Quantitative EPXMA imaging of rapidly frozen kidney proximal tubule primary cultures
The role of ions (elements), particularly calcium, in processes such as renal cell injury is equivocal. A variety of techniques are required not only to assess the total cell element content or cytoplasmic free ion concentration, but also to identify the sites involved in binding, uptake and release of those ions/elements. The need to correlate such changes with alterations in cell metabolism has given rise to the use of in vitro models; for example, isolated nephron segments in suspension have been employed to study the effects of anoxia/hypoxia in the absence of any hemodynamic effects. Our previous studies have shown that changes in Ca compartmentation in isolated rabbit proximal tubules after short-term anoxia appear to be subtle. However, the effects of longer periods of anoxia and/or recovery have not been studied due to lack of an in vitro model. A preparation of primary kidney proximal tubule cultures has been recently described, which accurately reflects the in vivo metabolic features of proximal tubules, for extended periods of time in vitro. We have therefore applied ultrarapid freezing and electron-probe x-ray microanalysis (EPXMA) imaging to characterize the ultrastructure and subcellular element composition of these new primary proximal tubule cultures.
Spencer, AJ; Hawkey, LA; LeFurgey, A; Dickman, KG; Mandel, LJ; Ingram, P
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