The Dutchman - scorer of his side's second in their dramatic last-16 second-leg win over Inter Milan - could not complete the full training session on Saturday due to a knock, the seriousness of which has not been ascertained.
However, Depay is just one of three concerns for Simeone to take into account for Wednesday, as Mario Hermoso is struggling with a thigh issue, while Thomas Lemar is on the mend from a ruptured Achilles.
With Depay likely taking his place on the bench either way, Antoine Griezmann and Alvaro Morata are primed to link arms in Simeone's two-man strikeforce, and the hosts' three-man midfield also picks itself.
Indeed, Rodrigo de Paul, Koke and Marcos Llorente are expected to line up in the engine room, with Nahuel Molina and Samuel Lino operating as the widest men in the 3-5-2 system.
Former Borussia Dortmund man Axel Witsel will likely reunite with his old employers as part of Jan Oblak's rigid defensive shield, which should also comprise Jose Gimenez and Reinildo, although Cesar Azpilicueta is a viable alternative.
Atletico Madrid possible starting lineup:
Oblak; Witsel, Gimenez, Reinildo; Molina, Llorente, Koke, De Paul, Lino; Griezmann, Morata