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Purpose: DNA damage is one of the main consequences of exposure to ionizing irradiation (IR). Recent studies indicate that IR can modulate the expression of immune system-related genes. However, the effects of IR on the expression of genes and pathways of the B7-CD28 superfamily remain poorly defined. The aim of this study was to evaluate the modulation of genes and pathways related to the B7-CD28 superfamily in response to IR., Materials and Methods: In this study, we used transcriptome data available from the Gene Expression Omnibus (GEO) database to investigate the modulation of the response of genes and pathways of samples of human peripheral blood irradiated with doses of 150, 300, and 600 cGy. The data were obtained at 6 and 24 h after irradiation. The relationship between genes and pathways was established through the Reactome database. The behavior of these pathways was analyzed using mathematical methods based on relative activity and diversity. Analysis of variance (ANOVA) followed by multiple comparisons tests (Bonferroni and Tamhanes) was used to identify differentially expressed genes. Data on transcriptomes were analyzed through ViaComplex V.1.0 and IBM SPSS Statistics 22., Results: For the pathways investigated in this study, we observed that the effects produced by these doses significantly modified the behavior of five pathways associated with the immune system. Also, the dose of 300 cGy might trigger signaling for the activation of T cells through the negative regulation ( p  < .05) of the co-inhibitory PDCD1LG2 gene. Positive regulation caused by 300 cGy ( p  < .05) of the CD80 receptor was observed by us, which might be related to a stimulatory signal. According to our findings, this dose induced the production of cytokines and genes that are associated with the activation and differentiation of T cells., Conclusions: Our findings indicate that the irradiation modulated the organization of the biological system, suggesting that 300 cGy is more efficient in activating the immune system.