The Experts below are selected from a list of 16569 Experts worldwide ranked by ideXlab platform
Yixue Xue - One of the best experts on this subject based on the ideXlab platform.
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MiR-429 Regulated by Endothelial Monocyte Activating Polypeptide-II (EMAP-II) Influences Blood-Tumor Barrier Permeability by Inhibiting the Expressions of ZO-1, Occludin and Claudin-5
Frontiers Media S.A., 2018Co-Authors: Yixue Xue, Jian Zheng, Liangyu Chen, Xiaobai LiuAbstract:The blood-tumor barrier (BTB) hinders delivery of chemotherapeutic drugs to tumors in the brain; previous studies have shown that the BTB can be selectively opened by endothelial monocyte activating polypeptide-II (EMAP-II), but the specific mechanism involved remains elusive. In this study, we found that microRNA-429 (miR-429) expression in glioma vascular endothelial cells (GECs) was far lower than in human brain microvascular endothelial cells (ECs). miR-429 had lower expression in GECs and glioma tissues compared to ECs or normal tissues of the brain. Furthermore, miR-429 had lower expression in high grade glioma (HGG) than in low grade glioma (LGG). In in vitro BTB models, we also found that EMAP-II significantly increased BTB permeability, decreased expression of ZO-1, occludin and Claudin-5 in GECs, in a time- and dose-dependent manner. EMAP-II greatly increased miR-429 expression in GECs of the BTB models in vitro. Overexpression of miR-429 in GECs significantly decreased the transepithelial electric resistance (TEER) values in BTB models, and led to enhanced horseradish peroxidase (HRP) flux. Overexpression of miR-429 in GECs significantly decreased the expression of tight junction (TJ)-associated proteins (ZO-1, occludin and Claudin-5), and decreased the distribution continuity. Silencing of miR-429 in GECs increased the expression of TJ-associated proteins and the distribution continuity. The dual-luciferase reporter assay revealed that ZO-1 and occludin were target genes of miR-429, and we demonstrated that miR-429 overexpression markedly down-regulated protein expression of p70S6K, as well as its phosphorylation levels. The dual-luciferase reporter assay also showed that p70S6K was a target gene of miR-429; miR-429 overexpression down-regulated expression and phosphorylation levels of p70S6K, and also decreased phosphorylation levels of S6 and increased BTB permeability. Conversely, silencing of miR-429 increased the expression and phosphorylation levels of p70S6K, and increased phosphorylation levels of S6, while decreasing BTB permeability. In conclusion, the results indicated that EMAP-II caused an increase in miR-429 expression that directly targeted TJ-associated proteins, which were negatively regulated; on the other hand, miR-429 down-regulated the expression of TJ-associated proteins by targeting p70S6K, also negatively regulated. As a result, the BTB permeability increased
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long non coding rna neat1 regulates permeability of the blood tumor barrier via mir 181d 5p mediated expression changes in zo 1 occludin and Claudin 5
Biochimica et Biophysica Acta, 2017Co-Authors: Junqing Guo, Heng Cai, Yunhui Liu, Jian Zheng, Xiaobai Liu, Zhongyou Que, Wei Gong, Yana Gao, Wei Tao, Yixue XueAbstract:The blood-tumor barrier (BTB) constitutes an efficient organization of tight junctions that limits the delivery of chemotherapeutic drugs to brain tumor tissues and impacts the treatment of glioma. Long non-coding RNAs (lncRNAs) are non-protein coding RNAs regulating gene expression, some lncRNAs play a crucial role in BTB permeability. However, the function of lncRNAs in BTB permeability is still largely unclear. Here, we have identified lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1), was remarkably up-regulated in glioma endothelial cells (GECs) obtained from an in vitro BTB model. Knockdown of NEAT1 impaired the integrity and increased the permeability of the BTB, accompanied by downregulation of expression of the tight junction proteins ZO-1, occludin and Claudin-5 in GECs. Both bioinformatics data and results of luciferase reporter assays demonstrated that NEAT1 influenced BTB permeability by binding to miR-181d-5p. Knockdown of NEAT1 also down-regulated the expression of sex determining region Y-box protein 5 (SOX5), which was defined as a direct and functional downstream target of miR-181d-5p. SOX5 interacts with the promoter region of ZO-1, occludin and Claudin-5 in GECs. In conclusion, knockdown of NEAT1 increased BTB permeability by binding to miR-181d-5p and then reducing tight junction protein expression by targeting SOX5. These results suggest an important role for NEAT1 in regulating BTB permeability and provide an additional strategy for treating glioma.
