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Arteriosclerosis, Thrombosis, and... Jul 2024Tight control of cytoplasmic Ca in endothelial cells is essential for the regulation of endothelial barrier function. Here, we investigated the role of Cavβ3, a subunit...
BACKGROUND
Tight control of cytoplasmic Ca in endothelial cells is essential for the regulation of endothelial barrier function. Here, we investigated the role of Cavβ3, a subunit of voltage-gated Ca (Cav) channels, in modulating Ca signaling in brain microvascular endothelial cells (BMECs) and how this contributes to the integrity of the blood-brain barrier.
METHODS
We investigated the function of Cavβ3 in BMECs by Ca imaging and Western blot, examined the endothelial barrier function in vitro and the integrity of the blood-brain barrier in vivo, and evaluated disease course after induction of experimental autoimmune encephalomyelitis in mice using Cavβ3 (Cav β3-deficient) mice as controls.
RESULTS
We identified Cavβ3 protein in BMECs, but electrophysiological recordings did not reveal significant Cav channel activity. In vivo, blood-brain barrier integrity was reduced in the absence of Cavβ3. After induction of experimental autoimmune encephalomyelitis, Cavβ3 mice showed earlier disease onset with exacerbated clinical disability and increased T-cell infiltration. In vitro, the transendothelial resistance of Cavβ3 BMEC monolayers was lower than that of wild-type BMEC monolayers, and the organization of the junctional protein ZO-1 (zona occludens-1) was impaired. Thrombin stimulates inositol 1,4,5-trisphosphate-dependent Ca release, which facilitates cell contraction and enhances endothelial barrier permeability via Ca-dependent phosphorylation of MLC (myosin light chain). These effects were more pronounced in Cavβ3 than in wild-type BMECs, whereas the differences were abolished in the presence of the MLCK (MLC kinase) inhibitor ML-7. Expression of cDNA in Cavβ3 BMECs restored the wild-type phenotype. Coimmunoprecipitation and mass spectrometry demonstrated the association of Cavβ3 with inositol 1,4,5-trisphosphate receptor proteins.
CONCLUSIONS
Independent of its function as a subunit of Cav channels, Cavβ3 interacts with the inositol 1,4,5-trisphosphate receptor and is involved in the tight control of cytoplasmic Ca and Ca-dependent MLC phosphorylation in BMECs, and this role of Cavβ3 in BMECs contributes to blood-brain barrier integrity and attenuates the severity of experimental autoimmune encephalomyelitis disease.
PubMed: 38957986
DOI: 10.1161/ATVBAHA.124.321141 -
Plant & Cell Physiology Jul 2024The INDUCER OF CBF EXPRESSION 1/C-REPEAT BINDING FACTOR (ICE1/CBF) pathway plays a crucial role in plant responses to cold stress, impacting growth and development....
The INDUCER OF CBF EXPRESSION 1/C-REPEAT BINDING FACTOR (ICE1/CBF) pathway plays a crucial role in plant responses to cold stress, impacting growth and development. Here, we demonstrated that ATBS1-INTERACTING FACTOR 2 (AIF2), a non-DNA-binding basic helix-loop-helix transcription factor, positively regulates freezing tolerance through the ICE1/CBF-induced cold tolerance pathway in Arabidopsis. Cold stress transcriptionally upregulated AIF2 expression and induced AIF2 phosphorylation, thereby stabilizing the AIF2 protein during early stages of cold acclimation. The AIF2 loss-of-function mutant, aif2-1, exhibited heightened sensitivity to freezing before and after cold acclimation. In contrast, ectopic expression of AIF2, but not the C-terminal-deleted AIF2 variant, restored freezing tolerance. AIF2 enhanced ICE1 stability during cold acclimation and promoted the transcriptional expression of CBFs and downstream cold-responsive genes, ultimately enhancing plant tolerance to freezing stress. MITOGEN-ACTIVATED PROTEIN KINASES 3 and 6 (MPK3/6), known negative regulators of freezing tolerance, interacted with and phosphorylated AIF2, subjecting it to protein degradation. Furthermore, transient co-expression of MPK3/6 with AIF2 and ICE1 downregulated AIF2/ICE1-induced transactivation of CBF2 expression. AIF2 interacted preferentially with BIN2 and MPK3/6 during the early and later stages of cold acclimation, respectively, thereby differentially regulating AIF2 activity in a cold acclimation time-dependent manner. Moreover, AIF2 acted additively in a gain-of-function mutant of BRASSINAZOLE-RESISTANT 1 (BZR1; bzr1-1D) and a triple knockout mutant of BRASSINOSTEROID-INSENSITIVE 2 (BIN2) and its homologs (bin2bil1bil2) to induce CBFs-mediated freezing tolerance. This suggests that cold-induced AIF2 coordinates freezing tolerance along with BZR1 and BIN2, key positive and negative components, respectively, of brassinosteroid signaling pathways.
