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Here are some pictures of human iPS cells in culture. After several passages, some cells look really weird, can you help me understand what is going on? What am I looking at? What are those blastocyst-like structures? (bluish picture) and what about that cell full of vacuole-like thing?

P.S. do not pay attention to the scale bar, it's incorrect.

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Extra info:

I did not induce them, I got them already dedifferentiated so I can't be specific on that part of the protocol. However, I am growing them in Essential 8 Flex Medium (with supplements as specified from the kit protocol), I add rock inhibitor 4h before passaging them. Passages are done by dissociating colonies using EDTA only (no enzymes). Most of the expanded cultures look normal, some showed the formation of embryonic bodies (the last picture added) but the blastocysts are very different and not really expected.

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    $\begingroup$ I am not entirely sure but there is some vacuole formation so maybe they are going towards the apoptosis. However to see clear phase contrast between Feeder layer and iPS cells the light diffraction may give you hallow formation around the radius of the cells. (Source: Dad) $\endgroup$ – SamD97 Sep 21 '16 at 15:06
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    $\begingroup$ What further changes on them happened later-on? $\endgroup$ – Always Confused Sep 22 '16 at 18:44
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    $\begingroup$ This question would be easier to give an answer to if you provided specifics on your protocol for inducing and culturing iPS cells, any variations from typical protocols, any experimental treatments, etc. The greatly elevated vacuole formation suggests cytotoxic stress. Elevated autophagy can be cytoprotective if the cell is moderately stressed, but it is a precursor to apoptosis when the stress cannot be alleviated by autophagy. Depending on the scale (please clarify), the blue picture may depict an "embryoid body" (essentially a blastocyst) - see "identity" section in your own wiki link. $\endgroup$ – Demosthenes' pars triangularis Sep 22 '16 at 20:28
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    $\begingroup$ I did not induce them, I got them already dedifferentiated so I can't be specific on that part of the protocol. However, I am growing them in Essential 8 Flex Medium (with supplements as the kit specified), I add rock inhibitor 4h before passaging them. Passages are done by dissociating colonies using EDTA only (no enzymes). Most of the expanded cultures look normal, some showed the formation of embryonic bodies (the last picture added) but the blastocysts are very different and not really expected. $\endgroup$ – alec_djinn Sep 23 '16 at 9:59
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    $\begingroup$ @alec_djinn Just a couple examples, though I may be mistaken. (ref) (ref) $\endgroup$ – CKM Jun 4 '17 at 0:12
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they might be embryoid bodies:

https://en.m.wikipedia.org/wiki/Embryoid_body

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  • $\begingroup$ looks like it, at least in the last picture. $\endgroup$ – alec_djinn Sep 5 '17 at 7:00
  • $\begingroup$ ips cells fascinate me to no end. I wish I could culture them at home. $\endgroup$ – 4D Neuron Sep 5 '17 at 22:52

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