KLHDC7B as a novel diagnostic biomarker in urine exosomal mRNA promotes bladder urothelial carcinoma cell proliferation and migration, inhibits apoptosis.

Bladder cancer is a common kind of urinary system cancer, in which bladder urothelial carcinoma (BLCA) comprises approximately 90% of all bladder cancer types. In our previous study, we discovered KLHDC7B in urine exosomal messenger RNA (mRNA) as a prospective molecular marker for bladder cancer detection. To systematically study the role and mechanism of KLHDC7B in BLCA, we focused on the most common type of BLCA in this study. First, we used RNA sequencing to discover that KLHDC7B was considerably increased in BLCA patients' urine exosomes compared to healthy controls. Then, we validated this result in an independent cohort and identified it as an effective tool for diagnosing and distinguishing high-grade and low-grade BLCA. Finally, we studied the role and mechanism of KLHDC7B in BLCA at the cellular level, providing a functional basis for its expression as a novel laboratory diagnostic biomarker for BLCA exosomal mRNA, which has important theoretical and clinical significance.

Large-Scale Quantitative Proteomic Analysis during Different Stages of Somatic Embryogenesis in Larix olgensis.

Larix olgensis is an economically important tree species native to northeastern China. The use of somatic embryogenesis (SE) is efficient and enables the rapid production of varieties with desirable qualities. Here, isobaric labeling via tandem mass tags was used to conduct a large-scale quantitative proteomic analysis of proteins in three critically important stages of SE in L. olgensis: the primary embryogenic callus, the single embryo, and the cotyledon embryo. We identified 6269 proteins, including 176 shared differentially expressed proteins across the three groups. Many of these proteins are involved in glycolipid metabolism, hormone response/signal transduction, cell synthesis and differentiation, and water transport; proteins involved in stress resistance and secondary metabolism, as well as transcription factors, play key regulatory roles in SE. The results of this study provide new insights into the key pathways and proteins involved in SE in Larix. Our findings have implications for the expression of totipotency, the preparation of synthetic seeds, and genetic transformation.

Expression Landscape and Functional Roles of HOXA4 and HOXA5 in Lung Adenocarcinoma.

BACKGROUND: The role of HOXA family genes in the occurrence and progression of a variety of human cancers has been scatteredly reported. However, there is no systematic study on the differential expression, prognostic significance and potential molecular mechanism of HOXA4 and HOXA5 in LUAD. METHODS: In-house immunohistochemistry (IHC), multi-center microarrays, RT-qPCR and RNA-seq data were incorporated for comprehensively evaluating the expression and prognostic value of HOXA4 and HOXA5 in LUAD. The mechanism of HOXA4 and HOXA5 in the formation and development of LUAD was analyzed from multiple aspects of immune correlations, upstream transcriptional regulation, functional states of single cells and co-expressed gene network. The functional roles of HOXA4 and HOXA5 in LUAD were validated by in vitro experiments. RESULTS: As a result, in 3201 LUAD samples and 2494 non-cancer lung samples, HOXA4 and HOXA5 were significantly downexpressed (P < 0.05). The aberrant expression of HOXA5 was significantly correlated with the clinical progression of LUAD (P < 0.05). HOXA5 showed remarkable prognostic value for LUAD patients (P < 0.05). The expression of HOXA4 and HOXA5 in LUAD were negatively correlated with tumor purity and positively correlated with the infiltration of various immune cells such as B cells, T cells and macrophages. HOXA4 and HOXA5 overexpression had notable inhibitory effect on the proliferation, migration and invasion of LUAD cells. CONCLUSIONS: In conclusion, the identified downexpressed HOXA4 and HOXA5 had significant distinguishing ability for LUAD samples and affected the cellular functions of LUAD cells. The low expression of HOXA5 indicated worse overall survival of LUAD patients. Therefore, the two HOXA family genes especially HOXA5 may serve as potential biomarkers for LUAD.

Three steps in one pot: biosynthesis of 4-hydroxycinnamyl alcohols using immobilized whole cells of two genetically engineered Escherichia coli strains.

