After his retirement from football on 2008, Anastasiou joined Panathinaikos as assistant on ten Cate side. On 2011 he left Greece for Netherlands as assistant coach for Jong Ajax, reserve team of Ajax. After two years, Anastasiou joined English club Reading in January 2013 as a first team coach; he had first met manager Brian McDermott in June 2012 whilst completing his UEFA Pro License. His stay at Reading lasted just two months, and he left the club following the departure of McDermott on 11 March 2013.
He was named manager of Panathinaikos in May 2013. On February 2015 was named Coach of the Year 2014. On May 24, Anastasiou managed his 100th game at the side of Panathinaikos, against Atromitos.