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roundabout 4 regulates blood tumor barrier permeability through the modulation of zo 1 occludin and Claudin 5 expression
Journal of Neuropathology and Experimental Neurology, 2015Co-Authors: Heng Cai, Wenjing Liu, Yixue Xue, Xiuli Shang, Jing Liu, Ping Wang, Libo Liu, Yunhui LiuAbstract:The blood-tumor barrier (BTB) restricts the delivery of chemotherapeutic drug molecules to tumor tissues. We found that the endothelial cell (EC) receptor molecule Roundabout 4 (Robo4) is endogenously expressed in human brain microvascular ECs and that it is upregulated in a BTB model of glioma cocultured ECs. Knockdown of Robo4 in this BTB model increased permeability; short hairpin RNA targeting Robo4 (shRobo4) led to decreased transendothelial electric resistance values, increased BTB permeability, and downregulated expression of the EC tight junction proteins ZO-1, occludin, and Claudin-5. Roundabout 4 influenced BTB permeability via binding with its ligand, Slit2. Short hairpin RNA targeting Robo4 also increased matrix metalloproteinase-9 (MMP-9) activity and expression in glioma cocultured ECs; pretreatment with the MMP inhibitor GM6001 partially blocked the effects of shRobo4 on the transendothelial electric resistance values and ZO-1 and occludin expression. Short hairpin RNA targeting Robo4 also upregulated the phosphorylation of Src and Erk1/2; the Src inhibitor PP2 and the Erk1/2 inhibitor PD98059 blocked shRobo4-mediated alteration in ZO-1 and occludin expression. Together, our results indicate that knockdown of Robo4 increased BTB permeability by reducing EC tight junction protein expression, and that the Src-Erk1/2-MMP-9 signal pathways are involved in this process. Thus, Robo4 may represent a useful future therapeutic target for enhancing BTB permeability.
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kruppel like factor 4 regulates blood tumor barrier permeability via zo 1 occludin and Claudin 5
Journal of Cellular Physiology, 2014Co-Authors: Ping Wang, Yunhui Liu, Lini Zhao, Yixue XueAbstract:Blood-tumor barrier (BTB) constitutes an efficient organization of tight junctions which significantly reduce permeability for chemotherapy drugs. Kruppel-like factor 4 (KLF4), a member of the Kruppel-like family, has been documented in endothelial cells and may serve as an essential regulator of endothelial barrier function. However, our knowledge about the expression and function of KLF4 in the endothelial cells of BTB still remains unclear. In this study, we sought to investigate the role of KLF4 in regulation of BTB function as well as the potential molecular mechanisms. Quantitative RT-PCR, Western blot, and immunofluorescence assays demonstrated that KLF4 was down-regulated in the glioma endothelial cells (GECs) which were obtained through endothelial cells co-cultured with glioma cells. Short hairpin RNA targeting KLF4 impaired the integrity of BTB detected by trans-endothelial electric resistance assay, and meanwhile reduced the expression of ZO-1, occludin and Claudin-5, demonstrated by quantitative RT-PCR, Western blot, and immunofluorescence assays. Depletion of KLF4 increased BTB permeability to small molecules detected by permeability assays. Furthermore, luciferase assays and chromatin immunoprecipitation assays showed that KLF4 up-regulated the promoter activities and interacted with “CACCC” DNA sequence presented in the promoters of ZO-1, occludin, and Claudin-5. GATA-1, GATA-6, Sp1, and Sp3 factors participated in KLF4 regulation of promoter activities through binding to the promoters of tight junctions related proteins. Collectively, our results indicated that KLF4 is a key transcriptional regulator of BTB function by regulating expressions of tight junction related proteins, which would draw growing attention to KLF4 as a potential target for glioma therapy. J. Cell. Physiol. 229: 916–926, 2014. © 2013 Wiley Periodicals, Inc.
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kruppel like factor 4 regulates blood tumor barrier permeability via zo 1 occludin and Claudin 5
Journal of Cellular Physiology, 2014Co-Authors: Ping Wang, Yunhui Liu, Lini Zhao, Yixue XueAbstract:Blood-tumor barrier (BTB) constitutes an efficient organization of tight junctions which significantly reduce permeability for chemotherapy drugs. Kruppel-like factor 4 (KLF4), a member of the Kruppel-like family, has been documented in endothelial cells and may serve as an essential regulator of endothelial barrier function. However, our knowledge about the expression and function of KLF4 in the endothelial cells of BTB still remains unclear. In this study, we sought to investigate the role of KLF4 in regulation of BTB function as well as the potential molecular mechanisms. Quantitative RT-PCR, Western blot, and immunofluorescence assays demonstrated that KLF4 was down-regulated in the glioma endothelial cells (GECs) which were obtained through endothelial cells co-cultured with glioma cells. Short hairpin RNA targeting KLF4 impaired the integrity of BTB detected by trans-endothelial electric resistance assay, and meanwhile reduced the expression of ZO-1, occludin and Claudin-5, demonstrated by quantitative RT-PCR, Western blot, and immunofluorescence assays. Depletion of KLF4 increased BTB permeability to small molecules detected by permeability assays. Furthermore, luciferase assays and chromatin immunoprecipitation assays showed that KLF4 up-regulated the promoter activities and interacted with "CACCC" DNA sequence presented in the promoters of ZO-1, occludin, and Claudin-5. GATA-1, GATA-6, Sp1, and Sp3 factors participated in KLF4 regulation of promoter activities through binding to the promoters of tight junctions related proteins. Collectively, our results indicated that KLF4 is a key transcriptional regulator of BTB function by regulating expressions of tight junction related proteins, which would draw growing attention to KLF4 as a potential target for glioma therapy.
