Effects of rosiglitazone,a PPARγ ligand,on chemoprevention of MNNG-induced gastric cancer in rats

AIM: To examine the chemo-preventive effects of peroxisome proliferator-activated receptor γ(PPARγ) ligand rosiglitazone (RSG) on a rat model of gastric carcinogenesis induced by chemical carcinogen N-methyl-N’-nitro-N-nitrosoguanidine (MNNG). We also attempted to identify novel anti-cancer mechanisms of rosiglitazone.METHODS: Ninety male Wistar rats were randomly allocated into six groups: group A (control group); group B (MNNG group); group C, D and E (RSG group, given different concentrations of rosiglitazone). The treatment procedures were terminated at 40th week. Stomach was harvested and gastric carcinoma was verified by histology. The gastric cancer incidence in different groups was calculated. To elucidate the mechanisms underlying the chemo-preventive effects of PPARγ ligand, we examine the gene expression profiles of MNNG induced gastric cancer and the rosiglitazone treated gastric cancer with Uniset Rat I Bioarray microarray.RESULTS: Incidence of gastric cancer in group A-E was 0% (0/10), 70% (14/20), 15% (3/20), 30% (6/20) and 30% (6/20), respectively. Gastric cancer incidence in group C, D and E was significantly lower than that in group B (P<0.01). A gene that showed prominent responses in rosiglitazone treated group was identified. The hypertension-related, calcium-regulated gene (HCaRG) was significantly upregulated in rat gastric carcinoma in rosiglitazone treated group when compared to MNNG group. The expression of HCaRG was down-regulated in human gastric cancerous tissue. CONCLUSION: PPARγ ligand rosiglitazone has a potent chemo-preventive effect against gastric cancer development in rats. Upregulation of HCaRG may be one of the mechanisms underlying the chemo-preventive effect of rosiglitazone in gastric cancer.     

Effects of simvastatin on proliferation of esophageal cancer cells through NF-κB p65 pathway

AIM To investigate the effects of simvastatin （SIM） on the proliferation of esophageal cancer Eca109 cells through NF-κB p65 pathway. METHODS Esophageal cancer Eca109 cells were cultured in vitro and treated with SIM at different concentrations （2 , 4, 8, 16 and 32 μmol/L）. The proliferation of Eca109 cells was measured by MTT assay and plate colony formation assay. The proliferation-related protein levels of cyclin D1, c-Myc, NF-κB p65 （p65） and IκB-α in the esophageal cancer Eca109 cells were determined by Western blot. ELISA was used to detect the levels of tumor necrosis factor α （TNF-α） and interleukin-6 （IL-6） in the culture supernatant of Eca109 cells. RESULTS Compared with control group, the inhibitory effects of SIM at 2 , 4 , 8 and 16 μmol/L on the viability of Eca109 cells were increased in a concentration-dependent and time-dependent manners （P<0.05）. Simultaneously, the inhibitory rates of SIM at 16 and 32 μmol/L on the viability of Eca109 cells showed no significant difference （P>0.05）. The inhibitory rates of SIM at 16 μmol/L on the viability of esophageal cancer cells was 50.61% at 48 h, which was closed to half of the inhibitory dose IC₅₀, and it was used as the optimum concentration and time for follow-up experiments. Compared with control group, the plate colony formation rate of Eca109 cells, the protein levels of cyclinD1, c-Myc, nuclear p65, TNF-α and IL-6 in 8 and 16 μmol/L groups were decreased, while the levels of cytosolic p65 and IκB-α proteins were increased （P<0.05）. No significant difference of plate colony formation rate in Eca109 cells, and the protein levels of cyclin D1, c-Myc, nuclear and cytoplasmic p65, IκB-α, TNF-α and IL-6 between 16 μmol/L SIM group and 32 μmol/L SIM group was observed （P>0.05）. CONCLUSION Simvastatin inhibits the proliferation of esophageal cancer Eca109 cells, and its mechanism may be related to up-regulation of IκB-α, down-regulation of cyclinD1 and c-myc, inhibition of p65 nuclear translocation, and the expression of TNF-α and IL-6 downstream of NF-κB p65 pathway.     

