Downstream Signaling Pathways Research Articles

Target Ligand Separation and Identification of Isoforsythiaside as a Histone Lysine-Specific Demethylase 1 Covalent Inhibitor Against Breast Cancer Metastasis.

Histone lysine-specific demethylase 1 (LSD1) is hyperactive in breast cancer, which is associated with the metastasis of the tumor. Current irreversible LSD1 inhibitors are all synthesized by covalently binding to the flavin adenine dinucleotide cofactor, which often have side effects due to the high affinity for a variety of targets. Here, we identified isoforsythiaside (IFA), a natural phenylpropanoid glycoside isolated from Forsythia suspensa, as a novel covalent inhibitor of LSD1. The target ligand fishing technique and LC-MS/MS analysis identified that IFA could covalently bind to the Ser817 residue of LSD1 by α,β-unsaturated ketone moiety to block the amine oxidase-like domain of LSD1. Moreover, RBMS3/Twist1/MMP2, the downstream signaling pathway of LSD1, was activated after IFA treatment to inhibit the metastasis of MDA-MB-231 cells in vitro and in vivo. This study provided novel molecular templates for development of LSD1 covalence-binding inhibitor and laid a foundation for developing agents against breast carcinoma metastasis for targeting LSD1.

Functional Evaluation of a Novel Homozygous ADCY3 Variant Causing Childhood Obesity

Adenylate cyclase 3 (ADCY3) is a transmembrane protein predominantly expressed in the primary cilia of neurons. It plays a vital role in converting ATP to cAMP, a secondary messenger that regulates various downstream signaling pathways such as carbohydrates and lipids metabolism. Homozygous loss-of-function variants in the ADCY3 gene lead to severe early-onset obesity and insulin resistance whereas gain-of-function variants protect against obesity. To describe a novel pathogenic ADCY3 variant implicated in early-onset obesity and functionally characterize this variant via in vitro and in silico validation, we identified a novel homozygous nonsense variant c.2520C>G, p.Thr840X in the ADCY3 gene using gene panel sequencing in a four-year-old girl. She was born to first-cousin consanguineous parents. The patient presented with severe obesity, and exhibited hepatomegaly and insulin resistance, with other biochemical and hormonal tests being normal. In vitro and in silico functional analyses showed downregulation and impaired activation of the ADCY3 protein. Our findings contribute to existing research that supports the role of ADCY3 in the genetic pathogenesis of early-onset obesity. In vitro and in silico functional characterization of the novel p.Thr840X variant showed impaired enzymatic activity leading to receptor loss of function, consistent with the patient’s phenotype. Genetic testing is essential in severe early-onset obesity and early diagnosis could benefit patients with personalized treatment strategies.

The discovery and simulation analysis of a novel mutation c.40 G < T (V14F) in the NRAS gene in patients with colorectal cancer in Saudi Arabia

BackgroundColorectal cancer (CRC) is the most diagnosed cancer in men and the second most common cancer in women and remains associated with high morbidity and mortality in Saudi Arabia. The current understanding of genetic heterogeneity of colorectal cancer biology encourages the identification of the genetic causes of CRC in the Saudi population. MethodsIn this study, we ascertained 89 CRC patients’ tumor samples from Saudi Arabia and investigated the molecular alterations of the NRAS proto-oncogene, GTPase (NRAS) gene by sequencing the collected CRC tumor tissue samples to identify gene mutations. The impact of mutations was analyzed using different bioinformatics tools including Missense3D algorithm of Swiss Model platform, molecular dynamics simulations using YASARA DYNAMICS and Protein Variation Effect Analyzer (PROVEAN) tool. ResultsWe identified a novel mutation c.40 G > T, in one patient in whom valine was replaced by phenylalanine (V14F). Notably, we also identified another mutation in the same codon c.40 G > A where valine is replaced by isoleucine (V14I). Our in-silico analysis revealed that this novel mutation alters the binding affinity of the NRAS gene substantially, and as a result, could have lethal consequences on the downstream signaling genes and pathways including MAPK and PI3K involved in regulating CRC growth and progression. ConclusionsThese findings provide insights into the molecular etiology of CRC in general and particularly in the Saudi population. Thus, these findings in NRAS mutation testing may also guide further treatment modalities, and more personalized therapy may be optimized.