BACKGROUND: 4-Hydroxycinnamyl alcohols are a class of natural plant secondary metabolites that include p-coumaryl alcohol, caffeyl alcohol, coniferyl alcohol and sinapyl alcohol, and have physiological, ecological and biomedical significance. While it is necessary to investigate the biological pathways and economic value of these alcohols, research is hindered because of their limited availability and high cost. Traditionally, these alcohols are obtained by chemical synthesis and plant extraction. However, synthesis by biotransformation with immobilized microorganisms is of great interest because it is environmentally friendly and offers high stability and regenerable cofactors. Therefore, we produced 4-hydroxycinnamyl alcohols using immobilized whole cells of engineered Escherichia coli as the biocatalyst. RESULTS: In this study, we used the recombinant E. coli strain, M15-4CL1-CCR, expressing the fusion protein 4-coumaric acid: coenzyme A ligase and the cinnamoyl coenzyme A reductase and a recombinant E. coli strain, M15-CAD, expressing cinnamyl alcohol dehydrogenase from Populus tomentosa (P. tomentosa). High performance liquid chromatography and mass spectrometry showed that the immobilized whole cells of the two recombinant E. coli strains could effectively convert the phenylpropanoic acids to their corresponding 4-hydroxycinnamyl alcohols. Further, the optimum buffer pH and the reaction temperature were pH 7.0 and 30 °C. Under these conditions, the molar yield of the p-coumaryl alcohol, the caffeyl alcohol and the coniferyl alcohol was around 58, 24 and 60%, respectively. Moreover, the highly sensitive and selective HPLC-PDA-ESI-MSn method used in this study could be applied to the identification and quantification of these aromatic polymers. CONCLUSIONS: We have developed a dual-cell immobilization system for the production of 4-hydroxycinnamyl alcohols from inexpensive phenylpropanoic acids. This biotransformation method is both simple and environmental-friendly, which is promising for the practical and cost effective synthesis of natural products. Graphical abstract Biotransformation process of phenylpropanoic acids by immobilized whole-cells.

MicroRNA-185 inhibits angiogenesis in human microvascular endothelial cells through targeting stromal interaction molecule 1.

Angiogenesis is a vital biological mechanism representing the adaptive response to a variety of pathological stimuli such as hypoxia. It is regulated by several pro-angiogenic and anti-angiogenic microRNAs. Studies have demonstrated an altered microRNA-185 (miR-185) expression in endothelial cells under hypoxic conditions; however, its role in angiogenesis has not been elucidated. We investigated the role of miR-185 in angiogenesis and found that miR-185 had an inhibitory effect on cell proliferation, migration, and tube formation. Stromal interaction molecule 1 (STIM1) appeared to be a direct target of miR-185 by computational prediction; this was confirmed by luciferase reporter assay. Silencing of the STIM1 gene was found to mimic miR-185-mediated inhibition of angiogenesis. STIM1 overexpression eliminated the anti-angiogenic effect of miR-185. Our study results suggest a direct interaction between miR-185 and STIM1 mRNA in microvascular endothelial cells. MicroRNA-185 acted as a negative regulator of angiogenesis in microvascular endothelial cells through downregulation of the STIM1 protein.

Overexpression of artificially fused bifunctional enzyme 4CL1-CCR: a method for production of secreted 4-hydroxycinnamaldehydes in Escherichia coli.

