| A large number of epidemiological evidences indicate that the incidence of cardiovascular and cerebrovascular diseases is high in winter,and the mortality rate of patients with cardio-cerebral vascular disease is significantly higher than that of other seasons.Meanwhile,a large number of epidemiological prospective studies indicate that the incidence of elevated blood lipids in humans in winter or cold regions.A significant increase,mechanism study may be related to the cold stimulation through the UCP1 pathway to activate the adipose tissue heat-related,but there are also related mechanisms found that cold blood lipids after stimulation,and for different species of mice,blood lipid results vary,combined with the above study The results are intended to carry out basic experiments to explore the mechanism of cold stimulation and blood lipids and clinical observation studies.MethodsExperiment?(Clinical Trial)Recruitment and screening of 9 subjects who meet the enrollment conditions,signed the test informed consent,the subjects were allowed to sit at 25? for 10 minutes,and baseline levels of blood pressure,blood lipids,and angiotensin ? were measured before cold exposure.Cold exposure was performed at 180C for 2 hours.After the end of cold exposure,indexes such as blood pressure,blood lipids,and angiotensin ? were rechecked,and the results were statistically analyzed.Experiment?(basic experiment)According to the different requirements of the experiment,ApoE-/-mice,C57BL/6J and LDLR-/-mice were randomly divided into groups and kept at 30? and 4?respectively.The body weight and food intake of mice were recorded and the corresponding stimulation conditions were completed.,Collect samples to be tested:first separate the serum and store it in a 1.5ml centrifuge tube,and store at-80?;according to the follow-up needs for molecular biology or pathological staining,the obtained tissue should be stored in liquid nitrogen or more than 4%Polyoxymethylene solution or 1%PBS solution was fixed.Blood lipid levels were measured by biochemical methods;antibodies were used in paraffin sections of each site to measure the expression of the target protein by immunohistochemistry;cholesterol efflux and influx rates in adipose tissue of C57BL/6J mice were detected by radioisotope assay.As well as the cholesterol efflux rate of APOE-/-mice adipose tissue,the cholesterol status was observed;the changes of adipocyte morphology,coloration,etc.were observed by HE staining to clarify the browning of adipose tissue and activation of brown adipose tissue;and by Western Blot method The expression levels of LDLR in liver of UCP1,LDLR protein and C57BL/6J mice were detected in adipose tissue of each site.Finally,the data were statistically analyzed.ResultsExperimentI(Clinical Trial)Short-term cold stimulation can increase the levels of TC,LDL-C,and TG in humans,and there is a significant statistical difference in the elevation of TC levels.The levels of LDL-C and TG have a rising trend,but the results are not statistically different;short-term cold stimulation can The level of angiotensin ?(Ang ?)and blood pressure(BP)increased and the level of diastolic blood pressure increased statistically.Ang ? and systolic blood pressure had a tendency to increase,but the results were not statistically different.Experiment?(basic experiment)?Effect of persistent cold stimulation at 4 weeks on browning of white adipose tissue and activation of brown adipose tissue in miceBy HE staining of adipose tissue of C57BL/6J and LDLR-/-mice,it was found that compared with the control group at 30?,the 4? group of cold stimulation group was sWAT and scapular adipose tissue in 4?mice(iBAT)fat cells decreased in size and darkened;immunohistochemical staining of UCP1 positive staining was deepened,and Western Blot detected a significant increase in the expression of ApoE-/-mice and C57BL/6J UCPl protein;however,epididymal adipose tissue was observed in the 4? group(Compared with 30?,only a very slight cell volume became smaller,and HE staining was only mildly stained;the positive staining of UCP1 immunohistochemistry did not increase.?Four weeks of cold stimulation on body weight and blood lipid levels in miceCompared with the control group at 30?,4? continuous cold stimulation,namely 4?group,significantly reduced the body weight of C57BL/6J mice;4? group significantly decreased the levels of TC,LDL-C and TG in C57BL/6J mice,and the decline level had Significant statistical difference;4-week continuous cold stimulation increased TC and LDL-C levels in ApoE-/-mice,and significantly decreased TG levels;4? group decreased TG levels in LDLR-/-mice,and there was a statistical difference,TC The level of LDL-C decreased slightly,with no significant effect,and there was no statistical difference in the decrease.?Four-week continuous cold stimulation increased adipose tissue and liver LDLR levels in C57BL/6J miceCompared with the control group at 30?,cold stimuli at 4 weeks increased the expression of LDLR protein in adipose tissue of C57BL/6J mice,but the results were not statistically different.Compared with 30? control group,4 weeks.Continuous cold stimulation increased the level of liver LDLR protein in C57BL/6J mice,and the results were statistically significant.?Effect of persistent cold stimulation at 4 weeks on cholesterol status in adipose tissue of C57BL/6J miceCompared with the control group at 30?,continuous cold stimulation at 4 weeks reduced the cholesterol efflux rate in the adipose tissue of C57BL/6J and APOE-/-mice.There was a statistically significant difference in the decrease of inguinal adipose tissue and epididymal adipose tissue of the mice,and scapular fat was observed.The decrease in tissue was not obvious;however,compared with the control group at 30?,continuous cold stimulation at 4 weeks increased the cholesterol influx rate of adipose tissue in C57BL/6J mice.The results were statistically different.The inflow rate of adipose tissue in the scapular area was lower than that in the inguinal adipose tissue and epididymis.Adipose tissue increased significantly.ConclusionsExperiment I(Clinical Trial)?Short-term cold stimulation increased the level of TC in humans,and short-term cold stimulation had no significant effect on human TG and LDL-C levels.?Short-term cold stimulation increased systolic blood pressure effect was not significant,short-term cold stimulation can significantly increase diastolic blood pressure.?Short-term cold stimulation has a tendency to increase the level of angiotensin ?(Ang?),but there is no statistical difference.Experiment ?(basic experiment)?4-week cold stimulation could cause browning in the inguinal white adipose tissue(BRITE)and activation of brown adipose tissue in the scapular apoE-/-mice and C57BL/6J mice,but had no significant effect on epididymal white adipose tissue.?Four weeks of cold stimulation can significantly reduce the blood lipid levels of C57BL/6J mice,increase the levels of TC/LDL-C in ApoE-/-mice,and decrease the TG levels in LDLR-/-mice.No TC and LDL-C levels were observed.Obvious effect.?After 4 weeks of continuous cold stimulation,the expression of LDLR protein in adipose tissue and liver of C57BL/6J mice was increased.?Four weeks of continuous cold stimulation reduced the cholesterol efflux rate of C57BL/6J and APOE-/-mice adipose tissue,and increased the C57BL/6J mice cholesterol influx rate.?After 4 weeks of cold stimulation,body weight of C57BL/6J mice was reduced,but there was no significant effect on body weight of ApoE-/-and LDLR-/-mice.Cold stimulation at 4 weeks increased the food intake of C57BL/6J mice. |
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