Yunhui Liu - One of the best experts on this subject based on the ideXlab platform.
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Mechanism of piR-DQ590027/MIR17HG regulating the permeability of glioma conditioned normal BBB
BMC, 2018Co-Authors: Xue Leng, Libo Liu, Yunhui Liu, Jian Zheng, Xiaobai Liu, Shuyuan Shen, Jiajia Chen, Lini ZhaoAbstract:Abstract Background The blood-brain barrier (BBB) strongly restricts the entry of anti-glioma drugs into tumor tissues and thus decreases chemotherapy efficacy. Malignant gliomas are highly invasive tumours that use the perivascular space for invasion and co-opt existing vessels as satellite tumor form. Because regulation of the effect of noncoding RNA on BBB function is attracting growing attention, we investigated the effects of noncoding RNA on the permeability of glioma conditioned normal BBB and the mechanism involved using PIWI-associated RNA piR-DQ590027 as a starting point. Methods The mRNA levels of MIR17HG, miR-153, miR-377, ZO-1, occludin, and Claudin-5 were determined using real-time PCR. Transient cell transfection was performed using Lipofectamine 3000 reagent. TEER and HRP flux were applied to measure the permeability of glioma conditioned normal BBB. Western blotting and immunofluorescence assays were used to measure ZO-1, occludin, and Claudin-5 levels. Reporter vector construction and a luciferase reporter assay were performed to detect the binding sites of MIR17HG and piR-DQ590027, MIR17HG and miR-153 (miR-377), and FOXR2 and miR-153 (miR-377). RNA immunoprecipitation was used to test the interaction between miR-153 (miR-377) and its target proteins. Chromatin immunoprecipitation was performed to detect the interaction between the transcription factor FOXR2 and ZO-1, occludin, and Claudin-5. Results piR-DQ590027 was expressed at low levels in glioma-conditioned ECs (GECs) of the in vitro glioma conditioned normal BBB model. Overexpression of piR-DQ590027 down-regulated the expressions of ZO-1, occludin, and Claudin-5 and increased the permeability of glioma conditioned normal BBB. MIR17HG had high expression in GECs but miR-153 and miR-377 had low expression. piR-DQ590027 bound to and negatively regulated MIR17HG. FOXR2 was a downstream target of miR-153 and miR-377; MIR17HG bound separately to miR-153 and miR-377 and negatively regulated their ability to mediate FOXR2 expression. FOXR2 associated with the promoter regions of ZO-1, occludin, and Claudin-5 in GECs to promote their transcription. Conclusion The piR-DQ590027/MIR17HG/miR-153 (miR-377)/FOXR2 pathway plays an important role in regulating glioma conditioned normal BBB permeability and provides a new target for the comprehensive treatment of glioma
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long non coding rna neat1 regulates permeability of the blood tumor barrier via mir 181d 5p mediated expression changes in zo 1 occludin and Claudin 5
Biochimica et Biophysica Acta, 2017Co-Authors: Junqing Guo, Heng Cai, Yunhui Liu, Jian Zheng, Xiaobai Liu, Zhongyou Que, Wei Gong, Yana Gao, Wei Tao, Yixue XueAbstract:The blood-tumor barrier (BTB) constitutes an efficient organization of tight junctions that limits the delivery of chemotherapeutic drugs to brain tumor tissues and impacts the treatment of glioma. Long non-coding RNAs (lncRNAs) are non-protein coding RNAs regulating gene expression, some lncRNAs play a crucial role in BTB permeability. However, the function of lncRNAs in BTB permeability is still largely unclear. Here, we have identified lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1), was remarkably up-regulated in glioma endothelial cells (GECs) obtained from an in vitro BTB model. Knockdown of NEAT1 impaired the integrity and increased the permeability of the BTB, accompanied by downregulation of expression of the tight junction proteins ZO-1, occludin and Claudin-5 in GECs. Both bioinformatics data and results of luciferase reporter assays demonstrated that NEAT1 influenced BTB permeability by binding to miR-181d-5p. Knockdown of NEAT1 also down-regulated the expression of sex determining region Y-box protein 5 (SOX5), which was defined as a direct and functional downstream target of miR-181d-5p. SOX5 interacts with the promoter region of ZO-1, occludin and Claudin-5 in GECs. In conclusion, knockdown of NEAT1 increased BTB permeability by binding to miR-181d-5p and then reducing tight junction protein expression by targeting SOX5. These results suggest an important role for NEAT1 in regulating BTB permeability and provide an additional strategy for treating glioma.