Identification of PSMD2 as a promising biomarker for pancreatic cancer patients based on comprehensive bioinformatics and in vitro studies

BackgroundPancreatic cancer patients have limited treatment options and extremely poor prognosis. Dysregulations of proteasome 26S subunit, non-ATPases (PSMDs) contribute to the development of various cancers, whereas the significance of PSMDs in pancreatic cancer is poorly understood. In the present study, we intended to explore the therapeutic potential of PSMDs in pancreatic cancer. MethodsBased on TCGA database, the expression of PSMDs was analyzed in pancreatic cancer patients. Multivariate Cox regression and Kaplan–Meier survival analyses were conducted to investigate the prognostic value of PSMDs. The correlations between the expression of PSMD2/14 and immune cell infiltration, immune checkpoint genes’ expression, enrichment of signaling pathways, and the sensitivity of chemotherapies were also evaluated. Knockdown and overexpression experiments were performed to explore the biological function of PSMD2/14. Immunoblotting was conducted to detect the downstream signaling pathway of PSMD2. ResultsMost of the PSMDs, except for PSMD5 and PSMD6, were significantly upregulated in pancreatic cancer tissues. Patients with higher grade tumor had increased mRNA levels of PSMD1/2/5/7/8/11/12/14. Survival and multivariate Cox regression analyses indicated that PSMD2 and PSMD14 were biomarkers of worse prognosis. High expression of PSMD2 and PSMD14 was positively correlated with the levels immune checkpoint genes but not with the infiltration of specific immune cell types. In vitro knockdown of PSMD2, but not PSMD14, increased the apoptosis, gemcitabine’s toxicity and inhibited the growth capacity of MIA cells. Conversely, decreased apoptosis and gemcitabine sensitivity along with accelerated cell proliferation ability were observed in PSMD2-overexpressing PANC-1 cells. Mechanistically, PSMD2 activated the AKT/mTOR signaling pathway, consistent with the findings from KEGG and GSEA analysis. The AKT specific inhibitor MK-2206 exhibited higher cytotoxicity in MIA and PANC-1 cells with high PSMD2 expression. Importantly, MK-2206 largely reversed the oncogenic functions of PSMD2 on the growth and proliferation of PANC-1 cells. ConclusionIn summary, our study provided a comprehensive bioinformatics analysis of PSMDs in pancreatic cancer. We identified that PSMD2 acted as a tumor-promoting protein in pancreatic cancer through the activation of the AKT/mTOR pathway. The overexpression of PSMD2 may be a potential biomarker that predicts the response of pancreatic cancer patients to AKT inhibitor treatments.

Targeting GM2 Ganglioside Accumulation in Dementia: Current Therapeutic Approaches and Future Directions.

Dementia in neurodegenerative diseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), and dementia with Lewy bodies (DLB) is a progressive neurological condition affecting millions worldwide. The amphiphilic molecule GM2 gangliosides are abundant in the human brain and play important roles in neuronal development, intercellular recognition, myelin stabilization, and signal transduction. GM2 ganglioside's degradation requires hexosaminidase A (HexA), a heterodimer composed of an α subunit encoded by HEXA and a β subunit encoded by HEXB. The hydrolysis of GM2 also requires a non-enzymatic protein, the GM2 activator protein (GM2-AP), encoded by GM2A. Pathogenic mutations of HEXA, HEXB, and GM2A are responsible for autosomal recessive diseases known as GM2 gangliosidosis, caused by the excessive intralysosomal accumulation of GM2 gangliosides. In AD, PD and DLB, GM2 ganglioside accumulation is reported to facilitate Aβ and α-synuclein aggregation into toxic oligomers and plaques through activation of downstream signaling pathways, such as protein kinase C (PKC) and oxidative stress factors. This review explored the potential role of GM2 ganglioside alteration in toxic protein aggregations and its related signaling pathways leading to neurodegenerative diseases. Further review explored potential therapeutic approaches, which include synthetic and phytomolecules targeting GM2 ganglioside accumulation in the brain, holding a promise for providing new and effective management for dementia.

Roles of PDGF/PDGFR signaling in various organs.

