This study was aimed to explore the effects of oleanolic acid on PTEN expression and apoptosis of Jurkat cells. The inhibitory rate was measured by Cell Counting Kit-8. The apoptotic nucleus morphous was observed by Hoechst 33258 staining. The apoptosis rate of Jurkat cells were determined by flow cytometry with Annexin V/PI double staining. PTEN mRNA and protein were detected by quantitative real-time PCR and Western blot respectively. The results showed that oleanolic acid inhibited the proliferation of Jurkat cells in time- and dose-dependent manners. The 50% growth inhibition (IC(50)) at 12, 24 and 48 hours were about 85.35 µmol/L, 53.66 µmol/L and 33.18 µmol/L respectively. Flow cytometric assay showed that the apoptotic rates of Jurkat cells treated with oleanolic acid (0, 40, 80 and 160 µmol/L) for 24 hours were 6.72%, 19.8%, 28.72% and 30.12% (p < 0.05). PTEN mRNA and protein expressions were up-regulated in Jurkat cells treated with oleanolic acid of concentration 80 µmol/L and 160 µmol/L for 24 hours. It is concluded that up-regulation of PTEN mRNA and PTEN protein may be involved in oleanolic acid-induced Jurkat cell apoptosis.
Apoptosis ; drug effects ; Cell Proliferation ; Humans ; Jurkat Cells ; Oleanolic Acid ; pharmacology ; PTEN Phosphohydrolase ; metabolism ; Up-Regulation

This study explored the effects of resveratrol(Res) on the expression of phosphatase and tensin homolog deleted on chromosome ten(PTEN) and the fibrosis of rat renal tubular epithelial cells in a high-glucose environment and the possible mechanism underlying the fibrosis reduction. After the pretreatment of rat renal tubular epithelial cells(NRK-52 E) cultured in a high-glucose condition with Res or PTEN inhibitor SF1670, they were divided into several groups, i.e., normal glucose(NG), normal glucose + SF1670(NS), high glucose(HG), high glucose + SF1670(HS), high glucose + Res at different concentrations(5, 10, 25 μmol·L~(-1)). The expression and distribution of E-cadherin and α-SMA in renal tubular epithelial cells were observed by immunofluorescence cytochemistry. The protein expression levels of PTEN, E-cadherin, α-SMA, p-Akt~((Thr308)) and collagen Ⅳ were determined by Western blot. Real-time PCR was employed to detect the expression of PTEN mRNA. Compared with the NG group, the HG group witnessed the reduced expression of PTEN mRNA, PTEN protein and E-cadherin protein, but saw the increased expression of α-SMA, p-Akt~((Thr308)) and collagen Ⅳ proteins. Besides, with the increase in Res concentration, the expression levels of PTEN mRNA, PTEN protein and E-cadherin protein gradually increased, while those of α-SMA, collagen Ⅳ, p-Akt~((Thr308)) proteins gradually decreased in the Res groups, showing a dose-effect dependence, compared with the HG group. No distinct difference was found between the NS group and the NG group. The expression level of E-cadherin was even lower and those of α-SMA, p-Akt~((Thr308)), and collagen Ⅳ were higher in the HS group than in the HG group, with no marked difference shown in the two groups in terms of PTEN mRNA and protein. Although the PTEN inhibitor did not affect PTEN, the expression changes of the other proteins were opposite to the results after Res treatment and the fibrosis was aggravated, which suggested that SF1670 promoted the fibrosis by inhibiting PTEN, activating Akt and increasing the synthesis of collagen Ⅳ and other extracellular matrix. The results show that Res can antagonize the high glucose-mediated fibrosis of renal tubular epithelial cells. This may be achieved via the up-regulation of PTEN and the inhibition of PI3 K/Akt signaling pathway.
Animals ; Epithelial Cells ; Fibrosis ; Glucose ; PTEN Phosphohydrolase/genetics* ; Rats ; Resveratrol/pharmacology*

