Construction of lncRNA-m6A gene-mRNA regulatory network to identify m6A-related lncRNAs associated with the progression of lung adenocarcinoma.

BACKGROUND: We evaluated the prognostic value of m6A-related long noncoding RNAs (lncRNAs) in lung adenocarcinoma (LUAD). METHODS: The expression levels of lncRNAs and mRNAs in LUAD and normal adjacent tissues from The Cancer Genome Atlas dataset were analyzed using the limma package. m6A enzyme-related differentially expressed lncRNAs and mRNAs were identified and used to construct a regulatory network. Survival analysis was performed and the correlation between lncRNAs, m6A regulators, and mRNAs was analyzed; followed by functional enrichment analysis. RESULTS: A comparison of LUAD samples and normal tissues identified numerous differentially expressed lncRNAs and mRNAs, demonstrating that a comprehensive network was established. Two lncRNAs and six mRNAs were selected as prognosis related factors including SH3PXD2A-AS1, MAD2L1, CCNA2, and CDC25C. The pathological stage and recurrence status were identified as independent clinical factors (P < 0.05). The expression levels of these RNAs in the different clinical groups were consistent with those in the different risk groups. The interactions of m6A proteins, two lncRNAs, and six mRNAs were predicted, and functional analysis showed that m6A target mRNAs were involved in the cell cycle, progesterone-mediated oocyte maturation, and oocyte meiosis pathways. CONCLUSIONS: These m6A target lncRNAs and mRNAs may be promising biomarkers for predicting clinical prognosis, and the lncRNA-m6A regulator-mRNA regulatory network could improve our understanding of m6A modification in LUAD progression.

Potential mitigation of environmental impacts of intensive plum production in southeast China with maintenance of high yields: Evaluation using life cycle assessment.

Introduction: Intensive plum production usually involves high yields but also high environmental costs due to excessive fertilizer inputs. Quantitative analysis of the environmental effects of plum production is thereby required in the development of optimum strategies to promote sustainable fruit production. Methods: We collected survey questionnaires from 254 plum production farms in Zhao'an county, Fujian province, southeast China to assess the environmental impacts by life cycle assessment (LCA) methodology. The farms were categorized into four groups based on yield and environmental impacts, i.e., LL (low yield and low environmental impact), LH (low yield but high environmental impact), HL (high yield but low environmental impact), and HH (high yield and high environmental impact). Results: The environmental impacts, i.e., average energy depletion, global warming, acidification, and eutrophication potential in plum production were 18.17 GJ ha-1, 3.63 t CO2 eq ha-1, 42.18 kg SO2 eq ha-1, and 25.06 kg PO4 eq ha-1, respectively. Only 19.7% of farmers were in the HL group, with 13.3% in the HH group, 39.0% in LL, and 28.0% LH. Plum yields of the HL group were 109-114% higher than the mean value of all 254 farms. Additionally, the HL group had a lower environmental impact per unit area compared to the overall mean value, with a reduction ranging from 31.9% to 36.7%. Furthermore, on a per tonne of plum production basis, the energy depletion, global warming potential, acidification potential, and eutrophication potential of HL farms were lower by 75.4%, 75.0%, 75.6%, and 75.8%, respectively. Overall, the total environmental impact index of LL, LH, HL, and HH groups were 0.26, 0.42, 0.06, and 0.21, respectively. Discussion: Excessive fertilizer N application was the main source of the environmental impacts, the potential to reduce fertilizer N rate can be achieved without compromising plum yield by studying the HH group. The results provide an important foundation for enhancing the management of plum production, in order to promote 'green' agricultural development by reducing environmental impacts.

SPARC expression in tumor microenvironment induces partial epithelial-to-mesenchymal transition of esophageal adenocarcinoma cells via cooperating with TGF-ß signaling.

