Antisense oligodeoxynucleotide inhibition of tumor necrosis factor-alpha expression is neuroprotective after intracerebral hemorrhage

Background and Purpose-Tumor necrosis factor-alpha (TNF-alpha) expression i s increased in brain after cerebral ischemia, although little is known abou t its abundance and role in intracerebral hemorrhage (ICH). A TNF-alpha -sp ecific antisense oligodeoxynucleotide (ORF4-PE) was used to study the exten t to which TNF-alpha expression influenced neurobehavioral outcomes and bra in damage in a collagenase-induced ICH model in rat. Methods-Male Sprague-Dawley rats were anesthetized, and ICH was induced by intrastriatal administration of heparin and collagenase. Immediately before or 3 hours after ICH induction, ORF4-PE was administered directly into the site of ICH. TNF-alpha mRNA and protein levels were measured by reverse tr anscriptase-polymerase chain reaction and immunoblot analyses. Cell death w as measured by terminal deoxynucleotidyl transferase-mediated uridine 5' tr iphosphate-biotin nick end labeling (TUNEL), Neurobehavioral deficits were measured for 4 weeks after ICH. Results-ICH induction (n=6) elevated TNF-alpha mRNA and protein levels (P<0 .01) at 24 hours after the onset of injury compared with sham controls (n=6 ). Immunohistochemical labeling indicated that ICH was accompanied by eleva ted expression of TNF-<alpha> in neutrophils, macrophages, and microglia. A dministration of ORF4-PE (2.0 nmol) directly into striatal parenchyma, 15 m inutes before (n=4) or 3 hours after (n=6) ICH, decreased levels of TNF-alp ha mRNA (P<0.001) and protein (P<0.01) in the brain tissue surrounding the hematoma compared with animals treated with saline alone (n=6). Mean+/-SEM striatal cell death (cells per high-powered field) was also reduced in anim als receiving ORF4-PE (34.1+/-5.0) compared with the saline-treated ICH gro up (80.3+/-7.50) (P<0.001). ORF4-PE treatment improved neurobehavioral defi cits observed at 24 hours (P<0.001) after induction of ICH (n=6) compared w ith the untreated ICH group (n=6). This improvement was maintained at 28 da ys after hemorrhage induction (P<0.001). Conclusions-These results indicate a pathogenic role for TNF-<alpha> during ICH and demonstrate that reducing TNF-alpha expression using antisense oli godeoxynucleotides is neuroprotective.