Quantification of glomerular TGF-β1 mRNA in patients with diabetes mellitus

Quantification of glomerular TGF-β1 mRNA in patients with diabetes mellitus. Transforming growth factor-β1 (TGF-β1) is a primary determinant of the mesangial expansion observed in diabetic nephropathy. In this study, we quantitated the levels of intraglomerular TGF-β1 mRNA in patients with diabetes mellitus using a competitive polymerase chain reaction (PCR) method. Renal biopsy specimens were obtained from 29 patients with non-insulin-dependent diabetes mellitus. Total RNA was extracted from the glomeruli and reverse transcribed into cDNA with reverse transcriptase. To prepare samples containing identical amounts of β-actin cDNA (8 pg), we performed competitive PCR by co-amplifying mutant templates of β-actin with a unique EcoRI site. We also used this competitive PCR method to measure TGF-β1 cDNA by co-amplifying mutant templates of TGF-β1. We observed higher expression of TGF-β1 mRNA in glomeruli of patients with diabetic nephropathy as compared with normal glomeruli. Intraglomerular TGF-β1 mRNA was elevated, even in the early stage of diabetic nephropathy. Moreover, levels of intraglomerular TGF-β1 mRNA correlated with values of HbA1c. These data suggest that hyperglycemia induces intraglomerular TGF-β1 mRNA expression in vivo, and that TGF-β1 overproduction may be associated with the progression of diabetic nephropathy.