Platelet-derived growth factors (PDGFs) ligands and their corresponding receptors, PDGF receptor (PDGFR)α and PDGFRβ, play a crucial role in controlling diverse biological functions, including cell growth, viability and migration. These growth factors bind to PDGFRs, which are receptor tyrosine kinases present on the surface of target cells. The interaction between PDGFs and PDGFRs induces receptor dimerization and subsequent activation through auto-phosphorylation, which in turn triggers a cascade of intracellular signaling pathways. PDGF/PDGFR signaling is essential for maintaining normal physiological functions, including tissue regeneration and growth. However, dysregulation of this signaling pathway leads to pathological conditions, including fibrosis, atherosclerosis, and cancer development in various organs. The pathological impact of PDGF/PDGFR signaling primarily stems from its capacity to promote excessive cell proliferation, enhanced migration, and increased extracellular matrix deposition, resulting in tissue overgrowth, scarring, and abnormal vessel formation. These processes are integral to the pathogenesis of fibrotic, neoplastic, and vascular disorders. Therefore, understanding these pathways is crucial for developing targeted treatments designed to inhibit PDGF/PDGFR signaling in these diseases. This review delves into the dual role of PDGF/PDGFR signaling in both physiological and pathophysiological contexts across different organs and provides insights into current pharmacological therapies designed to target the PDGF signaling pathway. INTRODUCTION Platelet-derived growth factors (PDGFs) are key signaling molecules that interact with specific cell to modulate various cellular responses. Upon binding to their receptors (PDGFRs), PDGFs initiate dimerization and tyrosine phosphorylation, which activates downstream signaling pathways. The PDGF signaling network comprises four ligands-PDGF-A, PDGF-B, PDGFC, and PDGF-D, that interact with two receptors, PDGFRα and PDGFRβ [1-6]. PDGFRα exhibits broader ligand specificity, binding to PDGF-A, PDGF-B, PDGF-C homodimers, and PDGFAB heterodimers, whereas PDGFRβ specifically binds to PDGFB and PDGF-D homodimers. Under both physiological and pathol.

A non-catalytic role of IPMK is required for PLCγ1 activation in T cell receptor signaling by stabilizing the PLCγ1-Sam68 complex

BackgroundPhospholipase C gamma 1 (PLCγ1) is an important mediator of the T cell receptor (TCR) and growth factor signaling. PLCγ1 is activated by Src family kinases (SFKs) and produces inositol 1,4,5-triphosphate (InsP3) from phosphatidylinositol 4,5-bisphosphate (PIP2). Inositol polyphosphate multikinase (IPMK) is a pleiotropic enzyme with broad substrate specificity and non-catalytic activities that mediate various functional protein-protein interactions. Therefore, IPMK plays critical functions in key biological events such as cell growth. However, the contribution of IPMK to the activation of PLCγ1 in TCR signaling remains mostly unelucidated. The current study aimed to elucidate the functions of IPMK in TCR signaling and to uncover the mode of IPMK-mediated signaling action in PLCγ1 activation.MethodsConcanavalin A (ConA)-induced acute hepatitis model was established in CD4+ T cell-specific IPMK knockout mice (IPMKΔCD4). Histological analysis was performed to assess hepatic injury. Primary cultures of naïve CD4+ T cells were used to uncover the role of mechanisms of IPMK in vitro. Western blot analysis, quantitative real-time PCR, and flow cytometry were performed to analyze the TCR-stimulation-induced PLCγ1 activation and the downstream signaling pathway in naïve CD4+ T cells. Yeast two-hybrid screening and co-immunoprecipitation were conducted to identify the IPMK-binding proteins and protein complexes.ResultsIPMKΔCD4 mice showed alleviated ConA-induced acute hepatitis. CD4+ helper T cells in these mice showed reduced PLCγ1 Y783 phosphorylation, which subsequently dampens calcium signaling and IL-2 production. IPMK was found to contribute to PLCγ1 activation via the direct binding of IPMK to Src-associated substrate during mitosis of 68 kDa (Sam68). Mechanistically, IPMK stabilizes the interaction between Sam68 and to PLCγ1, thereby promoting PLCγ1 phosphorylation. Interfering this IPMK-Sam68 binding interaction with IPMK dominant-negative peptides impaired PLCγ1 phosphorylation.ConclusionsOur results demonstrate that IPMK non-catalytically promotes PLCγ1 phosphorylation by stabilizing the PLCγ1–Sam68 complex. Targeting IPMK in CD4+ T cells may be a promising strategy for managing immune diseases caused by excessive stimulation of TCR.

