Therapeutic Effect Of Glucocorticoid On Brain Injury Induced By Diquat In Rats

Research backgroundIn recent years,with the rapid increase in herbicide market sales,the incidence of acute intoxication has also increased year by year.The nervous system damage caused by the poisoning of diquat occurs from time to time,which has a serious impact on the health of patients,but there is no clear treatment plan at present.Therefore,the treatment of toxic brain injury caused by diquat has become a new problem and challenge in the field of emergency and public health.Research purposeThis study explored the experimental study of toxic brain injury and whether glucocorticoid has therapeutic effect on toxic brain injury caused by diquat through rat model infected with diquat.The experimental results were analyzed,summarized and discussed in order to provide a basis for the pathogenesis and treatment of toxic brain injury caused by diquat.Research methodA total of 200 male Wistar rats(250g)were divided into 8 groups with 25 rats in each group.Blank control group and dexamethasone control group were set up.The rats in the DQ-L,DQ-M and DQ-H groups were exposed to low,medium and high doses of diquat.The rats in the DQ+GC-L,DQ+GC-M and DQ+GC-H groups were exposed to high doses of diquat and injected with low,medium and high doses of dexamethasone for 7 consecutive days.24h,3d,7d,14d,21d after gavage.At each point,5 rats from each group were sacrificed,and brain sections were used for HE staining to observe brain histopathology.Immunohistochemical staining of astrocytes;The demyelination was observed by LFB staining.Western Blot was used to detect the protein expression of p38MAPK,NF-kB and AQP4.Elisa was used to detect tumor necrosis factor-α(TNFα),interleukin-1β(IL-1β)and interleukin-6(IL-6)in brain tissue.Immunofluorescence was used to detect neuronal death.Electron microscopy was used to observe the morphological changes of mitochondria in brain slices at 7,14 and 21 days.Experimental results(1)General conditions:All the rats showed yellow hair,lethargy,irritability,etc.,and a few of them showed unstable walking,holding PAWS and other behaviors after toxic modeling.Symptoms such as shortness of breath and diarrhea appeared after 3 days.After 7 days,some rats developed bloody secretions around the nasal cavity.1421d hair color yellow degree with time deepening.During the 21d observation period.1 died in the DQ-L group,1 in the DQ-M group.2 in the DQ-H group.2 in the DQ+GCL group.1 in the DQ+GC-M group and 1 in the DQ+GC-H group,and no died in the other groups during the experimental period.During the experiment,the GC and Ctrl groups had a normal diet and drinking water,were active,and had good mental state without any symptoms of discomfort or poisoning.(2)Body weight monitoring results:The body weight of the GC group and Ctrl group increased slowly during the experiment.Compared with Ctrl group,the body weight of rats in DQ-L group.DQ-M group,DQ-H group,DQ+GC-L group,DQ+GCM group and DQ+GC-H group in the poisoned group continued to decrease,reaching the lowest point on about 7 days,and gradually increasing from 7 days to 21 days.From day 7 to day 21.the body weight of rats in DQ-L group,DQ-M group,DQ-H group,DQ+GC-L group,DQ+GC-M group and DQ+GC-H group increased slowly.(3)Brain histopathological changes:The Ctrl group and GC group showed no obvious abnormalities.In the DQ-L group,the nuclei of nerve cells were shrunk and dark stained,and the cells were arranged irregularly.In the DQ-M group,the cells were shrunk and dark stained,the cytoplasm was vacuolar,and the number of nerve cells in the CA2 region was small.In the DQ-H group,a small number of nerve cells showed nuclear shrinkage and deep staining in the first 7 days,and a large number of nerve cells showed nuclear shrinkage and a small amount of blood vessel congestion from 7 to 14 days.Compared with the DQ-H group,the damage was alleviated in the DQ+GC-L,DQ+GC-M,and DQ+GC-H groups.In the DQ+GC-L and DQ+GC-M groups,a small number of nerve cells were found to be irregular in shape and arranged disorderly,with occasional shrinkage and deep staining of nerve cells and vacuoles in cytoplasm.In the DQ+GC-H group,the nerve cells had regular morphology,obvious nucleoli,and loose arrangement,and no other obvious abnormalities were observed.