JNK Cascade Research Articles

Elucidation of significant regulatory proteins in the JNK pathway modulated by truncated pardaxin, PC6 through molecular dynamic simulation

ABSTRACT Cancer remains a critical global health issue, with current treatments often causing significant side effects. This has spurred the search for alternative therapies, such as bioactive peptides, which offer advantages such as higher selectivity against cancer cells and ease of modification. Pardaxin, a bioactive peptide, has shown anticancer effects on various cell lines via the JNK pathway. A truncated form of pardaxin, PC6, has demonstrated anticancer properties with reduced haemolytic effects though its cell death mechanism is unclear. This study aimed to investigate PC6-induced cell death, focusing on the JNK cascade compared to parental pardaxin (PAR). JNK cascade genes were sourced from the Gene Ontology database, followed by docking screening and protein-peptide network analysis. The network identified 65 genes involved in the JNK pathway, which were verified through gene enrichment analysis. Molecular dynamic simulation and binding affinity analysis showed that PC6 had a higher affinity (−88.69, −82.17, −54.93 kcal/mol) for MAPK8, MAPK9 and HRAS proteins than PAR (−68.88, −76.64, −38.63 kcal/mol). These findings suggest that PC6 specifically interacts with these proteins to induce apoptosis via the JNK cascade, highlighting its potential as an effective anticancer agent.

Abstract 2187: Influences of high-fat diet on tumor biology in a mouse model of colorectal cancer

Abstract Purpose: To test the influence of high-fat diet on tumor biology in the inducible Kras, Apc and p53 (iKAP) model (Boutin, et al Genes & Development 2017, 31:370). Methods: At weening, both male and female mice were fed either a high-fat (60%) or low-fat (10%) diet for 5-6 weeks prior to tumor induction. Mutant Kras expression was controlled using a tet-on system. Expression of mutant Kras was activated by the addition of doxycycline hyclate in the drinking water. Tumor growth was monitored by bi-weekly endoscopy and calculated as percent luminal occlusion. Tumor biology was assessed with proteomics and phospho-proteomics with comparisons by sex and diet. Results: In the absence of mutant Kras expression (no doxycycline in drinking water), tumors were larger in females on high-fat diet compared to low-fat diet. Tumor sizes were not significantly different in males or in the presence of mutant Kras expression. Proteomics analyses of tumor tissue identified significantly different patterns by diet in female mice, including altered levels of KIF4, DAB2iP, JNK cascade and JUN kinase activity (Table). Conclusion: Based on our preliminary data, high-fat diet may alter tumor kinase activities, even in a genetically engineered mouse model. Replication and validation studies are ongoing. Table of significantly upregulated biologic processes in female mice on high-fat diet compared to lo XXXXXXXXXXXXXXXX XXXXXXXXXXXXXXX Citation Format: Anindita Mahanty, Cara Wallingford, Revan Hammontree, Muturi Njoka, Zachary Clark, Michaella Rekowski, Michael Washburn, Jennifer S. Davis. Influences of high-fat diet on tumor biology in a mouse model of colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2187.

Mutant α-synuclein causes death of human cortical neurons via ERK1/2 and JNK activation

Synucleinopathies refer to a group of disorders characterized by SNCA/α-synuclein (α-Syn)-containing cytoplasmic inclusions and neuronal cell loss in the nervous system including the cortex, a common feature being cognitive impairment. Still, the molecular pathogenesis of cognitive decline remains poorly understood, hampering the development of effective treatments. Here, we generated induced pluripotent stem cells (iPSCs) derived from familial Parkinson’s disease (PD) patients carrying SNCA A53T mutation, differentiating them into cortical neurons by a direct conversion method. Patient iPSCs-derived cortical neurons harboring mutant α-Syn exhibited increased α-Syn-positive aggregates, shorter neurites, and time-dependent vulnerability. Furthermore, RNA-sequencing analysis, followed by biochemical validation, identified the activation of the ERK1/2 and JNK cascades in cortical neurons with SNCA A53T mutation. This result was consistent with a reverted phenotype of neuronal death in cortical neurons when treated with ERK1/2 and JNK inhibitors, respectively. Our findings emphasize the role of ERK1/2 and JNK cascades in the vulnerability of cortical neurons in synucleinopathies, and they could pave the way toward therapeutic advancements for synucleinopathies.

