Dear colleagues,
I've recently found an interesting paper (PMID 21763280) on the phenotypes of necrotic and apoptotic cells. As seen from Figure 1, cells treated with TNF and CHX undergo either apoptosis or necrosis when treated with either necrostatin-1 or zVAD, respectively. That's what everyone would expect. But surprisingly, when TNF/CHX-treated cells are exposed to both zVAD and necrostatin-1, their appearence seems to be normal, they do not seem to undergo any other type of cell death (autoschizis would be suitable, wouldn't it?). How would you explain the results? How do the cells cope with TNF/CHX? What would happen to them later? I don't really see a good reasonable explanation for the cells to live later on when treated with CHX.
Dear Peter,
About the interpretation of figure1: the addition of Nec1 to TNF+CHX has no additional effect on the outcome of TNF+CHX alone; Nec-1 does not block apoptosis in these conditions. Furthermore, i'm happy with the explicit publication of the annexin-staining not being reliable for distinguishing between apoptosis/necrosis.
Regarding your question, the combination of TNF+CHX+zVAD+Nec-1 together, it seems like cell death is indeed completely blocked at the investigated time point. The question is indeed, what happens to these cells later on, do they die at all? I think teh CHX-treatment will become toxic by itself over time, as we see in other cell types.
Regarding the block of necrosis, we have seen similar effects in L929 cells, where TNF by itself (no need for zVAD) induces necroptosis after 4-6 hours and the addition of Nec1 blocks necroptosis at 4-6 hours and later. However, at 12-14 hours post stimulus, the cells treated with TNF+Nec1 will start to die slowly, and this cell death is associated with caspase activity. It seems like the cells are stressed and poised to die, but since their canonical pathways are blocked, active cell death signaling cannot proceed. Over time, the cellular stress associated with death receptor activation and blocking canoncal cell death pathways will trigger the cells to die anyway, possibly through activation of ROS.
Similar effects are observed if MLKL or RIPK3 is knocked-down: a delayed switch to apoptosis, which is dependent on RIPK1-kinase activity (see Remijsen et al , CDD 2014, PMID 24434512). We have not identified a molecular mechanism behind this, but it is likely a switch to a different subtype of programmed cell death occurs (for a review, see vanden Berghe, Nat Rev Mol Cell Biol,2014, PMID 24452471)
Hope this helps,
sasker
Our article published in peer-reviewed Journal "Communicative & Integrative Biology". A few major points discussed in the paper:
(1) Brain is not the source of consciousness.
(2) Consciousness is ubiquitous in all living organisms, starting from bacteria to human beings.
(3) The individual cells in the multicellular organisms are also individually cognitive entities.
(4) Proposals like “artificial life”, “artificial intelligence”, “sentient machines” and so on are only fairytales because no designer can produce an artifact with the properties like internal teleology (Naturzweck) and formative force (bildende Kraft).
(5) The material origin of life and objective evolution are only misconceptions that biologists must overcome.
http://www.tandfonline.com/doi/full/10.1080/19420889.2015.1085138
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