Inhibition of Matrix Metalloproteinase-9 with Doxycycline Reduces Pancreatitis-Associated Lung Injury

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-α-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.