PPI Network Research Articles

12353 Differential Gene Expression And Pathway Analysis In Growth Hormone Secreting Pituitary Tumors According To Granulation Pattern

Abstract Disclosure: H. Shin: None. Differential gene expression and pathway analysis in growth hormone-secreting pituitary tumors according to granulation pattern. Hye Ju Shin [1], Kyung Won Kim[2], Chan Woo Kang[2], Eun Jig Lee[2], Cheol Ryong Ku[2]. [1]Department of Clinical Drug Discovery & Development, Yonsei University College of Medicine, Seoul, Republic of Korea, [2]Endocrinology, Institute of Endocrine Research, Department of Internal Medicine, Yonsei University College of Medicine, Seoul, Republic of Korea Objective: The purpose of this research is to explain clinical variations in patients with acromegaly by exploring gene expression differences according to granulation patterns in growth hormone (GH)-secreting pituitary tumors. Materials & methods: Transcriptome analysis was conducted on 6 normal pituitary tissues and 15 GH-secreting pituitary tumors, including 9 densely granulated somatotroph tumors (DGSTs) and 6 sparsely granulated somatotroph tumors (SGSTs). Results: The median age at diagnosis was 50 years, with a predominance of female patients (60%). We identified 3111 differentially expressed genes (DEGs) in tumors compared to normal pituitaries, with 1117 DEGs unique to a specific granulation within tumors. SGST showed enriched the pathways involved in neuronal development (axon development, axonogenesis, axon guidance, synapse and presynapse organization, gliogenesis, glial cell differentiation, nerve development, synaptic membrane adhesion, semaphorin-plexin signaling pathway involved in neuron projection guidance) and acute inflammatory response. No pathways were significantly downregulated in SGST compared with those in DGST. Three signaling pathways (JAK-STAT, phosphatidylinositol 3-kinase, and MAP cascade) and three neuronal development processes (generation of neurons, nerve development, and neurogenesis) were significantly enriched in both GO and GSEA analysis. The PPI networks constructed with the up- and downregulated DEGs consisted of 128 and 132 PPI pairs, respectively. There were 10 hub proteins, top highly connected DEGs: ANLN, TLE3, TLE2, DMD, ADRA1A, RBFOX1, MMP2, LMO1, RUNX1T1, and NPPA. Conclusion: The results indicate that granulation-specific gene expression may underpin diverse clinical characteristics in acromegaly. Overall, this study emphasizes the potential for further investigation into these transcriptomic variations and their roles in disease pathology, particularly the involvement of genes linked to neuronal development, inflammatory response, and JAK-STAT signaling in SGST. Presentation: 6/3/2024

TFF3 and PVRL2 co-targeting identified by multi-omics approach as an effective cancer immunosuppression strategy

BackgroundThe immunosuppressive tumour microenvironment (TME) plays a critical role in cancer progression and relapse by significantly influencing cancer pathogenesis through autocrine and paracrine signalling. Trefoil factor 3 (TFF3), a secreted protein, has been implicated in modulating the TME to promote cancer advancement. Herein, we investigated the potential association between TFF3 and key immunosuppressive TME components to distinguish a co-targetable oncotherapeutic strategy. MethodsThe TFF3-PVRL2 association were identified and investigated by integrating multiple bioinformatic-tools. The virtual compound screening for PVRL2 inhibitors was done with EasyVS. The TFF3-PVRL2 protein-level correlation was validated by immunoblotting, and the effectiveness of co-inhibiting TFF3 and PVRL2 was assessed using siRNA and AMPC (a TFF3 inhibitor). ResultsAnalysis of the TISIDB database revealed a positive correlation between TFF3 and PVRL2 mRNA levels across multiple cancer types. This correlation was confirmed at the protein level through immunoblot analysis. Further evaluation using TCGA pan-cancer datasets demonstrated that TFF3 and PVRL2 interact to establish an immunosuppressive TME, promoting cancer progression in BRCA, LUAD, PAAD, PRAD, and STAD. Enrichment analyses of positively correlated genes, PPI network hub proteins, and ceRNA networks involving TFF3 and PVRL2, conducted using LinkedOmics, STRING, and Cytoscape, provided insights into their potential co-functions in cancer. A cell-based assay was performed to evaluate the combined therapeutic efficacy of targeting both, TFF3 and PVRL2 and virtual screening identified potential drugs for inhibiting PVRL2. ConclusionPVRL2 has emerged as a promising immunoinhibitory target with significant associations with TFF3 and represents a key co-targetable molecule for effective oncotherapeutic strategies.

Exploration of the Relationship between Polycystic Ovary Syndrome and Recurrent Pregnancy Loss Based on Bioinformatics.