Secreted protein, acidic and rich in cysteine (SPARC) has been characterized as an oncoprotein in esophageal squamous cell carcinoma (ESCC), but its involvement in the pathological development of esophageal adenocarcinoma (ESAD) remains poorly understood. In this study, we aimed to explore the sources of SPARC in the tumor microenvironment (TME) and its functional role in ESAD. Bioinformatic analysis was conducted using data from The Cancer Genome Atlas (TCGA)-esophageal cancer (ESCA) and Genotype-Tissue Expression (GTEx). ESAD tumor cell line OE33 and OE19 cells were used as in vitro cell models. Results showed that SPARC upregulation was associated with unfavorable disease-specific survival (DSS) in ESAD. ESAD tumor cells (OE33 and OE19) had no detectable SPARC protein expression. In contrast, IHC staining in ESAD tumor tissues suggested that peritumoral stromal cells (tumor-associated fibroblasts and macrophages) were the dominant SPARC source in TME. Exogenous SPARC induced partial epithelial-to-mesenchymal transition of ESAD cells, reflected by reduced CDH1 and elevated ZEB1/VIM expression at both mRNA and protein levels. Besides, exogenous SPARC enhanced tumor cell invasion. When TGFBR2 expression was inhibited, the activation of TGF-ß signaling induced by exogenous SPARC was impaired. However, the activating effects were rescued by overexpressing mutant TGFBR2 resistant to the shRNA sequence. Copresence of exogenous SPARC and TGF-ß1 induced higher expression of mesenchymal markers and enhanced the invading capability of ESAD cells than TGF-ß1 alone. In conclusion, this study suggests a potential cross-talk between ESAD tumor stromal cells and cancer cells via a SPARC-TGF-ß1 paracrine network.

Bacteria not fungi drive soil chemical quality index in banana plantations with increasing years of organic fertilizer application.

BACKGROUND: Maintaining or improving soil chemical quality is critical for sustainable agricultural productivity and environmental safeguards. Organic fertilizer application, a common agricultural practice in banana cultivation, is often associated with greater microbial biomass and activity, which are linked to improvements in soil chemical quality. However, the effect of the duration of organic fertilizer application on soil chemical quality and whether it is microbially driven still needs to be investigated. We collected soil samples from banana plantations consistently applying organic fertilizers for 1 (Y1), 4 (Y4), 7 (Y7) and 10 (Y10) years. Soil chemical quality is expressed as total data set (TDS) and minimum data set (MDS) based on chemical indicators, and soil microorganisms are characterized by phospholipid fatty acid (PLFA). RESULTS: Based on TDS and MDS, the soil chemical quality indices in Y7 and Y10 treatments were significantly higher than that in Y1 and Y4 treatments. Soil total PLFA concentrations and the proportional abundance of fungi and arbuscular mycorrhizal fungi increased with prolonged banana cultivation. Total PLFA concentrations were significantly positive correlation with the soil chemical quality index. Soil gram-positive bacteria (G+), bacteria, protozoa and ratio of G+ to gram-negative bacteria (G-) were major drivers of soil chemical quality. CONCLUSION: The organic fertilizer application can significantly improve soil chemical quality, which is regulated by soil bacteria. Regular application of organic fertilizers is important in promoting soil quality and soil biological properties need to be incorporated into the assessment of soil health in banana plantations. © 2022 Society of Chemical Industry.

Clinical Study of 2 Radiotherapy Techniques for Semi-Hepatic Alternating Radiotherapy on Diffuse Liver Metastasis in Patients with Breast Cancer.