Advances in CRISPR-Cas9 Technology for Tumor Therapy

Genome editing technology is an emerging toolbox that can specifically target genes. At present, the existing gene editing technologies include ZFN, TALEN, and CRISPR system editing technology with clusters of regularly spaced short palindromic repeats. However, ZFN and TALEN are no longer the preferred tools for gene editing due to some of their problems, and their complex protein design, high cost, and high difficulty are the main reasons that limit their use. The CRISPR system, on the other hand, is able to achieve specific binding by base complementary pairing of sgRNA to the target sequence. Traditional cancer treatment methods have limited efficacy and high recurrence rates, and the disease can be fundamentally treated by inactivating oncogenes or activating tumor suppressor genes through gene editing technology and then changing downstream signaling pathways for cancer treatment. With CRISPR-Cas9 technology, it is possible to have gene knockouts, insertions, and mutations. Here, we will investigate and explore the feasibility of the CRISPR system for tumor treatment, including the following three topics: oncogene knockout, augmentation of tumor suppressor gene expression, and enhancement of engineered T cell viability for effective tumor Through these methods, the CRISPR system is of great help and promising tumor therapy.

Dose and time dependent morphodynamic changes in the ovary of nano-nickel treated rats A SEM study

AimsPresent study demonstrates dose and time dependent effects of NiONPs (<30 nm) on the ovaries of Wistar rat. MethodsFemale rats were gavaged NiONPs or NiOMPs (5 mg/kg b.w.) for 24 h, 15 days and 30 days, euthanized and ovaries thus removed were analyzed for nickel bioaconcentration and processed for scanning electron microscopy. Serum samples were analyzed to compare the effects of nickel nano & microparticles on progesterone and estradiol values. ResultsResults confirmed the bioaccumulation of Ni in ovarian tissue. Its concentration was higher in NiONPs treated rats than NiOMPs treated rats. Progesterone level increased whereas estradiol values decreased in NiONPs and NiOMPs treated rats. SEM results also exhibited dose dependent effects on the morphology of corpoluteal complex. The structural changes varied from formation of blebs to distorted microvilli and germinal epithelium. ConclusionIt is hypothesized that NiONPs/NiOMPs are biodegraded into smaller fragments that conjugate with amino acids and or alter downstream signaling pathways, generate ROS and modulate protein structure activity relationships. Finally, these processes manifest into morphological alterations in the ovary. Biopersistence of nickel in female reproductive system may compromise with fertility and reproductive performance of exposed population.

Modulation of persistent bladder pain in mice: The role of macrophage migration inhibitory factor, high mobility group box-1, and downstream signaling pathways

Background: Repeated intravesical activation of protease-activated receptor-4 (PAR4) serves as a model of persistent bladder hyperalgesia (BHA) in mice, which lasts several days after the final stimulus. Spinal macrophage migration inhibitory factor (MIF) and high mobility group box 1 (HMGB1) are critical mediators in the persistence of BHA. Objective: We aimed to identify effective systemic treatments for persistent BHA using antagonists or transgenic deletions. Methods: Persistent BHA was induced through transurethral instillations of a PAR4-activating peptide (PAR4-AP; 100 &amp;mu;M, 1 h; scrambled peptide, control) under anesthesia, administered on Days 0, 2, and 4. Lower abdominal hypersensitivity was measured on Days 0&amp;ndash;4 and 7&amp;ndash;9. Systemic injections from Days 2&amp;ndash;8 included ISO-1 (a MIF antagonist), ethyl pyruvate (an inhibitor of HMGB1 release), phosphate-buffered saline, or 10% DMSO (vehicle control) in C57BL/6 mice. To examine the role of HMGB1 receptors, Toll-like receptor-4 (TLR4)-null mice or systemic treatment with FPS-ZM1 (receptor for advanced glycation end product [RAGE] antagonist) were used. In addition, TIR-domain-containing adaptor-inducing interferon-&amp;beta; (TRIF)-null mice were tested to assess the involvement of TLR4 signaling pathways. Micturition volume and frequency were assessed on Day 9, and the bladder was histopathologically examined to assess inflammation and edema. Results: MIF antagonism significantly reversed persistent BHA, whereas HMGB1 antagonism led to a partial reduction of persistent BHA. TLR4 deficiency or systemic administration of FPS-ZM1 significantly mitigated persistent BHA, while TRIF-deficient mice experienced a faster onset of BHA. Only MIF or HMGB1 inhibition resulted in increased micturition volume. The histopathological examination revealed no changes in inflammation or edema. Conclusion: MIF and HMGB1, acting through TLR4 and RAGE, mediated persistent BHA, while TRIF might modulate its onset. Further exploration of downstream TLR4 signaling may uncover novel therapeutic targets for treating persistent bladder pain.

