Objective: To investigate the synergistic effects of Chuanxiong-Chishao herb-pair (CCHP) on : To investigate the synergistic effects of Chuanxiong-Chishao herb-pair (CCHP) on promoting angiogenesis promoting angiogenesis in silico in silico and in vivo in vivo. Methods: The mechanisms of action of an herb-pair, Chuanxiong- : The mechanisms of action of an herb-pair, ChuanxiongChishao, were investigated using the network pharmacological and pharmacodynamic strategies involving Chishao, were investigated using the network pharmacological and pharmacodynamic strategies involving computational drug target prediction and network analysis, and experimental validation. A set of network computational drug target prediction and network analysis, and experimental validation. A set of network pharmacology methods were created to study the herbs in the context of targets and diseases networks, pharmacology methods were created to study the herbs in the context of targets and diseases networks, including prediction of target profiles and pharmacological actions of main active compounds in Chuanxiong including prediction of target profiles and pharmacological actions of main active compounds in Chuanxiong and Chishao. Furthermore, the therapeutic effects and putative molecular mechanisms of Chuanxiong-Chishao and Chishao. Furthermore, the therapeutic effects and putative molecular mechanisms of Chuanxiong-Chishao actions were experimentally validated in a chemical-induced vascular insuf actions were experimentally validated in a chemical-induced vascular insuffi ciency model of transgenic zebra ciency model of transgenic zebrafi sh in vivo in vivo. The mRNA expression of the predicted targets were further analyzed by real-time polymerase chain . The mRNA expression of the predicted targets were further analyzed by real-time polymerase chain reaction (RT-PCR). reaction (RT-PCR). Results Results: The computational prediction results found that the compounds in Chuanxiong : The computational prediction results found that the compounds in Chuanxiong have antithrombotic, antihypertensive, antiarrhythmic, and antiatherosclerotic activities, which were closely have antithrombotic, antihypertensive, antiarrhythmic, and antiatherosclerotic activities, which were closely related to protecting against hypoxic-ischemic encephalopathy, ischemic stroke, myocardial infarction and heart related to protecting against hypoxic-ischemic encephalopathy, ischemic stroke, myocardial infarction and heart failure. In addition, compounds in Chishao were found to participate in anti-in failure. In addition, compounds in Chishao were found to participate in anti-infl ammatory effect and analgesics. ammatory effect and analgesics. Particularly, estrogen receptor Particularly, estrogen receptor α (ESRα) and hypoxia-inducible factor 1- ) and hypoxia-inducible factor 1-α (HIF-1α) were the most important ) were the most important potential protein targets in the predicted results. potential protein targets in the predicted results. In vivo In vivo experimental validation showed that post-treatment of experimental validation showed that post-treatment of tetramethylpyrazine hydrochloride (TMP•HCl) and paeoniflorin (PF) promoted the regeneration of new blood tetramethylpyrazine hydrochloride (TMP•HCl) and paeoniflorin (PF) promoted the regeneration of new blood vessels in zebra vessels in zebrafi sh involving up-regulating ESR sh involving up-regulating ESRα mRNA expression. Co-treatment of TMP•HCl and PF could mRNA expression. Co-treatment of TMP•HCl and PF could enhance the vessel sprouting in chemical-induced vascular insuf enhance the vessel sprouting in chemical-induced vascular insuffi ciency zebra ciency zebrafi sh at the optimal compatibility sh at the optimal compatibility proportion of PF 10 proportion of PF 10 μmol/L with TMP•HCl 1 mol/L with TMP•HCl 1 μmol/L. Conclusions Conclusions: The network pharmacological strategies : The network pharmacological strategies combining drug target prediction and network analysis identified some putative targets of CCHP. Moreover, combining drug target prediction and network analysis identified some putative targets of CCHP. Moreover, the transgenic zebra the transgenic zebrafi sh experiments demonstrated that the Chuanxiong-Chishao combination synergistically sh experiments demonstrated that the Chuanxiong-Chishao combination synergistically promoted angiogenic activity, probably involving ESRα signaling pathway.