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STOML2 Suppresses the Proliferation, Migration, and Inflammation of Airway Smooth Muscle Cells in Children with Asthma by Inhibiting NLRP3
No full textBackground: Asthma is a chronic disease of the airways which involves airway remodeling and inflammation. Stomatin like protein-2 (STOML2) regulates immunity and inflammation. This study aimed to elucidate the functional role of STOML2 in childhood asthma and the mechanisms by which it acts. Materials and Methods: Airway smooth muscle cells (ASMCs) treated with platelet-derived growth factor-BB (PDGF-BB) were used to mimic airway hyper-reactivity in vitro. STOML2 expression in serum from patients with asthma and in PDGF-BB-induced ASMCs was evaluated by quantitative real-time PCR (qRT-PCR) and western blot assays. Cell proliferation was detected by cell counting kit-8 (CCK-8) and colony formation assays. Cell migration and invasion were measured by wound healing and transwell assays. Enzyme-linked immunosorbent assay (ELISA) was applied to analyze the expression of interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α) in PDGF-BB-induced ASMCs. The levels of the nucleotide-binding oligomerization domain-like receptor (NLR) containing pyrin domain 3 (NLRP3) inflammasome were assessed by western blot assay. Results: STOML2 was decreased in the serum of children with asthma and in PDGF-BB-induced ASMCs (p < 0.01). Overexpression of STOML2 reduced the proliferation, migration, and invasion of PDGF-BB-induced ASMCs (p < 0.01). Increased STOML2 suppressed the secretion of inflammatory cytokines in PDGF-BB-induced ASMCs (p < 0.05). In addition, STOML2 overexpression inhibited the activation of the NLRP3 inflammasome. Conclusions: STOML2 regulated the inflammation and airway remodeling of ASMCs induced by PDGF-BB through inhibiting NLRP3, which might provide a novel direction for the treatment of asthma
Investigating the Efficacy of Cichorium Intybus L. as a Therapeutic Agent for Chronic Unpredictable Mild Stress-Induced Depression in Mice
No full textBackground: Oxidative stress, which is tightly linked to free radical generation, is cited as contributing to depression. Cichorium intybus L. (CI) is widely known as a viable source of abundant substances with antioxidant properties. In this study, we evaluated the possible antidepressant properties of CI in mice subjected to chronic unpredictable mild stress (CUMS). Method: Swiss albino mice were subjected to CUMS induction procedure for 14 days and then treated with imipramine (15 mg/kg) and CI (250 and 500 mg/kg) for 10 days. The treatment was followed by several behavioral parameter evaluations, such as sucrose preference test, forced swim test, open field test, and tail suspension test. Antioxidant enzyme levels, lipid peroxidation degree, and brain serotonin content of the sacrificed animals were assessed. Results: The neurochemical analysis of the mice exposed to CUMS revealed significantly decreased serotonin and antioxidant enzyme levels as well as heightened lipid peroxidation (p < 0.001). The mice with CUMS underwent significant behavioral changes, marked by protracted immobility, decreased exploratory behavior, and decreased preference for sucrose solution (p < 0.001). However, these behavioral changes could be reversed by administering CI, which also resulted in elevated antioxidant enzyme levels, diminished lipid peroxidation, and ultimately increased serotonin levels in the brain (p < 0.001). Conclusion: Our findings demonstrated that CI holds immense promise as a therapeutic agent for depression by strengthening the antioxidant defense system to mitigate oxidative stress, decrease lipid peroxidation, and preserve serotonin levels in the brain
Endothelial Cell-Derived Interleukin-6 Facilitated Colorectal Cancer Cell Migration and Invasion by Regulating Osteopontin in a Paracrine Manner
