug-in. RESULTS: Post-irradiated tumor tissues expressed significantly more immune-associated genes than contralateral brain tissues. GO and pathway analyses showed core DEGs were mainly enriched in the chemokine signaling and IL-6 signaling pathways, which could lead to immunosuppressive inflammatory monocyte infiltration and radioresistance. Chemokine signaling and IL-6 signaling pathway-associated genes were increased in the irradiated U87 cell strain. CONCLUSION: Chemokine signaling and IL-6 signaling pathways were activated after radiation in murine glioma and human glioma cell lines which could lead to changes in the immune microenvironment and treatment failure. The results of the present study could provide potential therapeutic targets especially when immune therapy and radiotherapy are combined to treat GBM patients.

Transcription levels and prognostic significance of the NFI family members in human cancers.

BACKGROUND: The nuclear factor I (NFI) is a family of transcription factors consisting of four distinct but closely related genes, NFIA, NFIB, NFIC and NFIX, which are important in the development of various tissues and organs in mammals. Recent study results have shown that NFI family may play a critical role in the progression of various human tumors and have been identified as key tumor suppressors and oncogenes for many cancers. However, the expression levels and distinctive prognostic values of the NFI family remain poorly explored in most cancers. MATERIALS AND METHODS: In the present study, the differences in mRNA expression of the NFI family in various cancers were investigated using the Oncomine and TCGA databases, and the mRNA expression, genetic alteration and DNA methylation of the NFI family members in various cancers were examined using cBioPortal for Cancer Genomics. In addition, the prognostic significance of the NFI family was assessed in multiple cancers using the Kaplan-Meier plotter (KM plotter) and SurvExpress databases. RESULTS: The mRNA expression levels in the NFI family were significantly downregulated in most cancers compared with normal tissues and DNA hypermethylation might downregulate the NFI family expression. Although NFIX expression was not downregulated in kidney, colorectal and prostate cancers. Furthermore, NFIB expression was upregulated in gastric cancer. Further survival analyses based on the KM plotter and SurvExpress databases showed dysregulations of the NFI genes were significantly correlated with survival outcomes in breast, lung, and head and neck cancers. Decreased expression levels of NFIA, NFIB and NFIC were associated with poor overall survival (OS) in head and neck cancer. Low mRNA expression of NFIA and NFIB was significantly associated with OS and first progression in lung adenocarcinoma, but not in lung squamous cell carcinoma. In addition, potential correlations between NFI family members and survival outcomes were also observed in liver, esophageal, kidney and cervical cancer. CONCLUSION: The results from the present study indicated certain members of the NFI family could be promising therapeutic targets and novel prognostic biomarkers for human cancers.

Difference in survival between right- versus left-sided colorectal neuroendocrine neoplasms.

Neuroendocrine neoplasms (NENs) are a heterogeneous group of tumors that arise from neuroendocrine cells, and in some cases are capable of producing agents that may cause characteristic hormonal syndromes (Cives and Strosberg, 2018). Such tumors were previously thought to be rare, but the rate of detection of NENs, especially from the gastrointestinal tract, is increasing with the widespread use of colonoscopy, cross-sectional imaging, and biomarkers (Gu et al., 2019). A study based on the Surveillance, Epidemiology, and End Results (SEER) database showed that the age-adjusted incidence of NENs increased 6.4-fold from 1973 (1.09 per 100 000) to 2012 (6.98 per 100 000) (Dasari et al., 2017), while there was a progressive increase in the incidence of colorectal NENs (Starzynska et al., 2017).

Long noncoding RNA MALAT1 knockdown inhibits progression of anaplastic thyroid carcinoma by regulating miR-200a-3p/FOXA1.

Long noncoding RNAs (lncRNAs) have been reported to play essential roles in progression of thyroid carcinoma. However, the roles of lncRNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in anaplastic thyroid carcinoma (ATC) process and its mechanism remain not been fully established. In this study, we focused on the effect of MALAT1 on cell proliferation, apoptosis, migration, invasion, and autophagy formation in ATC and explored the interaction between miR-200a-3p and MALAT1 or FOXA1. Moreover, murine xenograft model was established to investigate the roles and mechanism of MALAT1 in ATC progression in vivo. Results showed that MALAT1 expression was enhanced and miR-200a-3p was reduced in ATC tissues and cells. Knockdown of MALAT1 or overexpression of miR-200a-3p inhibited cell proliferation, migration and invasion but increased apoptosis and autophagy formation in ATC cells. Moreover, miR-200a-3p was directly bound to MALAT1 and its inhibition reversed the inhibitory effect of MALAT1 knockdown on progression of ATC. In addition, FOXA1 was indicated as a target of miR-200a-3p and its restoration attenuated the anti-cancer role of miR-200a-3p in ATC cells. Furthermore, MALAT1 functioned as a competing endogenous RNA (ceRNA) via sponging miR-200a-3p to derepress FOXA1 expression. Besides, interference of MALAT1 decreased tumor growth by upregulating miR-200a-3p and downregulating FOXA1. Collectively, MALAT1 knockdown suppressed ATC progression by regulating miR-200a-3p/FOXA1, providing a novel avenue for treatment of ATC.

