Electrical stability of erythrocytes in the presence of divalent cations

Erythrocytes suspended in a medium of low ionic strength lyse under the effect of an exponential electrical pulse. The percentage of haemolysed cells decreases several-fold in the presence of divalent cations. The protective action of the ions studied increases in the following order: Ca⁺⁺, Mg⁺⁺, Zn⁺⁺. It is assumed that divalent ions bind to the negative charges of the lipid and protein molecules and reduce their electrostatic repulsion, which results in stabilization of the membranes.     

The influence of a magnetic field on chromosome sets and cell division]

An effect of stable magnetic field on karyotype and cell division of human lymphocytes from peripheral blood was studied in tissue culture. Comparative investigations were carried out with lymphocytes, whoch were treated with magnetic field of different tension (0,179; 0,391 and 0,600 Ts) and continuance (30 sec, 30 and 60 min). Lymphocytes, treated with stable magnetic field, were suspended in donor plasma and immediately cultivated after the treatment. The stable magnetic field was found to have a distinct mutagenic effect on cultivated lymphocytes of human peripheral blood. The increase in structural impairements of chromosomes correlated with tension of magnetic field and continuance of its effect. In structural impairements of chromosomes the chromatid gaps and breaks were more often observed but chromosome ruptures and pericentric clearances occurred more rarely. The effect of stable magnetic field on lymphocytes proliferation was studie. Under weak and transient using of magnetic field the proliferation was stimulated, but in rigid conditions--mitosis and blastic transformation were decreased. Continuous effect of magnetic field with weak tension did not inhibite the blastic transformation but influenced on the survival rate of cells in tissue culture, enhancing their lethality.     

Characterization and potential antitumor effect of a heteropolysaccharide produced by the red alga Porphyridium sordidum

Taking into account the rising trend of the incidence of cancers of various organs, effective therapies are urgently needed to control human malignancies. However, almost all chemotherapy drugs currently on the market cause serious side effects. Fortunately, several studies have shown that some non‐toxic biological macromolecules, including algal polysaccharides, possess anti‐cancer activities or can increase the efficacy of conventional chemotherapy drugs. Polysaccharides are characteristic secondary metabolites of many algae. The efficacy of polysaccharides on the normal and cancer cells is not well investigated, but our investigations proved a cell specific effect of a newly isolated extracellular polysaccharide from the red microalga Porphyridium sordidum. The investigated substance was composed of xylose:glucose and galactose:manose:rhamnose in a molar ratio of 1:0.52:0.44:0.31. Reversible electroporation has been exploited to increase the transport through the plasma membrane into the tested breast cancer tumor cells MCF‐7 and MDA‐MB231. Application of 75 µg/mL polysaccharide in combination with 200 V/cm electroporation induced 40% decrease in viability of MDA‐MB231 cells and changes in cell morphology while control cells (MCF10A) remained with normal morphology and kept vitality.     

Bystander suppression of collagen-induced arthritis in mice fed ovalbumin

We wanted to assess whether B-cell and/or T-cell responses to collagen and thereby the course of collagen-induced arthritis could be suppressed by regulatory mechanisms associated with oral tolerance to an unrelated protein. DBA/1 mice were fed ovalbumin (OVA)-containing pellets ad libitum for 1 week and subsequently coimmunized twice, with a mixture of bovine collagen type II (BCII) and OVA in Freund's complete adjuvant. Mice fed OVA before coimmunization with BCII and OVA had significantly lower arthritic scores than mice immunized with BCII only. Their body weight increased during the study period and their anti-BCII antibody activity was significantly IgG_2a lower. The frequency of spleen cells producing IgG anti-BCII antibody was also reduced. Coimmunization per se slightly ameliorated the development of arthritis, resulting in an early, transient reduction. It resulted in significantly higher IgG₁ anti-BCII antibody activity and increased splenocyte secretion of IFN-γ and IL-10 in response to BCII. Our findings demonstrate that OVA-specific regulatory events induced by feeding OVA, i.e. bystander suppression, reduced the severity of arthritis in animals immunized with BCII and OVA. Anti-BCII specific antibody responses and cytokine secretion by types 1 and 2 T helper cells were also decreased.     