This study inquired about the role of tumor suppressor PTEN in the arterial remodeling of Ang II induced hypertension. The expression of PTEN of aorta was examined in the aortic-constricted hypertensive rats (hypertension group), in the aortic-constricted hypertensive rats treated with captopril(hypertension and captopril group), and in the rats having undergone sham operation (control group). At day 28 after surgery, the aortas were collected from the groups. The expression of PTEN mRNA was detected by RT-PCR. The expression and location of PTEN protein were determined by immunohistochemistry. The results showed that the expression of PTEN in aorta of the hypertension group was significantly lower than that of the hypertension and captopril group, and similarly lower than that of the control group. The intensity of PTEN-positive immunohistochemical production in aorta of the hypertension group was weaker than that of the hypertension and captopril group, and likewise, it was weaker than the control. PTEN-positive immunohistochemical production was located in VSMC of aorta. The findings indicated that the expression of PTEN is reduced in hypertensive aorta, that the reduced PTEN experession can be reversed by captopril treatment, that AngII and the increased mechanical strain may participate in regulating expression of PTEN, and that PTEN may play a role in the arterial remodeling induced by hypertension.
Angiotensin-Converting Enzyme Inhibitors ; pharmacology ; Animals ; Aorta, Abdominal ; metabolism ; Captopril ; pharmacology ; Constriction ; Genes, Tumor Suppressor ; Hypertension ; etiology ; metabolism ; Male ; PTEN Phosphohydrolase ; biosynthesis ; genetics ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate whether PTEN can increase sensitivity of Ishikawa cells, an endometrial carcinoma cell line, to doxorubicin.

METHODSIshikawa cells transfected by PTEN gene or not were separately treated with serial concentrations of doxorubicin. The sensitivity of cells to doxorubicin was determined by MTT assay. The cells were stained with Hoechst 33258 and examined under fluorescence microscope to determine cell apoptosis. Immunoprecipitation and Western blotting analysis were performed to evaluate the effects of doxorubicin on phosphorylation of Bad and Akt/PKB.

RESULTSDoxorubicin induced cell death of the PTEN-transfected and non-transfected Ishikawa cells in a dose-dependent manner, but the cell death was more significant in PTEN-expressing clones than in parental Ishikawa cells. A low concentration of doxorubicin (0.1 micromol/L) did not affect cell apoptosis in PTEN-null Ishikawa cells, but it induced cell apoptosis in PTEN-expressing clones. A high concentration of doxorubicin (1 micromol/L) induced cell apoptosis in both cell lines. However, the percentage of apoptotic cells was higher in PTEN-expressing clones than that in parental Ishikawa cells. In the PTEN-expressing clones, expression of phospho-Akt/PKB and phospho-Bad (Ser-136) was down regulated. Doxorubicin reduced the levels of phospho-Akt/PKB and phospho-Bad (Ser-136) in both cell lines, but the most significant reduction occurred in the PTEN-expressing clones.

CONCLUSIONPTEN significantly enhances chemosensitivity of Ishikawa cells to doxorubicin. With PTEN expression, doxorubicin may exert apoptosis-induction activity by downregulation of the PI3k/Akt/PKB signaling pathway in Ishikawa cells.

Adenocarcinoma ; genetics ; pathology ; Antibiotics, Antineoplastic ; pharmacology ; Apoptosis ; drug effects ; Cell Line, Tumor ; Doxorubicin ; pharmacology ; Endometrial Neoplasms ; genetics ; pathology ; Female ; Humans ; PTEN Phosphohydrolase ; biosynthesis ; genetics ; Transfection

Azacitidine ; administration & dosage ; analogs & derivatives ; pharmacology ; Cell Proliferation ; drug effects ; Daunorubicin ; administration & dosage ; pharmacology ; HL-60 Cells ; Humans ; PTEN Phosphohydrolase ; metabolism

AIMTo investigate the role of tumor suppressor PTEN in cardiac hypertrophy, the expression of PTEN mRNA in left ventricle of abdominal aorta constricted-induced cardiac hypertrophic rats which treated with and without captopril was analyzed.

METHODSSD rats were divided into control group, hypertrophy group and captopril group. The expression of PTEN mRNA in left ventricle was detected by RT-PCR in different groups in 4 weeks after operation.

RESULTS(1) Compared with control group, the expression of PTEN mRNA in left ventricle of hypertrophy group was reduced. (2) Compared with hypertrophy group, the expression of PTEN mRNA in left ventricle of captopril group was upregulated, which were similar to that of control group.

CONCLUSIONPTEN maybe plays a negative regulation role in the process of cardiac hypertrophy, and the role of PTEN is closely relative with renin-angiotensin system.