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roundabout 4 regulates blood tumor barrier permeability through the modulation of zo 1 occludin and Claudin 5 expression
Journal of Neuropathology and Experimental Neurology, 2015Co-Authors: Heng Cai, Wenjing Liu, Yixue Xue, Xiuli Shang, Jing Liu, Ping Wang, Libo Liu, Yunhui LiuAbstract:The blood-tumor barrier (BTB) restricts the delivery of chemotherapeutic drug molecules to tumor tissues. We found that the endothelial cell (EC) receptor molecule Roundabout 4 (Robo4) is endogenously expressed in human brain microvascular ECs and that it is upregulated in a BTB model of glioma cocultured ECs. Knockdown of Robo4 in this BTB model increased permeability; short hairpin RNA targeting Robo4 (shRobo4) led to decreased transendothelial electric resistance values, increased BTB permeability, and downregulated expression of the EC tight junction proteins ZO-1, occludin, and Claudin-5. Roundabout 4 influenced BTB permeability via binding with its ligand, Slit2. Short hairpin RNA targeting Robo4 also increased matrix metalloproteinase-9 (MMP-9) activity and expression in glioma cocultured ECs; pretreatment with the MMP inhibitor GM6001 partially blocked the effects of shRobo4 on the transendothelial electric resistance values and ZO-1 and occludin expression. Short hairpin RNA targeting Robo4 also upregulated the phosphorylation of Src and Erk1/2; the Src inhibitor PP2 and the Erk1/2 inhibitor PD98059 blocked shRobo4-mediated alteration in ZO-1 and occludin expression. Together, our results indicate that knockdown of Robo4 increased BTB permeability by reducing EC tight junction protein expression, and that the Src-Erk1/2-MMP-9 signal pathways are involved in this process. Thus, Robo4 may represent a useful future therapeutic target for enhancing BTB permeability.
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kruppel like factor 4 regulates blood tumor barrier permeability via zo 1 occludin and Claudin 5
Journal of Cellular Physiology, 2014Co-Authors: Ping Wang, Yunhui Liu, Lini Zhao, Yixue XueAbstract:Blood-tumor barrier (BTB) constitutes an efficient organization of tight junctions which significantly reduce permeability for chemotherapy drugs. Kruppel-like factor 4 (KLF4), a member of the Kruppel-like family, has been documented in endothelial cells and may serve as an essential regulator of endothelial barrier function. However, our knowledge about the expression and function of KLF4 in the endothelial cells of BTB still remains unclear. In this study, we sought to investigate the role of KLF4 in regulation of BTB function as well as the potential molecular mechanisms. Quantitative RT-PCR, Western blot, and immunofluorescence assays demonstrated that KLF4 was down-regulated in the glioma endothelial cells (GECs) which were obtained through endothelial cells co-cultured with glioma cells. Short hairpin RNA targeting KLF4 impaired the integrity of BTB detected by trans-endothelial electric resistance assay, and meanwhile reduced the expression of ZO-1, occludin and Claudin-5, demonstrated by quantitative RT-PCR, Western blot, and immunofluorescence assays. Depletion of KLF4 increased BTB permeability to small molecules detected by permeability assays. Furthermore, luciferase assays and chromatin immunoprecipitation assays showed that KLF4 up-regulated the promoter activities and interacted with “CACCC” DNA sequence presented in the promoters of ZO-1, occludin, and Claudin-5. GATA-1, GATA-6, Sp1, and Sp3 factors participated in KLF4 regulation of promoter activities through binding to the promoters of tight junctions related proteins. Collectively, our results indicated that KLF4 is a key transcriptional regulator of BTB function by regulating expressions of tight junction related proteins, which would draw growing attention to KLF4 as a potential target for glioma therapy. J. Cell. Physiol. 229: 916–926, 2014. © 2013 Wiley Periodicals, Inc.
