强力霉素抑制基质金属蛋白酶-9 可减少胰腺炎相关肺损伤。

Inhibition of matrix metalloproteinase-9 with doxycycline reduces pancreatitis-associated lung injury.

机构信息

Department of General and Visceral Surgery, University of Freiburg, Freiburg, Germany.

出版信息

Digestion. 2009;80(2):65-73. doi: 10.1159/000212080. Epub 2009 Jun 3.

DOI:10.1159/000212080

PMID:19494493

Abstract

BACKGROUND/AIMS: Severe lung injury, responsible for up to 15% of mortality in acute necrotizing pancreatitis patients, is promoted by neutrophil (PMN) migration into the lung. We have previously demonstrated that pulmonary injury in acute pancreatitis is mediated by PMN-derived matrix metalloproteinase-9 (MMP-9). This study was conducted to evaluate the ability of the broad-spectrum MMP inhibitor doxycycline to prevent secondary pulmonary injury in acute pancreatitis.

METHODS

Eighteen rats were randomized into three groups: severe pancreatitis (SAP), severe pancreatitis + doxycycline (SAP+Dox) (30 mg/kg body mass) or control. Acute pancreatitis was induced by intraductal glycodesoxycholic acid and i.v. stimulation with cerulein. Lung sections were histologically graded for edema, microthrombi, atelectasis and hemorrhage. Active MMP-9 in lung tissue was measured with fluorescent assay (ELISA). Naphtol-AS-D-chloroacetate esterase staining was used to determine pulmonary PMN infiltration. The inhibitory effect of doxycycline on MMP-9-induced transmigration was confirmed in a Matrigel transmigration assay.

RESULTS

Addition of doxycycline significantly reduced TNF-alpha-induced PMN transmigration across Matrigel membrane (12.6 +/- 2.6 vs. 20.1 +/- 3.9 PMNs; p < 0.05). SAP+Dox showed decreased concentration of active MMP-9 in lung tissue (37.89 +/- 1.75 vs. 46.29 +/- 3.68 ng/ml; p < 0.05) and as a result decreased pulmonary infiltration of PMNs (21.2 +/- 5.1 vs. 32.5 +/- 6.8; p < 0.05). Histological evaluation revealed decreased pulmonary edema (1.83 +/- 0.41 vs. 2.33 +/- 0.51, p < 0.05), atelectasis (1.67 +/- 0.52 vs. 2.33 +/- 0.52; p < 0.05) and pulmonary hemorrhage (2.5 +/- 0.55 vs. 1.83 +/- 0.41; p < 0.05) in SAP+Dox vs. SAP. These findings were paralleled by reduced pulmonary expression of active MMP-9.

CONCLUSIONS

Inhibition of MMP-9 activity with doxycycline reduced pancreatitis-associated lung injury and expression of MMP-9 in pulmonary tissue. Doxycycline reduced PMN migration in vitro and in vivo and therefore might represent a novel strategy for the prevention of secondary pulmonary complications in acute pancreatitis.

摘要

背景/目的：中性粒细胞（PMN）向肺部迁移导致急性坏死性胰腺炎患者的死亡率高达 15%，严重的肺损伤是其主要原因。我们之前的研究表明，急性胰腺炎中的肺损伤是由PMN 衍生的基质金属蛋白酶-9（MMP-9）介导的。本研究旨在评估广谱 MMP 抑制剂强力霉素预防急性胰腺炎继发肺损伤的能力。

方法

将 18 只大鼠随机分为三组：重症胰腺炎（SAP）组、重症胰腺炎+强力霉素（SAP+Dox）组（30mg/kg 体重）和对照组。通过胰管内甘氨脱氧胆酸诱导和静脉内给予 Cerulein 刺激诱导急性胰腺炎。对肺组织切片进行水肿、微血栓、肺不张和出血的组织学分级。用荧光测定法（ELISA）测量肺组织中的活性 MMP-9。萘基-AS-D-氯乙酸酯酶染色用于确定肺 PMN 浸润。在 Matrigel 迁移测定中证实了强力霉素对 MMP-9 诱导的迁移的抑制作用。

结果

强力霉素的加入显著降低了 TNF-α诱导的 PMN 通过 Matrigel 膜的迁移（12.6±2.6 对 20.1±3.9 PMNs；p<0.05）。SAP+Dox 组肺组织中活性 MMP-9 的浓度降低（37.89±1.75 对 46.29±3.68ng/ml；p<0.05），PMN 浸润减少（21.2±5.1 对 32.5±6.8；p<0.05）。组织学评估显示，SAP+Dox 组肺水肿（1.83±0.41 对 2.33±0.51，p<0.05）、肺不张（1.67±0.52 对 2.33±0.52；p<0.05）和肺出血（2.5±0.55 对 1.83±0.41；p<0.05）均较 SAP 组减少。这些发现与肺组织中活性 MMP-9 的表达减少相平行。