Inferred Holocaust: Never mind all the researchers that GLaDOS gassed; "Bring Your Daughter To Work Day" implies that their children became victims too, or were tested to death. GLaDOS also manages to sneak in a reference to the multiple disasters of the Half-Life universe during the Final Battle and in "Still Alive". It's implied during Portal 2 that Chell herself was the daughter of one of the scientists at Aperture and she was forcibly taken as a test subject by GLaDOS during her massacre.
The "High-Energy Pellet" is the same sprite and sound effects as the Combine's energy orbs. Its effect on living beings is pretty much the same as well, although Gordon's suit seems to block its damage.
Chell's leg-springs were part of an unused model for an enemy in Half-Life 2.
The sentry turrets have largely the same behavior as Half-Life 2's combine turrets.
Throw It In: In this interview, Word of God reveals that the blood splatters weren't meant to be in the game at all. However, they weren't even noticed until too late in development, and were so ingrained into the engine that, by that point, it would have been better to just leave them in the game. Valve went out of their way to remove the blood splatters for the sequel, however, hence its lower rating compared to the original.
In earlier concepts for a sequel to Portal, Cave Johnson was positioned to play a significantly larger role, including being subject to Brain Uploading (which, in the final release, he wanted, but he likely died before it could happen), and undergoing an existential crisis concerning the fact.