There has been a widespread and rapid rise in annual average temperature over the Korean peninsula, for example, 0.41℃ from 1981 to 2010 and 0.5℃ from 2001 and 2010. Particularly, subtropical weather such as humid and high temperature in Korea affects the productivity of Holstein dairy cows that are more sensitive to heat stress. miRNA profiles are widely used to investigate physiological changes in dairy cows exposed to heat stress (HS), however the profiles were limited to economic traits including milk production., In this study, we analyzed miRNA expression from whole blood and compared the expression between HS and non-HS cows to understand regulation of biological process in HS cows. The whole blood was collected from the jugular vein of the same cows (n=9) at summer (THI >86; average rectal temp., 39.6℃) and autumn (<63; 38.1℃ ), respectively. Further, decline in milk yield was recorded in the cows during HS condition, To analyze the profiles of miRNAs, RNA-seq was performed on an Illumina Hiseq 2500 sequencer. The data identified 79 differentially expressed miRNAs that were significantly changed between HS and non-HS cows (fold change≥1.5, P<0.05); 43 miRNAs were up-regulated (for example, bta-miR-15b, bta-miR-19a, bta-miR-19b, bta-miR-20a, bta-miR-29c, bta-miR-301a, bta-miR-301b, bta-miR-374b, bta-miR-199b) and 36 miRNAs down regulated in HS cows. Target- gene analysis using miRmap DAVID showed that the selected miRNAs might be associated with Leukocyte transendotheical migration, adherens junction, chemokine signaling pathways. Interestingly, splicesome, base excison repair, and TGF-beta and Glucagon signaling pathway were also detected in the analysis. In summary, our study provides expression profiles of miRNAs that are differently responded to HS and the profiles can be used to understand physiological changes and as potential markers for evaluation of HS cows.