The microstructural evolution at the fracture surface in response to very-high-cycle fatigue (VHCF) under stress ratios (R) of −1 and 0.5 in an AlSi10Mg alloy produced by Laser Powder Bed Fusion was investigated. The results show that appreciable growth of the Si precipitates at the cellular network boundaries in the as-built microstructure was observed under R = −1. Moreover, significant amorphization of the initial crystalline Si precipitates occurred in the vicinity of the fracture surface under this condition. A layer of fine Al grains was developed in the “fish-eye” region of the fracture surface. These microstructural responses are rationalized by the generation of lattice defects including dislocations and sub-grain boundaries during cyclic pressing of crack surfaces under R = −1, which mediates the Si solute diffusion and re-precipitation in the alloy as well as the amorphization of initial Si crystalline precipitation. In contrast, far fewer dislocations were observed near the crack surfaces under R = 0.5, which is attributed to the absence of cyclic pressing of crack surfaces and severe plastic deformation in this scenario. This work provides insights into the stress ratio dependence of the microstructural evolution in the fatigued Al alloys. The obtained knowledge is useful for future understanding of the fatigue failure in Al alloys produced by additive manufacturing.