| ObjectivesSafflower yellow (SY) is the main effective ingredient of Carthamus tinctorius L. Our previous studies showed that Hydroxysafflor yellow A (HSYA), which was the main active ingredient in safflower yellow, could inhibit the proliferation and adipogenesis of murine3T3-L1preadipocytes. HSYA also notably reduced the amount of intracellular lipid and triglyceride content in adipocytes. But no data has been demonstrated about its effect on losing weight and improving insulin sensitivity. In this study, we investigate the effect of SY on body weight, body fat mass, insulin sensitivity and explore the tissue targets in obese mice induced by high fat diet.Methods8-week-old male ICR mice (n=35) were randomly divided into standard food group (SF group, n=10) and high fat diet group (n=25). Mice were weighed after8weeks and then divided into high fat diet group (HFD group, n=13) and safflower yellow intervention group (SY group, n=12). Safflower yellow (80mg/kg) was intraperitoneally injected into mice in SY group and saline in equal volume was intraperitoneally injected into mice in SF and HFD groups. After8weeks intervention, the intraperitoneal injection of glucose tolerance test (IPGTT) and insulin tolerance test (IPITT) in mice were performed, and body weight, body fat mass in different location, serum insulin levels and other biochemical parameters were measured. Subcutaneous and visceral white adipose tissue, interscapular brown adipose tissue and skeletal muscle tissue were dissected. Real-time quantitative polymerase chain reaction (RT-qPCR) and western blot technoglogies were used to investigate mRNA and protein expression of related genes in these tissues of mice.Results1. Effects of SY on body weight, body fat mass and insulin sensitivity of obese mice.Body weight of mice fed with high fat diet (HFD) for8weeks were significantly increased and reached to1.24folds of mice fed with standard food (SF)(P<0.05), implying obese mice model was established successfully. After administration obese mice with SY for8weeks, subcutaneous and perirenal fat tissue mass significantly decreased to57.4%and59.8%of that in HFD group, respectively (P<0.05). IPITT test showed that glucose levels at0,30,60minutes and the area under curve (AUC) of0-30,0-60,0-90minutes in SY group were significantly reduced when compared with those in HFD group (P<0.05).2. Effects of SY on the expression of genes involved in insulin signaling pathways in adipose tissue and skeletal muscle tissue of obese mice.2.1Effects of SY on the mRNA and protein levels of genes involved in insulin signaling pathways in mesenteric white adipose tissue of obese mice.The mRNA levels of IRS1, AKT, GSK3β and FOXO1in mesenteric white adipose tissue of HFD-induced obese mice were significantly decreased to37.1%,26.2%,16.3%and11.3%of that in SF mice (P<0.05). After administration obese mice with SY for8weeks, the mRNA levels of these genes were significantly increased to1.9-fold,2.8-fold,3.3-fold and5.9-fold of that in HFD-induced obese mice (P<0.05). In consistent with the changes in transcription level, the protein levels of AKT and GSK3β in mesenteric white adipose tissue of obese mice were significantly decreased to25.0%and25.8%, respectively, in comparison with SF mice (P<0.05). While these proteins were significantly increased to3.0-fold and5.2-fold of these in HFD-induced obese mice, respectively, after administration these mice with SY for8weeks (P<0.05).2.2Effects of SY on the mRNA and protein levels of genes involved in insulin signaling pathways in subcutaneous white adipose tissue of obese mice.The mRNA levels of AKT and FOXO1in inguinal subcutaneous white adipose tissue of HFD-induced obese mice were significantly decreased to54.7%and34.1%of that in SF mice (P<0.05), but the mRNA levels of IRS1and GSK3β were not changed (P>0.05). After administration obese mice with SY for8weeks, the mRNA levels of AKT、GSK3β and IRS1were significantly increased to1.6-fold,1.5-fold and3.3-fold of that in HFD-induced obese mice (P<0.05), but the mRNA levels of FOXO1were not changed (P>0.05). However, inconsistent with the changes in transcription level, the protein levels of AKT and GSK3β in subcutaneous white adipose tissue had no significantly differences among the three groups (P>0.05).2.3Effects of SY on the mRNA levels of genes involved in insulin signaling pathways in skeletal muscle tissue of obese mice.The mRNA levels of GLUT4and FOXO1in skeletal muscle tissue of HFD-induced obese mice were not significantly changed as compared with SF mice (P>0.05). After administration obese mice with SY for8weeks, the mRNA levels of these genes were significantly increased to8.7-fold and14.2-fold of