Protective effect of SIRT1 on rat mesangial cells by decreasing high glucose-induced acetylation of NF-κB p65

AIM: To investigate the effects of silent information regulator 1 (SIRT1) on high glucose-induced acetylation of NF-κB p65 subunit and its protective role in rat mesangial cells. METHODS: Rat mesangial cells were cultured in DMEM supplemented with 10% FBS and were divided into control group, mannitol group, high glucose group, resveratrol group and SIRT1 RNAi group. The cell viability was determined by MTT assay. The mRNA expression of SIRT1, monocyte chemoattratant protein 1 (MCP-1), vascular cell adhesion molecule 1 (VCAM-1-1), tumor necrosis factor α (TNF-α), transforming growth factor β1 (TGF-β1) was analyzed by real-time quantitative PCR. The protein expression of SIRT1 and the acetylation of NF-κB p65 subunit were determined by Western blotting. The protein concentrations of MCP-1, VCAM-1, TNF-α, TGF-β1 and malondialdehyde (MDA) were detected by ELISA. RESULTS: The cell viability, superoxide dismutase (SOD) activity, and the expression of SIRT1 at mRNA and protein levels were decreased by high glucose treatment as compared with control group. The acetylation of NF-κB p65 subunit was significantly increased after interfered with high glucose, resulting in the increase in the secretion of MCP-1, VCAM-1, TNF-α and TGF-β1. Resveratrol decreased high glucose-induced acetylation of NF-κB p65 subunit. However, silencing SIRT1 significantly enhanced the acetylation of NF-κB p65 subunit and the expression of MCP-1, VCAM-1, TNF-α and TGF-β1. CONCLUSION: SIRT1 remarkably inhibits the inflammatory reactions by deacetylating NF-κB p65, suggesting that SIRT1 is a possible target for preventing diabetic nephropathy.     

Dihydroartemisinin enhances antitumor effect of 5-fluorouracil against gastric cancer by down-regulating SIRT1 expression

AIM: To investigate the effect of dihydroartemisinin (DHA) adjuvant treatment on enhancing the antitumor effect of 5-fluorouracil (5-FU) against gastric cancer. METHODS: The gastric cancer BGC-823 cells were divided into control group, DHA group, 5-FU group, 5-FU+DHA group and 5-FU+DHA+SIRT1 plasmid group. The viability of BGC-823 cells treated with DHA and 5-FU was measured by MTT assay. The expression of SIRT1 and NADPH oxidase, activation of caspase-9 and caspase-3, and phosphorylation of ASK1 and JNK in the BGC-823 cells treated with DHA and 5-FU were determined by Western blot. The production of ROS and the apoptosis of the BGC-823 cells treated with DHA and 5-FU were analyzed by flow cytometry. RESULTS: Dihydroartemisinin significantly inhibited the expression of SIRT1 and increased NADPH oxidase protein level (P<0.05). DHA increased the sensitivity of BGC-823 cells to 5-FU, thus decreasing the IC₅₀ of 5-FU to the gastric cancer cells. However, transfection with SIRT1 plasmid decreased the cytotoxicity of DHA and 5-FU co-treatment to the BGC-823 cells. DHA promoted the production of ROS and phosphorylation of ASK1 and JNK induced by 5-FU in the BGC-823 cells (P<0.05). However, ROS scavenger N-acetylcysteine (NAC) or JNK specific inhibitor SP600125 inhibited the cell death and activation of caspase-9 and caspase-3 induced by DHA and 5-FU co-treatment (P<0.05). In addition, NAC significantly inhibited the phosphorylation of JNK in the BGC-823 cells co-treated with DHA and 5-FU. However, treatment with SP600125 did not influence the ROS production in the BGC-823 cells, indicating that JNK was the downstream target of ROS pathway. CONCLUSION: Combination of DHA with 5-FU induces caspase-dependent apoptosis in gastric cancer cells through the SIRT1/NADPH oxidase/ROS/JNK signaling pathway.     

