1. Y axis of graphs in A and B should have the same number of decimal places. Remove two zeros from y axis of graph A.
2. Would it be possible to normalize the levels of Aβ and sAPPα towards the levels of a canonical secreted protein or soluble fragment of a plasma membrane protein processed in similar ways as APP, respectively, not affected by DMSO and DAPT (in graphs A and B)? Without these controls, it is difficult to anticipate whether changes in graphs A and B might translate DAPT changing secretion and affecting secretases activity in general or whether the observed effects are entirely specific for Aβ and sAPPα.
3. Some "statistical significance" asterisks are missing in graphs E, F and G.
4. Knockdown efficiency of each siRNA should be provided.
5. Is it significant the difference DMSO vs. DAPT in ADAM10, ADAM17 and ADAM10+ADAM17 knockdown conditions? By eye a reasonable difference seems to exist indicating that DAPT is still able to increase sAPPα when ADAM10/17 are depleted from cells. This suggests that DAPT might influence other mechanisms apart from affecting α-secretase activity. However, the existence of residual activity of some ADAM10/17 remaining in cells after their knockdown, or the existence of other ADAM proteins coping for the lack of ADAM10/17, cannot be excluded as possible explanations for the observed effect.
1. Why did the authors use 10 µM of DAPT and not a lower concentration?
2. Why did the authors treat HeLa cells for 4 hours in (A), HEK cells for 24 hours in (B), and HeLa cells for 24 hours in (E) and (F) with DAPT? Why the difference?
3. Data of the viability test mentioned in (A) and (B) should be included.
4. The legends should include the statistical test and number of samples analyzed.