Dataset Information

Cholera toxin induces malignant glioma cell differentiation via the PKA/CREB pathway.

ABSTRACT: Malignant gliomas are one of the leading causes of cancer deaths worldwide, but chemoprevention strategies for them are few and poorly investigated. Here, we show that cholera toxin, the traditional biotoxin and well known inducer of accumulation of cellular cAMP, is capable of inducing differentiation on malignant gliomas in vitro with rat C6 and primary cultured human glioma cells. Cholera toxin-induced differentiation was characterized by typical morphological changes, increased expression of glial fibrillary acid protein, decreased expression of Ki-67, inhibition of cellular proliferation, and accumulation of cells in the G(1) phase of the cell cycle. Cholera toxin also triggered a significant reduction in the G(1) cell-cycle regulatory proteins cyclin D1 and Cdk2 along with an overexpression of cell-cycle inhibitory proteins p21(Cip1) and p27(Kip1). Abrogation of cAMP-dependent protein kinase A activity by protein kinase A inhibitor or silencing of cAMP-responsive element binding proteins by RNA interference resulted in suppressed differentiation. These findings imply the attractiveness of cholera toxin as a drug candidate for further development of differentiation therapy. Furthermore, activation of the protein kinase A/cAMP-responsive element binding protein pathway may be a key and requisite factor in glioma differentiation.

SUBMITTER: Li Y

PROVIDER: S-EPMC1940034 | biostudies-literature | 2007 Aug

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Cholera toxin induces malignant glioma cell differentiation via the PKA/CREB pathway.

Li Yan Y Yin Wei W Wang Xia X Zhu Wenbo W Huang Yijun Y Yan Guangmei G

Proceedings of the National Academy of Sciences of the United States of America 20070806 33

Malignant gliomas are one of the leading causes of cancer deaths worldwide, but chemoprevention strategies for them are few and poorly investigated. Here, we show that cholera toxin, the traditional biotoxin and well known inducer of accumulation of cellular cAMP, is capable of inducing differentiation on malignant gliomas in vitro with rat C6 and primary cultured human glioma cells. Cholera toxin-induced differentiation was characterized by typical morphological changes, increased expression of ...[more]

PMID: 17679696

Similar Datasets

Project description:BackgroundLactate has emerged as a critical regulator within the tumor microenvironment, including glioma. However, the precise mechanisms underlying how lactate influences the communication between tumor cells and tumor-associated macrophages (TAMs), the most abundant immune cells in glioma, remain poorly understood. This study aims to elucidate the impact of tumor-derived lactate on TAMs and investigate the regulatory pathways governing TAM-mediated tumor-promotion in glioma.MethodsBioinformatic analysis was conducted using datasets from TCGA and CGGA. Single-cell RNA-seq datasets were analyzed by using UCSC Cell Browser and Single Cell Portal. Cell proliferation and mobility were evaluated through CCK8, colony formation, wound healing, and transwell assays. Western blot and immunofluorescence staining were applied to assess protein expression and cell distribution. RT-PCR and ELISA were employed to identify the potential secretory factors. Mechanistic pathways were explored by western blotting, ELISA, shRNA knockdown, and specific inhibitors and activators. The effects of pathway blockades were further assessed using subcutaneous and intracranial xenograft tumor models in vivo.ResultsElevated expressions of LDHA and MCT1 were observed in glioma and exhibited a positive correlation with M2-type TAM infiltration. Lactate derived from glioma cells induced TAMs towards M2-subtype polarization, subsequently promoting glioma cells proliferation, migration, invasion, and mesenchymal transition. GPR65, highly expressed on TAMs, sensed lactate-stimulation in the TME, fueling glioma cells malignant progression through the secretion of HMGB1. GPR65 on TAMs triggered HMGB1 release in response to lactate stimulation via the cAMP/PKA/CREB signaling pathway. Disrupting this feedback loop by GPR65-knockdown or HMGB1 inhibition mitigated glioma progression in vivo.ConclusionThese findings unveil the intricate interplay between TAMs and tumor cells mediated by lactate and HMGB1, driving tumor progression in glioma. GPR65, selectively highly expressed on TAMs in glioma, sensed lactate stimulation and fostered HMGB1 secretion via the cAMP/PKA/CREB signaling pathway. Blocking this feedback loop presents a promising therapeutic strategy for GBM.