Quercetin reduces microvascular permeability induced by ischemia/reperfusion injury through SIRT1 signaling

AIM To investigate whether quercetin (Que) attenuates microvascular injury after myocardial ischemia/reperfusion (I/R), and whether its mechanism is related to the up-regulation of silent information regulator 1 (SIRT1). METHODS In vivo, SD rats (220~250 g) were randomly divided into sham group (only threading without ligation), I/R group (ischemia 30 min/reperfusion 60 min), I/R+Que group (10 mg/kg Que injected intravenously 15 min before reperfusion) and I/R+Que+EX527 group (1 mg/kg EX527 injected intravenously 15 min before Que). The protein expression of SIRT1 in each group was determined by Western blot. The effect of Que on the morphological changes of myocardial microvascular was estimated by HE staining. The contents of endothelial damage markers and inflammatory factors in peripheral blood serum of the rats in each group were measured by ELISA. In vitro, Transwell chamber was used to culture H5V cells and then the influence of Que on the permeability of endothelial cells was evaluated. The expression of myosin light chain phosphatase(MLCP), zonula occludens-1 (ZO-1) and cytoskeleton was determined by Western blot and immunofluorescence method. RESULTS The results of Western blot showed that SIRT1 was significantly reduced by I/R in the myocardial tissue. Que increased the expression of SIRT1 after I/R, whereas EX527 significantly inhibited this effect (P<0.05). Que reduced the incidence of I/R arrhythmia, alleviated microvascular damage and inhibited the infiltrating of inflammatory cells,as well as decreased levels of the endothelial injury markers (E-selectin and VCAM-1) and proinflammatory factors (PAF, IL-1α and IL-6). While EX527 significantly reduced these effects of Que by inhibiting SIRT1. Cultured H5V cells showed that hypoxia/reoxygenation resulted in increased permeability of endothelial cells and decreased expression of ZO-1 and MLCP. However, Que, which up-regulated expression of SIRT1, significantly reduced the permeability of endothelial cells significantly and increased the expression of ZO-1 and MLCP. Meanwhile, the cytoskeleton remodeling basically disappeared. EX527, which down-regulated expression of SIRT1, significantly inhibited the above effects of Que. CONCLUSION Que attenuates myocardial I/R induced microvascular injury. The up-regulation of SIRT1 is involved in this mechanism.     

miR-34a induces senescence of bone marrow-derived mesenchymal stem cells under high glucose condition by SIRT1

AIM:To detect the effect and potential mechanism of microRNA-34a (miR-34a) on the senescence of bone marrow-derived mesenchymal stem cells (BMSCs) under high glucose condition. METHODS:BMSCs were isolated and cultured from 60~80 g male SD rats. The BMSCs were divided into 5 groups:normal glucose(NG) group, high glucose(HG) group, HG+miR-34a mimic group, HG+miR-34a NC group and HG+miR-34a inhibitor group. In order to confirm whether miR-34a regulated the senescence of BMSCs under high glucose condition by regulating the expression of silent information regulator 1(SIRT1), in addition to the above groups, HG+siRNA-SIRT1 group, HG+siRNA-NT group and HG+miR-34a inhibitor+siRNA-SIRT1 group were added. The expression of miR-34a and SIRT1 mRNA was detected by RT-qPCR. CCK-8 assay and senescence-associated β-galactosidase assay were used to detect cell viability and senescence, respectively. The protein expression of SIRT1, forkhead box O3a (FOXO3a) and P21 in the BMSCs was analyzed by Western blot. RESULTS:The expression of miR-34a in HG group was increased significantly compared with NG group (P<0.01), and long-term exposure of the BMSCs to high glucose lead to decreased cell viability and increased senescence (P<0.05). Compared with HG+miR-34a NC group, the cell viability in HG+miR-34a mimic group was decreased significantly (P<0.01), the senescence of BMSCs was increased significantly (P<0.01), the protein expression of SIRT1 was decreased significantly (P<0.01) and the protein expression of FOXO3a was increased significantly (P<0.01). However, inhibition of miR-34a expression showed the opposite effect to miR-34a mimic. Similar to the HG+miR-34a mimic group, the protein expression of P21 and FOXO3a in HG+siRNA-SIRT1 group were significantly higher than that in HG group (P<0.01). After adding siRNA-SIRT1 into HG+miR-34a inhibitor group, the inhibitory effect of the miR-34a inhibitor on the expression of P21 and FOXO3a in BMSCs were partly weakened (P<0.05). CONCLUSION:miR-34a regulate the senescence of BMSCs under high glucose condition by regulating the expression of SIRT1.     

