Tsitsipas, the 2019 champion, was trailing 2-1 in the opening set when he had a lengthy conversation with his trainer before slowly getting to his feet, shaking his head and walking over to shake Rune's hand.
That was after just 17 minutes of play and led to a chorus of jeers and whistles from the disappointed crowd in Turin — which instead had to settle for an exhibition match between alternates Taylor Fritz and Hubert Hurkacz.
Tsitsipas, who lost to home favourite Jannik Sinner 6-4 6-4 on Sunday, had cut short a practice session on Friday due to an apparent physical issue but the Greek player said then he was "absolutely fine."
Speaking at a press conference later, Tsitsipas revealed he has been struggling with a back problem and is withdrawing from the tournament, with Hurkacz set to take his place.
"My apologies to all the fans and the crowd that came to support me today and watch the match," he said.
"I'm really gutted that I wasn't able to finish the match. It's a shame also not to give that spot to someone else to at least try and do something with it. My doctors and the countless visits that I had in the last few days suggested that I play, gave me the green light to go and try it for myself.
"Unfortunately, I felt terrible on the court. I did what I could do in the best possible way to be ready and fit for this match, but it didn't work out for me. It kills me not to be able to finish this tournament, the one that I've prepped for for so long."
It counts as a first victory at the tournament for 20-year-old Rune, who was edged out by Novak Djokovic in his opening match.
"It was very unfortunate," said Rune. "I think we could all see in his first service game that he wasn't serving anywhere near full.
"For sure I knew something was up. We play a long season and it's important to take care of the body. I wish him all the best."
with PA