Recurrent Pregnancy Loss (RPL) and Polycystic Ovary Syndrome (PCOS) are both common diseases involving women of childbearing age, and their pathogenesis is still not sufficiently known. This study aimed to explore the relationship between RPL and PCOS in bioinformatics. Two expression chips, GSE86241 (obtained from 8 PCOS patients and 9 healthy controls) and GSE73025 (obtained from 5 RPL patients and 5 healthy controls), were downloaded from the Gene Expression Omnibus (GEO) database. We used the GEO database to analyze the gene expression profiles of PCOS and RPL to identify the intersection of abnormal miRNA expression, predicted the target genes of the intersecting miRNAs from miRDB, miRTarBase, and TargetScan databases, and then incorporated the miRNA-mRNA modulation network. By using the string database, the PPI network was built, which could screen the Hub genes and enrich them for analysis. Ultimately, the critical miRNA-mRNA regulatory network was set on the basis of the relationship between hub genes and miRNA. A total of 39 significantly altered miRNAs of PCOS and 137 significantly altered miRNAs of RPL were obtained, three miRNAs (miR-767-5p, miR-3196, and miR-187-3p), five signaling pathways (PI3K-Akt, p53, Toll-like receptor, C-type lectin receptor, and TNF signaling pathways), and six Hub genes (CASP8, PIK3R1, ADAMTS2, ADAMTS3, COL3A1, and MDM2) were found to be related to the development and progression of two diseases. More importantly, all Hub genes were regulated by miR-767-5p. This research clarifies the possible relationship between miRNA and mRNA with PCOS and RPL for the first time. It provides a basis for illustrating the pathogenic mechanism and a target of therapies for these two diseases.

New insight into the molecular mechanism of TCM Bufei Huoxue formula for chronic obstructive pulmonary disease based on network pharmacology and experimental verification.

To unveil the mechanism of the Bufei Huoxue formula (BHF) for chronic obstructive pulmonary disease (COPD) through integrated network pharmacology (NP) and experimental verification. LC-MS was first applied to the analysis of both in vitro and in vivo samples from BHF for chemical profiling. Then a ligand library was prepared for NP to reveal the mechanism of BHF against COPD. Finally, verification was performed using an animal model related to the results from the NP. A ligand library containing 170 compounds from BHF was obtained, while 357 targets related to COPD were filtered to construct a PPI network. GO and KEGG analysis demonstrated that bavachin, paeoniflorin, and demethylation of formononetin were the major compounds for BHF against COPD, which mainly by regulating the PI3K/Akt pathway. The experiments proved that BHF could alleviate lung injury and attenuate the release of TNF-α and IL-6 in the lung and BALF in a dose-dependent manner. Western blot further demonstrated the down-regulated effect of BHF on p-PI3K. BHF provides a potent alternative for the treatment of COPD, and the mechanism is probably associated with regulating the PI3K/AKT pathway to alleviate inflammatory injury by bavachin, paeoniflorin, and demethylation of formononetin.

Mechanisms of Berberine in anti-pancreatic ductal adenocarcinoma revealed by integrated multi-omics profiling

This study integrates pharmacology databases with bulk RNA-seq and scRNA-seq to reveal the latent anti-PDAC capacities of BBR. Target genes of BBR were sifted through TargetNet, CTD, SwissTargetPrediction, and Binding Database. Based on the GSE183795 dataset, DEG analysis, GSEA, and WGCNA were sequentially run to build a disease network. Through sub-network filtration acquired PDAC-related hub genes. A PPI network was established using the shared genes. Degree algorithm from cytoHubba screened the key cluster in the network. Analysis of differential mRNA expression and ROC curves gauged the diagnostic performance of clustered genes. CYBERSORT uncovered the potential role of the key cluster on PDAC immunomodulation. ScRNA-seq analysis evaluated the distribution and expression profile of the key cluster at the single-cell level, assessing enrichment within annotated cell subpopulations to delineate the target distribution of BBR in PDAC. We identified 425 drug target genes and 771 disease target genes, using 57 intersecting genes to construct the PPI network. CytoHubba anchored the top 10 highest contributing genes to be the key cluster. mRNA expression levels and ROC curves confirmed that these genes showed good robustness for PDAC. CYBERSORT revealed that the key cluster influenced immune pathways predominantly associated with Macrophages M0, CD8 T cells, and naïve B cells. ScRNA-seq analysis clarified that BBR mainly acted on epithelial cells and macrophages in PDAC tissues. BBR potentially targets CDK1, CCNB1, CTNNB1, CDK2, TOP2A, MCM2, RUNX2, MYC, PLK1, and AURKA to exert therapeutic effects on PDAC. The mechanisms of action appear to significantly involve macrophage polarization-related immunological responses.