JNK Cascade-Induced Apoptosis-A Unique Role in GqPCR Signaling.

The response of cells to extracellular signals is mediated by a variety of intracellular signaling pathways that determine stimulus-dependent cell fates. One such pathway is the cJun-N-terminal Kinase (JNK) cascade, which is mainly involved in stress-related processes. The cascade transmits its signals via a sequential activation of protein kinases, organized into three to five tiers. Proper regulation is essential for securing a proper cell fate after stimulation, and the mechanisms that regulate this cascade may involve the following: (1) Activatory or inhibitory phosphorylations, which induce or abolish signal transmission. (2) Regulatory dephosphorylation by various phosphatases. (3) Scaffold proteins that bring distinct components of the cascade in close proximity to each other. (4) Dynamic change of subcellular localization of the cascade's components. (5) Degradation of some of the components. In this review, we cover these regulatory mechanisms and emphasize the mechanism by which the JNK cascade transmits apoptotic signals. We also describe the newly discovered PP2A switch, which is an important mechanism for JNK activation that induces apoptosis downstream of the Gq protein coupled receptors. Since the JNK cascade is involved in many cellular processes that determine cell fate, addressing its regulatory mechanisms might reveal new ways to treat JNK-dependent pathologies.

Glucagon-like Peptide-1 Acts as Signaling Mediator to Modulate Human Sperm Performance via Targeting Akt, JNK and IRS-1 Cell Signaling Cascades: Novel Insights into Sperm Physiopathology.

Recent evidence suggests that the male gonad is a potential target of glucagon-like peptide-1 (GLP-1). We investigated the effects of glucagon-like peptide-1 (GLP-1) on sperm function and the molecular mechanisms through which it may act. Semen samples of healthy men were incubated in the presence or absence of a GLP-1 mimetic analog, exendin-4 (Exe). In a different analysis, sperm were exposed to tumor necrosis factor (TNF-α) alone and, in some tubes, TNF-α was added after previous exposure to exendin-4 (Exe). Sperm parameters and protein-kinase B (p-Akt), insulin receptor substrate-1 (p-IRS-1 Ser312), and c Jun N-terminal protein kinase (p-JNK Thr183/Tyr185) were considered and evaluated. Sperm parameters, when incubated for 4 h in a simple defined balanced salt solution lacking protein, declined progressively with incubation time. The maximum decline was associated with a significant decrease in phosphorylated protein kinase B (p-Akt), concomitantly to an increase in insulin receptor substrate-1 (p-IRS-1 Ser312) and c Jun N-terminal protein kinase (p-JNK Thr183/Tyr185). Preincubation with exendin-4 (Exe) prevented this decline and maintained sperm motility (progressive-PM and total-TM). TNF-α exposure resulted in decreased sperm motility (PM and TM) and viability (V) in a concentration-dependent manner. Exe addition attenuated this TNF-α negative effect on sperm parameters. Glucagon-like peptide-1 (GLP-1) also acts by reducing levels of the "negative" kinases p-IRS-1Ser312 and p-JNK. An imbalance involving these three kinases in sperm, as it occurs in somatic cells, is a novel scenario that may participate in sperm physiopathology.

Effect of age on the risk of immune-related adverse events in patients receiving immune checkpoint inhibitors.