that in HFD-induced obese mice (P<0.05).3. Effects of SY on the expression of genes involved in the browning of white adipose tissue in adipose tissue and associated with energy metabolism of obese mice.3.1Effects of SY on the mRNA and protein levels of genes involved in the browning of white adipose tissue in mesenteric white adipose tissue of obese mice.The mRNA levels of PGC1α and UCP1in mesenteric white adipose tissue of HFD-induced obese mice were significantly decreased to12.1%and5.5%of that in SF mice (P<0.05). After administration obese mice with SY for8weeks, the mRNA levels of these genes were significantly increased to6.2-fold and8.6-fold of that in HFD-induced obese mice (P<0.05). However, inconsistent with the changes in transcription level, the protein levels of PGC1α and UCP1in mesenteric white adipose tissue had no significantly differences among the three groups (P>0.05).3.2Effects of SY on the mRNA and protein levels of genes involved in the browningof white adipose tissue and associated with energy metabolism in subcutaneouswhite adipose tissue of obese mice.3.2.1Effects of SY on the mRNA and protein levels of genes involved in the browning of white adipose tissue in subcutaneous white adipose tissue of obese mice. The mRNA levels of PGC1α and FNDC5in inguinal subcutaneous white adipose tissue of HFD-induced obese mice were not significantly changed as compared with SF mice (P>0.05). After administration obese mice with SY for8weeks, the mRNA levels of these genes were significantly increased to2.5-fold and6.4-fold of that in HFD-induced obese mice (P<0.05). In consistent with the changes in transcription level, the protein levels of PGC1α in subcutaneous white adipose tissue of obese mice were significantly decreased to13.0%in comparison with SF mice (P<0.05). While the protein was significantly increased to3.0-fold of that in HFD-induced obese mice after administration these mice with SY for8weeks (P<0.05). 3.2.2Effects of SY on the mRNA levels of characteristic genes of classical brown adipose tissue in subcutaneous white adipose tissue of obese mice.The mRNA levels of PRDM16, which was specifically expressed in classical brown adipose tissue, in inguinal subcutaneous white adipose tissue of HFD-induced obese mice were not significantly changed as compared with SF mice (P>0.05). After administration obese mice with SY for8weeks, the mRNA levels of PRDM16were significantly increased to2.1-fold of that in HFD-induced obese mice (P<0.05). Additionally, the mRNA levels of CIDEA, another characteristic gene of classical brown adipose tissue, in inguinal subcutaneous white adipose tissue of HFD-induced obese mice were significantly decreased to31.8%of that in SF mice (P<0.05). While the gene was not significantly changed after administration these mice with SY for8weeks (P>0.05).3.2.3Effects of SY on the mRNA levels of PPARy2in subcutaneous white adipose tissue of obese mice.The mRNA levels of PPARy2in inguinal subcutaneous white adipose tissue of HFD-induced obese mice were not significantly changed as compared with SF mice (P>0.05). After administration obese mice with SY for8weeks, the mRNA levels of PPARy2were significantly increased to1.8-fold of that in HFD-induced obese mice (P<0.05).3.3Effects of SY on the mRNA levels of genes specifically expressed in brown adipose tissue in interscapular brown adipose tissue of obese mice.The mRNA levels of PRDM16in interscapular brown adipose tissue of HFD-induced obese mice were not significantly changed as compared with SF mice (P>0.05). After administration obese mice with SY for8weeks, the mRNA levels of PRDM16were significantly increased to4.5-fold of that in HFD-induced obese mice (P<0.05). Moreover, the mRNA levels of CIDEA in interscapular brown adipose tissue of HFD-induced obese mice were significantly decreased to47.3%of that in SF mice (P<0.05). While the gene was significantly increased to2.0-fold of that in HFD-induced obese mice after administration these mice with SY for8weeks (P<0.05).Conclusion Safflower yellow significantly reduced subcutaneous and perirenal fat mass of obese mice by promoting the browning of subcutaneous white adipose tissue. Safflower yellow could also reduce fasting blood glucose and increase insulin sensitivity in HFD-induced obese mice through activating IRS1/AKT/GSK3P insulin signaling pathway in visceral white adipose tissue, promoting PPARy2expression in subcutaneous white adipose and GLUT4expression in skeletal muscle tissue. Our study provides an important experimental evidence for developing safflower yellow as an potential loss weight drug, and it has a good application prospect. |
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