The 35-year-old American was hit by a car while training south of the Italian seaside resort of Rimini on Wednesday and was rushed to hospital with severe chest and head injuries.
“Nicky Hayden's clinical condition remains extremely critical,” said a statement from the Maurizio Bufalini Hospital in Cesena.
“The young man, who is still in the intensive care unit of Cesena's Bufalini Hospital, has suffered a serious polytrauma with subsequent serious cerebral damage.”
Hayden, nicknamed the “Kentucky Kid”, won the 2006 MotoGP world title before moving to the Superbike World Championship for the 2016 season.
His Honda team said that as well as having his fiancée Jackie by his side, Hayden has now been joined by his brother Tommy and mother Rose who arrived in Italy from the United States earlier on Thursday.
Hayden won just three grands prix in MotoGP – all in 2005-06 – but he famously took the world title that season after fighting it out with Valentino Rossi.
Nine-time world champion Rossi took time out from preparing for this weekend's French MotoGP at Le Mans to pay tribute to Hayden.
“Nicky is one of the best friends I have had in the paddock,” said the Italian, who lost the 2006 title to his friend by just five points.
“We were teammates in his rookie year in 2003, when he was a young guy making his debut in his first European experience.
“After a few years we fought for the title down to the last race in Valencia and, unfortunately for me, he won and became MotoGP world champion. After the race we shook hands and hugged.”
Rossi and Hayden later became teammates at Ducati. Wednesday's accident in Italy came just under a month after veteran Italian professional cyclist Michele Scarponi was killed after being hit by a van while out on a training ride on roads near his home.