Surface exposure of HSP70 and HSP90 in RBAc-PDT-treated HeLa cells.

<p>Kinetic of HSP70 and HSP90 membrane translocation (ecto-HSP70 and ecto-HSP90) was performed by Western blot of electroblotted to nitrocellulose membrane SDS-PAGE of membrane proteins fraction (30 µg protein/lane) of RBAc-PDT treated HeLa cells (10⁻⁵ M RBAc, 1 h, 1.6 J/cm², 90 sec) in the presence of 3-MA (10 mM) and Nec-1 (300 µ) (apoptotic cells) and in the presence of z-VAD-FMK (20 µM) and 3-MA (10 mM) (autophagic cells) at the indicated time intervals. Monoclonal antibodies against HSP70 (70 kDa) and HSP90 (90 kDa) were used. The amount of HSP70 and HSP90 proteins was reported as band intensity ratio (treated/untreated) measured by densitometer analysis. E-cadherin (97 kDa) and β-actin (45 kDa) expression is shown as a control. Actin is absent in the membrane proteins fraction. The data are the mean ± SD of three independent experiments. The values for ecto-HSP70 of the apoptotic cells are always significantly different (p<0.05) with respect to autophagic cells. The values of apoptotic and autophagic cells values are significantly different (p<0.05) with respect to untreated cells for both ecto-HSP70 and ecto-HSP90. One representative Western Blot is shown out of the three independent experiments performed.