Objective: To compare the effects of 2 techniques of semi-hepatic alternating radiotherapy on diffuse hepatic metastasis in patients with breast cancer. Methodology: A total of 68 breast cancer patients with diffuse liver metastasis were randomly divided into Group A (semi-hepatic alternating radiotherapy) and Group B (semi-hepatic sequential radiotherapy). In Group A (semi-hepatic sequential radiotherapy), the liver was divided into the first semi-liver and second semi-liver and alternatively treated with semi-hepatic intensity-modulated radiation therapy (IMRT). The interval between the 2 instances of semi-hepatic radiotherapy was 6 h. The average radiotherapy dose to the semi-livers was both 2 Gy/fraction, once a day, 5 times per week, with a total dose of 30 Gy for 15 days. The total radiation therapy time in Group A was 15 days in Group B (semi-hepatic sequential radiotherapy), the livers were divided into the first semi-liver and second semi-liver and treated with semi-hepatic sequential IMRT, The first semi-liver was first treated in the initial stage of radiation therapy, the average radiotherapy dose to the semi-liver was 2 Gy/fraction, once a day, 5 times per week, with a total dose of 30 Gy for 15 days. The second semi-liver was treated next in the second stage of radiation therapy, the average radiotherapy dose to the semi-liver was 2 Gy/fraction, once a day, 5 times per week, with a total dose of 30 Gy for 15 days. The total radiation therapy time in group B was 30 days. Results: The objective response rate (complete response + partial response) of Group A and Group B were 50.0% and 48.5%, respectively (p = .903). The median survival time after metastasis (median survival of recurrence) of Group A and Group B was 16.7 months and 16.2 months, respectively (p = .411). The cumulative survival rates of 6 months, 1 year, 2 years, and 3 years of Group A and Group B were 90.6% (29 of 32) and 84.8% (28 of 33) (p = .478), 65.6% (21 of 32) and 60.6% (20 of 33) (p = .675), 31.2% (10 of 32) and 27.3% (9 of 33) (p = .725), and 15.6% (5 of 32) and 0 (0 of 33) (p = .018), respectively. The differences between the 2 groups showed no statistical significance in terms of cumulative survival rates in 1 year, 2 years, however, the 3-year survival rate was significantly different. The main toxic reactions were digestive tract reactions, abnormal liver functions, and myelosuppression. The incidence of I to II degree gastrointestinal reactions was 78.13% (25 of 32) in Group A and 72.73% (24 of 33) in Group B (p = .614). The incidence of I to II abnormal liver function was 53.13% (17 of 32) in Group A and 48.48% (16 of 33) in Group B (p = .708). The differences between the 2 groups showed no statistical significance. The incidence of I to II myelosuppression was 59.38% (19 of 32) in Group A and 51.52% (17 of 33) in Group B (p = .524), respectively. The differences between the 2 groups showed no statistical significance in terms of adverse effects. Conclusion: Semi-hepatic alternating IMRT was an effective palliative treatment for diffuse liver metastasis in patients with breast cancer. Semi-hepatic alternating radiotherapy showed a trend of prolonged survival time when compared with semi-hepatic sequential radiotherapy. Compared with the former, the latter showed a trend of lower incidences of side effects without any statistical differences. Moreover, the side effects from the 2 radiotherapy techniques can be controlled through appropriate management, which is worthy of further exploration and applications.

Clinicopathological characteristics and prognosis of pulmonary spindle cell carcinoma (PSCC): a population-based retrospective study.

BACKGROUND: In consideration of the scarceness and importance of histological analysis, the clinic pathological features of pulmonary spindle cell carcinoma (PSCC) were comprehensively analyzed in the present work to improve the treatment and deepen our understanding of the disease. METHODS: Data of the PSCC patients from 2008 to 2013 in the Surveillance, Epidemiology, and End Results (SEER) database were acquired, analyzed and contrasted to that of the subjects with non-small cell lung cancer (NSCLC). Overall survival (OS) was evaluated based on the Kaplan-Meier method, univariate analysis (UVA) and multivariate analysis (MVA) were applied for the Cox proportional hazards regression. The risk factors related to 1-, 3-, and 5-year OS in PSCC subjects were identified. RESULTS: The data of 171 subjects considered to suffer from PSCC were collected and compared with that of 41,438 NSCLC patients. There was a poor differentiation in 72.9% of PSCC, and 44.4% were at the stage IV of American Joint Committee on Cancer (AJCC). The median OS time of PSCC was 8 months [95% confidence interval (CI): 6.23-10.72] with 5-year OS as 23.9% (95% CI: 21.5-25.7%). Tumors of PSCC were significantly undifferentiated, which exhibited the higher rate to be resected by surgery, with more lymph node metastases and distant metastases than that of NSCLC (P<0.001). It was demonstrated in UVA and MVA that the N stage, M stage, and surgery served independently as the risk factors of OS. The calibration variable for the nomogram was 0.735 (lower than 0.8). CONCLUSIONS: There were specific clinicopathologic features in PSCC. The results revealed that there was an independent correlation between N stage, M stage, or surgery with OS. However, 1-, 3- and 5-year OS could not be precisely predicted by the nomogram.

Long non-coding RNA SNHG1 stimulates ovarian cancer progression by modulating expression of miR-454 and ZEB1.

