Stereological Estimation of Ito Cells from Rat Liver Using the Optical Fractionator - a Preliminary Report

In the last two decades, much light has been shed on hepatic fibrosis, and the activation / proliferation of Ito cells (IC) emerged to play a central role. Therefore, it is essential to have solid quantitative data in nonpathological statuses; yet, this data is scarce and confined to "number per area" or semiquantitative information. Moreover, the supposed heterogeneous distribution of IC in the hepatic lobule was never analysed with design-based (unbiased) stereology. In the present study, the total number (N) of IC in rat liver was estimated for the first time, by combining immunocytochemistry with the optical fractionator. Quantification was extended to the hepatocytes, to disclose the IC index, an often-used ratio in hepatology. Systematic uniform random liver sections were obtained from male Wistar rats (n = 3), and immunostained against glial fibrillary acidic protein (GFAP), a known specific marker for hepatic IC. For the first time, these were marked against GFAP in thick (30 μm) paraffin sections. The estimated N of IC was 224E; with a coefficient of error of 0.04 or 0.06, depending on the particular equation used (based on the so-called "quadratic approximation"). The IC index was 91 IC/1000 hepatocytes. Concerning the lobular heterogeneity, it was proved the liver harbours a larger total number of periportal IC and hepatocytes.