Our challenge is to clarify the relationship between crack roughness and microstructure. Roughness-induced crack closure (RICC) is known to be one of the main factors decelerating fatigue crack growth. Two factors trigger RICC: 1) nanometer-scale roughness on the crack surface (nano-roughness) and 2) degree of crack deflection (micro-roughness). These factors affect the friction stress acting on the crack planes and the stress intensity factor range for crack closure. For instance, S. Suresh and R. O. Ritchie discussed the effects of geometrical mismatch between fatigue crack planes on crack closure. We further attempt to measure multi-scale crack roughness to quantitatively estimate RICC with respect to both friction and crack closure effects. In this study, we examine the multi-scale crack roughness of a lamellar-structured Fe-9Mn-3Ni-1.4Al-0.01C steel sample as a case study. We verify the effect of crack surface friction. Here, we assume that the basic effect of nano-roughness on friction is significant when the inclination angle of micro-roughness against the loading direction is less than the angle of nano-roughness. If the inclination angle of micro-roughness against the loading direction is larger than the angle of nano-roughness, the nano-roughness effect does not occur because the crack surfaces do no contact with each other. To further discuss the underlying effects of multi-scale roughness, we will present more details on the microstructure- and mechanical condition-related roughness parameters and their quantification techniques.