Combinatorial Chemistry & High Throughput Screening - Online First

Transcription factors (TF) are the central regulatory hubs of signaling pathways in eukaryotic cells. Here, we explored the abnormal expression of TF in septic-induced lung injury by sequencing.

The levels of target proteins were detected using Western Blot and Elisa. Cell function was evaluated using CCK8 and transwell assays. A double luciferase reporter assay was performed to detect interactions between target molecules.

We found that TF glioma-associated oncogene (GLI) family zinc finger 3 (GLI3) was abnormally low expressed in a lipopolysaccharide (LPS) induced acute lung injury (ALI) cell model. In an in vitro model, GLI3 overexpression promoted the proliferation and migration and inhibited apoptosis of lung epithelial cells in LPS-induced inflammatory environment. Importantly, GLI3 overexpression inhibited the secretion of inflammatory factors IL-1β, IL-6, and TNF-α. Additionally, miR-143-3p inhibited the expression of GLI3. MiR-143-3p inhibitor alleviated the cell damage caused by LPS, while knocking down GLI3 counteracted this effect, indicating that miR-143-3p downregulated GLI3 and inhibited its anti-inflammatory effect. Secreted frizzled related protein-1 (SFRP1) was upregulated in LPS-treated cells and SFRP1 promoter interacted with GLI3, suggesting that SFRP1 was a target of TF GLI3. Co-transfection with GLI3 knockdown and SFRP1 overexpression plasmids attenuated the secretion of inflammatory factors IL-1β, IL-6, and TNF-α caused by GLI3 knockdown in LPS-treated cells, indicating that SFRP1 plays an anti-inflammatory role as a GLI3 target in the ALI cell model.

miR-143-3p caused degradation of GLI3 mRNA and thus inhibited the transcription of SFRP1, leading to decreased proliferation and increased levels of inflammatory factors, providing new potential targets for the clinical diagnosis and treatment of ALI.

Benzoimidazo[1,2-a]pyrimidines are important compounds that have many useful effects in the body. They can help fight cancer, fungal infections, inflammation, and viruses. They can also help with various other health conditions. They can act as antineoplastic, antitubercular, parasitical activity, benzodiazepine receptor agonists, calcium channel blockers, potent P38 MAP kinase inhibitors, TIE-2 and/or VEGFR2 inhibitory activities, protein kinase inhibitors, and T cell activation. There are different methods to make the benzoimidazo[1,2-a]pyrimidines. Some of them dealth with the one-pot threecomponent condensation reactions of β-dicarbonyl compounds, aldehyde and 1H-benzo[d]imidazol-2-amine in the presence of catalyst. Although the synthesis of this group of compounds has been done before, and the products have been identified from the spectroscopic point of view, the kinetics and reaction mechanism have not been investigated. The strength of these calculations is that evaluation of the activation energy of various steps suggests possible mechanisms, probable mechanisms, and valuable synthetic intermediates.

In this report, seven possible mechanisms for synthesizing the benzoimidazo[1,2-a]pyrimidines have been investigated using density functional theory (DFT) at the B3LYP/6-311G** level of theory. Each synthetic route involves condensation of the benzaldehyde, indanedione and 1H-benzo[d]imidazol-2-amine molecules to yield the proposed product. The calculations showed that the suggested method has six steps; its initiation step includes the Knoevenagel reaction between indanedione and aldehyde, and the rate determining state is dehydration in the fifth step.

Six potential pathways for the reaction will occur. Then, we focused on the best pathway and studied it in detail. The ways that three chemicals-indanedione (R1), benzaldehyde (R2), and 1H-benzo[d]imidazol-2-amine (R3) react with each other were studied using ab-initio program by ChemBio3D, Gauss View, and Gaussian 09. The Density Functional Theory (DFT) using the B3LYP basis set was used to improve the arrangement of molecules involved in the three-part creation of a specific compound called 12-phenyl-5H-benzo[4,5]imidazo[1,2-a]indeno[1,2-d]pyrimidin-13(12H)-one (P).

During the study of the six mechanisms, the proposed pathway 2 is the best mechanism for this reaction because its rate-determining step has the lowest activation energy value. This route consists of 6 steps, the fifth step of which is related to the conversion of IM4 to IM5 (relative ∆E: 109.80 Kj/mol), during which a dehydration reaction is performed, and this step occurs by passing through transition state TS5 (Total Energy (Hart./particles: -1194.747403).

Autoimmune thyroiditis (AIT) is one of the most common autoimmune diseases and often causes hypothyroidism in patients. As a traditional formulation in my country, Buzhong Yiqi decoction (BZYQD) has significant effects in improving clinical symptoms of AIT and reducing autoantibody titers, but its specific mechanism of action needs to be further explored. The purpose of this study was to explore the effective targets and related mechanisms of Buzhong Yiqi decoction in AIT mice based on transcriptome sequencing technology.