Induction of apoptosis by electrotransfer of positively charged proteins as Cytochrome C and Histone H1 into cells

Cytochrome C (Cyt. C) is a mitochondrial protein inducing apoptosis when it is accumulated in the cytosol by a currently unknown mechanism, but regulated by the bcl-2 family of proteins. The linker Histone H1 is another basic protein with highly conservative structure, composition, and equal molecular weight, not changed during the evolution. An attempt was made to understand better the apoptotic processes by electroloading of leukemic cells, such as K562, HL-60, and SKW3, and human lymphocytes with positively charged proteins, such as Cyt. C, Histone H1, and methylated BSA albumin (mBSA). The triggering apoptotic processes followed by MTT test, FACS analysis, and DNA fragmentation after the electrotransfer of these proteins into the cells were observed. Histone H1 and mBSA induce the release of Cyt. C from rat liver mitochondria. Cytochrome C release was higher when mitochondria were in "high-energy" state. It is supposed that release of Cyt. C from mitochondria is due to the mechanical rupture of the outer mitochondrial membrane, rich in negatively charged groups, predominately due to cardiolipin. The reason for the morphological rupture of the outer mitochondial membrane could be the rigidification and segregation of the membrane and the destroyed membrane asymmetries of both monolayers in the presence of positively charged proteins at higher linear charges such as Histone H1. We suggested that Histone H1, at a given moment of activated signaling for apoptosis, could be not transported to the nucleus and could lead to the release of Cyt. C from the mitochondria in the cytoplasm. It is temping to speculate that Histone H1 has other physiological extranuclear functions involved in apoptosis.     

An assay for dielectrophoresis: Applications to electromagnetically induced membrane adhesion and fusion

A new assay for dielectrophoresis, consisting of a platinum wire placed along the axis of symmetry of a hollow metal cylinder, was constructed. The main advantage of this device is that because of the simple axisymmetrical electric field the experimental data can be easily interpreted. Preliminary experimental data for adhesion and fusion of red blood cells are reported. The most interesting observation was that fusion of cell membranes can be induced without applying DC electrical pulses, merely because of the dielectrophoretic effect. A possible explanation of this phenomenon is suggested.     

B30.2-like domain proteins: update and new insights into a rapidly expanding family of proteins

The B30.2 domain is a conserved region of around 170 amino acids associated with several different protein domains, including the immunoglobulin folds of butyrophilin and the RING finger domain of ret finger protein. We recently reported several novel members of this family as well as previously undescribed protein families possessing the B30.2 domain. Many proteins have subsequently been found to possess this domain, including pyrin/marenostrin and the midline 1 (MID1) protein. Mutations in the B30.2 domain of pyrin/marenostrin are implicated in familial Mediterranean fever, and partial loss of the B30.2 domain of MID1 is responsible for Opitz G/BBB syndrome, characterized by developmental midline defects. In this study, we scrutinized the available sequence data bases for the identification of novel B30.2 domain proteins using highly sensitive database-searching tools. In addition, we discuss the chromosomal localization of genes in the B30.2 family, since the encoded proteins are likely to be involved in other forms of periodic fever, autoimmune, and genetic diseases.      

Mechanism of Formation of Novel Covalent Drug·DNA Interstrand Cross-Links and Monoadducts by Enediyne Antitumor Antibiotics

The potent enediyne antitumor antibiotic C1027 has been previously reported to induce novel DNA interstrand cross-links and drug monoadducts under anaerobic conditions [Xu et al. (1997) J. Am. Chem. Soc. 119, 1133-1134]. In the present study, we explored the mechanism of formation of these anaerobic DNA lesions. We found that, similar to the aerobic reaction, the diradical species of the activated drug initiates anaerobic DNA damage by abstracting hydrogen atoms from the C4', C1', and C5' positions of the A1, A2, and A3 nucleotides, respectively, in the most preferred 5'GTTA1T/5'ATA2A3C binding sequence. It is proposed that the newly generated deoxyribosyl radicals, which cannot undergo oxidation, likely add back onto the nearby unsaturated ring system of the postactivated enediyne core, inducing the formation of interstrand cross-links, connecting either A1 to A2 or A1 to A3, or drug monoadducts mainly on A2 or A3. Comparative studies with other enediynes, such as neocarzinostatin and calicheamicin gamma1I under similar reaction conditions indicate that the anaerobic reaction process is a kinetically competitive one, depending on the proximity of the drug unsaturated ring system or dioxygen to the sugar radicals and their quenching by other hydrogen sources such as solvent or thiols. It was found that C1027 mainly generates interstrand cross-links, whereas most of the anaerobic lesions produced by neocarzinostatin are drug monoadducts. Calicheamicin gamma1I was found to be less efficient in producing both lesions. The anaerobic DNA lesions induced by enediyne antitumor antibiotics may have important implications for their potent cytotoxicity in the central regions of large tumors, where relative anaerobic conditions prevail.     

Mechanistic aspects of biosynthesis of silver nanoparticles by several <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> strains

Extracellular production of metal nanoparticles by several strains of the fungus Fusarium oxysporum was carried out. It was found that aqueous silver ions when exposed to several Fusarium oxysporum strains are reduced in solution, thereby leading to the formation of silver hydrosol. The silver nanoparticles were in the range of 20–50 nm in dimensions. The reduction of the metal ions occurs by a nitrate-dependent reductase and a shuttle quinone extracellular process. The potentialities of this nanotechnological design based in fugal biosynthesis of nanoparticles for several technical applications are important, including their high potential as antibacterial material.