Comprehensive analyses of solute carrier family members identify SLC12A2 as a novel therapy target for colorectal cancer.

As the largest transporter family impacting on tumor genesis and development, the prognostic value of solute carrier (SLC) members has not been elucidated in colorectal cancer (CRC). We aimed to identify a prognostic signature from the SLC members and comprehensively analyze their roles in CRC. Firstly, we downloaded transcriptome data and clinical information of CRC samples from GEO (GSE39582) and TCGA as training and testing dataset, respectively. We extracted the expression matrix of SLC genes and established a prognostic model by univariate and multivariate Cox regression. Afterwards, the low-risk and high-risk group were identified. Then, the differences of prognosis traits, transcriptome features, clinical characteristics, immune infiltration and drug sensitivity between the two groups were explored. Furthermore, molecular subtyping was also implemented by non-negative matrix factorization (NMF). Finally, we studied the expression of the screened SLC genes in CRC tumor tissues and normal tissues as well as investigated the role of SLC12A2 by loss of function and gain of function. As a result, we developed a prognostic risk model based on the screened 6-SLC genes (SLC39A8, SLC2A3, SLC39A13, SLC35B1, SLC4A3, SLC12A2). Both in the training and testing sets, CRC patients in the high-risk group had the poorer prognosis and were in the more advanced pathological stage. What's more, the high-risk group were enriched with CRC progression signatures and immune infiltration. Two groups showed different drug sensitivity. On the other hand, two distinct subclasses (C1 and C2) were identified based on the 6 SLC genes. CRC patients in the high-risk group and C1 subtype had a worse prognosis. Furthermore, we found and validated that SLC12A2 was steadily upregulated in CRC. A loss-of-function study showed that knockdown of SLC12A2 expression restrained proliferation and stemness of CRC cells while a gain-of-function study showed the contrary results. Hence, we provided a 6-SLC gene signature for prognosis prediction of CRC patients. At the same time, we identified that SLC12A2 could promote tumor progression in CRC, which may serve as a potential therapeutic target.

Chlamydospore dormancy and predatory activity of nematophagous fungus Duddingtonia flagrans.

The chlamydospores of Duddingtonia flagrans are an essential survival and reproductive structure and also an effective ingredient for the biocontrol of parasitic nematodes in livestock. In this study, entering and exiting dormancy conditions and predatory activity of the fungal chlamydospores were conducted. During this fungal growth process, the cultivation time is negatively correlated with spore germination rates. After the spores were processed by vacuum drying for 168 h, their germination rate dropped to 0.94%. In contrast, the percentage of living spores remained 54.82%, suggesting that the spores entered structural dormancy in the arid environment. Meanwhile, the efficacies of the spore against Haemonchus contortus larvae were 93.05% (0 h), 92.19% (16 h), 92.77% (96 h), and 86.45% (168 h), respectively. After dormant spores were stored at 4°C, -20°C, and 28°C (RH90 ~ 95%) for 7 days, their germination rate began to increase significantly (p < 0.05). For in vitro predation assay under the condition of 28°C (RH90 ~ 95%), the predation rate was significantly higher on the 7th day after incubation than that on the 3rd day (p < 0.05). During the period when spores were stored at room temperature for 8 months, their germination rate decreased in the first 5 months and then increased slowly to reach a peak in the 7th month. However, the reduction rate of H. contortus L3 in feces captured by spores remained above 71% for the first 7 months. These results will help us increase the end products yield and the quality of biological control of parasitic nematodes in livestock.

NAPSB as a predictive marker for prognosis and therapy associated with an immuno-hot tumor microenvironment in hepatocellular carcinoma.