(4)Results of myelin staining of LFB:The LFB histochemical staining of the uninfected group showed bright blue,and the density of myelin sheath area in the infected group was significantly reduced.The density of myelin area in DQ-L group,DQ-M group and DQ-H group decreased,and lamellar demyelination area of different sizes could be seen,the tissue arrangement was destroyed and the gap was large,and the walking of nerve fibers was disturbed.In the DQ-H group,the density of myelin area decreased to the lowest blue value in this study.In the DQ+GC-L,DQ+GC-M and DQ+GC-H groups treated with dexamethasone intervention,the density of myelin sheath area was increased,the tissue arrangement was closer and the space was smaller than that in the DQ-H group.(5)Expression of inflammatory factors in brain tissue:Compared with Ctrl group and GC group,the contents of TNF-α,IL-6 and IL-1β in the brain tissue of the DQ-L group,DQ-M group,DQ-H group,DQ+GC-L group,DQ+GC-M group and DQ+GCH group were significantly increased at each time point after successful modeling(P<0.05).Compared with the DQ-H group,the contents of TNF-α,IL-6 and IL-1β in the brain tissue of the DQ+GC-L group,DQ+GC-M group and DQ+GC-H group decreased at 24h,3d,7d and 14d,and the differences were statistically significant(P<0.05).(6)At each time point,compared with the Ctrl group,the expressions of p38MAPK/NF-κB pathway-related proteins p65,p-p65,p38,p-p38 and AQP4 in the GC group had no significant changes(P>0.05).Compared with the Ctrl group,the expression levels of p38MAPK/NF-κB pathway-related proteins p65,p-p65,p38,pp3 8 and AQP4 in the exposed groups were significantly increased(P<0.05).Compared with the DQ-H group,the expression of p65,p-p65 and p38 in the brain tissue of the DQ+GC-L group decreased at 24h(P<0.05),and the expression of p-p65,p38,p-p38 and AQP4 in the brain tissue of the DQ+GC-M group decreased.The difference was statistically significant(P<0.05).The expression of p38MAPK/NF-κB pathwayrelated proteins p65,p-p65,p38,p-p38 and AQP4 in brain tissue of DQ+GC-H group was significantly decreased(P<0.05).Compared with the DQ-H group,the expression levels of p65,p-p65,p38,p-p38 and AQP4 in the DQ+GC-M and DQ+GC-H groups were significantly decreased on the 3rd and 7th days(P<0.05).Compared with the DQ-H group,the DQ+GC-L group,DQ+GC-M group and DQ+GC-H group had significant reductions in the expression of p65,p-p65,p38,p-p38 and AQP4 in brain tissue at 14 and 21 days(P<0.05).(7)Immunofluorescence results of neurons in brain tissue:The immunofluorescence results of all Ctrl group and GC group showed no neuronal necrosis and apoptosis at 24h,3d,7d,14d and 21d,showing obvious bright blue areas.Compared with the control group,the immunofluorescence results of DQ-L group,DQ-M group and DQ-H group showed stronger green fluorescence intensity with the increase of poison dose.The DQ+GC-L group,DQ+GC-M group and DQ+GC-H group treated with dexamethasone intervention increased with the increase of intervention dose.The area of green fluorescence decreases.(8)Electron microscopy results in brain mitochondria:The electron microscopy results of the Ctrl group and the GC group showed that there was no obvious damage to the mitochondria at 7,14,and 21 days,and most of the structure was normal.Mitochondria in brain tissue were damaged to varying degrees in the DQ-L,DQ-M,DQ-H,DQ+GC-L,DQ+GC-M and DQ+GC-H groups.The damage was the most severe in the DQ-H group,showing severe swelling of mitochondria,dissolution of matrix,disappearance of cristae,and vacuolation.Compared with the DQ-H group,the damage of mitochondria in brain tissue was alleviated in the DQ+GC-L,DQ+GC-M,and DQ+GC-H groups.Most of the mitochondria were mild to moderate swelling,the membrane was intact,the matrix became shallow and dissolved,the cristae were broken and shortened,and the matrix was more dissolved and vacuolated in severe cases.(9)Immunohistochemical results of brain astrocytes:During 24h-21 d,immunohistochemical staining results of Ctrl and GC groups showed that the nuclei in the visual field were bright purple,and the brown areas were small.The brown area in the visual field of DQ-L group,DQ-M group and DQ-H group increased with the increase of poison dose.Compared with DQ-H group,the brown area of DQ+GC-L,DQ+GC-M and DQ+GC-H groups decreased.Research conclusions(1)Diquat can cause central nervous system damage and toxic brain injury in rats.(2)Glucocorticoid has a certain therapeutic effect on toxic brain injury in rats exposed to diquat.(3)p38MAPK/NF-κB signaling pathway is involved in the inflammatory response of toxic brain injury in rats poisoned by diquat.Glucocorticoids can affect the occurrence and development of inflammation in the body by affecting proteins related to p38MAPK/NF-κB signaling pathway.