No full textBackground: Patients with metastatic colorectal cancer (CRC) generally suffer from poor prognosis. Therefore, there is an urgent need to further delineate the mechanism of metastasis in CRC, which is of clinical significance for the development of more targeted therapy. This study aimed to explore the potential role of endothelial cells-derived interleukin-6 (IL-6) in CRC metastasis. Methods: Human umbilical vein endothelial cell (HUVEC)-conditioned medium and CRC-conditioned medium were prepared, with the latter being the control. Enzyme-linked immunosorbent assay was employed to detect IL-6 levels in HUVEC-conditioned or CRC-conditioned culture media. CRC cells were separately cultured in both media to investigate the effect of HUVECs on CRC metastasis using cell counting kit-8 assay, scratch test and Transwell assay. The role of HUVEC-derived IL-6 in CRC cells was investigated using rescue assay. The expressions of the osteopontin protein and the proteins in glycogen synthase kinase-3β (GSK3β)-β-catenin signaling pathways were determined by Western blotting. To confirm the mediating role of osteopontin in the relationship between IL-6 and CRC, the expression of osteopontin in CRC cells was up-regulated. Results: IL-6 level was higher in HUVEC-conditioned medium than in CRC-conditioned medium (p < 0.001). CRC cells in the HUVEC-conditioned medium showed enhanced viability, migration and invasion (p < 0.05). HUVEC-secreted IL-6 facilitated malignant phenotypes and activated osteopontin and GSK3β-β-catenin signaling pathway of CRC cells in a dose-dependent manner (p < 0.05). Moreover, osteopontin overexpression contributed to CRC metastasis in vitro and activated GSK3β-β-catenin signaling pathway (p < 0.001), while such effect was abrogated by IL-6 silencing (p < 0.001). Conclusion: HUVEC-derived IL-6 facilitated the metastasis of CRC cells by regulating osteopontin in a paracrine manner
Exploration of Pathogenesis of Aortic Dissection through Whole Genome DNA Methylation
No full textBackground: Aortic Dissection (AD) is the most common aortic emergency, and its exact etiology is still not fully understood. With advancements in molecular biology, a more profound understanding of the causes of AD will result in more significant breakthroughs in prevention and treatment. This work aimed to explore the role of genome-wide Deoxyribonucleic acid (DNA) methylation in exploring the pathogenesis and outcome mechanism of AD from the perspective of molecular biology. Methods: Aortic tissue was collected from 8 AD patients and 8 non-AD volunteers to analyze DNA methylation characteristics in AD patients through the whole genome DNA methylation method. Based on bioinformatics, epigenetic characteristics during AD were studied. Finally, the accuracy of the chip results was verified by pyrosequencing. Results: A total of 1563 sites were counted. Compared with the normal group, there were 942 methylation upregulated sites and 621 downregulated sites in the AD group. Differential methylation sites detected by the chip were distributed in Transmission Start Site (TSS) 1500, 5′ Untranslated Region (UTR), Genebody, 3′UTR, Cytosine, Phosphoric acid, and Guanine (CpG) island, and off-island CpG sites, mainly located in Genebody. After Gene Ontology (GO) enhancement analysis and pathway analysis of biological pathways, it was found that some methylated genes were closely related to cell differentiation, growth, maturation, aging, and death, affecting AD development. Conclusion: Whole genome DNA methylation plays a positive role in exploring the pathogenesis and outcome mechanism of AD from molecular biology. This helps explore new diagnostic markers and intervention targets for AD and improve the clinical diagnosis and treatment of AD in the future
Mendelian Randomization Analysis of Immune Cells and Asthma
Get PDFBackground: The immune cells play a substantial role in the development and advancement of asthma. Therefore, we utilized mendelian randomization (MR) analysis to investigate the correlation between immune cells and asthma. Objective: Given that immune cells play a crucial role in the onset and progression of the condition, this study aimed to elucidate the unclear links between immune cells and asthma. Methods: The publicly available genetic data regarding asthma were obtained from the IEU database, and genetic variation points were selected as instrumental variables (IVS). Moreover, genetic information concerning immune cells was obtained from published literature. We used five different methods for dual sample mendelian randomization (MR) analysis, including inverse variance weighted (IVW), weighted median (WMI), MR-Egger regression, simple mode (SM), and weighted mode (WM). Furthermore, sensitivity analysis was utilized to examine the heterogeneity, horizontal pleiotropy, and stability of the outcomes. Results: IVW results showed that B-cell Activating factor of the TNF family receptor (BAFF–R) on B cell, BAFF–R on IgD- CD27- B cell, BAFF–R on IgD+ CD24- B cell, BAFF–R on IgD+ CD38dim B cell, CD33br HLA DR+ CD14dim myeloid cell, CD25 on B cell, CD25 on IgD+ CD24- B cell, CD25 on IgD+ CD38- naive B cell, CD25 on naive-mature B cell, CD25 on transitional