The Mechanism Study Of Naochuxue Prescription On Hematoma Absorption After Intracerebral Hemorrhage Based On Poxuehuayu Method

Objective: To discuss cerebral hemorrhage prescription’s the regulating effects and molecular mechanism of hematoma absorption and its products,which is conducted by activating blood and dissolving stasis method.And the discussion is based on cerebral hemorrhage model of rats and the co-culture of microglia and RBC.What’s more,to offer a more reliable foundation of clinical experiment for activating blood and dissolving stasis method.Methods:1.Setting up a rat model of Intracerebral hemorrhage induced by collagenase injection.Divide the rats into six groups: blank group,sham,model group,low-dose group,medium-dose group and high-dose group.Each group is consist of ten rats.Gavage them with 1d、 3d、5 d 、10 d three hours after modeling.Observe their neurological score,maximum section area ratio and the volume of cerebral hematoma.Observe the pathologic condition of perihematoma,through chromosome HE;Test the protein expression of CD47、CD36、CD163、PPARγ、Nrf2、TLR4、NF-κb、MyD88、TRIF、TNFα、IL-1β in perihematoma by Western blot.2.Set up an in vitro model of intracerebral hemorrhage based on the co-culture of microglia and RBC.Divide them into normal group,model group,simvastatin group,low-dose group,medium-dose group and high-dose group of cerebral hemorrhage dose.Co-culture RBC and microglia by the ratio of 40:1,and provide drug therapy at the same time.Observe the consuming situation between microglia and RBC by IFA.Test microglia’s consuming ability by FCM;Test protein expression of CD36、CD47、PPARγ、Nrf2、TLR4、NF-κb、MyD88、TRIFby Western Blot;Test CD36 DNA expression by RT-PCR;Test protein expression of TNFα、IL-1β、IL-6 by Elisa.Results:1.In animal experiments,compare with blank group and sham,the Longa5 score raised,the function of motor nerve decreased,the maximum area ratio of coronal hematomaand hematoma volume both increased in model group.By comparing with model group,the Longa5 score,the maximum area ratio and hematoma volume all decreased in other drug therapy groups.2.In animal experiments,compare with blank group,the expression of CD36、CD163、PPARγ、Nrf2 increased;The expression of CD47、TLR4、NF-κb、TRIF、MyD88、TNFα、IL-1β decreased;By comparing with model group,the expression of of CD36 mRNA in perihematoma increased.3.In the co-culture experiment between microglia and RBC.compared to model group,simvastatin group and other drug therapy groups can accelerate microglia’s consuming of RBC.4.In the co-culture experiment between microglia and RBC,the expression of CD36、CD163、PPARγ、Nrf2 raised in model group;Meanwhile,the expression of CD47、TLR4、NF-κb、TRIF、MyD88、TNFα、IL-1β decreased;By comparing with model group,the expression of CD36、CD163、PPARγ、Nrf2 increased;And the expression of CD47、TLR4、NF-κb、TRIF、MyD88、TNFα、IL-1β decreased in drug therapy groups.By comparing with blank group,the expression of CD36 mRNA increased in model group;By comparing with model group,the expression of CD36 mRNA raised in the perihemetoma of grug therapy groups.Conclusion:1.The prescription do have an effect on accelerating the ethology of hematoma animals,and also mcroglia’s consuming of RBC.Therefore,it promotes the absorption of hematoma and speed up the nerve recovery hematoma animals.2.The prescription can promote the expression of CD36 signal’s related pathways indexes such as CD36、PPARγ、Nrf2 in In vitro model of hematoma.Thus it restrained the expression of CD47、TLR4、NF-κb、TRIF、MyD88、TNFα、IL-1β.It proved that,by stimulating CD36 signal pathway,the prescription accelerates the absorption of hematoma,and also restrains the inflammatory pathways of TLR4/MYD88,lowers the damage of NST and improves the DIND.3.Through in vitro model of hematoma,we verified that the prescription’s acceleration of hematoma absorption by CD36.