Partially cholesterol-substituted 8-arm poly(ethylene glycol)-block-poly(L-lactide) (8-arm PEG-b-PLLA-cholesterol) has been prepared as a novel star-shaped, biodegradable copolymer derivative. The amphiphilic 8-arm PEG-b-PLLA-cholesterol aqueous solution (polymer concentration, above 3 wt%) exhibits instantaneous temperature-induced gelation at 34 °C, but the virgin 8-arm PEG-b-PLLA does not, irrespective of concentration. Moreover, an extracellular matrix (ECM)-like micrometer-scale network structure has been created with favorable porosity for three-dimensional proliferation of cells inside the hydrogel. This network structure is mainly attributed to specific self-assembly between cholesterol groups. The 10 and 20 wt% hydrogels are eroded gradually in phosphate buffered saline at 37 °C over the course of a month, and after that the gel becomes completely dissociated. Moreover, L929 cells encapsulated into the hydrogel are viable and proliferate three-dimensionally inside the hydrogels. Thus, in-vitro cell culture studies demonstrate that 8-arm PEG-b-PLLA-cholesterol is a promising candidate as a novel injectable cellular scaffold.