B cell, CD33 on basophil, CD33 on CD14+ monocyte, CD33dim HLA DR+ CD11b- myeloid cell, CD33 on CD66b+ myeloid cell, CD38 on IgD- CD38dim B cell, CD86 on myeloid dendritic cells (DC), HLA DR on CD14- CD16+ monocyte, IgD+ CD38br lymphocyte, and transitional lymphocyte may be the risk factors of asthma. Moreover, CD11b on CD14+ monocyte, CD24 on IgD+ CD38br B cell, CD28 on CD45RA+ CD4+ Treg, CD45 on NK, HLA DR+ CD3- NK, HLA DR+ NK cell, IgD- CD38- B cell, PDL–1 on CD14+ CD16- monocyte, and plasmacytoid dendritic cells (DC) were identified as protective factors for asthma. Conclusion: This study explored the causal relationship between asthma and immune cells and identified immune cells correlated with asthma development. These immune cells may become new biomarkers or therapeutic targets, provide better treatment options for the prevention and treatment of asthma, and promote the understanding of asthma
In-Silico Identification of Novel Anti-Inflammatory Inhibitor Molecules Based on a Combination Strategy of Bioinformatic Approaches
No full textBackground: Cyclooxygenase-2 (COX-2) is an enzyme responsible for inflammatory responses and is associated with pain, fever, pyretic diseases, and tumor development. The present study aimed to identify new compounds that could be developed as selective inhibitors for COX-2 isoenzyme by using in-silico approaches. This study was conducted to search for potential anti-inflammatory compounds. Methods: The physiochemical and drug-like properties of natural compounds were determined by SWISSADME and toxicity by using ProTox-II. In-silico docking was performed to determine the affinity of various compounds with a target protein. Molecular dynamic analysis of the protein was carried out using Normal mode analysis (NMA) with internal coordinates normal mode analysis server (iMODS). Non-protein contacts were visualized using Protein Contacts Atlas. Protein-protein interactions (PPIs) were estimated using the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING). The free binding energies of the docked protein-ligand complexes were determined using Fast Amber Rescoring (farPPI). Prospective binding pockets in the protein were identified using DoGSiteScorer. Amino acid variation between the query and the database sequence were determined using BlastP analysis. Results: All the compounds in the present study followed Lipinskis rule of five. ProTox-II analysis revealed that syringic acid, eugenol, and gingerenone A were inactive for all toxicities, whereas isosilychristin was active for immunotoxicity. Isosilychristin was found to be the most effective compound with the highest binding affinity of –10.1 kcal/mol. Molecular dynamic analysis provided insights about the mobility, flexibility and stability of the protein molecule. Protein Contact Atlas analyzed the network of residue-residue interactions at the atomic level. Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) provides insights about protein-protein interactions (PPIs), extent of their similarities and functional associations. ShinyGO findings revealed that ovarian steroidogenesis was the predominant pathway. Homology modelling revealed that 96.90% of the residues were found to lie in the Ramachandran favored region. Protein Structure Analysis (ProSA) determined the overall quality of a protein molecule, with a Z-score of –8.91. Molecular Mechanics/Generalized Born Surface area (MM/GBSA) approach described stronger binding affinities between the protein and the ligand molecules. The findings obtained from DoGSiteScorer revealed binding pockets properties with COX-2/isosilychristin complex having the highest surface-volume ratio. BlastP findings revealed that our query sequence showed 100% similarity with prostaglandin G/H synthase 2 (accession number NP_000954.1) and with Crystal Structure of Aspirin Acetylated Human Cyclooxygenase-2 (5IKQ_A and 5F1A_A). Conclusions: The findings of this study revealed that isosilychristin could be a promising inhibitor to target COX-2
Berberine Alleviates 5-Fluorouracil-Induced Inflammatory Response in Macrophages and Intestinal Mucositis in Mice
