Vimentin and CD10/CALLA expression in podocytes. (A) Western blot analysis for vimentin expression in HGEC. Total protein was extracted from cells cultured in the presence of 5 mM (control) (lane 1) or 25 mM (lane 2) glucose. Lane 3 represents vimentin expression in HGEC continuously exposed to 25 mM and reverted to 5 mM glucose for 18 weeks (HGEC:25 mM-to-5 mM/18w). Lanes 4 and 5 represent HGEC continuously exposed to 5 mM glucose and reverted to 25 mM for 6 and 18 weeks respectively (HGEC:5 mM-to-25 mM/6w, HGEC:5 mM-to-25 mM/18w). Lanes 6 and 7 represent HGEC:25 mM/6w-to-5 mM/4w and HGEC:25 mM/18w-to-5 mM/4w respectively. For quantification of vimentin the 57 kDa band was used. Blots were reprobed with anti-tubulin antibody, to verify protein loads, against which the data were normalized. (B) Results were expressed as the mean ± SD of three independent experiments. (C) Representative experiments from flow cytometric analysis of CD10/CALLA. Thick grey lines represent HGEC:5 mM, thin grey lines represent the corresponding isotype control antibody and black lines represent stimuli. (D) Percentage of positive cells was calculated for the histogram section indicated as M1 (**p<0.01 vs. HGEC:5 mM).
Tsotakos et al. BMC Cell Biology 2013 14:28 doi:10.1186/1471-2121-14-28