Raynaud's phenomenon (RP) is an episodic vasospasm of the peripheral arteries caused by an exaggerated reaction to cold temperature or emotional stress. Restoring the angiogenesis capability of the acral lesional skin is a critical strategy to treat RP. Local injection of botulinum toxin-A (BTX-A) has also been reported for treatment of RP. However, since the exact mechanisms of BTX-A action are still unclear, its administration for treatment of RP is not widely used. In the present study, BTX-A was found to promote angiogenesis and relieve RP in the patient. To elucidate its mechanisms against angiogenesis, BTX-A was used to treat human umbilical vein endothelial cells (HUVECs), one of the most popular in vitro models of angiogenesis, and RNA sequencing was used to investigate differentially expressed genes. A total of 413 genes were upregulated, and 1634 were downregulated, with fold-changes >2.0 in HUVECs treated with BTX-A. Gene ontology annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis showed BTX-A affected expression of angiogenesis-associated, angiogenesis-associated signaling pathway-related, metabolic pathway, and epigenetic regulation-related genes. These results demonstrate potential biomarkers of BTX-A action, thereby providing potential therapeutic mechanism(s) by which BTX-A relieves RP symptoms.