The Neuroprotective Mechanism Of Liangxue Tongyu Prescription In Regulating Intestinal Flora For Treatment Of Intracerebral Hemorrhage

Objective:To study the neuroprotective mechanism of Liangxue Tongyu Recipe in regulating intestinal microbial therapy for cerebral hemorrhage.Methods:SD rats were randomly divided into normal group,sham operation group,model group,Liangxue Tongyu group and probiotic group to establish a model of cerebral hemorrhage.On the first day,the third day and the fifth day after treatment,the neuroprotective index was determined by Longa five-point scale and balance beam test;the hematoma content was determined by hemoglobin indirect conversion method.Microbial environmental genomics sequencing was performed on the feces of cerebral hemorrhage rats,and the effects of Liangxue Tongyu Recipe on intestinal microbes were analyzed.Western blot was used to detect the expression of BDNF protein in brain tissue of rats with cerebral hemorrhage.The effect of Liangxue Tongyu Recipe on neuron protection was studied.The serum levels of TLR4,IL-1 and IL-6 were determined by ELISA.TLR4 was detected by RT-PCR.The effect of Liangxue Tongyu Recipe on inflammatory immune function was studied.RT-PCR was used to detect the expression of CuZnSOD mRNA in brain tissue,and the content of MDA in brain tissue and serum was determined by thiobarbituric acid reaction.The effect of Liangxue Tongyu Recipe on oxidative stress was studied.Results:1.(1)Impact on neuroprotective index:Behavioral score:The neurological deficit of the Liangxue Tongyu group and the probiotic group was significantly improved.The Longa and balance beam test scores on the 3rd and 5th day after administration were significantly lower.Model group(P<0.05,P<0.01);(2)Hematoma content in brain tissue:On day 3 and day 5,the content of cerebral hematoma in Liangxuetongyu group and probiotic group was significantly lower than that in model group,which significantly promoted hematoma absorption(P<0.05).2.(1)Beta diversity analysis of intestinal microbiota:The distribution of intestinal microbiota distribution model group was similar on the 3rd day and the 5th day;the first day of the Liangxue Tongyu group was similar to the first day of the model group;The distributions on the 3rd and 5th day of the blood-passing group were the most similar,and the point group distribution was similar to that of the sham-operated group.It is indicated that Liangxue Tongyu Recipe can improve the composition of intestinal flora.(2)Alpha diversity analysis of intestinal microbial species richness and uniformity:Chao index,Ace index,Shannon index,Simpson index and indexing grade curve showed that the sham operation group 5th day,Liangxue Tongyu group 1st day and 5th day have the highest intestinal microbial species richness and uniformity.On the first day,the third day of the model group and the first day of the sham operation group,the species richness and uniformity were the lowest.This indicates that cerebral hemorrhage can cause the decrease of intestinal microbial richness and disorder of microbial distribution,and the treatment of Liangxue Tongyu Recipe can restore the richness of intestinal microflora and promote the uniform distribution of intestinal microbes.3.(1)The expression of BDNF protein in brain tissue by Western Blot showed that the expression of BDNF protein in the Liangxue Tongyu group was significantly higher than that in the model group on the 1st,3rd,and 5th day of the probiotic group(P<0.01).Liangxue Tongyu Recipe and probiotics all exert neurotrophic effects.(2)The content of TLR4 in serum by ELISA showed that the third day of Liangxue Tongyu group and the first day of probiotic group were significantly lower than the model group(p<0.05).The detection of IL-1? in serum showed that the first day of the Liangxue Tongyu group and the first day of the probiotic group were significantly lower than the model group(p<0.05).The detection of IL-6 in serum showed that the first and third days of the Liangxue Tongyu group and the first and third days of the probiotic group were significantly lower than the model group(p<0.01).Both Liangxue Tongyu Recipe and probiotics inhibited the inflammatory response.(3)The expression of TLR4 in brain tissue by RT-PCR showed that the 5th day of Liangxue Tongyu group was significantly lower than that of the model group(p<0.05).The detection of IL-1?expression in brain tissue showed that the first day of the Liangxue Tongyu group and the first day of the probiotic group were significantly lower than the model group(p<0.01).There was no significant difference in the expression of IL-6 in brain tissue(p>0.05).Both Liangxue Tongyu Recipe and probiotics inhibited the inflammatory response.4.(1)RT-PCR detection of CuZnSOD expression in brain tissue showed that the first,third,and fifth days of the Liangxue Tongyu group and the first group of the probiotic group were significantly lower than the model group(p<0.01).Liangxue Tongyu Recipe and probiotics all reduced oxidative stress;(2)MDA content in brain tissue:The first,third and fifth days of Liangxue Tongyu group and the first and third days of probiotic group were significantly lower than those of the model group(p<0.05);serum MDA content:the third and fifth days of Liangxue Tongyu group and the third group of probiotics group were significantly lower than the model group(p<0.05),Liangxue Tongyu Recipe and probiotics all reduced oxidative stress damage.Conclusion:Liangxue Tongyu Recipe can improve neurological deficits and promote hematoma absorption,thus reducing brain damage.Liangxue Tongyu Recipe can restore the richness of intestinal microbiota and promote the uniform distribution of intestinal microbes,thus restoring the normal composition of intestinal flora.Liangxue Tongyu Recipe can increase the expression of BDNF,inhibit the expression of TLR4,IL-1?,IL-6 inflammatory factors,thereby nourishing neurons,exerting anti-inflammatory effects and protecting nerve function.Liangxue Tongyu Recipe can alleviate oxidative stress and exert neuroprotective effects.