Effects of signal regulatory proteinα1 on proliferation of hepatocellular carcinoma: a preliminary study

BACKGROUND: Signal regulatory protein alpha1 (Sirpα1) is a negative regulatory factor, and inhibits receptor tyro- sine kinase-dependent cell proliferating signal. This study was undertaken to observe the effect of signal regulatory proteinα1 ( Sirpα1) on gankyrin, cyclin D1, CDK4 and Fas expression in Sk-hep1 mouse hepatoma carcinoma cell line. METHODS: BOSC 23 packed cells were respectively trans- fected by means of recombinated retrovirus including pLX- SN, pLXSN-Sirpα1 and pLXSN-Sirpα1Δ4Y2 with lipofec- tin, and various plasmid virus media (viral titer 2.1 × 106 CFU/ml) were collected and infected respectively in 80% confluent Sk-hepl cells. Transfected Sk-hep1 cells were se- lectively screened with G418 (1200 μg/ml), and Sk-hep1 cell lines transfected with various plasmids were obtained. The protein expressions of gankyrin, cyclin D1, CDK4 and Fas in various Sk-hep1 lines were determined by Western blotting. Various Sk-hep1 lines were recovered to culture with 10% fetal bovine serum at 12 hours and 24 hours after starving culture with free serum for 72 hours, and cells were collected to determine the percentage of S phase cells of proliferating cycle by flow cytometry. RESULTS: Sirpα1 transfection remarkably downregulated gankyrin and cyclin D1 expression. Sirpα1Δ4Y2 downregu- lation of gankyrin expression was greater than that of Sirpα1(P <0.05), but no significant effect of Sirpα1 and Sirpα1Δ4Y2 on CDK4 and Fas protein expression was ob- served in transfected Sk-hep1 lines (P >0.05). The per- centage of S phase cells significantly decreased in Sk-hep1 cells transfected with Sirpα1 and Sirpα1Δ4Y2 plasmids (vs pLXSN Sk-hep1, P <0.05). The percentage of S phase cells in various Sk-hep1 cells increased when recovering to culture with 10% fetal bovine serum at 12 hours, but the percentage of S phase cells in Sk-hep1 cells transfected with Sirpα1 was the lowest ( vs pLXSN and Sirpα1Δ4Y2 Sk- hepl, P<0.05). The percentage of S phase cells in trans- fected pLSXN Sk-hep1 cells was the largest (vs Sirpα1 and Sirpα1Δ4Y2 Sk-hepl, P <0. 05). There was no significant difference between the transfected Sirpα1 Sk-hepl cells and Sirpα1Δ4Y2 Sk-hep1 cells (P>0.05). CONCLUSIONS: Sirpα1 decreases gankyrin and cyclin D1 expression, and inhibits proliferation of liver carcinoma cells. It may be one of the forms for an Sirpα1 negative regulation of carcinogenesis and development of hepatocel- lular carcinoma.

Effects of Electroacupuncture at the Conception Vessel on Proliferation and Differentiation of Nerve Stem Cells in the Inferior Zone of the Lateral Ventricle in Cerebral Ischemia Rats

Objective： To observe the effects of electroacupuncture （EA） at the Conception Vessel on proliferation and differentiation of the nerve stem cells in the inferior zone of the lateral ventricle in cerebral ischemia rats. Methods： The model rats were prepared by occlusion of the middle cerebral artery for 2 hours and then by reperfusion. They were randomly divided into two groups： a control group and an EA group. Changes in differentiation and proliferation of the nerve stem cells were observed 7, 14 and 28 days after successful modeling. Results： As compared with the 7-day control group （C-7d group）, there was no significant difference （P〉0.05） in the numbers of 5-bromodeoxyuridine （Brdu） positive cells, Brdu/GFAP, Brdu/Nestin and Brdu/Nse double-labeled cells in the inferior zone of the lateral ventricle in the EA group 7 days after modeling. However, in the 14-day EA group （R-14d group） and the 28-day EA group （R-28d group）, the numbers of Brdu positive cells and Brdu/GFAE Brdu/Nestin, Brdu/Nse double-labeled cells significantly increased as compared respectively with the 14-day control （C-14d group） and the 28-day control （C-28d） group （P〈0.05 or P〈0.01）. Conclusions： EA at the Conception Vessel promotes differentiation and proliferation of the nerve stem cells in the inferior zone of the lateral ventricle in the cerebral ischemia rats, and may stimulate differentiation of the proliferous nerve stem cells towards the astrocvtes.

