DUSP1 Expression Research Articles

MKP-1 is required to limit myeloid-cell mediated oral squamous cell carcinoma progression and regional extension

Mitogen-activated protein kinases (MAPKs) require MAPK phosphatases (MKPs) for deactivation of MAPK intracellular signaling. MKP-1 (encoded by Dusp1) is a key negative regulator of MAPKs and prior reports have indicated that MKP-1 regulates oral cancer-associated inflammation and leukocyte infiltration. ObjectiveTo determine the significance of myeloid-based expression of MKP-1 in oral cancer. MethodsThe Cancer Genome Atlas (TCGA) was used to address DUSP1 expression in oral squamous cell carcinoma (OSCC). Syngeneic and carcinogen-induced mouse models using global and myeloid-specific Dusp-1 deficient mice with immunophenotypic, histologic, and transcriptomic analyses and in vitro migration assays. ResultsData from TCGA indicates the DUSP1 expression is inversely related to oral cancer burden and nodal involvement. Using murine models of OSCC, the role of MKP-1 signaling in tumor associated macrophages (TAMs) was assessed. Dusp1-deficient mice had increased tumor burden and TAM infiltrate with increased M2 macrophage polarization. Transcriptomic signatures of TAMs from Dusp1-deficent mice indicated a pro-metastatic phenotype as well as concomitant differences in myeloid-associated genes, cytokine/chemokine signaling, and Notch signaling consistent with tumor progression. In vitro and in vivo assays revealed mouse OSCC cells had a higher migration rate using TAM cell-free supernatant from Dusp1 deficiency mice compared to controls with enhanced regional cervical lymph node metastasis, respectively. To validate TAM studies using implantable mouse models, an OSCC progression model with conditional myeloid-specific Dusp-1 deficient mice demonstrated enhanced OSCC disease progression, characterized by advanced onset, histological stage, and tumor burden. ConclusionMyeloid-based Dusp1-deficiency increases OSCC burden and metastasis through alteration in TAM recruitment, gene profile, and polarity suggesting that MKP-1 could be a viable target to reprogram TAM to limit local/regional OSCC extension.

Initial results from a dose finding study of TNO155, a SHP2 inhibitor, in adults with advanced solid tumors.

3005 Background: SHP2 transduces signals from activated receptor tyrosine kinases to downstream pathways including MAPK. TNO155 is a selective, allosteric, oral inhibitor of SHP2. Methods: CTNO155X2101 (NCT03114319) is an ongoing first-in-human, open-label dose escalation/expansion trial of TNO155 in adults with advanced solid tumors. The primary objective is to characterize the safety and tolerability of TNO155 and identify regimen(s) for future study. Secondary assessments included pharmacokinetics, pharmacodynamics, and preliminary clinical efficacy. Here we present data from TNO155 single agent escalation. Results: As of 10/26/2020, 118 patients received TNO155 in variable schedules: once (QD; 1.5–70 mg; n = 55) or twice daily (BID; 30–50 mg; n = 25) in a 2 weeks on/1 week off (2w/1w) cycle; or QD in a 3w/1w cycle (30–60 mg; n = 32), or continuously (40 or 50 mg QD; n = 6). The most common cancer diagnoses treated were colorectal (54%), gastrointestinal stromal tumor (16%), non-small cell lung (12%), and head & neck (8%). The median number of prior antineoplastic therapies was 4 (range 1–10). Overall 109 patients (92%) have discontinued study treatment, 94 (80%) for progressive disease and 6 (5%) for adverse events (AEs). TNO155 showed rapid absorption (median day 1 Tmax ̃1.1 hours), an effective median T½ of ̃34 hours, and near dose-proportional exposure at day 14 (power model: AUCτ beta = 1.09 [90% CI 1.02–1.16]). AEs were mostly Grade 1/2 and generally consistent with on-target effects of SHP2 inhibition. The most common treatment-related AEs (all grades) were increased blood creatine phosphokinase (n = 33, 28%), peripheral edema (n = 31, 26%), diarrhea (n = 31, 26%), and acneiform dermatitis (n = 27, 23%). The most common treatment-related Grade ≥3 AEs were decreased platelets (n = 5, 4%), increased aspartate aminotransferase, diarrhea, and decreased neutrophils (each n = 4, 3%). The best observed response was stable disease (SD) per RECIST 1.1, reported in 24 (20%) patients, with a median duration of SD of 4.9 months (range 1.7–29.3). Evidence of SHP2 inhibition, as measured by change in DUSP6 expression by qPCR in paired pre- vs. on-treatment tumor samples, was seen in the majority of patients treated with TNO155 doses ≥20 mg/day (≥25% reduction, 38/42 [90%]; ≥50% reduction, 25/42 [60%]). Analysis of tumor whole-transcriptome RNA sequencing data is ongoing. Conclusions: TNO155 shows favorable pharmacokinetic properties and promising early safety and tolerability data at doses with evidence of target inhibition. The optimal dose using several schedules is still under evaluation. Studies of TNO155 in combination with other agents, including nazartinib (mutant-selective EGFR inhibitor[i]), adagrasib (KRAS G12Ci), spartalizumab (anti-PD-1 antibody), ribociclib (CDK4/6i), and dabrafenib (BRAFi) with LTT462 (ERKi), are ongoing (NCT03114319, NCT04330664, NCT04000529, NCT04294160). Clinical trial information: NCT03114319.