Ovarian cancer (OC) is highly prevalent and is associated with high mortality rates due to metastasis and relapse. In this study, we assessed the role of long non-coding RNA (lncRNA) small nucleolar RNA host gene 1 (SNHG1) in OC to gain further insight into mechanisms that contribute to its aggressiveness. We analyzed the correlation between SNHG1, miR-454 and zinc finger E-box-binding homeobox 1 (ZEB1) using a dual-luciferase reporter assay. Alterations in cell metastasis and invasiveness were observed using wound-healing and Transwell invasion assays, respectively. Tumor xenografts allowed us to monitor liver metastasis of mice injected with A2780 cells. We found that SNHG1 is overexpressed in OC. Downregulation of SNHG1 promoted miR-454 expression and reduced ZEB1 levels. In addition, knockdown of SNHG1, also reduced the aggressiveness of A2780 and SK-OV3 cells. Furthermore, SNHG1 downregulation by siRNA hindered cell migration and invasion; however, this effect was reversed by co-transfection of miR-454 into A2780 and SK-OV3 cells. Moreover, SNHG1 increased ZEB1 expression by downregulating miR-454 and activated Akt signaling, thereby promoting epithelial-mesenchymal transition and enhancing the invasiveness of OC cells. Tumor xenograft analyses confirmed that SNHG1 affects OC proliferation and metastasis in vivo. In summary, our data demonstrate that SNHG1 plays crucial roles in tumor progression and may be a useful maker for OC prognosis.

Evaluation of the Efficacy and Toxicity of Radiotherapy for Type III-IV Portal Vein Tumor Thrombi.

BACKGROUND: Type Ⅲ and Ⅳ portal vein tumor thrombi (PVTT) cannot be removed through surgery, and no effective therapeutic procedure is available. Type Ⅲ/Ⅳ PVTT can be downstage to type I/II PVTT by using Radiotherapy, and can further be can be removed surgically. Thus, radiotherapy may be an effective treatment for type Ⅲ/Ⅳ PVTT. This study aims to evaluate the efficacy and toxicity of radiotherapy for type III-IV PVTT. METHODS: This prospective study was conducted from August 1, 2017, to September 30, 2019, for patients with type Ⅲ and Ⅳ PVTT. Patients received radiotherapy with a target dose of 50Gy/25f or 59.5Gy/17 f. Advanced radiological technique such as image fusion technique for CT image and MRI image were utilized to produce more precise lesion localization, and limit the dose to organs at risk in order to get a better downstage rate and less adverse complications. RESULTS: Nine (9) patients with type Ⅲ PVTT and 5 patients with type Ⅳ PVTT were included in this study. 12 patients received a radiotherapy dose of 50Gy/25f, 2 patients received 59.50Gy/17 f. After radiotherapy, 92.9% of patients with PVTT were successfully downstage to type II/I. In patients with primary hepatocellular carcinoma, 8 patients (accounting 88.9%) achieved down-stage. 5 patients with other types of tumors achieved downstage which accounts 100%. In addition, none of the 14 patients observed radiation hepatitis and radiation liver failure. And none of the patients developed gastrointestinal ulcers and thrombocytopenia. CONCLUSION: Radiotherapy is a suitable treatment measure for type Ⅲ and Ⅳ PVTT to get downstage and make the opportunity for surgery. Image fusion technology for precise lesion location such as CT-MRI image fusion, and strict dose limitation of organ at risk, contributed to the improvement of radiotherapy efficiency and the significant decrease in adverse complications.

Identification of Hypoxia-Immune-Related Gene Signatures and Construction of a Prognostic Model in Kidney Renal Clear Cell Carcinoma.

Introduction: Kidney renal clear cell carcinoma (KIRC), a kind of malignant disease, is a severe threat to public health. Tracking the information of tumor progression and conducting a related dynamic prognosis model are necessary for KIRC. It is crucial to identify hypoxia-immune-related genes and construct a prognostic model due to immune interaction and the influence of hypoxia in the prognosis of patients with KIRC. Methods: The hypoxia and immune status of KIRC patients were identified by utilizing t-SNE and ImmuCellAI for gene expression data. COX and Lasso regression were used to identify some hypoxia-immune-related signature genes and further construct a prognostic risk model based on these genes. Internal and external validations were also conducted to construct a prognostic model. Finally, some potentially effective drugs were screened by the CMap dataset. Results: We found that high-hypoxia and low-immune status tend to induce poor overall survival (OS). Six genes, including PLAUR, UCN, PABPC1L, SLC16A12, NFE2L3, and KCNAB1, were identified and involved in our hypoxia-immune-related prognostic risk model. Internal verification showed that the area under the curve (AUC) for the constructed models for 1-, 3-, 4-, and 5-year OS were 0.768, 0.754, 0.775, and 0.792, respectively. For the external verification, the AUC for 1-, 3-, 4-, and 5-year OS were 0.768, 0.739, 0.763, and 0.643 respectively. Furthermore, the decision curve analysis findings demonstrated excellent clinical effectiveness. Finally, we found that four drugs (including vorinostat, fludroxycortide, oxolinic acid, and flutamide) might be effective and efficient in alleviating or reversing the status of severe hypoxia and poor infiltration of immune cells. Conclusion: Our constructed prognostic model, based on hypoxia-immune-related genes, has excellent effectiveness and clinical application value. Moreover, some small-molecule drugs are screened to alleviate severe hypoxia and poor infiltration of immune cells.

