The mechanism of Gejie Zhilao Pill in treating tuberculosis based on network pharmacology and molecular docking verification.

Introduction: Gejie Zhilao Pill (GJZLP), a traditional Chinese medicine formula is known for its unique therapeutic effects in treating pulmonary tuberculosis. The aim of this study is to further investigate its underlying mechanisms by utilizing network pharmacology and molecular docking techniques. Methods: Using TCMSP database the components, potential targets of GJZLP were identified. Animal-derived components were supplemented through the TCMID and BATMAN-TCM databases. Tuberculosis-related targets were collected from the TTD, OMIM, and GeneCards databases. The intersection target was imported into the String database to build the PPI network. The Metascape platform was employed to carry out Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Heatmaps were generated through an online platform (https://www.bioinformatics.com.cn). Molecular docking was conducted between the core targets and core compounds to explore their binding strengths and patterns at the molecular level. Results: 61 active ingredients and 118 therapeutic targets were identified. Quercetin, Luteolin, epigallocatechin gallate, and beta-sitosterol showed relatively high degrees in the network. IL6, TNF, JUN, TP53, IL1B, STAT3, AKT1, RELA, IFNG, and MAPK3 are important core targets. GO and KEGG revealed that the effects of GJZLP on tuberculosis mainly involve reactions to bacterial molecules, lipopolysaccharides, and cytokine stimulation. Key signaling pathways include TNF, IL-17, Toll-like receptor and C-type lectin receptor signaling. Molecular docking analysis demonstrated a robust binding affinity between the core compounds and the core proteins. Stigmasterol exhibited the lowest binding energy with AKT1, indicating the most stable binding interaction. Discussion: This study has delved into the efficacious components and molecular mechanisms of GJZLP in treating tuberculosis, thereby highlighting its potential as a promising therapeutic candidate for the treatment of tuberculosis.

ELANE is a promising prognostic biomarker that mediates pyroptosis in gastric cancer.

Background: Gastric cancer (GC) is a typical malignant tumor and the main cause of cancer-related deaths. Its pathogenesis involves multiple steps, including pyroptosis, although these steps are still uncertain. Pyroptosis, also known as gasdermin-mediated programmed necrosis, participates in various pathological processes in tumors, including GC. ELANE, which encodes neutrophil elastase, is closely associated with GC. Additionally, ELANE has been implicated in GC cell pyroptosis, but this has not been confirmed. Therefore, investigating the link between ELANE and pyroptosis in GC is warranted. This research uses bioinformatics and experiments to examine the relationship between ELANE, pyroptosis, and GC prognosis. Methods: The GEO and TCGA databases, along with pyroptosis-related genes, were applied to identify pyroptosis-related differentially expressed genes (DEGs). ELANE was selected via primary screening. Using the median expression level of ELANE as the threshold, pyroptosis-related DEGs were divided into low- and high-ELANE groups. Based on the DEGs in these two groups, GO, KEGG and GSEA analyses were conducted to elucidate the mechanisms of ELANE in GC. Furthermore, we plotted ROC and Kaplan-Meier curves to analyze the clinical and pathological features of ELANE expression. The Nomograms tool was applied to calculate the predictive value of ELANE for the clinical outcomes of GC cases. Immunohistochemical analysis was performed to detect the level of ELANE in GC tissues and to validate whether ELANE was involved in pyroptosis in GC cells through cell experiments. Finally, the immune infiltration of ELANE was investigated, and interaction networks (proteins-ELANE, microRNA-ELANE, and small-molecule drug-ELANE) were constructed. Results: We aimed to investigate the expression of the ELANE gene in GC and study the relationship among ELANE, pyroptosis, and the prognosis of patients with GC. Differential expression analysis of gene-expression datasets from TCGA-STAD and GSE49051 revealed that the expression of the ELANE gene was significantly up-regulated in GC. Using STRING network analysis, we identified multiple proteins involved in the occurrence and development of GC, including interactions between ELANE and GSDMC, a member of the gasdermin protein family. Survival analysis showed that ELANE expression levels significantly affected overall survival (OS), disease-free survival (DFS), and progression-free survival (PFS) in patients with GC. Additionally, ROC analysis demonstrated that ELANE was effective in distinguishing GC patients from normal controls (AUC = 0.812). Immunohistochemical analysis showed that ELANE was highly expressed in gastric cancer tissues and was closely related to age, tumor grade, and stage. The cell experiments further confirmed that the high expression of ELANE in gastric cancer cells was associated with pyroptosis. Comprehensive analysis indicated that ELANE could be used as a potential prognostic marker for GC and plays an important role in pyroptosis. Conclusion: High ELANE expression is related to poor survival and prognosis of patients with GC. It participates in pyroptosis and immune infiltration in GC. Therefore, ELANE is a promising prognostic biomarker for pyroptosis in GC.

