The 28-year old American midfielder has hardly featured for the Trotters over the past two years and was again struck down when attempting to make a comeback on Monday night.
"It's disappointing news for us all," Freedman told reporters. "We don't know the exact results (of the scan) at the moment, but we feel that it's the same injury.
"He's going to fly back to America to see his specialist on Monday. Hopefully, he can read his scans and tell us good news, but if not he will need to go into the knee and have a look at it.
"He's very positive and no matter what the outcome of this is, he will carry on and keep on going.
"I want to be very clear to everybody that Stuart Holden will be here in some capacity at this football club - he is such a fantastic lad and is a great ambassador for this football club.
"Stuart will hopefully be fine and will be back playing, but if he's not then he'll be back here during the summer trying to get fit to get back in some capacity."
Holden sustained a serious knee injury while playing against Manchester United in 2011 - the season he won Bolton's Player of the Year award - and the problem has haunted him ever since.
He suffered another knee injury while playing for the US at last summer's CONCACAF Gold Cup and has been restricted to less than 20 appearances in all forms of the game since the summer of 2011.
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