Changes in electric charge and phospholipids composition in human colorectal cancer cells

Cancer cells perform their malicious activities through own cell membranes that screen and transmit inhibitory and stimulatory signals out of the cells and into them. This work is focused on changes of phospholipids content (PI—phosphatidylinositol, PS—phosphatidylserine, PE—phosphatidylethanolamine, PC—phosphatidylcholine) and electric charge that occur in cell membranes of colorectal cancer of pT3 stage, various grades (G2, G3) and without/with metastasis. Qualitative and quantitative composition of phospholipids in the membrane was determined by HPLC (high-performance liquid chromatography). The surface charge density of colorectal cancer cell membranes was measured using electrophoresis. The measurements were carried out at various pH of solution. It was shown that the process of cancer transformation was accompanied by an increase in total amount of phospholipids as well as an increase in total positive charge at low pH and total negative charge at high pH. A malignant neoplasm cells with metastases are characterized by a higher PC/PE ratio than malignant neoplasm cells without metastases. (Mol Cell Biochem 276: 113–119, 2005)     

Efficient use of synchrotron radiation for macromolecular diffraction data collection

In recent years, number of X-ray synchrotron beam lines dedicated to collecting diffraction data from macromolecular crystals has exceeded 50. Indeed, today most protein and nucleic acid crystal structures are solved and refined based on the synchrotron data. Collecting diffraction data on a synchrotron beam line involves many technical points, but it is not a mere technicality. Even though the available hardware and software have become more advanced and user-friendly, it is always beneficial if the experimenter is aware of the problems involved in the data collection process and can make informed decisions leading to the highest possible quality of the acquired diffraction data. Various factors, important for the success of data collection experiments and their relevance for different kinds of applications, are discussed.     

Selenium level in benign and cancerous prostate

The dietary microelement selenium (Se) has been proposed as a potential chemopreventive agent for prostate cancer. This element is present in various amounts in all tissues. Little information is available on Se level in patients with prostate gland disorders. The levels of Se in prostatic gland of patients with prostate cancer, benign prostate hyperplasia, and healthy controls were examined. The Se level for benign prostate hyperplasia (156±30.6 ng/g) was the same as in the control group (157±26.0 ng/g), but in the gland of prostate cancer patients (182±34.1 ng/g wet weight), the Se level was significantly (p<0.01) higher than in both healthy controls and benign prostate hyperplasia. Thus, the Se level in human healthy controls is lower than in kidney and liver but higher compared with other tissues.     

Antiradical and anti-H2O2 properties of polyphenolic compounds from an aqueous peppermint extract

Polyphenolic compounds such as eriocitrin, luteolin-7-O-rutinoside, diosmin, hesperidin, narirutin, isorhoifolin, rosmarinic and caffeic acids were identified in an aqueous extract (Ex) obtained from peppermint leaves (Menthae x piperitae folium). The content of polyphenols in Ex was as follows: eriocitrin 38%, luteolin-7-O-rutinoside 3.5%, hesperidin 2.9%, diosmin 0.8%, isorhoifolin 0.6%, narirutin 0.3%, rosmarinic acid 3.7% and caffeic acid 0.05%. The strongest antiradical activity (determined as DPPH* scavenging features) was observed for luteolin-7-O-rutinoside, eriocitrin and rosmarinic acid. Caffeic acid and hesperidin revealed a lower antiradical activity while isorhoifolin, narirutin and diosmin showed the lowest activity. The strongest anti-H2O2 activity was observed for eriocitrin, a little lower for rosmarinic acid. The rate of hydrogen peroxide scavenging activity displayed by luteolin-7-O-rutinoside and caffeic acid was lower than that of rosmarinic acids. Hesperidin appeared to be a very weak scavenger of hydrogen peroxide. Almost no anti-H2O2 activity was demonstrated for diosmin, narirutin and isorhoifolin. Among examined flavonoids, the strongest antiradical and anti-H2O2 activity was shown for compounds with two hydroxy groups bound to the Bring in ortho position in relation to each other. Replacement of one hydroxy group in the Bring with a methoxy group or removing one hydroxy group leads to decrease of antiradical and anti-H2O2 activity of flavonoids. Our results suggest that eriocitrin is a powerful peppermint antioxidant and a free radical scavenger.     

Oocyte nucleus controls progression through meiotic maturation

We analyzed progression through the meiotic maturation in oocytes manipulated to replace the prophase oocyte nucleus with the nucleus from a cumulus cell, a pachytene spermatocyte or the pronucleus from a fertilized egg. Removal of the oocyte nucleus led to a significant reduction in histone H1 kinase activity. Replacement of the oocyte nucleus by a pronucleus followed by culture resulted in premature pseudomeiotic division and occasional abnormal cytokinesis; however, histone H1 kinase activity was rescued, microtubules formed a bipolar spindle, and chromosomes were condensed. In addition to the anomalies observed after pronuclear transfer, those after transfer of the nucleus from a cumulus cell or spermatocyte included a dramatically impaired ability to form the bipolar spindle or to condense chromosomes, and histone H1 kinase activity was not rescued. Expression of a cyclin B-YFP in enucleated oocytes receiving the cumulus cell nucleus rescued histone H1 kinase activity, but spindle formation and chromosome condensation remained impaired, indicating a pleiotropic effect of oocyte nucleus removal. However, when the cumulus cell nucleus was first transformed into pronuclei (transfer into a metaphase II oocyte followed by activation), such pronuclei supported maturation after transfer into the oocyte in a manner similar to that of normal pronuclei. These results show that the oocyte nucleus contains specific components required for the control of progression through the meiotic maturation and that some of these components are also present in pronuclei.     

