10. Killing Her Oldest Friend - Black Widow (Vol. 5) #20
As an assassin trained from childhood to love nobody and kill everyone, it's safe to say that the Black Widow was somewhat lacking in the friend department for most of her life. There was one exception to this rule, however: that of Marina, a girl who Natasha trained alongside in the Red Room, and with whom she formed a firm friendship.
So, obviously, the end of the fifth volume of Black Widow has Natasha being forced to kill Marina, as her superiors felt that the woman had grown too fond of her undercover life in Cuba and thus would betray them. Despite knowing that it would be denying her friend a long-deserved chance at happiness - and being aware that there was a very good chance Marina could successfully change identities instead - she shoots her in the shower, with an expression that suggests that she also knows this is a terrible thing to have done.
Just to emphasise that this isn't a heroic move, she then kills the cat, despite the fact it notably lacks the vocal faculties required to grass on the assassin.