recombinant human α klotho  (R&D Systems)

Journal: Journal of molecular medicine (Berlin, Germany)

Article Title: Klotho suppresses high phosphate-induced osteogenic responses in human aortic valve interstitial cells through inhibition of Sox9

doi: 10.1007/s00109-017-1527-3

Figure Lengend Snippet: a Calcified aortic valve tissues (from donors of 56 to 77 years old) have lower levels of Klotho compared to normal aortic valve tissues (from donors of 28 to 56 years old). Data are mean ± SE. n = 4 per group; *P < 0.05 vs. normal. b Representative images (×40 objective for original magnification) of immunofluorescence staining show that Klotho (red) is present in the interstitial cells and interstitial spaces of aortic valve tissue, and diseased aortic valves have lower levels of Klotho. 4′,6-Diamidino-2-phenylindole (DAPI, blue) was used for nuclear counterstaining. Alexa 488-tagged wheat germ agglutinin (WGA, green) was used to outline plasma membrane. c AV I C s o f normal valves were treated with high Pi (2 or 3 mmol/l) for 72 h. Representative immunoblots and densitometric data show that high Pi reduces Klotho protein levels in human AVICs. Data are mean ± SE. n = 4 experiments using distinct cell isolates; *P < 0.05 vs. untreated control

Article Snippet: Chemicals and reagents Antibodies against Klotho and recombinant human Klotho protein (expressed by mouse myeloma cell line; endotoxin free) were purchased from Abcam (Cambridge, MA).

Techniques: Immunofluorescence, Staining, Clinical Proteomics, Membrane, Western Blot, Control

Journal: Journal of molecular medicine (Berlin, Germany)

Article Title: Klotho suppresses high phosphate-induced osteogenic responses in human aortic valve interstitial cells through inhibition of Sox9

doi: 10.1007/s00109-017-1527-3

Figure Lengend Snippet: a AV I C s o f normal human valves were pretreated with different concentrations of recombinant Klotho (0.25 or 0.5 µg/ml) 1 h prior to high Pi treatment for 72 h. Representative immunoblots and densitometric data show that Klotho suppresses high Pi-induced Runx2 and ALP upregulation in human AVICs. b AVICs of normal human valves were treated with high Pi (3 mmol/l), in the presence or absence of Klotho (0.5 µg/ml) for 14 days in the conditioning medium. Representative images (×10 objective for original magnification in the higher power images) of alizarin red staining and spectrophotometric data show that recombinant Klotho suppresses high Pi-induced calcium deposit formation. Data are mean ± SE. n = 4 experiments using distinct cell isolates; *P < 0.05 vs. untreated control; #P < 0.05 vs. Pi alone

Article Snippet: Chemicals and reagents Antibodies against Klotho and recombinant human Klotho protein (expressed by mouse myeloma cell line; endotoxin free) were purchased from Abcam (Cambridge, MA).

Techniques: Recombinant, Western Blot, Staining, Control

Journal: Journal of molecular medicine (Berlin, Germany)

Article Title: Klotho suppresses high phosphate-induced osteogenic responses in human aortic valve interstitial cells through inhibition of Sox9

doi: 10.1007/s00109-017-1527-3

Figure Lengend Snippet: Klotho reduces Runx2 and ALP levels in AVICs of calcified valves. AVICs of calcified human aortic valves were treated with different concentrations of recombinant Klotho (0.25 or 0.5 µg/ml) for 72 h. Representative immunoblots and densitometric data show that recombinant Klotho reduces the levels of Runx2 and ALP in diseased AVICs in the absence of high Pi stimulation. Data are mean ± SE. n = 4 experiments using distinct cell isolates; *P < 0.05 vs. untreated control

Article Snippet: Chemicals and reagents Antibodies against Klotho and recombinant human Klotho protein (expressed by mouse myeloma cell line; endotoxin free) were purchased from Abcam (Cambridge, MA).