BACKGROUND: 4-Hydroxycinnamaldehydes are important intermediates in several secondary metabolism pathways, including those involved in the biosynthesis of phenolic acids, flavonoids, terpenoids and monolignols. They are also involved in the biosynthesis and degradation of lignins, which are important limiting factors during the processes of papermaking and biofuel production. Access to these aromatic polymers is necessary to explore the secondary biometabolic pathways they are involved in. Coniferaldehyde, sinapaldehyde, p-coumaraldehyde and caffealdehyde are members of the 4-hydroxycinnamaldehyde family. Although coniferaldehyde and sinapaldehyde can be purchased from commercial sources, p-coumaraldehyde and caffealdehyde are not commercially available. Therefore, there is increasing interest in producing 4-hydroxycinnamaldehydes. Here, we attempted to produce 4-hydroxycinnamaldehydes using engineered Escherichia coli. RESULTS: 4-Coumaric acid: coenzyme A ligase (4CL1) and cinnamoyl coenzyme A reductase (CCR) were fused by means of genetic engineering to generate an artificial bifunctional enzyme, 4CL1-CCR, which was overexpressed in cultured E. coli supplemented with phenylpropanoic acids. Three 4-hydroxycinnamaldehydes, p-coumaraldehyde, caffealdehyde and coniferaldehyde, were thereby biosynthesized and secreted into the culture medium. The products were extracted and purified from the culture medium, and identically characterized by the HPLC-PDA-ESI-MSn. The productivity of this new metabolic system were 49 mg/L for p-coumaraldehyde, 19 mg/L for caffealdehyde and 35 mg/L for coniferaldehyde. Extracellular hydroxycinnamoyl-coenzyme A thioesters were not detected, indicating that these thioesters could not pass freely through the cellular membrane. The fusion enzyme 4CL1-CCR can catalyze sequential multistep reactions, thereby avoiding the permeability problem of intermediates, which reveals its superiority over a mixture of individual native enzymes. Moreover, we have described a highly sensitive and selective method for separation and identification of phenylpropanoic acids and their corresponding cinnamaldehydes in the present paper. The feasibility of this method has been proven in the application of the method to the analysis of the metabolites of whole-cell catalysts. CONCLUSIONS: We have established a bioconversion pathway for the microbial production of valuable 4-hydroxycinnamaldehydes from phenylpropanoic acids. This biotransformation method is both convenient and environmentally friendly, and provides new insights into the biosynthesis of natural plant secondary products.

[Stromal interaction molecule 1 silencing attenuates the proliferation and migration capacities of endothelial progenitor cells].

OBJECTIVE: The purpose of this study is to explore the impact of stromal interaction molecule 1 (STIM1) knockdown on the proliferation and migration capacities of endothelial progenitor cells (EPCs). METHODS: The rat bone marrow derived EPCs were obtained and divided into three groups: adenovirus negative control (NSC) group, rat STIM1 adenovirus vector transfection (si/rSTIM1) group and rat and human recombinant STIM1 adenovirus transfection (si/rSTIM1+hSTIM1) group. The STIM1 expressions in each group were detected by reverse transcription PCR after transfection. The cell proliferation was tested by [(3)H] thymidine incorporation assay ((3)H-TdR). Cell cycle was analyzed by flow cytometry. The cells migration activity was detected by Boyden assay. Calcium ion concentration was detected by confocal laser scanning microscopy. RESULTS: 48 h after transfection, the expression level of STIM1 in si/rSTIM1 group was significantly lower than that in NSC group (0.21 ± 0.12 vs. 1.01 ± 0.01, P < 0.05), and number of EPCs at G1 phase in si/rSTIM1 group ((93.31 ± 0.24)%) was significantly higher than that in NSC group ((78.03 ± 0.34)%, P < 0.05), and EPCs' migration activity in si/rSTIM1 group (10.03 ± 0.33) was significantly lower than that in NSC group (32.11 ± 0.54, P < 0.05), and EPCs calcium ion concentration in EPCs in si/rSTIM1 group (38.03 ± 0.13) was significantly lower than that in NSC group (98.11 ± 0.34, P < 0.05), while there was no significant difference between si/rSTIM1+hSTIM1 group and NSC group on the above four indexes. CONCLUSION: Silencing STIM1 could attenuate EPCs proliferation and migration capacities by modulating the calcium ion concentration in EPCs.

Glycometabolic disorder induced by chronic exposure to low-concentration imidacloprid in zebrafish.

