Journal: Journal of Biological Chemistry

Article Title: The Angiotensin II Type 1 Receptor (AT1R) Closely Interacts with Large Conductance Voltage- and Ca2+-activated K+ (BK) Channels and Inhibits Their Activity Independent of G-protein Activation

doi: 10.1074/jbc.m114.595603

Figure Lengend Snippet: FIGURE 2. AT1R expression is essential for ANG-II inhibitory effect on BK channels. A and C, representative traces before and after application of 1 M ANG-II in BK and AT1R-IRES-BK transfected cells, respectively. B and D, corresponding mean % values S.D. (n 6 cells, four independent transfections for BK alone, and n 3 cells, one transfection for AT1R BK). Only when AT1R is expressed, ANG-II reduced BK currents to 66 10% of its original value. E, example of time course of ANG-II action on BK currents. A similar time course was collected for other drugs. F, current traces before and after 1 M ANG-II in AT1R-IRES-BK transfected cells pretreated with 10 M losartan. G, corresponding mean % values (n 5 cells, one transfection). H, example of immunoblot analysis of pERK1/2 in lysates from cells expressing AT1R and BK that were untreated (lane 1), treated with ANG-II alone (lane 2), treated with losartan alone (lane 3), and pretreated with losartan and stimulated with ANG-II (lane 4). pERK1/2 levels were higher in ANG-II treated cells (lane 2) and returned to near baseline in cells preincubated with losartan (lane 4) (n 3 experiments). Loading and expression controls were ERK1/2, AT1R, and BK blots. All lanes were loaded with 30 g of protein. Antibody concentrations were 186 ng/ml pERK1/2 pAb, 2.85 ng/ml ERK1/2 pAb, 250 ng/ml c-Myc pAb, and 525 ng/ml BK mAb. Error bars indicate S.D. values. *, p 0.05. Ctrl, control; Los, losartan.

Article Snippet: For biochemistry and immunocytochemistry we used C-terminal c-Myc-tagged AT1R (AT1R-c-Myc) in pCMV6Entry (Origene), N-terminal FLAG-tagged AT1R (FLAGAT1R) in pcDNA3 (that includes the signal peptide, KTIIALSYIFCLVFA, to promote membrane expression), and BK -subunit without tag (BK) or with an N-terminal c-Myc tag (c-Myc-BK) (21) in pcDNA3.

Techniques: Expressing, Transfection, Western Blot, Control

Journal: Journal of Biological Chemistry

Article Title: The Angiotensin II Type 1 Receptor (AT1R) Closely Interacts with Large Conductance Voltage- and Ca2+-activated K+ (BK) Channels and Inhibits Their Activity Independent of G-protein Activation

doi: 10.1074/jbc.m114.595603

Figure Lengend Snippet: FIGURE 3. BK and AT1R co-localization in HEK293T cells and in arterial myocytes. A–C, live labeling of BK (green) and AT1R (red) in transiently expressing HEK293T cells displaying co-localization at the plasma membrane. HEK293T cells were co-transfected with N-terminally tagged FLAG-AT1R and c-Myc-BK. Anti-FLAG mAb and anti-c-Myc pAb were used for labeling. D–F, labeling of BK (red) and AT1R (green) after permeabilization showing total expression and subplasmalemma co-localization in rat renal arterial smooth muscle cells. Anti-BK mAb and anti-AT1R pAb were used for labeling. G, a representative three- dimensional cross-correlation plot of BK 3 AT1R as a function of pixel shift in the x and y axes (arrows in A and B indicate the cell used for the plot). The cross-correlation surface has a peak at zero pixel shift that decays abruptly by shifting the image few pixels indicative of specific co-localization. H, quantifica- tion of co-localization in HEK293T cells and smooth muscle cells (SMCs) by PPI analysis (see “Experimental Procedures”). In HEK293T cells, PPI is 0.68 0.08 for BK 3 AT1R and 0.87 0.08 for AT1R 3 BK (n 16 cells, three independent transfections). In rat renal arterial SMCs, PPI is 0.62 0.08 for AT1R 3 BK and 0.71 0.08 for BK 3 AT1R (n 18 cells, two independent cell isolations). Error bars indicate S.D. values.