Influence of Tangshenfang on diabetic liver injury and the expression of SIRT1 and PGC-1α in the liver tissue

AIM: To observe the effect of Tangshenfang (TS) on the liver protection and the levels of silent information regulator 1 (SIRT1) and peroxisom proliferator-activated receptor γ coactivator-1α (PGC-1α) in the liver tissue. METHODS: The rat model of diabetes mellitus (DM) was established by intravenous injection of streptozotocin (STZ;30 mg/kg) after having the high fat/high glucose diets for 1 month. The diabetic rats were randomly divided into DM group, DM with high-dose TS (TS^Hi) group, medium-dose TS (TS^Med) group and low-dose TS (TS^Low)group. The normal rats were served as control group. There were 8 rats in each group. After treatment with TS for 12 weeks, the serum biochemical indices including fasting blood glucose (FBG), triglyceride (TG), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were tested. Fasting insulin (FINS) was also detected by radioimmunoassay, and homeostatic model assessment for insulin resistance (HOMA-IR) was calculated. The serum levels of tumor necrosis factor-α (TNF-α) and interleukin-1 (IL-1) were measured by ELISA. The activity of SOD and content of MDA in the liver tissues were measured by the methods of hydroxylamine and thiobarbituric acid. The liver pathological changes were observed under light microscope with HE and Masson staining. The protein expression of SIRT1and PGC-1α in the liver tissues was determined by Western blot. RESULTS: In DM group, serum FBG, TG, ALT, AST, FINS, HOMA-IR, TNF-α and IL-1 were obviously increased compared with the control group (P<0.01). The fatty changes, local necrosis, inflammation and fibrosis in the liver tissues were observed. The content of MDA in liver increased, while the activity of SOD decreased markedly. The protein expression of SIRT1 and PGC-1α was decreased (P<0.05). In TS treatment groups, all these changes in DM rats were markedly reversed by TS, and the protein expression of SIRT1 and PGC-1α in the liver tissues was markedly increased. CONCLUSION: TS may protect the rats from diabetic liver injury by increasing the expression of SIRT1 and PGC-1α, and thereby improving insulin resistance and oxidative stress.     

Baicalin inhibits high glucose-induced apoptosis of mouse glomerular mesangial cells via miR-141/Sirt1 signaling pathway

AIM:To investigate the therapeutic mechanism of baicalin for diabetic nephropathy involving microRNA-141 (miR-141)/silent information regulator 1 (Sirt1) signaling pathway. METHODS:Mouse glomerular mesan-gial cell line SV40-MES-13 was treated with high glucose (HG, 25 mmol/L glucose) to establish diabetic nephropathy cell model. Baicalin at 100 μmol/L was used to treat glomerular mesangial cells. qPCR and Western blot were performed to determine the expression levels of miR-141 and Sirt1. The regulatory relationship between miR-141 and Sirt1 was detected by dual-luciferase assay. The apoptosis of glomerular mesangial cells was analyzed by flow cytometry. RESULTS:Compared with control group, the cells treated with HG showed increased levels of miR-141 and apoptosis, and Sirt1 expression was decreased (P<0.01). Baicalin and miR-141 inhibitor suppressed the HG-induced effect on the levels of miR-141, Sirt1 and apoptosis. Knockdown of Sirt1 expression reversed the effect of miR-141 inhibitor on the levels of miR-141, Sirt1 and apoptosis. Over-expression of miR-141 reversed the effect of baicalin on the glomerular mesangial cells treated with HG. Up-regulation of Sirt1 abolished the effect of miR-141 over-expression on the glomerular mesangial cells. CONCLUSION:Baicalin inhibits the apoptosis of mouse glomerular mesangial cells via miR-141/Sirt1 signaling pathway, thus attenuating diabetic nephropathy.     

Role of thioredoxin-ASK1 in doxorubicin-induced apoptosis of neonatal rat cardiac myocytes