Reduction of Matrix Metallopeptidase 13 and Promotion of Chondrogenesis by Zeel T in Primary Human Osteoarthritic Chondrocytes.

Objectives: Zeel T (Ze14) is a multicomponent medicinal product. Initial preclinical data suggested a preventive effect on cartilage degradation. Clinical observational studies demonstrated that Ze14 reduced symptoms of osteoarthritis (OA), including stiffness and pain. This study aimed to explore these effects further to better understand the mode of action of Ze14 on human OA chondrocytes in vitro. Methods: Primary chondrocytes were obtained from the knees of 19 OA patients and cultured either as monolayers or in alginate beads. The cultures were treated with 20% or 10% (v/v) Ze14 or placebo. For RNA-seq, reads were generated with Illumina NextSeq5000 sequencer and aligned to the human reference genome (UCSC hg19). Differential expression analysis between Ze14 and placebo was performed in R using the DESeq2 package. Protein quantification by ELISA was performed on selected genes from the culture medium and/or the cellular fractions of primary human OA chondrocyte cultures. Results: In monolayer cultures, Ze14 20% (v/v) significantly modified the expression of 13 genes in OA chondrocytes by at least 10% with an adjusted p-value < 0.05: EGR1, FOS, NR4A1, DUSP1, ZFP36, ZFP36L1, NFKBIZ, and CCN1 were upregulated and ATF7IP, TXNIP, DEPP1, CLEC3A, and MMP13 were downregulated after 24 h Ze14 treatment. Ze14 significantly increased (mean 2.3-fold after 24 h, p = 0.0444 and 72 h, p = 0.0239) the CCN1 protein production in human OA chondrocytes. After 72 h, Ze14 significantly increased type II collagen pro-peptide production by mean 27% (p = 0.0147). For both time points CCN1 production by OA chondrocytes was correlated with aggrecan (r = 0.66, p = 0.0004) and type II collagen pro-peptide (r = 0.64, p = 0.0008) production. In alginate beads cultures, pro-MMP-13 was decreased by Ze14 from day 7–14 (from −16 to −25%, p < 0.05) and from day 17–21 (−22%, p = 0.0331) in comparison to controls. Conclusion: Ze14 significantly modified the expression of DUSP1, DEPP1, ZFP36/ZFP36L1, and CLEC3A, which may reduce MMP13 expression and activation. Protein analysis confirmed that Ze14 significantly reduced the production of pro-MMP-13. As MMP-13 is involved in type II collagen degradation, Ze14 may limit cartilage degradation. Ze14 also promoted extracellular matrix formation arguably through CCN1 production, a growth factor well correlated with type II collagen and aggrecan production.

