Claudin‐5 regulates blood‐brain barrier permeability by modifying brain microvascular endothelial cell proliferation, migration, and adhesion to prevent lung cancer metastasis

Ma, Shunchang; Li, Qi; Peng, Jiayi; Zhouwen, Jian-Long; Diao, Jinfu; Niu, Jianxing; Wang, Xi; Guan, Xiudong; Jia, Wang; Jiang, Wen Guo

doi:10.1111/cns.12764

Cited by 87 publications

(66 citation statements)

References 40 publications

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“…Recently, it was shown that the overexpression of claudin-5 in hCMEC/ D3 cells led to a reduction in the permeation of A549 cancer cells by approximately two-thirds, but did not lead to a noticeable improvement in electrical impedance. This suggested that additional mechanisms were missing to functionally incorporate claudin-5 at the tight junctions in the cell membrane [27]. Single cell RNA-seq data of mouse brain derived cells from the Betsholtz group [46] showed that fibroblast-like cells and not brain capillary endothelial cells expressed claudin-1.…”

Section: Discussionmentioning

confidence: 99%

“…In case of the ABC transporters the highest abundant was ABCC1 (478) followed by ABCC4 (299), ABCB1 (106), ABCC3 (93), ABCC5 (92), ABCG2 (29) and ABCC2 (9). Quite high and moderate expression was found for JAM3 (320), ZO-1 (315), CDH5 (294) and Occludin (145), whereas low expression was found for Tricellulin (27) and JAM-2 (0.23).…”

Section: Human Brain Endothelial Hcmec/d3 Cells Express Several Tightmentioning

confidence: 99%

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The pivotal role of micro-environmental cells in a human blood–brain barrier in vitro model of cerebral ischemia: functional and transcriptomic analysis

Gerhartl

Pracser

Vladetic

et al. 2020

Fluids Barriers CNS

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Background: The blood-brain barrier (BBB) is altered in several diseases of the central nervous system. For example, the breakdown of the BBB during cerebral ischemia in stroke or traumatic brain injury is a hallmark of the diseases' progression. This functional damage is one key event which is attempted to be mimicked in in vitro models. Recent studies showed the pivotal role of micro-environmental cells such as astrocytes for this barrier damage in mouse stroke in vitro models. The aim of this study was to evaluate the role of micro-environmental cells for the functional, paracellular breakdown in a human BBB cerebral ischemia in vitro model accompanied by a transcriptional analysis. Methods: Transwell models with human brain endothelial cell line hCMEC/D3 in mono-culture or co-culture with human primary astrocytes and pericytes or rat glioma cell line C6 were subjected to oxygen/glucose deprivation (OGD). Changes of transendothelial electrical resistance (TEER) and FITC-dextran 4000 permeability were recorded as measures for paracellular tightness. In addition, qPCR and high-throughput qPCR Barrier chips were applied to investigate the changes of the mRNA expression of 38 relevant, expressed barrier targets (tight junctions, ABC-transporters) by different treatments. Results: In contrast to the mono-culture, the co-cultivation with human primary astrocytes/pericytes or glioma C6 cells resulted in a significantly increased paracellular permeability after 5 h OGD. This indicated the pivotal role of micro-environmental cells for BBB breakdown in the human model. Hierarchical cluster analysis of qPCR data revealed differently, but also commonly regulated clustered targets dependent on medium exchange, serum reduction, hydrocortisone addition and co-cultivations. Conclusions: The co-cultivation with micro-environmental cells is necessary to achieve a functional breakdown of the BBB in the cerebral ischemia model within an in vivo relevant time window. Comprehensive studies by qPCR revealed that distinct expression clusters of barrier markers exist and that these are regulated by different treatments (even by growth medium change) indicating that controls for single cell culture manipulation steps are crucial to understand the observed effects properly.

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Section: Discussionmentioning

confidence: 99%

Section: Human Brain Endothelial Hcmec/d3 Cells Express Several Tightmentioning

confidence: 99%

The pivotal role of micro-environmental cells in a human blood–brain barrier in vitro model of cerebral ischemia: functional and transcriptomic analysis

Gerhartl

Pracser

Vladetic

et al. 2020

Fluids Barriers CNS

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“…Jia reported that down-regulating CLDN5 increased tumor invasion and potential metastatic abilities [32]. Ma discovered CLDN5 was closely related to brain metastases from lung cancer [33]. In 2019, Jia indicated that high-dose bevacizumab likely increased lung tumor invasion and potential metastatic abilities through down-regulating CLDN5 [34].Moreover, CLDN5 showed a close relationship with mental illness, such as depression [35], schizophrenia [36], brain edema following fatal heat stroke [37] and tumor brain metastasis [38].…”

Section: Discussionmentioning

confidence: 99%

Genes co-related with poor prognosis of patients with lung cancer via bioinformatical approaches