AIM: To investigate the role of thioredoxin(Trx)-apoptosis signal-regulating kinase 1(ASK1) in doxorubicin-induced apoptosis of neonatal rat cardiac myocytes (NRCMs). METHODS: Primary cardiomyocytes were isolated from newborn Sprague-Dawley rats with the purity of NRCMs >95%. NRCMs were pretreated with the indicated concentrations of ebselen 2 h prior to the addition of doxorubicin, then treated with doxorubicin at concentration of 1 μmol/L for another 24 h. The viability of the cells was examined by MTT assay.Reactive oxygen species(ROS) levels were measured by a ROS-specific probe 2',7'-dichlorodihydrofluorescein diacetate (H₂DCFDA). Apoptotic cardiomyocytes were determined by Hoechst 33258 nuclear staining. The activity of caspase-3 was detected with a caspase-3 colorimetric assay kit. The protein levels of poly(ADP-ribose) polymerase 1(PARP1), ASK1, p-ASK1, p38 and p-p38 were determined by Western blotting. Immunoprecipitation and immunoblotting were performed to detect whether the Trx-ASK1 was dissociated. RESULTS: Doxorubicin induced significant apoptosis of NRCMs. The levels of ROS were significantly increased. Ebselen significantly decreased the apoptosis. Compared with control group, increased activity of caspase-3 was showed in doxorubicin group (P<0.01). Increased protein levels of PARP1, ASK1 and p38 were observed (P<0.01). The increase in the dissociated Trx-ASK1 was also found. Compared with doxorubicin group, ebselen decreased the activity of caspase-3 (P<0.01), the levels of PARP1,ASK1 and p38 proteins (P<0.05), and the dissociated Trx-ASK1. CONCLUSION: Doxorubicin induces significant apoptosis of NRCMs. ASK1 is partly dissociated from Trx, and starts the ASK1-mediated apoptotic signaling. The process is significantly attenuated by pretreatment with ebselen. Trx-ASK1 plays an important role in doxorubicin-induced apoptosis of cardiomyocytes.     

Over-expression of SIRT1 gene inhibits high glucose-stimulated H9c2 cardiomyocyte apoptosis and ROS level through PI3K/AKT signaling pathway

AIM: To investigate the effects of silent information regulator 1 (SIRT1) over-expression on the apoptosis and the level of reactive oxygen species (ROS) in high glucose-induced H9c2 cardiomyocytes. METHODS: H9c2 cardiomyocytes were transfected with empty plasmid (pcDNA3.1-NC) and SIRT1 over-expression plasmid (pcDNA3.1-SIRT1), and then stimulated by high glucose. The H9c2 cells were divided into control group, high glucose group, high glucose + pcDNA3.1-NC group and high glucose + pcDNA3.1-SIRT1 group. The expression of SIRT1 at mRNA and protein levels in each group was determined by qPCR and Western blot. The viability of the cells was measured by MTT assay. The apoptotic rate was analyzed by flow cytometry. The protein levels of phosphatidylinositol 3-kinase (PI3K), phosphorylated PI3K, AKT and phosphorylated AKT were examined by Western blot. RESULTS: SIRT1 was significantly decreased in high glucose-induced H9c2 cardiomyocytes, the cell viability was significantly decreased compared with control group, while the ROS levels and apoptotic rate were increased, and the phosphorylated PI3K and AKT protein levels were down-regulated (P<0.05). Over-expression of SIRT1 significantly promoted the viability of H9c2 cardiomyocytes induced by high glucose, decreased the ROS levels and apoptotic rate, and up-regulated phosphorylated PI3K and AKT protein levels (P<0.05). CONCLUSION: SIRT1 over-expression reverses the decrease in the viability of high glucose-stimulated H9c2 cardiomyocytes, and the increases in apoptotic rate and oxidative stress by regulating PI3K/AKT signaling pathway.     

miR-193b enhances cytotoxicity of doxorubicin by targeting Mcl-1 in breast cancer

AIM: To investigate the effect of microRNA(miR)-193b on doxorubicin therapy in breast cancer in vitro.METHODS: miR-193b level in plasma was detected by real-time PCR in the patients with breast cancer or the healthy controls. MTT assay was performed to measure the inhibitory effect of miR-193b plus doxorubicin on the growth of MDA-MB-231 cells. Bioinformatics, real-time PCR and Western blot were performed to determine whether the expression of Mcl-1 was regulated by miR-193b. Mcl-1 expression vector was constructed, and the role of Mcl-1 vector toward miR-193b plus doxorubicin-induced cytotoxicity in MDA-MB-231 cells was observed by MTT assay.RESULTS: Down-regulation of miR-193b was found in breast cancer patients. The miR-193b plus doxorubicin group showed a higher growth inhibition than cisplation group in MDA-MB-231 cells. The expression of Mcl-1 at both mRNA and protein levels was down-regulated after miR-193b transfection. The growth inhibition of MDA-MB-231 cells treated with miR-193b plus doxorubicin was significantly decreased after the transfection of Mcl-1 expression vector.CONCLUSION: miR-193b sensitizes doxorubicin-induced cytotoxicity by targeting Mcl-1 in breast cancer.     

