Acetaldehyde activation of poly(ADP-ribose)polymerase in hepatocytes of mice treated in vivo

The activities of poly(ADP-ribose) polymerase and of DNA polymerases alpha and beta and the level of cytochrome P450 were determined in mouse parenchymal liver cells 5 h after treatment with 0.1, 0.3, 1.0, and 3.0 mumole of acetaldehyde. Injection with 1.0 and 3.0 mumole of acetaldehyde induced an increase in poly(ADP-ribose) polymerase activity and in the P450 level, but had no effect on DNA polymerases. The stimulation of poly(ADP-ribose) polymerase activity can be used as an index of induced DNA damage. The possibility of using this experimental approach with other cells derived from mice treated in vivo with different xenobiotics is discussed.     

Amino acid sequence of k Sci, the Bence Jones protein isolated from a patient with light chain deposition disease

Light chain Sci was isolated from the urine of a patient affected by light chain deposition disease with an apparent exclusive localization to the kidney. Sci protein is an intact light chain: it consists of 214 amino acid residues and has an Mr of 23.65. Its complete primary structure has been determined by sequence analysis of the corresponding tryptic peptides and by partially sequencing the intact protein. Sequence comparison shows that Sci protein is strictly related to the light chains of kIIIa family (88% structural identity) which are usually expressed in autoimmune rheumatoid syndromes. Computer graphics model suggests a perturbation in k Sci three-dimensional structure due to the unusual replacement of residues 53 and 77.     

A novel lantibiotic acting on bacterial cell wall synthesis produced by the uncommon actinomycete Planomonospora sp

Important classes of antibiotics acting on bacterial cell wall biosynthesis, such as beta-lactams and glycopeptides, are used extensively in therapy and are now faced with a challenge because of the progressive spread of resistant pathogens. A discovery program was devised to target novel peptidoglycan biosynthesis inhibitors capable of overcoming these resistance mechanisms. The microbial products were first screened according to their differential activity against Staphylococcus aureus and its L-form. Then, activities insensitive to the addition of a beta-lactamase cocktail or d-alanyl-d-alanine affinity resin were selected. Thirty-five lantibiotics were identified from a library of broth extracts produced by 40,000 uncommon actinomycetes. Five of them showed structural characteristics that did not match with any known microbial metabolite. In this study, we report on the production, structure determination, and biological activity of one of these novel lantibiotics, namely, planosporicin, which is produced by the uncommon actinomycete Planomonospora sp. Planosporicin is a 2194 Da polypeptide originating from 24 proteinogenic amino acids. It contains lanthionine and methyllanthionine amino acids generating five intramolecular thioether bridges. Planosporicin selectively blocks peptidoglycan biosynthesis and causes accumulation of UDP-linked peptidoglycan precursors in growing bacterial cells. On the basis of its mode of action and globular structure, planosporicin can be assigned to the mersacidin (20 amino acids, 1825 Da) and the actagardine (19 amino acids, 1890 Da) subgroup of type B lantibiotics. Considering its spectrum of activity against Gram-positive pathogens of medical importance, including multi-resistant clinical isolates, and its efficacy in vivo, planosporicin represents a potentially new antibiotic to treat emerging pathogens.     

The toxicity of tritium: the effects of tritiated amino-acids on preimplanted mouse embryos

The effects of tritiated amino-acids, arginine, lysine, histidine and aspartic acid on the growth and development of two-cell mouse embryos, cultured in vitro, were investigated. The LD50 for the dibasic amino acids, measured on the third day of growth, ranged from 30 to 130 nCi/ml. This was compared with the DNA precursor, thymidine, for which the LD50 was 80 nCi/ml.     

3D strain map of axially loaded mouse tibia: a numerical analysis validated by experimental measurements

A combined experimental/numerical study was performed to calculate the 3D octahedral shear strain map in a mouse tibia loaded axially. This study is motivated by the fact that the bone remodelling analysis, in this in vivo mouse model should be performed at the zone of highest mechanical stimulus to maximise the measured effects. Accordingly, it is proposed that quantification of bone remodelling should be performed at the tibial crest and at the distal diaphysis. The numerical model could also be used to furnish a more subtle analysis as a precise correlation between local strain and local biological response can be obtained with the experimentally validated numerical model.     

Detailed finite element modelling of deep needle insertions into a soft tissue phantom using a cohesive approach

Detailed finite element modelling of needle insertions into soft tissue phantoms encounters difficulties of large deformations, high friction, contact loading and material failure. This paper demonstrates the use of cohesive elements in high-resolution finite element models to overcome some of the issues associated with these factors. Experiments are presented enabling extraction of the strain energy release rate during crack formation. Using data from these experiments, cohesive elements are calibrated and then implemented in models for validation of the needle insertion process. Successful modelling enables direct comparison of finite element and experimental force–displacement plots and energy distributions. Regions of crack creation, relaxation, cutting and full penetration are identified. By closing the loop between experiments and detailed finite element modelling, a methodology is established which will enable design modifications of a soft tissue probe that steers through complex mechanical interactions with the surrounding material.     

