Development and Validation of a Proximity Labeling Fusion Protein Construct to Identify the Protein-Protein Interactions of Transcription Factors.

Dynamic interactions between transcription factors govern changes in gene expression that mediate changes in cell state accompanying injury response and regeneration. Transcription factors frequently function as obligate dimers whose activity is often modulated by post-translational modifications. These critical and often transient interactions are not easily detected by traditional methods to investigate protein-protein interactions. This chapter discusses the design and validation of a fusion protein involving a transcription factor tethered to a proximity labeling ligase, APEX2. In this technique, proteins are biotinylated within a small radius of the transcription factor of interest, regardless of time of interaction. Here we discuss the validations required to ensure proper functioning of the transcription factor proximity labeling tool and the sample preparation of biotinylated proteins for mass spectrometry analysis of putative protein interactors.

Saikosaponin A Recovers Impaired Filaggrin Levels in Inflamed Skin by Downregulating the Expression of FRA1 and c-Jun.

Filaggrin (FLG) is an essential structural protein expressed in differentiated keratinocytes. Insufficient FLG expression contributes to the pathogenesis of chronic inflammatory skin diseases. Saikosaponin A (SSA), a bioactive oleanane-type triterpenoid, exerts anti-inflammatory activity. However, the effects of topically applied SSA on FLG expression in inflamed skin remain unclear. This study aimed to evaluate the biological activity of SSA in restoring reduced FLG expression. The effect of SSA on FLG expression in HaCaT cells was assessed through various biological methods, including reverse transcription PCR, quantitative real-time PCR, immunoblotting, and immunofluorescence staining. TNFα and IFNγ decreased FLG mRNA, cytoplasmic FLG protein levels, and FLG gene promoter-reporter activity compared to the control groups. However, the presence of SSA restored these effects. A series of FLG promoter-reporter constructs were generated to investigate the underlying mechanism of the effect of SSA on FLG expression. Mutation of the AP1-binding site (mtAP1) in the -343/+25 FLG promoter-reporter abrogated the decrease in reporter activities caused by TNFα + IFNγ, suggesting the importance of the AP1-binding site in reducing FLG expression. The SSA treatment restored FLG expression by inhibiting the expression and nuclear localization of FRA1 and c-Jun, components of AP1, triggered by TNFα + IFNγ stimulation. The ERK1/2 mitogen-activated protein kinase signaling pathway upregulates FRA1 and c-Jun expression, thereby reducing FLG levels. The SSA treatment inhibited ERK1/2 activation caused by TNFα + IFNγ stimulation and reduced the levels of FRA1 and c-Jun proteins in the nucleus, leading to a decrease in the binding of FRA1, c-Jun, p-STAT1, and HDAC1 to the AP1-binding site in the FLG promoter. The effect of SSA was evaluated in an animal study using a BALB/c mouse model, which induces human atopic-dermatitis-like skin lesions via the topical application of dinitrochlorobenzene. Topically applied SSA significantly reduced skin thickening, immune cell infiltration, and the expression of FRA1, c-Jun, and p-ERK1/2 compared to the vehicle-treated group. These results suggest that SSA can effectively recover impaired FLG levels in inflamed skin by preventing the formation of the repressor complex consisting of FRA1, c-Jun, HDAC1, and STAT1.

Protein Kinases in Obesity, and the Kinase-Targeted Therapy.