BACKGROUND: Napsin B Aspartic Peptidase, Pseudogene (NAPSB) was associated with CD4 + T cell infiltration in pancreatic ductal adenocarcinoma. However, the biological role of NAPSB in hepatocellular carcinoma (HCC) remains to be determined. METHODS: The expression of NAPSB in HCC as well as its clinicopathological association were analyzed using data from several public datasets. qRT-PCR was used to verify the relative expression of NAPSB in patients with HCC using the Zhongnan cohort. Kaplan-Meier analyses, and univariate and multivariate Cox regression were conducted to determine the prognosis value of NAPSB on patients with HCC. Then enrichment analyses were performed to identify the possible biological functions of NAPSB. Subsequently, the immunological characteristics of NAPSB in the HCC tumor microenvironment (TME) were demonstrated comprehensively. The role of NAPSB in predicting hot tumors and its impact on immunotherapy and chemotherapy responses was also analyzed by bioinformatics methods. RESULTS: NAPSB was downregulated in patients with HCC and high NAPSB expression showed an improved survival outcome. Enrichment analyses showed that NAPSB was related to immune activation. NAPSB was positively correlated with immunomodulators, tumor-infiltrating immune cells, T cell inflamed score and cancer-immunity cycle, and highly expressed in immuno-hot tumors. High expression of NAPSB was sensitive to immunotherapy and chemotherapy, possibly due to its association with pyroptosis, apoptosis and necrosis. CONCLUSIONS: NAPSB was correlated with an immuno-hot and inflamed TME, and tumor cell death. It can be utilized as a promising predictive marker for prognosis and therapy in HCC.

Metabolism-Associated Molecular Classification of Colorectal Cancer.

The high heterogeneity of colorectal cancer (CRC) is the main clinical challenge for individualized therapies. Molecular classification will contribute to drug discovery and personalized management optimizing. Here, we aimed to characterize the molecular features of CRC by a classification system based on metabolic gene expression profiles. 435 CRC samples from the Genomic Data Commons data portal were chosen as training set while 566 sample in GSE39582 were selected as testing set. Then, a non-negative matrix factorization clustering was performed, and three subclasses of CRC (C1, C2, and C3) were identified in both training set and testing set. Results showed that subclass C1 displayed high metabolic activity and good prognosis. Subclass C2 was associated with low metabolic activities and displayed high immune signatures as well as high expression of immune checkpoint genes. C2 had the worst prognosis among the three subtypes. Subclass C3 displayed intermediate metabolic activity, high gene mutation numbers and good prognosis. Finally, a 27-gene metabolism-related signature was identified for prognosis prediction. Our works deepened the understanding of metabolic hallmarks of CRC, and provided valuable information for "multi-molecular" based personalized therapies.

Identification of NEO1 as a prognostic biomarker and its effects on the progression of colorectal cancer.

BACKGROUND: Due to the high morbidity and poor clinical outcomes, early predictive and prognostic biomarker identification is desiderated in colorectal cancer (CRC). As a homologue of the Deleted in Colorectal Cancer (DCC) gene, the role of Neogenin-1 (NEO1) in CRC remained unveiled. This study was designed to probe into the effects and potential function of NEO1 in CRC. METHODS: Online databases, Gene Set Enrichment Analysis (GSEA), quantitative real-time PCR and western blotting were used to evaluate NEO1 expression in colorectal cancer tissues. Survival analysis was performed to predict the prognosis of CRC patients based on NEO1 expression level. Then, cell proliferation was detected by colony formation and Cell Counting Kit 8 (CCK-8) assays. CRC cell migration and invasion were examined by transwell assays. Finally, we utilized the Gene Set Variation Analysis (GSVA) and GSEA to dig the potential mechanisms of NEO1 in CRC. RESULTS: Oncomine database and The Cancer Genome Atlas (TCGA) database showed that NEO1 was down-regulated in CRC. Further results validated that NEO1 mRNA and protein expression were both significantly lower in CRC tumor tissues than in the adjacent tissues in our clinical samples. NEO1 expression was decreased with the progression of CRC. Survival and other clinical characteristic analyses exhibited that low NEO1 expression was related with poor prognosis. A gain-of-function study showed that overexpression of NEO1 restrained proliferation, migration and invasion of CRC cells while a loss-of-function showed the opposite effects. Finally, functional pathway enrichment analysis revealed that NEO1 low expression samples were enriched in inflammation-related signaling pathways, EMT and angiogenesis. CONCLUSION: A tumor suppressor gene NEO1 was identified and verified to be correlated with the prognosis and progression of CRC, which could serve as a prognostic biomarker for CRC patients.

Identification of PIGU as the Hub Gene Associated with KRAS Mutation in Colorectal Cancer by Coexpression Analysis.