NKX2‐1 controls lung cancer progression by inducing DUSP6 to dampen ERK activity

Lung cancer remains a leading cause of cancer death, with unclear mechanisms driving the transition to aggressive cancer with poor prognosis. The RAS/RAF/MEK/ERK pathway is hyper-activated in ~50% of human lung adenocarcinoma (LUAD). An initial activating mutation induces homeostatic feedback mechanisms that limit ERK activity. Additional, undefined genetic hits overcome the feedback, leading to high ERK activity that drives malignant progression. At detection, the majority of LUADs express the homeobox transcription factor NKX2-1, which also limits malignant progression. NKX2-1 constrains LUAD, in part, by maintaining a state of well-differentiation. We asked if loss of NKX2-1 might also contribute to the release of ERK activity that drives tumor progression. Using human tissue samples and cell lines, xenografts, and genetic mouse models, we show that NKX2-1 induces the ERK phosphatase DUSP6. In cells with silenced NKX2-1, re-introduction of NKX2-1 induces DUSP6 and inhibits cell proliferation and migration and tumor growth and metastasis. Our CRISPR knockout studies show that DUSP6 is necessary for NKX2-1-mediated inhibition of ERK activity and tumor progression. Further, DUSP6 expression is sufficient to inhibit RAS-driven LUAD. We conclude that NKX2-1 silencing, and thereby DUSP6 silencing, is a mechanism by which early LUAD can unleash ERK hyperactivation for tumor progression.

SARS-CoV-2 Switches 'on' MAPK and NFκB Signaling via the Reduction of Nuclear DUSP1 and DUSP5 Expression.

Mitogen-activated protein kinases (MAPK) and NF-kappaB (NF-κB) pathway regulate many cellular processes and are essential for immune cells function. Their activity is controlled by dual-specificity phosphatases (DUSPs). A comprehensive analysis of publicly available gene expression data sets of human airway epithelial cells (AECs) infected with SARS-CoV-2 identified DUSP1 and DUSP5 among the lowest induced transcripts within these pathways. These proteins are known to downregulate MAPK and NF-κB pathways; and their lower expression was associated with increased activity of MAPK and NF-κB signaling and enhanced expression of proinflammatory cytokines such as TNF-α. Infection with other coronaviruses did not have a similar effect on these genes. Interestingly, treatment with chloroquine and/or non-steroidal anti-inflammatory drugs counteracted the SARS-CoV-2 induced reduction of DUSP1 and DUSP5 genes expression. Therapeutically, impeding this evasion mechanism of SARS-CoV-2 may help control the exaggerated activation of these immune regulatory pathways during a COVID-19 infection.

Prostaglandin E2 induces dual-specificity phosphatase-1, thereby attenuating inflammatory genes expression in human osteoarthritic synovial fibroblasts

Characteristic features of osteoarthritis (OA) are joint pain and cartilage degeneration. The degeneration is caused by excess induction of matrix metalloproteinases (MMPs) and the pain is caused by nerve growth factor (NGF)-dependent nerve invasion into synovial tissue in addition to nociceptive pain by prostaglandin (PG)E2. The objective of this study was to clarify the suppressive mechanism of PGE2 on the regulation of MMPs and NGF by focusing on mitogen-activated protein kinases (MAPKs) and their endogenous phosphatase, dual-specificity phosphatase (DUSP)-1 in human synovial fibroblasts. PGE2 strongly increased DUSP-1 and suppressed IL-1β-induced MAPKs phosphorylation. Inhibition of MAPKs by selective inhibitors differentially regulated the IL-1β-induced expression of MMPs and NGF expression. IL-1β-induced MAPKs phosphorylation was prolonged and enhanced in DUSP-1 knockdown cells and the expression of MMPs and NGF was also increased. This study revealed that PGE2 has novel biological activity that suppresses NGF and MMPs expression by inducing DUSP-1 expression.

Single-cell RNA-seq reveals dynamic change in tumor microenvironment during pancreatic ductal adenocarcinoma malignant progression

BackgroundPancreatic ductal adenocarcinoma (PDAC) is most aggressive among all gastrointestinal tumors. The complex intra-tumor heterogeneity and special tumor microenvironment in PDAC bring great challenges for developing effective treatment strategies. We aimed to delineate dynamic changes of tumor microenvironment components during PDAC malignant progression utilizing single-cell RNA sequencing. MethodsA total of 11 samples (4 PDAC I, 4 PDAC II, 3 PDAC III) were used to construct expression matrix. After identifying distinct cell clusters, subcluster analysis for each cluster was performed. New cancer associated fibroblasts (CAFs) subset was validated by weighted gene co-expression network analysis, RNA in situ hybridization and immunofluorescence. FindingsWe found that ductal cells were not dominant component while tumor infiltrating immune cells and pancreatic stellate cells gradually accumulated during tumor development. We defined several new Treg and exhausted T cell signature genes, including DUSP4, FANK1 and LAIR2. The analysis of TCGA datasets showed that patients with high expression of DUSP4 had significantly worse prognosis. In addition, we identified a new CAFs subset (complement-secreting CAFs, csCAFs), which specifically expresses complement system components, and constructed csCAFs-related module by weighted gene co-expression network analysis. The csCAFs were located in the tissue stroma adjacent to malignant ductal cells only in early PDAC. InterpretationWe systematically explored PDAC heterogeneity and identified csCAFs as a new CAFs subset special to PDAC, which may be valuable for understanding the crosstalk inside tumor. FundingThis study was supported by The Natural Science Foundation of China (NO.81572339, 81672353, 81871954) and the Youth Clinical Research Project of Peking University First Hospital (2018CR28).

