| Intracerebral hemorrhage (ICH) is a serious disease that has high mortality and disability ratio without effective treatment. So it is still a hot point to research the treatments of ICH. For a long time, the brain is a immunological privilege area without any lymphatic tissue and can't provoke immunological reaction. These years people get to know this theory is not true. Central neural system has immune function and ICH can produce inflammation reaction, too. The proinflammatory cytokines, known as an important symbol of inflammation, play a key role in ICH pathology. So how to inhibit cytokines over-expression after ICH is a new field need to explore. Consequently, theories can be used to guide therapies to alleviate the destroy of blood-brain barrier (BBB), reduce brain edema, decrease the death of neurocytes and astrocytes, inhibit neurons apoptosis, etc. Several studies reported that mild hypothermia can increase survival ratio and life quality of ICH patients. The purpose of this experiment is to study the effect of skull local mild hypothermia on cytokines and transcription factors in tissue surround the hematoma of ICH. The rat model of experimental ICH was made by the method of injection of autologous arterial blood into the caudate nucleus. A total of 36 male Wistar rats were divided into 3 groups at random: the sham-operation group, the normothermia group and the mild hypothermia group. After anesthesia the normothermia group and the mild hypothermia group received an injection of 50μl of autologous arterial blood into the basal ganglia. An head cooling therapeutic instrument was introduced into the mild hypothermia group immediately after the injection and then maintained mild hypothermia for 48hrs. The cooling pad of the instrument was placed on the operated side of the rats'head and immobilized. The temperature was modulated at 15℃—18℃. The rat's head and the cooling pad were covered with a wet towel in order to keep the temperature. A temperature sensor was placed at the center of the hematoma, ranged the intracerebral temperature from 32 ℃to 34℃. Kept the rats'body temperature between 37℃and 38 ℃. The sham-operation group underwent the same operation but didn't receive injection. Animals of all groups were anesthetized and killed by perfusion with 4% paraformaldehyd. Fixed cerebral tissues were cut coronally through the needle entry site, and slices including the hematoma site were embedded in paraffin. The coronal slides of cerebral tissue were stained by HE and with rabbit polyclonals anti-TNFα, anti-NF-κB/p65 antibodies by immunohistochemical technique. Counted the positive cells for statistic analysis. The result showed: ①The activity of the rats following suffering experimental ICH was decreased. The hemiplegia and circle motion could be observed among the ICH rats. ②The round hematoma in the area of caudate nucleus was existed by the gross observation of the normothermia group and the mild hypothermia group, and the diameter of the hematoma was about 3mm, which showed the experimental ICH model was successful. The operated side of the brain was swelling, edema of the normothermia group. Herniation could be seen in some samples. In the mild hypothermia group, the damage was less than the normothermia group. In the sham-operation group, only the needle hole could be seen. ③HE: The pathological changes of the normothermia grouparound the hematoma included tissue softening, edema, swelling and necrosis of neuron and astrocyte, nerve fibers rupture and demyelination, inflammatory cells emerging, small vessels hemorrhage. Of the mild hypothermia group edema and swelling around the hematoma was alleviated, necrotic cells were fewer, a few small vessels hemorrhage and astrocyte reaction can be seen. There were few inflammatory cells near the needle tunnel in the sham-operation group. ④Immunohistochemistry: Immunohistochemical labeling indicated that ICH was accompanied by elevated expression of TNF-αand NF-κB. The expression sites of the normothermia group focused on the circumambience of the hematoma, cortex, callose, choroid plexus of the operated side. Compared with the sham-operation group, the quantity of positive cells has significant deviation (P<0.01). The expressional level of NF-κB and TNF-αwas also reduced in the mild hypothermia group (P<0.01) compared with the normothermia group, having significant deviation. According to the result, inflammatory cells emerging and astrocytes hyperplasia appeared around the hematoma after ICH 48hrs. At the same time, TNF-αand NF-κB increased remarkably. It indicated the presence of inflammatory response in brain at acute stage of ICH. The remedy of mild hypothermia could decrease the expression both of TNF-αand NF-κB and the quantity of inflammatory cells around the hematoma, alleviate astrocytes hyperplasia. The degree of brain edema in the mild hypothermia group was lower than that of the normothermia group. These results revealed the remedy of mild hypothermia could inhibit inflammatory response and protect brain. It can be concluded that the remedy of mild hypothermia inhibit inflammatory response by inhibiting the expression of NF-κB.
|
PDF Full Text Request