Sun Jieyun, Zhang Peng, Yang Xinying, Xu Di, Du Bin, Chen Junliang, Wu Minchen, Pang Qingfeng
Wuxi Medical School, Jiangnan University, Wuxi 214000, Jiangsu, China. Corresponding author: Pang Qingfeng, Email:
Zhonghua Wei Zhong Bing Ji Jiu Yi Xue. 2017 Jan;29(1):25-29. doi: 10.3760/cma.j.issn.2095-4352.2017.01.006.
To observe the effect of biliverdin (BV) on the lung ischemia/reperfusion injury (LIRI), and to investigate the mechanism of BV in treatment of LIRI.
Thirty-two male Sprague-Dawley (SD) rats were randomly divided into control group, LIRI group, glutathione (GSH) group and BV-treated group, with 8 rats in each group. The rat LIRI model was reproduced by isolated lung perfusion system. Fifteen minutes after perfusion balance, lungs of control group were perfused continuously for 90 minutes, and the lungs in other groups were reperfused for 90 minutes after 1-hour ischemia, while the perfusion was added 10 μmol/L BV in BV-treated group and 4 mmol/L GSH in GSH group respectively. During the perfusion, the respiratory related indicators, such as tidal volume (VT), static compliance (Cst), arterial partial pressure of oxygen (PaO), airway resistance (Raw), were dynamically observed. After perfusion, the wet/dry weight ratio (W/D) of lungs was determined. Pathological changes in lung tissue were observed under light microscope. The cell apoptosis was observed by TdT-mediated dUTP nick end labeling (TUNEL) method, and the apoptosis index was calculated. The protein expression levels of heme oxygenase-1 (HO-1), phosphorylated c-Jun N-terminal kinase (p-JNK), and caspase-3 were determined by Western Blot.
Compared with control group, VT, Cst and PaO in LIRI group were significantly decreased from reperfusion for 30 minutes, and Raw was significantly increased. The pathological results showed that there was different degree of hyperemia edema, inflammatory cells infiltration and bronchial endothelium injury in the lung tissue of LIRI group. The W/D ratio of LIRI group was significantly higher than that of control group (8.98±2.34 vs. 5.89±0.52, P < 0.05). TUNEL results showed that the tan apoptosis cells of LIRI group were more than control group, and the apoptosis index was significantly higher than that of the control group [(13.88±2.35)% vs. (2.26±0.60)%, P < 0.05]. Compared with control group, the protein expression levels of HO-1, p-JNK, and caspase-3 in LIRI group were significantly increased [HO-1 (gray value): 0.55±0.13 vs. 0.16±0.02, p-JNK (gray value): 0.46±0.08 vs. 0.16±0.05, caspase-3 (gray value): 0.65±0.13 vs. 0.26±0.03, all P < 0.05]. Compared with LIRI group, the respiratory indicators in BV-treated group were improved significantly, the lung tissue injury was significantly reduced, the W/D ratio was significantly decreased (6.39±0.45 vs. 8.98±2.34, P < 0.05), and the cell apoptosis index was significantly reduced [(4.49±1.10)% vs. (13.88±2.35)%, P < 0.05], as well as the protein expression levels of HO-1, p-JNK, and caspase-3 were significantly lowered [HO-1 (gray value): 0.19±0.03 vs. 0.55±0.13, p-JNK (gray value): 0.31±0.06 vs. 0.46±0.08, caspase-3 (gray value): 0.33±0.05 vs. 0.65±0.13, all P < 0.05], which indicating that BV could alleviate LIRI via anti-apoptosis. The improvement effect of BV on PaO, apoptosis index, protein expressions of HO-1 and caspase-3, and JNK phosphorylation were better than positive drug GSH, indicating that BV could protect LIRI ideally.
BV alleviates LIRI via its anti-JNK pathway and its anti-apoptosis property.
观察胆红素（BV）对肺缺血/再灌注损伤（LIRI）的影响，并探讨BV治疗LIRI的机制。
将32只雄性Sprague-Dawley（SD）大鼠随机分为对照组、LIRI组、谷胱甘肽（GSH）组和BV治疗组，每组8只。采用离体肺灌注系统复制大鼠LIRI模型。灌注平衡15分钟后，对照组肺持续灌注90分钟，其他组在缺血1小时后再灌注90分钟，其中BV治疗组在灌注时加入10μmol/L BV，GSH组加入4mmol/L GSH。灌注过程中，动态观察潮气量（VT）、静态顺应性（Cst）、动脉血氧分压（PaO）、气道阻力（Raw）等呼吸相关指标。灌注后，测定肺组织湿/干重比（W/D）。光镜下观察肺组织病理变化。采用TdT介导的dUTP缺口末端标记（TUNEL）法观察细胞凋亡，并计算凋亡指数。采用蛋白质免疫印迹法检测血红素加氧酶-1（HO-1）、磷酸化c-Jun氨基末端激酶（p-JNK）和半胱天冬酶-3的蛋白表达水平。
与对照组比较，LIRI组再灌注30分钟起VT、Cst和PaO显著降低，Raw显著升高。病理结果显示，LIRI组肺组织有不同程度的充血水肿、炎性细胞浸润及支气管内皮损伤。LIRI组W/D比值显著高于对照组（8.98±2.34比5.89±0.52，P<0.05）。TUNEL结果显示，LIRI组棕黄色凋亡细胞多于对照组，凋亡指数显著高于对照组[（13.88±2.35）%比（2.26±0.60）%，P<0.05]。与对照组比较，LIRI组HO-1、p-JNK和半胱天冬酶-3蛋白表达水平显著升高[HO-1（灰度值）：0.55±0.13比0.16±0.02，p-JNK（灰度值）：0.46±0.08比0.16±0.05，半胱天冬酶-3（灰度值）：0.65±0.13比0.26±0.03，均P<0.05]。与LIRI组比较，BV治疗组呼吸指标明显改善，肺组织损伤明显减轻，W/D比值显著降低（6.39±0.45比8.98±2.34，P<0.05），细胞凋亡指数显著降低[（4.49±1.10）%比（13.88±2.35）%，P<0.05]，HO-1、p-JNK和半胱天冬酶-3蛋白表达水平显著降低[HO-1（灰度值）：0.19±0.03比0.55±0.13，p-JNK（灰度值）：0.31±0.06比0.46±0.08，半胱天冬酶-3（灰度值）：0.33±0.05比0.65±0.13，均P<0.05]，表明BV可通过抗凋亡减轻LIRI。BV对PaO、凋亡指数、HO-1和半胱天冬酶-3蛋白表达及JNK磷酸化的改善作用优于阳性药物GSH，表明BV对LIRI有理想的保护作用。
BV通过抗JNK途径及抗凋亡作用减轻LIRI。