A Ser252Trp substitution in mouse FGFR2 results in hyperplasia of embryonic salivary gland parenchyma

ObjectivesMutations in the fibroblast growth factor receptor 2 (FGFR2) gene are responsible for several severe forms of craniosynostotic disorders, such as Apert and Crouzon syndromes. Patients with craniosynostotic disorders caused by a mutation in Fgfr2 present with several clinical symptoms, including hypersalivation. Here we used a transgenic mouse model of Apert syndrome (Fgfr2+/S252W mice) to evaluate the morphology of the submandibular glands at embryonic day 15.5 (E15.5), the time point reported to mark the start of lumen formation. MethodsFgfr2+/S252W mice were generated by crossing ACTB-Cre+/+ and Fgfr2+/Neo-S252W mice. After measuring body weight, the submandibular glands were collected at E15.5. H&E staining, immunostaining, and RT-qPCR were performed to investigate the development of the submandibular gland. ResultsThe number of ducts and acini in Fgfr2+/S252W mice was significantly higher than in control littermates; however, lumen formation was not affected. The mRNA expression of Fgf1, Fgfr1, Mmp2, Bmp4, Bmp7, Dusp6, and Etv5 in Fgfr2+/S252W mice was significantly higher compared to control littermates. Immunoreactivity for FGF3, FGF1, BMP4, and F4/80 was detected in the parenchyma of Fgfr2+/S252W mice. The area of apoptotic cells stained with TUNEL in Fgfr2+/S252W mice was significantly larger than that of the control littermates. ConclusionsThese results suggested that increased FGFR1 signaling and apoptosis in the submandibular glands of Fgfr2+/S252W mice occurred at E15.5, leading to parenchymal hyperplasia. This study demonstrated that a Ser252Trp substitution in mouse FGFR2 resulted in hyperplasia of the submandibular gland parenchyma during development.

Cefoxitin treatment of MRSA leads to a shift in the IL-12/IL-23 production pattern in dendritic cells by a mechanism involving changes in the MAPK signaling

Methicillin resistant Staphylococcus aureus (MRSA) constitute a serious health care problem worldwide. This study addresses the effect of β-lactam treatment on the ability of clinically relevant MRSA strains to induce IL-12 and IL-23. MRSA strains induced a dose-dependent IL-12 response in murine bone-marrow-derived dendritic cells that was dependent on endocytosis and acidic degradation. Facilitated induction of IL-12 (but not of IL-23) called for activation of the MAP kinase JNK, and was suppressed by p38. Compromised peptidoglycan structure in cefoxitin-treated bacteria – as denoted by increased sensitivity to mutanolysin –caused a shift from IL-12 towards IL-23. Moreover, cefoxitin treatment of MRSA led to a p38 MAPK-dependent early up-regulation of Dual Specificity Phosphatase (DUSP)-1. Compared to common MRSA, characteristics associated with a persister phenotype increased intracellular survival and upon cefoxitin treatment, the peptidoglycan was not equally compromised and the cytokine induction still required phagosomal acidification. Together, these data demonstrate that β-lactam treatment changes the MRSA-induced IL-12/IL-23 pattern determined by the activation of JNK and p38. We suggest that accelerated endosomal degradation of the peptidoglycan in cefoxitin-treated MRSA leads to an early expression of DUSP-1 and accordingly, a reduction in the IL-12/IL-23 ratio in dendritic cells. This may influence the clearance of S. aureus.