The action of protein kinases and protein phosphatases is essential for multiple physiological responses. Each protein kinase displays its own unique substrate specificity and a regulatory mechanism that may be modulated by association with other proteins. Protein kinases are classified as dual-specificity kinases and dual-specificity phosphatases. Dual-specificity phosphatases are important signal transduction enzymes that regulate various cellular processes in coordination with protein kinases and play an important role in obesity. Impairment of insulin signaling in obesity is largely mediated by the activation of the inhibitor of kappa B-kinase beta and the c-Jun N-terminal kinase (JNK). Oxidative stress and endoplasmic reticulum (ER) stress activate the JNK pathway which suppresses insulin biosynthesis. Adenosine monophosphate (AMP)-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) are important for proper regulation of glucose metabolism in mammals at both the hormonal and cellular levels. Additionally, obesity-activated calcium/calmodulin dependent-protein kinase II/p38 suppresses insulin-induced protein kinase B phosphorylation by activating the ER stress effector, activating transcription factor-4. To alleviate lipotoxicity and insulin resistance, promising targets are pharmacologically inhibited. Nifedipine, calcium channel blocker, stimulates lipogenesis and adipogenesis by downregulating AMPK and upregulating mTOR, which thereby enhances lipid storage. Contrary to the nifedipine, metformin activates AMPK, increases fatty acid oxidation, suppresses fatty acid synthesis and deposition, and thus alleviates lipotoxicity. Obese adults with vascular endothelial dysfunction have greater endothelial cells activation of unfolded protein response stress sensors, RNA-dependent protein kinase-like ER eukaryotic initiation factor-2 alpha kinase (PERK), and activating transcription factor-6. The transcriptional regulation of adipogenesis in obesity is influenced by AGC (protein kinase A (PKA), PKG, PKC) family signaling kinases. Obesity may induce systemic oxidative stress and increase reactive oxygen species in adipocytes. An increase in intracellular oxidative stress can promote PKC-ß activation. Activated PKC-ß induces growth factor adapter Shc phosphorylation. Shc-generated peroxides reduce mitochondrial oxygen consumption and enhance triglyceride accumulation and lipotoxicity. Liraglutide attenuates mitochondrial dysfunction and reactive oxygen species generation. Co-treatment of antiobesity and antidiabetic herbal compound, berberine with antipsychotic drug olanzapine decreases the accumulation of triglyceride. While low-dose rapamycin, metformin, amlexanox, thiazolidinediones, and saroglitazar protect against insulin resistance, glucagon-like peptide-1 analog liraglutide inhibits palmitate-induced inflammation by suppressing mTOR complex 1 (mTORC1) activity and protects against lipotoxicity.

A novel biomarker GATM suppresses proliferation and malignancy of cholangiocarcinoma cells by modulating the JNK/c-Jun signalling pathways.

Background: Cholangiocarcinoma (CCA) is the second most common primary malignancy of the liver and is associated with poor prognosis. Despite the emerging role of glycine amidinotransferase (GATM) in cancer development, its function in CCA remains elusive. This study investigated the biological significance and molecular mechanisms of GATM in CCA. Method: GATM expression was measured using immunohistochemistry and western blotting. Cell proliferation, migration, and invasion were assessed through CCK-8, EdU, clone formation, wound healing, and Transwell assays. Rescue experiments were performed to determine whether the JNK/c-Jun pathway is involved in GATM-mediated CCA development. Immunoprecipitation and mass spectrometry were performed to screen for proteins that interact with GATM. The role of GATM in vivo was investigated according to the xenograft experiment. Result: GATM expression was downregulated in CCA tissues and cells (p < 0.05) and had a significant suppressive effect on CCA cell proliferation, migration, and invasion in vitro as well as on tumour growth in vivo (p < 0.05); conversely, GATM knockdown promoted these phenotypes (p < 0.05). Notably, GATM inhibited the JNK/c-Jun pathway, and JNK activation abrogated GATM's antitumor effects (p < 0.05). Isocitrate dehydrogenase 1 (IDH1) interacts with GATM, and IDH1 knockdown significantly attenuated GATM protein degradation. Overexpression of IDH1 restored the biological function of CCA by reversing the inhibition of JNK/c-Jun pathway phosphorylation by GATM (p < 0.05). Conclusion: GATM acts as a tumour suppressor in CCA by regulating the phosphorylation of the JNK/c-Jun pathway. IDH1 interacted with GATM to regulate CCA progression.

A weekly 4-methylpyrazole treatment attenuates the development of non-obese metabolic dysfunction-associated steatotic liver disease (MASLD) in male mice: Role of JNK.