Identifying patients at increased risk of immune-related adverse events (irAEs) facilitates safe application of immune checkpoint inhibitors (ICIs). This retrospective study aimed to determine the effect of age on the risk of irAEs in patients receiving ICIs and to identify potential mechanisms underlying age-related irAE risk differences. We analyzed reports of FDA Adverse Event Reporting System from July 1, 2014, to September 30, 2021. The information component ratio (ICΔ) was used to compare the irAE risk between older adults (> 65years) and younger adults (25-65years), of which the 95% confidential interval lower limit (ICΔ025) exceeding zero indicated significantly increased risk. We found that older adults had a significantly higher overall irAE risk than younger adults (ICΔ025 0.38), which was observed in almost all organ systems. We further analyzed the correlation between age-related irAE risks and age-related transcriptional changes to identify potential genes and pathways underlying age-related irAE risk differences. We found that genes significantly correlated with ICΔ were enriched in processes including extracellular matrix organization, regulation of myeloid leukocyte mediated immunity, and regulation of c-Jun N-terminal kinase (JNK) cascade. In addition, single-cell RNA sequencing analysis confirmed that genes involved in collagen-containing extracellular matrix and JNK cascade were significantly upregulated in myeloid cells from ICI-associated colitis tissues compared with ICI-treated colon tissues without colitis. In conclusion, older adults receiving ICIs have higher irAE risks than younger adults. Upregulation of genes involved in JNK cascade and collagen-containing extracellular matrix in myeloid cells may contribute to increased irAE risks in older adults.

Abstract 4015: EGFR activation via c-Jun axis promotes adaptive resistance to third generation ALK-TKI lorlatinib in ALK-rearranged non-small cell lung cancer cells

Abstract Anaplastic lymphoma kinase-tyrosine kinase inhibitors (ALK-TKIs), including lorlatinib have shown dramatic efficacy in patients with ALK-rearranged lung cancer; however, complete response in these patients is rare, as a small population of tumor cells survives due to adaptive resistance. In this preclinical study, we investigated the adaptive resistant mechanisms to lorlatinib and its therapeutic strategies to overcome, using in vitro and in vivo experiments. We have identified EGFR as an adaptive resistance mechanism of lorlatinib using phospho-RTK array and siRNA library. Cell line-based assay showed that exposure of lorlatinib accelerated EGFR signal activation as an adaptive response via activation of JNK cascade in ALK-rearranged lung cancer cells. Knockdown for EGFR suppressed the survival signal of ALK lung cancer cells when treated with lorlatinib. The combination of lorlatinib and erlotinib, a first-generation EGFR-TKI, was shown to enhance cell growth inhibition by inducing cell apoptosis via suppressing the anti-apoptotic factor Bcl-xl compared with lorlatinib monotherapy. Moreover, cell line-derived xenograft model indicated that the combination of lorlatinib plus erlotinib enhanced anti-tumor effects compared with lorlatinib monotherapy. Taken together, our results displayed that EGFR activation plays an important role in the initial adaptive resistance to lorlatinib for ALK-rearranged lung cancer. The combination of lorlatinib and erlotinib is expected as the promising strategy for those tumors. Citation Format: Yuki Katayama, Tadaaki Yamada, Kenji Morimoto, Hiroaki Ozasa, Shinsaku Tokuda, Koichi Takayama. EGFR activation via c-Jun axis promotes adaptive resistance to third generation ALK-TKI lorlatinib in ALK-rearranged non-small cell lung cancer cells. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4015.

JNK Activation Correlates with Cognitive Impairment and Alteration of the Post-Synaptic Element in the 5xFAD AD Mouse Model

The c-Jun N-terminal kinases (JNKs) are a family of proteins that, once activated by stress stimuli, can alter neuronal functions and survival. The JNK cascade plays a crucial role in the post-synaptic neuronal compartment by altering its structural organization and leading, at worst, to an overall impairment of neuronal communication. Increasing evidence suggests that synaptic impairment is the first neurodegenerative event in Alzheimer's disease (AD). To better elucidate this mechanism, we longitudinally studied 5xFAD mice at three selected time points representative of human AD symptom progression. We tested the mice cognitive performance by using the radial arm water maze (RAWM) in parallel with biochemical evaluations of post-synaptic enriched protein fraction and total cortical parenchyma. We found that 5xFAD mice presented a strong JNK activation at 3.5 months of age in the post-synaptic enriched protein fraction. This JNK activation correlates with a structural alteration of the post-synaptic density area and with memory impairment at this early stage of the disease that progressively declines to cause cell death. These findings pave the way for future studies on JNK as a key player in early neurodegeneration and as an important therapeutic target for the development of new compounds able to tackle synaptic impairment in the early phase of AD pathology.