The health risks induced by chronic exposure to low concentrations of imidacloprid (IMI) to zebrafish were investigated in this study. The results indicated that the growth of zebrafish was inhibited after being exposed to 10, 100, and 500 µg/L of IMI for 90 days. Moreover, the blood glucose levels in the IMI-exposed groups were significantly higher compared to the control group. Investigation into the development of zebrafish larvae revealed that IMI exposure hindered the development of the liver and pancreatic islets, organs crucial for glucose metabolism. In addition, the IMI-exposed groups exhibited reduced liver glycogen and plasma insulin levels, along with changes in the activity of enzymes and the transcription levels of genes associated with liver glucose metabolism. These findings suggest that IMI induces glycometabolic disorders in zebrafish. The analysis of intestinal flora revealed that several key bacteria associated with an elevated risk of diabetes were significantly altered in IMI-exposed fish. Specifically, a remarkable decrease was found in the abundance of the genera Aeromonas and Shewanella, which have been found closely related to the development of pancreatic islets. This implies that the alteration of key bacteria in the fish gut by IMI, which in turn affects the development of organs such as the pancreatic islets, may be the initial trigger for abnormalities in glucose metabolism. Our results revealed that chronic exposure to low concentrations of IMI led to glycometabolic disorder in fish. Therefore, considering the pervasive existence of IMI residues in the environment, the health hazards posed by low-concentration IMI to fish cannot be overlooked.

Absorption and Distribution of Imidacloprid and Its Metabolites in Goldfish (Carassius auratus Linnaeus).

Imidacloprid (IMI) is the first-generation neonicotinoid insecticide. But, the long-term use of IMI as a pesticide has caused severe water pollution. Recently, the toxicity of IMI to aquatic organisms has received increasing attention. This study aimed to investigate the absorption and distribution of IMI in various tissues (gills, intestine, liver, muscle, brain and gonads) of goldfish through short-term and continuous exposure tests over 28 days. The results of short-term exposure indicated that the concentration of IMI and its metabolites in tissues at the transfer stage decreased steadily after 1 day of 40 mg/L IMI water treatment and was below the detection limit after 3 days. Continuous exposure for 28 days at various treatment concentrations showed that the concentrations of IMI and its metabolites differed significantly among the different tissues of the goldfish. In the 20 mg/L treatment group (S1), the highest concentration of IMI was found in the liver (12.04 µg/gtissue), followed by the intestine (9.91 µg/gtissue), muscle (6.20 µg/gtissue), gill (6.11 µg/gtissue), gonads (5.22 µg/gtissue) and brain (2.87 µg/gtissue). In the 40 mg/L treatment group (S2), the order of the tissue concentrations was similar to that of the S1 group, with the highest concentration observed in the liver (12.04 µg/gtissue), followed by the intestine (9.91 µg/gtissue), muscle (6.20 µg/gtissue), gill (6.11 µg/gtissue), gonads (5.22 µg/gtissue) and brain (2.87 µg/gtissue). Furthermore, the study detected 5-hydroxy-IMI, IMI urea and 6-chloronicotinic acid in IMI metabolites in all tissues, while IMI was detected only in the intestine and liver. Overall, the results of this study contribute to a better understanding of the metabolic behavior of IMI in organisms and provide new data to support the assessment of IMI toxicity in fish.

Dissipation behavior and dietary risk assessment of cyclaniliprole and its metabolite in cabbage under field conditions.

Cyclaniliprole, a novel diamide insecticide, can successfully control Spodoptera litura (Fabricius, 1775) in cabbage. Understanding the residual level of cyclaniliprole in crops and the risk related to its dietary intake is imperative for safe application. Here, we established a simplified, sensitive method for simultaneous analysis of cyclaniliprole and its metabolite NK-1375 (3-bromo-2-((2-bromo-4H-pyrazolo[1,5-d]pyrido[3,2-b]-[1,4]oxazin-4-ylidene)amino)-5-chloro-N-(1-cyclopropylethyl)benzamide) in cabbage by liquid chromatography-tandem mass spectrometry (LC-MS/MS) to investigate their dissipation behavior and residual characteristics. Cyclaniliprole showed rapid dissipation in cabbage and had a half-life of 1.8-2.7 days. The highest residue of total cyclaniliprole (sum of cyclaniliprole and NK-1375) in cabbage from different pre-harvest intervals (3 and 5 days) was 0.25 mg/kg. Our results confirmed the generally low dietary risk quotient of cyclaniliprole (0.243-1.036%) among different age and gender groups in China. Therefore, cyclaniliprole did not pose an unacceptable risk to consumers. This study contributes to setting cyclaniliprole maximum residue limit in cabbage by assessing its dissipation fate and food safety risks.