Clarifying therapeutic mechanisms of Guanxinshutong capsule for cardiac fibrosis treatment by network pharmacology, molecular docking, molecular dynamics simulation, and in vivo experiments

PurposeGuanxinshutong capsule (GXST) is a Chinese patent medicine used for the treatment of cardiac fibrosis (CF). Nevertheless, the precise mechanism of action of GXST remains unclear. In this study, we applied network pharmacology, molecular docking, molecular dynamics simulation, and in vivo experiments to assess the effects and mechanisms of GXST in the treatment of CF. MethodsThe TCMSP database provided the GXST targets and active substances. The GeneCards, OMIM, and TTD databases were searched for targets relevant to cardiac fibrosis. To filter core targets, a PPI network was set up. Metascape was utilized to conduct enrichment analysis for the KEGG and GO pathways. To evaluate the interaction between putative targets and active drugs, molecular docking was utilized. MDS was performed for the optimal complex of the core ligand protein identified through molecular docking. Induction of cardiac fibrosis in SD rats using isoproterenol, and the protective effects of GXST on the heart were evaluated using echocardiography, H&E staining, and Masson staining. ResultsThrough network pharmacology analysis, we have identified 83 active components and 73 key target proteins related to cardiac fibrosis from GXST. Subsequent molecular docking and molecular dynamics simulation results suggest that quercetin, luteolin, and kaempferol can inhibit cardiac fibrosis by regulating the expression of TNF-α, IL6, and IL1B proteins. These compounds and proteins may be key factors in the treatment of CF with GXST. Further, in vivo experiments demonstrate that GXST can alleviate myocardial injury and improve cardiac function in CF rats. ConclusionThis study successfully predicted the effective components, potential targets, and related pathways involved in treating CF by GXST. GXST can improve cardiac function and alleviate pathological damage in the hearts of SD rats. This provides a new strategy for future investigations into the mechanisms of GXST in the treatment of CF.

Overexpression of Bmp4 induces microphthalmia by disrupting embryonic neural retina

Microphthalmia, mostly an autosomal dominant disorder, is a worldwide severe congenital ocular malformation that causes visual impairment. Our investigation unveiled a total of 30 genes associated with microphthalmia. Employing the CytoHubba and PPI network, we identified Bmp4 as the most pivotal hub gene. Subsequently, the conditional overexpression of Bmp4 in the retina caused highly distinctive microphthalmia, manifested by retinal disorganization with ganglion cell misalignment. Significant reduction in the number and abnormal distribution location of retinal cells in microphthalmia model mice. Elevated Bmp4 was associated with an increase in retinal apoptosis and a decrease in proliferating cells, which exacerbates the development of microphthalmia. Here we identify Bmp4 as an extremely important gene responsible for microphthalmia and the involved mechanisms. Overexpression of Bmp4 induces retinal cell ectopic expression and developmental defects, highlighting the importance of a well-balanced Bmp4 level in shaping the embryonic retina during early development.

Exploration of Pharmacological Mechanism of Cinnamomum tamala Essential Oil in Treating Inflammation based on Network Pharmacology, Molecular Modelling, and Experimental Validation.

Cinnamomum tamala (Buch.-Ham.) T.Nees & Eberm., also known as Indian Bay leaf, holds a distinctive position in complementary and alternative medicinal systems due to its anti-inflammatory properties. However, the active constituents and key molecular targets by which C. tamala essential oil (CTEO) exerts its anti-inflammatory action remain unclear. The present study used network pharmacology and experimental validation to investigate the mechanism of CTEO in the treatment of inflammation. GC-MS analysis was used to identify the constituents of CTEO. The key constituents and core targets of CTEO against inflammation were obtained by network pharmacology. The binding mechanism between the active compounds and inflammatory genes was ascertained by molecular docking and molecular dynamics simulation analysis. The pharmacological mechanism predicted by network pharmacology was verified in lipopolysaccharide-stimulated murine macrophage (RAW 264.7) cell lines. Forty-nine constituents were identified by GC-MS analysis, with 44 constituents being drug-like candidates. A total of 549 compounds and 213 inflammation-related genes were obtained, revealing 68 overlapping genes between them. Compound target network analysis revealed cinnamaldehyde as the core bioactive compound with the highest degree score. PPI network analysis demonstrated Il-1β, TNF-α, IL8, IL6 and TLR4 as key hub anti-inflammatory targets. KEGG enrichment analysis revealed a Toll-like receptor signalling pathway as the principally regulated pathway associated with inflammation. A molecular docking study showed that cinnamaldehyde strongly interacted with the Il-1β, TNF-α and TLR-4 proteins. Molecular dynamics simulations and MMPBSA analysis revealed that these complexes are stable without much deviation and have better free energy values. In cellular experiments, CTEO showed no cytotoxic effects on RAW 264.7 murine macrophages. The cells treated with LPS exhibited significant reductions in NO, PGE2, IL-6, TNF-α, and IL-1β levels following treatment with CTEO. Additionally, CTEO treatment reduced the ROS levels and increased the antioxidant enzymes such as SOD, GSH, GPx and CAT. Immunofluorescence analysis revealed that CTEO inhibited LPS-stimulated NF-κB nuclear translocation. The mRNA expression of TLR4, MyD88 and TRAF6 in the CTEO group decreased significantly compared to the LPS-treated group. The current findings suggest that CTEO attenuates inflammation by regulating TLR4/MyD88/NF- κB signalling pathway.

