Glomerular epithelial alterations resulting from sialic acid surface coat removal

PM Andrews - Kidney international, 1979 - Elsevier
PM Andrews
Kidney international, 1979Elsevier
Methods Fisher F inbred rats were used throughout the course of this investigation. The
kidneys from these rats were prepared in vitro study by techniques de-scribed in detail in
another investigation [13]. Briefly, rat kidneys are cleared of blood by in situ vascular
perfusion of oxygenated, normothermic (370 C) Tyrode's balanced salt solution (TBSS)[15].
The kidneys then are excised from the body, cut into thin slices (1 mm), and immersed in
oxygenated normothermic TBSS or Medium 199 with Earle's salts (Gibco) for various timed …
Methods
Fisher F inbred rats were used throughout the course of this investigation. The kidneys from these rats were prepared in vitro study by techniques de-scribed in detail in another investigation [13]. Briefly, rat kidneys are cleared of blood by in situ vascular perfusion of oxygenated, normothermic (370 C) Tyrode's balanced salt solution (TBSS)[15]. The kidneys then are excised from the body, cut into thin slices (1 mm), and immersed in oxygenated normothermic TBSS or Medium 199 with Earle's salts (Gibco) for various timed intervals. Glomeru-lar epithelial cells exposed on surfaces of kidney slices can be maintained routinely in the above manner for 12 hours without any resulting degeneration or major alteration of their fine structure [13]. For each kidney studied in vitro, the first slice cut was fixed immediately by immersion in phosphate buffered 3% glutaraidehyde (pH, 7.2; 410 mOsm). The remaining rat kidney slices were immersed in either 150 ml of oxygenated normothermic TBSS (controls) or in 150 ml of oxygenated normothermic TBSS containing 0.067 U/mi neuraminidase (Sigma type V prepared from Clostridium perfringens). At time intervals of 15 mm, 30 mm, 1 hour, 3 hours, and 6 hours during incubation, three slices were re-moved from each solution and fixed by immersion in phosphate buffered 3% glutaraldehyde. The above experiment was repeated twice with TBSS and once with 199 medium plus Hepes buffer in place of the TBSS.
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