Delay line length selection in generating fast random numbers with a chaotic laser.

The chaotic light signals generated by an external cavity semiconductor laser have been experimentally demonstrated to extract fast random numbers. However, the photon round-trip time in the external cavity can cause the occurrence of the periodicity in random sequences. To overcome it, the exclusive-or operation on corresponding random bits in samples of the chaotic signal and its time-delay signal from a chaotic laser is required. In this scheme, the proper selection of delay length is a key issue. By doing a large number of experiments and theoretically analyzing the interplay between the Runs test and the threshold value of the autocorrelation function, we find when the corresponding delay time of autocorrelation trace with the correlation coefficient of less than 0.007 is considered as the delay time between the chaotic signal and its time-delay signal, streams of random numbers can be generated with verified randomness.

Dissipation and dietary exposure risk assessment of pyraclostrobin, fluxapyroxad, difenoconazole, and azoxystrobin in the Fritillaria field ecosystem.

Fritillaria (Beimu in Chinese) is a well-known traditional Chinese medicinal herbal and valuable health food, which has attracted more and more attention. In this study, an efficient method was developed to determine pyraclostrobin, fluxapyroxad, difenoconazole, and azoxystrobin in plants, fresh Fritillaria, dry Fritillaria, and soil via liquid chromatography-tandem mass spectrometry. The average recoveries of the method were 78.9-109.7% with relative standard deviations of 0.94-11.1%. The dissipation half-lives of the four fungicides were 4.4-7.7 days in the Fritillaria plant and 11.6-18.2 days in the soil. The terminal residues of four fungicides were 0.033-0.13 mg/kg in fresh Fritillaria, 0.096-0.42 mg/kg in dry Fritillaria, and 0.12-0.74 mg/kg in soil. In the risk assessment of dietary exposure, all the chronic hazard quotient and acute hazard quotient index values were far below 100%, which were both acceptable to consumers. Accordingly, 7 days was recommended as the pre-harvest interval for the four fungicides in Fritillaria. This work could guide the safe use of these fungicides in Fritillaria and also give a reference for the Chinese government to establish the maximum residue limits (MRLs).

Guadipyr, a new insecticide, induces microbiota dysbiosis and immune disorders in the midgut of silkworms (Bombyx mori).

Guadipyr, which combines neonicotinoid and semicarbazone functional groups in one molecule, exhibits good activity on several pests and high acute and chronic toxicity to silkworms (Bombyx mori). In this report, the effects of low-dose guadipyr on the midgut microbiota and immune system of silkworms were studied. Results showed that the structure and richness of the midgut microbiota of silkworms were altered after being treated with 5.25 mg/L (1/10 of LC50) of guadipyr. The abundance of Pseudomonas was evidently increased, whereas Curvibacter was substantially reduced, which might be related to the growth and immunity of silkworms. The expression of key genes in the Toll, IMD, and JAK/STAT pathways, which ultimately led to the downregulation of antimicrobial peptide genes (AMPs), such as CecA, Defensin1, Leb, and glv2, was reduced upon guadipyr exposure. Simultaneously, the suppression of steroid hormone 20-hydroxyecdysone receptor and response genes, such as BR-C Z4, was detected in the exposed groups. The decreased expression of these immune regulatory pathway-related and 20-hydroxyecdysone signal pathway-related genes indicated that the immune system of silkworms was affected by low-dose guadipyr. Our results revealed the negative effects of guadipyr on silkworms and highlighted the unneglectable toxicity of low-dose guadipyr to this economic insect. Given the risk, it is necessary to control the application of guadipyr in or around the mulberry fields.