Downregulation of GPR4 and TCF7 Promotes Apoptosis and Inhibits Growth and Invasion of Ovarian Cancer Cells.

BACKGROUND: G Protein-coupled Receptor 4 (GPR4) has been reported to play essential roles in regulating the proliferation, migration and angiogenesis of vascular endothelial cells. GPR4 is also suggested to play significant roles in the growth and angiogenesis of ovarian cancer. OBJECTIVE: To explore the functions of GPR4 and Transcription Factor 7 (TCF7) in ovarian cancer. METHODS: The expression levels of genes involved in Wnt signaling were validated by quantitative Real-Time- PCR (q-RT-PCR). The effects of GPR4 and TCF7 on ovarian cancer cell invasion and apoptosis were determined using soft agar, transwell assay and flow cytometric assay. Protein levels of beta-catenin, MMP-2 and MMP-9 were evaluated by Western blotting. RESULTS: In this study, we found that GPR4 and TCF7 had the capacity to control cell division by altering cell cycle distribution, anchorage-independent growth, and directional cell motility of ovarian cancer cell A2780. Also, we showed that the knockdown of GPR4 and TCF7 in ovarian cancer cell A2780 induced significant inhibitition of cell growth and invasion, as well as the promotion of apoptosis. Downregulation of TCF7 resulted in the decreased MMP-2 and MMP-9 levels. CONCLUSION: The results implicate that GPR4 behaves like an oncogene and may function through WNT pathway molecule TCF7. Downregulation of GPR4 and TCF7 essentially inhibited cell growth and invasion and enhanced apoptosis of ovarian cancer cells, which may lay a foundation for ovarian cancer treatment.

The effect of CCL5 on the immune cells infiltration and the prognosis of patients with kidney renal clear cell carcinoma.

Background: Kidney renal clear cell carcinoma (KIRC) is the most representative subtype of renal cancer. Immune infiltration was associated with the survival time of patients with tumors. C-C chemokine ligand 5 (CCL5) can promote the malignant process of tumor and be related to infiltration immune cells in some cancers, but not reported in KIRC. Methods: The expression profile and clinical data were obtained from The Cancer Genome Atlas (TCGA) database. The correlation between the expression level of CCL5 and clinical features in KIRC was analyzed. Gene Set Enrichment Analysis (GSEA) was utilized to explore the functions and pathways of CCL5 in KIRC. Then, the analysis between the survival and immune infiltration cells was carried out, as well as the non-parametric tests between the CCL5 expression and the ratios of immune infiltration cells. Results: The correlations between the expression levels of CCL5 in KIRC and clinical features including survival time, pathological stage, grade, and status of the patient, have been identified. Meanwhile, GSEA analysis has shown relationships between the expression of CCL5 and immune pathways. The immune infiltrated cells were correlated with the prognosis of KIRC, especially regulatory T cells (Tregs), mast cells, and dendritic cells. And Tregs was associated with the CCL5 expression. Conclusion: The increased expression of CCL5 is related to poor prognosis and clinical features. Meanwhile, CCL5 is related to Tregs ratios and CCL5 may act as a typical chemokine to recruit Tregs in KIRC. CCL5 could be used as a biomarker for the prognosis prediction and a potential therapeutic target for patients with KIRC.

