Ag/TiN-Ag composite multilayers were deposited on Si and Ti-6Al-4V substrates using a multi-target magnetron co-sputtering system. X-ray diffraction (XRD) and transmission electron microscopy (TEM) indicated that the Ag/TiN-Ag composite multilayers exhibited a weaker TiN textures with the smaller modulation period (Λ), which proved the smaller Λ can block the growth of columnar grain and particle, inducing a smoother surface and smaller roughness parameters (Ra: 0.82nm; Rq: 1.2nm). Ag existence in the surrounding TiN in top TiN-Ag composite layer and Ag layer insertion into TiN layer prohibited the emergence and propagation of cracks and prevented the growth and dislocation slip of defects, thus limited TiN grain size in a nanoscale size (5–10nm), which enhanced the hardness and adhesion strength of the multilayers with smaller Λ. The hardness, elastic modulus and critical load reached 35.983GPa, 426.937GPa and 38mN, respectively, when Λ was 10nm. The population of living cells growing on the composite multilayers with Λ=10nm was far higher than those of other tested groups. This work proved the Ag/TiN-Ag composite multilayer with smaller Λ was beneficial for improvement of hardness, adhesion strength and living cells growth.