OBJECTIVE： To optimize the formulation of Methotrexate thermosensitive hydrogels， and to study the in vitro release property of it. METHODS： Taking PLGA-PEG-PLGA as matrix and mannitol as viscosity modifiers， Methotrexate thermosensitive hydrogels were prepared. Using the absolute value of the deviation of gelation temperature between 35 ℃， its formulation was optimized by orthogonal test using the concentrations of PLGA-PEG-PLGA， methotrexate and mannitol as factors. The dialysis bag method was used to investigate accumulative release rate of Methotrexate thermosensitive hydrogels and methotrexate solution within 336 h in vitro. RESULTS： The optimal formulation was as PLGA-PEG-PLGA of 20％， methotrexate of 0.10％ and mannitol of 0.10％； gelation temperature were 35.1， 35.0， 35.0 ℃. The average drug-loading rate was more than 90％. 12 d accumulative release rate was more than 90％ （r＝3）. Methotrexate solution has been substantially completely relased within 240 h. CONCLUSIONS： Methotrexate thermosensitive hydrogels with sustained-release are prepared successfully， and the preparation technology is simple and practical.