IFT20 Confers Paclitaxel Resistance by Triggering β-arrestin-1 to Modulate ASK1 Signaling in Breast Cancer.

System paclitaxel-based chemotherapy is the first-line treatment regimen of defense against breast cancer, but inherent or acquired chemotherapy resistance remains a major obstacle in breast cancer therapy. Elucidating the molecular mechanism of chemoresistance is essential to improve the outcome of patients with breast cancer. Here, we demonstrate that intraflagellar transport 20 (IFT20) is positively associated with shorter relapse-free survival in patients with system paclitaxel-based chemotherapy. High-expressed IFT20 in breast cancer cells increases resistance to cell death upon paclitaxel treatment; in contrast, IFT20 knockdown enhances apoptosis in breast cancer cells in response to paclitaxel. Mechanistically, IFT20 triggers β-arrestin-1 to bind with apoptosis signal-regulating kinase 1 (ASK1) and promotes the ubiquitination of ASK1 degradation, leading to attenuating ASK1 signaling and its downstream JNK cascades, which helps cells to escape from cell death during paclitaxel treatment. Our results reveal that IFT20 drives paclitaxel resistance through modulating ASK1 signaling and identifies IFT20 as a potential molecular biomarker for predicting the response to paclitaxel therapeutic in breast cancer. IFT20 drives paclitaxel resistance through modulating ASK1 signaling and IFT20 may act as a potential molecular biomarker for predicting the response to paclitaxel therapeutic in breast cancer.

Whole-transcriptome sequencing revealed differentially expressed mRNAs and non-coding RNAs played crucial roles in NiONPs-induced liver fibrosis

Nickel oxide nanoparticles (NiONPs) induced liver fibrosis, while its mechanisms associated with transcriptome remained unclear. This study aimed to investigate the roles of differentially expressed (DE) messenger RNAs (mRNAs) and non-coding RNAs (ncRNAs) in NiONPs-induced liver fibrosis, and further confirm whether JNK/c-Jun pathway enriched by the DE RNAs was involved in the regulation of the disease. A liver fibrosis rat model was established by intratracheal perfusion of NiONPs twice a week for 9 weeks. Whole-transcriptome sequencing was applied to obtain expression profiles of mRNAs, long non-coding RNAs (lncRNAs), microRNAs (miRNAs), and circular RNAs (circRNAs) in the model rat and control liver tissues. Comparing the RNA expression profiles of the model and control liver tissues, we identified 324 DE mRNAs, 129 DE lncRNAs, 24 DE miRNAs and 33 DE circRNAs, and the potential interactions among them were revealed by constructing two co-expression networks, including lncRNA–miRNA–mRNA and circRNA–miRNA–mRNA networks. Using RT-qPCR, we verified the sequencing results of some RNAs in the networks and obtained similar expression profiles, indicating our sequencing results were reliable and referable. Through Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, we predicted the biological functions and signaling pathways potentially related to NiONPs-induced liver fibrosis, such as “positive regulation of JNK cascade”, “inflammatory response”, “transcription factor binding”, and MAPK, Wnt, PI3K-Akt signaling pathways. JNK/c-Jun pathway, a subclass of MAPK signal, was selected for further investigation because it was significantly enriched by fibrosis-related DE genes and activated in animal models. In vitro, we detected the cytotoxicity of NiONPs on LX-2 cells and treated the cells with 5 μg/ml NiONPs for 12 h. The results showed NiONPs induced the up-regulated protein expression of fibrotic factors collagen-1a1 (Col-1a1) and matrix metalloproteinas2 (MMP2) and JNK/c-Jun pathway activation. While these effects were reversed after JNK/c-Jun pathway was blocked by SP600125 (JNK pathway inhibitor), indicating the pathway was involved in NiONPs-induced excessive collagen formation. In conclusion, our results revealed the DE mRNAs and ncRNAs played crucial roles in NiONPs-induced liver fibrosis, and JNK/c-Jun pathway mediated the development of the disease.