PubMed: 38957969
DOI: 10.1093/pcp/pcae072 -
Dental Research and Oral Health 2024Topoisomerase IIα (TOP2A), is an enzyme involved in DNA replication, transcription, recombination, and chromatin remodeling and is found in a variety of cancers....
BACKGROUND
Topoisomerase IIα (TOP2A), is an enzyme involved in DNA replication, transcription, recombination, and chromatin remodeling and is found in a variety of cancers. However, the role of TOP2A regulation in oral cancer progression is not fully explained. We investigated the effect of TOP2A inhibition on cell survival, metabolism, and cancer stem cell self-renewal function in oral cancer cells.
METHODS
Oral carcinoma cell line SCC25 was cultured in complete DMEM/F12 media and treated with 5μM of Etoposide (Topoisomerase II inhibitor) for 48h. The critical parameters of cellular metabolism, including extracellular acidification rate (ECAR) and mitochondrial oxidative phosphorylation based on the oxygen consumption rate of cancer cells were assessed using Seahorse assay. Western blotting was performed to assess the proteins that are associated with proliferation (Survivin, IL-6) and cancer stem cell function (Oct4, Sox2) in cell lysates prepared from control and etoposide treated groups. Statistical analysis was performed using One-way ANOVA with Dunnett's multiple comparisons test.
RESULTS
The protein expression of TOP2A was significantly (P<0.05) inhibited by etoposide. Additionally, TOP2A inhibition decreased the mitochondrial respiratory parameters including basal respiration, maximal respiration and ATP production. However, TOP2A inhibition has no impact on glycolytic function. Moreover, the proliferative marker survivin and IL-6 showed a significant (P<0.05) decrease after TOP2A inhibition. Conversely, the protein expression of cancer stem cell markers Oct-4 and Sox 2 were not altered.
CONCLUSION
These results indicate that inhibition of TOP2A is more efficacious by decreasing the mitochondrial metabolic reprogramming and thereby downregulating the key anti-apoptotic and pro-survival mediators. Thus, TOP2A represents an ideal therapeutic target and offers a potential treatment strategy for OSCC.
PubMed: 38957610
DOI: 10.26502/droh.0076 -
Journal of Molecular and Cellular... Jun 2024Type 2 diabetes mellitus (T2DM) is a metabolic disease and comorbidity associated with several conditions, including cardiac dysfunction leading to heart failure with...
Type 2 diabetes mellitus (T2DM) is a metabolic disease and comorbidity associated with several conditions, including cardiac dysfunction leading to heart failure with preserved ejection fraction (HFpEF), in turn resulting in T2DM-induced cardiomyopathy (T2DM-CM). However, the molecular mechanisms underlying the development of T2DM-CM are poorly understood. It is hypothesized that molecular alterations in myopathic genes induced by diabetes promote the development of HFpEF, whereas cardiac myosin inhibitors can rescue the resultant T2DM-mediated cardiomyopathy. To test this hypothesis, a Leptin receptor-deficient homozygous (Lepr ) mouse model was used to define the pathogenesis of T2DM-CM. Echocardiographic studies at 4 and 6 months revealed that Lepr db/db hearts started developing cardiac dysfunction by four months, and left ventricular hypertrophy with diastolic dysfunction was evident at 6 months. RNA-seq data analysis, followed by functional enrichment, revealed the differential regulation of genes related to cardiac dysfunction in Lepr heart tissues. Strikingly, the level of cardiac myosin binding protein-C phosphorylation was significantly increased in Lepr mouse hearts. Finally, using isolated skinned papillary muscles and freshly isolated cardiomyocytes, (mavacamten, MYK-461), a prescription heart medicine used for symptomatic obstructive hypertrophic cardiomyopathy treatment, was tested for its ability to rescue T2DM-CM. Compared with controls, MYK-461 significantly reduced force generation in papillary muscle fibers and cardiomyocyte contractility in the db/db group. This line of evidence shows that 1) T2DM-CM is associated with hyperphosphorylation of cardiac myosin binding protein-C and 2) MYK-461 significantly lessened disease progression , suggesting its promise as a treatment for HFpEF.