Article Snippet: For biochemistry and immunocytochemistry we used C-terminal c-Myc-tagged AT1R (AT1R-c-Myc) in pCMV6Entry (Origene), N-terminal FLAG-tagged AT1R (FLAGAT1R) in pcDNA3 (that includes the signal peptide, KTIIALSYIFCLVFA, to promote membrane expression), and BK -subunit without tag (BK) or with an N-terminal c-Myc tag (c-Myc-BK) (21) in pcDNA3.

Techniques: Labeling, Expressing, Clinical Proteomics, Membrane, Transfection

Journal: Journal of Biological Chemistry

Article Title: The Angiotensin II Type 1 Receptor (AT1R) Closely Interacts with Large Conductance Voltage- and Ca2+-activated K+ (BK) Channels and Inhibits Their Activity Independent of G-protein Activation

doi: 10.1074/jbc.m114.595603

Figure Lengend Snippet: FIGURE 4. Constitutive inhibition of BK channel activity by AT1R expression. A, voltage stimulation protocol. B and C, representative inside-out patch recordings of type 1 and 3 currents in HEK293T cells co-transfected with BK and AT1R. Fitted V1⁄2 values were 0.4 and 36.8 mV, respectively. V1⁄2 values were calculated using instantaneous tail currents, I, to obtain FPo (G/Gmax I/Imax) as a function of the preceding test pulse voltage and fitting the data to a Boltzmann distribution as in F (see “Experimental Procedures”). D, examples of normalized BK tail currents (I/Imax) in cells expressing BK alone or BK AT1R. The scheme (top trace) shows the pulse protocol. Current traces demonstrate that peak I/Imax (FPo) magnitude followed the trend BK type 1 AT1R BK type 2 AT1R BK type 3 AT1R BK. E, BK activation kinetics was slowed down by AT1R co-expression in the order type 3 type 2 type 1 (test pulse 60 mV) consistent with a larger decrease in Po in type 3 AT1R BK channels. F, average voltage activation curves (FPo versus voltage) of BK expressed alone or in combination with AT1R. The error bars are within symbols and indicate S.E. Continuous lines are the means of the fitted curves of each experiment. Average V1⁄2 values were 0.4 3.2 for BK (n 24 cells), 0.3 2 for AT1R BK type 1 (n 22 cells), 13 2 for AT1R BK type 2 (n 17 cells), and AT1R BK type 3 35 2 (n 12 cells). Type 3 channels displayed the highest inhibition by AT1R co-expression (FPo is lower at a given potential when compared with BK expressed alone). G, V1⁄2 distribution in all patches (n 79) from cells co-expressing BK and AT1R. Ca2 concentration facing the intracellular side of the channels was 6.7 M.

Article Snippet: For biochemistry and immunocytochemistry we used C-terminal c-Myc-tagged AT1R (AT1R-c-Myc) in pCMV6Entry (Origene), N-terminal FLAG-tagged AT1R (FLAGAT1R) in pcDNA3 (that includes the signal peptide, KTIIALSYIFCLVFA, to promote membrane expression), and BK -subunit without tag (BK) or with an N-terminal c-Myc tag (c-Myc-BK) (21) in pcDNA3.

Techniques: Inhibition, Activity Assay, Expressing, Transfection, Activation Assay, Concentration Assay

Journal: Journal of Biological Chemistry

Article Title: The Angiotensin II Type 1 Receptor (AT1R) Closely Interacts with Large Conductance Voltage- and Ca2+-activated K+ (BK) Channels and Inhibits Their Activity Independent of G-protein Activation

