High glucose reduces albumin uptake in cultured proximal tubular cells (LLC-PK1).
In this study, we clarify that high glucose inhibits albumin uptake in cultured LLC-PK1 cells. LLC-PK1 cells cultured for 6 days with 5.5-27.8 mM D-glucose were challenged by fluorescein isothiocyanate (FITC)-conjugated human albumin (HA). FITC-HA binding and uptake were inhibited by >5.5mM glucose (5.5 mM > (P < 0.01) 11.0 mM > (P < 0.05) 16.7 mM approximately= 27.8 mM). Analysis of FITC-HA binding and uptake at 5.5 and 16.7 mM D-glucose (high glucose, HG) showed decreased affinity (K(m) for binding: 35.5 mg/l versus 52.6 mg/l, K(m) for uptake; 41.3 mg/l versus 55.6 mg/l) and maximal velocity (B(max)--0.33 microg versus 0.27 microg/30 min/mg protein; U(max)--4.40 microg versus 3.48 microg/60 min/mg protein) at HG. A comparison of the time courses of FITC-HA binding and uptake at 5.5 mM glucose and at HG showed that HG suppressed them beyond 15 min (P < 0.005-0.001). Phlorizin (>0.25 mM) completely reversed the HG-induced inhibition of FITC-HA binding and uptake. High glucose decreased mRNA of GLUT-1 and SGLT-1, but did not influence that of SGLT-2. The simultaneous presence of Vitamin E (10(-6)M), Vitamin C (10(-6)M) and reduced glutathione (0.25 mM) reversed the suppressed FITC-HA binding and uptake by HG, while any one or two of these molecules, and various inhibitors of advanced glycation end products, failed to do so. In conclusion, a high glucose milieu causes inhibition of albumin binding and uptake in proximal tubular cells by increasing metabolic oxidative stress through excessive glucose flux via the sodium glucose transporter.
Ishibashi
Fukashi
F
Ishibashi Clinic, 1-9-41-2 Kushido Hatsukaichi Hiroshima 738-0033, Japan. ishiclic@urban.ne.jp
D000818:Animals; D002478:Cells, Cultured; D003470:Culture Media; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D005456:Fluorescent Dyes; D005947:Glucose / Q000494:pharmacology*; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000166:cytology*; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D018384:Oxidative Stress; D012709:Serum Albumin / Q000378:metabolism*; D013552:Swine; D013997:Time Factors
D003470:Culture Media; D005456:Fluorescent Dyes; D009004:Monosaccharide Transport Proteins; D012709:Serum Albumin; D016650:Fluorescein-5-isothiocyanate; D005947:Glucose