PubMed: 38957358
DOI: 10.1016/j.jmccpl.2024.100075 -
Journal of Cellular and Molecular... Jul 2024This research explores the role of microRNA in senescence of human endothelial progenitor cells (EPCs) induced by replication. Hsa-miR-134-5p was found up-regulated in...
This research explores the role of microRNA in senescence of human endothelial progenitor cells (EPCs) induced by replication. Hsa-miR-134-5p was found up-regulated in senescent EPCs where overexpression improved angiogenic activity. Hsa-miR-134-5p, which targeted transforming growth factor β-activated kinase 1-binding protein 1 (TAB1) gene, down-regulated TAB1 protein, and inhibited phosphorylation of p38 mitogen-activated protein kinase (p38) in hsa-miR-134-5p-overexpressed senescent EPCs. Treatment with siRNA specific to TAB1 (TAB1si) down-regulated TAB1 protein and subsequently inhibited p38 activation in senescent EPCs. Treatment with TAB1si and p38 inhibitor, respectively, showed angiogenic improvement. In parallel, transforming growth factor Beta 1 (TGF-β1) was down-regulated in hsa-miR-134-5p-overexpressed senescent EPCs and addition of TGF-β1 suppressed the angiogenic improvement. Analysis of peripheral blood mononuclear cells (PBMCs) disclosed expression levels of hsa-miR-134-5p altered in adult life, reaching a peak before 65 years, and then falling in advanced age. Calculation of the Framingham risk score showed the score inversely correlates with the hsa-miR-134-5p expression level. In summary, hsa-miR-134-5p is involved in the regulation of senescence-related change of angiogenic activity via TAB1-p38 signalling and via TGF-β1 reduction. Hsa-miR-134-5p has a potential cellular rejuvenation effect in human senescent EPCs. Detection of human PBMC-derived hsa-miR-134-5p predicts cardiovascular risk.
Topics: MicroRNAs; Humans; Endothelial Progenitor Cells; Cellular Senescence; Leukocytes, Mononuclear; Middle Aged; Adaptor Proteins, Signal Transducing; Male; Cardiovascular Diseases; p38 Mitogen-Activated Protein Kinases; Female; Aged; Neovascularization, Physiologic; Transforming Growth Factor beta1; Adult; Risk Factors
PubMed: 38957039
DOI: 10.1111/jcmm.18523 -
Korean Circulation Journal Jun 2024Although the clinical consequences of advanced heart failure (HF) may be similar across different etiologies of cardiomyopathies, their proteomic expression may show...
BACKGROUND AND OBJECTIVES
Although the clinical consequences of advanced heart failure (HF) may be similar across different etiologies of cardiomyopathies, their proteomic expression may show substantial differences in relation to underlying pathophysiology. We aimed to identify myocardial tissue-based proteomic characteristics and the underlying molecular pathophysiology in non-ischemic cardiomyopathy with different etiologies.
METHODS
Comparative extensive proteomic analysis of the myocardium was performed in nine patients with biopsy-proven non-ischemic cardiomyopathies (3 dilated cardiomyopathy [DCM], 2 hypertrophic cardiomyopathy [HCM], and 4 myocarditis) as well as five controls using tandem mass tags combined with liquid chromatography-mass spectrometry. Differential protein expression analysis, Gene Ontology (GO) analysis, and Ingenuity Pathway Analysis (IPA) were performed to identify proteomic differences and molecular mechanisms in each cardiomyopathy type compared to the control. Proteomic characteristics were further evaluated in accordance with clinical and pathological findings.