Multi-omics analysis of pyroptosis regulation patterns and characterization of tumor microenvironment in patients with hepatocellular carcinoma.

Background: Hepatocellular carcinoma (HCC) is a primary malignant tumor of the liver, and pyroptosis has been identified as a novel cellular program that plays a role in numerous diseases including cancer. However, the functional role of pyroptosis in HCC remains unclear. The purpose of this study is to explore the relationship between the two found hub genes and provide targets for clinical treatment. Methods: The Cancer Genome Atlas (TCGA) database was used to collect the gene data and clinically-related information of patients with HCC. After the differentially expressed genes (DEGs) were identified, they were intersected with the genes related to pyroptosis, and a risk prediction model was established to predict the overall survival (OS). Subsequently, drug sensitivity analysis, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Gene Set Enrichment Analysis (GSEA), and Gene Set Variation Analysis (GSVA) was used to analyze the biological characteristics of the DEGs. Different immune cell infiltration and related pathways were analyzed, and hub genes were identified by protein-protein interaction (PPI). Finally, the expression of hub genes was verified by real-time quantitative PCR (qRT-PCR) and immunohistochemistry. Results: We conducted a comprehensive bioinformatics analysis to investigate the molecular mechanisms of pyroptosis in hepatocellular carcinoma (HCC). A total of 8,958 differentially expressed genes were identified, and 37 differentially expressed genes were associated with pyroptosis through intersection. Moreover, we developed an OS model with excellent predictive ability and discovered the differences in biological function, drug sensitivity, and immune microenvironment between high-risk and low-risk groups. Through enrichment analysis, we found that the differentially expressed genes are related to various biological processes. Then, 10 hub genes were identified from protein-protein interaction networks. Finally, midkine (MDK) was screened from the 10 hub genes and further verified by PCR and immunohistochemistry, which revealed its high expression in HCC. Conclusion: We have developed a reliable and consistent predictive model based on the identification of potential hub genes, which can be used to accurately forecast the prognosis of patients, thus providing direction for further clinical research and treatment.

The functions of N6-methyladenosine (m6A) RNA modifications in colorectal cancer.

Colorectal cancers (CRC), which includes colon cancer (CC) and rectal cancer (RC), are some of the most common malignant tumors that are prone to distant metastasis. Its high incidence rate and high mortality rate have attracted much attention. In recent years, epigenetics has attracted increasing attention and has been the focus of many research studies. N6-methyladenosine(m6A) RNA modifications can modify eukaryotic mRNA to impact metabolism. The changes in the m6A regulatory genes are related to the occurrence and development of CRC and play an important role in the pathogenesis of CRC. The effect of m6A RNA modification is regulated by its related regulatory factors ("writer", "eraser", "reader"). In this review, we comprehensively analyzed the effect of m6A methylation on CRC and the relationship between the expression of related regulatory factors and the development and occurrence of CRC. Then, we summarized the roles of m6A and its regulatory factors in CRC and its potential clinical value, which provides a basis for further research on the mechanism of m6A methylation in CRC.

Ano1 is a Prognostic Biomarker That is Correlated with Immune Infiltration in Colorectal Cancer.

BACKGROUND: Anoctamin 1 (ANO1) has been observed to be overexpressed in gastrointestinal and pulmonary epithelial cells, as well as in a number of cancers. Although Ano1 is involved in the prognosis of colorectal cancer (CRC), its mechanism of action in metastatic CRC has not been fully elucidated. METHODS: The expression of Ano1 was assessed in samples obtained from The Cancer Genome Atlas (TCGA) database. Then, we used Gene Ontology (GO) enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, Gene set enrichment analysis (GSEA), Gene set variation analysis (GSVA), and Weighted Correlation Network Analysis (WGCNA) to determine the functions of Ano1. Additionally, random survival forest, Cox multivariate analysis, Kaplan Meier analysis, and ROC were used to determine the predictive value of Ano1 on clinical outcomes in CRC patients. Finally, HE staining, immunohistochemical (IHC) analysis and qRT-PCR were used to explore the expression of the Ano1 gene in CRC tissue. RESULTS: The expression level of Ano1 in CRC was significantly elevated, and the prognosis was poor. The modules with a higher proportion of upregulated genes tended to be positively correlated with Ano1-high. KNG1, GNG4, F2, POSTN, THBS2, SPP1 and FGA were identified as hub proteins of the PPI network. The heatmap showed that the expression level of the Ano1-high group was significantly negatively correlated with immune infiltrate. The overexpression of the Ano1 gene in CRC tissue samples was also confirmed by HE staining, immunohistochemical (IHC) analysis and qRT-PCR. CONCLUSION: High expression of Ano1 is closely related to a poor prognosis in patients with colorectal cancer. Ano1 may participate in the metastasis and progression, as well as the immune regulation of CRC. In summary, Ano1 can act as a potential prognostic biomarker and a novel target for CRC therapy.