Shi¹,

Han²,

Wang³

et al. 2020

Preprint

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Lung cancer is one of the most common malignant tumors with high mortality worldwide. Recently, researchers reported that molecular markers on lung cancer could be used as diagnostic and prognostic targets. However, these molecules were not ideal in specificity and high selectivity.Therefore, exploring more reliable biomarkers to improve the prognosis and clarify the underlying mechanism is urgently needed both for clinic and basic research. This study aimed to identify significant genes with poor prediction for lung cancer and their underlying mechanisms. Firstly, we used gene expression datasets available from GEO (Gene Expression Omnibus) database. There were 109 lung cancer samples and 27 normal samples in the selected datasets. First, DEGs (Different Expressed Gene set) of lung cancer and normal lung samples were screen out with GEO2R tool, and we displayed them by Venn diagram software and Heatmap. Secondly, we used DAVID (Database for Annotation, Visualization and Integrated Discovery) to analyze KEGG (Kyoto Encyclopedia of Gene and Genome) pathway and GO (Gene Ontology). Third, PPI (Protein-Protein Interaction) of these DEGs was conducted by Cytoscape with STRING (Search Tool for the Retrieval of Interacting Genes). Our results showed that the expression trends of 21 up-regulated genes and 116 down-regulated were similar in selected three datasets. Analyzed by MCODE (Molecular Complex Detection) plug-in, 11 up-regulated and 16 down-regulated genes were selected. To further verify gene expression differences, GEPIA (Gene Expression Profiling Interactive Analysis) was implemented and we found 26 of 27 genes were found differently expressed in lung cancer compared with normal lung tissues. Furthermore, Kaplan-Meier analysis was used and we found 23 of 26 genes for overall survival indicated much less survival time. At last, three genes, CDH5, CLDN5, PECAM1, were found to be significantly decreased in lung cancer tissue proved through re-analysis of DAVID, which mainly co-related with leukocyte transendothelial migration. In conclusion, three significant down-regulated deferentially expressed genes with poor prognosis on lung cancer were identified basing on integrated bioinformatical methods.These down-expressed genes may be as a potential prognosis targets for patients with lung cancer.

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“…Cell viability was evaluated via cell proliferation assay kit as described in the specifications (WI, USA). For gene knockdown, negative control small interfering RNA (siRNA) (5′-UUCUCCGA ACGUGUCACGUTT-3′) and Tau siRNA (5′-CCGCCAGG AGUUCGAAGUGAU-3′), Claudin-1 siRNA (5′-GCAUG GUAUGGCAAUAGAA-3′), Claudin-5 siRNA (5′-CCAAC AUUGUCGUCCGCGATT-3′), Claudin-7 siRNA (5′-AUUA GGGCUCGAGUGGCCUGCAAGG-3′), and α-tubulin siRNA (5′-AGAUGUCAAUGCUGCCAUU-3′) sequences were designed in reference to the literatures [37][38][39][40][41] . Cells were plated in six-well plates at a density of 1 × 10 6 per well and cultured for 24 h; the medium was replaced with fresh medium containing 10% FBS when cells reach~70% confluency.…”

Section: Cell Culture Viability Assay and Transfectionmentioning

confidence: 99%

Sulforaphane metabolites inhibit migration and invasion via microtubule-mediated Claudins dysfunction or inhibition of autolysosome formation in human non-small cell lung cancer cells

Zheng

Lin

et al. 2019

Cell Death Dis

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Both sulforaphane-cysteine (SFN-Cys) and sulforaphane-N-acetyl-L-cysteine (SFN-NAC) inhibited cancer migration and invasion, but the underlying mechanisms were not clear. Here we uncovered via tissue microarray assay that high expression of invasion-associated Claudin-5 was correlated to malignant grades in human non-small cell lung cancer (NSCLC) cells. Further, SFN-Cys (10 µM) induced the accumulated phosphorylation of ERK1/2, leading to downregulation of Claudin-5 and upregulation of Claudin-7, and the decrease of Claudin-1 in SK-1 cells and increase of Claudin-1 in A549 cells; knockdown of Claudin-5 significantly reduced invasion, whereas knockdown of Claudin-7 increased invasion; knockdown of Claudin-1 reduced invasion in SK-1 cells, whereas it increased invasion in A549 cells, indicating that SFN-Cys regulated Claudins and inhibited invasion depending on Claudin isotypes and cell types. Furthermore, immunofluorescence staining showed that SFN-Cys triggered microtubule disruption and knockdown of α-tubulin downregulated Claudin-1, 5, and 7, and inhibited migration and invasion, indicating that microtubule disruption contributed to invasive inhibition. Co-immunoprecipitation and confocal microscopy observation showed that SFN-Cys lowered the interaction between α-tubulin and Claudin-1 or 5, or 7. Meanwhile, Western blotting and immunofluorescence staining showed that SFN-NAC (15 µM) downregulated α-tubulin resulting in microtubule disruption; knockdown of α-tubulin increased SFN-NAC-induced LC3 II accumulation in SK-1 cells. Combined with the inhibitor of autolysosome formation, Bafilomycin A1 (100 nM), SFN-NAC inhibited invasion via accumulating LC3 II and blocking formation of autolysosome. Further, SFN-NAC upregulated microtubule-stabilizing protein Tau; knockdown of Tau reduced LC3 II/LC3 I inhibiting migration and invasion. These results indicated that SFN-Cys inhibited invasion via microtubule-mediated Claudins dysfunction, but SFN-NAC inhibited invasion via microtubule-mediated inhibition of autolysosome formation in human NSCLC cells.

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Claudin‐5 regulates blood‐brain barrier permeability by modifying brain microvascular endothelial cell proliferation, migration, and adhesion to prevent lung cancer metastasis

Cited by 87 publications

References 40 publications

The pivotal role of micro-environmental cells in a human blood–brain barrier in vitro model of cerebral ischemia: functional and transcriptomic analysis

The pivotal role of micro-environmental cells in a human blood–brain barrier in vitro model of cerebral ischemia: functional and transcriptomic analysis

Genes co-related with poor prognosis of patients with lung cancer via bioinformatical approaches

Sulforaphane metabolites inhibit migration and invasion via microtubule-mediated Claudins dysfunction or inhibition of autolysosome formation in human non-small cell lung cancer cells

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