Effect of GLP-1 receptor agonist on lipolysis in adipose tissue of obese mice and its underlying mechanism

AIM: To investigate the effects of glucagon-like peptide-1(GLP-1) receptor agonist exendin-4 on white adipose tissue (WAT) and the underlying mechanisms. METHODS: Male C57BL/6J mice (8 weeks) were challenged by high-fat diet for 12 weeks, and were randomly divided into saline group and exendin-4 group. The mRNA expression of sirtuin 1(SIRT1), adipose triglyceride lipase (ATGL), TNF-α and adiponectin of WAT was detected by real-time PCR. 3T3-L1 adipocytes or mouse embryonic fibroblasts cells were treated with exendin-4 for 24 h. The protein levels of SIRT1, ATGL and hormone-sensitive lipase (HSL) were determined by Western blot.RESULTS: Exendin-4 significantly decreased epididymal fat weight, fasting blood glucose and serum triglyceride levels (P<0.05), and reduced body weight and serum TNF-α level. The mRNA expression of SIRT1, ATGL and adiponectin in WAT was all significantly up-regulated by exendin-4, which were contrary to the down-regulation of TNF-α mRNA expression (P<0.05). Exendin-4 promoted the protein expression of SIRT1, ATGL, and HSL in 3T3-L1 adipocytes in a dose-dependent manner. Less lipid droplets with up-regulation of lipolytic protein expression were observed when combined with SIRT1 agonist treatment, which were suppressed by SIRT1 inhibitor. Deletion of SIRT1 led to larger adipocytes with more lipid droplets, and the effect of exendin-4 on the lipolysis disappeared when SIRT1 was deficient.CONCLUSION: Exendin-4 promotes lipolysis in WAT of obese mice via activation of SIRT1.     

Effect of pioglitazone on pre-adipocytes differentiation and GILZ expression

AIM: To investigate the roles of pioglitazone on differentiation and expression of GILZ in 3T3-L1 pre-adipocytes. METHODS: The morphological changes during 3T3-L1 cell differentiation were observed. The cells were treated with pioglitazone at concentrations of 1×10^-4～1×10^-2 mmol/L for 48 h, then the relative content of triglyceride were analyzed by oil red O staining at 2nd, 4th and 6th day during adipogenesis. The mRNA expression of peroxisome proliferator-activated receptor γ2 (PPARγ2) and lipoprotein lipase (LPL) was measured by real-time PCR. GILZ protein expression was determined by Western blot after the cells were treated with pioglitazone at concentrations of 1×10^-4 ～1×10^-2 mmol/L for 48 h.RESULTS: Oil-red O staining showed that the relative contents of triglyceride in adipocytes were increased with the increase in the pioglitazone concentration. Compared with the control, the relative contents of triglyceride in group 1×10^-3 mmol/L and group 1×10^-2 mmol/L were significantly increased (P < 0.05). The mRNA expression of PPARγ2 and LPL was also increased with the increase in the pioglitazone concentration. When pioglitazone concentration was more than 1×10^-3 mmol/L, compared with the control, the mRNA expression of PPARγ2 and LPL significantly increased (P < 0.01). The protein expression of GILZ was decreased with the increase in the pioglitazone concentration.CONCLUSION: Pioglitazone down-regulates GILZ expression, and up-regulate PPARγ2 expression and the downstream functional factor such as LPL.     

Effects of ERK1/2/PPARα/SCAD signal pathways on physiological cardiac hypertrophy and pathological cardiac hypertrophy

AIM：To investigate the different effects of ERK1/2/PPARα/SCAD (short-chain acyl-CoA dehydrogenase) signal pathways on the cardiac hypertrophy induced by insulin-like growth factors 1 (IGF-1) or phenylephrine (PE). METHODS：The neonatal rat cardiomyocytes induced by IGF-1 were used as the model of physiological cardiac hypertrophy, and those induced by PE were used as the model of pathological cardiac hypertrophy. The surface area of the cardiomyocytes, the expression of p-ERK1/2, PPARα and SCAD, the activity of SCAD and the content of free fatty acid in the cardiomyocytes were measured. RESULTS：Compared with the control cells, the surface area of the cardiomyocytes induced by IGF-1 and PE were both increased. Compared with the controls, the expression of SCAD and PPARα, and the activity of SCAD in the cardiomyocytes induced by IGF-1 were increased, while the expression of p-ERK1/2 was decreased. However, the cardiomyocytes treated with PE showed decreased expression of SCAD and PPARα, decreased activity of SCAD and increased expression of p-ERK1/2. Meanwhile, the decrease in free fatty acid in IGF-1-induced cardiomyocytes and the increase in PE-induced cardiomyocytes indicated that the fatty acid utilization was increased in the cardiomyocytes induced by IGF-1, but decreased in the cardiomyocytes induced by PE. CONCLUSION：The changes of p-ERK1/2, PPARα and SCAD in the cardiac hypertrophy induced by IGF-1 or PE indicate that the effects of ERK1/2/PPARα/SCAD signal pathways are different between physiological cardiac hypertrophy and pathological cardiac hypertrophy, and that SCAD may be a molecular marker of these 2 different cardiac hypertrophies and a potential therapeutic target for pathological cardiac hypertrophy.     