Project description:Differentiation-inducing therapy has been proposed to be a novel potential approach to treat malignant gliomas. Glial fibrillary acidic protein (GFAP) is a well-known specific astrocyte biomarker and acts as a tumor suppressor gene (TSG) in glioma pathogenesis. Previously we reported that a traditional biotoxin cholera toxin could induce malignant glioma cell differentiation characterized by morphologic changes and dramatic GFAP expression. However, the molecular mechanisms underlying GFAP induction are still largely unknown. Here we demonstrate that an oncogenic pathway interleukin-6/janus kinase-2/signal transducer and activator of transcription 3 (IL-6/JAK2/STAT3) cascade mediates the cholera toxin-induced GFAP expression. Cholera toxin dramatically stimulated GFAP expression at the transcriptional level in C6 glioma cells. Meanwhile, phosphorylation of STAT3 and JAK2 was highly induced in a time-dependent manner after cholera toxin incubation, whereas no changes of STAT3 and JAK2 were observed. Furthermore, the IL-6 gene was quickly induced by cholera toxin and subsequent IL-6 protein secretion was stimulated. Importantly, exogenous recombinant rat IL-6 can also induce phosphorylation of STAT3 concomitant with GFAP expression while JAK2 specific inhibitor AG490 could effectively block both cholera toxin- and IL-6-induced GFAP expression. Given that the methylation of the STAT3 binding element can suppress GFAP expression, we detected the methylation status of the critical recognition sequence of STAT3 in the promoter of GFAP gene (-1518 ∼ -1510) and found that it was unmethylated in C6 glioma cells. In addition, neither DNA methyltransferase1 (DNMT1) inhibitor 5-Aza-2'-deoxycytidine (5-AZa-CdR) nor silencing DNMT1 can stimulate GFAP expression, indicating that the loss of GFAP expression in C6 cells is not caused by its promoter hypermethylation. Taken together, our findings suggest that activation of a pro-survival IL-6/JAK2/STAT3 cascade contributes to cholera toxin-induced GFAP expression, which implies that a survival-promoting signal may also play a differentiation-supporting role in malignant gliomas.

Project description:BackgroundMicroglia, the innate immune cells in the central nervous system, play an essential role in brain homeostasis, neuroinflammation and brain infections. Dysregulated microglia, on the other hand, are associated with neurodegenerative diseases, yet the mechanisms underlying pro-inflammatory gene expression in microglia are incompletely understood.MethodsWe investigated the role of the actin-associated protein tropomyosin 1 (TPM1) in regulating pro-inflammatory phenotype of microglia in the retina by using a combination of cell culture, immunocytochemistry, Western blot, qPCR, TUNEL, RNA sequencing and electroretinogram analysis. TREM2-/- mice were used to investigate whether TPM1 regulated pro-inflammatory responses downstream of TREM2. To conditionally deplete microglia, we backcrossed CX3CR1CreER mice with Rosa26iDTR mice to generate CX3CR1CreER:Rosa26iDTR mice.ResultsWe revealed a vital role for TPM1 in regulating pro-inflammatory phenotype of microglia. We found that TPM1 drove LPS-induced inflammation and neuronal death in the retina via the PKA/CREB pathway. TPM1 knockdown ameliorated LPS-induced inflammation in WT retinas yet exaggerated the inflammation in TREM2-/- retinas. RNA sequencing revealed that genes associated with M1 microglia and A1 astrocytes were significantly downregulated in LPS-treated WT retinas but upregulated in LPS-treated TREM2-/- retinas after TPM1 knockdown. Mechanistically, we demonstrated that CREB activated by TPM1 knockdown mediated anti-inflammatory genes in LPS-treated WT retinas but pro-inflammatory genes in LPS-treated TREM2-/- retinas, suggesting a novel role for TREM2 as a brake on TPM1-mediated inflammation. Furthermore, we identified that TPM1 regulated inflammation downstream of TREM2 and in a microglia-dependent manner.ConclusionsWe demonstrate that TPM1 mediates inflammation downstream of TREM2 via the PKA/CREB signaling pathway. Our findings suggest that TPM1 could be a potential target for therapeutic intervention in brain diseases.