Effects of Spinach Powder Fat-Soluble Extract on Proliferation of Human Gastric Adenocarcinoma Cells

Four kinds of assays were used to study the effect of a fat-soluble extract of spinach powder (SPFE) on the proliferation of human gastric adenocareinoma cell line (SGC-7901) in vitro.These studies included: (Ⅰ) cell growth assay, (Ⅱ) colony forming assay, (Ⅲ) MTT colorimetric assay, and (Ⅳ) 3H-TdR incorporation assay. The concentrations of SPFE expressed as the level of β-carotene in the medium were 2×10-8, 2×10-7 and 2×10-6 mol/L β-carotene in assays (Ⅰ)～(Ⅲ), but 4×10- 8, 4×10-7 and 4×10-6 mol/L β-caretene in assay (Ⅳ) respectively. The results indicated that SPFE inhibited the prolifendion and colony forming ability of SGC-7901 cells. And in MTT assay, SPFE inhibited the viability of SGC7901 cells, but no inhibitory effect of SPFE was observed on the viability of lymphocytes in peripheral blood of healthy people. Finally, in the 3H-TdR incorporation test, both SPFE and β-carotene showed significant inhibitory effects on DNA synthesis in SGC-7901 cells, but SPFE was more effective than β-carotene.

Effects of caffeic acid phenethyl ester on proliferation of vascular smooth muscle cells in rats

Objective： To investigate the inhibitory effect of caffeic acid phenethyl ester（CAPE） on the proliferation of vascular smooth muscle cells （VSMC） activated by lipopolysaccharide （LPS） and to clarify its mechanism. Methods： VSMC activated by LPS （1 mg-L^-1） were treated with CAPE at different concentrations. The inhibitory effecfs of CAPE on the proliferation of VSMC were determined by methabenzthiazuron（MTT） colorimetry. The effects of CAPE on the expression of proliferating cell nuclear antigen （PCNA） and Survivin protein in VSMC were evaluated by immunocytochemistry staining technique （SABC method）. Cell cycle was analyzed by flow cytometry（FCM） with propidiumiodide （PI） labeling method. The relative expression level of Survivin mRNA was measured with real-time quantified RT-PCR technique. Results. CAPE exerted significant inhibitory effects on. proliferation of VSMC at concentrations ranging from 5 mg·L^-1 to 80 mg·L^-1, decreased the rate of cells positive for PCNA and Survivin protein and repressed the expressioh of Survivin mRNA in a dose- and time-dependent manner （P 〈 0.05）. FCM analysis displayed that CAPE up-regulated the ratio of G0/G1 stages and reduced the percentage of VSMC in S stage （P 〈 0.05）. Conclusion： CAPE can significantly inhibit the proliferation of VSMC activated by LPS in a dose- and time-dependent manner, which may be carded out through regulating cell cycle and repressing the expression of PCNA and Survivin.

Glycosylation-independent binding to extracellular domains 11-13 of mannose-6-phosphate/insulin-like growth factor-2 receptor mediates the effects of soluble CREG on the phenotypic proliferation of vascular smooth muscle cells

Background The present study aimed to investigate the detailed mode and specific sites for their binding as well as the functional relevance of this binding in the phenotypic proliferation of vascular smooth muscle cells(SMCs). Methods CREG knocked-down SMCs were employed to evaluate the biological activity of wtCREG and mCREG.Expressions of SMC differentiation markers SM myosin heavy chain(SM-MHC),SM-actin,heavy caldesmon and myocardin were determined by Western blotting using specific antibodies. Cellular growth of SMCs was assessed by bromide dewuridine (BrdU) incorporation and cell cycle analysis on fluorescence-activated cell sorting(FACS).A solid-phase binding assay was used to study the binding of CREG to extracellular domains of M6P/IGF2R.The cellular co-localization of the two recombinant CREGs with M6P/IGF2R was detected on SMC surface by immunoprecipitation and immunofluorescence analysis.Results The molecular weight of wtCREG was around 30 kD while that of the mCREG was～25 kD.Treatment of wtCREG with PNGase F reduced its molecular weight from～30 kD to～25 kD,whereas PNGase F treatment had no effect on the molecular weight of mCREG.Both wtCREG and mCREG proteins enhanced SMC differentiation,inhibited BrdU incorporation,and arrested cell cycle progression when added to the culture medium.In CREG knocked-down SMCs,the amount of CREG detected by immunoblotting in M6P/IGF2R immunoprecipitates was significantly reduced when compared to normal cells.Both recombinant CREGs co-immunoprecipitated with M6P/IGF2R, although slightly reduced amount of the mutant CREG was detected in M6P/IGF2R immunoprecipitates.Immunostaining revealed that His-tagged CREGs co-localized with IGF2R on the cell surface in a glycosylation-independent manner.In vitro binding assay showed that CREGs bound to M6P/ IGF2R extracellular domains 7-10 and 11-13 in a glycosylation -dependent and -independent manner,respectively.Further blocking experiments using soluble M6P/IGF2R fragments and M6P/IGF2R neutralizing antibody indicated that the biological activities of recombinant CREGs in SMC growth and the up-regulation of SMC differentiation markers were all abolished by treatment with the M6P/IGF2R neutralizing antibody. However,although the growth inhibitory effect of wtCREG was nearly abolished by D7-10 or D11-13,the effect of mCREG was only reversed by Dll-13,indicating that the binding to domains 11-13 is required for CREG to modulate the proliferation of SMCs.Conclusions These data suggest that solubleCREG proteins can exert their biological function via binding to the extracellular domains 7-10 and 11-13 of cell surface M6P/IGF2R in both a glycosylation-dependent and -independent manner.