Integrative analyses of immune-related biomarkers and associated mechanisms in coronary heart disease

Various studies showed that the effect of immune activation is pro-atherogenic and coronary heart disease (CHD) should therefore be considered an autoimmune disease. This study aimed to identify potential immune-related biomarkers, pathways, and the potential regulatory networks underlying CHD. Differentially expressed genes (DEGs) between CHD and control samples were determined by analyzing GSE71226 and GSE9128. The overlapping differential expression immune-related genes (DE-IRGs) for CHD were identified by analyzing the ImmPort database and two GEO databases. A total of 384 DE-IRGs were identified. Subsequently, comprehensive enrichment analyses suggested that DE-IRGs were enriched in immune-related pathways, including autoimmune thyroid disease, the intestinal immune network for IGA production, and downstream signaling events of B cell receptors. The signature of DE-IRGs was validated using an external independent dataset GSE20681 (AUC = 0.875). Furthermore, we conducted protein–protein interaction network analysis and identified eight hub genes, which were most enriched in regulation of defense response, NF-κB signaling pathway, regulation of JNK cascade, and regulation of cytokine production. Moreover, networks of miRNAs-mRNAs and transcription factors (TFs)-mRNA underlying the integrated data were established, involving eight miRNAs and 76 TF-targeting hub genes. Ultimately, 17 SNPs in miRNA-mediated gene networks were identified. We screened potential immune-related genes in CHD and constructed miRNA-mRNA-TF and SNP-miRNA networks, which not only provide inspired insights into the occurrence and the molecular mechanisms of CHD but also lay a foundation for targeting potential biomarkers using immunotherapy and for understanding the molecular mechanisms of CHD.

Molecular signatures in diabetic foot ulcer by integrated gene expression profiling via bioinformatics analysis

Introduction and Aim: Diabetic foot ulcers (DFUs) are a common and debilitating diabetic consequence leading to lower-limb amputations, long-term disability, and reduced lifespan. There is a lack of clinical diagnosis expertise because of no adequate diagnostic signs for DFU. As a result, the current research aims to find out how differently expressed genes (DEGs) affect the DFU.
 Materials and Methods: Bioinformatics analysis was used to evaluate DEGs using the GSE132187 dataset of the NCBI-GEO database, which contained samples from three hyperglycemic and three normoglycemic macrophage-like cell lines. Following the discovery of DEGs, Gene Ontology (GO) and KEGG pathway enrichment analysis were used to investigate how genes are classified into preset bins based on their functional properties. To discover hub DEGs in DFU, a protein-protein interaction (PPI) network was built and five topological parameters such as degree, stress, Closeness centrality, betweenness centrality, and radiality were evaluated.
 Results: We found 547 DEGs using the GSE132187 dataset, comprising 79 upregulated DEGs and 468 downregulated DEGs. There were 434 nodes and 1724 edges in the PPI network. The giant network uncovered six modules that are significantly enriched in biological processes like positive JNK cascade regulation, positive interferon-gamma production regulation, negative cell proliferation regulation, cellular response to zinc ion, cellular response to lipopolysaccharide, wound healing, and inflammatory response.
 Conclusion: Bioinformatics analysis revealed the major differentially expressed hub-genes implicated in DFUs. These findings suggested that these genes could be used as a DFU prognostic, diagnostic, or therapeutic targets.

Whole exome sequencing of high-risk neuroblastoma identifies novel non-synonymous variants.