Construction of recombination vector of pcDNA3.1(+)-MER-Syk（L）and the effect of Syk(L) on the proliferation of breast cancer cells

AIM: To explore the effect of 4-hydroxytomacifen (4-OHT) on MER-Syk(L) cellular localization and the function of Syk(L) on cell proliferation in breast cancer cells.METHODS: pcDNA3.1(+)-MER-Syk(L) vector was constructed and the cell line MDA-MB-231, which expressed MER-Syk(L) stably, was established. Western blotting and immunofluorescence techniques were used to detect localization of MER-Syk(L) fusion protein in MDA-MB-231 cells cultured with or without 4-OHT. MTT assay was used to explore the proliferation ability of MDA-MB-231 stable cells.RESULTS: (1) MER-Syk(L) fusion protein, not MER-Syk(S) and MER protein, translocated from cytoplasm to nucleus in the presence of 4-OHT. (2) Nuclear not cytoplasmic MER-Syk(L) fusion protein inhibited MDA-MB-231 stable cell growth. (3) With or without the treatment of 4-OHT, MER-Syk（S）and MER protein always located in cytoplasm and did not suppress cell growth.CONCLUSION: With 4-OHT, MER-Syk(L) fusion protein translocates to nucleus and inhibits cell growth.     

Reversal of multidrug resistance by transfection of tumor necrosis factor α and MDR1 antisense RNA into multidrug resistant breast cancer cell line

AIM: To study the reversal effects of multidrug resistance by transfecting tumor necrosis factor α (TNF-α) cDNA and multidrug resistant 1 (MDR1) gene antisense RNA into multidrug resistant breast cancer cell line MCF-7/ADR. METHODS: The recombinant vector of enhanced green fluorescent protein (EGFP) with MDR1 antisense RNA and recombinant vector of red fluorescent protein (DsRed2) with TNF-α cDNA were constructed by RT-PCR and DNA recombinant techniques. The recombinant vectors were transfected into multidrug resistant breast cancer cell line MCF-7/ADR. The cell growth curves, cell apoptosis rates, MDR1 gene expression at mRNA and P-gp levels, and the sensitivity to ADR were determined before and after the transfection. RESULTS: After the transfection, cells showed lower growth rate, higher apoptosis rate, lower MDR1 expression at mRNA and P-gp levels, and the sensitivity to ADR increased significantly. CONCLUSION: Transfection of TNF-α cDNA and MDR1 antisense RNA into multidrug resistant breast cancer cells may have good effects on reversal of multidrug resistance.     

PCDH10 inhibits proliferation of breast cancer MDA-MB-231 cells by NF-κB/cyclin D1 signaling pathway

AIM To investigate the inhibitory effect of protocadherin 10 （PCDH10） on proliferation of human breast cancer cells and its possible mechanism. METHODS RT-PCR was used to measure the mRNA expression levels of PCDH10 in 4 human breast cancer cell lines and normal mammary epithelial MCF-10A cells. The breast cancer cell line MDA-MB-231 with stable PCDH10 overexpression was constructed by recombinant lentivirus （pLV-PCDH10） infection, and blank control （blank） group and negative control （pLV-NC） group were also set up. The cell proliferation ability was detected using methyl thiazolyl tetrazolium （MTT） and colony formation experiments. The cell cycle was detected by flow cytometry. Western blot was used to determine the expression of cyclin D1, cyclin-dependent kinase 4 （CDK4）, nucear factor-κB p65 subunit （NF-κB p65） and NF-κB inhibitor α （IκBα）. RESULTS The mRNA expression levels of PCDH10 in 4 breast cancer cell lines were lower than that in normal mammary epithelial MCF-10A cells （P<0.05）. A breast cancer cell line MDA-MB-231 with stable PCDH10 overexpression was constructed successfully. Compared with negative control group, PCDH10 overexpression significantly inhibited cell proliferation, induced cell cycle arrest at G₁ phase, down-regulated the expression of cyclin D1 and CDK4, and decreased phosphorylation of NF-κB p65 and IκBα（P<0.05）. CONCLUSION PCDH10 inhibits the proliferation and blocks cell cycle progression of breast cancer cells by targeting NF-κB/cyclin D1 signaling pathway.     

Effect of dominant negative IκBα transfection on NF-κB pathway and cell motility in breast cancer

AIM: To observe the effect on NF-κB pathway and cell motility in breast cancer cell lines after transfection of dominant negative IκBα plasmid.METHODS: After stable transfection of mutant IκBα plasmid into highly metastatic breast cancer lines MDA-MB-231 and MDA-MB-435, we detected NF-κB binding activity by EMSA, cell growth ability by cell growth curve, colony forming test, and cell motility by millicell-PCF chamber.RESULTS: Constitutive activities in MDA-MB-231 and MDA-MB-435 were observed. Stable transfection of a dominant negative ⅠκBα resulted in downregulation of NF-κB binding activity, thus inhibited cell mobility without significant effect on cell growth.CONCLUSION: Cell migration ability is inhibited in highly invasive breast cancer cells by inhibition of NF-κB pathway in vitro.     