The GSK3\u03b2-\u03b2-catenin-TCF1 pathway improves naive T cell activation in old adults by upregulating miR-181a

MicroRNAs play an important role in the regulation of T cell development, activation, and differentiation. One of the most abundant microRNAs in lymphocytes is miR-181a, which controls T cell receptor (TCR) activation thresholds in thymic selection as well as in peripheral T cell responses. We previously found that miR-181a levels decline in T cells in the elderly. In this study, we identified TCF1 as a transcriptional regulator of pri-miR-181a. A decline in TCF1 levels in old individuals accounted for the reduced miR-181a expression impairing TCR signaling. Inhibition of GSK3ß restored expression of miR-181a by inducing TCF1 in T cells from old adults. GSK3ß inhibition enhanced TCR signaling to increase downstream expression of activation markers and production of IL-2. The effect involved the upregulation of miR-181a and the inhibition of DUSP6 expression. Thus, inhibition of GSK3ß can restore responses of old T cells by inducing miR-181a expression through TCF1.

Simvastatin suppresses renal cell carcinoma cells by regulating DDX5/DUSP5

Objectives Renal cell carcinoma (RCC) has the highest mortality rate of genitourinary cancers and the incidence of RCC has risen steadily. Simvastatin has been reported to exhibit anti-tumor activity in a variety of cancers; however, its roles and molecular mechanisms in RCC remain unclear. Our aim was to evaluate the inhibitory effect of simvastatin on RCC. Methods We used a variety of methods to test the changes of RCC cell lines’ viability, migration, invasion, cell cycle and apoptosis after treatment with simvastatin. Results We found that simvastatin not only inhibited RCC cell viability, migration, and invasion, but also regulated the cell cycle and induced apoptosis. We also observed abnormal expression of DDX5 and DUSP5 in RCC cell lines. Mechanistic investigation showed that simvastatin significantly suppressed DDX5 and promoted DUSP5 expression. Conclusion Together, these results provide a novel mechanism underlying simvastatin-induced inhibition of RCC via regulation of the DDX5/DUSP5 axis.

Upregulation of DUSP6 impairs infectious bronchitis virus replication by negatively regulating ERK pathway and promoting apoptosis

Elucidating virus-cell interactions is fundamental to understanding viral replication and identifying targets for therapeutic control of viral infection. The extracellular signal-regulated kinase (ERK) pathway has been shown to regulate pathogenesis during many viral infections, but its role during coronavirus infection is undetermined. Infectious bronchitis virus is the representative strain of Gammacoronavirus, which causes acute and highly contagious diseases in the poultry farm. In this study, we investigated the role of ERK1/2 signaling pathway in IBV infection. We found that IBV infection activated ERK1/2 signaling and the up-regulation of phosphatase DUSP6 formed a negative regulation loop. Pharmacological inhibition of MEK1/2-ERK1/2 signaling suppressed the expression of DUSP6, promoted cell death, and restricted virus replication. In contrast, suppression of DUSP6 by chemical inhibitor or siRNA increased the phosphorylation of ERK1/2, protected cells from apoptosis, and facilitated IBV replication. Overexpression of DUSP6 decreased the level of phospho-ERK1/2, promoted apoptosis, while dominant negative mutant DUSP6-DN lost the regulation function on ERK1/2 signaling and apoptosis. In conclusion, these data suggest that MEK-ERK1/2 signaling pathway facilitates IBV infection, probably by promoting cell survival; meanwhile, induction of DUSP6 forms a negative regulation loop to restrict ERK1/2 signaling, correlated with increased apoptosis and reduced viral load. Consequently, components of the ERK pathway, such as MEK1/2 and DUSP6, represent excellent targets for the development of antiviral drugs.

Single-Cell RNA-Seq Reveals Dynamic Change in Tumor Microenvironment During Pancreatic Ductal Adenocarcinoma Malignant Progression

