| BackgroundIntracerebral hemorrhage(ICH)is devastating subtype of stroke,accounting for about 10-20%of all stroke types.More than 2 million people in the world are affected by it every year,and the mortality within 30 days is as high as 43%?51%,and the prognosis is poor.Most survivors have neurological dysfunction related to Hemorrhage areas,and it is the most common acute and severe disease in neurology department.Compared with ischemic stroke,for example have a higher mortality,morbidity,and as the growth of the age,is also increasing year by year in research shows that the hemoglobin of brain tissue has the very high toxicity,and free hemoglobin can cause inflammatory responses,aggravating brain injury after ICH due to hemoglobin in the Central nervous system(CNS)that cannot be recycled,thus increase the metabolism of hemoglobin may be an effective strategy to improve hematoma removal.Heme Oxygenase(HO)is the speed limit of heme degradation enzyme mammalian cells exist in the two HO isozyme activity:induction type(HO-1)and type of activity(HO-2)HO-1 subtype is mainly expressed in vascular structure,but in the CNS of normal expression level is very low,can rapidly be induced after acute ICH brain damage in the increase of the transgenic mice HO-1 for permanent middle cerebral artery occlusion plays a protective role,however,after ICH HO-1 role is still controversial.Overexpression of HO-1 in astrocytes has been reported to improve prognosis after ICH,while other studies have shown that inhibition of HO reduces brain damage after ICHPurposeIn this study,we used Cobalt protoporphyrin(Copp)inducer and Zinc protoporphyrin(Znpp)inhibitor to investigate the exact early role of ho-1 in collagenase-induced ICH model in rats to provide experimental evidence for clinical treatment of ICHMethods(1)128 adult male SD rats(about 8 weeks)were randomly divided into Sham group,ICH group,Copp pretreatment ICH group and Znpp group,with 32 rats in each group,and each group was divided into four time points,8 at each time point(1,2,3,7 days).(2)The Sham group was only injected with the same dose of normal saline,and the rest groups were injected with type ? bacterial collagenase into the striatum system of the rat brain for ICH model.(3)The rats were euthanized one day,two days,three days and seven days after surgery.(4)Histological assessment of brain injury in patients with ICH,including HE staining of brain injury area,Fluoro-Jade B staining for neuronal death,TUNEL staining for brain cell death,Immunofluorescence staining of microglial/macrophage-activated Ibal and Detection of HO-1 expression by immunohistochemistry.Results(1)HE staining showed that there was no obvious ICH and brain injury in the Sham group on days 1,2,3 and 7,indicating that normal saline basically could not cause ICH in rats.Compared with the control group,the ICH group showed significant bleeding on day 2 and 3(p<0.05).In the Copp group,ICH and brain injury on day 1,2,3 and 7 were all greater than that in the ICH group(p<0.05),especially on day 2 and 3,the area of brain injury was significantly greater than that in the ICH group(p<0.01),which was statistically significant.In the Znpp group,ICH and brain injury on day 1,2,3 and 7 were all smaller than that in the ICHgroup(p<0.05),especially on day 2,the area of brain injury was significantly smaller than that in the ICH group(p<0.01),which was statistically significant.(2)Fluoro-Jade B staining showed no significant changes in the positive cells around the hematoma at 1,2,3,7 days in the Sham group.Compared with the control group,the positive cells around the hematoma in the ICH group were more obvious on day 2 and 3(p<0.05).In the Copp group,the positive cells around the hematoma on day 1,2,3 and 7 were all greater than those around the hematoma in the ICH group(p<0.05),and the number of positive cells around the hematoma on day 2 and 3 was significantly greater than that at other time points(p<0.01),showing statistical significance.In the Znpp group,the number of positive cells around the hematoma on day 1,2,3 and 7 was lower than that of the ICH group,especially the number of positive cells around the hematoma on day 2 and 3 was significantly lower than that at other time points(p<0.01),which was statistically significant.(3)TUNEL staining showed no significant changes in the positive cells around the hematoma in the Sham group.Compared with the control group,the positive cells around the hematoma on day 2 and 3 in the ICH group were more obvious and gradually increased.(p<0.05);In the Copp group,the positive cells around the hematoma on day 1,2,3 and 7 were all greater than those around the hematoma in the ICH group(p<0.05),and the number of positive cells around the hematoma on day 2 and 3 was significantly greater than that at other time points(p<0.01),showing statistical significance.The number of positive cells around the hematoma in Znpp group on day 1,2,3 and 7 was lower than that of the ICH group(p<0.05),and the number of positive cells around the hematoma on day 2 and 3 was significantly lower than that at other time points(p<0.01),indicating statistical significance.(4)Iba1 staining showed no significant changes in the positive cells around the hematoma in the Sham group.Compared with the control group,the positive cells around the hematoma on day 2,3 and 7 were more obvious in the ICH group(p<0.05).The number of positive cells around the hematoma on day 1,2,3 and 7 in the cobalt-protoporphyrin group was greater than that in the ICH group,especially the number of positive cells around the hematoma on day 2,3 and 7 was significantly greater than that at other time points(p<0.05),showing statistical significance.In the zinc protoporphyrin group,the number of positive cells around the hematoma on day 1,2,3 and 7 was lower than that of the ICH group,especially the number of positive cells around the hematoma on day 2 and 3 was significantly lower than that at other time points(p<0.05),which was statistically significant.(5)HO-1 immunohistochemical staining showed no significant changes in the positive cells around the hematoma in the Sham group.Compared with the control group,the positive cells around the hematoma in the ICH group were more obvious on day 2 and 3(p<0.05).In the Copp group,the positive cells around the hematoma on day 1,2,3 and 7 were all greater than those around the hematoma in the ICH group(p<0.05),and the number of positive cells around the hematoma on day 2 and 3 was significantly greater than that at other time points(p<0.01),showing statistical significance.The positive cells around the hematoma in Znpp group on day 1,2,3 and 7 were all smaller than those around the hematoma in the ICH group(p<0.05),and the number of positive cells around the hematoma on day 2 and 3 was significantly smaller than that at other time points(p<0.01),showing statistical significance.Conclusion(1)After ICH,the death cells of microglia around the hematoma increased significantly,indicating that they were related to the pathophysiological process of ICH.(2)HO-1 in the acute stage aggravates ICH and brain injury by activating microglia/macrophages and causing inflammatory responses to increase the death of neurons and the accumulation of iron.(3)Early inhibition of HO-1 expression can be used as a new therapeutic strategy to reduce cerebral edema,reduce inflammatory damage,and improve functional prognosis of ICH patients. |
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