BACKGROUND: 4-methylpyrazole (4MP, fomepizole) is a competitive inhibitor of alcohol dehydrogenase (ADH) preventing the metabolism of ethylene glycol and methanol, respectively, into their toxic metabolites. 4MP seems also to possess a potential in the treatment of intoxication from other substance, for example, acetaminophen, and to modulate JNK-dependent signalling. Here, we determined if a treatment with 4MP once weekly affects the development of diet-induced non-obese metabolic dysfunction-associated steatotic liver disease (MASLD) in C57BL/6 mice. METHODS: Male C57BL/6 mice (6-8 weeks old, n = 7-8/group) were pair-fed either a liquid control diet (C) or a liquid sucrose-, fat- and cholesterol-rich diet (SFC) for 8 weeks while being concomitantly treated with 4MP (50 mg/kg bw i.p.) or vehicle once a week. Liver damage, inflammatory markers and glucose tolerance were assessed. Moreover, in endotoxin-challenged J774A.1 cells pretreated with 4MP, pro-inflammatory markers were assessed. RESULTS: The concomitant treatment of SFC-fed mice with 4MP attenuated the increase in JNK phosphorylation and pro-inflammatory markers like IFNγ, IL-6 and 3-nitrotyrosine protein adducts in liver tissue found in vehicle-treated SFC-fed mice, while not affecting impairments of glucose tolerance or the increase in portal endotoxin levels. Moreover, a pretreatment of endotoxin-stimulated J774A.1 cells with 4MP significantly attenuated the increases in JNK phosphorylation and pro-inflammatory mediators like IL-6 and Mcp1. CONCLUSIONS: Taken together, our results suggest that a treatment with 4MP once weekly attenuates the activation of JNK and dampens the development of non-obese MASLD in mice.

Update on JNK inhibitor patents: 2015 to present.

INTRODUCTION: c-Jun N-terminal kinase (JNK) regulates various biological processes through the phosphorylation cascade and is closely associated with numerous diseases, including inflammation, cardiovascular diseases, and neurological disorders. Therefore, JNKs have emerged as potential targets for disease treatment. AREAS COVERED: This review compiles the patents and literatures concerning JNK inhibitors through retrieving relevant information from the SciFinder, Google Patents databases, and PubMed from 2015 to the present. It summarizes the structure-activity relationship (SAR) and biological activity profiles of JNK inhibitors, offering valuable perspectives on their potential therapeutic applications. EXPERT OPINION: The JNK kinase serves as a novel target for the treatment of neurodegenerative disorders, pulmonary fibrosis, and other illnesses. A variety of small-molecule inhibitors targeting JNKs have demonstrated promising therapeutic potential in preclinical studies, which act upon JNK kinases via distinct mechanisms, encompassing traditional ATP competitive inhibition, covalent inhibition, and bidentate inhibition. Among them, several JNK inhibitors from PregLem SA, Celegene SA, and Xigen SA have accomplished the early stage of clinical trials, and their results will guide the development and indications of future JNK inhibitors.

NF-κB p105-mediated nuclear translocation of ERK is required for full activation of IFNγ-induced iNOS expression.

Inducible nitric oxidase (iNOS) encoded by Nos2 is a representative IFNγ-inducible effector molecule that plays an important role in both innate and adaptive immunity. In the present study, we demonstrated that full-length NF-κB p105 (p105), which is a precursor of NF-κB p50 (p50), is required for full activation of IFNγ-induced iNOS expression in the RAW264.7 mouse macrophage cell line. In comparison to wild-type (WT) RAW264.7 cells, p105 KO RAW264.7 (p105 KO) cells completely lost IFNγ-induced iNOS expression. Despite the limited effect of exogenous expression of p50 in p105 KO cells on IFNγ-induced Nos2 promoter activity, p105 expression fully restored IFNγ-induced Nos2 promoter activity to a level comparable to that of WT cells, suggesting an important role for full-length p105 in IFNγ-induced iNOS expression. While the expression and phosphorylation of JAK1 and STAT1 were rather enhanced in p105 KO cells, the phosphorylation of c-Jun downstream of MAPK signaling was decreased. IFNγ-induced phosphorylation of ERK, a kinase for IFNγ-induced c-Jun phosphorylation, was not significantly reduced in p105 KO cells, although the nuclear activity of ERK was significantly decreased due to its reduced translocation to the nucleus. Expression of iNOS, nuclear translocation of ERK, and phosphorylation of c-Jun were restored by stable supplementation of p105 in p105 KO cells. These results suggest that p105 is required for the nuclear translocation of ERK and the subsequent phosphorylation of c-Jun, which are necessary for full activation of IFNγ-induced iNOS expression. Reduced nuclear translocation of ERK in p105 KO cells was also observed in the activation of ERK following serum starvation, further suggesting that the involvement of p105 in ERK nuclear translocation is not limited to IFNγ-stimulated cells but is a more general function of p105.