Colorectal cancer (CRC) patients with KRAS mutation are refractory and usually have poor prognosis. We aimed to identify the hub gene associated with KRAS mutant CRCs. Weighted gene coexpression network analysis (WGCNA) was used to calculate the key module and the hub genes in GSE39582. Combined with the protein-protein interaction (PPI) network and survival analysis, the real hub gene was identified and further validated. With the highest module significance value and correlation coefficient, the blue module was selected as the key module, 19 genes were identified as the hub gene candidates. The above genes were significantly downregulated in KRAS mutant CRCs compared with the wild type. Four genes (AAR2, PSMA7, NELFCD, and PIGU) were further screened as the potential hub genes by the PPI network. Low expression of PIGU for KRAS mutant patients had a poor prognosis. Therefore, PIGU was identified as the hub gene. PIGU expression was also downregulated in other two CRC datasets. "MAPK SIGNALING PATHWAY" was enriched in PIGU lowly expressed samples. PIGU was identified and validated to be closely related to KRAS mutation. It could be a potential prognosis biomarker and a novel treatment target for KRAS mutant CRC patients.

MicroRNA-375-3p enhances chemosensitivity to 5-fluorouracil by targeting thymidylate synthase in colorectal cancer.

Resistance to chemotherapy is a major challenge for the treatment of patients with colorectal cancer (CRC). Previous studies have found that microRNAs (miRNAs) play key roles in drug resistance; however, the role of miRNA-373-3p (miR-375-3p) in CRC remains unclear. The current study aimed to explore the potential function of miR-375-3p in 5-fluorouracil (5-FU) resistance. MicroRNA-375-3p was found to be widely downregulated in human CRC cell lines and tissues and to promote the sensitivity of CRC cells to 5-FU by inducing colon cancer cell apoptosis and cycle arrest and by inhibiting cell growth, migration, and invasion in vitro. Thymidylate synthase (TYMS) was found to be a direct target of miR-375-3p, and TYMS knockdown exerted similar effects as miR-375-3p overexpression on the CRC cellular response to 5-FU. Lipid-coated calcium carbonate nanoparticles (NPs) were designed to cotransport 5-FU and miR-375-3p into cells efficiently and rapidly and to release the drugs in a weakly acidic tumor microenvironment. The therapeutic effect of combined miR-375 + 5-FU/NPs was significantly higher than that of the individual treatments in mouse s.c. xenografts derived from HCT116 cells. Our results suggest that restoring miR-375-3p levels could be a future novel therapeutic strategy to enhance chemosensitivity to 5-FU.

Inhibition of Heat Shock Protein 90 as a Novel Platform for the Treatment of Cancer.

Heat shock protein 90 (Hsp90) plays an essential role in various physiological and pathological processes. It activates client proteins to participate in tumor progression. Blocking Hsp90 could enable effective antitumor effects in many tumor types, such as multiple myeloma and colon cancer. Recently, it has motivated an interest in Hsp90 inhibitors that bind to the N-terminal or C-terminal ATP pocket as antitumor drugs. We reviewed the data from experimental and clinical trials on Hsp90 inhibitors in the treatment of different malignancies to explore and summarize their antitumor mechanisms.

Coexpression network analysis identified that plakophilin 1 is associated with the metastasis in human melanoma.