Neuroblastoma (NBL), one of the main death-causing cancers in children, is known for its remarkable genetic heterogeneity and varied patient outcome spanning from spontaneous regression to widespread disease. Specific copy number variations and single gene rearrangements have been proven to be associated with biological behavior and prognosis; however, there is still an unmet need to enlarge the existing armamentarium of prognostic and therapeutic targets. We performed whole exome sequencing (WES) of samples from 18 primary tumors and six relapse samples originating from 18 NBL patients. Our cohort consists of 16 high-risk, one intermediate, and one very low risk patient. The obtained results confirmed known mutational hotspots in ALK and revealed other non-synonymous variants of NBL-related genes (TP53, DMD, ROS, LMO3, PRUNE2, ERBB3, and PHOX2B) and of genes cardinal for other cancers (KRAS, PIK3CA, and FLT3). Beyond, GOSeq analysis determined genes involved in biological adhesion, neurological cell-cell adhesion, JNK cascade, and immune response of cell surface signaling pathways. We were able to identify novel coding variants present in more than one patient in nine biologically relevant genes for NBL, including TMEM14B, TTN, FLG, RHBG, SHROOM3, UTRN, HLA-DRB1, OR6C68, and XIRP2. Our results may provide novel information about genes and signaling pathways relevant for the pathogenesis and clinical course in high-risk NBL.

Dock10 Regulates Cardiac Function under Neurohormonal Stress.

Dedicator of cytokinesis 10 (Dock10) is a guanine nucleotide exchange factor for Cdc42 and Rac1 that regulates the JNK (c-Jun N-terminal kinase) and p38 MAPK (mitogen-activated protein kinase) signaling cascades. In this study, we characterized the roles of Dock10 in the myocardium. In vitro: we ablated Dock10 in neonatal mouse floxed Dock10 cardiomyocytes (NMCMs) and cardiofibroblasts (NMCFs) by transduction with an adenovirus expressing Cre-recombinase. In vivo, we studied mice in which the Dock10 gene was constitutively and globally deleted (Dock10 KO) and mice with cardiac myocyte-specific Dock10 KO (Dock10 CKO) at baseline and in response to two weeks of Angiotensin II (Ang II) infusion. In vitro, Dock10 ablation differentially inhibited the α-adrenergic stimulation of p38 and JNK in NMCM and NMCF, respectively. In vivo, the stimulation of both signaling pathways was markedly attenuated in the heart. The Dock10 KO mice had normal body weight and cardiac size. However, echocardiography revealed mildly reduced systolic function, and IonOptix recordings demonstrated reduced contractility and elevated diastolic calcium levels in isolated cardiomyocytes. Remarkably, Dock10 KO, but not Dock10 CKO, exaggerated the pathological response to Ang II infusion. These data suggest that Dock10 regulates cardiac stress-related signaling. Although Dock10 can regulate MAPK signaling in both cardiomyocytes and cardiofibroblasts, the inhibition of pathological cardiac remodeling is not apparently due to the Dock10 signaling in the cardiomyocyte.

Salvianolic acid B inhibits autophagy and activation of hepatic stellate cells induced by TGF-β1 by downregulating the MAPK pathway.

In liver fibrosis, transforming growth factor-β1 (TGF-β1) can stimulate autophagy and activation of hepatic stellate cells (HSCs). Autophagy, playing a crucial role in HSCs activation, is related to liver fibrosis. Increasing evidence have suggested that antifibrosis effects of salvianolic acid B (Sal B) and their mechanisms of action, however, remain unclear. The aim of the article is to understand the role of Sal B in HSCs autophagy in liver fibrosis. Herein, we demonstrated that inducing TGF-β1 led to dramatic increase in autophagosome formation and autophagic flux in JS1 and LX2, which was mediated through the ERK, JNK, and p38 MAPK cascades. TGF-β1 significantly increased the protein of autophagy and liver fibrosis, including LC3BⅡ, ATG5, α-SMA, and Col.I; Sal B inhibits JS1 autophagy and activation by inhibiting the formation of autophagosomes and autophagic flux. Sal B significantly decreased the LC3BⅡ, ATG5, α-SMA, and Col.I protein expressions; pretreatment with autophagy inhibitors, chloroquine (CQ) and 3-methyladenine (3-MA) or silencing ATG7 further increase these reductions. However, pretreatment with autophagy agonist, rapamycin (Rapa), or overexpressed ATG5 attenuated this decrease. To further assess the importance of this mechanism, the antibody chip was used to detect the change of phosphorylation protein expression of the MAPK signaling pathway after treating JS1 with Sal B. Eleven differentially expressed proteins were verified. Sal B inhibits activation and autophagy of JS1 induced by TGF-β1 through downregulating the ERK, p38, and JNK signaling pathways, as demonstrated by downregulating p-ERK, p-JNK, and p-p38 MAPK protein expressions. In conclusion, Sal B inhibits autophagy and activation induced by TGF-β1 of HSCs possibly by downregulating the MAPK pathway.