Regulatory effect of NOX-4 on PI3K signaling pathway in TGF-β1-induced collagenⅠsynthesis from lung cancer cells

AIM: To investigate the regulatory effect of NADPH oxidase-4 (NOX-4) on PI3K signaling pathway in transforming growth factor-β1 (TGF-β1)-induced collagen type I (collagen I)synthesis from lung cancer cells and the mechanisms. METHODS: Human lung cancer A549 cells were cultured in vitro and stimulated with TGF-β1. The expression of NOX family and collagen family at mRNA and protein levels as well as the PI3K class I catalytic subunits and the activation of PI3K signaling pathway was measured. A549 cells were pre-treated with NOX-4 inhibitor diphenyleneiodonium (DPI), and the expression of collagen I at mRNA level as well as the PI3K class I catalytic subunits and the activation of PI3K signaling pathway was measured upon TGF-β1 stimulation. RESULTS: TGF-β1 stimulated the expression of NOX-4 and collagen I at mRNA and protein levels as well as the expression of PIK3CD and the activation of PI3K signaling pathway at a dose-and time-dependent manner. NOX-4 inhibitor DPI partly reversed TGF-β1-induced collagen I expression. Inhibition of NOX-4 down-regulated the degree of TGF-β1-stimulated activation of PI3K signaling pathway without effect on the expression of PIK3CD. CONCLUSION: NOX-4 participates in TGF-β1-induced collagenⅠsynthesis from lung cancer cells via regulating the activation of PI3K signaling pathway. TGF-β1/NOX-4/PI3K signaling pathway axis acts as a regulatory role in collagenⅠsynthesis from lung cancer cells.     

The Effect of Conditions of Storage on the Respiration of Apples: III. The Effect of Modulation of Temperature on the Respiration of Cox’s Orange Pippin Apples,and on the Extent of Low Temperature Injury

When apples which develop low temperature breakdown (LTB) at 32° F. are moved from 32° F. to 65° F. for 3 to 5 days at about the 7th to 8th week of storage, they subsequently develop within a given period of storage less LTB than apples kept at 32° F. continuously.

The respiration of apples susceptible to LTB increases steadily during storage at 32° F. If these apples are warmed to 65° F. during the period of exposure to 32° F., the subsequent rate of respiration at 32° F. is lower than before warming, and continues at a lower rate than for apples kept at 32° F. continuously.

If the apples are moved to 38° F., without an intermediate treatment at 65° F., the rate of respiration is higher than for apples at 38° F. continuously, and this higher rate persists.

If there is an intermediate wanning period at 65° F., the respiration of apples moved from 32° to 38° F. is of the same order as that for apples kept at 38° F. continuously.

The respiratory quotient of apples at 32° F. or at 38° F., which is indicative of the type of respiratory activity, is typical for the temperature at which it is measured, and is not affected by the warming treatment. The effects of wanning on both the incidence of LTB and respiration are similar for apples stored in air and in 2% oxygen: 98% nitrogen.     

Modelling the effects of timing and rates of application of benzyladenine as a secondary thinner of ‘Fuji’ apple after ethephon

A trial on ‘Fuji’ apples at the Grove Research Station in southern Tasmania during the 1991/92 season studied the thinning effect and interactions between ethephon and benzyladenine (BA) when BA was applied as a secondary thinner after a full bloom (FB) application of ethephon. The spray treatments were a factorial combination of eight application times of BA (11,13,15,17,19, 21, 23 or 25 days after full bloom (AFB)) with ten concentrations (20,40,60,80,100,120,140,160,180 or 200 mg I“1). An unsprayed control and an ethephon control were included. Target thinning results were achieved 19- 23 d AFB with concentrations of 140-160 mg I“1. A corollary of this successful thinning was an increase in fruit weight and size. Return bloom was improved significantly where thinning was successful. There was no effect on fruit firmness, soluble solids or lateral branching. Drawbacks were an increase in the incidence of russet and a reduction in pip number at the optimum thinning times and concentrations.     