Construct a circRNA/miRNA/mRNA regulatory network to explore potential pathogenesis and therapy options of clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is the most representative subtype of renal cancer. CircRNA acts as a kind of ceRNA to play a role in regulating microRNA (miRNA) in many cancers. However, the potential pathogenesis role of the regulatory network among circRNA/miRNA/mRNA is not clear and has not been fully explored. CircRNA expression profile data were obtained from GEO datasets, and the differentially expressed circRNAs (DECs) were identified through utilizing R package (Limma) firstly. Secondly, miRNAs that were regulated by these circRNAs were predicted by using Cancer-specific circRNA database and Circular RNA Interactome. Thirdly, some related genes were identified by intersecting targeted genes, which was predicted by a web tool (miRWalk) and differentially expressed genes, which was obtained from TCGA datasets. Function enrichment was analyzed, and a PPI network was constructed by Cytoscape software and DAVID web set. Subsequently, ten hub-genes were screened from the network, and the overall survival time in patients of ccRCC with abnormal expression of these hub-genes were completed by GEPIA web set. In the last, a circRNA/miRNA/mRNA regulatory network was constructed, and potential compounds and drug which may have the function of anti ccRCC were forecasted by taking advantage of CMap and PharmGKB datasets. Six DECs (hsa_circ_0029340, hsa_circ_0039238, hsa_circ_0031594, hsa_circ_0084927, hsa_circ_0035442, hsa_circ_0025135) were obtained and six miRNAs (miR-1205, miR-657, miR-587, miR-637, miR-1278, miR-548p) which are regulated by three circRNAs (hsa_circ_0084927, hsa_circ_0035442, hsa_circ_0025135) were also predicted. Then 497 overlapped genes regulated by these six miRNAs above had been predicted, and function enrichment analysis revealed these genes are mainly linked with some regulation functions of cancers. Ten hub-genes (PTGER3, ADCY2, APLN, CXCL5, GRM4, MCHR1, NPY5R, CXCR4, ACKR3, MTNR1B) have been screened from a PPI network. PTGER3, ADCY2, CXCL5, GRM4 and APLN were identified to have a significant effect on the overall survival time of patients with ccRCC. Furthermore, one compound (josamycin) and four kinds of drugs (capecitabine, hmg-coa reductase inhibitors, ace Inhibitors and bevacizumab) were confirmed as potential therapeutic options for ccRCC by CMap analysis and pharmacogenomics analysis. This study implies the potential pathogenesis of the regulatory network among circRNA/miRNA/mRNA and provides some potential therapeutic options for ccRCC.

Long non-coding RNA SNGH7 Is activated by SP1 and exerts oncogenic properties by interacting with EZH2 in ovarian cancer.

Long non-coding RNAs (lncRNAs) are key regulators or a range of diseases and chronic conditions such as cancers, but how they function in the context of ovarian cancer (OC) is poorly understood. The Coding-Potential Assessment Tool was used to assess the likely protein-coding potential of SNHG7. SNHG7 expression was elevated in ovarian tumour tissues measured by qRT-PCR. The online database JASPAR was used to predict the transcription factors binding to SNHG7. Twenty-four-well Transwell plates were used for invasion assays. RNA immunoprecipitation was performed to determine RNA-protein associations. EdU assay was introduced to detect cell proliferation. Chromatin immunoprecipitation was performed to confirm the directly interaction between DNA and protein. We discovered that in the context of OC there is a significant up-regulation of the lncRNA SNHG7. Knocking down this lncRNA disrupted both OC cell invasion and proliferation, while its overexpression had the opposite effect. SP1 binding sites were present in the SNHG7 promoter, and chromatin immunoprecipitation (ChIP) confirmed direct SP1 binding to this region, activating SNHG7 transcription. We found that at a mechanistic level in OC cells, KLF2 is a probable SNHG7 target, as we found that SHNCCC16 directly interacts with EZH2 and thus represses KLF2 expression. In summary, this research demonstrates that lncRNA SNHG7 is an SP1-activated molecule that contributes to OC progression by providing a scaffold whereby EZH2 can repress KLF2 expression.

Association between G-protein coupled receptor 4 expression and microvessel density, clinicopathological characteristics and survival in hepatocellular carcinoma.