Identification of Potential Hub Genes in Alopecia Areata.

Alopecia areata (AA) is an immune-mediated chronic alopecia disease, but its specific pathogenesis is unclear. Gene expression data for AA patients (AAs) and healthy controls (HCs) were retrieved from the GEO database, and the differentially expressed genes (DEGs) between AAs and HCs were identified. Then, GO, KEGG and GSEA analysis were performed. A PPI network for the DEGs was then constructed to screen for hub genes, which were validated by three additional datasets. Subsequently, the potential miRNAs interacting with the hub genes were obtained through TarBase and miRNet. The differentially expressed lncRNAs (DElncRs) were obtained for subcellular localisation analysis, and the DElncRs located in the cytoplasm were further screened to identify miRNAs that interact with them. The shared miRNAs interacting with the hub genes and lncRNAs were used to construct a network of mRNA-miRNA-lncRNA interactions. Lastly, ROC analysis was performed to evaluate the potential diagnostic value of the hub genes and DElncRs identified. A total of 173 DEGs were obtained, mainly enriched in cytokines, chemokines, hair follicle development and hair cycle related signalling pathways. Through PPI screening and validation based on 3 additional datasets, 24 hub genes were finally yielded. Of them, five hub genes were upregulated and the potential miRNAs that interact with these five hub genes were identified. Additionally, 26 DElncRs were obtained, including 9 upregulated lncRNAs located in the cytoplasm that were predicted to interact with the miRNAs. Finally, an mRNA-miRNA-lncRNA regulatory network was constructed using five hub genes, four lncRNAs and their shared five miRNAs. The regulatory relationship between CD8A, mir-185-5p and FOXD2-AS1 might be crucial in AA pathogenesis, with CD8A and FOXD2-AS1 exhibiting diagnostic potential. CD8A and FOXD2-AS1 may serve as potential therapeutic targets in AA.

To Explore the Mechanism of Maiwei Dihuang Decoction in the Treatment of Non-small Cell Lung Cancer based on Network Pharmacology Combined with LC-MS.

To explore the mechanism of Maiwei Dihuang decoction in the treatment of non-small cell lung cancer (NSCLC) by using network pharmacology and LC-MS technology. The effective components in Maiwei Dihuang decoction were detected by liquid chromatography- mass spectrometry (LC-MS). Use the SuperPred database to collect the relevant targets of the active ingredients of Mai Wei Di Tang, and then collect the relevant targets of nonsmall cell lung cancer from GeneCards, DisgenNET and OMIM databases. On this basis, PPI network construction, GO enrichment analysis and KEGG pathway annotation analysis were carried out for target sites. Finally, AutoDock Vina is used for molecular docking. We further screened 16 effective Chinese herbal compounds through LC-MS combined with ADME level. On this basis, we obtained 77 core targets through protein interaction network analysis. Through GO, KEGG analysis and molecular docking results, we finally screened out the potential targets of Maiwei Dihuang Decoction for NSCLC: TP53, STAT3, MAPK3. Maiwei Dihuang decoction may play a role in the treatment of NSCLC by coregulating TP53/STAT3/MAPK3 signal pathway.