Combination of Urine Exosomal mRNAs and lncRNAs as Novel Diagnostic Biomarkers for Bladder Cancer.

BACKGROUND: The recent discovery of miRNAs and lncRNAs in urine exosomes has emerged as promising diagnostic biomarkers for bladder cancer (BCa). However, mRNAs as the direct products of transcription has not been well evaluated in exosomes as biomarkers for BCa diagnosis. The purpose of this study was to identify tumor progression-related mRNAs and lncRNAs in urine exosomes that could be used for detection of BCa. METHODS: RNA-sequencing was performed to identify tumor progression-related biomarkers in three matched superficial tumor and deep infiltrating tumor regions of muscle-invasive bladder cancer (MIBC) specimens, differently expressed mRNAs and lncRNAs were validated in TCGA dataset (n = 391) in the discovery stage. Then candidate RNAs were chosen for evaluation in urine exosomes of a training cohort (10 BCa and 10 healthy controls) and a validation cohort (80 BCa and 80 healthy controls) using RT-qPCR. The diagnostic potential of the candidates were evaluated by receiver operating characteristic (ROC) curves. RESULTS: RNA sequencing revealed 8 mRNAs and 32 lncRNAs that were significantly upregulated in deep infiltrating tumor region. After validation in TCGA database, 10 markedly dysregulated RNAs were selected for further investigation in urine exosomes, of which five (mRNAs: KLHDC7B, CASP14, and PRSS1; lncRNAs: MIR205HG and GAS5) were verified to be significantly dysregulated. The combination of the five RNAs had the highest AUC to disguising the BCa (0.924, 95% CI, 0.875-0.974) or early stage BCa patients (0.910, 95% CI, 0.850 to 0.971) from HCs. The expression levels of these five RNAs were correlated with tumor stage, grade, and hematuria degrees. CONCLUSIONS: These findings highlight the potential of urine exosomal mRNAs and lncRNAs profiling in the early diagnosis and provide new insights into the molecular mechanisms involved in BCa.

Glutaredoxin S25 and its interacting TGACG motif-binding factor TGA2 mediate brassinosteroid-induced chlorothalonil metabolism in tomato plants.

The presence of pesticide residues in fresh fruits and vegetables poses a serious threat to human health. Brassinosteroids (BRs) can reduce pesticide residues in plants, but the underlying mechanisms still remain unclear. Here, we identified a tomato glutaredoxin gene GRXS25 which was induced by 24-epibrassinolide (EBR) and chlorothalonil (CHT) in a way dependent on apoplastic reactive oxygen species (ROS). Silencing of GRXS25 in tomato abolished EBR-induced glutathione S-transferases (GSTs) gene expression and activity, leading to an increased CHT residue. Yeast two-hybrid and bimolecular fluorescence complementation assays showed protein-protein interaction between GRXS25 and a transcription factor TGA2. Electrophoretic mobility shift and chromatin immunoprecipitation assays indicated that TGA2 factor bound to the TGACG-motif in the GST3 promoter. While silencing of TGA2 strongly compromised, overexpression of TGA2 enhanced expression of GST genes and CHT residue metabolism. Our results suggest that BR-induced apoplastic ROS trigger metabolism of pesticide residue in tomato plants through activating TGA2 factor via GRXS25-dependent posttranslational redox modification. Activation of plant detoxification through physiological approaches has potential implication in improving the food safety of agricultural products.

Expression of vimentin in nasopharyngeal carcinoma and its possible molecular mechanism: A study based on immunohistochemistry and bioinformatics analysis.