Evaluation of the activity of aqueous extracts of some organic waste materials on in vitro-cultured shoots of ‘M9’ apple rootstock

Summary

The present work investigated the effects of different aqueous extracts of organic waste compounds on growth, proliferation and photosynthetic activity in ‘M9’ (Malus domestica Borkh.) shoot cultures, with the aim of determining the feasibility of using in vitro cultures as a tool for the rapid evaluation of organic amendments in agriculture. Aqueous extracts of the following organic waste compounds: cow manure (CM), sugarbeet industrial waste (SB), mixed grape, poultry and municipal solid waste (GPM), and citrus pruning and industrial waste (CPI) were prepared at a rate of 1:10 (w/v) compound:distilled water. The basal media used in the proliferation phase were: (i) PM1, modified Murashige and Skoog (MS) enriched with 4.4 µM 6-benzyladenine (BA); (ii) PM2, as PM1 but with a reduced cytokinin concentration (1 µM BA) to evaluate possible hormone effects; and (iii) PM3, 4.4 µM BA with reduced salt strength (0.33 MS) to induce nutrient deficiency. Hormone-free medium with half-strength MS salts was used for rooting. All media were enriched with each extract at 0, 0.2, 2, 20 or 200 ml l^–1. Photosynthetic activity was measured on PM3 medium enriched with SB or CM. Standard culture conditions were 22° ± 2°C, with a 16 h photoperiod at 30 µmoles photosynthetically active radiation (PAR) m^–2 s^–1, but at 80 µmoles PAR m^–2 s^–1 to determine photosynthetic activity. Shoot weight increase in PM1 was not affected by the GPM and CPI extracts, while the growth trends of CM- and SB-treated shoots were described by a second-degree function with maxima at 2 ml l^–1 and 0.2 ml l^–1, respectively. Shoot proliferation for SB was represented by a quadratic curve (maximum at 2 ml l^–1), was linearly reduced as GPM increased, but was not affected by CM or CPI. Treatments did not significantly affect rooting percentage and root length; however root number was increased by SB at 2 ml l^–1.CO₂ fixation increased linearly with both SB and CM, despite reduced growth at the highest levels of extract.     

Evaluation of the Effect of Different Harvest Time on the Fruit Quality of Foşa Nut

This study was carried out in Arsin (Trabzon/Turkey) in 2011. The effects of different harvest time and altitudes on the quality of the nuts have been investigated. The study was performed on Fo?a hazelnut and the harvest process has been conducted at three terms, which are on normal harvest time and ten days before and after harvest time. The harvested nuts were dried in the shade on the concrete floor until their moisture content decreased to 5?%. Some properties of nuts including yield, fruit weight, internal weight, shell thickness, and protein, oleic, and linoleic acid amounts have been investigated. As evaluated all of the fruit properties it can be concluded that 11 August is the most suitable harvest date for coast zone. On the other hand, no significant differences were obtained in the point of protein, oleic, and linoleic acid amounts for different harvest time and altitudes.     

Fate of photosynthates from spur leaves of ‘Nijisseiki’ pear during the period of rapid fruitgrowth

Summary

Spurs of ‘Nijisseiki’ pear (Pyrus pyriflora Nakai) were allowed to assimilate _10033;CO₂ at 87.d after anthesis (DAA) and at 108 DAA during the period of rapid fruit growth. Then the spurs were sampled periodically until fruit harvest time to trace the time course and amount of movement of assimilates from spur leaves to individual organs in the spur. The amount of _10033;C absorbed by fruit within 3.d after labelling was constant until harvest, regardless of the labelling date. However, the total amount of _10033;C in the spur decreased continually until harvest.Of the total amount of _10033;C recovered in the spur labelled at 87 DAA, by harvest, 43.2% of 13C was found in the fruit flesh (cortex of receptacle), 5% in the core (pith of receptacle 1 pericarp 1 seeds), 5.6% in spur stem, 5.4% in the source leaves, and 40.8% was respired and exported from the spur. Of total amount of initial _10033;C labelled at 108 DAA, at harvest, the proportion of _10033;C translocated to flesh, core, stem, respired, and remaining in leaves was 61%, 6.1%, 3.0%, 24% and 5.4%, respectively. Photosynthates fixed by the spur early in the rapid growth stage of the fruit contribute more to the formation of starch and structural materials and less to solublecarbohydrates in fruit than do those fixed later.     

Effect of Culture Parameters on the Growth of Sparassis crispa Mycelium

The effect of carbon and nitrogen sources, pH, temperature and substrate water content on the growth and vigor of Sparassis crispa mycelium was evaluated. Mycelial growth rates were highest when rice starch served as the carbon source and when peptone, yeast extract or soybean meal were adopted as the nitrogen source. The optimum pH and substrate water content values were 5.25 and 62.5% respectively. Mycelial growth was optimal at 20.2 ℃ and inhibited at 35 ℃, and temperatures of 40 ℃ and above killed the fungus.     

Is Nutrient Contamination of Groundwater Causing Eutrophication of Groundwater-Fed Meadows?

There is an ongoing debate as to whether nutrient contamination of groundwater under agricultural fields may cause nutrient-enrichment and subsequent eutrophication in discharge areas. Often, there is only circumstantial evidence to support this supposition (proximity of agricultural fields, direction of water flow, highly productive vegetation). Research on solute transport along a flow path is necessary to evaluate the risk for eutrophication. In this paper we present results of such a study. Two transects were established in a discharge meadow, a few meters downstream from fertilized cornfields. Highly productive vegetation in parts of the meadow suggested nutrient-enrichment caused by inflow of contaminated groundwater. This supposition was supported by an analysis of groundwater flow paths, residence times and chloride as tracer for pollution. However, the fate of nutrients along the flow path indicated otherwise. While we found high concentrations of DIN (dissolved inorganic nitrogen), P and K under the cornfields, DIN and P concentrations drop below detection limit when groundwater enters the meadow. Only K progressed into the meadow but did not enter the root zone. We conclude that (1) polluted groundwater from the cornfields did not cause the nutrient-enrichment, as indicated by the highly productive vegetation. Restoration projects in discharge areas should not focus upon measures in upstream areas if only circumstantial evidence is available. Solute transport should be considered as well. (2) Because K clearly showed to be the most mobile nutrient, its importance for nutrient-enrichment in discharge wetlands merits more attention in future research.     

Effects of α-MSH on the Ca2+ channels of primary DMNV cells and effects of anti-apoptosis on primary DMNV cells after activation of melanocortin receptors

AIM: α-MSH is elevated in patients with inflammatory bowel disease and has been implicated as an inflammatory mediator. The purpose of this study was to investigate effects of α-MSH on the Ca2+ channels of primary DMNV cells, the effects of gastrointestinal inflammation on the dorsal motor nucleus of the vagus in rats, as well as the effects of proinflammatory cytokines and α-MSH on neurons from the dorsal motor nucleus of the vagus in vitro. METHODS: In vitro studies the primary culture of neurons from the dorsal motor nucleus of the vagus was performed. Single-cell cytoplasmic calcium transients were determined using the fluorescence dye fura-2-AM. Cell proliferation and apoptosis were measured by enzyme-linked immunosorbent assay. RESULTS: MC4R mRNA was expressed in the DMNV cells of normal rats. Activation of MC4R promoted the calcium influx of primary DMNV cells. The addition of α-MSH to thrombin or trypsin resulted in significant decreases in apoptosis compared to thrombin or trypsin alone. CONCLUSION: Functionally active α-MSH receptors are linked to Ca2+ channels in DMNV neurons. In cultured DMNV cells, α-MSH attenuates neuronal apoptosis and reverses inhibition of cellular proliferation.     

Evaluation of the sensitivity to ionising γ-radiation of a Cymbidium hybrid

Gamma-rays are an important mutagenic agent that can induce new, useful genetic variations in plants. However, γ-irradiation can also cause damage that negatively affects the use of such mutagens in plant breeding. The aim of the present study was to investigate the effects and damage caused by γ-radiation in a Cymbidium hybrid, RB001. The relative growth rate of protocorm-like bodies (PLBs) was reduced by 50% at a γ-ray dose of approximately 40 Gy. Malondialdehyde concentrations increased significantly with increasing radiation dose. However, almost no difference was observed between untreated control PLBs and PLBs treated with 200 Gy 8 weeks after γ-irradiation. The activities of several antioxidant defence enzymes increased gradually with increasing γ-ray dose, 24 h after irradiation. These enzymes showed different responses between 1 and 4 weeks, but no difference 8 weeks after irradiation. The ‘comet assay’ and flow cytometry were performed. Clear differences in radiation-induced damage were observed between control and 200 Gy-treated PLBs at 24 h. However, PLBs had a tendency to recover from 4 weeks after irradiation, and the integrity of their DNA was similar in samples treated with 10–200 Gy. These results indicated that γ-rays caused little DNA damage and the plants could recover. This demonstrated the feasibility of using physiological responses, the ‘comet assay’, and flow cytometry to detect DNA damage after γ-irradiation.