Lung matrix metalloproteinase activation following partial hepatic ischemia/reperfusion injury in rats

Abstract

Purpose: Warm hepatic ischemia-reperfusion (I/R) injury can lead to multiorgan dysfunction. The aim of the present study was to investigate whether acute liver I/R does affect the function and/or structure of remote organs such as lung, kidney, and heart via modulation of extracellular matrix remodelling.

Methods: Male Sprague-Dawley rats were subjected to 30 min partial hepatic ischemia by clamping the hepatic artery and the portal vein. After a 60 min reperfusion, liver, lung, kidney, and heart biopsies and blood samples were collected. Serum hepatic enzymes, creatinine, urea, Troponin I and TNF-alpha, and tissue matrix metalloproteinases (MMP-2, MMP-9), myeloperoxidase (MPO), malondialdehyde (MDA), and morphology were monitored.

Results: Serum levels of hepatic enzymes and TNF-alpha were concomitantly increased during hepatic I/R. An increase in hepatic MMP-2 and MMP-9 activities was substantiated by tissue morphology alterations. Notably, acute hepatic I/R affect the lung inasmuch as MMP-9 activity and MPO levels were increased. No difference in MMPs and MPO was observed in kidney and heart.

Conclusions: Although the underlying mechanism needs further investigation, this is the first study in which the MMP activation in a distant organ is reported; this event is probably TNF-alpha-mediated and the lung appears as the first remote organ to be involved in hepatic I/R injury.

Figure 1

Serum levels of TNF-α in animals submitted to ischemia/reperfusion (I/R). Sham operated group (control, n = 15) has been compared with I/R group (n = 17): *P < 0.05. These are the mean results of 32 different experiments ± S.E.M.

Figure 2

Bar graphs of MMP-2 and MMP-9 activity in ischemic liver lobe (a), lung (b), heart (c), kidney cortex (d), and medulla (e). Sham-operated group (control, n = 15) has been compared with I/R group (n = 17): *P < 0.05. Data are shown as mean values ± SEM.

Figure 3

Representative light micrographs of the left liver lobe of sham-operated rats (a) and of the left lobe of rats submitted to ischemia/reperfusion (I/R) (b and c). Hematoxylin and eosin staining. The sham animal shows normal hepatocyte and sinusoid morphology (a). In animals submitted to I/R the lower magnification picture (b) shows decreased eosinophilia of hepatocytes, hepatocyte vacuolation, disarrangement of hepatocyte cords, and altered sinusoidal dilatation (b). An area of extensive hepatocyte necrosis and plate disintegration (black stars) is shown under higher magnification in (c), example of grade 3. P: portal vein; CL: centrolobular vein.

Figure 4

Representative light micrographs of kidney samples obtained from sham-operated rats (a, b, and c) and from rats submitted to hepatic ischemia/reperfusion (I/R) (d, e, and f). Cortex (a, b), outer medulla (c, d), and inner medulla (e; f) are illustrated. Hematoxylin and eosin staining. With respect to the normal morphology of the cortex of sham animals (a), the cortex of animals submitted to hepatic I/R (b) shows dilated interstitium and injury to a few tubules. The insets in (a) show a normal distal convolute tubule (DCT) and in (b) patchy areas of dilatation. With respect to the normal morphology of the outer medulla of sham animals (c), the outer medulla of animals submitted to hepatic I/R shows extended areas of interstitial fluid (asterisk) apparently displacing thick limbs of Henle's loop (THL). Respect to the inner medulla of sham animals (e), the inner medulla of animals submitted to I/R (f), shows slightly dilated thin limbs of Henle's loop (tHL) and increased cellularity in the stromal.

Figure 5

Changes of lung granulocytes in response to hepatic I/R injury. Lung samples were obtained from sham-operated rats, not submitted to hepatic I/R, and from rats whose hepatic lobes were submitted to 30 min of ischemia and hence reperfused for 60 min. The number of granulocytes was calculated per microscopic field. *P < 0.05. Data are shown as mean values ± SEM.

Figure 6

Representative sections of lung tissue from sham-operated animals (a, b, and c) and from animals submitted to hepatic ischemia/reperfusion (I/R) (d, e, and f). Hematoxylin and eosin staining. The sections from sham animals show the typical morphology of bronchi (B), blood vessels, and alveoli (alv) with associated capillaries; alveolar capillaries are recognizable in high magnification (c), by erythrocytes in the lumen. In the lung tissues of animals submitted to hepatic I/R alveoli appear to be dilated. (d, e) Dilated lymph vessels (L) surrounding arteries (A) and an abundant number of inflammatory cells (arrowheads) in the lumen and stroma of blood vessels (f).