Astrocytic-Inducible Nitric Oxide Synthase in the Ischemic Developing Human Brain

Abstract

Variability in the expression of apoptotic and inflammatory mediators with time after an ischemic insult and their role in the expansion of cerebral infarcts are still controversial. This study examines DNA degradation and the expression of activated caspase-3 and iNOS, inducible nitric oxide (iNOS) in the human developing brain. Autopsy specimens from children with a neuropathologic diagnosis of focal ischemic infarct were included in the study. The specimens were classified based on the clinical history as acute (Ͻ24 h, n ϭ 5), subacute (24 -72 h, n ϭ 8), or old (Ͼ72 h, n ϭ 6) infarcts. Immunohistochemical staining for caspase-3, iNOS and TUNEL were then performed on all infarcts alongside age-matched controls. TUNEL staining was detected in regions of all infarcts. Expression of iNOS was significantly higher than that of caspase-3 in the penumbra of subacute infarcts (p ϭ 0.02). Glial fibrillary acidic protein and iNOS staining co-localized in the penumbra of acute and subacute infarcts. These results suggest that cell death continues to occur for more than 3 d post ischemic insult. Cell death in the penumbra of subacute infarcts is partially caspase-3 independent and may be attributed to nitric oxide. Astrocytes are a source of iNOS and may play a role in the evolution of pediatric brain injury days after the initial insult.

X-Linked Inhibitor of Apoptosis Protein Expression After Ischemic Injury in the Human and Rat Developing Brain

“…Core of chronic infarcts (wks-mo/y) were basically cavities with surrounding astrogliosis and in some cases persistent histiocytes. Normal brain parenchyma directly adjacent to the ischemic "core" is the "penumbra" as defined previously (10). None of the age-matched control brain sections showed any pathologic changes.…”

“…The neuronal expression of XIAP in human ischemic brain, we found, is in keeping with previous studies showing XIAP in cerebral neurons of neonatal mice (12), adult rat (17), and mice (18) brain after ischemic injury. We have previously shown that apoptotic cell death persists for several days in the ischemic human developing brain and that the number of caspase-3 positive cells significantly decreased in the penumbra of subacute compared with acute infarcts (10). Increasing XIAP expression in the penumbra of subacute infarcts may play a role in inhibiting both caspase-dependent and -independent apoptosis in ischemic developing brain (5)(6)10).…”

“…More recently, we have shown delayed apoptosis occurring for several days after stroke in the human developing brain (10). We also showed that activated caspase-3 continued to be detected for more than 72 h after stroke, suggesting that antiapoptotic agents are a plausible intervention with a prolonged therapeutic window (10).…”

“…Other limitations in using human tissue include the variability of brain regions affected by ischemia and the uncertainty in clinically determining the age of the infarct. We have previously shown that there was no significant difference in caspase-3 expression between infra-and supratentorial ischemic lesion in the human developing brain (10). All of the included patients were critically ill (Table 1) and heavily sedated.…”

“…Tissue preparation and neuropathologic examination of postmortem pediatric human brain specimens were described previously (10). Paraffinembedded 5 m sections stained with hematoxylin and eosin (H and E) were reviewed by the neuropathologist (C.H.)…”