G-protein coupled receptor 4 (GPR4) acts as a proton-sensing receptor and plays a role in regulating angiogenesis. Endoglin/CD105 is a marker of cell proliferation in vascular endothelial cells, particularly in tumor vasculature cells. Although there have been several studies investigating angiogenesis in hepatocellular carcinoma (HCC), none have investigated the association between GPR4 and microvessel density (MVD)-CD105 in this type of cancer. In the present study, CD105 and GPR4 were found to be expressed in benign and malignant liver tissues by immunofluorescence staining and laser confocal microscopy. Compared with levels in benign tissues, CD105 and GPR4 were highly expressed in neoplastic tissues. Furthermore, the average fluorescence intensity of GPR4 and MVD-CD105 was positively correlated. GPR4 and CD105 were found to be co-localized in the vascular endothelium in tumor tissues. Furthermore, the expression of GPR4 was higher in the marginal region of tumor tissues compared with the central region. These findings suggest that the expression of GPR4 in tumor microvessels in HCC may be implicated in tumor angiogenesis and development. Furthermore, the association between the expression of GPR4 and the clinicopathological features of patients with HCC further suggests a role for GPR4 in tumor angiogenesis and growth. Overall, these results suggest the potential of GPR4 as a prognostic factor and as an antiangiogenic target in patients with HCC.

Organic fertilizer application and Mg fertilizer promote banana yield and quality in an Udic Ferralsol.

Low soil fertility, high rates of fertilizer application and low yields and quality are major problems in intensive banana production in acid soils of south China. A field experiment was carried out for two years to determine the optimum management practices for maximizing soil health and banana yield and quality. The experiment consisted of an unamended control (CK) and lime (Lime), calcium magnesium phosphate fertilizer (CMP), organic fertilizer (OF), and organic fertilizer combined with calcium magnesium phosphate fertilizer (OFC) treatments. Soil nutrient concentrations and banana shoot biomass, nutrient uptake, yield and fruit quality were determined. Application of lime and CMP was found to increase soil pH and nutrient availability and increase banana yield. Yet, the banana biomass and yields in the Lime and CMP treatments were significantly lower than those in the OF and OFC treatments in which soil organic matter (SOM) content increased. Total soluble solids and soluble sugar contents increased in the CMP and organic fertilizer treatments. A consistent increase in Mg concentrations in banana leaves over the two years in the CMP and organic fertilizer treatments indicates that Mg is essential for banana production and quality. Short-term adding Mg from banana corms increased total soluble solids and soluble sugar content. The application of organic fertilizer combined with CMP or Mg solution is therefore recommended to increase soil health and promote the yield and quality of banana in intensively managed plantations in subtropical regions.

Effect of co-transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of hepatocellular carcinoma Huh7.

OBJECTIVE: To investigate the effects of co-transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of hepatocellular carcinoma Huh7. METHODS: Hepatocellular carcinoma Huh7 was cultured in vitro and lipidosome was used to transfect miR-520c-3p and miR-132, respectively or together. The effects of transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of Huh7 were detected by CCK8 and Annexin V staining and flow cytometry, and the expression level of the targeted gene of over-expressed miR-520c-3p and miR-132 was determined by Western blot and realtime PCR. RESULTS: Compared with the control group, the proliferation ability of Huh7 of the single transfected and co-transfected miR-520c-3p and miR-132 decreased significantly, and the apoptosis ratio increased distinctly (P < 0.05). Besides, the effect of the co-transfection group was better than that of the single transfection group. The protein levels of GPC3 (Glypican-3) and YAP (Yes-associated protein), the target genes transfected only by miR-520c-3p and miR-132, respectively, reduced obviously (P < 0.05), which was similar with the co-infected cells, but cells transfected by miR-132 only showed a decrease of YAP. CONCLUSIONS: The co-transfection of miR-520c-3p and miR-132 can target-regulate the expression of GPC3 and YAP, enhance the exhibition effect on proliferation of hepatocellular carcinoma Huh7 and induce cell apoptosis synergistically.

Change of the peripheral blood immune pattern and its correlation with prognosis in patients with liver cancer treated by sorafenib.