Effects of Silencing BAK1 and BCL2 Gene Expression on Proliferation, Invasion and Metastasis of HCC HepG2 Cells

Objective:To investigate the effects of silencing BAK1 and BCL2 gene expression on proliferation,invasion and metastasis of hepatocellular carcinoma(HCC)HepG2 cells.Methods:30 HCC HepG2 cells were randomly divided into groups and received the corresponding treatments,namely,control group,NC-siRNA group,BAK1-siRNA group,BCL2-siRNA group and BAK1+BCL2 group,with 6 strains in each group.ThenqRT-PCR,CCK8,Transwell chamber invasion and scratch assay were used to detect the expression,proliferation,invasion and metastasis of BAK1 and BCL2 genes in HepG2 cells.Results:The mRNA expression,cell proliferation rate,cell migration rate and cell invasion ability of BAK1 and BCL2 in HepG2 cells were lowest in the BAK1+BCL2 siRNA group,followed by BCL2-siRNA group,BAK1-siRNA group,NC-shRNA group and control group(P<0.05).The proliferation rate of HepG2 cells in the BAK1+BCL2 siRNA group decreased significantly with time(P<0.05).Conclusion:Silencing the expression of BAK1 and BCL2 genes can inhibit the proliferation and invasion of HCC HepG2 cells and promote their apoptosis.

EFFECTS OF BLEOMYCIN A5 COMBINED WITH CALMODU-LIN INHIBITOR ON THE PROLIFERATION OF S-180 CELLS IN VITRO

The effects of bleomycin A5 (BLM A5) alone and combined with calmodulin inhibitor N-(4-aminobutyl)-5-chloro-2-naphthalene sulfonamide (W-13) on the proliferation on S-180 cells in vitro were studied. IC50 of BLM used alone for the cells was about 2.63 μg/ml, but it was reduced to 1/3.8 and 1/9.5 of 2.63 μg/ml when plus W-13 1, 5 μg/ml respectively. The results indicated that nontoxic doses of W-13 enhanced the hinibition of cell proliferation under the condition of BLM 0.5 - 2.5 μg/ ml. In colony forming test, the survival fraction of S-180 cells treated with BLM plus W-13 was decreased to 1/87 - 240 of that of the cells treated with BLM alone. The results suggest that W-13 can enhance antitumor activity of BLM in vitro and may be used as an synergist of BLM A5 in vivo.

A new cerebroside and its anti-proliferation effect on VSMCs from the radix of Cyperus rotundus L.

A new cerebroside,1-O-(β-D-glucopyranosyloxy)-(2S,3R,4E,8Z)-2-[(2′R)-2’-hydroxylignoceranoylamino]-4,8-tetradecene-3- diol was isolated from the 60%EtOH extract of traditional Chinese medical plant Cyperus rotundus L.Its structure was determined on the basis of spectroscopic data.This new compound showed anti-proliferation effect on vascular smooth muscle cells(VSMCs).

EFFECT OF ADDED DIETARY CALCIUM ON ESOPHAGEAL EPITHELIAL-CELL PROLIFERATION IN SUBJECTS AT HIGH RISK FOR ESOPHAGEAL CANCER: A DOUBLE-BLIND INTER-VENTION STUDY

A randomized double-blind intervention trial was carried determine whether oral calcium supplementation could lower the proliferation of epithelial cells of the esophagus. 41 subjects identified with precancerous lesions by histopathology were randomized to receive oral supplementation of their conventional diets with 0.6 g of calcium as calcium carbonate or placebo. Both at the entry to the study and at the end of the treatment, seven months later, the subjects were examined, with an emphasis on the frequency and distribution of proliferating epithelial cells of the esophagus. Patterns of cell proliferation was defined by dividing the esophageal epithelium into cell columns oriented perpendicularly to the basal cell layer and by comparing the numbers and fractions of tritiated thymidine-labeled epithelial cells in the various cell columns and cell compartments.Before dietary supplementation with calcium, the profile of proliferating epithelial cells in the esophageal compartments in calcium group is similar to that in the placebo group, which is comparable to that previously observed in subjects with high risk for esophageal cancer. Seven months after supplementation having been started, in calcium group, proliferation was significantly reduced and the profile of the esophageal columns approached to that previously observed in subjects at low risk for esophageal cancer, however, in the placebo group, the proliferation and profile maintain at the same level as that before supplementation. Our findings indicate that oral calcium supplementation induces a more quiescent equilibrium in epithelial-cell proliferation in the esophageal mucosa of the subjects at high-risk for esophageal cancer, similar to that observed in subjects at low risk.

The effect of neuropeptides on proliferation of rat bone marrow mesenchymal stem cells

Objective To investigate the effects and mechanism of calcitonin gene-related peptide(CGRP)and substance P (SP) on proliferation of rat bone marrow mesenchymal stem cells.Methods The rBMSCs were isolated using whole bone marrow

Effect of hypoxia-inducible factor-1α on proliferation and invasion of prostate cancer PC-3 cell in hypoxic situation

Objective We transfected recombinant expression plasmid of pcDNA3. 1-HIF-1α into prostate cancer cells, to research effect of HIF-1α on proliferation of prostate cancer cell PC-3. Methods We selected a stable expression cell line with G418 we selected by transfection

Experimental study on effect of endothelin in the proliferation and collagen synthesis of human scar-derived fibroblasts

Objective To investigate the role of endothelin(ET) in the proliferation and collagen synthesis of human scar-derived fibroblasts and the moduktion of its antagonists such as nitric oxide(NO), tetrandrine ( Tet). Methods With the cultured fibroblasts from the scarring tissue, the cell pdiferation was determined by[3H]-TdR incorporation, while the collagen synthesis was evaluated by[3H]-proline incorporation. Results The ET-1 was significantly increasing the proliferation and collagen synthesis of human scar-derived fibroblasts. The values of [3H]-TdR absorption in the 2.5 ng/ml,25 ng/ml and 100 ng/ml of ET-1 groups were 1.8 times,4 times and 4.9 times more than in the control group, respectively(P【0. 01),while the values of the [3H]-proline incorporation were 1.1 times,3.1 times and 3.8 times respectively(P【0.01). The fibroblasts, treated with 50 μg/ml of S-nitroso-N-acetyl penicillamine(SNAP), were no detectable effect on the basal level of DNA synthesis,but produced decreasing effect on the

Cloning and expression of human arresten gene and effect of its recombinant protein on endothelial cell proliferation

To clone human arresten gene and investigate biological activity of the recombinant protein.Methods Human arresten gene was obtained from the plasmid pGEMArr and subcloned into the BamHⅠ and Pst Ⅰ restriction sites of prokaryotic expression vector pRSET containing T7 promoter.The recombinant plasmid pRSETAN was subsequently transformed into the strain E.coli BL21(DE3),and the target gene was expressed under induction of IPTG.The expressed protein was extracted,purified by Ni 2+ chelation affinity chromatography and refoled.The effect of the recombinant protein on proliferation of human umbilical vein endothelial cells (HUVECs) was also analyzed with the MTT assay.Results Endonuclease digesting and DNA sequencing confirmed that the arresten gene was correctly inserted into the expression vector.The recombinant protein was hightly expressed in the form of inclusion body in the host bacteria after induction.SDS-PAGE analysis revealed that the recombinant protein with a molecular weight of 26×103 amounted to 27% of the total bacterial proteins.The purity of the expected protein could reach over 96% through affinity chromatography.After renaturation,the recombinant protein could reach over 96% through affinity chromatography.After renaturation,the recombinant protein could significantly suppress proliferation of human umbilical vein endothelia cells(HUVECs) induced by vascular endothelial growth factor(VEGF).Conclusion Human arresten gene was successfully cloned into the expression vector pRSET and expressed at high level in Escherichia coli.Purified and refolded arresten protein could effectively inhibit proliferation of vascular endothelia cells.2 refs.

Effects of the Reynolds number on the mean skin friction decomposition in turbulent channel flows

As the Reynolds number increases, the skin friction has been identified as the dominant drag in many practical applications. In the present paper, the effects of the Reynolds number on the mean skin friction decomposition in turbulent channel flows up to Reτ= 5 200 are investigated based on two different methods, i.e., the FukagataIwamoto-Kasagi(FIK) identity(FUKAGATA, K., IWAMOTO, K., and KASAGI, N.Contribution of Reynolds stress distribution to the skin friction in wall-bounded flows.Physics of Fluids, 14(11), L73–L76(2002)) and the Renard-Deck(RD) identity(DECK,S., RENARD, N., LARAUFIE, R., and WEISS, P.′E. Large-scale contribution to mean wall shear stress in high-Reynolds-number flat-plate boundary layers up to Reθ= 13 650.Journal of Fluid Mechanics, 743, 202–248(2014)). The direct numerical simulation(DNS) data provided by Lee and Moser(LEE, M. and MOSER, R. D. Direct numerical simulation of turbulent channel flow up to Reτ≈ 5 200. Journal of Fluid Mechanics,774, 395–415(2015)) are used. For these two skin friction decomposition methods, their decomposed constituents are discussed and compared for different Reynolds numbers.The integrands of the decomposed constituents are locally analyzed across the boundary layer to assess the actions associated with the inhomogeneity and multi-scale nature of turbulent motion. The scaling of the decomposed constituents and their integrands are presented. In addition, the boundary layer is divided into three sub-regions to evaluate the contributive proportion of each sub-region with an increase in the Reynolds number.

Maintaining proton homeostasis is an essential role of the Warburg effect in proliferating cells

Non-proliferating cells efficiently generate adenosine 5’-triphosphate (ATP) through mitochondrial oxidative phosphorylation.By contrast, proliferating cells, including cancer cells, tend to rely on aerobic glycolysis, an inefficient way to generate energy, and this phenomenon is termed 'the Warburg effect'1,2.However, the advantage of the Warburg effect provided for proliferating cells has been unclear3.Here we propose that aerobic glycolysis may maintain proton homeostasis to benefit proliferating cells.

Measuring the dragging effect of natural resources: Evidence from the Yangtze River Delta metropolitan areas

This paper extends the resource drag studies by empirically investigating how spatial factors affect the regional economic growth. Using spatial panel econometric models, this paper estimates the dragging effect of energy resources of the Yangtze River Delta metropolitan areas. We fi nd that the growth drag of energy in the Yangtze River Delta is about 6% on average, which means that energy constraints decrease the economic growth by 6% annually, higher than the national level that has been previously measured in the literature. This result has taken into account the impact of neighboring cities' economic development, so as to obtain a more accurate estimate. Based on these measurement results, we propose some policy recommendations.

Impact of the Drag Force and the Magnus Effect on the Trajectory of a Baseball

We consider the impact of drag force and the Magnus effect on the motion of a baseball. Quantitatively we show how the speed-dependent drag coefficient alters the trajectory of the ball. For the Magnus effect we envision a scenario where the rotation of the ball confines the Magnus force to the vertical plane;gravity, drag force and the Magnus force make a trio-planar system. We investigate the interplay of these forces on the trajectories.

EFFECTIVE DIFFUSION AND EFFECTIVE DRAG COEFFICIENT OF A BROWNIAN PARTICLE IN A PERIODIC POTENTIAL

We study the stochastic motion of a Brownian particle driven by a constant force over a static periodic potential. We show that both the effective diffusion and the effective drag coefficient are mathematically well-defined and we derive analytic expressions for these two quantities. We then investigate the asymptotic behaviors of the effective diffusion and the effective drag coefficient, respectively, for small driving force and for large driving force. In the case of small driving force, the effective diffusion is reduced from its Brownian value by a factor that increases exponentially with the amplitude of the potential. The effective drag coefficient is increased by approximately the same factor. As a result, the Einstein relation between the diffusion coefficient and the drag coefficient is approximately valid when the driving force is small. For moderately large driving force, both the effective diffusion and the effective drag coefficient are increased from their Brownian values, and the Einstein relation breaks down. In the limit of very large driving force, both the effective diffusion and the effective drag coefficient converge to their Brownian values and the Einstein relation is once again valid.

Frame Dragging Effect on Properties of Rotating Neutron Stars with Strong Magnetic Field

The general relativistic frame dragging effect on the properties,such as the moments of inertia and the radiiof gyration of fast rotating neutron stars with a uniform strong magnetic field,is calculated accurate to the first orderin the uniform angular velocity.The results show that compared with the corresponding non-rotating static sphericalsymmetric neutron star with a weaker magnetic field,a fast rotating neutron star(millisecond pulsar)with a strongermagnetic field has a relative smaller moment of inertia and radius of gyration.