BACKGROUND AND AIMS: Malignant melanoma is a fatal cancer with high metastatic characteristics. Approximately 80% of skin cancer deaths are caused by metastatic melanoma. It has been established that the metastatic ability of melanoma is regulated by an intricate gene interconnection network. Thus, the aim of this study was to identify and validate hub genes associated with metastatic melanoma and to further illustrate its potential mechanisms. METHODS: The method of weighted gene coexpression network analysis (WGCNA) was applied to explore potential regulatory targets and investigate the relationship between the key module and hub genes associated with the metastasis ability of melanoma. RESULTS: In the turquoise module, 26 hub genes were initially selected, and 6 of them were identified as "real" hub genes with high connectivity in the protein-protein interaction network. In terms of validation, PKP1 had the highest correlation with metastasis among all the "real" hub genes. Data obtained from the GEPIA database and the Gene Expression Omnibus database showed a lower expression of PKP1 in melanoma tissues compared to normal skin tissues. The results also showed that PKP1 was downregulated in metastatic melanomas (n = 367) compared with primary melanomas (n = 103) in The Cancer Genome Atlas (TCGA) database (n = 470). Furthermore, an ROC curve showed that PKP1 expression had good power in the diagnostics of both primary melanoma (p = 5.30e-06, AUC = 0.8) and metastatic melanoma (p = 1.13e-10, AUC = 0.925). We also found that PKP1 could distinguish low- and high-grade of metastatic melanomas and was associated with inflammatory melanoma. Moreover, in a tumor-bearing mouse model, melanoma tissues also showed lower mRNA expression of PKP1 than the adjacent normal skin. Finally, Gene Set Enrichment Analysis indicated that the calcium signaling was significantly enriched in metastatic melanoma with highly expressed PKP1. CONCLUSIONS: PKP1 was identified as a new potential tumor suppressor in human melanoma, likely through regulating calcium signaling pathways.

Soft fibrin matrix downregulates DAB2IP to promote Nanog-dependent growth of colon tumor-repopulating cells.

Colon cancer stem cells (CSCs) have been shown to be responsible for the recurrence and metastasis of colorectal cancer (CRC). As a crucial microenvironmental factor, extracellular matrix (ECM) stiffness is known to affect the stemness of CSCs. Recently, fibrin deposition in the stroma of CRC was demonstrated to be responsible for tumor development. In this study, we used salmon fibrin gel to provide a 3D ECM for colon cancer cells and investigated its effects on cell growth as well as the underlying mechanisms. Compared with stiff 420 Pascal (Pa) and 1 050 Pa gels, 90 Pa soft fibrin gel was most efficient at isolating and enriching tumor colonies. The size and number of colony formation negatively correlated with gel stiffness. Specifically, these tumor colonies exhibited efficient tumorigenicity, upregulated stem cell markers, and had anti-chemotherapeutic properties and were thus named tumor-repopulating cells (TRCs). More importantly, the self-renewal molecule Nanog was sharply induced in 3D-cultured colon TRCs; further, Nanog siRNA significantly inhibited colony formation, suggesting the indispensable role of Nanog in TRC growth. A subsequent mechanistic study illustrated that Nanog expression could be modulated through fibrin gel stiffness-induced DAB2IP/PI3K/FOXA1 signaling in colon TRCs.

[Histone Deacetylation Down-regulates the Expression of BNIP3 in Renal Cell Carcinoma].

OBJECTIVES: To investigate the down-regulation mechanism of (bcl-2/adenovirus E1B 19 kDa interacting protein 3 (BNIP3) expression in renal cell carcinoma (RCC). METHODS: RCC cell lines 786-O, ACHN and A498 were treated with different concentrations of histone deacetylase inhibitor TSA. Thereafter, the proliferation of RCC cells was determined with CCK-8 assay, cell apoptosis was observed by flow cytometry, and the expression levels of BNIP3 were determined by Q-PCR and Western blot, and the acetylation status of histone H3 in the promoter of BNIP3 was detected by ChIP. RESULTS: After the treatment with TSA, the proliferation of the three RCC cell lines was significantly inhibited (P<0.05), the early apoptosis of cells obviously increased, and the expression levels of BNIP3 mRNA (P<0.05) and protein were up-regulated. The histone H3 in BNIP3 promoter of both 786-O and ACHN was deacetylated, while the histone H3 in BNIP3 promoter of A498 was acetylated. CONCLUSIONS: Histone deacetylation may be the important mechanism of BNIP3 silencing in RCC.

[Immediate Inguinal Lymph Node Dissection Improves Survival in Patients of Penile Cancer].