Background: Pancreatic ductal adenocarcinoma (PDAC) is most aggressive among all gastrointestinal tumors. The complex intra-tumor heterogeneity and special tumor microenvironment in PDAC bring great challenges for developing effective treatment strategies. We aimed to delineate dynamic changes of tumor microenvironment components during PDAC malignant progression utilizing single-cell RNA sequencing. Methods: A total of 11 samples (4 PDAC I, 4 PDAC II, 3 PDAC III) were used to construct expression matrix. After identifying distinct cell clusters, subcluster analysis for each cluster was performed. New cancer associated fibroblasts (CAFs) subset was validated by weighted gene co-expression network analysis, RNA in situ hybridization and immunofluorescence. Findings: We found that ductal cells were not dominant component while tumor infiltrating immune cells and pancreatic stellate cells gradually accumulated during tumor development. We defined several new Treg and exhausted T cell signature genes, including DUSP4, FANK1 and LAIR2. The analysis of TCGA datasets showed that patients with high expression of DUSP4 had significantly worse prognosis. In addition, we identified a new CAFs subset (complement-secreting CAFs, csCAFs), which specifically expresses complement system components, and constructed csCAFs-related module by weighted gene co-expression network analysis. The csCAFs were located in the tissue stroma adjacent to malignant ductal cells only in early PDAC. Interpretation: We systematically explored PDAC heterogeneity and identified csCAFs as a new CAFs subset special to PDAC, which is valuable for understanding the crosstalk inside tumor. Funding Statement: This study was supported by The Natural Science Foundation of China (NO.81572339, 81672353, 81871954) and the Youth Clinical Research Project of Peking University First Hospital (2018CR28). Declaration of Interests: We declare there are no any competing financial interests in relation to this work. Ethics Approval Statement: According to the article (GSA: https://bigd.big.ac.cn/gsa under accession number CRA00116064), all patient samples were obtained from the department of general surgery of Peking Union Medical College Hospital (PUMCH) and patients had signed the informed consent. This study was approved by Ethics Committee.

SETD8 potentiates constitutive ERK1/2 activation via epigenetically silencing DUSP10 expression in pancreatic cancer.

Constitutive ERK1/2 activation has been frequently observed in pancreatic adenocarcinoma (PDAC). How ERK1/2 activation status been potentiated and maintained by epigenetic mechanisms has seldom been discussed in PDAC. In this study, we first examined the expression status of p-ERK1/2 in PDAC tissues by immunohistochemical staining and then screened possible epigenetic factors that displayed different expression status between p-ERK1/2 high and low groups by RNA profiling, and found that SETD8 displayed an increased expressional pattern in p-ERK1/2high patient group. Then the impact of SETD8 on the proliferation of PDAC cells were investigated on the basis of gain or loss-of-function assays. RNA sequencing assays were performed to screen potential SETD8 downstream targets that contribute to ERK1/2 activation. Mass spectrometry and transcriptional analysis, including dual-luciferase assay and chromatin immunoprecipitation assay (ChIP), were used to explore the molecular mechanisms that governing SETD8-mediated ERK1/2 activation. In vitro cell line studies and in vivo xenograft mouse model studies indicated that SETD8 promoted cell proliferation and increased tumor formation capacity of PDAC cell lines. Mechanism explorations uncovered that SETD8 suppressed the expression of DUSP10, which was responsible for dephosphorylation of ERK1/2. Mass spectrometry and transcriptional analysis results demonstrated that STAT3 interacted with SETD8 and recruited SETD8 to the promoter region of DUSP10, leading to epigenetic silencing of DUSP10 and the resultant activation of ERK1/2. In conclusion, SETD8 interacts with STAT3 on DUSP10 promoter region and epigenetically silences DUSP10 expression. Decreased DUSP10 expression in PDAC potentiates activation of ERK1/2 phosphorylation, resulting in unfavorable prognosis of PDAC.

Transcriptional regulation in the rat prefrontal cortex and hippocampus after a single administration of psilocybin

Psilocybin is a serotonergic psychedelic found in "magic mushrooms" with a putative therapeutic potential for treatment-resistant depression, anxiety, obsessive-compulsive disorder, and addiction. In rodents, psilocybin acutely induces plasticity-related immediate early genes in cortical tissue; however, studies into the effects on subcortical regions, of different doses, and the subsequent translation of corresponding proteins are lacking. We examined the acute effects of a single administration of psilocybin (0.5-20 mg/kg) on the expression of selected genes in the prefrontal cortex and hippocampus. In total, 46 target genes and eight reference genes were assessed using real-time quantitative polymerase chain reaction. Corresponding protein levels of the three most commonly regulated genes were assessed using Western blotting. In the prefrontal cortex, psilocybin increased the expression of Cebpb, c-Fos, Dups1, Fosb, Junb, Iκβ-α, Nr4a1, P11, Psd95, and Sgk1, and decreased the expression of Clk1. In the hippocampus, psilocybin strongly increased the expression of Arrdc2, Dusp1, Iκβ-α, and Sgk1 in a dose-dependent manner, and decreased the expression of Arc, Clk1, Egr2, and Ptgs2. Protein levels of Sgk1, Dusp1, and Iκβ-α showed only partial agreement with transcriptional patterns, stressing the importance of assessing downstream translation when investigating rapid gene responses. The present study demonstrates that psilocybin rapidly induces gene expression related to neuroplasticity, biased towards the prefrontal cortex, compared to the hippocampus. Our findings provide further evidence for the rapid plasticity-promoting effects of psilocybin.