No full textBackground: The primary reason for the development of chemotherapeutic intestinal mucositis induced by 5-fluorouracil (5-FU) is the activation of macrophages in the mucosa. This study aimed to explore how berberine can alleviate inflammation in macrophages and the resulting intestinal mucosal inflammation in mice induced by 5-FU. Methods: Tohoku Hospital Pediatrics-1 (THP-1) cell inflammatory response and mouse intestinal mucositis were induced by 5-FU and treated with berberine. The levels of inflammation-related factors and autophagy related proteins in THP-1 cells were detected by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blotting. The concentrations of double-stranded DNA (dsDNA) and interleukin-1β (IL-1β) in mice serum and the small intestine hematoxylin-eosin (HE) staining were used to evaluate intestinal mucosal damage. Immunoglobulin A (IgA), an indicator of mucosal immunity, was detected in mice serum by enzyme-linked immunosorbent assay (ELISA). We used quantitative polymerase chain reaction (qPCR) to detect the relative contents of four important strains (Bifidobacterium, Lactobacillus, Escherichia coli, and Enterococcus) in the colon contents of mice. Additionally, we employed liquid chromatograph-mass spectrometer/mass spectrometer (LC-MS/MS) technique to measure the concentrations of three main short-chain fatty acids (acetic acid, propionic acid, and butyric acid) in the mices plasma. Furthermore, we employed ultra-performance liquid chromatography-quadrupole-time of flight-mass spectrometer (UPLC-Q Tof-MS) technique to analyze the non-targeted metabolomics of mouse serum. Results: Berberine has been found to inhibit the expressions of NOD-Like Receptor Thermal Protein Domain Associated Protein 3 (NLRP3), Caspase-1, and IL-1β in THP-1 cells (p < 0.05). Additionally, it effectively suppresses the expression of autophagy related proteins LC3 and Beclin-1 in THP-1 cells (p < 0.05). Furthermore, in a mouse model study, berberine significantly enhances the levels of beneficial bacteria Bifidobacterium and Lactobacillus, as well as the concentrations of three main short-chain fatty acids (SCFAs) in the plasma of mice (p < 0.05). Moreover, it reduced the concentration of pro-inflammatory factor dsDNA and increased the mucosal immunity index IgA concentration in blood (p < 0.05). The untargeted metabolomics results demonstrated that berberine could regulate the inflammatory response of mice by impacting the metabolism of taurine, glycerol phospholipid, arachidonic acid, and primary bile acid biosynthesis. Conclusions: Berberine has demonstrated its ability to effectively suppress the inflammatory reaction of THP-1 cells induced by 5-FU. Furthermore, it also influences the “intestinal flora-metabolite-inflammation” pathway by regulating the composition of the intestinal flora, increasing the production of SCFAs, reducing the expression of inflammatory factors, and preserving the structural integrity of the intestinal mucosa
Relationship between IL-6 and IL-10 Gene Polymorphisms and Susceptibility to Pneumonia and Serum Levels of IL-6, IL-10, and C-Reactive Protein in Children
Get PDFBackground: Cytokines are molecules that play a crucial role in the development of childhood pneumonia. This study aims to investigate the relationship between gene polymorphisms of interleukin (IL)-6 and IL-10 and the susceptibility to childhood pneumonia. Methods: A total of 200 child patients with pneumonia and 200 healthy children were assigned to the disease group and control group, respectively. Peripheral blood was collected from both groups, and the polymorphisms were analyzed by sequencing. Additionally, the serum levels of IL-6, IL-10, and C-reactive protein were analyzed. Results: The frequencies of certain alleles and genotypes of IL-6 and IL-10 gene polymorphisms were found to be higher in the disease group compared to the control group (p < 0.05). Specifically, the allele C of IL-6 gene polymorphism rs762371056, allele A of IL-6 gene polymorphism rs762759043, and allele A of IL-10 gene polymorphism rs570186303 were more frequent in the disease group. Additionally, the disease group exhibited higher frequencies of genotype CC of rs762371056, genotype AA of polymorphism rs762759043, and genotype AA of rs570186303 (p < 0.05). The two groups also differed in the distributions of the dominant model of rs762759043 and the recessive model of rs570186303 (p < 0.05). Besides, there were differences in the distributions of haplotype CA of rs762371056 and rs762759043 and haplotypes AA, AG and GG of rs570186303 and rs575853731 between the two groups (p < 0.05). Child patients with genotype TC had higher levels of serum IL-6, while the levels of serum IL-10 were lower in child patients carrying genotype AA. Child patients with genotype AA had a distinctly higher level of serum C-reactive protein (p < 0.05). Conclusions: IL-6 and IL-10 gene polymorphisms have been found to be correlated with susceptibility to childhood pneumonia
