Estrous cycle stage-dependent expression of acute tolerance to morphine analgesia in rats

Both baseline pain sensitivity and the response to antinociceptive treatment are sensitive to an animal's sex and estrous cycle stage. Sex differences are also observed in the development of antinociceptive tolerance induced by repetitive exposure to opiate drugs such as morphine. Conventional tolerance study protocols do not assess the impact of the estrous cycle stage. The present study aimed to compare the development of acute tolerance to morphine-induced antinociception in male and female (cycling and ovariectomized) Wistar rats using the tail-flick test. Acute tolerance was induced by two consecutive subcutaneous injections of morphine (10 mg/kg) or saline separated by an interval of 6 h. It was found that rats pretreated with morphine were tolerant to the second morphine dose. Tolerance was most pronounced in proestrous female rats and, to a lesser degree, in male rats. It was absent in ovariectomized rats as well as during the estrus, metestrus and diestrus phases. Thus, the estrous cycle exerts dramatic effects on the induction of acute tolerance to morphine-induced antinociception. These results suggest that pain management strategies can be optimized through the use of sex- and estrous cycle-specific techniques.     

Effects of cyclooxygenase 2 inhibitors on biological traits of nasopharyngeal carcinoma cells

AIM: To investigate effects of cyclooxygenase 2 (COX-2) inhibitors on nasopharyngeal carcinoma (NPC) cells and on angiogenesis in vitro. METHODS: Human NPC cell lines (CNE1, CNE2 and SUNE) were treated with nimesulide or celecoxib. MTT assay and colony formation assay were performed to observe antiproliferation activity of COX-2 inhibitors to NPC cell lines. Cell cycle arrest and apoptosis of NPC cell strains were tested by flow cytometry assay, microscopic morphology observation, and DNA fragmentation assay. The effect of COX-2 inhibitor on angiogenesis was tested by chick chorioallantoic membrane (CAM) model. RESULTS: Nimesulide (Nim) and celecoxib (Cel) could antiproliferate NPC cell lines with IC50 182 μmo1/LNim-SUNE, 78 vmol/LNim-CNE1, 175μmoI/LNim-CNE2, 7.2 μmol/LCet-SUNE, 8.1 μmoI/LCet-CNE1, and 7.6 μmol/LCet-CNE2. The antiproliferation presented dose-dependent (Nim 5-400 μmol/L, Cel 0.5-80 μmol/L) and time-dependent manner (Nim IC50 562 μmol/L24 h, 316 μmol/L 48 h,50.1 μmol/L240h). Nim and Cel arrested SUNE and CNE1 cell cycle at phase G2/M (cell aggregation rate 28.9 %-45.1%y.Nim25-200μmol-12h-SUNM,18.9 %-26.2 %Nim25-200μmol-24h-SUNM, 28.8%-35.6 %Nim25-200μmol-48h-SUNM,30.4 %-16.4 %Nim25-200μmol-12h-SUNM,21.2 %-19.7 %Nim25-200μmol-24h-SUNM, 31.1%- 19.9%Nim25-200μmol-12h-SUNM, 20.5 %-34.1%Nim25-200μmol-12h-SUNM,25.2 %-26.9 %Nim25-200μmol-12h-SUNM, 11.5 %-7.1% Nim25-200μmol-12h-SUNM,Apoptosis shape and apoptosis strap displayed in NPC cells after treatment with Nim and Cel. Nim had a feature of anti-angiogenesis on CAM model. CONCLUSION:Nim and Cel could suppress proliferation of squamous epithelium NPC cell (SUNE, CNE1 and CNE2) through blocking cell cycle and inducing cell apoptosis. Nim could apparently suppress CAM angiogenesis induced bySUNE cell.     

Metabolic sensitivity of pancreatic tumour cell apoptosis to glycogen phosphorylase inhibitor treatment

Inhibitors of glycogen breakdown regulate glucose homeostasis by limiting glucose production in diabetes. Here we demonstrate that restrained glycogen breakdown also inhibits cancer cell proliferation and induces apoptosis through limiting glucose oxidation, as well as nucleic acid and de novo fatty acid synthesis. Increasing doses (50-100 microM) of the glycogen phosphorylase inhibitor CP-320626 inhibited [1,2-(13)C(2)]glucose stable isotope substrate re-distribution among glycolysis, pentose and de novo fatty acid synthesis in MIA pancreatic adenocarcinoma cells. Limited oxidative pentose-phosphate synthesis, glucose contribution to acetyl CoA and de novo fatty acid synthesis closely correlated with decreased cell proliferation. The stable isotope-based dynamic metabolic profile of MIA cells indicated a significant dose-dependent decrease in macromolecule synthesis, which was detected at lower drug doses and before the appearance of apoptosis markers. Normal fibroblasts (CRL-1501) did not show morphological or metabolic signs of apoptosis likely due to their slow rate of growth and metabolic activity. This indicates that limiting carbon re-cycling and rapid substrate mobilisation from glycogen may be an effective and selective target site for new drug development in rapidly dividing cancer cells. In conclusion, pancreatic cancer cell growth arrest and death are closely associated with a characteristic decrease in glycogen breakdown and glucose carbon re-distribution towards RNA/DNA and fatty acids during CP-320626 treatment.     

Clusterin overexpression in both malignant and nonmalignant prostate epithelial cells induces cell cycle arrest and apoptosis

Expression of the castration-induced clusterin protein is incompatible with the survival of human prostate cancer cells in tissues and in cell culture. To investigate the fate of human prostate epithelial cells, when engineered to maintain expression of clusterin protein, we have used an IRES-hyg vector and hygromycin selection. PC-3 prostate tumour cells were substantially more sensitive to clusterin expression than nonmalignant PNT1a cells, showing multiple phenotypic changes including cell cycle arrest and increased apoptosis. The results strengthen the hypothesis that clusterin expression is proapoptotic. Expression of exogenous clusterin in both cell types resulted in its relocation from the cytoplasm and a nuclear accumulation of the protein, as was also seen in the same cells when apoptosis was induced by etoposide treatment. To survive clusterin expression, the PC-3 tumour cells developed apoptosis-inhibitory properties. This could have significance for the resistance of prostate cancers to chemo/radiotherapy, where clusterin overexpression is observed.     

Status of selenium in prostate cancer prevention

The complete, 13 years, results of the Nutritional Prevention of Cancer Trial have been analysed, causing some speculation over the robustness of the previously reported findings of reduction of cancer risks by supplements of selenium (Se) to a cohort of older Americans. These analyses confirmed that Se supplementation was associated with marked reductions in risks to total (all-site except skin) carcinomas and to cancers of the prostate and colon-rectum. Of those deep-site treatment effects, the most robust was for prostate cancer, which was more frequent, and was confirmed by serum prostate-specific antigen level. Recent subgroup analyses showed Se supplementation reduced risk of cancer mostly among subjects who entered the trial with plasma Se levels in the bottom tertile of the cohort. Other recent findings have demonstrated that Se treatment can promote apoptosis in prostate cancer cells and, possibly, impair their proliferation through antiangiogenic effects. Thus, a body of basic understanding is developing by which one can understand and evaluate the results of the Nutritional Prevention of Cancer and future clinical trials. This understanding also requires inclusion of the mechanisms of Se transport and cellular uptake, so that appropriate inferences can be made from findings from cell culture systems, which tended to use effective Se doses much larger than relevant to cells in vivo. Also needed is information on the chemical speciation of Se in foods, so that Se delivery can be achieved in ways that are effective in reducing cancer risk and is also safe, accessible and sustainable.     

Reduction of TSG101 protein has a negative impact on tumor cell growth

TSG101 was defined originally as a tumor-suppressor gene, raising the expectation that absence of the encoded protein should lead to increased tumor cell growth and, perhaps, increased tumor cell aggressiveness. We have used the RNA interference (RNAi) technique to downregulate TSG101 in PC3 (prostate cancer) and MDA-MB-231 (breast cancer) cells. An approximately 85% selective downregulation at the protein level was achieved in both cell lines over a period of 12 days as detected by Western blotting. This treatment resulted in inhibition of tumor cell growth, with a decreased level of TSG101 causing partial cell cycle arrest at the G(1)/S boundary and a reduction in the rate at which cells passed from G(2) through mitosis and back into G(1). In both cell lines, the percentage of cells in S-phase was reduced significantly at day 4 after the TSG101 siRNA transfection (27% vs. 41% in MDA-MB-231 cells; 22% vs. 39% in PC3 cells). Additionally, RNAi-mediated downregulation of TSG101 reduced the colony formation capacities of both cancer cell lines. Rather more surprisingly, TSG101 downregulation affected the migratory activity of the MDA-MB-231 cells, independent of any effect on proliferation. Thus, in a Transwell assay, after 4-hr incubation, 36.0% of control MDA-MB-231 cells had migrated to the lower chamber vs. 7.3% of TSG101-downregulated cells (p < 0.001; scrambled control, 36.5%). These results show that the TSG101 gene does not comply with the usual characteristics of a tumor-suppressor gene; rather, its expression may be necessary for activities associated with aspects of tumor progression.     

p53 polymorphic variants at codon 72 exert different effects on cell cycle progression

Two common polymorphic forms of the p53 tumor suppressor protein are widely distributed throughout the human population. These encode either proline or arginine at position 72, and this difference results in a marked alteration in the primary structure of the protein. A number of previous studies have shown significant differences in the biochemical properties of the p53 protein, depending on the particular polymorphic form. There is little information, however, on their respective biologic activities. In this study, we have used an inducible switch system for expressing both polymorphic forms of p53 within Saos-2 cells. Cell cycle analysis postinduction of p53 function reveals striking differences in how the 2 forms of p53 bring about a cessation of cell growth. Thus, the Arg72 form of p53 is significantly more efficient than the Pro72 form at inducing apoptosis. In contrast, the Pro72 form appears to induce a higher level of G1 arrest than the Arg72 form. These results demonstrate significant differences in how the codon 72 polymorphism affects the biological activity of p53.     

Transgenic overexpression of a dominant negative mutant of FADD that, although counterselected during tumor progression, cooperates in L-myc-induced tumorigenesis

Activation of the so-called death receptors, e.g., CD95/Fas/Apo-1, is a potent stimulus to trigger apoptosis. Overexpression of the C-terminal FADD deletion mutant FADD-DN blocks death receptor-induced apoptosis, but despite this antiapoptotic activity, lck FADD-DN transgenic mice do not develop lymphomas. To analyze whether functional inactivation of FADD cooperates with Myc overexpression in tumorigenesis, lck FADD-DN transgenic mice were crossed with Emicro L-myc transoncogenic animals. While no tumors were detected in single transgenic FADD-DN or L-myc mice within 15 months, 5 of 17 (29%) FADD-DN/L-myc double transgenic animals developed lymphomas with an average latency period of 47 weeks. Protein analysis of FADD-DN/L-myc tumors showed, however, undetectable levels of FADD-DN protein. FADD-DN protein expression was again lost in 16 of 17 FADD-DN/p53 k.o. T-cell lymphomas, though no significant acceleration of tumorigenesis in P53-deficient lck FADD-DN mice compared to p53 k.o. animals was observed. These data suggest a strong counterselection against the FADD-DN protein during tumor progression, which could be explained by the cell cycle inhibitory activity of FADD-DN. Such counterselection would have to be compensated for by other antiapoptotic mutations, and indeed, strong upregulation of the antiapoptotic Bcl-2 family member Bcl-xL was found in one of the tumors. This in vivo mouse model demonstrates that an antiapoptotic protein involved in the onset of tumorigenesis is selected against and consequently lost during tumor progression because of its additional antiproliferative activity.     

Comparison of estimated and measured maximal oxygen uptake during exercise testing in patients with chronic obstructive pulmonary disease

BACKGROUND: Maximal oxygen uptake (VO(2max)) and exercise modalities such as walking and standard pulmonary function testing are measurements that have been used by the surgical community as an indication of a patient's current exercise capacity to predict operative outcomes. There are equations available in published reports that allow an estimate of VO(2max) to be made by measuring a combination of the distance walked as well as lung function in patients with chronic obstructive -pulmonary disease (COPD). AIMS: The aim of the present study was to determine if estimates of VO(2max) and measured VO(2max) based on predictive equations are useful in individuals with COPD. METHODS: Twenty-eight male patients (mean age 68 years) with a mean forced expiratory volume in 1 s of 1.3 L were enrolled in the study after determining that they could perform a maximal exercise study. The estimated VO(2max) using equations reported by Chuang et al. and Cahalin et al. was cross-validated with the measured VO(2max) determined during cardiopulmonary exercise testing. RESULTS: The mean estimated VO(2max) using the pre-diction equation did not differ from the mean measured VO(2max) (1.13 vs 1.18 L/min, respectively; P = 0.25). However, the scattered relationship between the measured and the estimated VO(2max) did not support the use of this equation to predict an individual's performance. The prediction equations currently available in published reports significantly underestimate the measured VO(2max) (P < 0.05-10(-12)). CONCLUSIONS: It is recommended that VO(2max) is measured rather than estimated using the prediction equations when a VO(2max) measurement is used for clinical decision-making.     

Altered cellular distribution of iron in rat cerebral cortex during the oestrous cycle

Summary. Iron levels in blood, liver and the substantia nigra fluctuate during the oestrous cycle but it is not known whether the cellular distribution also changes. This study shows that during dioestrus, when serum levels of oestradiol are low, the amount of histochemically detectable iron in the cerebral cortex is significantly lower than in proestrus when oestradiol levels are highest. During dioestrus iron is concentrated within neurones, and the transition to proestrus is associated with a shift in iron localisation from neurones to vascular endothelial cells and oligodendrocytes. These data raise the possibility that changes in the concentration of serum oestrogen (or other reproductive hormones) during the oestrous cycle can influence the intercellular transport of iron in the brain.Current address: Neuroplasticity Department, European Neuroscience Institute, Göttingen, Germany