Value of endonuclease domain containing 1 in progression of prostate cancer

AIM: To analyze the difference of endonuclease domain containing 1 (ENDOD1) expression between benign prostatic hyperplasia (BPH) tissues and prostate cancer (PCa) tissues and to investigate the effect of ENDOD1 on the biological function of human prostate cancer cells. METHODS: The BPH samples (n=20) and PCa samples (n=21) were processed and analyzed according to the instruction of immunohistochemical (IHC) staining. The mRNA and protein levels of ENDOD1 in the normal prostate epithelial cells and prostate cancer cells were evaluated by RT-qPCR and Western blot, respectively. The recombinant plasmids pCMV-N-Flag-ENDOD1 was constructed and was transfected into the human prostate cancer cells. The proliferation, apoptosis, migration and invasion abilities of the prostate cancer cells were evaluated by MTT assay, flow cytometry, Transwell migration and Matrigel invasion assays, respectively. RESULTS: The analysis of variance of the immunoreactivity score showed that PCa tissues with high Gleason score displayed significantly lower ENDOD1expression than that with low Gleason score and BPH (P<0.05). The expression of ENDOD1 at mRNA and protein levels in PC3 cells and DU145 cells was significantly lower than that in the LNCap cells (P<0.05). The proliferation of DU145 transfected with ENDOD1 was inhibited. The flow cytometry indicated that ENDOD1 over-expression in the DU145 cells resulted in a notable increase in G₀/G₁ phase arrest (P<0.05), but the apoptotic rates showed no statistical difference. The results of Transwell assay showed that migration and invasion abilities of the cells were also inhibited after transfection with over-expressing ENDOD1 plasmid (P<0.05). CONCLUSION: The expression of ENDOD1 significantly decreased in prostate cancer with high Gleaon score. Meanwhile, the ENDOD1 is specifically down-regulated in androgen independent prostate cancer (AIPC) cell lines. Over-expression of ENDOD1 remarkably inhibits the proliferation, migration and invasion abilities of AIPC.     

Fucoidan sulfate promotes apoptosis of gastric cancer cells by regulating expression of HDAC1

AIM:To study the effects of fucoidan sulfate on the apoptosis of gastric cancer cells, and to explore its relationship with histone deacetylase 1 (HDAC1). METHODS:The effects of fucoidan sulfate on the viability of gastric cancer BGC-823 cells were measured by MTT assay. The cells were divided into si-HDAC1 group, si-NC group and fucoidan+pcDNA 3.1-HDAC1 group and fucoidan+pcDNA 3.1 group. The transfection were performed by liposome method. The apoptosis was analyzed by flow cytometry. The protein expression of HDAC1, Bcl-2 and Bax was determined by Western blot. The mRNA expression of HDAC1 was detected by RT-qPCR. RESULTS:Compared with the control cells, the inhibitory rates of the cell viability by fucoidan sulfate (0.2, 0.4, 0.6, 0.8 and 1.0 g/L) were increased signi-ficantly (P<0.05). The appropriate concentration was 0.6 g/L. Compared with control group, the expression of HDAC1 at mRNA and protein levels in fucoidan sulfate group was significantly down-regulated, while the apoptotic rate was significantly increased (P<0.05). Knock-down of HDAC1 expression significantly up-regulated the apoptotic rate, while over-expression of HDAC1 reversed the apoptosis-promoting effect of fucoidan sulfate on the BGC-823 cells. CONCLUSION:Fucoidan sulfate promotes apoptosis of gastric cancer cells. The mechanism may be related to the direct inhibition of HDAC1, which provides support for the clinical application of fucoidan sulfate.     

Berberine promotes anti-proliferative effects of doxorubicin in T24 bladder cancer cells by enhancing apoptosis

AIM: To observe the effect of berberine (Ber) on doxorubicin (DOX)-induced apoptosis in bladder cancer T24 cells. METHODS: The cells were exposed to DOX in the presence or absence of different concentrations of Ber. The viability of the cells was determined by CCK-8 assay. The apoptosis was measured by Hoechst 33258 staining and the protein levels of cleaved caspase-3, cleaved caspase-9, Bcl-2 and Bax were detected by Western blotting.RESULTS: Ber enhanced the inhibitory effect of DOX on the viability of T24 cells and promoted DOX-induced apoptosis in T24 cells. DOX increased the protein levels of cleaved caspase-3, cleaved caspase-9 and Bax, all of which were enhanced by treatment with Ber. In contrast, Ber exposure further decreased the expression of Bcl-2 in DOX-treated T24 cells.CONCLUSION: Ber enhances the anti-proliferative effects of DOX through promoting apoptosis in bladder cancer cells.     