Galangin alleviated Doxorubicin-induced cardiotoxicity by inhibiting ferroptosis through GSTP1/JNK pathway.

BACKGROUND: Doxorubicin (DOX) is a potent anticancer medication, but its significant cardiotoxicity poses a challenge in clinical practice. Galangin (Gal), a flavonoid compound with diverse pharmacological activities, has shown potential in exerting cardioprotective effects. However, the related molecular mechanism has not been fully elucidated. PURPOSE: Combined with bioinformatics and experimental verification methods to investigate Gal's potential role and underlying mechanisms in mitigating DOX-induced cardiotoxicity (DIC). METHODS: C57BL/6 mice received a single dose of DOX via intraperitoneal injection 4 days before the end of the gavage period with Gal. Myocardial injury was evaluated using echocardiography, myocardial injury biomarkers, Sirius Red and H&E staining. H9c2 cells were stimulated with DOX to mimic DIC in vitro. The potential therapeutic target of Gal was identified through network pharmacology, molecular docking and cellular thermal shift assay (CETSA), complemented by an in-depth exploration of the GSTP1/JNK signaling pathway using immunofluorescence. Subsequently, the GSTP1 inhibitor Ezatiostat (Eza) substantiated the signaling pathway. RESULTS: Gal administration considerably raised DOX-inhibited the left ventricular ejection fractions (LVEF), reduced levels of myocardial injury markers (c-TnI, c-TnT, CKMB, LDH, and AST), and alleviated DOX-induced myocardial histopathological injury and fibrosis in mice, thereby improving cardiac dysfunction. The ferroptosis induced by DOX was inhibited by Gal treatment. Gal remarkably ameliorated the DOX-induced lipid peroxidation, accumulation of iron and Ptgs2 expression both in H9c2 cells and cardiac tissue. Furthermore, Gal effectively rescued the DOX-inhibited crucial regulators of ferroptosis such as Gpx4, Nrf2, Fpn, and Slc7a11. The mechanistic investigations revealed that Glutathione S-transferase P1 (GSTP1) may be a potential target for Gal in attenuating DIC. Gal act on GSTP1 by stimulating its expression, thereby enhancing the interaction between GSTP1 and c-Jun N-terminal kinase (JNK), leading to the deactivation of JNK/c-Jun pathway. Furthermore, interference of GSTP1 with inhibitor Eza abrogated the cardioprotective and anti-ferroptotic effects of Gal, as evidenced by decreased cell viability, reduced expression of GSTP1 and Gpx4, elevated MDA levels, and promoted phosphorylation of JNK and c-Jun compared with Gal treatment. CONCLUSION: Gal could inhibit ferroptosis and protect against DIC through regulating the GSTP1/JNK pathway. Our research has identified a novel pathway through which Gal regulates DIC, providing valuable insights into the potential therapeutic efficacy of Gal in mitigating cardiotoxic effects.

RG108 attenuates acute kidney injury by inhibiting P38 MAPK/FOS and JNK/JUN pathways.

Acute kidney injury (AKI) is an important clinical syndrome characterised by a sudden decline in renal function, often accompanied by renal inflammation and tubular epithelial cell damage. It has been reported that inhibiting DNA methylation significantly suppress the progression of AKI. In the current study, we investigate the effect of the DNA methyltransferase (DNMT) inhibitor RG108 in cisplatin- and hypoxia-reoxygenation-induced AKI. The expression of kidney injury molecules and inflammatory factors was examined by immunofluorescence, Western blotting and Real-time PCR. The results demonstrated that RG108 treatment significantly reduced kidney inflammation and injury. Furthermore, RNA-seq analysis was performed to reveal the regulatory mechanism of RG108 in AKI. The expression of the FOS and JUN genes, which are downstream of the MAPK pathway, were significant increased in AKI. Meanwhile, the expression of FOS and JUN were both inhibited by RG108, which is similar to what we found treatment with a specific JNK inhibitor and a specific p38 MAPK inhibitor, and thus attenuated renal inflammation and injury. In conclusion, we suggest that RG108 inhibits P38 MAPK/FOS and JNK/JUN pathways and attenuates renal injury and inflammatory responses. In these results, RG108 may become a novel MAPK pathway inhibitor and a clinical candidate for the treatment of AKI.