Techniques: Recombinant, Western Blot, Control

Journal: Journal of molecular medicine (Berlin, Germany)

Article Title: Klotho suppresses high phosphate-induced osteogenic responses in human aortic valve interstitial cells through inhibition of Sox9

doi: 10.1007/s00109-017-1527-3

Figure Lengend Snippet: a AVICs were pretreated with different concentrations of recombinant Klotho (0.25 or 0.5 µg/ml) 1 h prior to high Pi treatment for 72 h. Immunoblots and densitometric data show that recombinant Klotho suppresses high Pi-induced Sox9 upregulation. b AVICs were pretreated with recombinant Klotho (0.5 µg/ml) 1 h prior to high Pi (3 mmol/l) stimulation for 3 h. Representative immunofluorescence images (×40 objective for original magnification) show that Klotho inhibits Sox9 (red) intranuclear translocation induced by high Pi. Alexa 488-tagged wheat germ agglutinin (WGA, green) was used to outline plasma membrane. 4′,6-Diamidino-2-phenylindole (DAPI, blue) was used for nuclear counterstaining. c AVICs were pretreated with different concentrations of recombinant Klotho (0.25 or 0.5 µg/ml) 1 h prior to high Pi treatment for 2 h. Immunoblots and densitometric data show that recombinant Klotho inhibits high Pi-induced phosphorylation of Akt and PKD. Data are mean ± SE. n = 4 experiments using distinct cell isolates; *P < 0.05 vs. untreated control; #P < 0.05 vs. Pi alone

Article Snippet: Chemicals and reagents Antibodies against Klotho and recombinant human Klotho protein (expressed by mouse myeloma cell line; endotoxin free) were purchased from Abcam (Cambridge, MA).

Techniques: Recombinant, Western Blot, Immunofluorescence, Translocation Assay, Clinical Proteomics, Membrane, Phospho-proteomics, Control

Journal: The Journal of Clinical Endocrinology and Metabolism

Article Title: α-Klotho Expression in Human Tissues

doi: 10.1210/jc.2015-1800

Figure Lengend Snippet: Confirmation of Transmembrane α-Klotho Protein Expression in Human Tissues and Cells

Article Snippet: As a control, we used recombinant human (rh) full-length α-Klotho protein (rh-α-Klotho) (5334-KL-025; R&D Systems).

Techniques: Expressing, Recombinant

Journal: Kidney & blood pressure research

Article Title: Upregulation of the creatine transporter Slc6A8 by Klotho.

doi: 10.1159/000368462

Figure Lengend Snippet: Fig. 1. Effect of Klotho coexpression on electrogenic creatine transport in Slc6A8-expressing Xenopus oocytes. A: Representative original tracings of creatine (2 mM) induced current in Xenopus oocytes injected with water (a), or with cRNA encoding Klotho alone (b),

Article Snippet: Where indicated, Brefeldin A (5μM, Sigma), recombinant human alpha Klotho protein (30ng/ml, 5334-KL-R&D Systems) and D-saccharic acid 1,4-lactone monohydrate (DSAL, 10μM, Sigma) were added.

Techniques: Expressing, Injection

Journal: Kidney & blood pressure research

Article Title: Upregulation of the creatine transporter Slc6A8 by Klotho.

doi: 10.1159/000368462

Figure Lengend Snippet: Fig. 3. Effect of brefeldin A in Slc6A8 expressing Xenopus oocytes with or without coexpression of Klotho. Arithmetic means ± SEM (n = 10-18) of the normalized creatine (2 mM) induced current in Xenopus oocytes injected with cRNA enconding CreaT, without (white bars) or with additional ex- pression of Klotho (black bars) in the absence (left bars) and pres- ence of 5 µM Brefeldin A for 12 hours (middle bars) or 24 hours (right bars) prior to the measure- ment. * (p<0.05), ** (p<0.005), *** (p<0.001) indicates statistically significant difference from the ab- sence of Klotho coexpression.

Article Snippet: Where indicated, Brefeldin A (5μM, Sigma), recombinant human alpha Klotho protein (30ng/ml, 5334-KL-R&D Systems) and D-saccharic acid 1,4-lactone monohydrate (DSAL, 10μM, Sigma) were added.

Techniques: Expressing, Injection