Forty NOD.H-2h4 mice were selected and 0.05% NaI was drinking ad libitum for 8 weeks to establish AIT mice, and drug intervention was performed according to groups for 8 weeks. The groups were as follows: control group, Model Group, Buzhong Yiqi Decoction group (9.56 g·kg-1) and Positive control group (Se yeast tablets, 3.033×10-5 g·kg-1), of which Buzhong Yiqi Decoction was the clinical equivalent dose. Thyroid tissues of the Model Group, blank group and Buzhong Yiqi Decoction group were subjected to transcriptome sequencing to analyze the expression of differential genes, and enrichment analysis was carried out. Hematoxylin and eosin staining (HE staining) was used to detect the pathological changes in thyroid tissues, and enzyme-linked immunosorbent assay (ELISA) was used to detect the content of serum thyroglobulin antibody (TGAb) to determine the intervention effect of Buzhong Yiqi Decoction; Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) was performed based on the transcriptome sequencing results to detect the expression of TLR8, JUN, TICAM2, TIRAP, and IL-1β mRNA in thyroid tissue.

According to the transcriptome results, compared with the blank group, there were 327 significantly up-regulated genes and 440 significantly down-regulated genes in the Model Group; compared with the Model Group, there were 502 significantly up-regulated genes and 380 significantly down-regulated genes in the Buzhong Yiqi Decoction group, mainly enriched in immune inflammation and other related pathways including Toll-like receptors. Animal experiments showed that compared with the control group, the model group had obvious lymphocyte infiltration in thyroid tissue under light microscope, a significant increase in inflammatory cells, a significant increase in TGAb content in serum, and a significant increase in TLR8, JUN, TICAM2, TIRAP, IL-1β mRNA expression was observed (P<0.05 or P<0.01). Compared with the Model Group, Buzhong Yiqi Decoction could significantly improve the inflammatory damage of thyroid tissue in AIT mice, reduce the content of TGAb in serum, and down-regulate the expression of TLR8, JUN, TICAM2, TIRAP, IL-1β mRNA (P<0.05 or P<0.01).

Buzhong Yiqi Decoction can effectively improve the inflammatory damage of AIT, and inhibiting the abnormal activation of the Toll-like receptor pathway may be one of its intervention mechanisms.

The objective of this research is to demonstrate that alignment-free bioinformatics approaches are effective tools for analyzing the similarity and dissimilarity of protein sequences. All numerical parameters representing sequences are expressed analytically, ensuring precision, clarity, and efficient processing, even for large datasets and long sequences. Additionally, a novel approach for identifying previously unknown virus strains is introduced.

A novel approach is proposed, integrating the unique features of our newly developed method, the 20D-Dynamic Representation of Protein Sequences, with the K-means clustering algorithm. The sequences are represented as clouds of material points in a 20-dimensional space (20D-dynamic graphs), with their spatial distribution being unique to each protein sequence. The numerical parameters, referred to as descriptors in molecular similarity theory, represent quantities characteristic of dynamic systems and serve as input data for the K-means clustering algorithm.

Examples of the application of the approach are presented, including projections of the 20D-dynamic graphs onto 3D spaces, which serve as a visual tool for comparing sequences. Additionally, cluster plots for the analyzed sequences are provided using the proposed method.

It has been demonstrated that the 20D-Dynamic Representation of Protein Sequences, combined with the K-means clustering algorithm, successfully classifies subtypes of influenza A virus strains.

Osteosarcoma (OS), a malignant tumor originating in bone or cartilage, primarily affects children and adolescents. Notably, substantial alterations in mitochondrial energy metabolism have been observed in OS; however, the specific contribution of mitochondrial-related genes (MRGs) to OS pathogenesis and prognosis remains unclear. Herein, we identified novel diagnostic biomarkers associated with mitochondrial-related processes in OS via comprehensive bioinformatics analysis.

OS mRNA expression profiles were retrieved from GSE16088 and GSE19276 databases. Mitochondrial-related differentially expressed genes (MitoDEGs) were identified by integrating differentially expressed analysis with mitochondrial-localized genes. A protein-protein interaction network was constructed, and machine learning algorithms (LASSO regression analysis and SVM-RFE) identified characteristic MitoDEGs. Subsequently, immune cell infiltration, microenvironment analysis, and single-cell RNA sequencing (scRNA-seq) analyzed differences in characteristic MitoDEGs, and RT-PCR was used for in vitro verification of characteristic MitoDEGs.