Ailanthone ameliorates pulmonary fibrosis by suppressing JUN-dependent MEOX1 activation.

Pulmonary fibrosis poses a significant health threat with very limited therapeutic options available. In this study, we reported the enhanced expression of mesenchymal homobox 1 (MEOX1) in pulmonary fibrosis patients, especially in their fibroblasts and endothelial cells, and confirmed MEOX1 as a central orchestrator in the activation of profibrotic genes. By high-throughput screening, we identified Ailanthone (AIL) from a natural compound library as the first small molecule capable of directly targeting and suppressing MEOX1. AIL demonstrated the ability to inhibit both the activation of fibroblasts and endothelial-to-mesenchymal transition of endothelial cells when challenged by transforming growth factor-ß1 (TGF-ß1). In an animal model of bleomycin-induced pulmonary fibrosis, AIL effectively mitigated the fibrotic process and restored respiratory functions. Mechanistically, AIL acted as a suppressor of MEOX1 by disrupting the interaction between the transcription factor JUN and the promoter of MEOX1, thereby inhibiting MEOX1 expression and activity. In summary, our findings pinpointed MEOX1 as a cell-specific and clinically translatable target in fibrosis. Moreover, we demonstrated the potent anti-fibrotic effect of AIL in pulmonary fibrosis, specifically through the suppression of JUN-dependent MEOX1 activation.

Integrating network analysis and experimental validation to reveal the mechanism of si-jun-zi decoction in the treatment of renal fibrosis.

Treating kidney diseases from the perspective of spleen is an important clinical method in traditional Chinese medicine (TCM) for anti-renal fibrosis (RF). Si-jun-zi decoction (SJZD), a classic formula for qi-invigorating and spleen-invigorating, has been reported to alleviate RF. This study aims to investigate the potential mechanism by which SJZD attenuates RF. The results demonstrated notable improvements in renal function levels, inflammation and fibrosis indices in UUO-mice following SJZD intervention. The main active ingredients identified were Quercetin, Kaempferol, Naringenin and 7-Methoxy-2-methyl isoflavone. Furthermore, STAT3, MAPK3, MYC were confirmed as key targets. Additionally, GO enrichment analysis demonstrated that SJZD delayed RF primarily by regulating oxidative stress and other biological mechanisms. KEGG enrichment analysis revealed the involvement of pathways such as Lipid and atherosclerosis signaling pathway, MAPK signaling pathway and other pathways in the reno-protective effects of SJZD. The molecular docking results revealed that the active ingredients of SJZD were well-bound and stable to the core targets. The experiments results revealed that Quercetin, Kaempferol, and Naringenin not only improved the morphology of TGF-ß-induced HK-2 cells but also reversed the expression of α-SMA, COL1A1 and MAPK, thereby delaying the progression of RF. The anti-RF effects of SJZD were exerted through multi-components, multi-targets and multi-pathways.

Dysregulation of c-Jun (JUN) and FBJ murine osteosarcoma viral oncogene homolog B (FOSB) in obese people and their predictive values for metabolic syndrome.

The incidences of metabolic syndrome (MetS), denoting insulin resistance-associated various metabolic disorders, are increasing. This study aimed to identify new biomarkers for predicting MetS and provide a novel diagnostic approach. Herein, the expression profiles of c-Jun (JUN) and FBJ murine osteosarcoma viral oncogene homolog B (FOSB) in individuals with obesity and patients with MetS from the Gene Expression Omnibus database. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) was used to evaluate the messenger RNA levels of JUN and FOSB in the peripheral blood of healthy volunteers (lean and obese) and patients with MetS (lean and obese), along with that in the adipose tissue and peripheral blood of obese mouse model. Furthermore, receiver operating characteristic (ROC) curve and logistic regression analyses were performed to determine the diagnostic value of JUN and FOSB in MetS. The expression profiles and RT-qPCR results showed that JUN and FOSB were highly expressed in individuals with obesity, obese mouse models, and patients with MetS. The ROC analysis results showed an area under the curve values of 0.872 and 0.879 for JUN, 0.802 and 0.962 for FOSB, and 0.946 and 0.979 for JUN-FOSB in the lean group and the group with obesity, respectively, in predicting MetS. Logistic regression analysis showed that the p-values of both JUN and FOSB as MetS-affecting factors were <0.05. Altogether, the findings of this study indicate that both JUN and FOSB, abnormally expressed in individuals with obesity, are good biomarkers of MetS.