Cba-miR-222-3p involved in photoperiod-induced apoptosis in testes of striped hamsters by targeting TRAF7.

The role of miRNAs in the regulation of seasonal reproduction in rodents, particularly in relation to photoperiod changes, is still poorly understood. Previous studies on miRNA transcriptomes of striped hamster (Cricetulus barabensis) testes have indicated that the photoperiodism of testes, especially apoptosis, may be influenced by miRNAs. As a functional miRNA, cba-miR-222-3p in striped hamster testes exhibits suppression under a short photoperiod. To elucidate the potential role of testicular cba-miR-222-3p in the seasonal reproduction of striped hamsters, we exposed male striped hamsters to different photoperiods or injected miRNA agomir into the testes and observed the effects of these treatments, particularly some indicators related to apoptosis. The results showed that the levels of apoptosis in the testes increased in short daylength, accompanied by a significant decrease in cba-miR-222-3p expression and an increase in TRAF7 expression. Dual luciferase reporter assays verified the targeting relationship between cba-miR-222-3p and TRAF7 predicted by bioinformatics. In addition, the expression of TRAF7 decreased in the testes, which injected miRNA agomir, leading to inhibition of apoptosis, and the expression of key genes (MEKK3, p38, p53) in the downstream MAPK signaling pathway of TRAF7 was suppressed. These results suggest that short daylength induces testicular apoptosis in striped hamsters, and one possible mechanism is that the decreased expression of miR-222-3p in testes reduces the repression of TRAF7 translation, thereby activating the MAPK pathway and affecting the level of testicular apoptosis. These findings reveal the potential role of miR-222-3p in animal reproduction and provide new insights into the regulation of rodent populations.

Aging alters the effect of adiponectin receptor signaling on bone marrow-derived mesenchymal stem cells.

Adiponectin receptor signaling represents a promising therapeutic target for age-related conditions such as osteoporosis and diabetes. However, the literature presents conflicting evidence regarding the role of adiponectin signaling in bone homeostasis and fracture repair across different health states, ages, and disease conditions. These inconsistencies may arise from the complex endocrine and paracrine feedback mechanisms regulating adiponectin, as well as the variability in adiponectin isoforms and receptor expressions. In this study, we observed differential expression of adiponectin receptors in the bone marrow (BM) of aged mice, characterized by elevated levels of adiponectin receptor 2 and reduced levels of receptor 1, as corroborated by both single-cell sequencing and invivo staining. Additionally, circulating levels of adiponectin and its local expression were significantly higher in aged mice compared to younger counterparts. Treatment with adiponectin receptor agonist, AdipoRon, enhanced bone regeneration and repair in young mice by promoting osteogenesis and reducing osteoclastogenesis. Conversely, in aged mice, AdipoRon treatment led to cellular senescence, delayed bone repair, and inhibited osteogenic activity. Notably, the adiponectin receptor 1-Wnt and adiponectin receptor 2-MAPK and mTOR signaling pathways were differentially activated in AdipoRon-treated BM mesenchymal stem cells from young and aged mice. Additionally, the NF-κB, and AKT pathways were consistently downregulated in BM macrophages of both age groups following AdipoRon administration. In conclusion, aging significantly modulates the impact of adiponectin receptor signaling on BM mesenchymal stem cells. This modulation is potentially attributable to changes in receptor transcription and distribution, as well as differential activation of downstream signaling pathways.

