Effect of intratracheal injection of JNK siRNA on ischemia-reperfusion injury in a rat model of lung transplantation

Abstract

Objective To investigate the effect of intratracheal injection of c-Jun N-terminal kinase (JNK) siRNA plasmid on ischemia-reperfusion (I/R) injury in a rat model of lung transplantation. Methods ExperimentⅠ Thirty-two male Wistar rats, weighing 250-280 g, were divided into 2 groups (n=16 each) using a random number table method: control group (group C) and JNK siRNA group.JNK siRNA plasmid 2 mg/kg (diluted to 0.2 ml in sterile phosphate buffer solution) was intratracheally injected in JNK siRNA group.Scrambled siRNA plasmid 2 mg/kg (diluted to 0.2 ml in sterile phosphate buffer solution) was intratracheally injected in group C. Six rats in each group were sacrificed at 48 h of administration, and left lung tissues were removed for determination of the expression of JNK and JNK mRNA (by Western blot and real-time polymerase chain reaction, respectively). The other 10 rats left in each group were used for left lung transplantation.Experiment Ⅱ Thirty male Wistar rats, weighing 250-280 g, were divided into 3 groups (n=10 each) using a random number table method: sham operation group (group S), transplanted lung I/R group (group I/R) and transplanted lung I/R+ JNK siRNA group (group I/R+ JNK siRNA). In group I/R and group I/R+ JNK siRNA, the left lung transplantation was performed, and the donor lungs were obtained from the living donors in group C and group JNK siRNA, respectively.At 15 min of mechanical ventilation and 30, 60, 90 and 120 min of reperfusion, arterial blood samples were obtained for blood gas analysis, PaO2 was recorded, and the oxygenation index (PaO2÷FiO2) was calculated.Arterial blood samples were obtained at 120 min of reperfusion in transplanted lungs for determination of concentrations of interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-α) and interferon-γ (IFN-γ) in serum (using enzyme-linked immunosorbent assay), and the rats were sacrificed and left lung tissues were removed for microscopic examination of the pathological changes which were scored and for determination of wet/dry lung weight ratio (W/D ratio), and nuclear factor kappa B (NF-κB) and activator protein-1 (AP-1) activities (using enzyme-linked immunosorbent assay). Results ExperimentⅠ Compared with group C, the expression of JNK and JNK mRNA was significantly down-regulated in group JNK siRNA (P<0.05). ExperimentⅡ Compared with group S, the oxygenation index was significantly decreased, and the concentrations of serum IL-8, TNF-α and IFN-γ, W/D ratio, lung injury score and activities of NF-κB and AP-1 were increased in I/R and I/R+ JNK siRNA groups (P<0.05). Compared with group I/R, the oxygenation index of receptors were significantly increased, and the concentrations of serum IL-8, TNF-α and IFN-γ, W/D ratio, lung injury score and activities of NF-κB and AP-1 were decreased in group I/R+ JNK siRNA (P<0.05). Conclusion Intratracheal injection of JNK siRNA can reduce transplanted lung I/R injury, and the mechanism may be related to inhibiting inflammatory responses of rats. Key words: JNK mitogen-activated protein kinases; Lung transplantation; Reperfusion injury; Injection, intratracheal