Cathepsin and calpain inhibitor E64d attenuates matrix metalloproteinase-9 activity after focal cerebral ischemia in rats.
OBJECTIVE
Matrix metalloproteinases (MMPs) and cysteine proteases (calpain and cathepsin B) play an important role in cell death and are upregulated after focal cerebral ischemia. Because there is a significant interaction between MMP-9 with calpain and cathepsin B, we investigated the role of E64d (a calpain and cathepsin B inhibitor) on MMP-9 activation in the rat focal ischemia model.


METHODS
Male Sprague-Dawley rats were subjected to 2 hours of middle cerebral artery occlusion by using the suture insertion method followed by 22 hours of reperfusion. In the treatment group, a single dose of E64d (5 mg/kg IP) was administrated 30 minutes before the induction of focal ischemia, whereas the nontreatment group received dimethyl sulfoxide only. The neurological deficits, infarct volumes, Evans blue extravasation, brain edema, and MMP-9 activation in the brain were determined.


RESULTS
Pretreatment with E64d produced a significant reduction in the cerebral infarction volume (353.1+/-19.8 versus 210.3+/-23.7 mm3) and the neurological deficits. Immunofluorescence studies showed MMP-9, calpain, and cathepsin B activation colocalized to both neurons and the neurovascular endothelial cells after ischemia, which was reduced by E64d.


CONCLUSIONS
These results suggest that E64d treatment provides a neuroprotective effect to rats after transient focal cerebral ischemia by inhibiting the upregulation of MMP-9.
Tsubokawa
Tamiji
T
Department of Physiology and Pharmacology, Loma Linda University School of Medicine, California, USA.
Solaroglu
Ihsan
I
Yatsushige
Hiroshi
H
Cahill
Julian
J
Yata
Kenichiro
K
Zhang
John H
JH
D000818:Animals; D001812:Blood-Brain Barrier / Q000187:drug effects; D001929:Brain Edema / Q000209:etiology / Q000473:pathology; D002154:Calpain / Q000037:antagonists & inhibitors*; D002401:Cathepsin B / Q000037:antagonists & inhibitors*; D002543:Cerebral Hemorrhage / Q000209:etiology; D015853:Cysteine Proteinase Inhibitors / Q000008:administration & dosage / Q000494:pharmacology / Q000627:therapeutic use*; D004353:Drug Evaluation, Preclinical; D004789:Enzyme Activation / Q000187:drug effects; D004790:Enzyme Induction / Q000187:drug effects; D005070:Evans Blue / Q000493:pharmacokinetics; D005119:Extravasation of Diagnostic and Therapeutic Materials; D020244:Infarction, Middle Cerebral Artery / Q000150:complications / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D007930:Leucine / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000627:therapeutic use; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000096:biosynthesis; D061965:Matrix Metalloproteinase Inhibitors; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors*; D018696:Neuroprotective Agents / Q000008:administration & dosage / Q000494:pharmacology / Q000627:therapeutic use*; D011292:Premedication; D051381:Rats; D017207:Rats, Sprague-Dawley; D015424:Reperfusion; D016037:Single-Blind Method
D015853:Cysteine Proteinase Inhibitors; D061965:Matrix Metalloproteinase Inhibitors; D009419:Nerve Tissue Proteins; D018696:Neuroprotective Agents; D005070:Evans Blue; D002154:Calpain; D002401:Cathepsin B; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9; D007930:Leucine; C108192:aloxistatin