Paraptosis—A Distinct Pathway to Cell Death

Cell death is a critical biological process necessary for development, tissue maintenance, and defense against diseases. To date, more than 20 forms of cell death have been identified, each defined by unique molecular pathways. Understanding these different forms of cell death is essential for investigating the pathogenesis of diseases such as cancer, neurodegenerative disorders, and autoimmune conditions and developing appropriate therapies. Paraptosis is a distinct form of regulated cell death characterized by cytoplasmic vacuolation and dilatation of cellular organelles like the mitochondria and endoplasmic reticulum (ER). It is regulated by several signaling pathways, for instance, those associated with ER stress, calcium overload, oxidative stress, and specific cascades such as insulin-like growth factor I receptor (IGF-IR) and its downstream signaling pathways comprising mitogen-activated protein kinases (MAPKs) and Jun N-terminal kinase (JNK). Paraptosis has been observed in diverse biological contexts, including development and cellular stress responses in neuronal, retinal, endothelial, and muscle cells. The induction of paraptosis is increasingly important in anticancer therapy, as it targets non-apoptotic stress responses in tumor cells, which can be utilized to induce cell death. This approach enhances treatment efficacy and addresses drug resistance, particularly in cases where cancer cells are resistant to apoptosis. Combining paraptosis-inducing agents with traditional therapies holds promise for enhancing treatment efficacy and overcoming drug resistance, suggesting a valuable strategy in anticancer therapy.

4-Anilinoquinazoline Derivatives as the First Potent NOD1-RIPK2 Signaling Pathway Inhibitors at the Nanomolar Range.

Inflammation is a defense mechanism that restores tissue damage and eliminates pathogens. Among the pattern recognition receptors that recognize danger or pathogenic signals, nucleotide oligomerization domains 1 and 2 (NOD1/2) have been identified to play an important role in innate immunity responses, and inhibition of NOD1 could be interesting to treat severe infections and inflammatory diseases. In this work, we identified the first selective NOD1 versus NOD2 pathway inhibitors at the nanomolar range based on a 4-anilinoquinazoline scaffold. We demonstrated that NOD1 inhibition occurs through the inhibition of receptor interacting protein kinase 2 (RIPK2), which is involved in its downstream signaling pathways. Compound 37 demonstrates no cytotoxicity, a selectivity for RIPK2 over epithelial and vascular endothelial growth factor receptors (EGFR/VEGFR), and a capacity to reduce pro-inflammatory cytokine IL-8 secretion. The structure of the RIPK2-compound 37 complex was resolved by crystallography. The 4-anilinoquinazoline scaffold offers novel perspectives to design NOD1-RIPK2 signaling inhibitors.