MitoDEGs in OS were significantly enriched in the pathways associated with mitochondrial function and immune regulation. Two MitoDEGs, UCP2 and PRDX4, were identified via LASSO and SVM-RFE. Correlation analysis demonstrated a close association between UCP2 and PRDX4 expression levels and immune cell infiltration, particularly in CD8+ T and native CD4+ T cells, as observed in both immune cell and scRNA-seq analyses. Furthermore, RT-PCR confirmed the expression levels of UCP and PRDX4 at the cellular level, which was consistent with the bioinformatics results.

This study identified UCP2 and PRDX4 as characteristic MitoDEGs and potential diagnostic biomarkers for OS using machine learning algorithms. These findings provide novel insights into the clinical applications of these biomarkers for OS diagnosis.

This study aimed to assess the safety and effectiveness of carbamazepine in treating trigeminal neuralgia in contrast to gabapentin. Hence, a systematic review and meta-analysis of randomised controlled trials had been carried out.

The relevant studies were searched in PubMed and filtered according to the inclusion and exclusion criteria. Independently, two reviewers chose the studies, evaluated the quality of the investigations, and retrieved the data. RevMan was used for analysis when the data were collected and entered into the data extraction sheet. In addition to heterogeneity, the overall estimate measures were computed as mean differences with a 95% confidence interval for continuous data and relative risk for dichotomous data. To investigate the impact of outliers on the result, a sensitivity analysis was performed. A funnel plot was used to qualitatively evaluate the publishing bias. A total of 1,650 participants from 19 randomised controlled trials were evaluated.

The meta-analysis revealed that the group receiving gabapentin therapy had a similar overall effective rate to the group receiving carbamazepine therapy (OR = 1.94, 95% CI 1.46, 2.57, P = 0.32). Additionally, our meta-analysis revealed that the group receiving gabapentin therapy witnessed a significantly lower risk of adverse reactions than the group receiving carbamazepine therapy (OR= 0.29, 95% CI 0.22, 0.387, P<0.00001).

In summary, the current trials comparing carbamazepine and gabapentin have had inadequate methodological quality. It is not possible to conclude that gabapentin is more effective than carbamazepine in terms of adverse effects based on the evidence that is currently available.

This study aimed to examine the impact of quercetin on a mouse model of endometriosis and elucidate its underlying mechanisms.

An endometriosis model was established using C57BL/6 mice, which were divided into three groups: 1) sham group, 2) model group, and 3) model group treated with daily gavage administration of 100 mg/kg/d quercetin. After three weeks, mice were euthanized, and histopathological examination was performed using hematoxylin and eosin (HE) staining. The microstructure of the lesions was examined using electron microscopy, and the expression level of S-adenosylmethionine (SAM) was measured using enzyme-linked immunosorbent assay (ELISA). Additionally, the expressions of related proteins, such as the peroxisome proliferator-activated receptor-γ (PPARγ) and methionine adenosyl-transferase 2A (MAT2A), were analyzed via Western blotting. Immunohistochemistry was employed to evaluate the expressions of Ki67, vimentin, vascular endothelial growth factor (VEGF), and Caspase-1.

The endometriosis mice model was successfully established and characterized by ectopic lesions displaying transparent or red vesicular or nodular features with a visible vascular network on the surface. In the model group, endometrial epithelial hyperplasia exhibited columnar morphology, increased mesenchymal cell numbers, and regular cell morphology. Conversely, in the medication group, endometrial stromal cell numbers were sparse, cell morphology was irregular, and numerous vacuoles were observed in the endometrial tissue, indicative of apoptotic cell morphology changes. In comparison to the sham group, no statistically significant alterations were observed in the expression levels of SAM (P > 0.05). Conversely, the expression of PPARγ exhibited a notable decline. MAT2A, PRMT5, cyclin D1, and C-MYC expressions were increased, and vimentin, Ki67, VEGF, and caspase-1 expressions were strongly positive, with statistically significant differences (P < 0.05). In contrast, compared to the model group, the quercetin intervention group exhibited significantly reduced ectopic lesion weights, increased PPARγ expression, and significantly reduced protein expression levels of MAT2A, PRMT5, SAM, cyclin D1, and C-MYC. Furthermore, expressions of vimentin, Ki67, VEGF, and caspase-1 were weakly positive, with statistically significant differences (P < 0.05).

Quercetin modulated the transcription of the MAT2A/PRMT5 gene by activating PPARγ activity, thereby influencing the ectopic implantation and growth of endometrial cells.

Trauma, resulting from mechanical factors, entails damage to human tissues or organs. Whether occurring during times of war or peace, trauma is prevalent, particularly skin defects arising from surgery or external injuries. The development and design of effective wound dressings have become paramount. Bingqing Gao(BQG), rooted in Chinese folk medicine, is employed explicitly in trauma treatment based on traditional Chinese medicine (TCM) theory. This study aims to elucidate how BQG facilitates full-thickness skin wound healing in Sprague Dawley (SD) rats.

