Adam17-tnf-alpha Pathway In Pathological Process Of Cerebral Ischemia And Tongluojiu'nao Recipe Of Intervention Mechanism

With the development of modern medicine, Collateral disease in traditional Chinese medicine boasts a new meaning.A great many central nervous system(CNS) disease such as ischemic stroke, AD belong to the category of Collateral disease in trditional Chinese medicine,'toxin attacks brain collaterals'is the key point in its progressive pathogenesis, and in the guide of collateral, effective clinical curative outcome has been acquired. The relationship between Collateral in trditional Chinese medicine and microcirculation is extremely close, especially, the concept of neurovascular unit (NVU) coming up provides Collateral in trditional Chinese medicine with morden medical basis.It is well known that TNF-αas a imflemation factor plays a important role in the pathgenesis of a great many disease such as ischemic stroke and so on. ADAM17/TACE is a key shadase that play a key role in release of TNF-α. ADAM17 and TNF-αcan be secreted by cerebral microvascular endothelial cells(CMEC), neuron and glial in CNS. ADAM17-TNF-αpathway is extremely immportant for the pathgenesis of CNS disease especially ischemic stroke.As a result, under guide of'collateral theory'in traditional Chinese medicine, we observe change characters of the ADAM17-TNF-αpathway in brain after hypoxia(cerebral ischemia and OGD) and intervention by injection of Tongluojiunao (TLJN) to explore the mechanism of neuroprotection by TLJN, and provide the modern biological basis for'cure from collateral disease'.Objective:Observe characteristic changes in time series of ADAM17, TNF-αin serum and brain tissue of middle cerebral artery occlusion (MCAO) rat model to explore the relationship between ADAM17, TNF-αand brain injury,in addition, through intervention by TLJN observe the neuroprotection of TLJN after cerebral ischemia, furthermore, to explore weather there is the relationship between the mechanism neuroprotection of TLJN and the pathway of ADAM17-TNF-αIn addition, in vitro observe characteristic changes in time series of ADAM17,TNF-αin normal, oxygen and gclucose deprivation (OGD) and TLJN intervention CMEC condition medium (CMEC-CM). The researchs above furthermore provide Collateral disease in traditional Chinese medicine with modern medical basis.Method:1. Appraise the neurofunction rate of MCAO rat model,and observe morphological stractural changes and information about brain injury of MCAO rat brain tissue to study neuroprotection of TLJN in different time, using brain tissue slice with HE staining and TTC staining2. Observe characteristic changes in different time of ADAM17, TNF-a of MCAO rat serum and brain tissue using ELISA and immunohistochemistry to explore the role of ADAM17-TNF-a pathway played in pathgenesis of ischemic stroke in vivo and weather TLJN plays its neuroprotective role through ADAM17-TNF-a pathway in ischamic stroke in vivo.3. Observe expression of ADAM17,TNF-a in CMEC-CM and influence on expression of ADAM 17,TNF-a in CMEC-CM intervented by TLJN.First, successfully cultivate and identify CMEC. Second,make OGD CMEC model. Third, add TLJN to intervene OGD CMEC. Finally, using ELISA to detect changes in time series of ADAM17, TNF-a in CMEC-CM.Result:1. Through appraising the neurofunction rate of MCAO rat model,and observing morphological stractural changes and information about brain injury of MCAO rat brain tissue, we found that with cerebral ischemia span extending, neuro function morphological stracture of cerebral tissue and cerebral infarct size become deteriorated. Through light microscope (LM), we found that neuron obviously decrease, cellular swell, karyorrhexis, and matrix swell in ischemic infarct focus. It also can be observed that interspace of blood vessel become large; endothelial cells obviously decrease, cell deflate and karyorrhexis; glia become swell and dropsy and inflaming cell infiltrating around ischemic infarct focus. Hemorrhagic focus also can be observed. TLJN can significantly extenuate cerebral infarct size, brain edema and obviously restore neuro function. Through light microscope, we found that TLJN can significantly extenuate pathological changes. Positive control drug injection of Edaravone has the similar effect but its effect is less than TLJN's.2. ADAM17 and TNF-a in serum and brain tissue of MCAO rat model significantly elevate in the earlier stage after cerebral ischemia and vary with time extending. In addition.TLJN can significantly reduce expression of ADAM17, TNF-a in brain in different time.3. Through observing characteristic expression in different time of AD AM17, TNF-a in CMEC-CM and influence on expression of ADAM17, TNF-a in CMEC-CM by TLJN, we found that OGD can promote CMEC to secrete more AD AM17, TNF-a in different time and TLJN can promote OGD cerebral microvascular endothelial cell to secrete less ADAM17, TNF-αin different time.Conclusions:1. TLJN can obviously extenuate cerebral edema, cerebral infarct area, pathological injury of cerebral tissue and restore neuro function. As a result, the founding above can indicate that TLJN boasts a better anti-neuroinjury on brain after cerebral ischemia. Positive control drug injection of Edaravone has the similar effect but its effect is less than TLJN's.2. After cerebral ischemia, ADAM17 and TNF-αin injured brain coherently and significantly elevate. ADAM17-TNF-αpathway may play a key role in pathgenesis of cerebral ischemia and the mechanism of anti-neuroinjury by TLJN may be related to inhibiting ADAM17-TNF-αpathway in injuried brain after cerebral ischemia.3. The mechanism of anti-neuroinjury after cerebral ischemia by TLJN through inhibition of ADAM17-TNF-αpathway in cerebral ischemic tissue may be closely related to both ADAM17 and TNF-αsecretion of CMEC.