CRKL Enhances YAP Signaling through Binding and JNK/JUN Pathway Activation in Liver Cancer.

The Hippo pathway transducers yes-associated protein (YAP) and WW-domain containing transcription regulator 1 (WWTR1/TAZ) are key regulators of liver tumorigenesis, promoting tumor formation and progression. Although the first inhibitors are in clinical trials, targeting the relevant upstream regulators of YAP/TAZ activity could prove equally beneficial. To identify regulators of YAP/TAZ activity in hepatocarcinoma (HCC) cells, we carried out a proximity labelling approach (BioID) coupled with mass spectrometry. We verified CRK-like proto-oncogene adaptor protein (CRKL) as a new YAP-exclusive interaction partner. CRKL is highly expressed in HCC patients, and its expression is associated with YAP activity as well as poor survival prognosis. In vitro experiments demonstrated CRKL-dependent cell survival and the loss of YAP binding induced through actin disruption. Moreover, we delineated the activation of the JNK/JUN pathway by CRKL, which promoted YAP transcription. Our data illustrate that CRKL not only promoted YAP activity through its binding but also through the induction of YAP transcription by JNK/JUN activation. This emphasizes the potential use of targeting the JNK/JUN pathway to suppress YAP expression in HCC patients.

Hsp90aa1/JUN/Ccl2 regulatory axis mediates migration and differentiation of NSPCs, promoting the onset and progression of early post-ischemic stroke epilepsy.

Early-onset epilepsy following ischemic stroke is a severe neurological condition, the pathogenesis of which remains incompletely understood. Recent studies suggest that Neural stem/progenitor cells (NSPCs) play a crucial role in the disease process, yet the precise molecular mechanisms regulating NSPCs have not been thoroughly investigated. This study utilized single-cell transcriptome sequencing and bioinformatics analysis to identify disease-related genes, which were subsequently validated in both in vitro and in vivo experiments. The findings revealed that Hsp90aa1 (heat shock protein 90 kDa alpha, class A member 1), Jun proto-oncogene (JUN), and CC Motif Ligation 2 (Ccl2) constitute an important regulatory axis influencing the migration and differentiation of NSPCs, potentially impacting the onset and progression of early-onset epilepsy post-ischemic stroke. Additionally, the expression of Hsp90aa1 was found to influence the likelihood of seizure occurrence and the severity of brain ischemia.

Loss of prohibitin 2 in Schwann cells dysregulates key transcription factors controlling developmental myelination.

Schwann cells are critical for the proper development and function of the peripheral nervous system (PNS), where they form a collaborative relationship with axons. Past studies highlighted that a pair of proteins called the prohibitins play major roles in Schwann cell biology. Prohibitins are ubiquitously expressed and versatile proteins. We have previously shown that while prohibitins play a crucial role in Schwann cell mitochondria for long-term myelin maintenance and axon health, they may also be present at the Schwann cell-axon interface during development. Here, we expand on this, showing that drug-mediated modulation of prohibitins in vitro disrupts myelination and confirming that Schwann cell-specific ablation of prohibitin 2 (Phb2) in vivo results in severe defects in radial sorting and myelination. We show in vivo that Phb2-null Schwann cells cannot effectively proliferate and the transcription factors EGR2 (KROX20), POU3F1 (OCT6), and POU3F2 (BRN2), necessary for proper Schwann cell maturation, are dysregulated. Schwann cell-specific deletion of Jun, a transcription factor associated with negative regulation of myelination, confers partial rescue of the developmental defect seen in mice lacking Schwann cell Phb2. Finally, we identify a pool of candidate PHB2 interactors that change their interaction with PHB2 depending on neuronal signals, and thus are potential mediators of PHB2-associated developmental defects. This work develops our understanding of Schwann cell biology, revealing that Phb2 may modulate the timely expression of transcription factors necessary for proper PNS development, and proposing candidates that may play a role in PHB2-mediated integration of axon signals in the Schwann cell.