Effect of abnormal expression of PDK4 on glycolysis and proliferation in prostate cancer cells

AIM To investigate the expression of pyruvate dehydrogenase kinase 4 （PDK4） in prostate cancer tissue and its effect on glycolysis and growth of prostate cancer cells. METHODS Immunohistochemistry was used to compare the expression differences of PDK4 protein in benign prostatic hyperplasia （BPH） and prostate cancer tissues. The expression levels of PDK4 in normal prostatic epithelial cells （RWPE-1） and different prostate cancer cell lines （PC3, LNCaP, DU145 and C4-2） were detected by RT-qPCR and Western blot. Recombinant plasmid carrying PDK4-shRNA was constructed, and the expression of PDK4 in prostate cancer PC3 cells was down-regulated by transfection with PDK4-shRNA. The changes in glycolysis level of PC3 cells before and after transfection were determined by cell glycolysis kit, and the effects of PDK4 on the viability and cell cycle distribution of PC3 cells were detected by CCK-8 assay and flow cytometry. RESULTS In prostate cancer tissues, the expression level of PDK4 protein was significantly higher than that in BPH tissues （P<0.05）, and the analysis of immunohistochemical score showed that prostate cancer tissues with high Gleason score displayed significantly higher PDK4 expression than those with low Gleason score （P<0.05）. Compared with normal prostatic epithelial cells, RT-qPCR and Western blot results indicated that the expression level of PDK4 was also significantly increased in prostate cancer cell lines （P<0.05）. In addition, CCK-8 assay results showed that the viability of prostate cancer PC3 cells was significantly decreased after knockdown of PDK4 expression （P<0.05）. The results of flow cytometry demonstrated that knockdown of PDK4 expression in PC3 cells resulted in a notable increase in G₀/G₁ phase arrest （P<0.05）. CONCLUSION PDK4 is highly expressed in prostate cancer tissues and cell lines, and significantly increases in prostate cancer with high Gleason score. In addition, down-regulation of PDK4 expression significantly inhibits glycolysis and growth of prostate cancer cells, resulting in cell cycle arrest at G₀/G₁ phase.     

SIRT1 promotes autophagy of pancreatic cancer cells induced by hypoxia via regulating FOXO1/RAB7 signaling pathway

AIM: To investigate the effect of SIRT1 on the autophagy of pancreatic cancer cells under hypoxia condition, and to analyze the underlying mechanism of regulating FOXO1/RAB7 signaling pathway. METHODS: Western blot and immunofluorescence methods were used to determine the expression of SIRT1 in the pancreatic cancer cells. The small interfering RNA targeting SIRT1 and SIRT1 over-expression plasmid were transfected into the pancreatic cancer Panc-1 cells. Confocal microscopy was used to detect the LC3 expression. Western blot was used to analyze the protein levels of LC3, p62 and FOXO1/RAB7 signaling pathway-related molecules. Co-immunoprecipitation was used to detected the protein interaction between SIRT1 and FOXO1. RESULTS: The expression level of SIRT1 in the nucleus of Panc-1 cells was increased under hypoxia condition. Compared with negative control under hypoxia condition, knock-down of SIRT1 expression attenuated the autophagy flux in the pancreatic cancer Panc-1 cells (P<0.05). Over-expression of SIRT1 increased the protein levels of FOXO1 and RAB7. On the contrary, knock-down of SIRT1 expression inhibited the protein levels of FOXO1 and RAB7. The protein interaction between SIRT1 and FOXO1 in the pancreatic cancer cells was observed. CONCLUSION: SIRT1 in pancreatic cancer Panc-1 cells under hypoxia condition is over-expressed in the nucleus. Down-regulation of SIRT1 inhibits autophagy and its mechanism may be related to FOXO1/RAB7 signaling pathway.     