doi: 10.1074/jbc.m114.595603

Figure Lengend Snippet: FIGURE 5. AT1R forms a complex with BK in HEK293T cells. A, cartoons of seven-transmembrane domain BK -subunit (left) and AT1R (right). B, AT1R pulls down BK (n 3 independent experiments). Lanes 1–4, IP using anti-c-Myc pAb recognizing AT1R-c-Myc in lysates from transfected () or untransfected () cells with AT1R-c-Myc and/or BK; lanes 5 and 6, IP with rabbit IgG or protein G beads using lysates from cells co-transfected with AT1R-c-Myc and BK; lane 7, IP using a mixture of lysates from cells independently expressing AT1R-c-Myc or BK. Loading was equal in all lanes (25 l). Lower panel, control immunoblot of BK expression in input cell lysates (30 g of protein/lane). Immunoblots were with 525 ng/ml anti-BK mAb. C, control showing effective IP of AT1R in same cell lysates or mixture of lysates. Loading was equal in all lanes (5 l). Lower panel, AT1R expression in input cell lysates (30 g of protein/lane) probed with 250 ng/ml anti-c-Myc pAb. D, BK also pulls down AT1R (n 2 independent experiments). Lanes 1–3, IP of lysates from cells expressing only AT1R, expressing only BK or co-expressing both proteins. IP was with anti-c-Myc pAb recognizing c-Myc-BK, and immunoblot was with 495 ng/ml anti-FLAG mAb recognizing AT1R. Lane 4, negative control using IgG to IP lysates from cells expressing both AT1R and BK. Lower panel, immunoblot of input lysates. E, control of effective IP of BK. Lower panel, expression of BK in corresponding input cell lysates. Immunoblot was with 525 ng/ml anti-BK mAb. IB, immunoblot; IE, independently expressing.

Article Snippet: For biochemistry and immunocytochemistry we used C-terminal c-Myc-tagged AT1R (AT1R-c-Myc) in pCMV6Entry (Origene), N-terminal FLAG-tagged AT1R (FLAGAT1R) in pcDNA3 (that includes the signal peptide, KTIIALSYIFCLVFA, to promote membrane expression), and BK -subunit without tag (BK) or with an N-terminal c-Myc tag (c-Myc-BK) (21) in pcDNA3.

Techniques: Transfection, Expressing, Control, Western Blot, Negative Control

Journal: Journal of Biological Chemistry

Article Title: The Angiotensin II Type 1 Receptor (AT1R) Closely Interacts with Large Conductance Voltage- and Ca2+-activated K+ (BK) Channels and Inhibits Their Activity Independent of G-protein Activation

doi: 10.1074/jbc.m114.595603

Figure Lengend Snippet: FIGURE 6. Molecular analysis of BK-AT1R association. A, BK topology (left) and truncated BK constructs that were used for immunocytochemistry (right). RCK, regulator of conductance for K. B, overlaid confocal images of cells co-expressing AT1R (red) and full-length or truncated BK (green) molecules: BK, BK1–711, BK1–441, BK1–343, and BK323–1113. Note that BK323–1113 appears intracellular. Larger yellow squares show 5 magnification of the regions in the smaller yellow squares. C, specific co-localization analysis by PPI (25) demonstrating that the first 1–343 amino acids of BK are sufficient for its association with AT1R. PPI values were 0.68 0.1 for BK to AT1R and 0.87 0.08 for AT1R to BK (n 16 cells, four independent experiments); 0.63 0.06 for BK1–711 to AT1R and 0.83 0.12 for AT1R to BK1–711 (n 9 cells, two independent experiments); 0.75 0.1 for BK1–441 to AT1R and 0.88 0.12 for AT1R to BK1–441 (n 13 cells, two independent experiments); 0.83 0.11 for BK1–343 to AT1R and 0.91 0.07 for AT1R to BK1–343 (n 15 cells, three independent experiments); and 0.19 0.08 for BK322–1113 to AT1R and 0.26 0.09 for AT1R to BK322–1113 (n 17 cells, three independent experiments). Error bars indicate S.D. values. *, p 0.05.

Article Snippet: For biochemistry and immunocytochemistry we used C-terminal c-Myc-tagged AT1R (AT1R-c-Myc) in pCMV6Entry (Origene), N-terminal FLAG-tagged AT1R (FLAGAT1R) in pcDNA3 (that includes the signal peptide, KTIIALSYIFCLVFA, to promote membrane expression), and BK -subunit without tag (BK) or with an N-terminal c-Myc tag (c-Myc-BK) (21) in pcDNA3.