ABA and GA3 increase carbon allocation in different organs of grapevine plants by inducing accumulation of non‐structural carbohydrates in leaves,enhancement of phloem area and expression of sugar transporters

Grape quality for winemaking depends on sugar accumulation and metabolism in berries. Abscisic acid (ABA) and gibberellins (GAs) have been reported to control sugar allocation in economically important crops, although the mechanisms involved are still unknown. The present study tested if ABA and gibberellin A₃ (GA₃) enhance carbon allocation in fruits of grapevines by modifying phloem loading, phloem area and expression of sugar transporters in leaves and berries. Pot‐grown Vitis vinifera cv. Malbec plants were sprayed with ABA and GA₃ solutions. The amount of soluble sugars in leaves and berries related to photosynthesis were examined at three points of berry growth: pre‐veraison, full veraison and post‐veraison. Starch levels and amylase activity in leaves, gene expression of sugar transporters in leaves and berries and phloem anatomy were examined at full veraison. Accumulation of glucose and fructose in berries was hastened in ABA‐treated plants at the stage of full veraison, which was correlated with enhancement of Vitis vinifera HEXOSE TRANSPORTER 2 (VvHT2) and Vitis vinifera HEXOSE TRANSPORTER 6 (VvHT6) gene expression, increases of phloem area and sucrose content in leaves. On the other hand, GA₃ increased the quantity of photoassimilates delivered to the stem thus increasing xylem growth. In conclusion, stimulation of sugar transport by ABA and GA₃ to berries and stems, respectively, was due to build‐up of non‐structural carbohydrates in leaves, modifications in phloem tissue and modulation in gene expression of sugar transporters.     

A partially structured species of beta 2-microglobulin is significantly populated under physiological conditions and involved in fibrillogenesis

The folding of beta(2)-microglobulin (beta(2)-m), the protein forming amyloid deposits in dialysis-related amyloidosis, involves formation of a partially folded conformation named I(2), which slowly converts into the native fold, N. Here we show that the partially folded species I(2) can be separated from N by capillary electrophoresis. Data obtained with this technique and analysis of kinetic data obtained with intrinsic fluorescence indicate that the I(2) conformation is populated to approximately 14 +/- 8% at equilibrium under conditions of pH and temperature close to physiological. In the presence of fibrils extracted from patients, the I(2) conformer has a 5-fold higher propensity to aggregate than N, as indicated by the thioflavine T test and light scattering measurements. A mechanism of aggregation of beta(2)-m in vivo involving the association of the preformed fibrils with the fraction of I(2) existing at equilibrium is proposed from these results. The possibility of isolating and quantifying a partially folded conformer of beta(2)-m involved in the amyloidogenesis process provides new opportunities to monitor hemodialytic procedures aimed at the reduction of such species from the pool of circulating beta(2)-m but also to design new pharmaceutical approaches that consider such species as a putative molecular target.     

Mandibuloacral dysplasia is caused by a mutation in LMNA-encoding lamin A/C

Mandibuloacral dysplasia (MAD) is a rare autosomal recessive disorder, characterized by postnatal growth retardation, craniofacial anomalies, skeletal malformations, and mottled cutaneous pigmentation. The LMNA gene encoding two nuclear envelope proteins (lamins A and C [lamin A/C]) maps to chromosome 1q21 and has been associated with five distinct pathologies, including Dunnigan-type familial partial lipodystrophy, a condition that is characterized by subcutaneous fat loss and is invariably associated with insulin resistance and diabetes. Since patients with MAD frequently have partial lipodystrophy and insulin resistance, we hypothesized that the disease may be caused by mutations in the LMNA gene. We analyzed five consanguineous Italian families and demonstrated linkage of MAD to chromosome 1q21, by use of homozygosity mapping. We then sequenced the LMNA gene and identified a homozygous missense mutation (R527H) that was shared by all affected patients. Patient skin fibroblasts showed nuclei that presented abnormal lamin A/C distribution and a dysmorphic envelope, thus demonstrating the pathogenic effect of the R527H LMNA mutation.     

Reduction of 1,4-quinone and ubiquinones by hydrogen atom transfer under UVA irradiation

1,4-Benzoquinone, coenzyme Q 0 and Q 10 were reacted with a series of hydrogen donors in the ESR cavity in the presence or absence of UVA irradiation. The signals of the radicals generated from the hydrogen donors or of those of the semiquinones were detected. The reaction mechanism was interpreted by a hydrogen atom transfer instead of the usual electron transfer mechanism on the basis of the redox potentials of the reactants and the Marcus theory. The hydrogen atom transfer is explained by the excited triplet state of quinones, which, on the basis of quantum mechanic calculations, may be reached even under visible light. In some cases, hydrogen atom transfer was also observed without irradiation, although to a lesser extent.