Dexmedetomidine protects cardiomyocytes against hypoxia/reoxygenation injury via multiple mechanisms

FIGURE 1. Dex ameliorated H/R‐induced injury in H9c2 cells. (A) H9c2 cells were treated with different doses of Dex for 24 h, and cell viabilities were measured. H9c2 cells were pretreated with different doses of Dex for 2 h, following which the cells were subjected to H/R treatment. (B) Cellular viabilities were assayed. (C) The release of LDH was measured. (D) Levels of IL‐1α were measured. (E) Levels of TNF‐α were measured. Data are presented as mean ± SD, *p < 0.05; **p < 0.01

FIGURE 2. Dex alleviated H/R‐induced oxidative stress via the upregulation of catalase in H9c2 cells. H9c2 cells were pretreated with different doses of Dex for 2 h, following which cells were subjected to H/R treatment. (A) ROS levels were measured. (B) MDA levels were measured. (C) SOD activities were measured. (D) CAT activities were measured. (E) The mRNA levels of catalase were measured. (F) H9c2 cells were pretreated with different doses of Dex for 2 h, following which cells were subjected to H/R treatment and protein levels of catalase were measured by Western blotting. (G) H9c2 cells were treated with different doses of Dex for 6 h and mRNA levels of catalase were measured. (H) H9c2 cells were treated with different doses of Dex for 6 h, and protein levels of catalase were measured. (I) H9c2 cells were transfected with si‐NC or si‐catalase for 24 h, following which the protein levels of catalase were measured by Western blotting. (J) H9c2 cells were transfected with si‐NC or si‐catalase for 4 h, following which cells were treated with Dex for 2 h. Next, cells were exposed to H/R conditions, and ROS levels were measured. (K) H9c2 cells were transfected with si‐NC or si‐catalase for…

FIGURE 3. Dex inhibited apoptosis and ER stress in H/R‐treated H9c2 cells. H9c2 cells were pretreated with indicated doses of Dex for 2 h, following which cells were subjected to H/R treatment. (A) Cellular apoptosis was measured. (B) Relative caspase‐3 activities were measured. (C) Cellular lysates were subjected to Western blotting. (D) Activation of Bax was assayed by immunoprecipitation, and release of mitochondrial proteins was assayed. (E) H9c2 cells were transfected with si‐NC or si‐Bcl‐2 for 24 h, and the levels of Bcl‐2 were measured. (F) H9c2 cells were transfected with si‐NC or si‐catalase for 4 h, following which cells were treated with Dex for 2 h and then exposed to H/R condition. Afterward, cell viabilities (left) and apoptosis (right) were measured. (G) Protein levels of caspase‐3 were measured. (H) Relative caspase‐3 activities were measured. (I) Protein levels of GRP78 and CHOP were measured. (J) H9c2 cells were pretreated with Dex (20 μM) with or without thapsigargin (50 nM) for 2 h, following which cells were exposed to H/R, GRP78, and CHOP were measured by Western blotting. (K) Cell viabilities (left) and apoptosis (right) were measured. (L) Caspase‐3 levels were…

FIGURE 4. Dex treatment led to the activation of JAK2/STAT3 in H9c2 cells. (A) H9c2 cells were pretreated with indicated doses of Dex for 2 h, following which cells were exposed to H/R conditions, total cellular lysates were subjected to Western blotting. (B) H9c2 cells were pretreated with Dex (20 μM) with or without AG490 (50 μM) for 2 h, following which cells were exposed to H/R conditions and cellular lysates were subjected to Western blotting. (C) Cell viabilities were measured. (D) Apoptosis was measured. (E) Relative caspase‐3 activities were measured. (F) The release of LDH was measured. (G) ROS levels were measured. (H) MDA levels were measured. (I) SOD activities were measured. (J) CAT activities were measured. (K) Levels of IL‐1α (left) and TNF‐α (right) were measured. Data are presented as mean ± SD, *p < 0.05; **p < 0.01

FIGURE 5. STAT3 is responsible for the transcription of catalase. (A) H9c2 cells were transfected with si‐NC or si‐STAT3 for 4 h, following which cells were treated with IL‐6 for another 6 h, and mRNA levels of catalase were measured. (B) H9c2 cells were transfected with si‐NC or si‐STAT3 for 4 h, following which cells were treated with IL‐6 (50 ng/ml) for another 12 h. The protein levels of STAT3 and catalase were measured. (C) Schematic structures of the catalase promoter constructs used to measure the luciferase activity (left) are shown. H9c2 cells were transfected with the reporter constructs for 4 h and treated with IL‐6 (50 ng/ml) for another 6 h, following which luciferase activities were measured. Data are presented as mean ± SD, *p < 0.05; **p < 0.01