recombinant human klotho protein  (PeproTech)

Journal: Frontiers in Pharmacology

Article Title: Klotho-derived peptide KP1 ameliorates SARS-CoV-2-associated acute kidney injury

doi: 10.3389/fphar.2023.1333389

Figure Lengend Snippet: Klotho deficiency is a common feature of mouse kidney with preexisting chronic kidney diseases or advanced age. ( A – E ) Representative Western blots show renal protein level of Klotho in mouse models of ischemia-reperfusion injury (IRI), unilateral ureteral obstruction (UUO), remnant kidney after 5/6 nephrectomy (5/6Nx), db/db with diabetic kidney disease (DKD) and advanced age (24 months old), respectively. ( F ) Quantitative data of Klotho protein levels in the kidney of various models as indicated are presented. ** p < 0.01 versus sham ( n = 6). ( G ) Representative micrographs of immunochemical staining show Klotho expression and localization in different models of mice as indicated. Arrows indicate positive staining. Scale bar, 50 µm.

Article Snippet: Plasmid DNA-lipid complexes were added to the cells and incubated for 4–6 h. After transfection, cells were treated with KP1 (10 μg/mL) or recombinant human Klotho (100 ng/mL) (#5334-KL; R&D Systems) for 48 h. Cells were then collected and subjected to Western blot analyses or TUNEL staining, respectively.

Techniques: Western Blot, Staining, Expressing

Journal: Frontiers in Pharmacology

Article Title: Klotho-derived peptide KP1 ameliorates SARS-CoV-2-associated acute kidney injury

doi: 10.3389/fphar.2023.1333389

Figure Lengend Snippet: Klotho-deficiency sensitizes mice to SARS-CoV-2 N protein-triggered tubular injury and apoptosis. (A) Experimental design. The blue arrow indicates the timing of injecting pcDNA3 empty vector or pSARS-CoV-2 N Protein (N) plasmid into normal (WT) or Klotho-deficient ( KL/KL ) mice, respectively. (B, C) Graphic presentation shows serum creatinine (SCr) and blood urea nitrogen (BUN) levels in different groups. ns, not significant. (D–H) Representative Western blot (D) and quantitative data show renal protein levels of N Protein (F) , KIM-1 (G) and NGAL (H) in different groups. * p < 0.05, ** p < 0.01, *** p < 0.001 ( n = 6). ( I – L ) Representative Western blot (I) and quantitative data show renal protein levels of cleaved PARP (J) , p53 (K) and FADD ( L ). * p < 0.05, ** p < 0.01, *** p < 0.001 ( n = 6). ( M ) Representative micrographs of immunochemical staining show renal expression of N protein and P53 are presented. Arrows indicate positive staining. Scale bar, 50 µm. ( N ) Quantitative data of N Protein and p53. ** p < 0.01, *** p < 0.001 ( n = 6).

Article Snippet: Plasmid DNA-lipid complexes were added to the cells and incubated for 4–6 h. After transfection, cells were treated with KP1 (10 μg/mL) or recombinant human Klotho (100 ng/mL) (#5334-KL; R&D Systems) for 48 h. Cells were then collected and subjected to Western blot analyses or TUNEL staining, respectively.

Techniques: Plasmid Preparation, Western Blot, Staining, Expressing

Journal: Frontiers in Pharmacology

Article Title: Klotho-derived peptide KP1 ameliorates SARS-CoV-2-associated acute kidney injury

doi: 10.3389/fphar.2023.1333389

Figure Lengend Snippet: KP1 ameliorates tubular cell injury and apoptosis induced by SARS-CoV-2 N protein in vitro . (A–C) Representative Western blot (A) and quantitative data show the expression of N Protein (B) and KIM-1 (C) . Cells were treated with KP1 or recombinant human Klotho for 48 h after transfecting SARS-CoV-2 N Protein plasmid. ** p < 0.01, *** p < 0.001 ( n = 6). (D–G) Representative Western blot (D) and quantitative data show the expression of PARP (E) , p53 (F) and FADD (G) . * p < 0.05, ** p < 0.01, *** p < 0.001 ( n = 6). (H, I) Representative TUNEL staining (H) and quantitative data (I) show that SARS-CoV-2 N protein induced HK-2 cell apoptosis, while KP1 or Klotho (KL) protected HK-2 cells from apoptosis. ** p < 0.01 ( n = 6). Arrow indicates apoptotic cell.

Article Snippet: Plasmid DNA-lipid complexes were added to the cells and incubated for 4–6 h. After transfection, cells were treated with KP1 (10 μg/mL) or recombinant human Klotho (100 ng/mL) (#5334-KL; R&D Systems) for 48 h. Cells were then collected and subjected to Western blot analyses or TUNEL staining, respectively.

Techniques: In Vitro, Western Blot, Expressing, Recombinant, Plasmid Preparation, TUNEL Assay, Staining

Journal: Frontiers in Pharmacology

Article Title: Klotho-derived peptide KP1 ameliorates SARS-CoV-2-associated acute kidney injury

doi: 10.3389/fphar.2023.1333389

Figure Lengend Snippet: KP1 ameliorates acute kidney injury induced by SARS-CoV-2 N protein and ischemia-reperfusion injury in vivo . (A) Experimental design. The blue arrow indicates the timing of injecting pcDNA3 empty vector or pSARS-CoV-2 N Protein plasmid (N). The green arrowheads indicate the timing of injecting KP1 at the concentration of 1 mg/day/kg. The black arrow indicates the timing of IRI. (B, C) Graphic presentations show SCr and BUN levels in different groups as indicated. * p < 0.05, ** p < 0.01, *** p < 0.001 ( n = 6). (D–H) Representative Western blot (D) and quantitative data show renal protein levels of Klotho (E) , N protein (F) , KIM-1 (G) and NGAL (H) . * p < 0.05, ** p < 0.01, *** p < 0.001 ( n = 6). (I, J) Representative micrographs of Periodic acid-Schiff (PAS) staining and immunochemical staining for N protein and KIM-1 are presented. Arrows indicate positive staining. Scale bar, 50 µm. Semi-quantification data are presented in Panel (J) . * p < 0.05, ** p < 0.01, *** p < 0.001 ( n = 6).

Article Snippet: Plasmid DNA-lipid complexes were added to the cells and incubated for 4–6 h. After transfection, cells were treated with KP1 (10 μg/mL) or recombinant human Klotho (100 ng/mL) (#5334-KL; R&D Systems) for 48 h. Cells were then collected and subjected to Western blot analyses or TUNEL staining, respectively.

Techniques: In Vivo, Plasmid Preparation, Concentration Assay, Western Blot, Staining