Influence of budesonide and fluticasone propionate in the anti-osteoporotic potential in human bone marrow-derived mesenchymal stem cells via stimulation of osteogenic differentiation

Osteoporosis is a prevalent bone condition with adverse effects observed in patients undergoing long-term glucocorticoid therapy, resulting in bone demineralization and tissue loss. There has been limited studies on the global response to dexamethasone in terms of comparing its expression profile to other common glucocorticoids during osteogenic differentiation. This study focused on the downregulated gene expression profile of glucocorticoid compounds; dexamethasone, budesonide, and fluticasone propionate, during osteogenic differentiation to elucidate the related target genes and pathways associated with the anti-osteoporotic potential of telomerase-immortalized human bone marrow-derived mesenchymal stem cells using a bioinformatics approach. Based on gene expression microarrays experiments and bioinformatics analysis, several key genes involved in the regulation of osteogenic differentiation and osteoporosis development in mesenchymal stem cells that were targeted by these specific glucocorticoids were determined. Network analysis using GeneCards, OMIM, and CTD databases were performed and osteoporosis-related genes were identified. LIMMA and moderated Welch test R packages were performed to determine significant downregulated differentially expressed genes for each glucocorticoid treatment. A total of 479 (dexamethasone), 84 (budesonide), and 889 (fluticasone propionate) differentially expressed genes were identified for each glucocorticoid, of which 35 common genes overlapped. Enrichment pathway analysis was conducted using Metascape, and protein-protein interaction networks were constructed using the STRING database and Cytoscape software to determine potential target genes involved with osteoporosis. Enrichment pathway analysis revealed genes involved in 3 Reactome pathways namely cytokine signaling in immune system, immune system and the interferon alpha/beta signaling pathways and identified 10 hub genes based on the PPI network to determine potential target pathways associated with osteoporosis. These findings provide preliminary insights into the relationship between the key target genes of dexamethasone, budesonide, and fluticasone propionate, and the pathways associated with regulated osteoporosis metabolism during osteogenic differentiation.

TRPM8 overexpression suppresses hepatocellular carcinoma progression and improves survival by modulating the RTP3/STAT3 pathway.

Hepatocellular carcinoma (HCC) is a malignant tumour associated with high morbidity and mortality rates worldwide. Recently, TRPM8 was reported to play an important role in tumour progression. However, the precise role of TRPM8 in HCC remains unclear. In this study, we explored the expression levels, molecular functions and underlying mechanisms of TRPM8 in HCC. Tissue samples were used to analyse the expression of TRPM8 to assess its diagnostic value for prognosis. Cell Counting Kit-8, EdU and colony formation assays were performed to evaluate the effects of TRPM8 on cell proliferation, whereas the Transwell assay was used to assess cell migration and invasion. The role of TRPM8 invivo was evaluated using a mouse subcutaneous xenograft tumour model. We performed PPI network analyses to understand the possible mechanisms of TRPM8 action. TRPM8 expression was decreased in HCC tissues and was correlated with histological grade and poor patient prognosis. Functionally, TRPM8 repressed the proliferation and metastasis of HCC cells both invitro and invivo by modulating the RTP3/STAT3 signalling pathway. Our findings underscore the critical role of the TRPM8-RTP3-STAT3 axis in maintaining the malignant progression of HCC. Moreover, our study demonstrates that AD80 is involved in anti-tumour processes by upregulating the expression of TRPM8.

Comprehensive investigation of network pharmacology, computational modeling, and pharmacokinetic assessment to evaluate the efficacy of flavonoids in rheumatoid arthritis.

Rheumatoid arthritis is a chronic autoimmune disease characterized by inflammation and joint damage, imposing a significant burden on affected individuals worldwide. Flavonoids, a class of natural compounds abundant in various plant-based foods, have shown promising anti-inflammatory and immunomodulatory effects, suggesting their potential as therapeutic agents for RA. In this study, we conducted a comprehensive investigation of identified LCMS compounds utilizing network pharmacology, computational modeling, in silico approaches, and pharmacokinetic assessment to evaluate the efficacy of flavonoids in RA treatment. The study identified 5 flavonoid structures with common targets via LCMS and Integration of network pharmacology approaches enabled a comprehensive evaluation of the pharmacological profile of flavonoids in the context of RA treatment, guiding the selection of promising candidates for further experimental validation and clinical development. The top 10 targets were AKT1, PI3KR1, CDK2, EGFR, CDK6, NOS2, FLT3, ALOX5, CCNB1, and PTPRS via PPI network. The investigation emphasized several pathways, including the AGE-RAGE signaling pathway, resistance to EGFR tyrosine kinase inhibitors, the PI3K-AKT signaling network, and the Rap 1 signaling pathway. In silico studies estimated binding affinities that ranged from - 7.0 to - 10.0 kcal/mol. Schaftoside and Vitexin showed no toxicity in computational approach and found suitable for further investigations. Overall, our study underscores the potential of flavonoids as therapeutic agents for RA and highlights the utility of integrative approaches combining network pharmacology, computational modeling, in silico methods, and pharmacokinetic assessment in drug discovery and development processes.

