Search for author "Zhaohua Cai"

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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 1. a, Hypoxia/reoxygenation increases the expression of FKN protein in ECs. Cultured HCAECs and HUVECs were exposed to hypoxia for 2 hours and...Show More

  Figure 1. a, Hypoxia/reoxygenation increases the expression of FKN protein in ECs. Cultured HCAECs and HUVECs were exposed to hypoxia for 2 hours and reoxygenation for up to 120 minutes. Western blots showed an increase in FKN protein in EC lysates beginning within 10 minutes of reoxygenation and continuing for at least 120 minutes. HUVECs were seeded into 12-well plates and grown to 100% confluence (lower left). For each time point of normoxia or hypoxia/reoxygenation, medium was harvested from 2 wells, pooled (600 μL), concentrated to ≈30 μL, and completely loaded on the gel in the appropriate lane. FKN in the media, representing the lower-molecular-weight s-FKN, increased within 15 minutes of reoxygenation and peaked at 30 minutes. b, s-FKN increases ICAM-1 mRNA in ECs. HUVECs were exposed to s-FKN for the indicated doses and times. Cells were harvested, total RNA was isolated, and real-time RT-PCR was performed for quantitation of ICAM-1 mRNA. ICAM-1 mRNA increased in a dose-dependent (top) and time-dependent (bottom) fashion.Show Less
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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 2. s-FKN upregulates ICAM-1 protein in the intact mouse heart. Hearts were perfused with or without 50 ng/mL mouse s-FKN for 3 hours. a, Wester...Show More

  Figure 2. s-FKN upregulates ICAM-1 protein in the intact mouse heart. Hearts were perfused with or without 50 ng/mL mouse s-FKN for 3 hours. a, Western blots were done with goat anti-mouse ICAM-1 antibody. b and c, Immunohistology shows that ICAM-1 is upregulated in capillary endothelium (brown stain) following perfusion with s-FKN compared with perfusion with control buffer. Image magnification for b and c, ×160. IB indicates immunoblotting.Show Less
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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 3. Vascular ECs express the FKN receptor CX3CR1. a, The cellular lysates from cultured HCAECs and HUVECs and fresh human leukocytes were used f...Show More

  Figure 3. Vascular ECs express the FKN receptor CX3CR1. a, The cellular lysates from cultured HCAECs and HUVECs and fresh human leukocytes were used for Western blots, using anti-human FKN and CX3CR1 antibodies. Both types of ECs (left 2 lanes) express both FKN and CX3CR1. Macrophages and neutrophils express CX3CR1 but not FKN itself. CX3CR1 in ECs has two 50-kDa bands as well as a 40-kDa band. Macrophages (MΦ) (fourth lane) express only a 40-kDa CX3CR1 protein, and neutrophils (PMNs) (right lane) also express it but in lesser amounts. b, RT-PCR assay for CX3CR1 mRNA. A pair of primers from the V28 cDNA sequence–spanning exons 2 and 4 was used to amplify reverse transcript products from total RNA from ECs and leukocytes. The RT-PCR products were run on 5% polyacrylamide gel and stained with ethidium bromide. ECs and leukocytes both express CX3CR1 mRNA. However, PMNs express less than ECs and MΦ. c, Confocal microscopic images of an HUVEC. Cultured HUVECs were fixed with methanol, blocked with goat serum, and incubated with anti-human CX3CR1 antibody. After washing, cells were incubated with secondary antibody labeled with green fluorescence. Top, Confocal scan sections of a single HUVEC. Bottom, Image of a single-scan section (2.24 μm) through the cell. The green staining, representing CX3CR1, is located primarily on the cell membrane and partially extends to the cytoplasm. The nucleus is stained blue with DAPI (2′,6′-diamidino-2-phenylindole).Show Less
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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 4. FKN activates ICAM-1 in ECs through its receptor, CX3CR1. a, HUVECs were transfected with control or CX3CR1 siRNA. The inset shows that siRN...Show More

  Figure 4. FKN activates ICAM-1 in ECs through its receptor, CX3CR1. a, HUVECs were transfected with control or CX3CR1 siRNA. The inset shows that siRNA for CX3CR1 knocks down the receptor protein compared with control. The transfected cells were exposed to s-FKN or control buffer for 2 hours, and ICAM-1 mRNA was measured by real-time RT-PCR. Cells transfected with control siRNA showed increased ICAM-1 mRNA expression following s-FKN stimulation (P<0.01), whereas cells transfected with CX3CR1 siRNA did not (P>0.5). b, HUVECs were transfected with siRNAs and exposed to 10 nmol/L s-FKN for 3 hours. ICAM-1 protein expression was determined by Western blotting. ICAM-1 protein increased during s-FKN stimulation in the control group but not in the CX3CR1 knockdown group.Show Less
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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 5. FKN increases adhesion of PMNs to HUVECs, and the increased adhesion is blocked by anti–ICAM-1 antibody. a, Fluorescein isothiocyanate (FITC...Show More

  Figure 5. FKN increases adhesion of PMNs to HUVECs, and the increased adhesion is blocked by anti–ICAM-1 antibody. a, Fluorescein isothiocyanate (FITC) and myeloperoxidase (MPO) activities, representing adherent PMNs, increased significantly in control cells during s-FKN simulation (**P<0.001) but not in ECs with CX3CR1 knockdown. b, ICAM-1 antibody blocked PMN adhesion to HUVECs during s-FKN stimulation, indicating that the increase in PMN adhesion induced by s-FKN is ICAM-1 dependent.Show Less
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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 6. FKN, acting through CX3CR1, activates the Jak2–Stat5 pathway. a, Western blots for Jak2 and Stat5 protein. HUVECs were transfected with cont...Show More