OBJECTIVE: To investigate the clinical outcome of immediate inguinal lymph node dissection on the survival of the patients with penile carcinoma. METHODS: A total of 67 patients of penile carcinoma whose inguinal lymph nodes (ILN) were initial clinically impalpable, received inguinal lymph node dissection (ILND) from Dec 2008 to April 2014. Among them, 33 patients received immediate ILND within 1 month after the resection of penile cancer, while 34 patients underwent delayed ILND which was performed when ILN was found clinically apparent during follow-up. The Kaplan-Meier survival analysis was performed. The prognostic factors was evaluated by log-rank test, including age, morphology, location, T stage, grade of primary tumor, clinical status of ILN before ILND, lymphatic pathology, time to ILND. Cox proportional hazard model was used to find the independent risk factors on survival. RESULTS: The median age was 50 year-old (range 26 to 84 year-old). The median follow-up time was 23 months (range 3-76 months). The 3-year and 5-year overall survival were 70.1% and 65.4%, respectively, The 5-year survival rate in immediate ILND and delayed ILND group were 93.1%, and 33.7% respectively. Positive ILN metastasis was found in 7 patients from immediate ILND group but 26 patients from delayed ILND group that the prognostic factors included T stage, tumor grade, clinical status of inguinal lymph nodes before ILND, and lymphatic pathology. Cox model found the status of inguinal lymph nodes was independent prognostic factor for the survival. CONCLUSION: Inguinal lymph node metastasis is the important prognostic indicator of the survival of penile cancer. Immediate ILND could improve survival for the patients with clinically impalpable lymph nodes.

[Progress in the application of laser ablation ICP-MS to surface microanalysis in material science].

In the present paper, apparatus and theory of surface analysis is introduced, and the progress in the application of laser ablation ICP-MS to microanalysis in ferrous, nonferrous and semiconductor field is reviewed in detail. Compared with traditional surface analytical tools, such as SEM/EDS (scanning electron microscopy/energy dispersive spectrum), EPMA (electron probe microanalysis analysis), AES (auger energy spectrum), etc. the advantage is little or no sample preparation, adjustable spatial resolution according to analytical demand, multi-element analysis and high sensitivity. It is now a powerful complementary method to traditional surface analytical tool. With the development of LA-ICP-MS technology maturing, more and more analytical workers will use this powerful tool in the future, and LA-ICP-MS will be a super star in elemental analysis field just like LIBS (Laser-induced breakdown spectroscopy).

[Element distribution analysis of welded fusion zone by laser-induced breakdown spectroscopy].

Over the past decade there has been intense activity in the study and development of laser-induced breakdown spectroscopy (LIBS). As a new tool for surface microanalysis, it caused widespread in materials science because of the advantage of rapid and high sensitivity. In the present paper, the distribution of Ni, Mn, C and Si near weld fusion line was analyzed on two kinds of weld sample. Line scanning mode analysis was carried out by three different kinds of methods, namely laser-induced breakdown spectroscopy (LIBS), scanning electron microscope/energy dispersive spectrometer (SEM/EDS) and electron probe X-ray microanalyser (EPMA). The concentration variation trend of Ni and Mn acquired by LIBS is coincident with SEM/EDS and EPMA. The result shows that the content of Ni and Mn was significantly different between weld seam and base metal on both the samples. The content of Ni and Mn was much higher in weld seam than in base metal, and a sharp concentration gradient was analyzed in the fusion zone. According to the distribution of Ni and Mn, all the three methods got a similar value of welded fusion zone width. The concentration variation trend of C and Si acquired by LIBS is not coincident with SEM/EDS and EPMA. The concentration difference between weld seam and base metal was analyzed by LIBS, but had not by SEM/EDS and EPMA, because of the low concentration and slight difference. The concentration gradient of C and Si in fusion zone was shows clearly by LIBS. For higher sensitivity performance, LIBS is much more adapted to analyze low content element than SEM/EDS and EPMA.

[Expression and significance of BNIP3 in clear cell renal cell carcinoma].

OBJECTIVE: To investigate the expression of proapoptosis protein BNIP3 in clear cell renal cell carcinoma (ccRCC) and its clinical significance. METHODS: The RCC tumor tissue samples from 30 pathologically diagnosed ccRCC and their adjacent pericarcinous tissues were adopted to detect the mRNA and protein expressions of BNIP3, von Hippel-Lindau (VHL), hypoxia inducible factor (HIF)-1alpha and vascular enothelial growth factor (VEGF) by real-time quantitative PCR (real-time PCR) and Western blot. The correlations of these genes expressions with clinicopathologic features were analyzed. RESULTS: The expression levels of BNIP3 and VHL were lower in ccRCC tissues than those in pericarcinous tissues (P < 0.05), but the mRNA expression levels of HiF-1alpha and VEGF were higher in ccRCC tissues than those in pericarcinous tissues (P < 0.05). The lower level expression of BNIP3 in ccRCC was not related with any clinicopathologic features. No significant correlation was observed between the BNIP3 mRNA and protein level with the expressions of VHL, HIF-1alpha and VEGF. CONCLUSION: In ccRCC, the expression of BNIP3 is decreased, which not correlated with the expression levels of VHL, HIF-1alpha and VEGF.