TRIM65 Promotes Invasion of Endometrial Stromal Cells by Activating ERK1/2/C-myc Signaling via Ubiquitination of DUSP6

Endometriosis (EM) is a benign gynecological disease that shares some characteristics with malignancy, such as proliferation and invasion. So far, the pathogenesis of EM is still unclear. In this study, we investigated whether TRIM65 can play a role in the development of EM. TRIM65 expression levels in eutopic, ectopic, and normal endometrium were detected by quantitative real-time PCR and Western blot. Cell proliferation and invasion of primary endometrial stromal (EMS) cells were detected by CCK-8 and Transwell analysis. The interaction between TRIM65 and DUSP6 or C-myc was measured by coimmunoprecipitation, ubiquitylation, dual luciferase, and chromatin immunoprecipitation analysis. We found that TRIM65 was identified as an up-regulated gene in ectopic endometrial tissues and EMS cells compared with control groups without EM. TRIM65 expression was positively correlated with the levels of p-ERK1/2, C-myc, matrix metalloproteinase-2, and integrin β1 in ectopic endometrial tissues in patients and mice. TRIM65 promoted the cell proliferation and invasion of EMS cells via the ERK1/2/C-myc pathway through ubiquitination of DUSP6. C-myc promoted TRIM65 expression through inducing TRIM65 promoter activity. Additionally, the increased expression of TRIM65, C-myc, matrix metalloproteinase-2, integrin β1, and p-ERK1/2 and the decreased expression of DUSP6 in ectopic endometrial tissues were significantly suppressed by inhibition of ERK1/2 signaling pathway in ectopic endometrial tissues in experimental mice model. In conclusion, TRIM65 promotes invasion of ectopic EMS cells by activating a feedback loop with the ERK1/2/C-myc signaling pathway and may be a potential therapeutic target for EM.

An integrative analysis of genome-wide methylation and expression in ameloblastoma: A pilot study.

DNA methylation regulates the expression of various genes involved in tumorigenesis. Ameloblastoma is a benign odontogenic jaw tumor. It is locally aggressive with a high level of recurrence. A delay in treatment can lead to severe facial disfigurement. To the best of our knowledge, this is the first integrated analysis of DNA methylation and gene expression in ameloblastoma with the aim to identify genes that may be regulated by DNA methylation. We used an Infinium MethylationEPIC array to measure genome-wide methylation and the Illumina HiSeq platform to obtain gene expression data in ameloblastoma tissues from five patients and dental follicles from three healthy subjects. An integration analysis was performed using City of Hope CpG Island Analysis Pipeline software. We identified 25,255 differentially methylated CpG sites and 17 differentially methylated CpG islands; six of the islands were negatively correlated with the expression of BAIAP2, DUSP6, FGFR2, FOXF2, NID2, and PAK6. Pyrosequencing and immunostaining techniques were further used to validate FGFR2, NID2, and PAK6. This analysis identifies a group of novel genes that may be regulated by DNA methylation and will possibly lead to new insights into the pathology and invasion mechanism of ameloblastoma.

M2 macrophage-derived exosomal miR-25-3p improves high glucose-induced podocytes injury through activation autophagy via inhibiting DUSP1 expression.