Exploring the globoid cell leukodystrophy protein network and therapeutic interventions.

Globoid cell leukodystrophy is a severe rare disorder characterized by white matter degradation, resulting in a progressive loss of physical and mental abilities and has extremely limited therapeutic interventions. Therefore, this study aimed to delve into the Globoid cell leukodystrophy associated intricate network of differentially expressed genes (p < 0.05, |Fc|> 1) to identify potential druggable targets and possible therapeutic interventions using small molecules. The disease-associated neuronal protein circuit was constructed and analyzed, identifying 53 nodes (minimum edge cutoff 1), among which five (FOS, FOSB, GDNF, GFRA1, and JUN) were discerned as potential core protein nodes. Although our research enumerates the potential small molecules to target various protein nodes in the proposed disease network, we particularly underscore T-5224 to inhibit c-Jun activity as JUN was identified as one of the pivotal elements within the disease-associated neuronal protein circuit. The evaluation of T-5224 binding energy (- 11.0 kcal/mol) from docking study revealed that the compound to exhibit a notable affinity towards Jun/CRE complex. Moreover, the structural integrity of complex was affirmed through comprehensive molecular dynamics simulations, indicating a stable hydrophilic interaction between T-5224 and the Jun/CRE complex, thereby enhancing protein compactness and reducing solvent accessibility. This binding energy was further substantiated by free binding analysis, revealing a substantial thermodynamics complex state (- 448.00 ± 41.73 kJ/mol). Given that this investigation is confined to a computational framework, we additionally propose a hypothetical framework to ascertain the feasibility of inhibiting the Jun/CRE complex with T-5224 against Globoid cell leukodystrophy, employing a combination of in vitro and in vivo methodologies as a prospective avenue of this study.

Ferroptosis-Related Genes in IgA Nephropathy: Screening for Potential Targets of the Mechanism.

Aims: Exploring key genes and potential molecular pathways of ferroptosis in immunoglobulin A nephropathy (IgAN). Methods: The IgAN datasets and ferroptosis-related genes (FRGs) were obtained in the Gene Expression Omnibus (GEO) and FerrDb database. Differentially expressed genes (DEGs) were identified using R software and intersected with FRGs to obtain differentially expressed FRGs (DE-FRGs). After that, the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis (PEA) and Gene Ontology (GO) functional annotation were performed on DE-FRGs. In the Search Tool for the Retrieval of Interacting Genes (STRING) website, we construct a protein-protein interaction (PPI) network. The PPI network was further investigated with screening hub genes with Cytoscape software. The core genes were then subjected to gene set enrichment analysis (GSEA). Finally, the samples were analyzed for immune infiltration in R, and the correlation between hub genes and immune cells was analyzed. Results: A total of 347 DEGs were identified. CD44, CDO1, CYBB, IL1B, RRM2, AKR1C1, activated transcription factor-3 (ATF3), CDKN1A, GDF15, JUN, MGST1, MIOX, MT1G, NR4A1, PDK4, TNFAIP3, and ZFP36 were determined as DE-FRGs. JUN, IL1B, and ATF3 were then screened as hub genes. GSEA and immune infiltration analysis revealed that the hub genes were closely associated with immune inflammatory responses such as NOD-like receptor signaling, IL-17 signaling, and TNF signaling. Conclusions: Our results show that JUN and ATF3 are possibly critical genes in the process of IgAN ferroptosis and may be related with immune cell infiltration.

Down-Regulated JDP2 Attenuated Trophoblast Invasion and Migration in Preeclampsia by Inhibiting Epithelial-Mesenchymal Transition through the Wnt/ß-Catenin Pathway.