[Segregation bands analysis of steel sample using laser-induced breakdown spectroscopy].

In the present paper, under optimum experimental condition, two middle-low alloy slab and homogeneous samples were analyzed under the condition of spatial resolution about 100 microm by scanning mode. Element 2D intensity distribution can be converted into 2D concentration distribution via establishing calibration curve. The results showed that there is a central segregation for C, Si, Mn, P, S and Cu for 86 # slab sample, and C, Si, P and Ti for 174 # slab sample, the width of segregation band was estimated, and it agrees well with metallographic analysis. Homogeneous sample was analyzed by scanning mode, the result showed that C, Si, Mn, P, S and so on are well distributed, and there is no segregation band existing. 2D distribution of element intensity or concentration can be used to indirectly reflect sample's homogeneity. Compared with traditional metallographic analysis, LIBS can not only show central segregation bands position and width, but also provide 2D concentration distribution for C, Si, Mn, P, S etc in detail. This method can be used to characterize segregation band position and its width rapidly, and provide theoretical guidance for improving metallurgical process.

[Automatic adjustment control system for DC glow discharge plasma source].

There are three important parameters in the DC glow discharge process, the discharge current, discharge voltage and argon pressure in discharge source. These parameters influence each other during glow discharge process. This paper presents an automatic control system for DC glow discharge plasma source. This system collects and controls discharge voltage automatically by adjusting discharge source pressure while the discharge current is constant in the glow discharge process. The design concept, circuit principle and control program of this automatic control system are described. The accuracy is improved by this automatic control system with the method of reducing the complex operations and manual control errors. This system enhances the control accuracy of glow discharge voltage, and reduces the time to reach discharge voltage stability. The glow discharge voltage stability test results with automatic control system are provided as well, the accuracy with automatic control system is better than 1% FS which is improved from 4% FS by manual control. Time to reach discharge voltage stability has been shortened to within 30 s by automatic control from more than 90 s by manual control. Standard samples like middle-low alloy steel and tin bronze have been tested by this automatic control system. The concentration analysis precision has been significantly improved. The RSDs of all the test result are better than 3.5%. In middle-low alloy steel standard sample, the RSD range of concentration test result of Ti, Co and Mn elements is reduced from 3.0%-4.3% by manual control to 1.7%-2.4% by automatic control, and that for S and Mo is also reduced from 5.2%-5.9% to 3.3%-3.5%. In tin bronze standard sample, the RSD range of Sn, Zn and Al elements is reduced from 2.6%-4.4% to 1.0%-2.4%, and that for Si, Ni and Fe is reduced from 6.6%-13.9% to 2.6%-3.5%. The test data is also shown in this paper.

[Laser-induced breakdown spectrometer--a new tool for quick analysis of on-the-spot sample in metallurgy].

In metallurgical steel making a quick analysis of on-the-spot sample is required to know the process of steel making. A new-type spectrometer, laser-induced breakdown spectrometer developed by ourselves, was presented in the present paper. A Nd : YAG laser with pulse width of nanometer was used as an ionization and excitation source. Emission from the plasma ap-peared when the laser beam was focused on the surface of sample. After it was spectrally resolved by a Paschen-Runge polychromator, detected by photomultiplier detectors, integrated by gate intensifier, and converted by analog-to-digital converter, the final result was transmitted to a computer in order to complete data processing. Compared to the common spectrometer on-the-spot in metallurgy, this instrument allows fast analyis without sample preparation (one minute or less), with high precision and sensitivity, so it is very suitable for the analysis of on-the spot sample in metallurgy. In recent years with the fast development of optical fiber, on-line analysis of liquid steel and dynamical control of metallurgical processing will come true by using this instrument.