Experimental Research On The Relationship And Mechanism Between Propofol And Obesity On Cognitive Function

PART ? THE RELATIONSHIP BETWEEN PROPOFOL AND OBESITY ON COGNITIVE FUNCTIONObjective: To observe the effect of propofol on cognitive function in normal and obese rats,and to explore the correlation between propofol and obesity on cognitive function.Methods: 1.One hundred and fifty six 21-day-old male SpragueDawley(SD)rats were randomly divided into two groups.The animals in the two groups were given different diets,36 rats in normal chow control group and 120 rats in high-fat diet group,for 4 weeks.Animals in high-fat diet group that were greater than chow control group mean body weight + 1.4*Standard Deviation were designated as obesity rats,according to the method that others have previously used.2.Six rats were randomly selected from each group.The serum was collected to detect the blood lipid level,and the bilateral perirenal,peritesticular and mesenteric fat were taken and weighed.The other rats in each group were divided into two subgroups.Normal-intralipid group(NL,n=12)was given an identical volume of vehicle control 10% intralipid.Normal-propofol group(NP,n=12)was given intraperitoneal injections of propofol.The obesity-intralipid(OL,n=12)and obesity-propofol(OP,n=12)groups were given intralipid and propofol,respectively.Propofol and intralipid were injected intraperitoneally at doses of 100 mg/kg and 10 ml/kg.Animals received one injection every day for seven consecutive days.3.Morris water maze experiment was carried out to test the spatial learning and memory ability.Results:1.There were 49 rats meeting the obese criterion after 4 weeks of high fat diet,the success rate of obesity modeling was 41.11%(49/120).2.Rats in obesity group gained more body weight than normal chow control(p<0.01),and the total wet weight of fat in the kidney,testis and mesentery of rats in obesity group increased significantly(p<0.05).Compared with normal control group,the level of high density lipoprotein(HDL)in obesity group was significantly lower(p<0.05),the level of low density lipoprotein(LDL),total cholesterol(TC)and triglyceride(TG)was significantly higher(p<0.05),in obesity group.3.Compared with NL group,the average time of escape latency in NP and OL group was longer on the first and second day(p<0.01);compared with NP and OL group,the average time of escape latency of OP group was longer on the second to fifth day significantly p<0.05).Compared with NP group and OL group,the number of platform location crossing was less(p<0.05)and the target quadrant time in OP group was significantly shorter(p<0.05).Conclusion: Propofol lastingly impairs spatial learning and memory function in obesity rats.Obesity or propofol anesthesia per se induces shortterm and reversible spatial learning and memory impairment.Obesity may be an important susceptibility basis for propofol to damage spatial learning and memory function.PART ? RESEARCH ON THE MECHANISM OF PROPOFOL IMPAIRED COGNITIVE FUNCTION IN OBESITY RATSObjective: To explore the mechanism of propofol induced cognitive impairment in obesity rats,and to study the mechanism of susceptibility to propofol induced cognitive impairment in obesity rats.Methods: The same group and treatment as part one.The experimental groups included: Normal-intralipid group(NL),normal-propofol group(NP),obesity-intralipid group(OL)and obesity-propofol group(OP).The treatment was the same as first part.Animals were sacrificed after 24 hours of propofol injection.The expression of cleaved caspase-3?Bcl-2?Bax?BDNF-PI3K/Akt/GSK-3? signal pathway proteins and p-tau were detected by western blot or immunohistochemistry.The ultrastructural changes of neurons were observed by transmission electron microscopy,the expression of ?