Significant correlation between HSPA4 and prognosis and immune regulation in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is an inflammation-associated tumor involved in immune tolerance and evasion in the immune microenvironment. Heat shock proteins (HSPs) are involved in the occurrence, progression, and immune regulation of tumors. Therefore, HSPs have been considered potential therapeutic targets. Here, we aimed to elucidate the value of HSP family A (Hsp70) member 4 (HSPA4) in the diagnosis and predicting prognosis of HCC, and its relationship with immune cell infiltration, immune cell biomarkers, and immune checkpoints. Gene mutation, DNA methylation, and the pathway involved in HCC were also analyzed. METHODS: The gene expression omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases were used to compare HSPA4 expression, and the results were confirmed by immunohistochemical staining of clinical samples. R package was used to analyze the correlation between HSPA4 and cancer stage, and to establish receiver operating characteristic (ROC) curve of diagnosis, time-dependent survival ROC curve, and a nomogram model. cBioPortal and MethSurv were used to identify genetic alterations and DNA methylation, and their effect on prognosis. The Tumor Immune Estimation Resource (TIMER) was used to analyze immune cell infiltration, immune cell biomarkers, and immune checkpoints. The STRING database was used to analyze protein-protein interaction network information. Gene Ontology (GO) analysis and the Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to investigate the functions of HSPA4 and its functional partner genes. RESULTS: Overexpression of HSPA4 was identified in 25 cancers. Overexpression of HSPA4 considerably correlated with cancer stage and alpha-fetoprotein (AFP) level in HCC. Patients with higher HSPA4 expression showed poorer prognosis. HSPA4 expression can accurately identify tumor from normal tissue (AUC = 0.957). The area under 1-, 3-, and 5-year survival ROCs were above 0.6. The HSPA4 genetic alteration rate was 1.3%. Among the 14 DNA methylation CpG sites, seven were related to the prognosis of HCC. HSPA4 was positively related to immune cell infiltration and immune checkpoints (PD-1 and CTLA-4) in HCC. The KEGG pathway enrichment analysis revealed HSPA4 enrichment in antigen processing and presentation together with HSPA8 and HSP90AA1. We verified the value of HSPA4 in the diagnosis and predicting prognosis of HCC. HSPA4 may not only participate in the occurrence and progression but also the immune regulation of HCC. Therefore, HSPA4 can be a potential diagnostic and prognostic biomarker and a therapeutic target for HCC.

Heat Shock Protein Beta 1 is a Prognostic Biomarker and Correlated with Immune Infiltrates in Hepatocellular Carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most serious malignancies. The main features of HCC are vascular invasion and drug resistance. Ferroptosis is a novel cell program that is involved in several diseases, such as cancer. Heat shock protein beta 1 (HSPB1) is a major component of heat shock proteins. A recent study showed that HSPB1 could be a new therapeutic target for colorectal cancer with 5-fluorouracil-acquired resistance. However, the functional role of HSPB1 in HCC remains unclear. AIM: The aim of this study is to clarify HSPB1 expression in HCC and its potential therapeutic and prognostic value. METHODS: We collected data on HSPB1 expression levels in HCC and normal liver tissues from The Cancer Genome Atlas and Gene Expression Omnibus databases. We then validated it using immunohistochemistry (IHC). Receiver operating characteristic and Kaplan-Meier survival curves were used to investigate the role of HSPB1 in the prognosis analysis of HCC. Further, we used the online Search Tool for the Retrieval of Interacting Genes/Proteins website, Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes to conduct enrichment analysis and identify the predictive signaling pathways. Meanwhile, we used the TIMER and GSVA package of R (v3.6.3) to analyze the association between HSPB1 and immunocyte infiltration. RESULTS: Compared to normal tissues, there was differential expression of HSPB1 in pan-cancers. HSPB1 expression was higher in HCC tissues than in normal tissues (p<0.05). There was an evident significant difference between HSPB1 mRNA levels and histologic grade, vascular invasion, and alpha-fetoprotein level (all p values<0.05). Univariate analysis indicated that HCC patients with high HSPB1 levels had shorter overall survival rates than those with low HSPB1 levels (p<0.05). MAPK14, HSPA8, MAPKAPK3, MAPKAPK5, and MAPKAPK2 are essential proteins that interact with HSPB1. There was a significant correlation between HSPB1 expression levels and immune cell infiltration, including CD4+ T cells (r=0.203, p<0.05). CONCLUSION: High HSPB1 expression is closely associated with a worse prognosis in HCC patients, and HSPB1 may be a target of immunotherapy in HCC.