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kruppel like factor 4 regulates blood tumor barrier permeability via zo 1 occludin and Claudin 5
Journal of Cellular Physiology, 2014Co-Authors: Ping Wang, Yunhui Liu, Lini Zhao, Yixue XueAbstract:Blood-tumor barrier (BTB) constitutes an efficient organization of tight junctions which significantly reduce permeability for chemotherapy drugs. Kruppel-like factor 4 (KLF4), a member of the Kruppel-like family, has been documented in endothelial cells and may serve as an essential regulator of endothelial barrier function. However, our knowledge about the expression and function of KLF4 in the endothelial cells of BTB still remains unclear. In this study, we sought to investigate the role of KLF4 in regulation of BTB function as well as the potential molecular mechanisms. Quantitative RT-PCR, Western blot, and immunofluorescence assays demonstrated that KLF4 was down-regulated in the glioma endothelial cells (GECs) which were obtained through endothelial cells co-cultured with glioma cells. Short hairpin RNA targeting KLF4 impaired the integrity of BTB detected by trans-endothelial electric resistance assay, and meanwhile reduced the expression of ZO-1, occludin and Claudin-5, demonstrated by quantitative RT-PCR, Western blot, and immunofluorescence assays. Depletion of KLF4 increased BTB permeability to small molecules detected by permeability assays. Furthermore, luciferase assays and chromatin immunoprecipitation assays showed that KLF4 up-regulated the promoter activities and interacted with "CACCC" DNA sequence presented in the promoters of ZO-1, occludin, and Claudin-5. GATA-1, GATA-6, Sp1, and Sp3 factors participated in KLF4 regulation of promoter activities through binding to the promoters of tight junctions related proteins. Collectively, our results indicated that KLF4 is a key transcriptional regulator of BTB function by regulating expressions of tight junction related proteins, which would draw growing attention to KLF4 as a potential target for glioma therapy.
Michal Toborek - One of the best experts on this subject based on the ideXlab platform.
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signaling mechanisms of hiv 1 tat induced alterations of Claudin 5 expression in brain endothelial cells
Journal of Cerebral Blood Flow and Metabolism, 2005Co-Authors: Ibolya E Andras, Maria A Deli, Bernhard Hennig, Avindra Nath, Jing Tian, Michal ToborekAbstract:Exposure of brain microvascular endothelial cells (BMEC) to human immunodeficiency virus-1 (HIV-1) Tat protein can decrease expression and change distribution of tight junction proteins, including Claudin-5. Owing to the importance of Claudin-5 in maintaining the blood-brain barrier (BBB) integrity, the present study focused on the regulatory mechanisms of Tat-induced alterations of Claudin-5 mRNA and protein levels. Real-time reverse-transcription-polymerase chain reaction revealed that Claudin-5 mRNA was markedly diminished in BMEC exposed to Tat. However, U0126 (an inhibitor of mitogen-activated protein kinase kinase1/2, MEK1/2) protected against this effect. In addition, inhibition of the vascular endothelial growth factor receptor type 2 (VEGFR-2) by SU1498, phosphatidylinositol-3 kinase (PI-3 K) by LY294002, nuclear factor-kappaB (NF-kappaB) by peptide SN50, and intracellular calcium by BAPTA/AM partially prevented Tat-mediated alterations in Claudin-5 protein levels and immunoreactivity patterns. In contrast, inhibition of protein kinase C did not affect Claudin-5 expression in Tat-treated cells. The present findings indicate that activation of VEGFR-2 and multiple redox-regulated signal transduction pathways are involved in Tat-induced alterations of Claudin-5 expression. Because Claudins constitute the major backbone of tight junctions, the present data are relevant to the disturbances of the BBB in the course of HIV-1 infection.
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hiv 1 tat protein alters tight junction protein expression and distribution in cultured brain endothelial cells
Journal of Neuroscience Research, 2003Co-Authors: Ibolya E Andras, Maria A Deli, Hong Pu, Bernhard Hennig, Avindra Nath, Michal ToborekAbstract:Disruption of the blood-brain barrier (BBB) is widely believed to be the main route of human immunodeficiency virus (HIV) entry into the central nervous system (CNS). Although mechanisms of this process are not fully understood, alterations of tight junction protein expression can contribute, at least in part, to this phenomenon. Tight junctions are critical structural and functional elements of cerebral microvascular endothelial cells and the BBB. The aim of the present study was to examine the effects of HIV-1 Tat protein on expression of tight junction proteins. Primary cultures of brain microvascular endothelial cells (BMEC) were employed in these experiments. A 24-hr exposure of BMEC to Tat 1 - 7 2 resulted in a decrease of Claudin-1, Claudin-5, and zonula occludens (ZO)-2 expression, whereas total levels of occludin and ZO-1 remained unchanged. In addition, a short (3-hr) exposure of BMEC to Tat 1 - 7 2 induced cellular redistribution of Claudin-5 immunoreactivity. Tat 1 - 7 2 -induced alterations of Claudin-5 expression also were confirmed in vivo where Tat 1 - 7 2 was injected into the right hippocampus of mice. These findings indicate that HIV-1 Tat protein can markedly affect expression and distribution of specific tight junction proteins in brain endothelium. Alterations of only distinct tight junction proteins suggest a finely tuned effect of Tat 1 - 7 2 on the BBB. Because tight junction proteins are critical for the barrier function of the BBB, such alterations can lead to disturbances of the BBB integrity and contribute to HIV trafficking into the brain.
