These are what we call the 'YMMV items.' Things that some people find in this work. We call them 'your mileage might vary' because not everyone sees these things in the same way. This starts discussions in the trope lists, a thing we don't want. Please use the discussion page if you'd like to discuss any of these items.
YMMV: Better Living Through Science and Ponies
Dude, Not Funny!: GLaDOS identifies Ditzy's speech disorder and "derpy" eyes as being unrelated to her intelligence, and says she's actually not stupid at all, she's above average even. But since she can't express herself properly, the other ponies must think of her as a mentally handicapped moron unfit for any task more complex than a mailmare because she's too stupid to do anything else. This can hit way too hard if you happen to be the precise type of person Ditzy is explained as.
Moral Event Horizon: Acknowledged in-story. Chell has managed to leave Apeture behind her and is now living a happy, glorious life as a pony in Ponyville. When GLaDOS still manages to ruin her life and abduct all her friends, Chell decides that she's finally crossed the line.
What she does to Ditzy Doo is arguably even worse. Ditzy is able to deal with all of the verbal abuse aimed at her, but when GLaDOS starts mocking her for having a daughter, Dinky, and telling her that Dinky's probably inherited the conditions that make Ditzy stand apart from other ponies, that's when Ditzy's heart splits in half.
All of GLaDOS' interactions with the ponies are a quite interesting look at how much worse her usual taunts are when directed at someone other than Chell.