Effects Of PAS-Na On ERK/p90RSK Pathway And Inflammatory Response In Hippocampal Of Lead-stained Rats

Objective To explore and research the effect of ERK/p90RSK pathway on lead induced hippocampal inflammatory response in rats and the intervention of PAS-Na.It provides a scientific basis for further elucidating the neurotoxic mechanism of lead and the protective effect of PAS-Na on lead poisoning mechanism.Methods A total of 72 SPF grade SD rats（male）were given adaptive feeding for one week.After they were determined to have no symptoms of discomfort,they were divided into the following 6 groups according to the completely random grouping principle:normal control group,lead staining group,low,medium and high PAS-NA（L-,M-,H-PAS）treatment group,and PAS-NA control group,with 12 rats in each group.Lead-stained group,L-,M-,H-the PAS treatment group,according to the 1 time/day,5 times per week,4weeks duration frequency,were respectively received intraperitoneal injection（i.p.）of（CH₃COO）₂Pb·3H₂O 6 mg/kg,and normal control group,PAS-Na control group were injected with saline of equal volume.Then,the PAS-Na control group,the L-,M-,and H-PAS treatment groups then undercut injections（s.c.）of 300,100,200 and 300 mg/kg PAS-Na in the back,once a day,5 times a week for 3 consecutive weeks.respectively,and the normal control group were injected with saline of equal volume.The experiment indicators:（1）Growth and development indicators:the rats were weighed once a week at a fixed time.At the end of the experiment,the rats were killed after anesthesia.The liver,kidney,testis,spleen and other organs of the rats were extracted,weighed and the coefficient ratio of each organ was calculated.（2）Spatial learning and memory:Six days before the the material is taken,all rats in each group were tested through Morris water maze experiment to detect changes in the rat rats’spatial learning and memory ability.（3）Pathological section to observe the degree of hippocampal injury caused by lead and the therapeutic effect of PAS-Na Western-blot assay to determine the expression levels of ERK1/2,P90RSK,NF-κB p65 and other related proteins in brain tissues enzyme-linked immunosorbent assay（ELISA）to determine the content of inflammatory factors in brain tissues.Results（1）Growth and development of rats:there was no significant difference in the initial body weight of rats in each experimental group.From the 1st week to the 4th week after the lead staining,the body weight of rats in each group increased with time,but the body weight of rats in the lead-stained group and the L-,M-and H-PAS treatment group were at a low level than the normal control group（P<0.05）.Compared with the normal control group,the PAS-Na control group showed no significant change in weight（P>0.05）.（2）The dirty factor of rats:compared with the normal control group,it has a significant increase in liver coefficient in lead-stained groups（P<0.01）,and there was no significant change in the kidney,testicle and spleen coefficients,with no statistically significant difference.After PAS-Na treatment,it had no difference between the PAS-Na treatment group and the normal control group（P>0.05）.Compared with the lead-stained group,the liver coefficient of the middle PAS-Na group decreased（P<0.05）.（3）Changes in spatial learning and memory ability of rats:the escape latency of rats in each group fluctuated,and the overall trend was shorter.Among them,the escape incubation period of the first and second days of lead staining group was longer than the normal control group,and the difference was significantly different.（P<0.05）.There was no significant difference in escape latency between the PAS-Na control group and the normal control group（P>0.05）.The average swimming distance of rats in the lead-stained group was longer than the normal control group on day 2 and day 5,and the difference was statistically significant（P<0.05）.On day 2 and day 5 of the test,the average swimming distance in H-and M-PAS treatment group was shorter than that of the lead-stained group,and the difference was statistically significant（P<0.05）.There was no significant difference in the mean swimming distance between the PAS-Na control group and the normal control group（P>0.05）.There was no statistically significant difference in the distance,time and times of piercing in the quadrant of the platform（P>0.05）.（4）Then the rat hippocampal pathomorphism changes:Then the rat hippocampus were observed under Optical microscope.We found that in the normal control group of nerve cells in the hippocampus of rats,and the size of the microglia,the morphological structure of all normal CA2 area closely lined up cones,cell number,size and shape are normal,tissue overall uniform color,the rules of the nuclei are round or oval,more euchromatin,less heterochromatin,nucleolus visible;In the lead-stained group,the vertebral cells in the CA2 region of the hippocampus were arranged in disorder,and the cell morphology was significantly changed.The overall staining of the tissue cells was deep,and the nuclei of the nuclei were irregular like fusiform and triangular,and heterochromatin increased,presenting basophilic hyperchromatism.The pathologic changes of rats treated with PAS-Na were significantly lighter than those treated with lead.（5）Compared with the normal control group,the phosphorylation ERK protein level in the lead-stained group increased significantly（P<0.05）.Compared with the lead-stained group,the level of the L-and H-PAS treatment group reduced significantly（P<0.05）.Compared with the M-PAS group,the phosphorylation level of ERK in the H-PAS group reduced significantly（P<0.05）.PAS-Na had no effect on PAS-Na control group（P>0.05）.Compared with the normal control group,the phosphorylation level of p90RSK protein in the lead-stained group was significantly increased（P<0.05）.Compared with the lead-stained group,p90RSK protein phosphorylation level was significantly reduced in the L-PAS group（P<0.05）.PAS-Na had no effect on PAS-Na control group（P>0.05）.（6）Expression of inflammatory cytokines:Compared with the normal control group,the phosphorylation level of NF-κB p65 was significantly increased in the lead-stained group（P<0.05）.Compared with the lead-stained group,the phosphorylation levels of NF-κB p65 in the L-and H-PAS treatment groups were reduced significantly（P<0.05）.PAS-Na had no effect on PAS-Na control group（P>0.05）.Compared with the normal control group,the content of IL-1 in the hippocampus of rats exposed to lead increased,with statistically significant difference（P<0.05）.Compared with the lead-stained group,the content of Il-1βin the hippocampus of rats treated with L-,M-and H-PAS treated group decreased significantly（P<0.05）.Compared with the normal control group,the Il-1βlevel in the PAS-Na control group decreased significantly,and the difference was statistically significant（P<0.05）.Compared with the normal control group,the IL-6 level in the hippocampal tissue of the lead-stained rats had no significant change（P>0.05）.Conclusion:（1）Lead contamination can cause weight loss in rats and has certain toxicity.（2）lead staining affects the learning and memory ability of rats,at the same time,PAS-Na treatment has a certain effect on the improvement of learning and memory ability in rats.（3）lead staining can cause pathological damage to the hippocampal tissue of rats,and PAS-Na has a certain improvement effect.（4）Lead staining can cause increased phosphorylation level of ERK/p90RSK pathway related proteins in the hippocampus of rats,and PAS-Na has a certain antagonistic effect on this change.（5）Lead exposure caused increased levels of inflammatory factors in the hippocampal tissues of rats,and PAS-Na had a certain antagonistic effect on hippocampal inflammation.