RESULTS
The principal component analysis score plot showed that the controls, DCM, and HCM clustered well. However, myocarditis samples exhibited scattered distribution. IPA revealed the downregulation of oxidative phosphorylation and upregulation of the sirtuin signaling pathway in both DCM and HCM. Various inflammatory pathways were upregulated in myocarditis with the downregulation of Rho GDP dissociation inhibitors. The molecular pathophysiology identified by extensive proteomic analysis represented the clinical and pathological properties of each cardiomyopathy with abundant proteomes.
CONCLUSIONS
Different etiologies of non-ischemic cardiomyopathies in advanced HF exhibit distinct proteomic expression despite shared pathologic findings. The benefit of tailored management strategies considering the different proteomic expressions in non-ischemic advanced HF requires further investigation.
PubMed: 38956938
DOI: 10.4070/kcj.2024.0033 -
Cellular & Molecular Biology Letters Jul 2024An increasing number of studies have demonstrated the association of circular RNAs (circRNAs) with the pathological processes of various diseases and their involvement...
BACKGROUND
An increasing number of studies have demonstrated the association of circular RNAs (circRNAs) with the pathological processes of various diseases and their involvement in the onset and progression of multiple cancers. Nevertheless, the functional roles and underlying mechanisms of circRNAs in the autophagy regulation of gastric cancer (GC) have not been fully elucidated.
METHODS
We used transmission electron microscopy and the mRFP-GFP-LC3 dual fluorescent autophagy indicator to investigate autophagy regulation. The cell counting kit-8 assay, colony formation assay, 5-ethynyl-2'-deoxyuridine incorporation assay, Transwell assay, and Western blot assay were conducted to confirm circPTPN22's influence on GC progression. Dual luciferase reporter assays validated the binding between circPTPN22 and miR-6788-5p, as well as miR-6788-5p and p21-activated kinase-1 (PAK1). Functional rescue experiments assessed whether circPTPN22 modulates PAK1 expression by competitively binding miR-6788-5p, affecting autophagy and other biological processes in GC cells. We investigated the impact of circPTPN22 on in vivo GC tumors using a nude mouse xenograft model. Bioinformatics tools predicted upstream regulatory transcription factors and binding proteins of circPTPN22, while chromatin immunoprecipitation and ribonucleoprotein immunoprecipitation assays confirmed the binding status.
RESULTS
Upregulation of circPTPN22 in GC has been shown to inhibit autophagy and promote cell proliferation, migration, and invasion. Mechanistically, circPTPN22 directly binds to miR-6788-5p, subsequently regulating the expression of PAK1, which activates protein kinase B (Akt) and extracellular signal-regulated kinase (Erk) phosphorylation. This modulation ultimately affects autophagy levels in GC cells. Additionally, runt-related transcription factor 1 (RUNX1) negatively regulates circPTPN22 expression, while RNA-binding proteins such as FUS (fused in sarcoma) and ELAVL1 (recombinant ELAV-like protein 1) positively regulate its expression. Inhibition of the autophagy pathway can increase FUS expression, further upregulating circPTPN22 in GC cells, thereby exacerbating the progression of GC.
CONCLUSION
Under the regulation of the transcription factor RUNX1 and RNA-binding proteins FUS and ELAVL1, circPTPN22 activates the phosphorylation of Akt and Erk through the miR-6788-5p/PAK1 axis, thereby modulating autophagy in GC cells. Inhibition of autophagy increases FUS, which in turn upregulates circPTPN22, forming a positive feedback loop that ultimately accelerates the progression of GC.
Topics: Humans; Stomach Neoplasms; RNA, Circular; Autophagy; MicroRNAs; p21-Activated Kinases; Cell Proliferation; RNA-Binding Protein FUS; Cell Movement; Cell Line, Tumor; Animals; ELAV-Like Protein 1; Core Binding Factor Alpha 2 Subunit; Gene Expression Regulation, Neoplastic; Mice, Nude; Mice; Neoplasm Invasiveness; Mice, Inbred BALB C
PubMed: 38956466
DOI: 10.1186/s11658-024-00610-9 -
Acta Pharmacologica Sinica Jul 2024Abnormal accumulation of hyperphosphorylated tau protein plays a pivotal role in a collection of neurodegenerative diseases named tauopathies, including Alzheimer's...
