Suppression of Smad-1 mRNA expression level by Smad-2 likely control dichotomy of NF-κB and Smads mediated activation.

The aim of this study was to find out how NF-κB and Smad-mediated signaling influenced the expression of astrogliogenic versus neurogenic markers of brain development in U4C cells which were either enriched (Tg Jak-1) or deprived in Jak-1 molecule (Jak-1 KO). Genetically modified U4C cells were transfected with NF-kB reporter plasmid in order to follow its activation when cells were cotransfected with different combinations of Smads constructs. In wild type cells no significant activation of NF-κB was observed while genetically modified cells exhibited somewhat different pattern of NF-κB activation depending on the Smad constructs combination used. The absence of NF-κB activation in Jak-1 transgenic cells transfected with Smad-1 plus Smad-3 was accompanied by the appearance of apoptotic cells as revealed by DAPI staining. Smad-1 expression was undetectable in Jak-1 transgenic cells and was downregulated in wild type cells upon transfection with Smad-2. The absence of p65 nuclear translocation in Smad-2 transfected cells and the presence of Smad-4 in nucleus of the same cells indicates dichotomy in NF-κB and Smads mediated signaling pathways. The significance of this study is that helps to elucidate the point of collaboration among three different signaling pathways - Jak-1 mediated cytokine signaling, NF-κB and Smads mediated pathways.

Alternative splicing of p53 and p73: the novel p53 splice variant p53delta is an independent prognostic marker in ovarian cancer.

Similar to p73, the tumor suppressor gene p53 is subject to alternative splicing. Besides p53DeltaE6 and p53beta, we identified p53zeta, p53delta and p53varepsilon, arising from alternative splicing of exon 6 and intron 9, respectively. p53 splice variants were present in 18 of 34 ovarian cancer cell lines (52.9%) and 134 of 245 primary ovarian cancers (54.7%). p53delta expression was associated with impaired response to primary platinum-based chemotherapy (P=0.032). Also, p53delta expression constituted an independent prognostic marker for recurrence-free and overall survival (hazard ratio 1.854, 95% confidence interval 1.121-3.065, P=0.016; and hazard ratio 1.937, 95% confidence interval 1.177-3.186, P=0.009, respectively). p53beta expression was associated with adverse clinicopathologic markers, that is, serous and poorly differentiated cancers (P=0.002 and P=0.008, respectively), and correlated with worse recurrence-free survival in patients exhibiting functionally active p53 (P=0.049). DeltaN'p73 constituted the main N-terminally truncated p73 isoform and was preferentially found in ovarian cancer cell lines showing functionally active p53, supporting our hypothesis that N-terminally truncated p73 isoforms can alleviate the selection pressure for p53 mutations by the inhibition of p53 protein function.

The effect of aprotinin on risk of acute renal failure requiring dialysis after on-pump cardiac surgery.

The use of aprotinin in cardiac surgery to reduce perioperative bleeding and transfusion is controversial. We assessed the effect of aprotinin on the risk of acute renal failure in 423 patients who underwent on-pump cardiac surgery between January 1, 2005 and December 31, 2006. Of these 423 patients, 318 (75.2%) received aprotinin (median dose=3.0 million KIU, standard deviation=2.8 million KIU; interquartile range: 2 million KIU to 4 million KIU). Aprotinin was more likely to be used in patients who did not cease aspirin before surgery, in urgent or emergency surgery, who had impaired left ventricular function, a longer period of bypass and aortic cross-clamp time, and with both coronary artery bypass graft and valvular surgery performed. The overall incidence of acute renal failure requiring dialysis was 2.8%. The use of aprotinin was not associated with a reduction in transfusion nor an increased risk of renal failure requiring dialysis, atrial fibrillation, cerebrovascular accident or mortality in the univarate analyses. In the multivariate analysis, only preoperative serum creatinine concentration (odds ratio [OR] 1.06 per 10 micromol/l increment in creatinine, 95% confidence interval [CI]: 1.01 to 1.14, P=0.029) and urgency of the surgery (urgent vs. scheduled surgery: OR 12.8, CI: 2.3 to 70.8, P=0.004; emergency vs. scheduled surgery: OR 23.1, CI: 3.0 to 180.2, P=0.003) were significantly associated with an increased risk of acute renal failure requiring dialysis. The use of low-dose aprotinin did not significantly reduce perioperative transfusion requirements and was not a significant risk factor for acute renal failure requiring dialysis in our patients.

Effects of habitat destruction and resource supplementation in a predator-prey metapopulation model.

