The Hatter's emotional state. He's literally dying of despair until Alice tells him the Red Queen has his family. This is especially true when Alice comes to see him after her time-travel adventures. He's in bed motionless and almost completely white. The Underlanders are gathered around him, convinced he's on his deathbed. Even after Alice begins talking to him, it takes him such a while to respond that the viewer might almost be convinced he's not going to recover.
And just prior to that, when Alice arrives in Underland, she's taken aback to see her friends are gathered and in a tremendously somber mood. The White Queen is weeping over the Hatter's condition. Remember, it was the Hatter who literally saved the White Queen on the day she lost her crown and he lost his family; she might have been killed if (as the first movie showed) he had not guided her horse away from the carnage. They have been close for a very long time and she's crushed by what's happening to him.
Iracebeth hitting her head as a child. While the modern-day Red Queen is not a very sympathetic character, it's surprisingly pitiful to see her as a little girl, lying on the snowy street and crying softly about how her head hurts.
Then there's young Mirana's reaction, how she watches in complete silence, tears in her eyes and knowing what happened was her fault.
The argument between Tarrant and his father Zanik following the disastrous coronation, also doubles as a Parting-Words Regret:
Zanik: If you are to be a hatter worthy of the Hightopp name, you must be sane, sober, disciplined. Everything you now are not! Tarrant: All I did was laugh, father. Couldn't help it. Her head is rather voluminous... Zanik: You cost the princess her crown! Do you know what this means to us? Tarrant: Why am I never good enough for you? Zanik: Why are you always such a disappointment to me? Tarrant:[beat] There. You've said it. You've said it, father. Well, if I'm such a disappointment, I don't suppose you'll be sorry if I leave home. Tyva: No, Tarrant... Tarrant: My hat please, mom. Tyva:[reluctantly hands Tarrant his hat] Tarrant... [Tarrant walks out]
Near the end of the film, as the whole of Underland turns to rust, the "animal" gang runs out of room to run away. As they say their goodbyes to each other, Bayard jumps in front of his friends to defend them at the last second, snarling in defiance as he is the first to be turned.
Cheshire simply smiling and quipping "Time's up..." as he's consumed by the rust. Not helped at all by the fact that it's perfectly in-character for him.
Mirana refuses to leave the frozen form of her sister behind and is turned to rust right along with her.
