Analysis of gene expression in glomeruli of BTBR ob/ob diabetic mice

Although diabetic nephropathy (DN) is the most common cause for end-stage renal disease (ESRD) in western societies, its pathogenesis still remains largely unclear. A different gene pattern of diabetic and healthy kidney cells is one of the probable explanations. Numerous signaling pathways have emerged as important pathophysiological mechanisms for diabetes-induced renal injury. Glomerular cells, as podocytes or mesangial cells, are predominantly involved in the development of diabetic renal lesions. While a lot of gene assays concerning DN are performed with whole kidney or renal cortex tissue, we isolated glomeruli from BTBR ob/ob and wildtype mice at 4 different timepoints (4, 8, 16, 24 weeks) and performed a mRNA microarray to identify differentially expressed genes (DEGs). In contrast to many other diabetic mouse models, these homozygous ob/ob leptin-deficient mice do not only develop a severe type II diabetes, but also diabetic kidney injury with all the clinical and especially histologic features defining human DN. The identified DEGs in diabetic glomeruli were used to investigate biological processes and pathways enriched at different disease stages. Glomeruli were isolated from 4, 8, 16, and 24 weeks old female BTBR wildtype (WT) and BTBR ob/ob (ob) mice. We used 3 animals of each group (WT and ob/ob) at 4 weeks and 4 animals of each group (WT and ob/ob) at 8, 16, and 24 weeks. After cervical dislocation kidneys were explanted and passed through a series of stainless steel sieves (150 µm, 100 µm, 70 µm, 50 µm) to isolate glomeruli. After confirmation of the purity of glomeruli, RNA was extracted for hybridization on Affymetrix microarray. Subsequent RNA extraction was conducted using the RNeasy Mini Kit