[Phenotypic and functional alterations of mesangial cells in patients with diabetic nephropathy].
OBJECTIVE
To explore the phenotypic and functional changes of glomerular mesangial cells (MC) in the development of diabetic nephropathy (DN).


METHODS
Renal biopsy specimens were obtained from patients with type II DN. MC from microdissected glomeruli were cultured in vitro. Both the cells deriving from minute piece of renal biopsy specimen of type 2 DN with overt proteinuria and from transplanted donors' kidneys were investigated. Cell volume, RNA/DNA ratio and the production of fibronectin and glucose transporter 1 (GLUT1) was analyzed by flow ctrometry. Cell proliferation was determined by 3H-Tdr incorporation assay and doubling time. Immunofluorescence staining and flow ctrometry were used to examine the expression of alpha-smooth muscle actin and extracellular matrix. The mRNA expression of GLUT1 was determined by Northern blotting and flow cytometry. Glucose uptake rate by MC was detected using the [3H]-2-DG. The activity of glutamine: fructose-6-P aminotransferase (GFAT), the key enzyme of hexosamine pathway, was measured by spetrophotometry method. Specimens from transplanted kidneys were used a s controls.


RESULTS
In specimens from patients with type II DN, increase in volume of MC (in arbitrary units) and RNA/DNA ratio (0.29 +/- 0.05 vs. 0.17 +/- 0.03, P < 0.01) accompanied with high 3H-Tdr incorporation rate (1,898 +/- 421 vs. 1,221 +/- 262, P < 0.05) and shorter doubling time (31.6 +/- 1.84 h vs. 35.0 +/- 5.37 h, P < 0.05) was found in comparison with the control. The synthesis of alpha-smooth muscle actin and extracellular matrix, including fibronectin and laminin, increased. Enhanced mRNA and protein expression of GLUT1 were verified in MC from DN. 2-DG uptake assay showed increased glucose uptake rate in MC from DN compared to the control(1,592 cpm x 10(5) cell-1 vs. 1,275 cpm x 10(5) cell-1, P < 0.05). Furthermore, MC from DN demonstrated a higher GFAT activity as compared to control.


CONCLUSIONS
Dramatic phenotypic and functional changes, including cell hypertrophy, increased cell turnover, excessive formation of ECM, enhanced expression of GLUT1 as well as alterration in cell glucose uptake, excessive flux of glucose metabolism through the hexosamine pathway, occur in MC from type II DN. The above mentioned phenotypic and functional changes may be the basis of pathogenesis of MC in DN.
Liu
Z
Z
Research Institute of Nephrology, Jinling Hospital, Nanjing University School of Medicine, Nanjing 210002, China.
Chen
Z
Z
Li
Y
Y
D000199:Actins / Q000378:metabolism*; D002455:Cell Division / Q000502:physiology; D048429:Cell Size / Q000502:physiology; D004247:DNA / Q000378:metabolism; D003928:Diabetic Nephropathies / Q000503:physiopathology*; D005109:Extracellular Matrix / Q000378:metabolism; D005920:Glomerular Mesangium / Q000378:metabolism* / Q000473:pathology; D005947:Glucose / Q000378:metabolism; D051272:Glucose Transporter Type 1; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing) / Q000378:metabolism*; D006801:Humans; D009004:Monosaccharide Transport Proteins / Q000096:biosynthesis*; D010641:Phenotype; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis
D000199:Actins; D051272:Glucose Transporter Type 1; D009004:Monosaccharide Transport Proteins; D012333:RNA, Messenger; C495677:SLC2A1 protein, human; D012313:RNA; D004247:DNA; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing); D005947:Glucose