Angiotensin-Converting Enzyme Inhibitors ; pharmacology ; Animals ; Captopril ; pharmacology ; Cardiomegaly ; metabolism ; pathology ; Male ; PTEN Phosphohydrolase ; metabolism ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley

The study was aimed to investigate the inducing effect of ursolic acid (UA) on the apoptosis of human T-cell leukemia/lymphoma (Jurkat), and whether the regulation of PTEN involved in the effect of UA on Jurkat cells. The Jurkat cells were treated with different concentrations of UA for different time. The cell proliferation was analyzed with cytotoxicity test (CCK8 method). Cell apoptosis was detected by fluorescence microscopy and flow cytometry. The expression of PTEN mRNA was detected by real-time quantitative PCR. The results indicated that UA could significantly inhibited the viability of Jurkat cells treated with 10-80 µmol/L and in dose- and time-dependent manner. UA could induce Jurkat cell apoptosis in a dose-dependent manner, which was statistical different from the control at the same time (P < 0.05). PTEN mRNA expression was up-regulated by UA, which was statistical different from the control (P < 0.05). It is concluded that UA may induce Jurkat cell apoptosis by up-regulating the PTEN mRNA expression.
Apoptosis ; drug effects ; Cell Proliferation ; Dose-Response Relationship, Drug ; Humans ; Jurkat Cells ; PTEN Phosphohydrolase ; genetics ; metabolism ; RNA, Messenger ; genetics ; Triterpenes ; pharmacology ; Up-Regulation

OBJECTIVETo study the effect of tumor suppressor gene PTEN on proliferation and cell cycle of hepatocellular carcinoma cell line HHCC.

METHODSFirstly, eukaryotic expression vectors of wild type and mutated type of PTEN gene were constructed, named as pEGFP-WT-PTEN and pEGFP-PTEN; G129R, respectively. Lipofectamine 2000 was used to transfect the constructed expression vectors into hepatocellular carcinoma cell line HHCC which was PTEN protein negative. G418 was used to select the cell clones constantly expressing PTEN protein. Flow cytometry was used to assay the cell cycle of HHCC transfected by above mentioned eukaryotic expression vectors and non-transfected cell line HHCC. Intrinsic 473-phosphorylated AKT representing the level of active AKT was assayed by Western blot. The non-transfected HHCC served as control.

RESULTSThe proliferation of HHCC constantly expressing PTEN protein was obviously inhibited compared with HHCC cells transfected with mutated PTEN gene and empty vectors, and non-transfected HHCC cells. The number of HHCC cells transfected with wild type PTEN gene at G(1) phase, G(2) phase and S phase was 70.8%, 6.8% and 22.4%, respectively. Compared with control group transfected with empty vector, the number of G(1) phase HHCC cells constantly expressing wild type-PTEN protein was significantly higher than that of control. The number of cells in G(2) and S phase was significantly lower than that of control. However, the number of cells in G(1) phase, G(2) phase and S phase of HHCC transfected with mutant PTEN was 63.2%, 10.1% and 26.7%, respectively. There was no significant difference compared with control group. Western blot result showed that the intrinsic level of 473-phosphorylated AKT of HHCC constantly expressing wild type PTEN protein was down-regulated, and that of HHCC transfected with mutated PTEN gene was equal to that of control.

CONCLUSIONWild type PTEN gene can inhibit the proliferation of hepatocellular carcinoma cells at G(1) phase. The mechanism is possibly related with intrinsic activity of AKT, which is down-regulated by wild type PTEN.

Carcinoma, Hepatocellular ; pathology ; Cell Division ; drug effects ; Cell Line, Tumor ; Genes, Tumor Suppressor ; Humans ; Liver Neoplasms ; pathology ; PTEN Phosphohydrolase ; genetics ; pharmacology

OBJECTIVETo investigate the effect of hepatitis B virus X protein (HBx) on adriamycin-induced apoptosis of hepatocellular carcinoma cells and the expressions of p53 and PTEN.

METHODSHepG2, HepG2/GFP, and HepG2/GFP-HBx cells were treated with adriamycin (2.5 microg/ml), and the apoptotic cell death was determined by observing the morphological changes and flow cytometry. The expressions of p53 and PTEN mRNA in the 3 cells were detected by RT-PCR, and the expressions of p53 and PTEN protein were analyzed by Western blotting.

