The effect of 2,3,4-trimethylpentane on the ultrastructure of proximal tubular cells in primary cell culture

Abstract

To aid in the assessment of the risk of Air Force personnel working with hydrocarbon fuels and compounds, an attempt was made to further characterize the nephropathy that results from exposure to hydrocarbons. The purpose of this study was to isolate and establish purified primary cultures of male rat proximal tubular cells suitable for experimental exposure to sublethal concentrations of solubilized 2,3, 4-trimethylpentane (TMP), a model hydrocarbon. Experiments were conducted to evaluate the cytotoxicity and metabolism of solubilized TMP in media containing or lacking the protein albumin.Proximal tubule cells in primary suspension culture were exposed to one of the following levels of TMP: 7.9, 12.0, 15.7, 19.1 or 25.5 mM. After 4 hours of exposure, pelleted cells were fixed for transmission electron microscopy by resuspension in 2% glutaraldehyde and 2.5% paraformaldehyde in 0.1M cacodylate buffer at pH 7.4. After a minimum fixation of at least 24 hours, the cells were post-fixed with 2% osmium tetroxide in 0,1M cacodylate buffer at pH 7.4. Cells were processed into Polybed 812 plastic capsules. Sections one micron thick, were cut in order to verify that cells were intact and suitable for thin sectioning. Thin sections (60- 90 nm) were cut on an ultramicrotome using a diamond knife. Thin sections, stained with uranyl acetate and lead citrate, were examined with a transmission electron microscope at 60 kV. Photographs of representative proximal tubule cells were taken at three levels of magnification.