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Fig. 9. Effect of tomosyn-1 silencing on the pool of secretory granules docked at the plasma membrane. INS-1E cells were grown on glass coverslips coated with laminin and poly-L-Lysine, and were transiently co-transfected with a plasmid encoding IAPP-Emerald and the pSUPER vector siRNA-i (control) or siRNA-a. Three days later the transfected cells were visualized by TIRF microscopy (A). Single IAPP-Emerald-EGFP-positive granules from nine control cells and nine cells transfected with siRNA-a were manually selected and counted. The number of IAPP-Emerald-EGFP-positive granules was then divided by the single cell area. (B) Results of the experiments in from A, expressed as means ± s.e.m. (C) Six control cells and six cells transfected with siRNA-a were incubated in the presence of stimulatory concentrations of glucose, K+, forskolin and IBMX. Single IAPP-Emerald-EGFP-positive granules were manually selected and counted after 0, 8 and 16 minutes of incubation in the presence of the stimuli. Data are expressed as means ± s.e.m. *P<0.01, significantly different (unpaired t-test).
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