Ping Wang - One of the best experts on this subject based on the ideXlab platform.
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roundabout 4 regulates blood tumor barrier permeability through the modulation of zo 1 occludin and Claudin 5 expression
Journal of Neuropathology and Experimental Neurology, 2015Co-Authors: Heng Cai, Wenjing Liu, Yixue Xue, Xiuli Shang, Jing Liu, Ping Wang, Libo Liu, Yunhui LiuAbstract:The blood-tumor barrier (BTB) restricts the delivery of chemotherapeutic drug molecules to tumor tissues. We found that the endothelial cell (EC) receptor molecule Roundabout 4 (Robo4) is endogenously expressed in human brain microvascular ECs and that it is upregulated in a BTB model of glioma cocultured ECs. Knockdown of Robo4 in this BTB model increased permeability; short hairpin RNA targeting Robo4 (shRobo4) led to decreased transendothelial electric resistance values, increased BTB permeability, and downregulated expression of the EC tight junction proteins ZO-1, occludin, and Claudin-5. Roundabout 4 influenced BTB permeability via binding with its ligand, Slit2. Short hairpin RNA targeting Robo4 also increased matrix metalloproteinase-9 (MMP-9) activity and expression in glioma cocultured ECs; pretreatment with the MMP inhibitor GM6001 partially blocked the effects of shRobo4 on the transendothelial electric resistance values and ZO-1 and occludin expression. Short hairpin RNA targeting Robo4 also upregulated the phosphorylation of Src and Erk1/2; the Src inhibitor PP2 and the Erk1/2 inhibitor PD98059 blocked shRobo4-mediated alteration in ZO-1 and occludin expression. Together, our results indicate that knockdown of Robo4 increased BTB permeability by reducing EC tight junction protein expression, and that the Src-Erk1/2-MMP-9 signal pathways are involved in this process. Thus, Robo4 may represent a useful future therapeutic target for enhancing BTB permeability.
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kruppel like factor 4 regulates blood tumor barrier permeability via zo 1 occludin and Claudin 5
Journal of Cellular Physiology, 2014Co-Authors: Ping Wang, Yunhui Liu, Lini Zhao, Yixue XueAbstract:Blood-tumor barrier (BTB) constitutes an efficient organization of tight junctions which significantly reduce permeability for chemotherapy drugs. Kruppel-like factor 4 (KLF4), a member of the Kruppel-like family, has been documented in endothelial cells and may serve as an essential regulator of endothelial barrier function. However, our knowledge about the expression and function of KLF4 in the endothelial cells of BTB still remains unclear. In this study, we sought to investigate the role of KLF4 in regulation of BTB function as well as the potential molecular mechanisms. Quantitative RT-PCR, Western blot, and immunofluorescence assays demonstrated that KLF4 was down-regulated in the glioma endothelial cells (GECs) which were obtained through endothelial cells co-cultured with glioma cells. Short hairpin RNA targeting KLF4 impaired the integrity of BTB detected by trans-endothelial electric resistance assay, and meanwhile reduced the expression of ZO-1, occludin and Claudin-5, demonstrated by quantitative RT-PCR, Western blot, and immunofluorescence assays. Depletion of KLF4 increased BTB permeability to small molecules detected by permeability assays. Furthermore, luciferase assays and chromatin immunoprecipitation assays showed that KLF4 up-regulated the promoter activities and interacted with “CACCC” DNA sequence presented in the promoters of ZO-1, occludin, and Claudin-5. GATA-1, GATA-6, Sp1, and Sp3 factors participated in KLF4 regulation of promoter activities through binding to the promoters of tight junctions related proteins. Collectively, our results indicated that KLF4 is a key transcriptional regulator of BTB function by regulating expressions of tight junction related proteins, which would draw growing attention to KLF4 as a potential target for glioma therapy. J. Cell. Physiol. 229: 916–926, 2014. © 2013 Wiley Periodicals, Inc.