TRAF4 promotes lung cancer development by activating tyrosine kinase of EGFR

Objective: To explore the role of tumor necrosis factor receptor-associated factor 4 (TRAF4) in promoting the abnormal activation of epidermal growth factor receptor (EGFR) and its effect on lung cancer cell proliferation, migration and invasion. Methods: Tumor tissues from patients who underwent lung adenocarcinoma resection at The First Affiliated Hospital of Second Military Medical University, from January 2015 to May 2017 were collected, and the expressions of TRAF4 and Ki-67 in lung cancer tissues were detected by immunohistochemistry, the mRNA levels of Cyclin D and Vimentin were detected by real-time fluorescence quantitative PCR (qRT-PCR). The effect of TRAF4 on the tumor growth ability of lung cancer A549 cells was investigated by the xenograft model, the effect of TRAF4 or EGFR on the tumor proliferation ability was detected by using cell counting kit 8 (CCK8) and BrdU assay, and the migration and invasion abilities of tumor cells were detected by Transwell assay. Different structural domain deletion expression vectors of TRAF4 and EGFR were constructed to transfect cells, and the interaction mode of TRAF4 and EGFR was investigated by immunoprecipitation assay. Results: The expression of TRAF4 in non-small cell lung cancer (NSCLC) tissues was positively correlated with the expressions of Ki-67, cyclin D, and vimentin (r2: 0.438, 0.695, and 0.736, respectively, all P＜0.01). Immunohistochemical assay of tumor tissues from NSCLC patients showed that tissues with high expression of TRAF4 were also high in Ki-67. Patients with high TRAF4 expression (TRAF4 positivity >30%) had a shorter progression-free survival (PFS) time than that of patients with low TRAF4 expression (TRAF4 positivity ≤30%) (median PFS of 12 and 19 months, respectively; P=0.034). Traf4-/- cells had a weakened proliferative capacity than traf4+/+ cells and formed tumors with smaller size (P＜0.05). The expression level of Ki-67 in the tumor tissues formed by traf4-/- cells [(45.6±8.7)%] was lower than that in the tumor tissues formed by traf4+/+ cells [(62.3±10.3)%, P=0.015], the mRNA levels of cyclin D (1.01±0.15) and vimentin (1.01±0.12) in the traf4-/- cells were lower than those of the traf4+/+ cells (3.41±0.32 and 3.12±0.18, respectively, both P＜0.05).The western blot results showed that, with the elevated intracellular expression level of TRAF4, phosphorylation level of EGFR was significantly increased in both wild-type EGFR and activation mutant EGFR-expression cells. The capacities of proliferation, migration and invasion of A549 cells was weakened after EGFR knockdown (all P＜0.01). Immunoprecipitation experiments showed that TRAF4 binds to the peptide segment of the near-membrane region of EGFR through the TRAF structural domain, and the mutual binding between EGFR molecules was enhanced under TRAF4 overexpression conditions. Increasing TRAF4 expression promoted EGFR molecular phosphorylation and activation of downstream signaling. Conclusions: TRAF4 expression is elevated in NSCLC tissues and tumor cells, which promotes tumor proliferation, migration and invasion. TRAF4 directly binds to EGFR molecules, enhances its own phosphorylation and activates the downstream signaling pathway by promoting the interaction between EGFR molecules.

Ginsenoside Rk3 Treats Corneal Injury Through the HMGB1/TLR4/NF-κB Pathway.

The cornea serves as a vital protective shield for the eye, safeguarding its intricate internal structures from external threats. Damage to the cornea compromises this protective function, triggering inflammation and potentially causing long-term harm. While ginsenoside Rk3 has demonstrated potential for repairing the corneal barrier and reducing inflammation, its effectiveness in treating corneal damage remains relatively unexplored. This comprehensive study uses both in vivo and in vitro models to investigate the therapeutic capabilities of ginsenoside Rk3. Using two models of corneal damage, a benzalkonium chloride-induced mouse model and a high osmolarity-induced human corneal epithelial cell model, we scrutinized the effects of ginsenoside Rk3 treatment. Our results showed that ginsenoside Rk3-treated mice manifested reduced corneal damage and inflammation compared with their untreated counterparts. Furthermore, mice treated with ginsenoside Rk3 exhibited an organized arrangement of corneal cells and diminished stromal layer thickness, indicating reparative properties of ginsenoside Rk3. Additionally, ginsenoside Rk3 increased the expression of tight junction proteins, suppressed inflammatory factors, and decreased HMGB1 protein expression, thereby modulating downstream signaling pathways. Collectively, our findings present compelling evidence that ginsenoside Rk3 is a promising therapeutic option for corneal injury. By repairing the corneal barrier, mitigating inflammation, and modulating specific protein levels, ginsenoside Rk3 opens new avenues for managing corneal damage.

Multifunctional role of the tumor associated monocytes/macrophages in the metastatic potential of inflammatory breast cancer.

Inflammatory breast cancer (IBC) is the most aggressive and lethal phenotype form of breast cancer, which afflicts young women at high incidence in North Africa compared to other continents of the world. IBC is characterized by highly metastatic behavior and possesses specific pathobiological properties different from non-IBC. IBC disease displays unusual common properties at typical presentation, including positive metastatic lymph-nodes, high infiltration of tumor associated monocytes/macrophages, rapid progression to distant metastasis, and possibly the production of a unique repertoire of growth factors, cytokines and chemokines, as well as a striking association with different polarized macrophages compared to non-IBC. Indeed, tumor associated monocytes/macrophages (TAM/M) play a crucial role in breast cancer development. Previously, we showed that cross talk between IBC cells and patient derived TAMs occurs via secretion of inflammatory mediators from TAMs that act on specific extracellular domain receptors activating down-stream signaling pathways that promote the epithelial-to-mesenchymal transition, cancer cell invasion, IBC stem cell properties, drug resistance, local and metastatic recurrence of residual tumor cells and other key markers of malignancy, including in vitro colony formation capacity. In this mini review, we will discuss the role of TAMs in IBC cancer metastatic potential and molecules involved. The review also discusses the recent discoveries in the field of IBC research.