Epigenetic Regulation of NGF-Mediated Osteogenic Differentiation in Human Dental Mesenchymal Stem Cells.

Nerve growth factor (NGF) is the best-characterized neurotrophin and is primarily recognized for its key role in the embryonic development of the nervous system and neuronal cell survival/differentiation. Recently, unexpected actions of NGF in bone regeneration have emerged as NGF is able to enhance the osteogenic differentiation of mesenchymal stem cells. However, little is known regarding how NGF signaling regulates osteogenic differentiation through epigenetic mechanisms. In this study, using human dental mesenchymal stem cells (DMSCs), we demonstrated that NGF mediates osteogenic differentiation through p75NTR, a low-affinity NGF receptor. P75NTR-mediated NGF signaling activates the JNK cascade and the expression of KDM4B, an activating histone demethylase, by removing repressive H3K9me3 epigenetic marks. Mechanistically, NGF-activated c-Jun binds to the KDM4B promoter region and directly upregulates KDM4B expression. Subsequently, KDM4B directly and epigenetically activates DLX5, a master osteogenic gene, by demethylating H3K9me3 marks. Furthermore, we revealed that KDM4B and c-Jun from the JNK signaling pathway work in concert to regulate NGF-mediated osteogenic differentiation through simultaneous recruitment to the promoter region of DLX5. We identified KDM4B as a key epigenetic regulator during the NGF-mediated osteogenesis both in vitro and in vivo using the calvarial defect regeneration mouse model. In conclusion, our study thoroughly elucidated the molecular and epigenetic mechanisms during NGF-mediated osteogenesis.

Anti-obesity and Anti-diabetic Effect of Ursolic Acid against Streptozotocin/High Fat Induced Obese in Diabetic Rats.

Obesity is occurring due to continue taken high fat diet; this is the fast-growing problem reaching epidemic proportion globally. Ursolic acid altered the abnormal glucose metabolism in diabetic rats. In this experimental protocol, we examine ursolic acid (UA) anti-obesity effect against streptozotocin (STZ) and high-fat diet-induced obesity in rats. Orally administered the ursolic acid (2.5, 5 and 10 mg/kg) dose to the hyperglycemic rats for 8 weeks and estimated the blood glucose level at different time intervals. Biochemical, hepatic, lipid, renal and antioxidant parameters were estimated. Traf-4, Mapk-8, Traf-6 and genes such as Ins-1, ngn-3 and Pdx-1 mRNA expression were estimated using qRT-PCR to scrutinize the molecular mechanism in MAPK downstream JNK cascade and insulin pathway signalling pathways. Ursolic acid significantly (p<0.001) down-regulated the blood glucose level at dose dependent manner. Its also reduced the plasma insulin level, non-essential fatty acid and increased the level of adiponectin as compared to obese control group rats. Ursolic acid treated group rats reduced the level of total cholesterol and triglycerides. Ursolic-acid-treated rats have been shown to decrease oxidative stress in pancreatic tissue by restoring the free radical effect of scavenging, suppress the Traf-6, Mapk-8 and Traf-4 mRNA expression, enhance the expression of Pdx-1, Ins-1 and Ngn-3 and ensure the regeneration of pancreas β cells and therefore pancreas insulin. The current result suggested the anti-obese effect of ursolic acid against high fat diet (HFD) induced obese rats via alteration of insulin and JNK signaling pathway.

Transforming growth factor β3 deficiency promotes defective lipid metabolism and fibrosis in murine kidney.