Unlocking the secrets of NPSLE: the role of dendritic cell-secreted CCL2 in blood-brain barrier disruption.

This study employed RNA-seq technology and meta-analysis to unveil the molecular mechanisms of neuropsychiatric systemic lupus erythematosus (NPSLE) within the central nervous system. Downloaded transcriptomic data on systemic lupus erythematosus (SLE) from the Gene Expression Omnibus (GEO) and analyzed differential genes in peripheral blood samples of NPSLE patients and healthy individuals. Employed WGCNA to identify key genes related to cognitive impairment and validated findings via RNA-seq. Conducted GO, KEGG, and GSEA analyses, and integrated PPI networks to explore gene regulatory mechanisms. Assessed gene impacts on dendritic cells and blood-brain barrier using RT-qPCR, ELISA, and in vitro models. Public databases and RNA-seq data have revealed a significant upregulation of CCL2 (C-C motif chemokine ligand 2) in the peripheral blood of both SLE and NPSLE patients, primarily secreted by mature dendritic cells. Furthermore, the secretion of CCL2 by mature dendritic cells may act through the RSAD2-ISG15 axis and is associated with the activation of the NLRs (Nod Like Receptor Signaling Pathway) signaling pathway in vascular endothelial cells. Subsequent in vitro cell experiments confirmed the high expression of CCL2 in peripheral blood dendritic cells of NPSLE patients, with its secretion being regulated by the RSAD2-ISG15 axis and inducing vascular endothelial cell pyroptosis through the activation of the NLRs signaling pathway. Clinical trial results ultimately confirmed that NPSLE patients exhibiting elevated CCL2 expression also experienced cognitive decline. The secretion of CCL2 by dendritic cells induces pyroptosis in vascular endothelial cells, thereby promoting blood-brain barrier damage and triggering cognitive impairment in patients with systemic lupus erythematosus.

Exploring multitarget potential of Piper nigrum fruit constituents for Alzheimer's disease: An AI-driven strategy

BackgroundAlzheimer's disease (AD) is a neurodegenerative disease that leads to development of cognition and memory dysfunctions. Currently, there is no known cure for AD, although limited medications are approved for the management of disease condition. Various plant-based leads give new hope for considering phytoconstituents as anti-AD drugs. The Piper nigrum L. fruit extract was reported to have anti-Alzheimer's activity. It creates an interest in the finding of active moieties that may be accountable for anti-AD activity. Hypothesis/PurposeIdentification of multitarget directed ligands isolated from Piper nigrum fruits through AI based studies. Study designThe phytochemical analysis of alkaloid fraction was carried out by LCMS, followed by the evaluation of constituents through in silico studies. MethodsThe fruits methanolic extract was prepared by cold maceration technique. The chemical profiling of the alkaloidal fraction was carried out using LCMS/MS analysis. The obtained compound's target hit genes were identified through network pharmacology studies using String, Metascape, and Cytoscape tools. Further, docking studies and MD simulations were carried out using AutoDock4 and Desmond-Maestro software. Then, electrochemical properties of hit compound P4 were determined using Gaussview6 software. ResultsFrom LCMS/MS analysis data, 29 compounds were considered based on compound intensity and accuracy (>95 %). Then, 41 common gene targets were identified from AD genes and compound-targeted genes. The 41 common genes in the PPI network suggested that AChE and BACE1 were the most abundant proteins. Further, docking studies revealed the hit compound P4 binding interaction and energies when compared to other 28 ligands. The molecular dynamics studies showed that P4-AChE and P4-BACE1 complexes were stable, and there were no RMSD and RMSF fluctuations were observed up to 100 ns. Further, PCA and MM-GBSA analysis data supported that complexes (P4-AChE and P4-BACE1) were stable. The DFT and surface properties indicated that compound P4 was ideal candidate for AD treatment and must be considered for further biological activity studies. ConclusionThe study identified compound P4 (dehydropipernonaline) from alkaloidal fraction of Piper nigrum fruits, suggesting it may be hit candidate for AD treatment.