Association between premature vascular smooth muscle cells senescence and vascular inflammation in Takayasu’s arteritis

ObjectivesArterial wall inflammation and remodelling are the characteristic features of Takayasu’s arteritis (TAK). It has been proposed that vascular smooth muscle cells (VSMCs) are the main targeted cells of inflammatory...

Up-Regulation of miR-625-5p Correlates with Suppressed Sox2, Increased Apoptosis, and Cell Cycle Arrest via The PI3K/AKT Signalling Pathway in Acute Myeloid Leukaemia

Background: Up-regulation of the microRNA-625 and abnormal expression of the Sox2 gene have been studied and seen in several tumors. Few reports have also shown the aberrant expression of miR-625 and Sox2 expression in various cancers. Several studies have also confirmed that phosphatidylinositol 3' -kinase /protein kinase B pathways regulate hematological malignancies, including Acute Myeloid Leukemia (AML). Thus, this study aimed to investigate the effects of mir-625 up-regulation on proliferation, apoptosis, and cell cycle by targeting the Sox2 gene via the downstream Akt signaling pathway and cell cycle regulators, such as p21, p27, and cyclin E in the KG-1 cell line. Materials and Methods: Cells obtained from the KG-1 cell line were cultured and transfected with plasmid DNA (miR-625) and scrambled as the control using the Lonza electroporation system. Flow cytometry was used to evaluate cell cycle, proliferation, and apoptosis. Relative gene expression was validated by qRT-PCR. All data were analyzed using graph pad prism 7.01 and REST 2009. Results: KG-1 cells transfected with the mir625-GFP construct showed decreased proliferation, increased apoptosis, and induced cell cycle arrest. Low levels of Sox2, p21, cyclin E, and up-regulation of p27 were confirmed and validated by qRT-PCR ( P &lt; 0.05 ). Conclusion: MiR-625 can be a promising approach to aid in the treatment of AML. However, further studies are required in this field

G-CSFR-induced leukocyte transendothelial migration during the inflammatory response is regulated by the ICAM1-PKCa axis: based on multiomics integration analysis

As an indispensable inflammatory mediator during sepsis, granulocyte colony-stimulating factor (G-CSF) facilitates neutrophil production by activating G-CSFR. However, little is known about the role of intracellular downstream signalling pathways in the induction of inflammation. To explore the functions of molecules in regulating G-CSFR signalling, RNA sequencing and integrated proteomic and phosphoproteomic analyses were conducted to predict the differentially expressed molecules in modulating the inflammatory response after G-CSFR expression was either up- or downregulated, in addition to the confirmation of their biological function by diverse experimental methods. In the integrated bioinformatic analysis, 3190 differentially expressed genes (DEGs) and 1559 differentially expressed proteins (DEPs) were identified in multiple-group comparisons (p < 0.05, FC > ± 1.5) using enrichment analyses, as well as those classic pathways such as the TNF, NFkappaB, IL-17, and TLR signalling pathways. Among them, 201 proteins, especillay intercellular cell adhesion molecule-1 (ICAM1) and PKCa, were identified as potential molecules involved in inflammation according to the protein–protein interaction (PPI) analysis, and the leukocyte transendothelial migration (TEM) pathway was attributed to the intervention of G-CSFR. Compared with the control and TNF-a treatment, the G-CSFR (G-CSFROE)-overexpressing led to an obvious increase in the number of leukocytes with the TEM phenotype. Mechanically, the expression of ICAM1 and PKCa was significantly up- and downregulated by G-CSFROE, which directly led to increased TEM; moreover, PKCa expression was negatively regulated by ICAM1 expression, leading to aberrant leukocyte TEM. Altogether, the ICAM1‒PKCa axis was found a meaningful target in the leukocyte TEM induced by G-CSFR upregulation.