<p style="text-align: center;">CHEN Hui , JIN Xin-Yue , SHEN&nbsp; Yu-Jun , W ANG Tian-Lin , OU Chen , LUSheng-Feng , CAI&nbsp; Yun , LI Qian , DING&nbsp; Ya-Juan&nbsp; &nbsp;and ZHU Bing-Mei</p><p style="text-align: center;">(1 Key Laboratory of Acupuncture and Medicine Research of Ministry of Education, Nanjing University of Chinese Medicine, Nanjing 210023, China, 2 Regenerative Medicine Research Center, West China Hospital, Sichuan University, Chengdu, Sichuan, China 610041)</p><p style="text-align: justify;"><strong>Abstract</strong> Background: To investigate whether Stat5 is a true player for cardioprotective effect on remote ischemic preconditioning (RIPC), and explored the role of Stat5 in electro-acupuncture (EA) pretreatment against myocardial I/R, and determine&nbsp; its molecular mechanisms.&nbsp;&nbsp;<strong>Methods</strong>: Stat5<span style="vertical-align: super;">fl/fl</span>mice and Stat5-cKOmice&nbsp; were&nbsp; randomly&nbsp; divided&nbsp; into&nbsp; 6&nbsp; groups:&nbsp; Stat5<span style="vertical-align: super;">fl/fl</span>+I/R&nbsp; and&nbsp; Stat5-cKO+I/R&nbsp; groups&nbsp; (30/180&nbsp; min&nbsp; left coronary&nbsp; artery&nbsp; occlusion/reperfusion);&nbsp; Stat5<span style="vertical-align: super;">fl/fl</span>+RIPC+I/R&nbsp; and&nbsp; Stat5-cKO+RIPC+I/R&nbsp; groups&nbsp; (applied RIPC with three 5-min cycles of femoral artery occlusion and reperfusion before I/R); Stat5<span style="vertical-align: super;">fl/fl</span>+EA+I/R and Stat5-cKO+EA+I/R groups (subjected to 20 min/day of EA treatment for 1 week before I/R). The heart tissue and blood were harvested at the end point of I/R, and the parameters were measured.&nbsp; <strong>Results</strong>: RIPC activated&nbsp; STAT5&nbsp; in&nbsp; the&nbsp; heart&nbsp; subjected&nbsp; to&nbsp; I/R.&nbsp; Decreased&nbsp; expressions&nbsp; of&nbsp; Cyt,&nbsp; cleaved&nbsp; caspase-3,&nbsp; and increased Bcl-xL and Bcl-2 were detected only in the presence of STAT5.RIPC increased cardiac HIF-1α, IL10, p-PI3K, and VEGF proteins only in the Stat5<span style="vertical-align: super;">fl/fl</span>mice with RIPC. EA reduced myocardial infarct size and TUNEL-positive&nbsp; cells&nbsp; in&nbsp; both&nbsp; presence&nbsp; and&nbsp; absence&nbsp; of&nbsp; STAT5,&nbsp; suggesting&nbsp; its&nbsp; Stat5-independency. RNA-seq data further provided evidence that both RIPC and EA pretreatment activated multiple pathways related their protective role against myocardial I/R injury.&nbsp; <strong>Conclusion</strong>: RIPC-induced protection against myocardial I/R injury was Stat5 dependent, and was through anti-apoptotic signaling and survival signaling. The cardioprotection induced by EA was Stat5-independent, but Stat5 does participate in the event through anti-apoptotic and survival signaling at certain extent.</p><p><strong>Key words</strong>:&nbsp; Acupuncture, RIPC, Electro-acupuncture pretreatment, Stat5</p><p><br/></p>