Expression Of Ppar-gamma 2 Of The Primary Cultured Rat Adipocyte Affected By PAES

Environment is material foundation of human survival and development. However, with the development of modern industry, many toxic artificial organic materials enter human body through various channels, and accumulate in the human body to cause lasting damage to human body. Phthalic acid ester (PAEs) is a global environmental pollutant, which can not only interfere with multiple links in human reproductive development, but also cause cancer. Recently, it is suggested PAEs in the human body have reached a high volume, and although the content of PAEs in different region are not the same, potential hazard of PAEs to human is very dangerous. And studies show that PAEs may also be relative to human obesity, because that PAEs would accumulate in fat tissue, then the hazard of PAEs to human may have surpassed the extent that we have assessed before. Therefore, it is necessary to conduct profound research about the relationship between the PAEs and obesity. This article establishes a model of primary culture of rat adipocytes, that after PAEs contaminated rat adipocytes, observe the changing of shape of adipocytes,lipochondria,correlated mRNA and different changes,then analyze effect of PAEs to rat adipocytes, to provide information for research for the relationships of PAEs and obesity.The main contents and the results of research are as follows:1. Establish a model of primary culture of rat adipocytes. Rats were executed by breaking neck. Then extract rats'adipose tissues, use digestive enzymes to digest adipose tissues, filter levitation liquid of adipose tissues through the stainless steel cell screen, remove the sediment after centrifuge cells, fluidize cells using serum free basal medium, culture adipocytes, observe the change of adipocytes to determine appropriate conditions of culture the rat adipocytes. Experimental results show that the appropriate conditions of primary adipocytes culture is taking adipose tissues in the groin and epididymis of rats, and digesting for 90min usingⅠtype collagenase, filtering with 200 meshes aperture cells sieve, centrifuging at 1000rpm speed twice to sedimentation heavy suspended cells.2. TO observe various changes of adipocytes and evaluate the effects by culturing adipocytes after contaminating using PAEs.Main experiment:①TO observe shape changes of adipocytes every day by microscope;②Dye adipocytes using Oil red O, and to observe the change of lipochondrion in adipocytes;③Oil red O dyeing extraction and colorimetric MTT method detect growth of adipocytes;④Extract the total RNA of adipocytes, through reverse transcription PCR, and analysis expressions of peroxisome proliferation agent receptor gamma-2(PPAR-γ2) mRNA. PPAR-γ2 is closely related with adipocytes growth. Experimental results show that,①through the microscope observation, cells processed by PAEs is found to be initial small circular, to spindle, finally to become the oval . Along with the increase of culturing time, lipochondrion in adipocytes began to appear, and gradually increase, finally part of lipochondrion fusion to a bigger lipochondrion, accord with the developing law of adipocytes.②Through the oil red O dyeing, we confirmed fat cells treated by PAEs have produced many lipochondrion.③Through the oil red O dyeing extraction and determined by MTT colorimetric method, found that as medium of concentration of PAEs increases, rat adipocytes produce a growing number of lipochondrion.④Through reverse transcription PCR, found that expression of PPAR-γ2 increase as well as concentration of PAEs.Conclusion: the effect to growth of mature adipocytes appears increasing of the number, increasing of lipochondrion, expression of PPAR-γ2 mRNA enhancing. By comprehensive evaluation of these effects, researchers believe that PAEs can be argued to promote the growth and development of adipocytes by enhancing PPAR-γ2 express. The results of this study will provide the scientific basis for the relationship with obesity, and provide a new idea for the the further research of the effects of PAEs to health.