One of the main components of atherosclerotic plaque is the production and accumulation of cholesterol crystals (CCs), which could serve as a biomarker of atherosclerosis. Atherosclerosis is a chronic inflammatory artery disease that is the root cause of myocardial infarction and stroke. Endothelial dysfunction is one of the main contributors to the development of atherosclerosis. The aim of the study was to investigate whether Human Umbilical Vein Endothelial Cells (HUVECs) can uptake CCs and to examine CCs-induced inflammatory response in HUVECs. Using molecular and functional techniques, the distinctive characteristic of CC-mediated immune response was discovered in HUVECs. CCs were mostly taken up by HUVECs by macropinocytosis and phagocytosis. CCs were found to induce Signal Transducer of Activators of Transcription (STAT) 3 phosphorylation and Interleukin (IL)-6 release in HUVEC. In addition, Caspase activation and recruitment domain 8 (CARD8) knockdown drastically reduced CCs uptake and CCs induced IL-6 expression in HUVECs. Moreover, the stem cell growth factor (SCGF)-b protein release was downregulated in response to CCs. IL-1A and Colony Stimulating Factor (CSF) 2 were identified as the topmost hub nodes interacting with all other differentially expressed proteins. A significant increase in neutrophil adhesion on HUVECS was found in response to CCs and conditioned medium from CCs-treated HUVECs. In conclusion, the study findings suggest that the CCs induceSTAT3-mediated IL-6 release and neutrophil adhesion, thereby promoting inflammation in HUVECs.