Abnormal accumulation of hyperphosphorylated tau protein plays a pivotal role in a collection of neurodegenerative diseases named tauopathies, including Alzheimer's disease (AD). We have recently conceptualized the design of hetero-bifunctional chimeras for selectively promoting the proximity between tau and phosphatase, thus specifically facilitating tau dephosphorylation and removal. Here, we sought to optimize the construction of tau dephosphorylating-targeting chimera (DEPTAC) and obtained a new chimera D14, which had high efficiency in reducing tau phosphorylation both in cell and tauopathy mouse models, while showing limited cytotoxicity. Moreover, D14 ameliorated neurodegeneration in primary cultured hippocampal neurons treated with toxic tau-K18 fragments, and improved cognitive functions of tauopathy mice. These results suggested D14 as a cost-effective drug candidate for the treatment of tauopathies.
PubMed: 38956416
DOI: 10.1038/s41401-024-01326-4 -
Scientific Reports Jul 2024Increasing evidence has shown that many environmental and toxic factors can cause testicular damage, leading to testicular ferroptosis and subsequent male reproductive...
Increasing evidence has shown that many environmental and toxic factors can cause testicular damage, leading to testicular ferroptosis and subsequent male reproductive disorders. Melatonin is a major hormone and plays an vital role in regulating male reproduction. However, there is a lack of research on whether Mel can alleviate testicular cell ferroptosis and its specific mechanism. In this study, the results indicated that Mel could enhance the viability of swine testis cells undergoing ferroptosis, reduce LDH enzyme release, increase mitochondrial membrane potential, and affect the expression of ferroptosis biomarkers. Furthermore, we found that melatonin depended on melatonin receptor 1B to exert these functions. Detection of MMP and ferroptosis biomarker protein expression confirmed that MT2 acted through the downstream Akt signaling pathway. Moreover, inhibition of the Akt signaling pathway can eliminate the protective effect of melatonin on ferroptosis, inhibit AMPK phosphorylation, reduce the expression of mitochondrial gated channel (VDAC2/3), and affect mitochondrial DNA transcription and ATP content. These results suggest that melatonin exerts a beneficial effect on mitochondrial function to mitigate ferroptosis through the MT2/Akt signaling pathway in ST cells.
Topics: Animals; Melatonin; Male; Ferroptosis; Proto-Oncogene Proteins c-akt; Signal Transduction; Mitochondria; Swine; Testis; Receptor, Melatonin, MT2; Membrane Potential, Mitochondrial
PubMed: 38956409
DOI: 10.1038/s41598-024-65666-1 -
Communications Biology Jul 2024Gastric cancer (GC) is the 5 most prevalent cancer and the 4 primary cancer-associated mortality globally. As the first identified m6A demethylase for removing RNA...
Gastric cancer (GC) is the 5 most prevalent cancer and the 4 primary cancer-associated mortality globally. As the first identified m6A demethylase for removing RNA methylation modification, fat mass and obesity-associated protein (FTO) plays instrumental roles in cancer development. Therefore, we study the biological functions and oncogenic mechanisms of FTO in GC tumorigenesis and progression. In our study, FTO expression is obviously upregulated in GC tissues and cells. The upregulation of FTO is associated with advanced nerve invasion, tumor size, and LNM, as well as the poor prognosis in GC patients, and promoted GC cell viability, colony formation, migration and invasion. Mechanistically, FTO targeted specificity protein 1 and Aurora Kinase B, resulting in the phosphorylation of ataxia telangiectasia mutated and P38 and dephosphorylation of P53. In conclusion, the m6A demethylase FTO promotes GC tumorigenesis and progression by regulating the SP1-AURKB-ATM pathway, which may highlight the potential of FTO as a diagnostic biomarker for GC patients' therapy response and prognosis.
Topics: Humans; Alpha-Ketoglutarate-Dependent Dioxygenase FTO; Stomach Neoplasms; Cell Line, Tumor; Ataxia Telangiectasia Mutated Proteins; Sp1 Transcription Factor; Aurora Kinase B; Male; Female; Gene Expression Regulation, Neoplastic; Disease Progression; Middle Aged; Signal Transduction; Prognosis; Mice; Animals
PubMed: 38956367
DOI: 10.1038/s42003-024-06477-y