Recognizing the role of Epstein-Barr virus in gastric cancer: transcriptomic insights into malignancy modulation.

BACKGROUND: Studies show that Epstein-Barr virus (EBV) infection can play a role in malignancy and increase the risk of gastric cancer (GC). The objective of this research was to pinpoint genes whose expression may be influenced by EBV and play a role in the development of GC. METHODS: Candidate genes potentially susceptible to expression modulation in the presence of EBV were identified through the analysis of GSE185627 and GSE51575 datasets. The association of candidate genes with GC and the survival rate of patients was investigated based on the cancer genome atlas (TCGA) data. Also, pathways related to candidate genes were examined through the MsigDB database. The PPI network was used to identify Hub genes. To corroborate the obtained results, we utilized the RT-qPCR method, employing GC samples from both EBV + and EBV-cases, as well as adjacent normal samples. RESULTS: Our results showed that genes upregulated by the EBV in the GC cell line, as well as in EBV + samples, are significantly linked to pathways involving the immune response, inflammation, and the P53 pathway. Conversely, genes downregulated by EBV are closely linked to pathways involving cell proliferation and mTORC1. Examining the candidate genes revealed that a considerable portion of genes susceptible to downregulation under the influence of EBV exhibit oncogenic roles based on TCGA data. Moreover, some of these genes are associated with an unfavorable prognosis. Protein-protein interaction network analysis of candidate genes highlighted IFI44L and OAS2 as potential hub genes in the EBV-GC axis. Our RT-qPCR results further validated these findings, demonstrating that the expression levels of IFI44L and OAS2 were higher in EBV + samples compared to both healthy and EBV-samples. CONCLUSION: Our study underscores the capacity of EBV to exert regulatory control over genes associated with GC malignancy. In addition to its inflammatory effects, EBV elicits transcriptomic changes that appear to attenuate the progression of GC.

Prognostic significance of CHCHD2P9 and ZNF204P in breast cancer: exploring their expression patterns and associations with malignancy-related genes.

BACKGROUND: Non-coding RNAs (ncRNAs) have a crucial impact on diverse cellular processes, influencing the progression of breast cancer (BC). The objective of this study was to identify novel ncRNAs in BC with potential effects on patient survival and disease progression. METHODS: We utilized the cancer genome atlas data to identify ncRNAs associated with BC pathogenesis. We explored the association between these ncRNA expressions and survival rates. A risk model was developed using candidate ncRNA expression and beta coefficients obtained from a multivariate Cox regression analysis. Co-expression networks were constructed to determine potential relationships between these ncRNAs and molecular pathways. For validation, we employed BC samples and the RT-qPCR method. RESULTS: Our findings revealed a noteworthy increase in the expression of AC093850.2 and CHCHD2P9 in BC, which was correlated with a poor prognosis. In contrast, ADAMTS9-AS1 and ZNF204P displayed significant downregulation and were associated with a favorable prognosis. The risk model, incorporating these four ncRNAs, robustly predicted patient survival. The co-expression network showed an effective association between levels of AC093850.2, CHCHD2P9, ADAMTS9-AS1, and ZNF204P and genes involved in pathways like metastasis, angiogenesis, metabolism, and DNA repair. The RT-qPCR results verified notable alterations in the expression of CHCHD2P9 and ZNF204P in BC samples. Pan-cancer analyses revealed alterations in the expression of these two ncRNAs across various cancer types. CONCLUSION: This study presents a groundbreaking discovery, highlighting the substantial dysregulation of CHCHD2P9 and ZNF204P in BC and other cancers, with implications for patient survival.

Genome editing approaches with CRISPR/Cas9: the association of NOX4 expression in breast cancer patients and effectiveness evaluation of different strategies of CRISPR/Cas9 to knockout Nox4 in cancer cells.

BACKGROUND: The increasing prevalence of cancer detection necessitated practical strategies to deliver highly accurate, beneficial, and dependable processed information together with experimental results. We deleted the cancer biomarker NOX4 using three novel genetic knockout (KO) methods. Homology-directed repair (HDR), Dual allele HITI (Du-HITI) and CRISPR-excision were utilized in this study. METHODS: The predictive value of the NOX4 expression profile was assessed using a combined hazard ratio (HR) with a 95% confidence interval (CI). With a 95% confidence interval, a pooled odd ratio (OR) was used to calculate the relationship between NOX4 expression patterns and cancer metastasis. There were 1060 tumor patients in all sixteen research that made up this meta-analysis. To stop the NOX4 from being transcribed, we employed three different CRISPR/Cas9-mediated knockdown methods. The expression of RNA was assessed using RT-PCR. We employed the CCK-8 assay, colony formation assays, and the invasion transwell test for our experiments measuring cell proliferation and invasion. Using a sphere-formation test, the stemness was determined. Luciferase reporter tests were carried out to verify molecular adhesion. Utilizing RT-qPCR, MTT, and a colony formation assay, the functional effects of NOX4 genetic mutation in CRISPR-excision, CRISPR-HDR, and CRISPR du-HITI knockdown cell lines of breast cancer were verified. RESULTS: There were 1060 malignant tumors in the 16 studies that made up this meta-analysis. In the meta-analysis, higher NOX4 expression was linked to both a shorter overall survival rate (HR = 1.93, 95% CI 1.49-2.49, P < 0.001) and a higher percentage of lymph node metastases (OR = 3.22, 95% CI 2.18-4.29, P < 0.001). In breast carcinoma cells, it was discovered that NOX4 was overexpressed, and this increase was linked to a poor prognosis. The gain and loss-of-function assays showed enhanced NOX4 breast carcinoma cell proliferation, sphere-forming capacity, and tumor development. To activate transcription, the transcriptional factor E2F1 also attaches to the promoter region of the Nanog gene. The treatment group (NOX4 ablation) had substantially more significant levels of proapoptotic gene expression than the control group (P < 0.01). Additionally, compared to control cells, mutant cells expressed fewer antiapoptotic genes (P < 0.001). The du-HITI technique incorporated a reporter and a transcription termination marker into the two target alleles. Both donor vector preparation and cell selection were substantially simpler using this approach than with "CRISPR HDR" or "CRISPR excision." Furthermore, single-cell knockouts for both genotypes were created when this method was applied in the initial transfection experiment. CONCLUSIONS: The NOX4 Knockout cell lines generated in this research may be used for additional analytical studies to reveal the entire spectrum of NOX4 activities. The du-HITI method described in this study was easy to employ and could produce homozygous individuals who were knockout for a specific protein of interest.

Molecular Basis of Bicalutamide Response Alteration of Androgen Receptor Caused by Single Nucleotide Polymorphisms: An In Silico Investigation.

The vast majority of drugs act through binding to their protein targets. Prediction of the interaction between small molecules and these receptors is a key element in the process of drug discovery. Advances in structural biology have enabled us to resolve the three-dimensional structure of proteins, which are the targets of the drugs. Pharmacogenetics also helped researchers to study the structural variations arise from the single nucleotide polymorphisms (SNPs) and to survey the effects these variations in drug design and development. These improvements led to the identification of structural changes caused by SNPs, which affect the drug interaction with their receptors, called drug response. In this study, the interaction between androgen receptor and bicalutamide was investigated using a computational analysis. The results of these analyses were then used for identification of nonsynonymous SNPs that are potentially involved in drug response alterations. The data show that amino acids Met895, Trp741, Arg752, Ile899, Leu707, Gly708, Gln711, Met745, Met749, Thr877, Phe764, Met742, Asn705 and Leu704 are the main residues involved in the interaction between androgen receptor and bicalutamide. The occurrence of nonsynonymous polymorphisms I843T, L708R, H690P, I870M, N757S, L713F, G744E, L678P, M788V, M781I, A722T, H875Y, I842V, and F827L in this receptor greatly affected its interaction with bicalutamide, and they were able to cause drug resistance. The results of this study could be useful in predicting the response to treatment in patients receiving bicalutamide.

miR-455-5p downregulation promotes inflammation pathways in the relapse phase of relapsing-remitting multiple sclerosis disease.

MicroRNA-455-5p (miR-455-5p) seems to have an anti-inflammatory role in the immune system since its expression is induced by IL-10 cytokine. Multiple sclerosis (MS) is a chronic demyelinating neurodegenerative disease of the central nervous system that is caused by an autoimmune inflammatory attack against the myelin insulation of neurons. The expression level of miR-455-5p and its role in MS pathogenesis has yet to be elucidated. We found that miR-455-5p expression was highly correlated with disease severity in MS patients. miR-455-5p expression inversely correlates with its inflammatory-predicted targets (MyD88 and REL) in relapse- and remitting-phase patients. Luciferase assays confirm that MyD88 and REL are direct targets of miR-455-5p. This study represents the first report of the miR-455-5p acts as an anti-inflammatory role in MS, at least partially through targeting MyD88 and REL. This study may provide important information for the use of miR-455-5p as a novel strategy to improve the severity of disease and control inflammation and attack in MS patients.

Functional SNP in microRNA-491-5p binding site of MMP9 3'-UTR affects cancer susceptibility.

MicroRNAs (miRNA) are small RNA molecules that negatively regulate gene expression through base pairing interactions between 3'-UTR of the target mRNAs and seed sequence of miRNA. Any changes in the recognition site could destroy binding sites or modify binding affinity, resulting in evasion from miRNA regulation. A putative binding site for miR-491-5p resides in 3'-UTR of MMP9, and a genetic variant (rs1056628 A → C) is present in this region. The role of MMP9 over expression well marked in various cancers. However, whether rs1056628 SNP in miR-491-5p binding site of MMP9 3'-UTR could abrogate its post-transcriptional regulation and affect cancer susceptibility remains largely unknown. To test this, the rs1056628 SNP was genotyped in 300 cases of lung, gastric and breast cancers and 200 age- and sex-matched healthy controls. The results showed that compared with the AA genotype, C was a risk genotype for all three cancers development and was also associated with gastric and breast cancers metastasis and invasion. Based on the base pairing analysis and secondary structure evaluation of MMP9 mRNA and miR-491-5p, we found that miR-491-5p had a higher binding affinity for A genotype than the C genotype. The Luciferase activity of MMP9 3'-UTR indicates differential regulation of two genetic variations of MMP9. Overexpression of miR-491-5p decreased MMP9 mRNA level in cell lines of gastric, breast and lung cancers and thus leads to decreasing of the invasion ability. Therefore, for the first time we imply that the C variant of MMP9 contributes to the likelihood of gastric, breast and lung cancers susceptibility via a novel mechanism of subtle gene regulation through miRNA binding capacity.

Unlocking the potential of Escherichia coli K-12: A novel approach for malignancy reduction in colorectal cancer through gene expression modulation.

Studies have noted the association between Escherichia coli K-12 (E. coli K-12) and the reduction of malignancy in colorectal cancer (CRC). However, the molecular mechanisms underlying this relationship have not been thoroughly explored. The aim of this study was to identify the genes influenced by E. coli K-12 and their connection to CRC. We identified the genes affected by E. coli K-12 using the GSE50040 dataset. Additionally, we investigated the relationship between the expression of genes affected by E. coli K-12 and CRC using the cancer genome atlas data. The association between the expression of E. coli K-12-affected genes and patient prognosis was investigated using clinical data. Pathways related to CRC and E. coli K-12-related genes were analyzed using the Enrichr tool. Furthermore, we employed a protein-protein interaction (PPI) network to identify hub genes associated with both E. coli K-12 and CRC. To validate our findings, we conducted RT-qPCR analysis on CRC samples and adjacent normal tissue. The results of GSE50040 showed that E. coli K-12 could change the expression of many genes related to CRC in colorectal cell lines. The results showed that E. coli K-12 reduces the expression of several genes linked to the main pathways used by cancer cells, such as the metastasis, WNT, cell proliferation pathway, and mTORC1. It was demonstrated that elevated BGN, FJX1, and LZTS1 expression is linked to a bad prognosis in patients and that E. coli K-12 may be able to lower this expression. Also, based on the PPI network, genes such as KLF4 and CXCL3 were identified as hub genes related to genes affected by E. coli K-12. When KLF4 and CXCL3 expression levels in cancer samples were compared to nearby normal tissue, a significant change in these genes' expression levels was found in CRC. Our findings demonstrated the potential relationship between oncogene genes and genes impacted by E. coli K-12. Also, our findings demonstrated that E. coli K-12 may regulate the expression of genes linked to a high death rate. In summary, the results of this study suggest that E. coli K-12 can be regarded as a significant probiotic with the potential to mitigate the risk of CRC development.

Impact of NOX4 Knockout by CRISPR/Cas9 on the MCF-7, HCA-7 and UM-RC-6 Cancer Cells.

Background: The second most common cause of mortality is cancer. Increased NOX4 expression is linked to cancer development and metastasis. However, the significance of NOX4 in cell growth and assault, remains unclear. Objective: This study aimed to evaluate the effect of NOX4 knockouts in MCF7, UM-RC-6, HCA-7 cell lines. Materials and Methods: The NOX4 gene was knocked out in MCF7, UM-RC-6, and HCA-7 cell lines through using CRISPR Cas-9 genetic engineering techniques. After transfection, the CRISPR Cas-9 cassette, the T7 endonuclease I, qPCR, and western blotting assay detected the NOX4 knockouts. MTT and Annexin assessed the percentage of cell proliferation and apoptosis. Real-time PCR was used to measure the expression of pro- and anti-apoptotic genes. Results: Occurrence of NOX4 gene knockout in the examined cell lines, was confirmed by q-PCR and Western blot (P<0.001). The NOX4-deleted cell lines with increased sub-G1 caused lowered cell proliferation and population at S / G2/ M phases. In Vitro, NOX4 silencing caused lowered expressions of anti-apoptosis genes BCL-2 and SURVIVIN (P<0.0001), leading to increased tendency of apoptosis in the cell lines (P<0.0001) of the apoptotic genes BAX, P53, FAS. Additionally, the MTT and Annexin results of the target gene NOX4 knockout inhibited proliferation, increased mortality rates (P<0.01), and increased apoptosis. Conclusion: The findings of this study indicate that using NOX4 as a target can have therapeutic value for creating potential treatments against breast, colorectal, and kidney cancers which shows a need for a deeper understanding of the biology of these cancers with direct clinical outcomes for developing novel treatment strategies.

Evaluation of the Immune Response Against Helicobacter pylori; in Infused BALB/c Mice by pcDNA3.1(+)-ureA

BACKGROUND: The purpose of the present study was to produce a pcDNA3.1(+)-ureA recombinant vector and evaluate the capacity of this vector to stimulate the immune response against H. pylori infection in infused BALB/c mice. MATERIALS AND METHODS: The pcDNA3.1(+)-ureA construct was prepared and transformed into E. coli, successfully. The animals we used in the study were allotted into three groups for infusion of 1) recombinant plasmid, 2) pcDNA3.1(+)-ureA + nanoparticles, and 3) pcDNA3.1(+). Blood and tissue specimens from each group of mice were collected at days 15, 30, and 45 after the last infusion and the expression levels of cytokines such as TGF-ß1, IL-4, and IFNγ genes comparing to GAPDH as well as the expression of ureA in the mice's thigh muscle were evaluated. RESULTS: The genes expression analysis showed that the IL4 expression significantly decreased (p<0.001) but IFNγ and TGF-ß1 expression increased in the blood of infused mice (p<0.001). Also, the urea expression level in pcDNA3.1(+)-urea and pcDNA3.1(+)-ureA+ nanoparticle 15, 30, and 45 days after the last infusion was significantly different (p<0.001) and its expressions at days 15 and 30 were significantly different (p<0.001), but 45 days after the last infusion it was not significantly different (p>0.05). CONCLUSION: The pcDNA3.1(+)-ureA recombinant vector with or without chitosan nanoparticles can stimulate the immune response in animal models against H. pylori infection. Also, after combining the recombinant vector with nanoparticles we observed a better immune response was observed. In future studies this recombinant construct can be used as a biomarker and therapeutic approaches in eukaryotic systems.