A novel role for NFIA in restoring radiosensitivity in radioresistant NSCLC cells by downregulating the AKT and ERK pathways.

Radioresistance remains the most challenging issue leading to radiotherapy failure in the treatment of non-small cell lung cancer (NSCLC). The nuclear factor IA (NFIA) is associated with tumor response to treatments in many cancers, but its role in NSCLC radioresistance remains unclear. Here, we established two radioresistant NSCLC cell lines, H226R and H460R, by dose-gradient irradiation to investigate the function of NFIA in NSCLC radioresistance. The results showed a dramatically reduced expression of NFIA in radioresistant cells accompanied with elevated phosphorylation of AKT and ERK, when compared with their parental cells. Overexpression of NFIA restored the sensitivity of radioresistant cells to radiation through increased ionizing radiation (IR)-induced apoptosis and DNA damage by downregulating p-AKT and p-ERK, whereas knockdown of NFIA promoted radioresistance of the parental cells. Our findings suggested that NFIA enhanced cell radiosensitivity by downregulating p-AKT and p-ERK in NSCLC. Our study fills a gap in the field of NFIA and radioresistance, and establishes a mechanistic foundation to improve radiotherapy efficiency in NSCLC patients.

Nutritional Interventions Improved Rumen Functions and Promoted Compensatory Growth of Growth-Retarded Yaks as Revealed by Integrated Transcripts and Microbiome Analyses.

Growth retardation reduces the incomes of livestock farming. However, effective nutritional interventions to promote compensatory growth and the mechanisms involving digestive tract microbiomes and transcripts have yet to be elucidated. In this study, Qinghai plateau yaks, which frequently suffer from growth retardation due to malnutrition, were used as an experimental model. Young growth-retarded yaks were pastured (GRP), fed basal ration (GRB), fed basal ration addition cysteamine hydrochloride (CSH; GRBC) or active dry yeast (ADY; GRBY). Another group of growth normal yak was pastured as a positive control (GNP). After 60-day nutritional interventions, the results showed that the average daily gain (ADG) of GRB was similar to the level of GNP, and the growth rates of GRBC and GRBY were significantly higher than the level of GNP (P < 0.05). Basal rations addition of CSH or ADY either improved the serum biochemical indexes, decreased serum LPS concentration, facilitated ruminal epithelium development and volatile fatty acids (VFA) fermentation of growth-retarded yaks. Comparative transcriptome in rumen epithelium between growth-retarded and normal yaks identified the differentially expressed genes mainly enriched in immune system, digestive system, extracellular matrix and cell adhesion pathways. CSH addition and ADY addition in basal rations upregulated ruminal VFA absorption (SLC26A3, PAT1, MCT1) and cell junction (CLDN1, CDH1, OCLN) gene expression, and downregulated complement system (C2, C7) gene expression in the growth-retarded yaks. 16S rDNA results showed that CSH addition and ADY addition in basal rations increased the rumen beneficial bacterial populations (Prevotella_1, Butyrivibrio_2, Fibrobacter) of growth-retarded yaks. The correlation analysis identified that ruminal VFAs and beneficial bacteria abundance were significantly positively correlated with cell junction and VFA absorption gene expressions and negatively correlated with complement system gene expressions on the ruminal epithelium. Therefore, CSH addition and ADY addition in basal rations promoted rumen health and body growth of growth-retarded yaks, of which basal ration addition of ADY had the optimal growth-promoting effects. These results suggested that improving nutrition and probiotics addition is a more effective method to improve growth retardation caused by gastrointestinal function deficiencies.

Circlular RNA BARD1 (Hsa_circ_0001098) overexpression in breast cancer cells with TCDD treatment could promote cell apoptosis via miR-3942/BARD1 axis.