Integration of bioinformatics and multi-layered experimental validation reveals novel functions of acetylation-related genes in intervertebral disc degeneration

BackgroundThe molecular mechanisms underlying intervertebral disc degeneration (IDD) remain poorly understood. The purpose of this work is to elucidate key molecules and investigate the roles of acetylation-related RNAs and their associated pathways in IDD. MethodDatasets GSE70362 and GSE124272 were obtained from the Gene Expression Omnibus (GEO) and combined to investigate differentially expressed genes (DEGs) associated with acetylation in IDD patients compared to healthy controls. Critical genes were pinpointed by integrating GO, KEGG and PPI networks. Furthermore, CIBERSORTx analysis was used to investigate the differences in immune cell infiltration between different groups and the biological processes (BP), cellular components (CC) and molecular functions (MF) were calculated by GSEA and GSVA. In addition, The single-cell database GSE165722 was incorporated to validate the specific expression patterns of hub genes in cells and identify distinct cell subtypes. This provides a theoretical basis for a more in-depth understanding of the roles played by critical cell subtypes in the process of IDD. Subsequently, tissues from IVD with varying degrees of degeneration were collected to corroborate the key DEGs using western blot, RT-qPCR, and immunofluorescence staining. ResultsBy integrating various datasets and references, we identified a total of 1620 acetylation-related genes. These genes were subjected to a combined analysis with the DEGs from the databases included in this study, resulting in the discovery of 358 acetylation-related differentially expressed genes (ARDEGs). A comparative analysis with differentially expressed genes obtained from three databases yielded 19 ARDEGs. The PPI network highlighted the top 10 genes (IL1B, LAMP1, PPIA, SOD2, LAMP2, FBL, MBP, SELL, IRF1 and KHDRBS1) based on their protein interaction relationships. CIBERSORTx immune infiltration analysis revealed a moderate positive correlation between the gene IL1β and Mast.cells.activated, as well as a similar correlation between the gene IRF1 and Mast.cells.activated. Single-cell dataset was used to identify cell types and illustrate the distribution of hub genes in different cell types. The two cell types with the highest AUCell scores (Neutrophils and Monocytes) were further explored, leading to the subdivision of Neutrophils into two new cell subtypes: S100A9-type Neutrophils and MARCKS-type Neutrophils. Monocytes were labeled as HLA-DRA9-type Monocytes and IGHG3-type Monocytes. Finally, molecular biology techniques were employed to validate the expression of the top 10 hub genes. Among them, four genes (IL1β, SOD2, LAMP2, and IRF1) were confirmed at the gene level, while two (IL1β and SOD2) were validated at the protein level. ConclusionIn this study, we carried out a thorough analysis across three databases to identify and compare ARDEGs between IDD patients and healthy individuals. Furthermore, we validated a subset of these genes using molecular biology techniques on clinical samples. The identification of these differently expressed genes has the potential to offer new insights for diagnosing and treating IDD.

Comparative assessment of acute neurotoxicity of real-world ultra-fine black carbon emitted from residential solid fuel combustion

Incomplete combustion of residential solid fuel is one of the main anthropogenic sources for black carbon (BC). Fresh BC, mainly enriched in ultra-fine fraction of particles, can directly cross blood-brain barrier and are reported to be associated with neurodegenerative diseases. Because of the difficulties in collection and purification of BC from ambient particles, there are still significant knowledge gaps in understanding neurotoxicity caused by real-world BC. The purpose of this study is to compare the neurotoxic effects caused by BCs emitted from combustion of six residential solid fuels, and try to reveal associated biological mechanisms in SH-SY5Y cells. Two straw BC (Wheat-BC and Corn-BC) showed highest neurotoxic effects followed by wood BC (Pine-BC and Aspen-BC) and coal BC (Xvzhou and Longkou Coal), as indicated by viability, lactic dehydrogenase, malondialdehyde, adenosine triphosphate and acetylcholine levels. Coal BC caused nearly no toxicity in human neuroblastoma (SH-SY5Y) cells within highest dose of 200 μg/mL. RNA sequence and bioinformatics analysis were applied to effectively identify differential genes and signaling pathways. Based on Gene Ontology (GO) annotation, Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment, Protein Protein Interaction network (PPI network) construction, we found biomass BC affected mitochondrial function, interfered with cellular metabolic processes, disturbed redox homeostasis, and finally resulted in cellular damages. Coal-BC mainly caused cytokine/chemokine related inflammatory responses. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blotting methods were further applied to find out related signaling pathways. Biomass BC activated IL6R/JAK3/STAT3 and JAK3/STAT6 pathways leading to oxidative stress and inflammatory responses. Coal BC activated JAK3/STAT3 pathway leading to chemokine related responses. This study revealed the heterogeneity in neurotoxicity of BCs from different combustion sources and provided important data for health risk assessment. BC-related neurotoxicity should be considered when making air pollution emission control strategies, with residential biomass receiving more policy attention.

Interplay of RNA m6A Modification-Related Geneset in Pan-Cancer.

