Knee osteoarthritis is a chronic disease caused by the deterioration of the knee joint due to various factors such as aging, trauma, and obesity, and the nonrenewable nature of the injured cartilage makes the treatment of osteoarthritis challenging. Here, we present a three-dimensional (3D) printed porous multilayer scaffold based on cold-water fish skin gelatin for osteoarticular cartilage regeneration. To make the scaffold, cold-water fish skin gelatin was combined with sodium alginate to increase viscosity, printability, and mechanical strength, and the hybrid hydrogel was printed according to a pre-designed specific structure using 3D printing technology. Then, the printed scaffolds underwent a double-crosslinking process to enhance their mechanical strength even further. These scaffolds mimic the structure of the original cartilage network in a way that allows chondrocytes to adhere, proliferate, and communicate with each other, transport nutrients, and prevent further damage to the joint. More importantly, we found that cold-water fish gelatin scaffolds were nonimmunogenic, nontoxic, and biodegradable. We also implanted the scaffold into defective rat cartilage for 12 weeks and achieved satisfactory repair results in this animal model. Thus, cold-water fish skin gelatin scaffolds may have broad application potential in regenerative medicine.