Holden sidelined with anterior cruciate ligament tear
The 27-year-old picked up the injury during the United States' 1-0 victory in the CONCACAF Gold Cup final against Panama on Sunday.
Holden recently signed a new contract and the Reebok Stadium and it is not yet known how long he will be sidelined for by his most recent injury.
Bolton's head of sports performance Mark Leather told the club's official website: "We've got the scans and the results will be analysed by our specialists.
"We can then put an action plan in place over the next week to 10 days when the initial swelling reduces."
Holden has been badly affected by knee injuries over the course of the past two years.
He first injured his knee in a Premier League fixture with Manchester United in March 2011 that would keep him out of action for six months.
Upon his return he required further surgery that sidelined him for 16 months.
Holden made a long-awaited return to action in January before making four appearances during a short loan spell at fellow Championship side Sheffield Wednesday.