OBJECTIVE: To study the change of the peripheral blood immune pattern and its correlation with prognosis in patients with liver cancer after treated by sorafenib. METHODS: Patients with advanced liver cancer admitted in our hospital were enrolled and treated with sorafenib. After two months of the treatment, their peripheral blood was collected. The immune cell subset and cytokines level were determined by flow cytometry and luminex technology. According to the reaction expressed by patients towards sorafenib, patients were divided into the response group and the no response group. The changes of the peripheral blood immune pattern and its correlation with prognosis of patients in the two groups were compared. RESULTS: Before and after treatment of sorafenib, there was no significant difference in the ratios of T cells, NK cells and their subtypes in peripheral blood of patients between the two groups; while after treatment the ratio of B cells and regulatory B cells (Breg) of patients in the response group was significant higher than that of the no response group (P < 0.05), and the prognosis conditions of patients with decreased ratio of Breg cells were better than other patients after undergoing chemotherapy. The levels of plasma cytokines IL-6, IL-10, IL-12, IL17, FIL-3L, IFN-γ, TNF-α, MCP-1 and VEGF showed no significant differences. CONCLUSIONS: After treatment of sorafenib, the prognosis conditions of patients of advanced liver cancer with a reduced Breg ratio are better than patients with an unaltered or increased Breg ratio. The ratio of Breg in peripheral blood may be considered as early biological indicator for the prediction of the curative effects of sorafenib.

Clinical research on alternating hyperfraction radiotherapy for massive hepatocellular carcinoma.

The delivery of high tumoricidal doses of radiation with low rates of toxicity is of particular significance for massive hepatocellular carcinoma (HCC) radiotherapy. In order to observe the efficacy and adverse reactions of alternating hyperfraction radiotherapy treatment of massive HCC, seventy-two cases of massive HCC were randomly divided into two groups, group A and group B. The liver lesions of group A were divided into sublesions and treated with alternating hyperfraction radiotherapy [intensity modulated radiotherapy (IMRT)]. The interval between radiotherapy to the sublesions was a minimum of six hours. The average radiotherapy dose to the sublesions was 2 Gy/fraction, once a day, five times per week, treating the gross tumor volume with a total dose of 40-50 Gy, and the clinical target volume with a total dose of 30-40 Gy. By contrast, the lesions of group B were not divided into sublesions for the IMRT treatment, but were treated with an otherwise identical protocol, by 2 Gy/fraction, once a day, five times per week, and with the same total dose. Patients were followed up with regular blood tests, liver function tests, measurements of serum α-fetoprotein levels and contrast-enhanced magnetic resonance imaging (MRI) of the liver. Treatment responses were assessed every 3 months by MRI. The results revealed that the overall response rates of the two groups were 82.9 and 81.3%, respectively (P=0.864). The alternating hyperfraction radiotherapy protocol resulted in enhanced survival (P=0.002). The median survival time of the two groups was 9.7 and 6.5 months, respectively. The overall 6-month, 1-year, 2-year and 3-year survival rates of the two groups were 62.9 and 59.4% (P=0.770), 48.6 and 21.9% (P=0.040), 17.1 and 0.0% (P=0.025) and 2.9 and 0.0% (P=1.000), respectively. The I-II degree of abnormal liver function and radiation-induced liver disease of group B was higher than that of group A (P=0.021 and 0.046, respectively). In addition, the incidence rate of radiation-induced liver injury of group A was lower than that of group B. Therefore, treatment of massive HCC with alternating hyperfraction radiotherapy improved the quality of life and prolonged the overall survival time, compared with conventional IMRT, suggesting that it was an effective radiation pattern.

Characterization of alkaline phosphatase labeled UidA(Gus) probe and its application in testing of transgenic tritordeum.

Hybridization is a very important molecular biology technique to measure the degree of genetic similarity between DNA sequences, and detect the foreign genes in transgenic organisms. To label a DNA or RNA probe plays a key role in hybridization. A method using nonradioactive material alkaline phosphatase to label UidA(Gus) DNA as probe has been studied. On that basis of Renz and our previous work, alkaline phosphatase-labeled DNA was used as a probe to examine the transformation of the foreign UidA(Gus) gene in transgenic tritordeum. Such DNA-enzyme complexes were characterized and examined carefully, the results showed that it was a sensitive, specific, safe and economical probe. For dot hybridization and Southern blot under full-stringency conditions with alkaline phosphatase as the detector and 5-bromo-4-chloro-3-indolyl phosphate (BCIP)-Nitro Blue Tetrazolium (NBT) as the substrate, dot hybridization showed that the UidA(Gus) gene was transformed into the target plants and inherited stable, Southern blot showed that at least two copies of UidA(Gus) gene were inserted into one line of our transgenic tritordeum. Histochemical staining with X-Gluc of transgenic tritordeum also certified that the foreign UidA(Gus) DNA were transformed into the transgenic tritordeum.