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kruppel like factor 4 regulates blood tumor barrier permeability via zo 1 occludin and Claudin 5
Journal of Cellular Physiology, 2014Co-Authors: Ping Wang, Yunhui Liu, Lini Zhao, Yixue XueAbstract:Blood-tumor barrier (BTB) constitutes an efficient organization of tight junctions which significantly reduce permeability for chemotherapy drugs. Kruppel-like factor 4 (KLF4), a member of the Kruppel-like family, has been documented in endothelial cells and may serve as an essential regulator of endothelial barrier function. However, our knowledge about the expression and function of KLF4 in the endothelial cells of BTB still remains unclear. In this study, we sought to investigate the role of KLF4 in regulation of BTB function as well as the potential molecular mechanisms. Quantitative RT-PCR, Western blot, and immunofluorescence assays demonstrated that KLF4 was down-regulated in the glioma endothelial cells (GECs) which were obtained through endothelial cells co-cultured with glioma cells. Short hairpin RNA targeting KLF4 impaired the integrity of BTB detected by trans-endothelial electric resistance assay, and meanwhile reduced the expression of ZO-1, occludin and Claudin-5, demonstrated by quantitative RT-PCR, Western blot, and immunofluorescence assays. Depletion of KLF4 increased BTB permeability to small molecules detected by permeability assays. Furthermore, luciferase assays and chromatin immunoprecipitation assays showed that KLF4 up-regulated the promoter activities and interacted with "CACCC" DNA sequence presented in the promoters of ZO-1, occludin, and Claudin-5. GATA-1, GATA-6, Sp1, and Sp3 factors participated in KLF4 regulation of promoter activities through binding to the promoters of tight junctions related proteins. Collectively, our results indicated that KLF4 is a key transcriptional regulator of BTB function by regulating expressions of tight junction related proteins, which would draw growing attention to KLF4 as a potential target for glioma therapy.
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specific role of tight junction proteins Claudin 5 occludin and zo 1 of the blood brain barrier in a focal cerebral ischemic insult
Journal of Molecular Neuroscience, 2011Co-Authors: Haixia Jiao, Yunhui Liu, Ping Wang, Zhenhua Wang, Yixue XueAbstract:Blood–brain barrier (BBB) leakage plays a key role in cerebral ischemia–reperfusion injury. It is quite necessary to further explore the characteristic and mechanism of BBB leakage during stroke. We induced a focal cerebral ischemia model by transient middle cerebral artery occlusion in male rats for defining the time course of BBB permeability within 120 h following reperfusion and evaluate the specific role of tight junction (TJ) associated proteins Claudin-5, occludin, and ZO-1 as well as protein kinase C delta (PKCδ) pathway in BBB leakage induced by reperfusion injury. We verified a bimodal increase in the permeability of the BBB following focal ischemia by Evans blue assay. Two peaks of BBB permeability appeared at 3 h and 72 h of reperfusion after 2 h focal ischemia, respectively. The leak at the endothelial cell was represented at the level of transmission electron microscopy. TTC staining results showed increased infarct size with time after cerebral ischemia reperfusion. The mRNA and protein expression levels of these three TJ associated proteins were significantly decreased compared with the sham-operated group within 120 h of reperfusion, corresponding to the time-dependent change of the biphasic pattern in BBB leakage. The redistribution of Claudin-5, occludin, and ZO-1 in ischemia brain microvascular endothelial cells was observed at the same time points. In addition, Western blot assay revealed PKCδ level was also significantly increased in a similar biphasic pattern to above results within 120 h after cerebral ischemia–reperfusion. This study demonstrates the timing of TJ associated proteins Claudin-5, occludin, and ZO-1 in light of BBB permeability associated with cerebral ischemia reperfusion, and suggests PKCδ pathway may participate in TJ barrier open and BBB leakage during reperfusion injury in a time-dependent manner.
Yuri Persidsky - One of the best experts on this subject based on the ideXlab platform.
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inhibition of glycogen synthase kinase 3β promotes tight junction stability in brain endothelial cells by half life extension of occludin and Claudin 5
PLOS ONE, 2013Co-Authors: Servio H Ramirez, Shongshan Fan, Holly Dykstra, Slava Rom, Aaron J Mercer, Nancy L Reichenbach, Larisa Gofman, Yuri PersidskyAbstract:Neuroinflammatory conditions often involve dysfunction of the Blood-Brain Barrier (BBB). Therefore, identifying molecular targets that can maintain barrier fidelity is of clinical importance. We have previously reported on the anti-inflammatory effects that glycogen synthase kinase 3β (GSK3β) inhibition has on primary human brain endothelial cells. Here we show that GSK3β inhibitors also promote barrier tightness by affecting tight junction (TJ) protein stability. Transendothelial electrical resistance (TEER) was used to evaluate barrier integrity with both pharmacological inhibitors and mutants of GSK3β. Inhibition of GSK3β produced a gradual and sustained increase in TEER (as much as 22% over baseline). Analysis of subcellular membrane fractions revealed an increase in the amount of essential tight junction proteins, occludin and Claudin-5, but not Claudin-3. This phenomenon was attributed to a decrease in TJ protein turnover and not transcriptional regulation. Using a novel cell-based assay, inactivation of GSK3β significantly increased the half-life of occludin and Claudin-5 by 32% and 43%, respectively. A correlation was also established between the enhanced association of β-catenin with ZO-1 as a function of GSK3β inhibition. Collectively, our findings suggest the possibility of using GSK3β inhibitors as a means to extend the half-life of key tight junction proteins to promote re-sealing of the BBB during neuroinflammation.