An immunomagnetic separation and bifunctional Au nanoparticle probe-based multiamplification electrochemical strategy.

A novel electrochemical magnetoimmunosensor for the rapid and sensitive detection of carcinoembryonic antigen (CEA) was fabricated based on a combination of high-efficiency immunomagnetic separation, bifunctional Au-nanoparticle (bi-AuNP) probes, and enzyme catalytic amplification. The reaction carrier magnetic beads (MBs) effectively reduced the toxicity of the complex sample to the working electrode, and the signal carrier bi-AuNP probes loaded a large amount of signal molecules, both of which enhanced the signal-to-noise ratio and further improved the detection sensitivity. A detection limit as low as 0.11 pg/mL was achieved for CEA detection based on the immunomagnetic separation and bi-AuNP probe-based multiamplification strategy, and the strategy was further successfully applied in human serum samples. The transducer was regenerated via a simple washing procedure, which enabled the detection of all samples on a single electrode with high reproducibility. The proposed strategy, which has the merits of high sensitivity, selectivity, and reproducibility exhibits great potential for detection in complex samples.

Synthesis of a new chitosan derivative and assay of Escherichia coli adsorption.

A new chitosan derivative is prepared using chitosan. Ethyl cholorocarbonate was first introduced to the hydroxyl group of phthaloylchitosan through a nucleophilic reaction. Hydrazine was then added to recover the amino groups of chitosan, and promote cross-linking. The structure of this new chitosan derivative was characterized by Fourier transform infrared (FT-IR) and proton nuclear magnetic resonance (1H NMR) spectroscopy, and its physical properties were determined by X-ray diffraction (XRD), differential scanning calorimetry (DSC), and thermogravimetric analysis (TGA). The thermal and chemical stabilities of the new derivative were improved compared with those of native chitosan. Assay of Escherichia coli adhesion on a film based on this chitosan derivative showed good adsorption and biofilm formation.

Superparamagnetic microsphere-assisted fluoroimmunoassay for rapid assessment of acute myocardial infarction.

Rapid assessment of acute myocardial infarction (AMI) was successfully demonstrated using an improved superparamagnetic polymer microsphere-assisted sandwich fluoroimmunoassay to detect two early cardiac markers-myoglobin and human heart-type fatty acid binding protein (H-FABP). This assay used a preparation of superparamagnetic poly(styrene-divinylbenzene-acrylamide) microspheres, glutaraldehyde-coupled capture antibodies (monoclonal anti-myoglobin 7C3 and anti-H-FABP 10E1) grafted onto the polymer microspheres, and a sequential sandwich fluoroimmunoassay using detection antibodies (FITC-labeled anti-myoglobin 4E2 and FITC-labeled anti-H-FABP 9F3). Characterization of the polymer microspheres by TEM, SEM and Fourier transform infrared spectroscopy (FT-IR) showed that the microspheres were uniformly round with an average diameter of 1.12 microm, and had a Fe(3)O(4)-polymer core-shell structure (shell thickness was about 84 nm) with 0.22 mmol/g amino groups on their surfaces. The magnetic behavior of the Fe(3)O(4)-polymer microspheres was superparamagnetic (M(s)=13 emu/g, H(c)=13.1 Oe). Fluorescence images of the post-immunoassay microspheres recorded using a confocal laser-scanning microscope showed that the average fluorescence intensity was correlated with the concentration of cardiac markers, in agreement with the results obtained by an F-4500 FL spectrophotometer; this indicated that the fluoroimmunoassay could be used to semi-quantitatively detect both myoglobin and H-FABP. The detection limit was 25 ng/mL for myoglobin and 1 ng/mL for H-FABP.