Microplastics (MPs) are currently a global environmental pollutants and health hazards that caused by MPs cannot be ignored. However, studies on MP toxicity in mammals are scare. Here, we investigated the effects of two doses (0.1 mg and 0.5 mg) of 5 µm polystyrene microplastic (PS-MP) particles on the hematological system of mice through traditional toxicology experiments and assessed the related potential biological mechanisms using transcriptome sequencing analysis. The toxicological examinations showed that the 0.5 mg dose significantly decreased white blood cell count, increased Pit count, and inhibited the growth of colony-forming unit CFU-G, CFU-M and CFU-GM. Compared with the control group, there were 41 differentially expressed genes (DEGs) in the 0.1 mg-treated group and 32 significantly changed genes in 0.5 mg-treated group. Of note, eight genes were found to be significantly altered in both the PS-MP-treated groups. Gene ontology analysis showed that DEGs were mainly involved in T cell homeostasis, response to osmotic stress, extracellular matrix and structure organization, and metabolic process of NADP and nucleotides. In addition, pathway analysis revealed that the Jak/Stat pathway, pentose and glucuronate interconversions, nicotinate and nicotinamide metabolism, biosynthesis of unsaturated fatty acids, and the pentose phosphate pathway were involved in PS-MP-induced toxicity in mice. These results indicated that PS-MP exposure can cause hematotoxicity to some extent, impact gene expression, and disturb related molecular and biological pathways in mouse bone marrow cells. Our study provides fundamental data on the hematotoxicity of PS-MPs in terrestrial mammals that will help to further assess the corresponding health risks in these mammals.