| Objtctive Obesity as the most public health problem in the world,has always been of concern to human beings.With the improvement of living standards,the problem of obesity has become increasingly severe.How to solve the excessive accumulation of body fat has become a hot spot in the current study.In this study,white-fat browning was used as the entry point.Real-time PCR and Western-blot were used to explore the effects of telocyanins extracted from Radix Rehmanniae on primary white adipocytes in C57/BL6 mice.The influence of browning process was analyzed on the molecular mechanism of acteoside affecting the browning process of white adipocytes.Methods1.Check the relevant literature on obesity and rehmannia in traditional Chinese medicine,explore the understanding of Chinese medicine on obesity and rehmannia,and the use of rehmannia in the treatment of obesity.2.Resuscitate 3T3-L1 preadipocytes and subculture.After three generations of adipocytes to log phase,the cells were randomly divided into 6 groups: experimental group 1(1 u M acteoside);experimental group 2(5 u M acteoside);experimental group 3(10 u M acteoside);experimental group 4(20 u M acteoside);experimental group 5(30 u M acteoside);control group(0 u M acteoside).Browning induction was carried out for 8 days under the same induction conditions.After oil red staining was used to identify the adipogenic effect of the cells and Real-time PCR was used to detect the UCP-1 m RNA expression level.3.4-week-old C57BL/6 male mice were used as experimental subjects.After ingestion,inguinal adipose tissue was taken and used as primary white adipocytes for cell subculture.After three generations of white adipocytes are incubated to logarithmic phase,they are randomly divided into four groups: experimental group 1(5 u M acteoside);experimental group 2(5u MDMSO + forsk);experimental group 3 5 u M acteoside + forsk);Control group(5 u M DMSO).Browning induction was carried out for 8 days under the same induction conditions.After oil red staining was used to identify the adipogenic effect of cells,the expression of UCP-1 m RNA and protein was detected by Real-time PCR and Western-Blot.4.4-week-old C57BL/6 male mice were used as experimental subjects.After the sacrifice,the adipose tissue in the interscapular area was taken as the primary brown adipocytes for cell subculture.When three generations of brown adipocytes are incubated to logarithmic phase,they are randomly divided into two groups: experimental group(5 u M acteoside)and control group(5 u M DMSO).Browning induction was carried out for 8 days under the same induction conditions.After oil red staining was used to identify the adipogenic effects of cells,the expression of UCP-1 m RNA was detected by Real-time PCR.Result1.3T3-L1 prelipid cells were induced to culture,oil red staining showed good fat formation.The results of real-time PCR showed that the expression of UCP-1 m RNA in experimental group 2 and experimental group 4 was significantly higher than that in control group(P<0.01);the expression of UCP-1 m RNA in experimental group 5 was higher than that in control group(P<0.05);There was no significant difference in UCP-1 m RNA expression between experimental group 1,3 and control group(P>0.05).2.The primary white adipocytes were induced by induction,oil red staining showed a good fat-forming effect.Real-time PCR results showed that UCP-1 m RNA expression in experimental group 1 was significantly higher than that in control group(P<0.01).Compared with the control group,the experimental group 1 was no significant difference in m RNA expression with AP2,Adipq,Leptin,Pgc1α,Cidea,Prdm16 m RNA expression(P>0.05).The expression of UCP-1 and Pgc1α m RNA in experimental group 3 was significantly higher than that in control group(P<0.01).The experimental group 3 was more effective than the control group.There was no significant difference between AP2,Adipq,Leptin,Cidea,and Prdm16(P<0.05).Western-Blot’s results showed that UCP-1 protein expression was significantly increased in experimental group 1 compared with control group(P<0.01).3.The primary cultured brown adipocytes were induced by induction,oil red staining showed good fat formation.Real-time PCR showed that there was no significant difference in UCP-1 m RNA expression between the experimental group and the control group(P>0.05).Conclusion1.5u M,20 u M acteoside had a significant effect on promoting the expression of UCP-1m RNA in 3T3-L1 adipocytes.2.5u M acteoside has a significant effect on promoting the expression of UCP-1m RNA and protein in white fat cells of C57/BL6 mice.3.The effect of 5u M acteoside on the expression of UCP-1 m RNA in brown adipocytes of C57/BL6 mice was not significant.4.Actinoside has the effect of promoting the browning of white fat.5.The active ingredient of Chinese herbs Ephedra in the treatment of weight loss may be actinoside、,which may be related to the browning of white fat. |
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