Techniques: Construct, Immunocytochemistry, Expressing

Journal: International immunopharmacology

Article Title: GTS-21 attenuates ACE/ACE2 ratio and glycocalyx shedding in lipopolysaccharide-induced acute lung injury by targeting macrophage polarization derived ADAM-17.

doi: 10.1016/j.intimp.2024.111603

Figure Lengend Snippet: Fig. 2. GTS-21 regulated the ACE/ACE2 ratio and glycocalyx shedding in LPS-ALI mice A-D ELISA kits were used to quantify the concentrations of major renin- angiotensin system (RAS) regulating enzymes (ACE, Ang II, ACE2, and Ang 1-7) in the lung tissue (n = 6 per group). E Protein levels of important RAS- regulating enzymes (ACE, AT1R, ACE2, and MasR) (n = 3 per group). The relative protein levels are displayed. F Levels of NF-κB phosphoproteins (p-P65 and p- IκB) (n = 3 per group). The relative protein levels are displayed. The immunofluorescence of HS, SDC-1, and surfactant protein D in the lungs is shown in G in representative images and densitometry (scar bar, 20 mm, n = 3 per group). H Evans Blue staining tested lung permeability (n = 3 for each group). Using ELISA, we assessed the BALF concentrations of HS I and HPA J; SP-D in serum K and BALF L (n = 5 for each group). M Relative protein levels of SDC-1, HPA, EXT-1, and SP-D in The lung tissues (n = 3 for each group). All data were shown as mean ± SD. * P < 0.05, ** P < 0.01.

Article Snippet: AT1R (cat. no. 25343–1-ap) and MasR (cat. no. 20080–1-ap) were bought from Proteintech.

Techniques: Enzyme-linked Immunosorbent Assay, Immunofluorescence, Staining, Permeability

Journal: International immunopharmacology

Article Title: GTS-21 attenuates ACE/ACE2 ratio and glycocalyx shedding in lipopolysaccharide-induced acute lung injury by targeting macrophage polarization derived ADAM-17.

doi: 10.1016/j.intimp.2024.111603

Figure Lengend Snippet: Fig. 5. Effect of macrophage polarization on ACE/ACE2 ratio and glycocalyx shedding in epithelial cells. A The experimental procedure for RAW264.7 cells su pernatant with different polarization states were added to the culture medium to test their effect on alveolar epithelial cells. B The expressions of key renin- angiotensin system regulatory enzymes (ACE, AT1R, ACE2, and MasR) and main component of NF-κB pathway (p-P65 and p-IκB) were detected. C Representa tive image and densitometry of immunofluorescence of the main component of glycocalyx (HS and SDC-1) and surfactant protein-D in MLE-12 cells (Scar bar, 20 μm). D Expression of the main components of the glycocalyx (SDC-1), key regulatory enzymes (HPA and EXT1), and SP-D was quantified in MLE-12 cells. Based on three independent experiments, all data were shown as mean ± SD. * P < 0.05, ** P < 0.01.

Article Snippet: AT1R (cat. no. 25343–1-ap) and MasR (cat. no. 20080–1-ap) were bought from Proteintech.

Techniques: Immunofluorescence, Expressing

Journal: International immunopharmacology

Article Title: GTS-21 attenuates ACE/ACE2 ratio and glycocalyx shedding in lipopolysaccharide-induced acute lung injury by targeting macrophage polarization derived ADAM-17.

doi: 10.1016/j.intimp.2024.111603

Figure Lengend Snippet: Fig. 7. Effect of ADAM-17 on ACE/ACE2 ratio and glycocalyx shedding in epithelial cells Experimental procedure of RAW264.7 cells with recombinant ADAM17 (rADAM-17) or siRNA ADAM-17 to elevate and inhibit the activity of ADAM-17. Macrophage supernatant was used to test its effect on epithelial cells. A The ex pressions of key renin-angiotensin system regulatory enzymes (ACE, AT1R, ACE2, and MasR) and main component of NF-κB pathway (p-P65 and p-IκB) were determined. B Representative image and densitometry of immunofluorescence of main component of glycocalyx (HS and SDC-1) and surfactant protein-D in the MLE- 12 cells (Scar bar, 20 μm). C The expression levels of the main components of the glycocalyx (SDC-1), key regulatory enzymes (HPA and EXT1), and SP-D were quantified in MLE-12 cells. Based on three independent experiments, all data were shown as mean ± SD. * P < 0.05, ** P < 0.01.

Article Snippet: AT1R (cat. no. 25343–1-ap) and MasR (cat. no. 20080–1-ap) were bought from Proteintech.