  Figure 6. FKN, acting through CX3CR1, activates the Jak2–Stat5 pathway. a, Western blots for Jak2 and Stat5 protein. HUVECs were transfected with control or CX3CR1 siRNA or loaded with an inhibitor of GPCRs (PTX) and then exposed to s-FKN for 7, 15, or 30 minutes. In control cells, Jak2 phosphorylation occurred within 7 minutes of s-FKN stimulation and quickly decreased by 15 minutes. Tyrosine Stat5 phosphorylation increased shortly following Jak2 phosphorylation and continued for at least 30 minutes. In contrast, in cells transfected with CX3CR1 siRNA or loaded with PTX, phosphorylation of Jak2 or Stat5 did not occur.Show Less
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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 7. a, Immunoprecipitation assay for Stat5 isoforms. HUVECs were exposed to 10 nmol/L s-FKN for 10 minutes, and the cells were lysed in radioimm...Show More

  Figure 7. a, Immunoprecipitation assay for Stat5 isoforms. HUVECs were exposed to 10 nmol/L s-FKN for 10 minutes, and the cells were lysed in radioimmunoprecipitation assay buffer.17 Anti-Stat5α and -Stat5β antibodies were then added, and the tubes were rotated overnight at 4°C. The complexes were pulled down with protein A agarose (Invitrogen) and run on NuPAGE gel for immunoblotting using anti–phosphorylated tyrosine Stat5 and anti-Stat5 antibodies. IP indicates immunoprecipitation. The Stat5 that became phosphorylated was the α isoform. b, Electrophoretic mobility-shift assay and competition assays show that Stat5α binds to the GAS element in the ICAM-1 promoter. ECs were exposed to 10 nmol/L s-FKN or control (C) medium for 10 minutes, and the cells were harvested to isolate nuclear protein (NE). For competition assays, unlabeled GAS, anti-Stat5 antibody, or anti-Stat5α or -β antibodies were used.Show Less
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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 8. Stat5 upregulates ICAM-1 in ECs. Double-stranded Stat5 siRNA (siSt5) and control siRNA (siC) were transfected into HUVECs. After 60 hours of...Show More

  Figure 8. Stat5 upregulates ICAM-1 in ECs. Double-stranded Stat5 siRNA (siSt5) and control siRNA (siC) were transfected into HUVECs. After 60 hours of transfection, the cells were exposed to 10 nmol/L s-FKN for 3 hours and harvested for Western blotting. After blotting with anti–ICAM-1 antibody (immunoblot: ICAM-1), the membrane was stripped and reblotted with anti-Stat5 antibody (immunoblot: Stat5); the house-keeping protein GAPDH was blotted with anti-GAPDH antibody to demonstrate similar amounts of loaded proteins. Compared with control siRNA cells, ICAM-1 protein expression after Stat5 knockdown was constitutively reduced. s-FKN stimulation resulted in an increase in ICAM-1 protein in control siRNA cells but not in double-stranded Stat5 siRNA cells.Show Less
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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 1. a, Hypoxia/reoxygenation increases the expression of FKN protein in ECs. Cultured HCAECs and HUVECs were exposed to hypoxia for 2 hours and...Show More

  Figure 1. a, Hypoxia/reoxygenation increases the expression of FKN protein in ECs. Cultured HCAECs and HUVECs were exposed to hypoxia for 2 hours and reoxygenation for up to 120 minutes. Western blots showed an increase in FKN protein in EC lysates beginning within 10 minutes of reoxygenation and continuing for at least 120 minutes. HUVECs were seeded into 12-well plates and grown to 100% confluence (lower left). For each time point of normoxia or hypoxia/reoxygenation, medium was harvested from 2 wells, pooled (600 μL), concentrated to ≈30 μL, and completely loaded on the gel in the appropriate lane. FKN in the media, representing the lower-molecular-weight s-FKN, increased within 15 minutes of reoxygenation and peaked at 30 minutes. b, s-FKN increases ICAM-1 mRNA in ECs. HUVECs were exposed to s-FKN for the indicated doses and times. Cells were harvested, total RNA was isolated, and real-time RT-PCR was performed for quantitation of ICAM-1 mRNA. ICAM-1 mRNA increased in a dose-dependent (top) and time-dependent (bottom) fashion.Show Less
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  Fractalkine Upregulates Intercellular Adhesion Molecule-1 in Endothelial Cells Through CX3CR1 and the Jak–Stat5 Pathway

  Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani, Lewis C. Becker

  Circulation Research November 2007, 101 (10) 1001-1008; DOI: https://doi.org/10.1161/CIRCRESAHA.107.160812

  

  By Xiao Ping Yang, Subhendra Mattagajasingh, Shaobo Su, Guibin Chen, Zheqing Cai, Karen Fox-Talbot, Kaikobad Irani and Lewis C. Becker

  Figure 2. s-FKN upregulates ICAM-1 protein in the intact mouse heart. Hearts were perfused with or without 50 ng/mL mouse s-FKN for 3 hours. a, Wester...Show More

  Figure 2. s-FKN upregulates ICAM-1 protein in the intact mouse heart. Hearts were perfused with or without 50 ng/mL mouse s-FKN for 3 hours. a, Western blots were done with goat anti-mouse ICAM-1 antibody. b and c, Immunohistology shows that ICAM-1 is upregulated in capillary endothelium (brown stain) following perfusion with s-FKN compared with perfusion with control buffer. Image magnification for b and c, ×160. IB indicates immunoblotting.Show Less

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