The number and size of rat tubular cells in diabetic growth, regression, and regrowth using the optical disector

Abstract

Diabetic renal growth, regression and regrowth was studied using stereological methods on perfusion-fixed rat kidneys. A razor blade tissue slicer was used for cutting the entire kidney into 2-mm-thick slices and every second slice was sampled systematically random. Every sampled slice from each kidney was embedded in a single capsule using glycolmethacrylate. From every plastic block one 35-μm-thick section and one 2-μm-thick section was cut and stained with PAS. The volume of kidney cortex was estimated on the thin sections using the Cavalieri principle. Total volume of proximal and distal tubules (V(tub)) were estimated by point-counting on the thin sections at a magnification of 332x using a projection microscope. Using an optical disector (Fig. 1a-d) on the thick sections, the numerical densities (NV) of proximal and distal tubular cells were estimated. The total number of tubular cells: N(total) = NV*V(tub). The mean volume of a tubular cell ( (tub)) including basement membrane and lumen was calculated as: (tub)=V(tub)/N(total). Lumen of the tubular cell was excluded by point-counting using the thin sections and the microscope mentioned in Fig. 1.