Data collection commenced using two approaches: retrieval from TCM system pharmacology databases (TCMSP) and literature mining to compile the practical chemical components and targets of BQG. A drug-target network was constructed. Subsequently, disease targets related to wound healing were collected to select core targets and pathways, building a drug-disease target protein-protein interaction (PPI) network using the ClusterONE algorithm to identify core genes. Gene Ontology (GO) and KEGG enrichment analyses were conducted based on the Metascape database. Finally, molecular docking validation was performed on the screened core targets and core components. In terms of in vivo experimentation, an SD rat full-thickness skin defect model was established, and varying doses of BQG were applied. Healing area, HE staining, Masson staining, ELISA, PCR, and other methods were employed to validate cytokines, differential proteins, and pathways. The study collectively discusses the mechanism and targets by which BQG promotes full-thickness skin wound healing in SD rats.

Through network pharmacology screening, we identified various active components, including resveratrol, Lithospermic acid B, sanguiinH-2, asernestioside A_qt, kaempferol, daidzein, quercetin, apigenin, and Medicarpin. The core targets encompass Interleukin-6 (IL-6), Protein Kinase B (AKT1), Vascular Endothelial Growth Factor A (VEGFA), Interleukin-1 beta (IL-1β), Tumor Protein 53 (TP53), Epidermal Growth Factor Receptor (EGFR), Tumor Necrosis Factor (TNF), Albumin (ALB), among others. Potential signaling pathways include Phosphoinositide 3-kinase (PI3K)/AKT, Tumor Necrosis Factor (TNF), Hypoxia-Inducible Factor-1 (HIF-1), and more. Molecular docking studies suggest a robust binding interaction between the active components of BQG and disease targets, indicating a potential regulation of cytokines through the PI3K/AKTsignaling pathway, thereby promoting wound healing. The results of the in vivo experiment revealed that, in comparison to the model group, both the rhb-FGF group and BQG-H group exhibit a noteworthy increase in the expression levels of PI3K and AKT genes. Concurrently, there is a significant decrease in the levels of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α. Additionally, there is a substantial increase in the levels of Transforming Growth Factor-beta (TGF-β) and Vascular Endothelial Growth Factor (VEGF).

Network pharmacology results indicate that BQG promotes wound healing through multiple components, targets, and pathways. In vivo experimental results suggest that BQG may activate the PI3K/AKTsignaling pathway, inhibit the production and release of related pro-inflammatory cytokines IL-1β, IL-6, and TNF-α, promote VEGF generation at the injury site, and enhance TGF-β signaling transduction, effectively regulates the inflammatory response at the site of injury, promotes vascular regeneration in the injury area, and induces the proliferation and migration of cells in the injury area, ultimately contributing to wound healing. This study establishes the foundation for a more profound understanding of the molecular mechanisms underlying BQG's promotion of wound healing and offers insights for future drug research on BQG.

With the accelerated aging of the population, aging has emerged as a major risk factor for osteoporosis (OP). This study aims to investigate the relationship and shared molecular mechanisms between OP and aging through various genetic approaches.

Single-cell data from the peripheral blood of osteoporosis patients, aging individuals, and healthy controls were integrated to analyze characteristic changes in cell subpopulations. Differentially expressed genes (DEGs) were then identified within core subpopulations, and Mendelian randomization (MR) analysis was employed to explore potential causal links between key genes and OP. Additionally, an OP model was established in rats, and mRNA levels of key genes were measured using RT-qPCR.

Through the integration, filtering, and analysis of scRNA-seq data, an increased proportion of CD4⁺ effector memory T (CD4⁺ TEM) cells were identified in OP and aging samples, marking them as a core subpopulation. Differential expression analysis identified 49 DEGs, and further analysis through Mendelian Randomization (MR) identified three key genes (KLRB1, NR4A2, and S100A4) significantly associated with OP. Notably, the upregulation of KLRB1 and S100A4 may enhance the interactions within T cells and with other cell subgroups. At the same time, the downregulation of NR4A2 could impede communication between T cells and other cell subpopulations. The RT-qPCR results indicated that NR4A2 was significantly downregulated in the OP group.

This study conducted a comprehensive analysis of the potential link between OP and aging, identifying CD4⁺ TEM cells as the core cell subgroup in OP and aging samples. It further revealed the causal relationship between KLRB1, NR4A2, and S100A4 and the occurrence of OP. The upregulation of KLRB1 and S100A4 may contribute to OP pathogenesis by promoting interactions between CD4+ TEM cells and other cell subgroups, providing new insights for molecular targeting and immunotherapy of OP.