m6A Demethylase FTO Promotes Lung Cancer Progression in Vitro and in Vivo by Inhibiting CLIC5
Get PDFBackground: Lung cancer (LC), a leading cause of cancer-related mortality worldwide, is a major health concern. The N6-methyladenosine (m6A) methylation, a pivotal RNA modification, plays a crucial role in the progression of LC. Therefore, this study aimed to identify m6A modification-related mechanisms underpinning LC progression. Methods: Expression levels of chloride intracellular ion channel 5 (CLIC5), fat mass and obesity-associated protein (FTO), and the proto-oncogene tyrosine-protein kinase SRC (SRC) in LC cells and xenograft tumors were assessed utilizing bioinformatics tools, quantitative reverse transcription polymerase chain reaction (qRT-PCR), and Western blot analysis. Furthermore, their interactions were predicted and validated using bioinformatics tools and/or Methylated RNA immunoprecipitation (MeRIP) assay. After CLIC5/FTO overexpression in LC cells, their roles in LC cell proliferation, migration, invasion, and tumorigenesis were examined employing cell counting kit-8 (CCK-8), 5-Ethynyl-2′-deoxyuridine (EdU), Transwell, and murine xenograft assays. Results: CLIC5 was downregulated in LC cells (p < 0.001). CLIC5 overexpression inhibited cell viability, proliferation, migration, invasion, and tumorigenesis in LC (p < 0.01). The m6A level was decreased while the FTO level was increased in LC cells (p < 0.001). FTO could interact with the m6A modification site on CLIC5, and the m6A methylation of CLIC5 was potentiated following FTO knockdown in LC cells (p < 0.001). Furthermore, FTO overexpression reversed the inhibitory effect of CLIC5 overexpression on the proliferation, migration, invasion, and tumorigenesis in LC (p < 0.05). Additionally, elevated SRC expression in LC could interact with CLIC5 (p < 0.001). CLIC5 overexpression diminished SRC expression, and the effect of CLIC5 on the SRC expression was reversed by FTO overexpression. Conclusions: The downregulation of CLIC5, induced by FTO-mediated m6A demethylation, facilitates LC progression both in vitro and in vivo
The Interplay between FTO-Mediated m6A Demethylation and Ferroptosis in the Development of Diabetic Retinopathy
No full textBackground: Diabetic vascular complications are mainly caused by endothelial dysfunction, which is influenced by inflammation and epigenetic modifications. In this study, we aimed to investigate the role of fat-, mass- and obesity-associated (FTO) protein, an N6-methyladenosine (m6A) demethylase, in diabetes-induced retinal vascular endothelial dysfunction. Methods: Human retinal vascular endothelial cells (RECs) were treated with high glucose (HG). The effect of HG on RECs was examined by various methods. The cell proliferation and apoptosis rate were determined using cell counting kit-8 (CCK-8) and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) methods. The oxidative stress represented by the levels of Fe2+, malondialdehyde (MDA), and reduced glutathione (GSH) was assessed using commercially available kits. Moreover, the levels of m6A methylation and FTO protein, and the levels of ferroptosis-related proteins were evaluated using corresponding kits and Western blot analysis, respectively. The cell transfection method was used to regulate FTO in RECs and to determine the consequent effect on HG-treated RECs. Additionally, the levels of reactive oxygen species (ROS), MDA, 4-hydroxynonenal (4-HNE), and the structural changes in mitochondria were observed using Transmission electron microscopy. Furthermore, Erastin was used to induce ferroptosis in RECs with FTO knockdown and the consequent effects on ferroptosis, m6A methylation, oxidative stress, proliferation and apoptosis of HG-treated RECs were evaluated. Results: HG treatment decreased proliferation (p < 0.01) and m6A methylation (p < 0.01), while increased apoptosis (p < 0.01), oxidative stress (p < 0.01), and ferroptosis (p < 0.01) in RECs. Furthermore, these HG-induced changes were ameliorated by FTO (p < 0.01). Additionally, it was found that the protective effect of FTO knockdown was disrupted by ferroptosis inducer (p < 0.01). Conclusion: This study revealed the pathological role of FTO-dependent RNA demethylation in the retinal endothelial homeostasis induced by diabetes. Our findings suggest that FTO mediates diabetes-induced vascular endothelial changes by regulating ferroptosis. This finding could provide a new strategy for intervening in FTO and its related inflammatory pathways to treat diabetic vascular complications