QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

Help viewing high resolution images Return to article

Fig. 4. Effect of FCCP, TNF and isoproterenol in the expression of ATGL, HSL and perilipin A. (A) Quantitative gene expression analysis by real time RT-PCR of ATGL in differentiated 3T3-L1 (DIFF) incubated for 1 (white columns) or 3 (black columns) days with 10 ng/ml TNF, 0.5 µM FCCP or 10 µM isoproterenol. For data normalization, TBP was used as reference gene. Results are expressed in relative fold increase as means ± s.d. for n=3 experiments, ^***P<0.001 and ^**P<0.01 significantly different from DIFF. (B) Western blot analysis for HSL, ATGL and perilipin A abundance was performed using the same conditions as above. (C) HSL phosphorylation (Ser563 and Ser660) status and abundance were analyzed after short incubation times using the same conditions as above. Equal protein loading was controlled by the immunodetection of -tubulin in all western blot experiments. Abundance and localization of HSL and perilipin A in adipocytes. (D,E) Differentiated cells (DIFF) were incubated on coverslips for 3 days with or without 10 ng/ml TNF, 0.5 µM FCCP or 10 µM isoproterenol before immunostaining and confocal microscopic analysis. At the end of the incubations cells were fixed, permeabilized and then incubated with the specific antibodies (green fluorescence) against HSL (D) and perilipin A (E). For nuclear staining TOPRO-3 was used (blue fluorescence).

Return to article