We developed a mean field, metapopulation model to study the consequences of habitat destruction on a predator-prey interaction. The model complements and extends earlier work published by Bascompte and Solé (1998, J. theor. Biol.195, 383-393) in that it also permits use of alternative prey (i.e., resource supplementation) by predators. The current model is stable whenever coexistence occurs, whereas the earlier model is not stable over the entire domain of coexistence. More importantly, the current model permits an assessment of the effect of a generalist predator on the trophic interaction. Habitat destruction negatively affects the equilibrium fraction of patches occupied by predators, but the effect is most pronounced for specialists. The effect of habitat destruction on prey coexisting with predators is dependent on the ratio of extinction risk due to predation and prey colonization rate. When this ratio is less than unity, equilibrial prey occupancy of patches declines as habitat destruction increases. When the ratio exceeds one, equilibrial prey occupancy increases even as habitat destruction increases; i.e., prey "escape" from predation is facilitated by habitat loss. Resource supplementation reduces the threshold colonization rate of predators necessary for their regional persistence, and the benefit derived from resource supplementation increases in a nonlinear fashion as habitat destruction increases. We also compared the analytical results to those from a stochastic, spatially explicit simulation model. The simulation model was a discrete time analog of our analytical model, with one exception. Colonization was restricted locally in the simulation, whereas colonization was a global process in the analytical model. After correcting for differences between nominal and effective colonization rates, most of the main conclusions of the two types of models were similar. Some important differences did emerge, however, and we discuss these in relation to the need to develop fully spatially explicit analytical models. Finally, we comment on the implications of our results for community structure and for the conservation of prey species interacting with generalist predators.

The efficacy of retroviral herpes simplex virus thymidine kinase gene transfer and ganciclovir treatment on the inhibition of melanoma growth in vitro and in vivo.

One approach to gene therapy of cancer is based on the insertion of a suicide gene into tumor cells and subsequent activation of the suicide mechanism. We used the herpes simplex virus thymidine kinase (HSVtk) gene followed by ganciclovir (GCV) treatment. The goal of our experiments was to determine the effectiveness of HSVtk gene therapy in malignant melanoma. B16BL6 murine melanoma cells retrovirally transduced with the HSVtk gene became sensitive to low concentrations of GCV. Analysis by RT-PCR showed HSVtk expression in transduced B16BL6tk+ cells. Apoptotic cell death was found in B16BL6tk+ cells treated with GCV (20 microM). The sensitivity of B16BL6tk+ cells to GCV was also examined in vivo. Tumors inoculated subcutaneously into C57BL6 mice regressed rapidly when treated with GCV (50 mg/kg twice a day) and disappeared completely after 14 days treatment. The mice remained in remission for 5 months. A bystander effect through which nontransduced B16BL6 cells were also inhibited by GCV administration when cocultured with B16BL6tk+ cells was expected. However, only slight killing of nontransduced cells was observed in vitro. Analysis of the bystander effect in vivo showed complete regression of tumors inoculated with a mixture of cells mostly consisting of B16BL6tk+ cells. A distant bystander effect was also examined. There was no regression of wild-type tumors raised at a distant site from primary B16BL6tk+ tumors. The failure of a more effective bystander effect indicates the need for further investigation of the possible use of combined gene therapy to treat melanoma.

Expression of erbB-3 protein in colorectal adenocarcinoma: correlation with poor survival.

BACKGROUND/AIMS: The family of erbB receptors includes four transmembrane glycoproteins with tyrosine kinase activity. These receptors are widely expressed in normal tissues, but they also have been implicated in the development of several human adenocarcinomas. c-erbB-3/HER-3 has been detected to a greater or lesser extent in many tissues from the digestive, urinary, reproductive and respiratory tracts. The overexpression of c-erbB-3/HER-3 protein has also been shown in 53%-88% of colorectal adenocarcinomas. In this study we investigated the expression of the c-erbB-3/ HER-3 gene product in colorectal tumour samples, and compared the results obtained with several clinicopathological parameters, including the survival of patients. METHODS: Paraffin-embedded tissue sections were analysed immunohistochemically, using monoclonal antibody RTJ1 to human erbB-3 protein. Antibody RTJ1 specificity was confirmed by immunoprecipitation followed by Western blotting analysis. Amplification of the erbB-3 oncogene was tested by dot-blot hybridization. RESULTS: Adenocarcinomas of the colon were positive for erbB-3 protein in 78% of samples examined. Dot-blot analysis showed no amplification of the erbB-3 gene in colon adenocarcinomas. Statistical analysis showed that patients with tumours that could not be stained for erbB-3 protein survived significantly longer (P<0.05) than patients with tumours staining positive for the erbB-3 protein. A Cox proportional-hazards model with stepwise variable selection identified age, sex and erbB-3 expression as important prognostic factors. CONCLUSION: These findings demonstrate that erbB-3 protein expression could serve as a prognostic factor in colorectal malignancies.

Effect of extracellular NADH on human tumor cell proliferation.

We investigated the antiproliferative effects of extracellular nicotinamide adenine dinucleotide against human malignant CaCo-2 (colon carcinoma), Hep-2 (laryngeal carcinoma), MCF-7 (breast carcinoma), CaSki (cervix carcinoma) cell lines, as well as against murine fibrosarcoma and normal human embryonal fibroblast (HEF). NADH was very potent in the growth inhibition of murine fibrosarcoma and human Hep-2 cells, regardless of the dose applied. During the observed period (4 or 5 days) only one dose of NADH was sufficient in reducing the growth rate for up to 92%. It had no effect on the growth of other cell lines tested. The identification of DNA-fragmentation and p53 and Ki-67 genes expression suggest that the mechanism of NADH action is different from disregulation of genes considered as check-points in cell cycle.

Legal standards and the significance of DNA evidence.

Most human biologists are aware of controversies regarding the use of DNA profiles in the courtroom. Much attention has been given to estimating the probability of obtaining matches between DNA samples from an innocent suspect and those from a crime scene, but considerably less attention has been given to the critical issue of determining the probability of guilt given a match. Using Bayes' rule and simple algebra, we develop a measure of the strength of DNA evidence that indicates the amount of incriminating evidence needed in combination with DNA match evidence to meet a given conviction standard. Based on current standards and practices, we use this measure to demonstrate that (1) the amount of non-DNA evidence needed to convict, given a DNA match, generally is quite small, even if errors can occur in the processing of DNA evidence; (2) DNA match evidence alone is insufficient to convict, even for the lowest recognized conviction standards; (3) failure to match DNA evidence samples should be exculpatory unless laboratory proficiency is poor; and (4) if errors in handling evidentiary samples occur (even rarely) that tend to produce a false DNA match, then the legal significance of DNA evidence is remarkably insensitive to estimates of chance match probability.

A Bayesian approach to the inference of diploid genotypes using haploid genotypes.

Morris and Spieth (1978) described a method of calculating unbiased estimates of diploid genotype frequencies given information on the genotypes of haploid cells derived from diploid individuals. They concluded that three haploids per diploid would minimize sampling variance of genotype frequencies, given a fixed total number of haploids examined. If the identity of individual diploid genotypes is needed, Morris and Spieth (1978) stated that more haploids should be collected per diploid. We extend this work by showing from a Bayesian perspective that the probability of misclassification of individuals depends not only on the number of haploids sampled, but also on the genetic structure of the population since misclassification error will increase as the frequency of heterozygotes increases. Since information on the genetic structure (allele frequencies, inbreeding coefficient) of a population is rarely known prior to the initiation of an empirical study, the usefulness of our Bayesian approach is in experimental design, by revealing the magnitude of possible misclassification errors given a particular choice of number of haploids.

p185neu is expressed in yolk sac during rat postimplantation development.

We have shown that the neu oncogene product (p185neu) is not present in the rat embryo before organogenesis. However, coincident with the onset of organogenesis, p185neu was detected in neural and connective tissue as well as in the secretory epithelium as was described by Kokai et al. (1987). In addition, p185neu is also expressed in the rat visceral yolk sac (VYS) endodermal cells but not in the mesenchymal and mesothelial layers of the same structure nor in the amnion. The first detectable sign of p185neu expression in VYS was found at d 11 of gestation and the levels of protein increased towards the end of pregnancy. In the yolk sac carcinoma (YSC), which is considered to be the malignant counterpart of the rat yolk sac, p185neu was observed only within columnar epithelial cells (the visceral component of the neoplasm) while parietal endoderm-like cells were devoid of detectable protein. From d 9 of pregnancy up to delivery some of the trophoblastic giant cells also showed a faint to moderate immunoreactivity. Results are presented which would indicate a possible role of p185neu in rat embryogenesis.

Collagenase derived from human fibrosarcoma is responsible for degradation of basement membranes.

A collagenase-like enzyme with the ability to degrade the proteins of artificial basement membranes (BM) was isolated from human fibrosarcoma. Secretion of the same peptide was observed from the primary fibrosarcoma cell cultures. This peptide degrades the artificial basement membranes derived from bovine corneal endothelial cells. Using electrophoretic methods it was found that the isolated and partially purified enzyme consists of eight bands of different molecular mass corresponding to the collagenase standard from Cl. histolyticum. Only two bands with molecular masses of 22,000 (pI 5.5) and 63,000 (pI 5.9) degrade BM.

A new procedure for the cryopreservation of equine embryos.

Early equine blastocysts and blastocysts were collected nonsurgically at six days post-ovulation. Thirty-two embryos were randomly assigned to a 2x2 factorial design. Factors were: 1) 0.5-ml straws or 1-ml glass ampules; and 2) plunging into liquid nitrogen (IN(2)) at -33 C or -38 C. Cryoprotectant, 10% glycerol in PBS plus 5% fetal calf serum (FCS) was added in two steps, 5% then 10%. Embryos were cooled at 4 C/min to -6 C and then seeded, 0.3 C/min to -30 or -35 C and 0.1 C/min to -33 or -38 C. Samples were thawed in 37 C water and glycerol removed in six steps, 10 min per step. Embryo quality and stage of development were evaluated prior to freezing, immediately post-thaw and after 24 h culture in Ham's F10 with 5% FCS. The mean post-thaw quality of embryos plunged at -33 C was superior (P<0.05) to that of embryos plunged at -38 C (2.0 vs 2.9). Embryos frozen in ampules and plunged at -38 C were of poorer quality (P<0.05) than those frozen in ampules and plunged at -33 C or frozen in straws and plunged at -33 C. After 24 h of culture, more embryos developed if frozen in straws compared to ampules, and plunging at -33 C resulted in higher quality embryos than plunging at -38 C. In Experiment 2, 23 embryos were packaged in straws and plunged at -33 C as described in Experiment 1. Six of the 23 surgically transferred frozen embryos were degenerate at thawing and the remaining 17 surgically transferred were via flank incision. Pregnancy rate at 50 days post-ovulation was 53% (nine of 17). Early blastocysts resulted in a higher (P<0.05) pregnancy rate (8 10 , 80%) than expanded blastocysts (1 7 , 14%).

Localized amyloid of the ureter--case report and review of the literature.

An unusual case of localized amyloidosis of the ureter is described and a review of the 12 previously reported cases is presented. No case had been diagnosed prior to surgery. Loin pain, with or without haematuria, associated with a sterile pyuria and an abnormal pyelogram were the commonest clinical signs. In the case reported an incidental small renal cell carcinoma was also present and the association of amyloidosis and neoplasia is discussed.

Fractionation and turnover of stable carbon isotopes in animal tissues: Implications for δ13C analysis of diet.

The use of stable carbon isotopes as a means of studying energy flow is increasing in ecology and paleoecology. However, secondary fractionation and turnover of stable isotopes in animals are poorly understood processes. This study shows that tissues of the gerbil (Meriones unguienlatus) have different δ13C values when equilibrated on corn (C4) or wheat (C3) diets with constant 13C/12C contents. Lipids were depleted 3.0‰ and hair was enriched 1.0‰ relative to the C4 diet. Tissue δ13C values were ranked hair>brain>muscle>liver>fat. After changing the gerbils to a wheat (C3) diet, isotope ratios of the tissues shifted in the direction of the δ13C value of the new diet. The rate at which carbon derived from the corn diet was replaced by carbon derived from the wheat diet was adequately described by a negative exponential decay model for all tissues examined. More metabolically active tissues such as liver and fat had more rapid turnover rates than less metabolically active tissues such as hair. The half-life for carbon ranged from 6.4 days in liver to 47.5 days in hair.The results of this study have important implications for the use of δ13C values as indicators of animal diet. Both fractionation and turnover of stable carbon isotopes in animal tissues may obscure the relative contributions of isotopically distinct dietary components (such as C3 vs. C4, or marine vs. terrestrial) if an animal's diet varies through time. These complications deserve attention in any study using stable isotope ratios of animal tissue as dietary indicators and might be minimized by analysis of several tissues or products covering a range of turnover times.

Arterial embolisation in renal carcinoma: a useful procedure?

Twenty-eight patients underwent embolisation over a 2-year period. III effects followed embolisation in 16 (57%), 2 of whom died. Attention is drawn to the intolerance of many patients to the length of the procedure. Of 24 patients undergoing operation, one case was inoperable and difficulty was encountered in 9. No perirenal plane of oedema was identified. No specimen examined histologically showed complete infarction. The tumour tissue was more resistant to infarction than normal kidney and 8 tumours (35%) appeared to have escaped completely. It is concluded that the role of embolisation is limited and that a combination of methods for achieving adequate arterial occlusion must be used for the optimum effect.