Background: N6-methyladenosine (m6A), is the most common modification found in mRNA and lncRNA in higher organisms and plays an important role in physiology and pathology. However, its role in pan-cancer has not been explored. Results: A total of 31 m6A modification regulators, including 12 writers, 2 erasers, and 17 readers are identified in the current study. The functional analysis of the regulators results in the enrichment of processes, primarily related to RNA modification and metabolism, and the PPI network reveals multiple interactions among the regulators. The mRNA expression analysis reveals a high expression for most of the regulators in pan-cancer. Most of the m6A regulators are found to be mutated across the cancers, with ZC3H13, VIRMA, and PRRC2A having a higher frequency rate. Significant correlations of the regulators with clinicopathological parameters, such as age, gender, tumor stage, and grade are identified in pan-cancer. The m6A regulators' expression is found to have significant positive correlations with the miRNAs in pan-cancer. The expression pattern of the m6A regulators is able to classify the tumors into different subclusters as well as into high- and low-risk groups. These tumor groups show differential patterns in terms of their immune cell infiltration, tumor stemness score, genomic heterogeneity score, expression of immune regulatory/checkpoint genes, and correlations between the regulators and the drugs. Conclusions: Our study provide a comprehensive overview of the functional roles, genetic and epigenetic alterations, and prognostic value of the RNA m6A regulators in pan-cancer.

Network Pharmacology and Molecular Docking to Explore the Mechanism of Action of Danxiong Huoxue Tablets for the Prevention and Treatment of Heterotopic Ossification

Background: Danxiong Huoxue tablets have been used clinically to prevent and treat Heterotopic Ossification (HO) after total hip arthroplasty, but its therapeutic mechanism is not clear. background: Danxiong Huoxue Tablets has been used clinically to prevent and treat heterotopic ossification (HO) after total hip arthroplasty, but its therapeutic mechanism is not clear. Objectives: This study investigated the potential mechanism of action of Danxiong Huoxue tablets against HO using network pharmacology and molecular docking methods. Methods: The TCMID and SymMap databases and the existing literature were used to screen the active ingredients and targets of Danxiong Huoxue tablets, and the targets of heterotopic ossification were obtained from Gene Cards, DisGeNET, and PharmGkb databases. The PPI network diagram was constructed using the String database and Cytoscape 3.9.1 software. Finally, the core targets were analyzed by GO and KEGG enrichment and validated by molecular docking. Results: A total of 39 active ingredients and 328 corresponding targets of Danxiong Huoxue tablets were identified, which were enriched in signaling pathways, such as PI3K/Akt and MAPK. Molecular docking verified that there was good binding activity between the core targets and the corresponding components. Conclusion: Danxiong Huoxue tablets may prevent and treat HO by acting on key targets, such as SRC, AKT1, JUN, and TNF, and regulating the PI3K/Akt signaling pathway and MAPK signaling pathway.

Prognostic value and molecular mechanisms of OAS1 in lung adenocarcinoma

BackgroundThe expression of 2’-5’-oligoadenylate synthetase 1 (OAS1) in lung cancer has been validated in numerous studies. However, the prognostic value of OAS1 expression in lung adenocarcinoma (LUAD) still remains unclear. This study aimed to reveal the prognostic value and associated molecular mechanisms of OAS1 expression in LUAD.MethodsGene expression data of LUAD were extracted from online databases. Gene and protein expression levels of OAS1 in LUAD and normal samples were revealed, followed by prognostic analysis of OAS1. Next, we conducted a thorough bioinformatics analysis to examine the enrichment of key functional and biological signaling pathways and their correlation with the abundance of immune cells. The independent prognoses, drug responses, and PPI networks associated with OAS1 were analyzed. OAS1 expression was evaluated in LUAD tissues and cell lines. OAS1 was knocked down by siRNA transfection, followed by CCK8, colony formation, and wound-healing assays.ResultsGene and protein expression levels of OAS1 in LUAD samples were significantly higher than those in normal samples (all P < 0.05). OAS1 stimulation were correlated with poor prognosis, lymph node metastasis, advanced tumor stage, immune cells, and immunomodulators. The prognostic value of OAS1 in LUAD was determined via univariate regression analysis. In total, 10 OAS1-associated genes were revealed via PPI analysis of OAS1, which were primarily enriched in functions, such as the negative regulation of viral genome replication. Transcriptional analysis revealed several OAS1-related interactions, including STAT3-miR-21-OAS1. STAT3 was overexpressed and miR-21 was expressed in LUAD cells. Upregulation of OAS1 protein was determined in LUAD tissues and cell lines. OAS1 knockdown significantly reduced proliferation and migration of LUAD cells.ConclusionsOAS1 overexpression influenced survival and immune cell infiltration in patients with LUAD, which might be a potential prognostic gene for LUAD. Moreover, OAS1 contributed to LUAD progression by participating in STAT3-miR-21-OAS1 axis.