| ObjectiveMyocardial ischemia is noxious stimulation and can activate nociceptors. Based on the results of preliminary studies, the purpose of this study was to detect proteins of the heart and dorsal root ganglion, which change in acute myocardial ischemia and capsaicin sensitive nerves denervation, and discover the new contact point between the heart and dorsal root ganglia.MethodsThe newborn male Sprague-Dawley rats within 48 h-72 h of birth were injected capsaicin (50mg/kg) in order to creat the rat mdel of capsaicin sensitive nerves denervation. The radiant heat tail flick latency was measured to evaluate the model. Male SD rats(270g±20g) were selected to ligate coronary artery in order to creat the rat mdel of acute myocardial ischemia. The color of heart and ECG were observed to evaluate the model.Rats were randomly divided into 8 groups: sham group of normal rats at 3h (N-Sham3h), coronary artery occlusion group of normal rats at 3h (N-CAO3h), sham group of normal rats at 6h(N-Sham6h), coronary artery occlusion group of normal rats at 6h (N-CAO6h), sham group of denervated rats at 3h (D-Sham3h), coronary artery occlusion group of denervated rats at 3h (D-CAO3h), sham group of denervated rats at 6h (D-Sham6h), coronary artery occlusion group of denervated rats at 6h (D-CAO6h).Proteomics were used to find proteins of the heart and dorsal root ganglia(T1-T5)in acute myocardial ischemia. Proteins which change were excised, digested and extracted for peptides. Mass spectrometry analysis(LC-MS/MS) were used to identify the proteins. According to results of identification, we design primers by Primer Premier 5.0 software. Real-time quantitative PCR were used to amplify and Pfaffl method were used to calculate the relative gene expression.ResultsThe radiant heat tail flick latency was significantly increased in group of denervated rats. After coronary artery occlusion, ST-segment of ECG were elevated and T wave of ECG were changed large, which consistent with the ECG characteristics of acute myocardial ischemia. In the heart, selected 7 proteins were identified as 5 proteins: Cu-Zn superoxide dismutase (SOD1), lens protein (Cryab), glutathione S-transferase-P (GST-P ), short-chain acyl-coenzyme A dehydrogenase (SCAD), NADP-isocitrate dehydrogenase (NADP-ICDH) precursors.In the dorsal root ganglia, 5 proteins were identified: GST-P, Cryab, ATP synthaseαsubunit (Atp5a1) precursors, cyclophilin A (Cyp A) and carbonate anhydride enzyme-2 (CA2).Cryab were identified both in the heart and the dorsal root ganglia. There were three forms of Cryab in CAO heart(number from PDQuest software): spot 5105 (30 KDa, pI 6.98), spot 5106 (32.5 KDa, pI 7.02), spot 5111 (30.2 KDa,, pI 7.27). Spot 5106 in the heart of the sham group were expressed and increased in CAO. Spot 5105 appeared in N-CAO6h and D-CAO6h rather than other groups. Spot 5111 appeared in N-CAO3h, N-CAO6h, D-CAO3h and D-CAO6h rather than sham groups.In N-CAO, significant changes were occurred in some proteins of the heart: SOD1 which reduced significantly in N-CAO6h compared with N-Sham6h(P<0.05), Cryab in spot 5106 and spot 5111 which increased significantly in N-CAO3h compared with N-Sham3h(P<0.05), Cryab in spot 5105 and spot 5106 and spot 5111 which increased significantly in N-CAO6h compared with N-Sham6h(P<0.05 or P<0.01), GST-P which reduced significantly in N-CAO6h compared with N-Sham6h(P <0.05), SCAD which increased significantly in N-CAO6h compared with N-Sham6h(P<0.05), and NADP-ICDH precursors which reduced significantly in N-CAO6h compared with N-Sham6h(P<0.05). In N-CAO, significant changes were occurred in some proteins of the dorsal root ganglion: Cryab which reduced significantly in N-CAO6h compared with N-Sham6h(P<0.05), and Atp5a1 precursors which reduced significantly in N-CAO3h compared with N-Sham3h(P<0.05). In D-CAO, significant changes were occurred in some proteins of the heart: Cryab in spot 5106 and spot 5111 which increased significantly in D-CAO3h compared with D-Sham3h(P<0.05), Cryab in spot 5105 and spot 5111 which increased significantly in D-CAO6h compared with D-Sham6h(P<0.05), GST-P which reduced significantly in D-CAO3h compared with D-Sham3h(P<0.01), SCAD which increased significantly in D-CAO6h compared with D-Sham6h(P<0.05), and NADP-ICDH precursors which reduced significantly in D-CAO6h compared with D-Sham6h(P<0.05). In D-CAO, significant changes were occurred in some proteins of the dorsal root ganglion: Cyp A which increased significantly in D-CAO3h compared with D-Sham3h(P<0.05) and increased significantly in D-CAO6h compared with D-Sham6h(P<0.05), and CA2 which reduced significantly in D-CAO3h compared with D-Sham3h(P<0.01). Compared with normal rats, significant changes of denervated rats were occurred in some proteins of the heart: Cryab and SOD1, and significant changes of denervated rats were occurred in some proteins of the dorsal root ganglion: Atp5a1 precursors and GST-P.Both in normal rats and denervated rats, real-time quantitative PCR results are consistent with the trend of protein: SOD1 and SCAD.ConclusionThe reduction of SOD1 in acute myocardial ischemia may increase superoxide; the level of SOD1 was low in the rat mdel of capsaicin sensitive nerves denervation, which suggested the ability to remove superoxide decreased. The protein level of heart SOD1 mainly depend on SOD1 mRNA levels. In normal rats, sensory nerve may be involved in regulation of heart SOD1. Cryab of the heart has three forms; Cryab of the dorsal root ganglia may reach the heart through the axon in myocardial ischemia. Myocardial ischemia resulted in the reduction of GST-P in hearts; the protein levels of GST-P in dorsal root ganglia were not affected by cardiac ischemia. SCAD of hearts increased in myocardial ischemia. NADP-ICDH precursors were used to generate a large number of NADP-ICDH. Myocardial ischemia caused the reduction of Atp5a1 precursors in DRG, which suggested it may generate ATP synthase.SOD1, Cryab, GST-P, SCAD and NADP-ICDH (mitochondria) precursor were the proteins of change in the heart induced by acute myocardial ischemia. Cryab and Atp5a1 precursor were the proteins of change in the dorsal root ganglia induced by acute myocardial ischemia. SOD1 and Cryab of the heart may be regulated by capsaicin sensitive sensory nerves. The role of capsaicin on sensory nerve was manifested in Cryab, Atp5a1 precursors, GST-P, Cyp A and CA2.
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