Regulation of matrix metalloproteinases and effect of MMP-inhibition in heart transplant related reperfusion injury.
OBJECTIVE
Metalloproteinases (MMPs) regulate extracellular matrix turnover and degrade basal lamina. Aim of the study was to examine the regulation of MMPs and the effect of an MMP inhibitor in transplant related ischemia/reperfusion injury.


METHODS
Heterotopic cardiac transplantation was performed after 4 h of ischemia in three groups of six rats: allografts (black hooded inbred strain, PVG donor/August Copenhagen Irish inbred strain recipient); allografts treated with a competitive MMP-inhibitor (Batimastat) 15 mg/kg every 24 h; isografts (PVG donor and recipient). Normal PVG hearts served as a control. Hearts were explanted after 72 h of reperfusion. Expression of MMP-2 and -9 was measured using gelatin zymography. Proteolytic activity was measured using a gelatinase activity assay. Myeloperoxidase activity and tumor necrosis factor-alpha (TNF-alpha) were measured as markers of inflammatory response. Immunostaining for collagen IV and laminin was used to study degradation of basal lamina.


RESULTS
There was a significant increase of MMP-2 expression in allografts (2271+/-571 microg/ml) as compared to normal (683+/-139 microg/ml) and the Batimastat-treated (259+/-140 microg/ml, P<0.05) groups. Although pro-MMP-2 expression was equally high in the untreated iso- and allograft group (75+/-23 versus 62+/-30 microg/ml) MMP-2 expression in the isograft hearts was significantly lower (359+/-267 microg/ml) suggesting activation of the pro-form by an immunologic mechanism. Pro-MMP-9 levels were significantly higher in the untreated iso- and allograft groups as opposed to normal hearts and MMP-inhibited hearts. MMP-9 was completely inhibited by Batimastat treatment. Collagenolytic activity was lower in the treated group as compared to untreated allografts (538+/-140 versus 384+/-97 microg/ml, P<0.05), demonstrating effective inhibition of MMPs by Batimastat. In the treated group a lesser extent of basement membrane component alterations could be demonstrated by laminin and collagen IV staining. There was a significant reduction in myeloperoxidase activity (P=0.027) as well as lower TNF-alpha levels (ns) in the in the Batimastat treated group.


CONCLUSIONS
Ischemia leads to an increase in MMP expression and degradation of basal lamina. This process is enhanced in allografts as compared to isografts suggesting additional activation of MMPs by immunologic mechanisms. MMP-inhibition is effective in preventing the proteolytic activity of MMPs and may alter the host rejection response by preserving extracellular matrix components and basement membranes.
Falk
V
V
Department of Cardiothoracic Surgery, Falk Cardiovascular Research Building, Stanford University School of Medicine, Stanford, CA, USA. falv@medizin.uni-leipzig.de
Soccal
P M
PM
Grünenfelder
J
J
Hoyt
G
G
Walther
T
T
Robbins
R C
RC
D000818:Animals; D001485:Basement Membrane; D016027:Heart Transplantation / Q000276:immunology / Q000502:physiology*; D008297:Male; D020782:Matrix Metalloproteinases / Q000276:immunology / Q000378:metabolism*; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors*; D015428:Myocardial Reperfusion Injury / Q000503:physiopathology*; D009195:Peroxidase / Q000378:metabolism; D010649:Phenylalanine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D051381:Rats; D011919:Rats, Inbred Strains; D013876:Thiophenes / Q000494:pharmacology*; D014184:Transplantation, Homologous; D014185:Transplantation, Isogeneic; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis; D015854:Up-Regulation
D013876:Thiophenes; D014409:Tumor Necrosis Factor-alpha; D010649:Phenylalanine; C080985:batimastat; D009195:Peroxidase; D020782:Matrix Metalloproteinases; D008666:Metalloendopeptidases