Project description:Dopamine (DA), a neurotransmitter in the nervous system, has been shown to modulate immune function. We have previously reported that five subtypes of DA receptors, including D1R, D2R, D3R, D4R and D5R, are expressed in T lymphocytes and they are involved in regulation of T cells. However, roles of these DA receptor subtypes and their coupled signal-transduction pathway in modulation of natural killer (NK) cells still remain to be clarified. The spleen of mice was harvested and NK cells were isolated and purified by negative selection using magnetic activated cell sorting. After NK cells were incubated with various drugs for 4 h, flow cytometry measured cytotoxicity of NK cells against YAC-1 lymphoma cells. NK cells expressed the five subtypes of DA receptors at mRNA and protein levels. Activation of D1-like receptors (including D1R and D5R) with agonist SKF38393 enhanced NK cell cytotoxicity, but activation of D2-like receptors (including D2R, D3R and D4R) with agonist quinpirole attenuated NK cells. Simultaneously, SKF38393 elevated D1R and D5R expression, cAMP content, and phosphorylated cAMP-response element-binding (CREB) level in NK cells, while quinpirole reduced D3R and D4R expression, cAMP content, and phosphorylated CREB level in NK cells. These effects of SKF38393 were blocked by SCH23390, an antagonist of D1-like receptors, and quinpirole effects were abolished by haloperidol, an antagonist of D2-like receptors. In support these results, H89, an inhibitor of phosphokinase A (PKA), prevented the SKF38393-dependent enhancement of NK cells and forskolin, an activator of adenylyl cyclase (AC), counteracted the quinpirole-dependent suppression of NK cells. These findings show that DA receptor subtypes are involved in modulation of NK cells and suggest that D1-like receptors facilitate NK cells by stimulating D1R/D5R-cAMP-PKA-CREB signaling pathway and D2-like receptors suppress NK cells by inhibiting D3R/D4R-cAMP-PKA-CREB signaling pathway. The results may provide more targets of therapeutic strategy for neuroimmune diseases.

Project description:Background and purposeNeuronal pyroptosis is a type of regulated cell death triggered by proinflammatory signals. CCR5 (C-C chemokine receptor 5)-mediated inflammation is involved in the pathology of various neurological diseases. This study investigated the impact of CCR5 activation on neuronal pyroptosis and the underlying mechanism involving cAMP-dependent PKA (protein kinase A)/CREB (cAMP response element binding)/NLRP1 (nucleotide-binding domain leucine-rich repeat pyrin domain containing 1) pathway after experimental intracerebral hemorrhage (ICH).MethodsA total of 194 adult male CD1 mice were used. ICH was induced by autologous whole blood injection. Maraviroc (MVC)-a selective antagonist of CCR5-was administered intranasally 1 hour after ICH. To elucidate the underlying mechanism, a specific CREB inhibitor, 666-15, was administered intracerebroventricularly before MVC administration in ICH mice. In a set of naive mice, rCCL5 (recombinant chemokine ligand 5) and selective PKA activator, 8-Bromo-cAMP, were administered intracerebroventricularly. Short- and long-term neurobehavioral assessments, Western blot, Fluoro-Jade C, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and immunofluorescence staining were performed.ResultsThe brain expression of CCL5 (chemokine ligand 5), CCR5, PKA-Cα (protein kinase A-Cα), p-CREB (phospho-cAMP response element binding), and NLRP1 was increased, peaking at 24 hours after ICH. CCR5 was expressed on neurons, microglia, and astrocytes. MVC improved the short- and long-term neurobehavioral deficits and decreased neuronal pyroptosis in ipsilateral brain tissues at 24 hours after ICH, which were accompanied by increased PKA-Cα and p-CREB expression, and decreased expression of NLRP1, ASC (apoptosis-associated speck-like protein containing a CARD), C-caspase-1, GSDMD (gasdermin D), and IL (interleukin)-1β/IL-18. Such effects of MVC were abolished by 666-15. At 24 hours after injection in naive mice, rCCL5 induced neurological deficits, decreased PKA-Cα and p-CREB expression in the brain, and upregulated NLRP1, ASC, C-caspase-1, N-GSDMD, and IL-1β/IL-18 expression. Those effects of rCCL5 were reversed by 8-Bromo-cAMP.ConclusionsCCR5 activation promoted neuronal pyroptosis and neurological deficits after ICH in mice, partially through the CCR5/PKA/CREB/NLRP1 signaling pathway. CCR5 inhibition with MVC may provide a promising therapeutic approach in managing patients with ICH.

Dataset Information

Cholera toxin induces malignant glioma cell differentiation via the PKA/CREB pathway.

Publications

Cholera toxin induces malignant glioma cell differentiation via the PKA/CREB pathway.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets