Effect of TMB-8 on the activation,proliferation and cell-cycle distribution of the mouse T lymphocytes in vitro

AIM: To study the effects of ［8-(diethylamino) octyl-3, 4, 5 -trimethoxybenzoate］ (TMB-8), an intracellular Ca2+ antagonist, on the activation, proliferation and cell-cycle distribution of the mouse T lymphocytes stimulated by concanvalin A (Con A) in vitro. METHODS: After stimulated with Con A, T cells were treated with different concentrations of TMB-8 alone and its combination with cyclosporine A (CsA). The expression of CD69, the early marker of CD3+ T cell activation, was measured by FACS. The proliferation-related index was determined by carboxyl fluorescin diacetate succinmidyl ester (CFDA-SE) flow cytometry. The cell-cycle distribution was analyzed by propidium iodide staining.RESULTS: After 6 h culture, the activation rate of CD69+ T cell in Con A group was (74.88±1.88)%. 10, 20 and 40 μmol/L of TMB-8 inhibited the expression of CD69 (P<0.01), especially in 40 μmol/L (52.55%±1.54%). After 48 h and 72 h culture, the PI of Con A group was 1.24±0.01, 2.05±0.07, respectively. TMB-8 with the concentration up to 5 μmol/L exerted a definite inhibitory effect on the proliferation with a maximal inhibition in 40 μmol/L(P<0.01). In the combination of 10 μmol/L of TMB-8 with 25 μg/L of CsA, an evident synergistic effect was observed (P<0.01). Moreover, the cell-cycle distribution analysis showed that after 48 h culture, the concentration of TMB-8 over 10 μmol/L showed an evident suppression in S phase.CONCLUSION: TMB-8 significantly inhibites the early steps of the Con A-induced T cell activation and proliferation, as well as the progression of T lymphocytes in S phase.     

Effect of isoflavone genistein on activation and proliferation of mouse T cells in vitro

AIM: To study the effect of genistein on activation and proliferation of T cells, and explore the molecular mechanism of genistein. METHODS: Fluorescence conjugated monoclonal antibodies and flow cytometry were used to detect the express of CD69 and CD25 by activated T cells in vitro in response to Concanavalin (ConA )and Phorbol 12, 13-dibutyrate(PDB) or T cell proliferation stained by CFSE in response to PDB / Ionomycin or ConA. RESULTS: Genistein inhibited the expression of CD69 and CD25 in activated T cells in response to Con A in a concentration-dependent manner and in response to PDB in a high concentration. Genistein inhibited proliferation of T cells in both groups in a concentration-dependent manner. CONCLUSION: Genistein inhibited activation and proliferation of T cells in vitro in response to polyclonal stimulus, and it may hold potential as a new immunosuppressant.     

Effect of proteasome inhibitor MG132 on proliferation and cell cycle distribution of NK/T cell lymphoma cells

AIM: To study the influence of MG132 on the proliferation and cell cycle distribution of NK/T cell lymphoma cells, and to investigate the potential role of proteasome inhibitor on the treatment of NK/T cell lymphoma. METHODS: NK/T cell lymphoma cells HANK1 were treated with proteasome inhibitor MG132, and the proliferation was evaluated by MTT assay. The morphological changes were observed under inverse microscope. The cell cycle distribution and apoptosis were detected using flow cytometry. RESULTS: The growth inhibitory rate of HANK1 cells was(57.72±7.44)% after cultured for 24 h with 1 μmol/L MG132 and was just(3.98±0.07)% after cultured for 24 h with 0.1 μmol/L MG132. The positive relationship between the concentration of MG132 and growth inhibitory rate was observed. On the other hand, after cultured for 24 h with 1μmol/L MG132, the cells in G1 and G2 phases were(72.33±3.44)% and(12.86±1.29)%, respectively, much higher than those in control group(63.63%±2.29% and 7.94%±1.91%, respectively). The early and late apoptosis rates in MG132 group were 33.57%±2.10% and 16.66%±0.47%, respectively, much higher than that in control group (7.18%±0.82% and 3.81%±1.06%, respectively). CONCLUSION: MG132 inhibits cell proliferation and induces cell cycle arrested at G1 and G2 phases, and cell apoptosis in NK/T cell lymphoma cells in a concentration dependent manner. Proteasome inhibitor may be a good drug to treat patients with advanced NK/T cell lymphomas.     

Inhibitory effect of berberine on the activation and proliferation of T lymphocytes

AIM: To investigate the effect of berberine (Ber) on the activation and proliferation of T lymphocytes and its mechanism of action. METHODS: Whole peripheral blood from normal subjects was stimulated with phytohemagglutinin (PHA) or phorbol ester (PDB) plus ionomycin (Ion) and the expression levels of CD69 and CD25 were evaluated with flow cytometry after the staining with appropriate fluorescent monoclonal antibody. The distribution of cell cycles was analyzed by propidium iodide staining and dead cells by 7-aminoactinomycin live staining. RESULTS: 100 μmol/L and 50 μmol/L of Ber had significant inhibition of the expression of CD69 on T cells stimulated with PDB plus Ion or PHA, while effect of 25 μmol/L Ber was not significant. And as time of action extended, the extent of inhibition decreased. For the expression of CD25, Ber at the concentrations as above all exerted significant inhibitory effect in a dose-dependent manner. Moreover, Ber could block lymphocytes cell cycle progression from G₀/G₁ phase to S and G₂/M phase without phase specificity. Besides, live staining analysis revealed that Ber did not have significant cytotoxicity on lymphocytes. CONCLUSIONS: Ber significantly inhibits the expression of early and mid activation antigens of T cells and also blocks the progression of lymphocytes cell cycles. These results suggest that Ber exerts immunosuppression effect through inhibiting the activation and proliferation of T cells.     

The inhibitory effect of quercetin on in vitro activation of T lymphocytes

AIM:To investigate the inhibitory effect of quercetin on in vitro activation of T lymphocytes by polyclonal activators with CD69 expression as an activation marker.METHODS:After being separated from lymphoid nodes of a C57BL/6 mouse, the lymphocytes were exposed to polyclonal activators (PDB or Con A) with or without quercetin. Then they were harvested at 2 h, 6 h and 24 h, respectively. The expressional rates of CD69 on T lymphocytes were assessed by two-color immunofluorescent staining and flow cytometry, and the inhibitory rates of quercetin at different time points were estimated.RESULTS:Quercetin had no effect on the expressional rate of CD69 on T lymphocytes under resting states. After the stimulation with PDB or Con A, the expressional rates of CD69 on T lymphocytes in the present of quercetin (10 μmol/L) showed significant decrease compared with those of control groups at different time points (P＜0.01). The inhibitory rate of quercetin on CD69 expression stimulated by PDB dropped sharply from 2 h to 24 h, whereas the inhibitory rate of quercetin on Con A action were relatively stable.CONCLUSION:Quercetin has inhibitory effects on the activation of T lymphocytes by Con A or PDB, suggesting that the action site of quercetin may be on PKCθ or its downstream. Furthermore,these inhibitory effect seems to be reversible.     

Effect of PRL-2 gene transfection on proliferation and cell cycle in hepatocellular cell line

AIM: To observe the changes of proliferation and cell cycle after PRL-2 gene effectively expressed in human hepatocellular cell line.METHODS: The PRL-2 vector was transfected into CL1 cell with lipofectamine reagent,the stable expression clones were screened by G418.The expression of PRL-2 mRNA was detected by real-time PCR.The expressive protein was identified by Western blotting.The subcellular localization was demonstrated by immunochemistry.The cell cycle was measured by flow cytometry.The population doubling time (TD) was analyzed by MTT assay.The expressions of cyclin A,cyclin D1,cyclin E,p16,p21Waf1 and p27Kip1 were detected by Western blotting.The p21Waf1 mRNA was determined by real- time PCR.RESULTS: The full length ORF of PRL-2 gene was inserted into the vector pcDNA3.1 (+),transfected into CL1 cells,and expressed successfully.Real-time PCR showed stable expression of PRL-2 mRNA.Western blotting confirmed the overexpression of PRL-2 protein.The subcellular localization of PRL-2 was in the plasmid.The proportion of cells in S-phase was increased.The population doubling time was reduced (P<0.01),a significant decrease was observed both in the mRNA and the protein expression of the p21Waf1 in comparison with untransfected or vector- transfected control cells (P<0.05).The expressions of cyclin D1,cyclin E,cyclinA,p16 and p27Kip1 were not appreciably different between the control and PRL-transfected cell lines.CONCLUSION: Eukaryocytic expression vector of PRL-2 has been successfully constructed,which shows stable and effective expression in CL1 cell line.PRL -2 increases cell proliferation by stimulating progression from G1 into S phase,which is primarily associated with decreased p21Waf1.     

Effect of HMGB2 on cell cycle and proliferation of lung adenocarcinoma

AIM: To investigate the effect of high-mobility group protein B2 (HMGB2) on cell cycle and proliferation of lung adenocarcinoma cells. METHODS: Cancer RNA-Seq Nexus (CRN) was used to analyze HMGB2 expression in lung adenocarcinoma tissues. OncoLnc was used to analyze the correlation between HMGB2 and prognosis of lung adenocarcinoma patients. Cancer Single-cell State Atlas (CancerSEA) was used to analyze the correlation between HMGB2 and 14 kinds of functional states of lung adenocarcinoma. siRNA was used to inhibit HMGB2 expression in human lung adenocarcinoma A549 cells. The silencing effects were verified by real-time PCR and Western blot, and the cell proliferation was detected by CCK8 and EdU assays. RESULTS: HMGB2 was over-expressed in the lung adenocarcinoma tissues. The overall survival of the patients with lung adenocarcinoma in HMGB2 high expression group was significantly lower than that of the patients with low expression of HMGB2 (log-rank test P=0.017 3). HMGB2 expression was positively correlated with cell cycle and proliferation of lung adenocarcinoma cells. The viability and proliferation ability of A549 cells after HMGB2 expression knock-down were significantly reduced (P < 0.05). CONCLUSION: The expression of HMGB2 is positively correlated with the cell cycle and proliferation of lung adenocarcinoma, and it can be used as a potential marker for evaluating the prognosis and therapeutic target of patients with lung adenocarcinoma.     

Effect of mycophenolic acid on T lymphocyte proliferation and cytokine expression in vitro

AIM: To investigate the effect of mycophenolic acid (MPA) alone or in combination with anti B7-1 mAb on proliferation of T lymphocytes and the possible mechanisms. METHODS: The proliferation of T lymphocytes was detected by BrdU incorporation method. The expressions of IL-2, IFN-γ and IL-10 in mRNA and protein levels were detected by RT-PCR and ELISA, respectively. RESULTS: (1) MPA markedly inhibited the T lymphocyte proliferation as compared with control (P<0.01). (2) MPA significantly inhibited the levels of IL-2 and IFN-γ (42.73 ng/L±14.64 ng/L vs 99.70 ng/L±9.15 ng/L, P<0.01; 7.87 ng/L±4.22 ng/L vs 82.42 ng/L±25.55 ng/L, P<0.05), and significantly increased content of IL-10 compared with control (770.95 ng/L±126.85 ng/L vs 545.71 ng/L±22.45 ng/L, P<0.05). MPA in combination with anti B7-1 mAb obviously enhanced the content of IL-10 compared with MPA alone (941.90 ng/L±56.61 ng/L vs 770.95 ng/L±126.85 ng/L, P<0.05). (3) The expression levels of IL-2 and IFN-γ mRNA in the MPA group were obviously lower than those in control (0.74±0.10 vs 1.17±0.15, 0.52±0.05 vs 0.75±0.12, P<0.01). MPA in combination with anti-B7-1 mAb showed a statistically significant increase in IL-10 mRNA expression (1.28±0.06 vs 0.84±0.09, P<0.01) as compared with MPA alone. CONCLUSION: MPA induces the changes of cytokine expressive spectrum and the Th1 and Th2 shift might be involved in the immunosuppressive effect. The combination of MPA with anti B7-1 mAb might have a synergic effect.     

Effect of microRNA-100 on proliferation activity and cell cycle of hepatocarcinoma cells

AIM：To investigate the effect of microRNA-100 (miR-100) on the proliferation activity and cell cycle of hepatocarcinoma cells. METHODS：Synthetic miR-100 mimic and its negative control were transfected into human hepatocarcinoma HepG2 cells by liposome method. After transfection, the cell counting kit-8 (CCK-8) was used to measure the cell proliferation activity. The cell cycle distribution was determined by flow cytometry. The expression of Polo-like kinase 1 (Plk1) at mRNA and protein levels was detected by quantitative real-time PCR (qRT-PCR) and Western blotting. RESULTS：The transfection efficiency mediated by cationic liposome was greater than 85%. The inhibitory rates of cell proliferation in HepG2 cells were (43.5±12.2)%, (46.5±3.7)% and (52.1±0.2)% at 24 h, 48 h and 72 h after transfected with miR-100 mimic, respectively, which were significantly increased as compared with the control cells. Moreover, the cell proliferation index in experimental group (35.8 ± 1.4) was higher than that in negative control group (39.2 ± 1.0) and simple liposome group (40.7 ± 2.0) at 72 h. At the same time, the mRNA and protein expression levels of Plk1 obviously decreased in HepG2 cells transfected with miR-100 at 72 h after transfection. CONCLUSION：miR-100 suppresses the proliferation activity of hepatocarcinoma cells by down-regulating Plk1 gene expression.     

Effects of human growth arrest-specific homeobox gene delivery on proliferation,migration and cell cycle distribution of vascular smooth muscle cells

AIM: To explore a new gene therapeutic strategy for vein graft restenosis by investigating the effects of adenovirus-mediated human growth arrest-specific homeobox (Ad5-hGax) gene delivery on the proliferation, migration and cell cycle distribution of serum-induced rabbit vascular smooth muscle cells (VSMCs). METHODS: The recombinant adenovirus vector containing hGax gene was constructed and transfected into rabbit VSMCs. The expression of hGax in VSMCs was detected by RT-PCR and immunofluorescent staining. Methyl thiazolyl tetrazolium (MTT) assay was used to assess the effect of hGax over-expression on serum-induced proliferation of VSMCs. Wound healing method was applied to examine the distance of serum-induce VSMCs migration. Flow cytometry was employed to analyze the cell cycle distribution. RESULTS: The recombinant adenovirus vector Ad5-hGax was successfully constructed. The results of RT-PCR and immunofluorescent staining revealed that the hGax -transfected cells contained a 174 bp specific fragment of hGax gene and target protein 48 h after transfection. The proliferation of serum-induced VSMCs was significantly inhibited by overexpression of hGax gene as compared with control group. The migration of serum-induced VSMCs was inhibited after hGax gene delivery. Flow cytometry showed that 72 h after serum induction, the cells in G₀/G₁ phase in Ad5-hGax group were significantly increased, whereas the cells in G₂/M+S phase were significantly decreased. CONCLUSION: Overexpression of hGax gene inhibits the proliferation and migration of serum-induced rabbit VSMCs, and arrests the cells in G₀/G₁ phase. It is likely that hGax gene is a potential target for the gene therapy of vein graft restenosis.     

NIH3T3 cell proliferation and cell cycle in the conditions of hypoxia or low nutrition and its response to bFGF

AIM: To investigate NIH3T3 cell proliferation and cell cycle in the condition of hypoxia or low nutrition and its response to bFGF, and to explore the pathophysiological changes of fibroblast under hypoxic or low nutrional conditions.METHODS: The cells were placed in anaerobic workstation where the mixture gas was given to simulate hypoxic environment. Partial oxygen pressure (PO2) of medium was controlled in 27 mmHg, 44 mmHg and 175mmHg. NIH3T3 cells were cultured with low nutritional medium contained new bovine serum (NCS) less than 10% to simulate low nutritional environment. MTT assay was used for observing cell activity and flow cytometry for cell cycle analysis.RESULTS: Under 44 mmHg PO2, no obvious difference was shown between hypoxia group and normal group. Under 27 mmHg PO2, the proliferation activity of NIH3T3 cells was significantly lower than that in normal group (P<0.01), as well as the cell numbers in G0-G1 phase increased (P<0.05), S phase decreased (P<0.01). bFGF had no effect on cell proliferation. In 0.5% NCS medium, the NIH3T3 cell proliferation speed decreased (P<0.01) and cell cycle was arrested at G0-G1 (P<0.01). The proliferation speed was improved by bFGF (P<0.01).CONCLUSION: In lower PO2 or lower nutrinal condition, fibroblast proliferation activity is inhibited by cell cycle arrest in G0/G1 phase. However the decreasing proliferation in low nutritional medium could be improved by external bFGF.     

Effect of tanshinone IIA on expression of cell cycle regulators and proliferation of pancreatic cancer cells

AIM：To observe the effect of tanshinone IIA on the expression of cell cycle regulators and the proliferation of pancreatic cancer cell line BX-PC-3. METHODS： The pancreatic cancer cell line BX-PC-3 was treated with tanshinone ⅡA at various concentrations for 48 h. The inhibition of proliferation was measured by MTT method. The change of the cell cycle was detected by flow cytometry. The protein levels of cyclin A and cyclin D2 were determined by Western blotting. RESULTS：Tanshinoone IIA significantly inhibited the proliferation of BX-PC-3 cells in a dose-dependent manner. The cancer cells were arrested in stage G0/G1 after treated with tanshinone IIA at low dose. The protein levels of cyclin A and cyclin D2 were decreased after drug intervention. CONCLUSION：Tanshinone IIA inhibits the proliferation of human pancreatic cancer cell line BX-PC-3 and the expression of cell cycle-promoting factors (cyclin A and cyclin D2), which may be the mechanism of attenuating the proliferation of pancreatic cancer cells.     

Effect of EGFP gene transfection on the cell cycle distribution of primary cultured human chondrocytes

AIM: To investigate the effect of enhanced green fluorescence protein (EGFP) gene transfection on the cell cycle distribution of primary cultured human chondrocytes in order to establish a tracking method of cultured human nasoseptal chondrocytes. METHODS: pEGFP-N1 plasmid was amplified in E.coli, and purified by high purity kit. Primary cultured human chondrocytes,which were initially obtained from the nasoseptal cartilage, were cultured in vitro and transferred with pEGFP-N1 by means of electroporation with Amaxa nucleofector device. Transfering process and transient expression were evaluated by laser scanning confocal microscope (LSCM), the transfer efficiency and the cell cycle distribution were evaluated by flow cytometry. RESULTS: There was significant expression of EGFP at 24 h after transferring. The transfection efficiency of pEGFP-N1 into primary cultured human chondrocytes reached 35.37％ at 48 h. It didn't affect the process of cell adherance and had no effect on the cell cycle distribution. CONCLUSION: Primary cultured human chondrocytes, which were transfected with pEGFP, are alive in vitro, and the transferring process doesn't affect the cell cycle distribution. These results suggest that pEGFP-N1 is an ideal transient expression vector for primary cultured human chondrocytes and it might be a well tracer in construction tissue engineered cartilage.     

Effects of erythromycin and its derivatives on the proliferation of T lymphocyte in vitro

AIM: To study the effects of erythromycin and its derivatives on the proliferation and the apoptosis of T lymphocytes and to evaluate the anti-inflammatory mechanism of erythromycin derivatives. METHODS: The effects of four erythromycin derivatives without antibacterial action on the proliferation T lymphocytes were evaluated by MTT. The effects of the erythromycin derivatives on apoptosis in T lymphocytes were observed by flow cytometry and TUNEL staining. RESULTS: The proliferation of T lymphocytes were inhibited by erythromycin derivative-Ⅰ and erythromycin, the IC50 was (425.2±32.1) μmol/L and (606.3±35.4) μmol/L, respectively. The IC50 value of erythromycin derivative-Ⅰ was lower than that of EM (P<0.01). Erythromycin derivative-Ⅰ (3-30 mg/L) and EM (30-100 mg/L) induced T lymphocytes apoptosis in a concentration-related manner by flow cytometry and TUNEL, while erythromycin derivative-Ⅰ induced cells to die at concentration of 100 mg/L. CONCLUSION: The anti-inflammatory activity of erythromycin derivatives may be due partly to their effects on the proliferation and apoptosis of T lymphocytes.     

Phagocytosis of viable apoptotic cells inhibits the activation of T lymphocytes

AIM: To investigate whether viable apoptotic cells and phagocytosis of them affect the activation of T lymphocytes. METHODS: Ultraviolet irradiation was used to induce apoptotic cells in vitro and the model of phagocytosis of these cells was established. Cytokine TGF_β1 was detected by ELISA. The rate of apoptotic cells and phagocytosis of them were assessed by flow cytometry. Furthermore, flow cytometry was also employed to examine the expression of activation signs, such as CD69, CD25, CD71, of T lymphocytes under the intervention of apoptotic cells and macrophage which ingested apoptotic cells, to reflect whether the apoptotic cells and the phagocytosis of these cells could influence the activation of lymphocytes stimulated by Con A. RESULTS: Ingestion of apoptotic cells increased TGF_β1 secretion. Only the macrophages that had ingested apoptotic cells could suppress the activation of lymphocytes. The expression of the markers of lymphocytes activation such as CD69, CD25, CD71 had been restrained. These inhibition effects were abolished by monoclonal anti-TGF_β1 antibody. CONCLUSION: The macrophages that have ingested apoptotic cells inhibit expression of CD69, CD25 and CD71 of T lymphocytes stimulated by ConA. This effect is dependent on the increase in TGF_β1 secretion in local site.     

Effect of norcantharidin on proliferation and invasion of human breast cancer cell line SKBR3 in vitro

AIM: To investigate the effect of norcantharidin（NCTD）on proliferation and invasion of human breast cancer cell line SKBR3 in vitro and its anticancer mechanisms.METHODS: MTT assay was used to determine SKBR3 cell proliferation. Light and FACScan were used to detect apoptosis and cell cycle. The invasiveness of SKBR3 was evaluated by the adhesion test，Matrigel experiment and the crossing-river test.RESULTS: NCTD had inhibitive effects on growth of SKBR3 cells in a dose and time-dependent manner, with the IC₅₀ value of 12.5 mg/L at 24 h．The cells treated with 10 mg/L NCTD for 24 h and 48 h showed typical apoptotic morphology and hypodiploid peak before G₁ phase. The cell cycle was arrested at G₂/M phase. The apoptosis percentage was up to 3.44% and 6.17%, and the G₂/M percentage was up to 35.82% and 38.70%. NCTD also could inhibit obviously the adhesion, movement and invasive capability simulating human basement membrane of SKBR3. Its effect was also in a dose-dependent manner. In the NCTD-treated group, crossing-river time was prolonged significantly and passing-membrane cells markedly decreased. CONCLUSION: NCTD in vitro inhibits not only the proliferation and growth of human breast cancer cells but also invasion and metastasis of the cells at relatively low concentration. NCTD shows prominent anti-tumor effects.     

Effect of L-arginine on intimal proliferation and expression of related cell cycle regulatory factors after vascular injury in rats

AIM:To investigate the effect of L-arginine (L-Arg) on intimal proliferation and expression of related cell cycle regulatory factors after vascular injury in rats. METHODS:Rats were divided into three groups :sham operation group, balloon injury group(this group included balloon 48 h,7 d and 14 d subgroup) and balloon+L-Arg group. Neointima area were calculated morphologiocally. The expression of cyclin dependent kinase-2(CDK2),cyclin E and proliferation cell nuclear antigen (PCNA) were measured by means of immunohistochemical technique and computer image analyzer. RESULTS:After vascular balloon injury, the level of plasma NO decreased, CDK2、cyclin E and PCNA expressed in the media at 48 h and in the neointima at 7 d and 14 d but with low and undetected expression in the media, the expression of CDK2, cyclin E and PCNA increased with the intima thickening. Compared with balloon 14 d group, the plasma NO level increased (P＜0.01), the neointima area reduced by 59.1%(P＜0.01) and the positive expression indexes of CDK2, cyclin E and PCNA decreased by 36.1%, 46.3% and 76.2% respectively in balloon+L-Arg group (P all＜0.01). CONCLUSION:L-Arg can effectively repress intima proliferation after vascular injury, which may be associated with its inhibiting the proliferation of vascular smooth muscle cell through downregulating the excessive expression of CDK2, cyclin E and PCNA.     

Effect of over-expression of transcription factor CDX2 on proliferation and cell cycle of human gastric cancer cell line SGC-7901

AIM: To study the effect and the molecular mechanism of CDX2 over-expression on the proliferation, growth and cell cycle of human gastric cancer cell line SGC-7901. METHODS: The SGC-7901 cells in LV-CDX2-GFP group were transfected with the recombinant lentivirus vector LV-CDX2-GFP, the cells in LV-GFP group were transfected with the negative control lentiviral vector for the negative control, and the cells in blank control group were without any treatment. The cell proliferation was detected by CCK-8 assay. The cell cycle distribution was analyzed by flow cytometry. The expression of CDX2, Bax, Bcl-2, cyclin D1 and survivin was determined by semi-quantitative RT-PCR and Wes-tern blotting. RESULTS: Compared with LV-GFP group and blank control group, the proliferation activity of the SGC-7901 cells was significantly lower (P<0.05), the G0/G1 phase proportion increased (P<0.05), the mRNA and protein levels of Bcl-2, cyclin D1 and survivin were reduced (P<0.05), and the mRNA and protein levels of Bax were up-regulated (P<0.05) in LV-CDX2-GFP group. No statistically significant difference of the above indexes was observed (P>0.05) between LV-GFP group and blank control group. CONCLUSION: Over-expression of CDX2 mediated by lentivirus inhibits the proliferation and growth of human gastric cancer SGC-7901 cells and arrestes the cell cycle at G0/G1 phase, which may be related to down-regulation of Bcl-2, cyclin D1 and survivin and up-regulation of Bax.     

Effect of lactacystin and β-lactacystin on the activation and proliferation of T-lymphocytes

AIM: To evaluate the effects of lactacystin (LAC) and β-lactacystin (β-LAC), proteasome inhibitor, on the proliferation and activation of T lymphocytes. METHODS: Flow cytometry was used to analyse the proliferation and the expression of CD69, CD25 and CD3 in PHA activated T-lymphocytes. Furthermore, the expression of PA28 and IL-2 mRNA were assayed by competitive RT-PCR. RESULTS: (1) LAC and β-LAC significantly decreased the incorporation in PHA activated T-lymphocytes. (2) Although LAC and β-LAC did not affect the expression of CD69 at any time, they significantly inhibited the expression of CD25 (48 h, 72 h, P＜0.05). (3) In comparison with control, LAC and β-LAC significantly down-regulated the expression of PA28 and IL-2 mRNA (48 h, 72 h, P＜0.05).CONCLUSIONS: LAC and β-LAC significantly inhibit the proliferation and activation of T lymphocytes. Mechanisms involved are inhibition of CD25 and down-regulation of PA28 and IL-2 mRNA expression.