Expression of histone chaperone ASF1B in prostate cancer cells and its effect on cell viability in vitro

AIM To investigate the expression of histone chaperone anti-silencing function 1B （ASF1B） in prostate cancer cells and its effect on cell viability in vitro. METHODS Human prostate cancer PC-3 cells were used, and knockdown of ASF1B was conducted by small interfering RNA （siRNA） transfection into the cells. The cells were divided into control group, siRNA negative control vector （mock） group and siRNA-ASF1B group. The viability of the PC-3 cells treated with ASF1B-siRNA for 12, 24 and 48 h was measured by MTT assay. Flow cytometry was used to analyze cell apoptosis and cell cycle distribution. The mRNA expression of apoptosis-related molecules was detected by RT-qPCR, and the expression levels of MAPK/JNK/ERK signaling pathway-related proteins were determined by Western blot. RESULTS The protein level of ASF1B in the normal cells （benign prostatic hyperplasia） was significantly lower than that in the PC-3 cells （P<0.01）. Compared with control group and mock group, the protein expression level of ASF1B in the PC-3 cells transfected with siRNA-ASF1B plasmid and the viability of the PC-3 cells were significantly decreased （P<0.01）. The apoptotic rate of the PC-3 cells transfected with siRNA-ASF1B plasmid was significantly higher than that in control group （P<0.01）, and the cell cycle was arrested at G₁ phase. The mRNA levels of p53, caspase-3, Bax and PARP-1 in the PC-3 cells transfected with siRNA-ASF1B plasmid were up-regulated compared with those in control group and Mack group （P<0.01）. In addition, the protein levels of MAP2K4 and p-JNK in the PC-3 cells in siRNA-ASF1B group were significantly higher than those in mock group （P<0.01）, while the protein level of p-ERK was significantly lower than that in mock group （P<0.01）. CONCLUSION ASF1B silencing induces G₁ arrest and promotes apoptosis of PC-3 cells. Activating MAPK/JNK/ERK signaling pathway may be a possible contributor to the anti-prostate cancer effect of siRNA-ASF1B.     

Protective effect of paeoniflorin on nerve cells in APP/PS1 mice and its mechanism

AIM: To investigate the therapeutic and preventive effects of paeoniflorin (PF) on APP/PS1 mice, and to explore the possible mechanism. METHODS: Fifteen male 5-month-old APP/PS1 non-dominant mice were chosen as normal control group, 15 male 5-month-old APP/PS1 double transgenic mice were used as model group, and 15 male 5-month-old APP/PS1 double transgenic mice treated with 5 mg/kg PF by intraperitoneal injection were allocated in administation group. The learning and memory ability of the mice in each group was detected by Morris water maze. The apoptosis was assessed by TUNEL fluorescence staining. The protein expression of PI3K, Akt, p-PI3K, p-Akt, caspase-3, caspase-9, Bcl-2 and Bax in cerebral cortex and hippocampus was detected by Western Blot. The protein expression levels and distribution of caspase-3 and caspase-9 were detected by immunohistochemistry. RESULTS: (1) Compared with normal control group, the learning and memory ability declined in APP/PS1 model group. Compared with APP/PS1 model group, PF obviously improve the ability of learning and memory in mice. (2) Compared with normal control group, the apoptosis of nerve cells in APP/PS1 model group significantly increased and distributed in wider areas, while that in PF group was reduced (P<0.05). (3) Compared with APP/PS1 model group, PF could significantly lower pro-apoptotic factors, caspase-3, caspase-9 and Bax (P<0.05), and increase the expression of anti-apoptotic factors, p-PI3K, p-Akt and Bcl-2 (P<0.05). CONCLUSION: PF can up-regulate the expression of Bcl-2 and down-regulate the expression levels of caspase-9, caspase-3 and Bax via the activation of PI3K/Akt pathway, thereby inhibiting the nerve cell apoptosis and protecting the nerve cells, so as to treat neurodegenerative diseases.