Control of Taenia solium taeniasis/cysticercosis: from research towards implementation

Theoretically, considering the biology of its transmission and reservoirs, global eradication of Taenia solium taeniasis and cysticercosis is feasible. Recently much progress has been made in research on diagnosis, treatment and prevention of human taeniasis and porcine cysticercosis, although more operational research is still needed. In spite of this, global eradication of T. solium infection is still unlikely in the near future. Major obstacles to practical implementation of control measures include low levels of sanitation and health education amongst endemic populations, ineffective health services infrastructure and inadequate socioeconomic development in these areas. The continued public health impact of neurocysticercosis, especially fatalities and epilepsy, force us to identify improved options for control. In order to implement control measures in highly endemic areas the active involvement of medical services in controlling T. solium infection and more effective collaboration between medical and veterinary services is necessary. A switch is suggested from total reliance on meat inspection to active diagnosis and treatment of human taeniasis, protection of pigs against infection, promotion of health education and improved surveillance preparing chemotherapeutic and/or sanitary interventions. This could be implemented in areas where active transmission causes substantial morbidity and mortality provided there is the political will, social support, better financing and an effective organizational framework.     

Expression of cell survival/death genes: Bcl-2 and Bax at the rate of colon cancer prognosis

The rate of tumour growth is dependent on the balance between proliferation and apoptosis at all stages of carcinogenesis. Apoptosis inhibition, in turn, depends partly on the balance between expression of two cell death regulatory genes, Bcl-2 and Bax. Colon cancer has long been associated with disturbances in apoptosis regulation. The aim of our study was to determine the expression levels of Bcl-2 and Bax mRNAs in 1 microg sample of total RNA obtained from normal colon and colon adenocarcinoma. This study was intended to evaluate possible differences in Bcl-2 and Bax mRNA levels at particular stages of colon adenocarcinoma classified according to Duke's system. The apoptotic frequency (represented by Bax mRNA copy number) was inversely proportional to the decrease of Bcl-2 gene expression. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) was performed to confirm apoptosis.     

Circular dichroism and aggregation studies of amyloid beta (11-8) fragment and its variants

Aggregation of Abeta peptides is a seminal event in Alzheimer's disease. Detailed understanding of Abeta assembly would facilitate the targeting and design of fibrillogenesis inhibitors. Here comparative conformational and aggregation studies using CD spectroscopy and thioflavine T fluorescence assay are presented. As a model peptide, the 11-28 fragment of Abeta was used. This model peptide is known to contain the core region responsible for Abeta aggregation. The structural and aggregational behaviour of the peptide was compared with the properties of its variants corresponding to natural, clinically relevant mutants at positions 21-23 (A21G, E22K, E22G, E22Q and D23N). In HFIP (hexafluoro-2-propanol), a strong alpha-helix inducer, the CD spectra revealed an unexpectedly high amount of beta-sheet conformation. The aggregation process of Abeta(11-28) variants provoked by water addition to HFIP was found to be consistent with a model of an alpha-helix-containing intermediate. The aggregation propensity of all Abeta(11-28) variants was also compared and discussed.     

The catalytic domain of Escherichia coli Lon protease has a unique fold and a Ser-Lys dyad in the active site

ATP-dependent Lon protease degrades specific short-lived regulatory proteins as well as defective and abnormal proteins in the cell. The crystal structure of the proteolytic domain (P domain) of the Escherichia coli Lon has been solved by single-wavelength anomalous dispersion and refined at 1.75-A resolution. The P domain was obtained by chymotrypsin digestion of the full-length, proteolytically inactive Lon mutant (S679A) or by expression of a recombinant construct encoding only this domain. The P domain has a unique fold and assembles into hexameric rings that likely mimic the oligomerization state of the holoenzyme. The hexamer is dome-shaped, with the six N termini oriented toward the narrower ring surface, which is thus identified as the interface with the ATPase domain in full-length Lon. The catalytic sites lie in a shallow concavity on the wider distal surface of the hexameric ring and are connected to the proximal surface by a narrow axial channel with a diameter of approximately 18 A. Within the active site, the proximity of Lys(722) to the side chain of the mutated Ala(679) and the absence of other potential catalytic side chains establish that Lon employs a Ser(679)-Lys(722) dyad for catalysis. Alignment of the P domain catalytic pocket with those of several Ser-Lys dyad peptide hydrolases provides a model of substrate binding, suggesting that polypeptides are oriented in the Lon active site to allow nucleophilic attack by the serine hydroxyl on the si-face of the peptide bond.