Effects of phytosterol ester on aortic aging and expression of related genes in rats

AIM: To explore the protective effect of phytosterol ester (PSE) on aortic aging in rats. METHODS: The female SD rats (12 months old, n=42) were randomly divided into control group, model group and PSE group. During the experiment, the rats in control group, model group and PSE group were treated with basic feed, high-fat diet (HFD) and HFD with 2% PSE (W/W) for 6 months, respectively. The morphological changes of the aorta were observed by HE staining and Masson staining, and the absolute area of smooth muscle cells and collagen fiber in the vascular wall were measured by image analysis. The levels of advanced glycosylation end products (AGEs), malondialdehyde (MDA), superoxide dismutase (SOD) and catalase (CAT) in the plasma were detected. The expression of silent information regulator 1 (SIRT1) and peroxisome proliferator-activated receptor γ (PPARγ) at mRNA and protein levels in the vascular tissue was determined by real time PCR and Western blot, respectively. RESULTS: PSE significantly lowered plasma TC and LDL-C, and increased plasma HDL-C level (P<0.05), but had no effect on plasma TG level. PSE significantly attenuated the thickening of intima and media of aging aortic, and decreased the migration of vascular smooth muscle cells (VSMC) and the amount of VSMC and collagen fiber in the aorta (P<0.05). PSE significantly reduced the contents of AGEs and MDA (P<0.05), but had no effect on the activity of SOD and CAT in the plasma. PSE also down-regulated the expression of PPARγ and up-regulated the expression of SIRT1 (P<0.05). CONCLUSION: PSE is able to attenuate the senescence process in the aorta by reducing the production of reactive oxygen species in plasma, and activating SIRT1, or inhibiting the expression of PPARγ in vascular tissues.     

Plumbagin enhances TRAIL-induced apoptosis in Kasumi-1 cells

AIM: To investigate the effect of plumbagin and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) on the apoptosis of leukemic Kasumi-1 cells. METHODS: Kasumi-1 cells were treated with plumbagin alone, recombinant soluble TRAIL(rsTRAIL) alone or the combination of plumbagin with rsTRAIL to induce apoptosis. The cell proliferation was analyzed by CCK-8 assay. Apoptosis was determined by flow cytometry with AnnexinⅤ/PI double staining and TUNEL staining. The expression of DR4 and DR5 at mRNA level was measured by real-time PCR. The expression of signal transduction proteins, such as DR5, caspase-3, caspase-8, caspasep-9, Bid, Bax and c-FLIP was detected by Western blotting. RESULTS: Both rsTRAIL and plumbagin induced the apoptosis in Kasumi-1 cells, and combination of plumbagin with rsTRAIL enhanced the apoptosis. The ratios of Annexin V-positive Kasumi-1 cells were (27.7±2.9)%, (25.6±3.1)% and (52.1±3.3)% in 100 μg/L rsTRAIL group, 2 μmol/L plumbagin group and the combination group, respectively, and the positive rate in combination group was significantly higher than those in other 2 groups. TUNEL assay demonstrated that the number of apoptotic cells in combination group was higher than that in the cells treated with rsTRAIL or plumbagin alone. Plumbagin up-regulated the expression of DR5 at mRNA level in Kasumi-1 cells, and up-regulation of DR5, activation of caspase-8 and down-regulation of c-FLIP at protein level were detected in the cells treated with plumbagin alone and the combination of plumbagin with rsTRAIL. CONCLUSION: Plumbagin enhances TRAIL-induced apoptosis in Kasumi-1 cells by up-regulating DR5, activating caspase-8 and down-regulating c-FLIP.     

Relationship between phosphoglycerate kinase 1 and clinical features of prostate cancer and its role in prognosis

AIM: To study the expression and prognostic functions of phosphoglycerate kinase 1 (PGK1) in prostate cancer. METHODS: The prostatic samples were collected from the patients with prostate cancer and benign prostatic hyperplasia (BPH) in TCM-Integrated Hospital of Southern Medical University from Jan 2013 to Dec 2013. The protein expression of PGK1 in the prostate specimens was detected by immunohistochemical analysis and Western blot. Furthermore, the correlations of PGK1 expression with the clinicopathological features and prognosis of prostate cancer were also evaluate. RESULTS: The expression of PGK1 in the prostate specimens was significantly up-regulated compared with the BPH individuals. In addition, the expression of PGK1 was significantly correlated with the local infiltration, Gleason score, TNM grade, bone metastasis, and serum prostate-specific antigen (PSA) concentration. Finally, bone metastasis, serum PSA level and PGK1 expression were independent risk factors for prostate cancer illustrated by Cox analysis, and high expression of PGK1 was correlated with poor prognosis. CONCLUSION: PGK1 expression is an independent risk factor for prostate cancer, and it might act as a prognostic biomarker for prostate cancer.