Objective: To observe the changes in the expressions of microRNA-126 in myocardial tissue and cardiac troponin I (cTnI) in serum of rats in the early stage of severe burn injury with analysis of their relationship, and to validate the relationship between microRNA-126 and myocardial damage in cellular level.
Methods: (1) Forty-eight SD rats were divided into sham injury group (n=8, without fluid therapy after sham injury) and burn injury group (n=40, inflicted with 30% TBSA full-thickness scald on the back, hereinafter referred to as burn, and received intraperitoneally injection of lactic acid Ringer's solution) according to the random number table. Blood was collected from abdominal aorta of rats in sham injury group at post injury hour (PIH) 1, and then these 8 rats were sacrificed for obtaining left ventricular tissue. Blood was respectively collected from abdominal aorta of 8 rats in burn injury group at PIH 3, 6, 12, 24, and 48, and then they were sacrificed and the left ventricular tissue was obtained at each time point. The expression of microRNA-126 in myocardial tissue was assessed by real-time fluorescent quantitative RT-PCR. Serum level of cTnI was assessed by ELISA. (2) Rat myocardial cell line H9C2 was divided into normal control group (NC, routinely cultured), stimulation group (S), negative transfection+stimulation group (NT+S), and transfection+stimulation group (T+S) according to the random number table. Cells in S group were treated with hypoxia for 24 h after being cultured with DMEM containing 10% burn serum obtained from rats in burn injury group at PIH 6 in experiment (1). Cells in NT+S group and T+S group were respectively transfected with the negative control of microRNA mimics and microRNA-126 mimics for 24 h, and then were given the same treatment as that of S group. The expression of microRNA-126 in myocardial cells was determined by real-time fluorescent quantitative RT-PCR (with the sample number of 3). Cell counting kit 8 was used to examine the vitality of myocardial cell (with the sample number of 4, denoted as absorbance value). Apoptotic rate of myocardial cells was determined by flow cytometer (with the sample number of 3). Data were processed with one-way analysis of variance and LSD-t test. The relationship between microRNA-126 expression in myocardial tissue and serum level of cTnI of rats was assessed by linear correlation analysis.
Results: (1) Compared with that of sham injury group at PIH 1, the expression levels of microRNA-126 in myocardial tissue of rats in burn injury group at PIH 3, 6, 12, 24, and 48 were significantly decreased (with t values from 5.68 to 9.79, P values below 0.01), reaching its nadir at PIH 24 (0.40 ± 0.08). Compared with that of sham injury group at PIH 1, the serum levels of cTnI of rats in burn injury group at PIH 3, 6, 12, 24, and 48 were significantly increased (with t values from 6.68 to 12.79, P values below 0.01), peaking at PIH 12 [(1 035 ± 177) pg/mL]. A significant negative correlation between the expression level of microRNA-126 in myocardial tissue and serum level of cTnI was observed in rats of burn injury group at each time point (r=-0.797, P<0.001). (2) Compared with those of NC group, the microRNA-126 expression levels in myocardial cells of S group and T+S group were respectively decreased and increased (with t values respectively 4.57 and 5.73, P<0.05 or P<0.01), the cell vitality levels were obviously decreased (with t values respectively 14.88 and 6.48, P values below 0.01), and the apoptotic rates were significantly increased (with t values respectively 13.82 and 6.96, P values below 0.01). Compared with that in NT+S group, the microRNA-126 expression level in myocardial cells of T+S group was significantly increased (t=6.77, P<0.01), the cell vitality level was obviously increased (t=8.23, P<0.001), and the apoptotic rate was significantly decreased (t=6.14, P<0.001).
Conclusions: Expression level of microRNA-126 in myocardial tissue of rat was decreased in the early stage of severe burn injury. It may participate in regulating myocardial damage and play a protective role.