-Catenin in hippocampal cells was observed by immunofluorescence.Results: 1.Compared with NP and OL group,there was a significant increase in apoptosis and expression of cleaved-caspase-3(p<0.01)in OP group.Compared with NL group,the ratio of Bcl-2/Bax in OL group decreased significantly(p<0.01);compared with NP group and OL group,the ratio of Bcl-2/Bax decreased significantly in OP group(p<0.01).2.Transmission electron microscopy showed that there was no significant change in hippocampal neurons in NL and NP groups.In the OL group,there was mild invagination in the plasma membrane and chromatin margination.In OP group,the stroma was deeply stained,nuclear pyknosis and apoptosis were obvious.In addition,there was no significant change in mitochondria of hippocampal neurons in NL and NP group,swelling and degeneration were found in OL group,while swelling,vacuolation and even fragmentation were found in OP group.3.Compared with NL group,the expression of CytC protein increased significantly in OL group(p<0.05);compared with NP group and OL group,the expression of Cyt-C protein increased significantly in OP group(p<0.01).4.Compared with NL group,the expression of pGSK-3? in NP group was significantly higher than that in NP group(p<0.05),the expression of BDNF-PI3K/Akt/GSK-3? signal pathway protein in OL group was decreased;while that in OP group was further decreased compared with OL and NP group.5.Immunofluorescence showed that ?-Catenin was mainly expressed in cytoplasm in NL group,while the expression of ?-Catenin transfered into nucleus in OP group.6.Compared with NP and OL group,phosphorylation of tau protein increased significantly in OP group(p<0.01).Conclusion: In obesity,hippocampal cells are more sensitive to propofol induced injury.Propofol exposure can induce apoptosis and phosphorylation of tau protein in the hippocampus of obesity rats.The slight mitochondrial damage and the inhibition of BDNF-PI3K/Akt/GSK-3? pathway in hippocampal cells under obesity may be the important vulnerable basis.PART ? THE EFFECT AND MECHANISM OF PROPOFOL AND PALMITIC ACID ON HIPPOCAMPAL NEURONSObjective: To observe the effect of propofol and palmitic acid treatment on apoptosis of hippocampal neurons,and to explore their correlation in hippocampal neuron injury.Methods: 1.18-20 days pregnant SD rats,hippocampal neurons were cultured with serum-free neurobasal medium added B27 nerve growth factor.2.Neuron specific antibody NF200 was used to identify neurons and calculate the culture purity.3.In order to select the appropriate concentration and time of propofol and palmitic acid treatment,different concentrations and time of palmitic acid and propofol treatment were set up on seventh 7th day of neurons culture.The toxicity of propofol and palmitic acid exposure on cells were detected by CCK8 kit.4.According to the results of CCK8,the exposure concentration and time of propofol and palmitic acid were determined.Neurons were treated with propofol and palmitic acid on the 7th day.The experimental groups were as follows: intralipid-control(IC),propofol-control(PC),palmitic-intralipid(PI),palmitic-propofol(PP).5.After the interventions,immunofluorescence and western blot were used to detected expression level of apoptosis and molecular pathway.Results: 1.The hippocampal neurons were successfully cultured.The purity of cultured neurons were over 95%.2.CCK8 showed that the exposure time of palmitic acid was 24 hours,concentration was 80 ?M,the exposure time of propofol was 2 hours,and concentration was 50 ?M,which had no effect significant on cell viability.3.The expression of cleaved caspase-3 in PP group increased significantly(p<0.01),there was no significant difference between the other groups(p>0.05).Compared with IC group,Bcl-2/Bax in PI group decreased significantly(p<0.01).Compared with PC group and PI group,Bcl-2/Bax in PP group decreased significantly(p<0.01).4.Immunofluorescence showed that the expression of Cyt-C was significantly increased in PP group.?-Catenin mainly expressed in cytoplasm in IC and PC group.The expression of ?-Catenin transfered into nucleus in OP group.5.The expression of p-tau significantly increased in PP group(p<0.05).6.Western blot showed that BDNF-PI3K/Akt/GSK-3? signal pathway did not change significantly in PC group,while the expression of protein in this signal pathway decreased in PI group.BDNFPI3K/Akt/GSK-3? signal pathway in OP group was further inhibited compared with PI and PC group.Conclusion: After palmitic acid exposure,propofol significantly increased apoptosis and phosphorylation of tau protein.The inhibition of BDNF-PI3K/Akt/GSK-3? pathway after palmitic acid exposure may be an important mechanism of propofol induced apoptosis of hippocampal neurons.