Homocysteine induces DNA synthesis and proliferation of vascular smooth muscle cells by interfering with MAPK kinase pathway

Hyperhomocysteinemia has been identified as an important and independent risk factor for cerebral, coronary and peripheral atherosclerosis. However the mechanisms by which homocysteine promote atherosclerotic plaque formation are not clearly defined. Earlier reports have suggested that homocysteine exert its effect via the H2O2 produced during its metabolism. To evaluate which signalling molecules are involved in homocysteine induced atherosclerotic changes during the pathogenesis of vascular diseases, we examined homocysteine induced smooth muscle cell proliferation in the presence of different signal transduction inhibitors. We show that MAPK kinase pathway is involved in homocysteine induced DNA synthesis and proliferation of vascular smooth muscle cells in the presence of the peroxide scavenging enzyme, catalase. Our data suggest that homocysteine induces smooth muscle cell growth through a pathway that is independent of H2O2, that involves MAPK kinase activation, and that results in accelerated atherosclerosis.     

Surgical resection for gastrointestinal stromal tumors (GIST): experience on 25 patients

Background

The benefit of pelvic lymphadenectomy in patients with cancer of the urinary bladder remains controversial. Though the inclusion of lymph node dissection in conjunction with radical cystectomy for patients with clinically negative nodes is well accepted, however, the extent of the nodal dissection remains contentious, particularly in patients with gross disease and T₁G₃ cancer. The extent of the primary bladder tumor, number of lymph nodes removed and the lymph node tumor burden are important prognostic variables in patients undergoing cystectomy. We analyzed the impact of the extent of lymphadenectomy during radical cystectomy on survival in the contemporary literature.

Methods

A Pubmed search was carried out for the literature published over the last 15 years using bladder cancer, radical cystectomy, survival, lymphadenectomy and complications as the key words. We have discussed the extent of lymphadenectomy on survival and its anatomical basis to determine the optimal number of lymph nodes to be removed and the concept of node density.

Results

Evidence from contemporary literature indicate significantly increased survival rates after cystectomy in patients with bladder cancer diagnosed with stages III or IV disease who have had relatively more lymph nodes examined, suggesting that even some patients with higher stage disease may benefit from extended pelvic lymphadenectomy at the time of cystectomy. Studies also indicate that more extensive lymphadenectomy significantly improved the prognosis of patients with bladder cancer, not only by providing prognostic information but perhaps it is also due to its inherent therapeutic value.

Conclusion

Extended lymph node dissection improves local control and survival. However, in the absence of controlled randomized trial this remains a dubitable issue.     

Calcium dysregulation and membrane disruption as a ubiquitous neurotoxic mechanism of soluble amyloid oligomers

Increasing evidence suggests that amyloid peptides associated with a variety of degenerative diseases induce neurotoxicity in their intermediate oligomeric state, rather than as monomers or fibrils. To test this hypothesis and investigate the possible involvement of Ca2+ signaling disruptions in amyloid-induced cytotoxicity, we made homogeneous preparations of disease-related amyloids (Abeta, prion, islet amyloid polypeptide, polyglutamine, and lysozyme) in various aggregation states and tested their actions on fluo-3-loaded SH-SY5Y cells. Application of oligomeric forms of all amyloids tested (0.6-6 microg ml-1) rapidly (approximately 5 s) elevated intracellular Ca2+, whereas equivalent amounts of monomers and fibrils did not. Ca2+ signals evoked by Abeta42 oligomers persisted after depletion of intracellular Ca2+ stores, and small signals remained in Ca2+-free medium, indicating contributions from both extracellular and intracellular Ca2+ sources. The increased membrane permeability to Ca2+ cannot be attributed to activation of endogenous Ca2+ channels, because responses were unaffected by the potent Ca2+-channel blocker cobalt (20 microm). Instead, observations that Abeta42 and other oligomers caused rapid cellular leakage of anionic fluorescent dyes point to a generalized increase in membrane permeability. The resulting unregulated flux of ions and molecules may provide a common mechanism for oligomer-mediated toxicity in many amyloidogenic diseases, with dysregulation of Ca2+ ions playing a crucial role because of their strong trans-membrane concentration gradient and involvement in cell dysfunction and death.     

Red wine polyphenols influence carcinogenesis, intestinal microflora, oxidative damage and gene expression profiles of colonic mucosa in F344 rats

Polyphenols from tea and other beverages such as red wine have been regarded with interest as possible chemopreventive agents against cancer. Here we report that red wine polyphenols (50 mg/kg) administered with the diet to F344 rats for 16 weeks inhibited colon carcinogenesis induced by azoxymethane (AOM, 7.4 mg/kg, total dose 74 mg/kg) or dimethylhydrazine (DMH, 30 mg/kg, total dose, 300 mg/kg). Polyphenol-treated animals had a consistently lower tumour yield compared to controls. In polyphenol-treated rats, the main bacterial strains in the faeces at sacrifice were Bacteroides, Lactobacillus and Bifidobacterium spp., whereas microorganisms predominantly identified in control-fed rats were Bacteroides, Clostridium and Propionibacterium spp. Wine polyphenols (57 mg/kg for 10 days, by gavage), administered to rats not treated with carcinogens, produced a significant decrease in the basal level of DNA oxidative damage of the colon mucosa as measured with the comet assay (average pyrimidine oxidation was reduced by 62% and purine oxidation by 57%, p<0.05). To further explore the molecular effects of wine polyphenols we used the microarray technology to study gene expression profiles: rats were treated with 50 mg/kg wine polyphenols for 14 days, mixed in the diet. Global expression analysis of 5707 genes revealed an extensive down-regulation of genes involved in a wide range of physiological functions, such as metabolism, transport, signal transduction and intercellular signalling. By analysing metabolic pathways with the GenMAPP software program we observed that two major regulatory pathways were down-regulated in the colon mucosa of polyphenols-treated rats: inflammatory response and steroid metabolism. We also found a down-regulation of many genes regulating cell surface antigens, metabolic enzymes and cellular response to oxidative stress. In conclusion, reduction of oxidative damage, modulation of colonic flora and variation in gene expression may all concur in the modulation of intestinal function and carcinogenesis by wine polyphenols.     

A microsatellite linkage map of the European sea bass Dicentrarchus labrax L

A genetic linkage map of the European sea bass (Dicentrarchus labrax) was constructed from 174 microsatellite markers, including 145 new markers reported in this study. The mapping panel was derived from farmed sea bass from the North Adriatic Sea and consisted of a single family including both parents and 50 full-sib progeny (biparental diploids). A total of 162 microsatellites were mapped in 25 linkage groups. Eleven loci represent type I (coding) markers; 2 loci are located within the peptide Y (linkage group 1) and cytochrome P450 aromatase (linkage group 6) genes. The sex-averaged map spans 814.5 cM of the sea bass genome. The female map covers 905.9 cM, whereas the male map covers only 567.4 cM. The constructed map represents the first linkage map of European sea bass, one of the most important aquaculture species in Europe.     

The role of pH dynamics and the Na+/H+ antiporter in the etiopathogenesis and treatment of cancer. Two faces of the same coin--one single nature

Looked at from the genetic point-of-view cancer represents a daunting and, frankly, confusing multiplicity of diseases (at least 100) that require an equally large variety of therapeutic strategies and substances designed to treat the particular tumor. However, when analyzed phenotypically cancer is a relatively uniform disease of very conserved 'hallmark' behaviors across the entire spectrum of tissue and genetic differences [D. Hanahan, R.A. Weinberg, Hallmarks of cancer, Cell 100 (2000) 57-70]. This suggests that cancers do, indeed, share common biochemical and physiological characteristics that are independent of the varied genetic backgrounds, and that there may be a common mechanism underlying both the neoplastic transformation/progression side and the antineoplastic/therapy side of oncology. The challenge of modern oncology is to integrate all the diverse experimental data to create a physiological/metabolic/energetic paradigm that can unite our thinking in order to understand how both neoplastic progression and therapies function. This reductionist view gives the hope that, as in chemistry and physics, it will possible to identify common underlying driving forces that define a tumor and will permit, for the first time, the actual calculated manipulation of their state. That is, a rational therapeutic design. In the present review, we present evidence, obtained from a great number of studies, for a fundamental, underlying mechanism involved in the initiation and evolution of the neoplastic process. There is an ever growing body of evidence that all the important neoplastic phenotypes are driven by an alkalization of the transformed cell, a process which seems specific for transformed cells since the same alkalinization has no effect in cells that have not been transformed. Seen in that light, different fields of cancer research, from etiopathogenesis, cancer cell metabolism and neovascularization, to multiple drug resistance (MDR), selective apoptosis, modern cancer chemotherapy and the spontaneous regression of cancer (SRC) all appear to have in common a pivotal characteristic, the aberrant regulation of hydrogen ion dynamics [S. Harguindey, J.L. Pedraz, R. García Ca?ero, J. Pérez de Diego, E.J. Cragoe Jr., Hydrogen ion-dependent oncogenesis and parallel new avenues to cancer prevention and treatment using a H+-mediated unifying approach: pH-related and pH-unrelated mechanisms, Crit. Rev. Oncog. 6 (1) (1995) 1-33]. Cancer cells have an acid-base disturbance that is completely different than observed in normal tissues and that increases in correspondence with increasing neoplastic state: an interstitial acid microenvironment linked to an intracellular alkalosis.     

On the mechanism of Rhodotorula gracilis D-amino acid oxidase: role of the active site serine 335

Serine 335 at the active site of D-amino acid oxidase from the yeast Rhodotorula gracilis (RgDAAO) is not conserved in other DAAO sequences. To assess its role in catalysis, it was mutated to Gly, the residue present in mammalian DAAO, an enzyme with a 35-fold lower turnover number with D-alanine. The spectral and ligand binding properties of the S335G mutant are similar to those of wild-type enzyme, suggesting an active site with minimally altered electrostatic properties. The S335G mutant is catalytically active, excluding an essential role of S335 in catalysis. However, S335-OH contributes to the high efficiency of the mutant enzyme since the catalytic activity of the latter is lower due to a decreased rate of flavin reduction relative to wild-type RgDAAO. Catalytic rates are pH-dependent and appear to converge to very low, but finite and similar values at low pH for both wild-type and S335G RgDAAO. While this dependence exhibits two apparent pKs with wild-type RgDAAO, with the S335G mutant a single, apparent pK approximately 8 is observed, which is attributed to the ionization of the alphaNH2 group of the bound substrate. Removal of S335-OH thus suppresses an apparent pK approximately 6. Both wild-type RgDAAO and the S335G mutant exhibit a substantial deuterium solvent kinetic isotope effect (> or =4) at pH<7 that disappears with increasing pH and reflects a pKapp=6.9 +/- 0.4. Interestingly, the substitution suppresses the activity towards d-lactate, suggesting a role of the serine 335 in removal of the substrate alpha-OH hydrogen.