ABSTRACTGlomerulosclerosis and tubulointerstitial fibrosis are pathological features of chronic kidney disease. Transforming growth factor β (TGFβ) is a key player in the development of fibrosis. However, of the three known TGFβ isoforms, only TGFβ1 has an established role in fibrosis, and the pathophysiological relevance of TGFβ2 and TGFβ3 is unknown. Because Tgfb3 deficiency in mice results in early postnatal lethality, we analyzed the kidney phenotype of heterozygous Tgfb3-knockout mice (Tgfb3+/−) and compared it with that of matched wild-type mice. Four-month-old Tgfb3+/− mice exhibited incipient renal fibrosis with epithelial–mesenchymal transition, in addition to glomerular basement membrane thickening and podocyte foot process effacement associated with albuminuria. Also evident was insulin resistance and oxidative stress at the renal level, together with aberrant renal lipid metabolism and mitochondrial function. Omics analysis revealed toxic species, such as diacylglycerides and ceramides, and dysregulated mitochondrial metabolism in Tgfb3+/− mice. Kidneys of Tgfb3+/− mice showed morphological alterations of mitochondria and overactivation of non-canonical MAPK ERK1/2 and JNK cascades. Our study indicates that renal TGFβ3 might have antifibrotic and renoprotective properties, opposing or counteracting the activity of TGFβ1. This article has an associated First Person interview with the first author of the paper.

Prognostic Significance of ROR2 Expression in Patients with Urothelial Carcinoma.

We investigated the association of receptor tyrosine kinase-like orphan receptor 2 (ROR2) expression with clinicopathological features and oncologic outcomes in large urothelial carcinoma (UC) of the upper tract (UTUC) and urinary bladder (UBUC) cohorts. Through transcriptomic profiling of a published dataset (GSE31684), ROR2 was discovered to be the most upregulated gene during UC progression, focusing on the JNK cascade (GO:0007254). Initially, the evaluation of ROR2 mRNA expression in 50 frozen UBUCs showed significantly upregulated levels in high-stage UC. Moreover, high ROR2 immunoexpression significantly correlated with high tumor stage, high tumor grade, lymph node metastasis, and vascular invasion (all p < 0.05). In multivariate analysis, after adjusting for standard clinicopathological features, ROR2 expression status was an independent prognosticator of cancer-specific survival and metastasis-free survival in UTUC and UBUC (all p < 0.01). In the subgroup analysis, it also significantly predicted bladder tumor recurrence in non-muscle invasive UBUC. Furthermore, the GO enrichment analysis showed that fatty acid, monocarboxylic acid, carboxylic acid metabolic processes, negative regulation of neutrophil migration, and negative regulation of granulocyte and neutrophil chemotaxis were significantly enriched by ROR2 dysregulation. In conclusion, high ROR2 immunoexpression was associated with aggressive pathological characteristics in UC and independently predicted worse prognosis, suggesting it could play roles in clinical risk stratification and therapy decisions.

20-hydroxyecdysone (20E) signaling regulates amnioserosa morphogenesis during Drosophila dorsal closure: EcR modulates gene expression in a complex with the AP-1 subunit, Jun

ABSTRACTSteroid hormones influence diverse biological processes throughout the animal life cycle, including metabolism, stress resistance, reproduction, and lifespan. In insects, the steroid hormone, 20-hydroxyecdysone (20E), is the central hormone regulator of molting and metamorphosis, and plays roles in tissue morphogenesis. For example, amnioserosa contraction, which is a major driving force in Drosophila dorsal closure (DC), is defective in embryos mutant for 20E biosynthesis. Here, we show that 20E signaling modulates the transcription of several DC participants in the amnioserosa and other dorsal tissues during late embryonic development, including zipper, which encodes for non-muscle myosin. Canonical ecdysone signaling typically involves the binding of Ecdysone receptor (EcR) and Ultraspiracle heterodimers to ecdysone-response elements (EcREs) within the promoters of responsive genes to drive expression. During DC, however, we provide evidence that 20E signaling instead acts in parallel to the JNK cascade via a direct interaction between EcR and the AP-1 transcription factor subunit, Jun, which together binds to genomic regions containing AP-1 binding sites but no EcREs to control gene expression. Our work demonstrates a novel mode of action for 20E signaling in Drosophila that likely functions beyond DC, and may provide further insights into mammalian steroid hormone receptor interactions with AP-1.