BACKGROUND: Although previous researchers have analyzed the expression level of vimentin in nasopharyngeal carcinoma (NPC), the sample size of each study was too small, and there was no further in-depth study utilizing microarray and RNA-sequencing data. More importantly, the role and molecular mechanism of vimentin in NPC have not yet been addressed comprehensively. Accordingly, the aim of the present research was to conduct a full exploration of the clinical significance of vimentin in NPC in a large sample size. MATERIALS AND METHODS: Immunohistochemistry was used to test the expression of vimentin in clinical samples. Data from relevant microarray and RNA-sequencing datasets were screened and extracted to explore the clinical role of vimentin in NPC. Subsequently, vimentin-related signaling pathways were investigated via in-silico approaches. RESULTS: The clinical immunohistochemistry detection showed the positive expression ratio of vimentin was 24.6% (14/57) of the NPC specimens, whereas vimentin expression was negative in nasopharyngitis (NPG) tissues (0/20, P = 0.016). The mRNA and protein levels of vimentin were both remarkably up-regulated in NPC based on 196 and 1566 cases, respectively. The protein level of vimentin was also a risky factor for the prognosis prediction of NPC with the hazard ratios (HR) being 3.831. Gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) analyses, the localization of vimentin was in both the cytoplasm and the cytoskeleton, and vimentin was involved in the regulation of molecular function, the execution phase of apoptosis, and the regulation of cellular component organization. CONCLUSION: The high expression of vimentin plays a pivotal role in the development and poor progression of NPC, which indicates that vimentin may be an effective predictive indicator for NPC.

The clinical significance of CHEK1 in breast cancer: a high-throughput data analysis and immunohistochemical study.

Breast cancer (BC) is a kind of malignant cancer that seriously threatens women's health. Research scientists have found that BC occurs as the result of multiple effects of the external environment and internal genetic changes. Cell cycle checkpoint kinase 1 (CHEK1) is a crucial speed limit point in the cell cycle. Alterations of CHEK1 have been found in various tumors but are rarely reported or verified in BC. By mining database information, a large amount of mRNA and protein data was collected and meta-analyzed. Also, in-house immunohistochemistry was carried out to validate the results of the CHEK1 expression levels. Relative clinical features of BC patients were calculated with the CHEK1 expression levels to determine their diagnostic value. The mRNA levels of CHEK1 were higher in 1,089 cases of BC tissues than in 291 cases of non-BC tissues. We observed that the mRNA levels of CHEK1 are related to the clinical stages of BC patients (P = 0.008) and are also significant for overall survival (HR = 1.6, P = 0.0081). Using the immunohistochemistry method, we calculated and confirmed, using Fisher's exact test (P < 0.001), that a high-level CHEK1 protein is exhibited in BC tissues. Overexpressed CHEK1 mRNA promotes the occurrence of BC. Also, up-regulated CHEK1 could serve as an independent risk biomarker in BC patients' prognoses.

Clinical and prognostic value of chaperonin containing T-complex 1 subunit 3 in hepatocellular carcinoma: A Study based on microarray and RNA-sequencing with 4272 cases.

Liver cancer is one of the few tumors with a steadily increasing morbidity and mortality; hepatocellular carcinoma (HCC) is the most common type of primary liver cancer. We combined the expression profiles of Chaperonin Containing T-complex 1 Subunit 3 (CCT3) in HCC tissues based on microarray and RNA-sequencing data. The CCT3 expression levels were extracted and examined based on 421 samples from The Cancer Genome Atlas (TCGA) (HCC, n = 371; non-HCC, n = 50) and 3851 samples from 31 microarray or RNA-sequencing datasets (HCC, n = 1975; non-tumor = 1876). We used a variety of meta-analytic methods, including SMD forest maps, sensitivity analysis, subgroup analysis and sROC curves, to confirm the final results. Meanwhile, database-derived immunohistochemistry data was used for validation. We also further explained the potential mechanism of CCT3 in HCC through signal pathway analyses and PPI network construction with the CCT3 co-expressed genes. The mRNA and protein expression of CCT3 in HCC tissues were higher than in non-HCC tissues. The expression of CCT3 differed between groups when grouped according to clinicopathological parameters, such as race, family history, and histological grade. The results of standardised mean difference (SMD) forest map and summary receiver operating characteristic (sROC) curve revealed that CCT3 was highly expressed in HCC tissues and had a high ability to distinguish between cancer tissues and non-cancer tissues. The main form of CCT3 gene alteration in HCC was mRNA up-regulation and amplification (23%), and the most common mutation type was missense. The mRNA expression of CCT3 in HCC was negatively correlated with DNA methylation. According to the Kyoto Encyclopedia of Genes and Genomes pathway analysis, CCT3 can influence HCC occurrence and development through cell cycle and DNA replication pathways. In summary, this study carries out the staging and prognostic analysis of HCC. It suggests that CCT3 might play an important part in the tumorigenesis and progression of HCC and may have a certain prognostic value in HCC. Moreover, CCT3 might represent a promising biomarker for HCC.

