Tenocytes were isolated from the rotator cuff tendons of healthy (HT) and glucocorticoid (GC)-treated rats (GCT) and were cultured on polystyrene wells (TCP) as control, and on 2 de-cellularized collagen matrices: porcine small intestinal submucosa (SIS), and human dermal matrix (Graftjacket 1 , GJ). At 3 and 7 days cell proliferation and synthesis were evaluated. Proliferation of HT tenocytes increased between experimental times for both tested membranes, but already at 3 days, HT tenocytes cultured on GJ showed the highest WST-1 value. The collagen-I (CICP) synthesis on GJ membrane did not change between experimental times and was significantly higher than TCP and SIS at 7 days. Proteoglycans (PG), and fibronectin (FBN) synthesis increased when HT were cultured on GJ, between experimental times, and both PG and FBN synthesis on GJ membrane were higher than TCP and SIS at 7 days. GC determined decreases in cell proliferation, CICP and PG syntheses at 3 days of culture on TCP when compared to HT tenocytes while a decrease in WST-1 was maintained at 7 days. CICP, PG and FBN (only at 3 days) syntheses were significantly higher in GCT tenocytes cultured on GJ. The negative effects on GC on GCT tenocytes cultured on membrane were particularly evident on SIS for CICP (À18%) and FBN (À67%) synthesis. The obtained results support the conclusion that GJ is more suitable than SIS as a scaffold for in situ tissue engineering and for the in vitro bioengineering of tendons to heal massive tears of the rotator cuff tendon. ß