Diabetic nephropathy (DN) is the primary reason of chronic kidney disease. The aim of our study is to explore the role and action mechanism of M2 macrophage-derived exosomes in high glucose (HG)-induced podocytes injury. Here, 30 mmol/L of HG was used to induce podocytes injury. Annexin V-FITC/PI double staining was performed to measure podocytes apoptosis, and western blot was carried out to ensure proteins expression. The shape of exosomes was identified using TEM. Besides, the expression of miR-25-3p was determined by qRT-PCR, FAM-labeled miR-25-5p combined with DiI-labeled exosomes were utilized to explore the uptake of podocytes to exosomes. Relationship between miR-25-3p and DUSP family members was ensued by luciferase activity assay. In the beginning, we found that M2 macrophage ameliorated HG-induced podocytes apoptosis and epithelial-mesenchymal transition through secreting exosomes. Subsequently, highly expressed miR-25-3p was found in M2 macrophage-derived exosomes that effectively improved HG-induced podocytes injury. Furthermore, inhibition of miR-25-3p in M2 macrophage inefficiently repressed HG-induced podocytes injury, thus we proposed that M2 macrophage attenuated podocytes injury through secreting exosomal miR-25-3p. Then, we used an autophagy inhibitor to stimulate podocytes, and demonstrated that M2 macrophage-derived exosomal miR-25-3p improved HG-induced podocytes injury through activating autophagy. Finally, DUSP1 was proved to be a downstream target and mediated the inhibition of exosomal miR-25-3p to HG-induced podocytes injury. Our results indicated that M2 macrophage could improve HG-induced podocytes injury via secreting exosomal miR-25-3p to activate autophagy of the cells through suppressing DUSP1 expression. We proved a newly potential therapy strategy for DN treatment.

KRAS K104 modification affects the KRASG12D-GEF interaction and mediates cell growth and motility

Mutant RAS genes play an important role in regulating tumors through lysine residue 104 to impair GEF-induced nucleotide exchange, but the regulatory role of KRAS K104 modification on the KRASG12D mutant remains unclear. Therefore, we simulated the acetylation site on the KRASG12D three-dimensional protein structure, including KRASG12D, KRASG12D/K104A and KRASG12D/K104Q, and determined their trajectories and binding free energy with GEF. KRASG12D/K104Q induced structural changes in the α2- and α3-helices, promoted KRAS instability and hampered GEF binding (ΔΔG = 6.14 kJ/mol). We found decreased binding to the Raf1 RBD by KRASG12D/K104Q and reduced cell growth, invasion and migration. Based on whole-genome cDNA microarray analysis, KRASG12D/K104Q decreased expression of NPIPA2, DUSP1 and IL6 in lung and ovarian cancer cells. This study reports computational and experimental analyses of Lys104 of KRASG12D and GEF, and the findings provide a target for exploration for future treatment.

Upregulation of JHDM1D-AS1 alleviates neuroinflammation and neuronal injury via targeting miR-101-3p-DUSP1 in spinal cord after brachial plexus injury

BackgroundNeuroinflammation in the spinal cord following acute brachial plexus injury (BPI) remains a vital cause that leads to motor dysfunction and neuropathic pain. In this study, we aim to explore the role of long non-coding RNA JHDM1D antisense 1 (JHDM1D-AS1) in mediating BPI-induced neuroinflammation and neuronal injury. MethodsA total brachial plexus root avulsion (tBPRA) model in adult rats and IL-1β-treated motor neuron-like NSC-34 cells and LPS-treated microglia cell line BV2 were conducted for in vivo and in vitro experiments, respectively. The expressions of JHDM1D-AS1, miR-101-3p and DUSP1, p38, NF-κB, TNF-α, IL-1β, and IL-6 were detected by RT-PCR and western blot seven days after tBPI. Immunohistochemistry (IHC) was used to detect neuronal apoptosis. CCK8 assay, Tunel assay and LDH kit were used for the detection of neuronal injury. The targeted relationships between JHDM1D-AS1 and miR-101-3p, miR-101-3p and DUSP1 were verified by RNA immunoprecipitation (RIP) and dual-luciferase reporter gene assay. ResultsWe found significant downregulated expression of JHDM1D-AS1 and DUSP1 but upregulated expression of miR-101-3p in the spinal cord after tBPI. Overexpression of JHDM1D-AS1 had a prominent neuroprotective effect by suppressing neuronal apoptosis and microglial inflammation through reactivation of DUSP1. Further exploration revealed that JHDM1D-AS1 may act as a competitive endogenous RNA targeting miR-101-3p, which bound on the 3′UTR of DUSP1 mRNA. In addition, overexpression of miR-101-3p could reverse the neuroprotective effects of JHDM1D-AS1 upregulation by blocking DUSP1. ConclusionsJHDM1D-AS1 exerted neuroprotective and anti-inflammatory effects in a rat model of tBPI by regulating miR-101-3p/DUSP1 axis.