Techniques: Recombinant, Activity Assay, Immunofluorescence, Expressing

Journal: Journal of the American Heart Association

Article Title: Connexin37‐Dependent Mechanisms Selectively Contribute to Modulate Angiotensin II‐Mediated Hypertension

doi: 10.1161/jaha.118.010823

Figure Lengend Snippet: Figure 5. Loss of Cx37 selectively alters the expression of some Ang II-stimulated signals. The levels of transcripts coding for Angiotensinogen (Agt), Angiotensin receptor 1a (Agtr1a), and Angiotensin receptor 1b (Agtr1b) were similar in the aortas of WT and Cx37/ mice. In contrast, the transcripts coding for Angiotensin receptor 2 (Agtr2), Connector enhancer of kinase suppressor of Ras 1 (Cnksr1), SH2 domain-containing tyrosine phosphatase 2 (Ptpn6), and Glutamyl aminopeptidase A (Enpep), which all contribute to control the Ang II pathway, were increased in the aortas of Cx37/ mice. N=9. Results are means+SEM. *P≤0.05, ***P≤0.001 vs WT mice, as given by t test. Ang II indicates angiotensin II; Cx37, Connexin37; WT, wild type.

Article Snippet: The membranes were then incubated overnight at 4°C with 1 of the following primary antibodies: rabbit polyclonal antibodies against AT1R (Alomone labs, AAR-011, 1:1000), AT2R (Alomone labs, AAR-012, 1:1000), MLC2 (Cell Signaling, 3672, 1:1000), P-MLC2 (Cell Signaling, 3674, 1:1000), ERK (Cell Signaling, 4695, 1:1000), P-ERK (Cell Signaling, 9101, 1:1000), AKT (Cell Signaling, 9272S, 1:1000), Cx40 (Chemicon, AB1726; 1:250), Cx37 (Biotrend Chemikalien, Cx37A11-A; 1:500), Cx43 (Cell Signaling, 3512S, 1:500) or Cx45 (Millipore, AB1745, 1:500); mouse monoclonal antibodies against eNOS (BD Biosciences, 610297, 1:500), PeNOS (BD Biosciences, 612392, 1:500), P-AKT (Cell Signaling, 4051, 1:500), and atubulin (Sigma-Aldrich, T5168; 1:2500).

Techniques: Expressing, Control

Journal: Journal of the American Heart Association

Article Title: Connexin37‐Dependent Mechanisms Selectively Contribute to Modulate Angiotensin II‐Mediated Hypertension

doi: 10.1161/jaha.118.010823

Figure Lengend Snippet: Figure 6. Loss of Cx37 selectively alters the expression of the AT2 receptors in the aortas of Cx37/ mice. A, Immunofluorescence showed a similar staining for AT1R (upper panels) in the aortas of all WT and Cx37/ mice. In contrast, the staining for AT2R was more intense in the aortas of Cx37/ than in WT mice. Bar=20 lm. L indicates lumen; M, media. B, Western blot showed that the aortas of WT and Cx37/ mice expressed similar levels of the AT1R protein. C, In contrast, the basal levels of the AT2R protein were higher in Cx37/ than in WT mice, and were not increased after the 2K1C surgery. WT sham N=21; Cx37/ sham N=17; WT 2K1C N=19; Cx37/ 2K1C N=17. D and E, Comparable observations were made for both AT1R (D) and AT2R (E) in mice infused daily with 1 mg Ang II/kg body weight WT NaCl N=4; Cx37/ NaCl, Cx37/ Ang II N=3, WT Ang II N=6. Results are means+SEM. °P≤0.05 vs WT mice, as given by 2-way ANOVA. 2K1C indicates 2-kidney, 1-clip; AngII, angiotensin II; AT1R, Ang II type 1 receptors; AT2R, Ang II type 2 receptors; Cx37, Connexin37; DAPI, 40,6-diamidino-2-phenylindole; Sham, sham- operated controls; WT, wild type.

Article Snippet: The membranes were then incubated overnight at 4°C with 1 of the following primary antibodies: rabbit polyclonal antibodies against AT1R (Alomone labs, AAR-011, 1:1000), AT2R (Alomone labs, AAR-012, 1:1000), MLC2 (Cell Signaling, 3672, 1:1000), P-MLC2 (Cell Signaling, 3674, 1:1000), ERK (Cell Signaling, 4695, 1:1000), P-ERK (Cell Signaling, 9101, 1:1000), AKT (Cell Signaling, 9272S, 1:1000), Cx40 (Chemicon, AB1726; 1:250), Cx37 (Biotrend Chemikalien, Cx37A11-A; 1:500), Cx43 (Cell Signaling, 3512S, 1:500) or Cx45 (Millipore, AB1745, 1:500); mouse monoclonal antibodies against eNOS (BD Biosciences, 610297, 1:500), PeNOS (BD Biosciences, 612392, 1:500), P-AKT (Cell Signaling, 4051, 1:500), and atubulin (Sigma-Aldrich, T5168; 1:2500).

