Hyperthermia aggravates focal cerebral ischemia-reperfusion injury in rats by down-regulating claudin-1 expression

Abstract

Objective To investigate the impact of hyperthermia on the expression of claudin-1 in focal cerebral ischemia-reperfusion injury in rats and its mechanism in ischemic cerebral injury.Methods A total of 100 male Sprague-Dawley rats were randomly divided into 3 groups：sham operation,normothermia and hyperthermia groups.Both the normothermia and hyperthermia groups were redivided into 3 time points：Ischemia （1 hour） and reperfusion for 3,6,and 24 h.A model of middle cerebral artery occlusion was induced by the intraluminal suture method.At 24 h after reperfusion,brain water content was measured by the wet and dry weight method.The volume of cerebral infarction was assessed by 2,3,5 triphenyl tetrazolium chloride staining.At 3,6,and 24 h after reperfusion,the claudin-1 expression in ischemic brain tissue was measured by Western blot and immunohistochemical staining Results At 24 h after reperfusion,the mean neurological function score in the normothermia group was significantly lower than that in the hyperthermia group （2.3 ± 0.48 vs.3.2 ± 0.63; t =3.576,P =0.002）.The brain water content on the operated sides in the sham operation,normothermia and hyperthermia groups was 79.31％ ± 0.60％,81.13％ ± 0.12％,and 84.4％ ± 0.55％,respectively.There were significant differences （F=147.115,P=0.000）.Western blot analysis showed that at 3,6,and 24 h after reperfusion,the expression levels of claudin-1 in the normothermia and hyperthermia groups were significantly lower than the sham operation group （all P =0.000）,and the expression levels of claudin-1 progressively decreased with the extension of ischemia-reperfusion time （all P ＜ 0.05）.At the same time point,the expression level of claudin-1 in the hyperthermia group was significantly lower than that in the normothermia group （all P ＜ 0.01）.At 3 and 6 h after reperfusion,the positive expression of claudin-1 among the cerebrovascular endothelial cells was observed in the sham operation,normothermia and hyperthermia groups,while at 24 h after reperfusion,no claudin-1 positive cells were observed.Compared to the sham operation group,at 3 h after reperfusion,the numbers of claudin-1 positive cell and claudin-1 IOD/area （integrated optical density/accumulated positive cell area） in the normothermia and hyperthermia groups begin to decrease,they decreased significantly at 6 h and disappeared at 24 h （P=0.000）.At 3 and 6 h after reperfusion,claudin-1 IOD/area in the hyperthermia group was significantly lower than that in the normothermia group （all P ＜ 0.01）.Conclusions During cerebral ischemia-reperfusion,hyperthermia may aggravate ischemic brain edema and brain injury by down-regulating the expression of claudin-1 in blood-brain barrier. Key words: Brain Ischemia; Hyperthermia, Induced; Claudin-1; Blood-Brain Barrier; Reperfusion Injury; Disease Models, Animal; Rats