RESULTSAdriamycin induced significant cell death in HepG2 and HepG2/GFP cells, which became rounded, shrunk, and detached after the treatment; but no significant cell death occurred in HepG2/GFP-HBx cells. Flow cytometry analysis showed that the apoptotic rate was significantly lower in HepG2/GFP-HBx cells (3.94%) than in HepG2 (59.03%) and HepG2/GFP cells (61.38%) at 36 h after the treatment (P<0.001), while no significant difference was observed between HepG2/GFP-HBx (3.94%) and the control cells (2.12%, 2.78%, and 2.55%) (P>0.05). RT-PCR showed lowered expression of PTEN mRNA in HepG2/GFP-HBx cells as compared to that in HepG2 and HepG2/GFP cells, while no significant difference was noted in p53 mRNA. Western blot analysis showed that PTEN protein decreased while p53 protein remain unchanged in HepG2/GFP-HBx cells.

CONCLUSIONHBx suppresses adriamycin-induced apoptosis of HepG2 cells and PTEN expression. The inhibitory effect of HBx on the cell apoptosis may be related to the inhibition of p53-PTEN pathway.

Apoptosis ; drug effects ; Carcinoma, Hepatocellular ; metabolism ; pathology ; Doxorubicin ; pharmacology ; Hep G2 Cells ; Humans ; Liver Neoplasms ; metabolism ; pathology ; PTEN Phosphohydrolase ; metabolism ; Trans-Activators ; metabolism ; Tumor Suppressor Protein p53 ; metabolism

OBJECTIVE: To investigate the influence and mechanism of atorvastatin on glycolysis of adriamycin resistant acute promyelocytic leukemia (APL) cell line HL-60/ADM. METHODS: HL-60/ADM cells in logarithmic growth phase were treated with different concentrations of atorvastatin, then the cell proliferation activity was measured by CCK-8 assay, the apoptosis was detected by flow cytometry, the glycolytic activity was checked by glucose consumption test, and the protein expressions of PTEN, p-mTOR, PKM2, HK2, P-gp and MRP1 were detected by Western blot. After transfection of PTEN-siRNA into HL-60/ADM cells, the effects of low expression of PTEN on atorvastatin regulating the behaviors of apoptosis and glycolytic metabolism in HL-60/ADM cells were further detected. RESULTS: CCK-8 results showed that atorvastatin could inhibit the proliferation of HL-60/ADM cells in a concentration-dependent and time-dependent manner (r=0.872, r=0.936), and the proliferation activity was inhibited most significantly when treated with 10 μmol/L atorvastatin for 24 h, which was decreased to (32.3±2.18)%. Flow cytometry results showed that atorvastatin induced the apoptosis of HL-60/ADM cells in a concentration-dependent manner (r=0.796), and the apoptosis was induced most notably when treated with 10 μmol/L atorvastatin for 24 h, which reached to (48.78±2.95)%. The results of glucose consumption test showed that atorvastatin significantly inhibited the glycolytic activity of HL-60/ADM cells in a concentration-dependent and time-dependent manner (r=0.915, r=0.748), and this inhibition was most strikingly when treated with 10 μmol/L atorvastatin for 24 h, reducing the relative glucose consumption to (46.53±1.71)%. Western blot indicated that the expressions of p-mTOR, PKM2, HK2, P-gp and MRP1 protein were decreased in a concentration-dependent manner (r=0.737, r=0.695, r=0.829, r=0.781, r=0.632), while the expression of PTEN protein was increased in a concentration-dependent manner (r=0.531), when treated with different concentrations of atorvastatin for 24 h. After PTEN-siRNA transfected into HL-60/ADM cells, it showed that low expression of PTEN had weakened the promoting effect of atorvastatin on apoptosis and inhibitory effect on glycolysis and multidrug resistance. CONCLUSION Atorvastatin can inhibit the proliferation, glycolysis, and induce apoptosis of HL-60/ADM cells. It may be related to the mechanism of increasing the expression of PTEN, inhibiting mTOR activation, and decreasing the expressions of PKM2 and HK2, thus reverse drug resistance.
Humans ; Atorvastatin/pharmacology* ; PTEN Phosphohydrolase/pharmacology* ; Sincalide/metabolism* ; Drug Resistance, Neoplasm/genetics* ; TOR Serine-Threonine Kinases/metabolism* ; Leukemia, Promyelocytic, Acute/drug therapy* ; Doxorubicin/pharmacology* ; Apoptosis ; RNA, Small Interfering/pharmacology* ; Glycolysis ; Glucose/therapeutic use* ; Cell Proliferation