Techniques: Expressing, Staining, Western Blot, Immunofluorescence

Journal: Journal of the American Heart Association

Article Title: Connexin37‐Dependent Mechanisms Selectively Contribute to Modulate Angiotensin II‐Mediated Hypertension

doi: 10.1161/jaha.118.010823

Figure Lengend Snippet: Figure 7. Loss of Cx37 does not alter the expression of several signals involved in the Ang II effects. The levels of transcripts coding for the chymase (Cma1), which converts Ang I to Ang II, the Regulator of G-protein signaling 2 (Rgs2), which negatively regulates AT1R function by modulating its Gaq subunit activity, the Dual specificity protein phosphatase 1 (Dusp1), an AT2R- associated phosphatase that contributes to AT2R-mediated inactivation of the ERK cascade, and Mas1, the receptor for Ang-(1–7) that promotes vasodilatation, were similar in the aortas of WT and Cx37/ mice. WT N=9; Cx37/ N=8. Results are means+SEM. Statistical analysis was performed using t test. Ang indicates angiotensin; AT1R, Ang II type 1 receptors; AT2R, Ang II type 2 receptors; Cx37, Connexin37; WT, wild type.

Article Snippet: The membranes were then incubated overnight at 4°C with 1 of the following primary antibodies: rabbit polyclonal antibodies against AT1R (Alomone labs, AAR-011, 1:1000), AT2R (Alomone labs, AAR-012, 1:1000), MLC2 (Cell Signaling, 3672, 1:1000), P-MLC2 (Cell Signaling, 3674, 1:1000), ERK (Cell Signaling, 4695, 1:1000), P-ERK (Cell Signaling, 9101, 1:1000), AKT (Cell Signaling, 9272S, 1:1000), Cx40 (Chemicon, AB1726; 1:250), Cx37 (Biotrend Chemikalien, Cx37A11-A; 1:500), Cx43 (Cell Signaling, 3512S, 1:500) or Cx45 (Millipore, AB1745, 1:500); mouse monoclonal antibodies against eNOS (BD Biosciences, 610297, 1:500), PeNOS (BD Biosciences, 612392, 1:500), P-AKT (Cell Signaling, 4051, 1:500), and atubulin (Sigma-Aldrich, T5168; 1:2500).

Techniques: Expressing, Activity Assay, Inactivation, Analysis

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: CCDC25 serum levels were detected using the dot blot assay. (A) Representative dot blot assay to detect CCDC25 serum levels. Samples A1 and B1 were the standard (1 ng/µl), samples A2-A8 and B2-B8 were duplicates of two-fold serial dilutions (0.5, 0.25, 0.125, 0.0625, 0.0313, 0.0156 and 0.0078 ng/µl), samples A9 and B9 were the positive control (pooled CCA serum samples) and samples C1-C2 were the blank control (saline solution). (B) The standard curve of CCDC25 levels. CCDC25, coiled-coil domain containing 25.

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques: Dot Blot, Positive Control, Control, Saline

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: Protein-ligand interaction map of CCDC25-related proteins using STITCH v.5.0 software. Stronger associations are presented as thicker lines. Protein-protein interactions were indicated using solid lines, and no chemical-protein interactions or interactions between chemical and chemical molecules were observed. CCDC25, coiled-coil domain containing 25.

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques: Software, Protein-Protein interactions

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: Serum CCDC25, CA19-9 and CEA levels in the CCA, BBD and HC groups. (A) The median level ± quartile deviation of the CCDC25 level was 0.0017±0.0008 ng/µl in the HC group, 0.15±0.03 ng/µl in the BBD group and 0.28±0.06 ng/µl in the CCA group. The serum CCDC25 level at 0.2 ng/µl was used as the cut-off value to distinguish between patients with CCA with low and high serum CCDC25 level. (B) Median level ± quartile deviation of the CEA level was 3.9±2.6 ng/ml in the BBD group and 5.7±5.1 ng/ml in CCA group and the cut-off level was 5 ng/ml. (C) The median level ± quartile deviation of the CA19-9 level was 53.4±26.3 U/ml in the BBD group and 73.5±29.8 U/ml in the CCA group and the cut-off level was 37 U/ml. CCDC25, coiled-coil domain containing 25; CA19-9, carbohydrate antigen 19-9; CEA, carcinoembryonic antigen; CCA, cholangiocarcinoma; BBD, benign biliary disease; HC, healthy control.

