High-cycle fatigue test after multiple overloading was performed to develop a structural integrity assessment method for hydrogen utilization machines suffered seismic load. To simulate the material used in hydrogen gas for a long period, the hydrogen charged specimen whose hydrogen concentration was 80ppm was used. Three kinds of notched specimens with different notch root radii were used. The fatigue test in air using uncharged specimen was also performed. The fatigue limit of p = 0.2mm specimen tested in air was decreased to almost one half by the application of 200 cycles overload with the amplitude of 1.25ω 0.2. The decrease was caused by the microcracks generated by the overload. The number of cycles of overload corresponded to only 0.67% of the low-cycle fatigue life for the overload. Since no reduction of the fatigue limit was observed by the overload of 0.75ω 0.2, there is a threshold value of overload that does not cause the reduction of high-cycle fatigue strength. The fatigue strength of hydrogen charged specimen tested in hydrogen gas was lower than that of uncharged specimen tested in air. Deeper microcracks were formed by the overload in the hydrogen charged specimen than in uncharged hydrogen.