Cellulose nanocrystals (CNCs) grafted with fluorescent and thermo-responsive poly (N-isopropylacryalamide) (PNIPAM) brushes were prepared for encapsulation and the release of 5-fluorouracil (5-FU). The successful grafting was evidenced by Fourier transform infrared (FTIR) spectroscopy and solid-state 13C nuclear magnetic resonance (13C NMR). Differential scanning calorimetry measurements suggested that the lower critical solution temperature of PNIPAM-grafted CNCs is close 32 °C. During polymerization, tuned fluorescence signatures were obtained by varying the dye dosages. At room temperature, the release amount of the loaded 5-FU was about 42% at a pH of 2.1, while this value approached 60% at a pH of 7.4. Both the cumulative release amount and the release rate were greatly increased when the temperature was raised to 37 °C. The novel PNIPAM-grafted CNCs with both fluorescence and stimuli-sensitive properties possess potential for application in intelligent drug delivery systems.