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques: Control

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: Distribution of the CCDC25 serum levels in patients with CCA. The frequency plot of the CCDC25 serum level in patients with CCA revealed two distinct populations, which were separated at concentration 0.2 ng/µl. Therefore, 0.2 ng/µl was used as the cut-off value to distinguish between low and high CCDC25 levels in serum samples obtained from patients with CCA. CCDC25, coiled-coil domain containing 25; CCA, cholangiocarcinoma.

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques: Concentration Assay

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: Correlation between CCDC25 serum levels and CA19-9 and CEA serum levels. (A) Correlation between CCDC25 serum level and CA19-9 serum level (r=0.04; P=0.81) and (B) CEA serum level (r=0.03; P=0.62), calculated using the Spearman's correlation test, both exhibited a negligible positive correlation. CCDC25, coiled-coil domain containing 25; CA19-9, carbohydrate 19-9 antigen; CEA, carcinoembryonic antigen .

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques:

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: ROC curve evaluation of serum CCDC25 levels in the CCA, BBD and HC groups. (A) ROC curve of the CCDC25 level between the CCA and HC groups represented 93% sensitivity and 100% specificity with a cut-off CCDC25 level of 0.11 ng/µl. (B) ROC curve of the CCDC25 level between the BBD and HC groups represented 98.1% sensitivity and 90.4% specificity with a cut-off CCDC25 level of 0.045 ng/µl. ROC, receiver operating characteristic; CCDC25, coiled-coil domain containing 25; CCA, cholangiocarcinoma; BBD, benign biliary disease; HC, healthy control. CCDC25, coiled-coil domain containing 25; AUC, area under the curve; CI, confidence interval.

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques: Control

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: ROC curve evaluation of CCDC25, CA19-9 and CEA serum levels in the BBD and CCA groups. (A) ROC curve of the CCDC25 level in the BBD and CCA groups represented 75% sensitivity and 84% specificity, and a cut-off CCDC25 level of 0.18 ng/µl (B) ROC curve of serum CA19-9 level in the BBD and CCA groups represented 52.4% sensitivity and 46.5% specificity, and a cut-off CA19-9 level of 105.4 U/ml. (C) ROC curve of serum CEA level in the BBD and CCA groups represented 54.4% sensitivity and 60% specificity, and a cut-off CEA level of 19.6 ng/ml. ROC, receiver operating characteristic; CCDC25, coiled-coil domain containing 25; CA19-9, carbohydrate 19-9 antigen; CEA, carcinoembryonic antigen; AUC, area under the curve; CI, confidence interval .

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques:

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: Receiver operating charactersitic curve analysis of the diagnostic value of CCDC25, CEA and CA19-9 in CCA.

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques: Diagnostic Assay, Comparison

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: Correlation between the serum CCDC25 level and the CCDC25 H-score. The Spearman's correlation test revealed a moderate positive correlation between the two variables. r 2 =0.52, P=0.01. CCDC25, coiled-coil domain containing 25.

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques:

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: Association of patient clinical data and low and high serum CCDC25 levels in patients with cholangiocarcinoma.

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques: Marker

Journal: Oncology Letters

Article Title: Serum coiled-coil domain containing 25 protein as a potential screening/diagnostic biomarker for cholangiocarcinoma

doi: 10.3892/ol.2019.11162

Figure Lengend Snippet: Overall survival time of patients with cholangiocarcinoma with low and high CCDC25 serum levels estimated using the Kaplan-Meier method. The overall survival times was significantly higher in patients with high serum CCDC25 levels compared with those with low levels (365 days vs. 242 days; P=0.031). Censored data represent incomplete information for a study participant, observation or value of a measurement.

Article Snippet: The CCDC25 concentration in each serum sample was calculated based on the standard curve prepared using the standard recombinant CCDC25 protein (cat. no. orb424527; Biorbyt Ltd.).

Techniques: