While the Argentine apparently is not suffering from any physical complaints, he only makes the bench for Wednesday's encounter.
Barcelona put the surprise switch down to a "technical decision" prior to kick-off, but declined to give further information.
Gerard Deulofeu therefore partners Luis Suarez up front, with Paulinho included in a packed Barca midfield to nullify the Juve threat.
The Blaugrana are otherwise at near full-strength, although Lucas Digne is included on the left side of defence and Jordi Alba joins Messi on the bench.
With Giorgio Chiellini ruled out for the hosts, meanwhile, Juventus coach Massimiliano Allegri has been forced to shuffle his defence.
Daniele Rugani starts alongside Mehdi Benatia in the middle, while there is no place on the bench for Benedikt Howedes, who is approaching full fitness after an injury that has delayed his Juve debut.
Juventus: Buffon; Barzagli, Rugani, Benatia, Alex Sandro; Pjanic, Khedira; Cuadrado, Dybala, Douglas Costa; Higuain.
Substitutes: Szczesny, De Sciglio, Asamoah, Marchisio, Matuidi, Bentancur, Mandzukic.
Barcelona: Ter Stegen; Semedo, Pique, Umtiti, Digne; Rakitic, Paulinho, Busquets, Iniesta; Suarez, Deulofeu.
Substitutes: Cillessen, Denis, Messi, Alcácer, J.Alba, Aleix Vidal, Vermaelen.