INTRODUCTION: Preeclampsia (PE) is an immensely prevalent condition that poses a significant risk to both maternal and fetal health. It is recognized as a primary cause of perinatal morbidity and mortality. Despite extensive research efforts, the precise impact of JDP2 on trophoblast invasion and migration in the context of preeclampsia remains unclear. MATERIALS AND METHODS: The present study aimed to investigate the differential expression of JDP2 between normal control and preeclampsia placentas through the use of quantitative polymerase chain reaction (qPCR), western blotting, and immunostaining techniques. Furthermore, the effects of JDP2 overexpression and silencing on the migration, invasion, and wound healing capabilities of HTR-8/SVneo cells were evaluated. In addition, this study also examined the impact of JDP2 on epithelial-mesenchymal transition (EMT)-associated biomarkers and the Wnt/ß-catenin pathway. RESULTS: In the present investigation, it was ascertained that Jun dimerization protein 2 (JDP2) exhibited a substantial decrease in expression levels in placentae afflicted with preeclampsia in comparison to those of normal placentae. Impairment in migration and invasion was noted upon JDP2 down-regulation, whereas augmentation of migration and invasion was observed upon JDP2 overexpression in HTR-8/SVneo cells. Subsequently, western blot and immunofluorescence assays were conducted, revealing marked alterations in EMT-associated biomarkers, such as E-cadherin, N-cadherin, and ß-catenin, thereby indicating that JDP2 can facilitate cell invasion by modulating the EMT process in HTR-8/SVneo cells. Finally, activation of Wnt/ß-catenin signaling was observed as a result of JDP2. After that, IWR-1, a Wnt inhibitor, was used in the recovery study. IWR-1 could inhibit the role of JDP2 in promoting migration and invasion in HTR-8/SVneo cells. CONCLUSION: Our findings elucidated the impact of JDP2 on trophoblast invasion and migration in preeclampsia by suppressing the EMT through the Wnt/ß-catenin signaling pathway, thereby offering a potential prognostic and therapeutic biomarker for this condition.

Recent Molecular Targets and Their Ligands for the Treatment of Alzheimer Disease.

Alzheimer's disease is a multifaceted neurodegenerative disease. Cholinergic dysfunction, amyloid ß toxicity, tauopathies, oxidative stress, neuroinflammation are among the main pathologies of the disease. Ligands targeting more than one pathology, multi-target directed ligands, attract attention in the recent years to tackle Alzheimer's disease. In this review, we aimed to cover different biochemical pathways, that are revealed in recent years for the pathology of the disease, as druggable targets such as cannabinoid receptors, matrix metalloproteinases, histone deacetylase and various kinases including, glycogen synthase kinase-3, mitogen-activated protein kinase and c-Jun N-terminal kinase, and their ligands for the treatment of Alzheimer's disease in the hope of providing more realistic insights into the field.

Bergenin protects against osteoarthritis by inhibiting STAT3, NF-κB and Jun pathways and suppressing osteoclastogenesis.

Osteoarthritis (OA) is a chronic degenerative disease characterized by articular cartilage destruction and subchondral bone reconstruction in the early stages. Bergenin (Ber) is a cytoprotective polyphenol found in many medicinal plants. It has been proven to have anti-inflammatory, antioxidant, and other biological activities, which may reveal its potential role in the treatment of OA. This study aimed to determine the potential efficacy of Ber in treating OA and explore the possible underlying mechanism through network pharmacology and validation experiments. The potential co-targets and processes of Ber and OA were predicted by using network pharmacology, including a Venn diagram for intersection targets, a protein‒protein interaction (PPI) network to obtain key potential targets, and GO and KEGG pathway enrichment to reveal the probable mechanism of action of Ber on OA. Subsequently, validation experiments were carried out to investigate the effects and mechanisms of Ber in treating OA in vitro and vivo. Ber suppressed IL-1ß-induced chondrocyte apoptosis and extracellular matrix catabolism by inhibiting the STAT3, NF-κB and Jun signalling pathway in vitro. Furthermore, Ber suppressed the expression of osteoclast marker genes and RANKL-induced osteoclastogenesis. Ber alleviated the progression of OA in DMM-induced OA mice model. These results demonstrated the protective efficacy and potential mechanisms of Ber against OA, which suggested that Ber could be adopted as a potential therapeutic agent for treating OA.