Breast cancer threatened the health of millions of people around the world. Here we explored the influence of TCDD on the expression of circRNA_BARD1 (circ_0001098) in breast cancer and studied the potential molecular mechanism of circRNA_BARD1. The data from GSE76608 was applied to analyze differentially expressed circRNAs and mRNAs. The expressions of circRNA_BARD1, BARD1, miR-3942-3p, miR-4760-3p and apoptosis-related protein p53 were detected by qRT-PCR or western blot. Circinteractome, TargetScan, CIRCNET and dual luciferase reporter assay were employed to uncover the target relationship between circRNA_BARD1/BARD1 and miR-3942-3p/miR-4760-3p. Flow cytometric analysis was used to reveal cell cycle and cell apoptosis. Immunofluorescence was applied to determinate γ-H2AX level. Xenograft assay and in vivo 3-D imaging was implemented to further verify the conclusions in vitro. CircRNA_BARD1 (circ_0001098) was up-regulated in breast cancer with the treatment of TCDD and the up-regulation of circRNA_BARD1 could restrain cell proliferation, block cell cycle and promote cell apoptosis. Moreover, the target relationship between circRNA_BARD1/BARD1 and miR-3942-3p was confirmed. In addition, miR-3942-3p overexpression promoted the disease progression and BARD1 up-regulation inhibited the disease progression in the breast cancer. Similarly, circRNA_BARD1 overexpression induced by TCDD suppressed the growth and metastasis of tumor in vivo. In conclusion, TCDD induced circ_0001098 overexpression and then suppressed breast cancer tumorigenesis via miR-3942-3p/BARD1 axis. The finding of TCDD-circRNA-miRNA-mRNA axis might bring a new perspective for cure strategy of breast cancer.

Subgroup analysis reveals molecular heterogeneity and provides potential precise treatment for pancreatic cancers.

BACKGROUND: The relationship between molecular heterogeneity and clinical features of pancreatic cancer remains unclear. In this study, pancreatic cancer was divided into different subgroups to explore its specific molecular characteristics and potential therapeutic targets. PATIENTS AND METHODS: Expression profiling data were downloaded from The Cancer Genome Atlas database and standardized. Bioinformatics techniques such as unsupervised hierarchical clustering was used to explore the optimal molecular subgroups in pancreatic cancer. Clinical pathological features and pathways in each subgroup were also analyzed to find out the potential clinical applications and initial promotive mechanisms of pancreatic cancer. RESULTS: Pancreatic cancer was divided into three subgroups based on different gene expression features. Patients included in each subgroup had specific biological features and responded significantly different to chemotherapy. CONCLUSION: Three distinct subgroups of pancreatic cancer were identified, which means that patients in each subgroup might benefit from targeted individual management.

Aggregation of lipid rafts activates c-met and c-Src in non-small cell lung cancer cells.

BACKGROUND: Activation of c-Met, a receptor tyrosine kinase, induces radiation therapy resistance in non-small cell lung cancer (NSCLC). The activated residual of c-Met is located in lipid rafts (Duhon et al. Mol Carcinog 49:739-49, 2010). Therefore, we hypothesized that disturbing the integrity of lipid rafts would restrain the activation of the c-Met protein and reverse radiation resistance in NSCLC. In this study, a series of experiments was performed to test this hypothesis. METHODS: NSCLC A549 and H1993 cells were incubated with methyl-ß-cyclodextrin (MßCD), a lipid raft inhibitor, at different concentrations for 1 h before the cells were X-ray irradiated. The following methods were used: clonogenic (colony-forming) survival assays, flow cytometry (for cell cycle and apoptosis analyses), immunofluorescence microscopy (to show the distribution of proteins in lipid rafts), Western blotting, and biochemical lipid raft isolation (purifying lipid rafts to show the distribution of proteins in lipid rafts). RESULTS: Our results showed that X-ray irradiation induced the aggregation of lipid rafts in A549 cells, activated c-Met and c-Src, and induced c-Met and c-Src clustering to lipid rafts. More importantly, MßCD suppressed the proliferation of A549 and H1993 cells, and the combination of MßCD and radiation resulted in additive increases in A549 and H1993 cell apoptosis. Destroying the integrity of lipid rafts inhibited the aggregation of c-Met and c-Src to lipid rafts and reduced the expression of phosphorylated c-Met and phosphorylated c-Src in lipid rafts. CONCLUSIONS: X-ray irradiation induced the aggregation of lipid rafts and the clustering of c-Met and c-Src to lipid rafts through both lipid raft-dependent and lipid raft-independent mechanisms. The lipid raft-dependent activation of c-Met and its downstream pathways played an important role in the development of radiation resistance in NSCLC cells mediated by c-Met. Further studies are still required to explore the molecular mechanisms of the activation of c-Met and c-Src in lipid rafts induced by radiation.