Prospective molecular mechanism of COL5A1 in breast cancer based on a microarray, RNA sequencing and immunohistochemistry.

Breast cancer (BC) has a complex etiology and pathogenesis, and is the most common malignant tumor type in females, in USA in 2018, yet its relevant molecular mechanisms remain largely unknown. The collagen type V α­1 chain (COL5A1) gene is differentially expressed in renal and ovarian cancer. Using bioinformatics methods, COL5A1 was determined to also be a significant gene in BC, but its association with BC has not been sufficiently reported. COL5A1 microarray and relevant clinical data were collected from the Gene Expression Omnibus, The Cancer Genome Atlas and other databases to summarize COL5A1 expression in BC and its subtypes at the mRNA and protein levels. All associated information was comprehensively analyzed by various software. The clinical significance of the mutation was obtained via the cBioPortal. Furthermore, Gene Ontology functional annotation and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment were also performed to investigate the mechanism of COL5A1 in BC. Immunohistochemistry was also conducted to detect and confirm COL5A1 expression. It was determined that COL5A1 was highly expressed in BC tissues, compared with normal tissues at the mRNA level [standard mean difference, 0.84; 95% confidence interval (CI), 0.60­1.07; P=0.108]. The area under the summary receiver operator characteristic curve for COL5A1 was 0.87 (95% CI, 0.84­0.90). COL5A1 expression was altered in 32/817 (4%) sequenced samples. KEGG analysis confirmed the most notable pathways, including focal adhesion, extracellular matrix­receptor interaction and regulation of the actin cytoskeleton. Immunohistochemical detection was used to verify the expression of COL5A1 in 136 selected cases of invasive BC tissues and 55 cases of adjacent normal tissues, while the rate of high expression of COL5A1 in BC was up to 90.4%. These results indicated that COL5A1 is highly expressed at the mRNA and protein levels in BC, and the prognosis of patients with BC with high COL5A1 expression may be reduced; therefore, COL5A1 may be used independently or combined with other detection factors in BC diagnosis.

Glutaredoxin GRXS16 mediates brassinosteroid-induced apoplastic H2O2 production to promote pesticide metabolism in tomato.

Brassinosteroids (BRs), a group of steroid phytohormones, are involved in multiple aspects of plant growth, development and stress responses. Despite recent studies on BRs-promoted pesticide metabolism in plants, the underlying mechanisms remain poorly understood. Here, we showed that 24-epibrassinolide (EBR) significantly enhanced the expression of RESPIRATORY BURST OXIDASE HOMOLOG1 (RBOH1) and H2O2 accumulation in the apoplast of chlorothalonil (CHT, a broad spectrum nonsystemic fungicide)-treated tomato plants. Silencing of RBOH1 significantly decreased the efficiency of EBR-induced CHT metabolism. Moreover, the EBR-induced upregulation in the transcripts of glutaredoxin gene GRXS16 was suppressed in RBOH1-silenced plants. Further studies indicated that silencing of GRXS16 compromised EBR-induced increases in glutathione content, activity of glutathione S-transferase (GST) and transcript of GST1, leading to an increase in CHT residue. By contrast, overexpression of tomato GRXS16 enhanced the basal levels of glutathione content and GST activity that eventually decreased CHT residues in transgenic plants. Our results reveal that BR-mediated induction of a modest oxidative burst is essential for the acceleration of glutathione-dependent pesticide metabolism via redox modulators, such as GRXS16. These findings shed new light on the mechanisms of BR-induced pesticide metabolism and thus have important implication in reducing pesticide residues in agricultural products.