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stat1 signaling modulates hiv 1 induced inflammatory responses and leukocyte transmigration across the blood brain barrier
Blood, 2008Co-Authors: Anathbandhu Chaudhuri, Yuri Persidsky, Bo Yang, Howard E Gendelman, Georgette D KanmogneAbstract:The relationship among neuroinflammation, blood-brain barrier (BBB) dysfunction, and progressive HIV-1 infection as they affect the onset and development of neuroAIDS is incompletely understood. One possible link is signal transducers and activators of transcription (STATs) pathways. These respond to proinflammatory and regulatory factors and could affect neuroinflammatory responses induced from infected cells and disease-affected brain tissue. Our previous works demonstrated that HIV-1 activates pro-inflammatory and interferon-alpha–inducible genes in human brain microvascular endothelial cells (HBMECs) and that these genes are linked to the Janus kinase (JAK)/STAT pathway. We now demonstrate that HIV-1 activates STAT1, induces IL-6 expression, and diminishes expression of Claudin-5, ZO-1, and ZO-2 in HBMECs. The STAT1 inhibitor, fludarabine, blocked HIV-1–induced IL-6, diminished HIV-1–induced Claudin-5 and ZO-1 down-regulation, and blocked HIV-1– and IL-6–induced monocyte migration across a BBB model. Enhanced expression and activation of STAT1 and decreased Claudin-5 were observed in microvessels from autopsied brains of patients with HIV-1–associated dementia. These data support the notion that STAT1 plays an integral role in HIV-1–induced BBB damage and is relevant to viral neuropathogenesis. Inhibition of STAT1 activation could provide a unique therapeutic strategy to attenuate HIV-1–induced BBB compromise and as such improve clinical outcomes.
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ethanol induced activation of myosin light chain kinase leads to dysfunction of tight junctions and blood brain barrier compromise
Alcoholism: Clinical and Experimental Research, 2005Co-Authors: James Haorah, David Heilman, Bryan Knipe, Jesse Chrastil, Jessica Leibhart, Anuja Ghorpade, Donald W Miller, Yuri PersidskyAbstract:Background: Brain endothelial cells form the blood-brain barrier (BBB) that regulates solute and macromolecule flux in and out of the brain, leukocyte migration, and maintains the homeostasis of the central nervous system. BBB dysfunction is associated with disruption of tight junctions (TJ) in the brain endothelium. We propose that alcohol abuse may impair BBB permeability through TJ modification. Methods: Primary cultured bovine brain microvascular endothelial cells (BBMEC) were treated with 50 mM ethanol (EtOH), and monolayer tightness was assessed by measurement of transendothelial electrical resistance (TEER). Changes in TEER were correlated with alterations in TJ protein distribution occludin, zonula occludens-1 (ZO-1), Claudin-5 using immunofluorescence (IF). Expression of myosin light chain (MLC) kinase (MLCK), ZO-1, Claudin-5, and phosphorylated MLC, occludin and Claudin-5 were determined by immunoprecipitation and Western blot. EtOH-induced changes in monocyte migration across in vitro BBB constructs were also examined. Results: EtOH induced a decrease in TEER of BBMEC monolayers that was reversed by EtOH withdrawal. Treatment of BBMEC with EtOH or its metabolite, acetaldehyde, prior to monocyte application resulted in a 2-fold increase in monocyte migration across the BBB. IF demonstrated decrease in Claudin-5 staining, occludin translocation from cell borders to cytoplasm and gap formation in EtOH-treated BBMEC monolayer. These changes paralleled significant increase in phosphorylation of MLC, occludin and Claudin-5. EtOH-treated BBMEC showed reduction of total occludin and Claudin-5 without changes in ZO-1 or MLC. TEER decrease, changes in occludin/Claudin staining, increase in MLC, occludin and Claudin-5 phosphorylation and enhanced monocyte migration across the BBB were all reversed by inhibition of MLCK. Inhibition of EtOH metabolism in BBMEC also reversed these events. Conclusion: These results suggest that EtOH activates MLCK leading to phosphorylation of MLC, occludin and Claudin-5. Cytoskeletal alterations (MLC) and TJ changes (occludin and Claudin-5 phosphorylation) result in BBB impairment (decrease in TEER). TJ compromise is associated with increased monocyte migration across the BBB.