Energy state and myosin heavy chain isoforms in single fibres of normal and transforming rabbit muscles

 Energy-rich phosphates, [ATP]/[ADP_free] ratios, and the myosin heavy chain (MHC) complement were determined in single fibres from normal rabbit muscles, and in fibres isolated from tibialis anterior muscle undergoing fast-to-slow conversion by chronic low-frequency stimulation (CLFS). In normal muscles, energy-rich phosphate contents and [ATP]/[ADP_free] ratios could thus be assigned to different MHC-based fibre types. Phosphocreatine (PCr) contents and [ATP]/[ADP_free] ratios differed markedly between fast- and slow-twitch fibres, as well as within the fast fibre subtypes. Both magnitudes were approximately twofold higher in the fastest (type IIB) fibres as compared to the slowest (type I) fibres. According to PCr contents and [ATP]/[ADP_free] ratios pure and hybrid fibres were aligned in an order similar to that determined by their contractile properties and myofibrillar ATPase activities. CLFS for up to 30 days induced pronounced decreases in PCr and [ATP]/[ADP_free] which attained levels twofold lower than in normal slow-twitch fibres. In both normal and stimulated muscles, PCr and [ATP]/[ADP_free] ratios were correlated, indicating their equilibrium in the different fibre types. The relationship detected between MHC isoform expression and the [ATP]/[ADP_free] ratio suggests that the drastic and persistent depression of the cellular energy state may act as an important signal initiating fast-to-slow transformation processes in muscle fibres. Received: 26 June 1998 / Accepted: 31 July 1998     

Delayed effects of long-term administration of granulocyte colony-stimulating factor to mice

We studied the effects of chronic administration of granulocyte colony-stimulating factor in nonmobilizing doses to mice. Over 18 months of the study, 55% animals of the treatment group died of unknown cause, blood diseases and tumors were found in 20% mice, and in 5% animals pathological changes were absent. Control mice had no diseases (normal values of total and differential leukocyte count). The diagnoses made over the first 7 months mainly included myeloproliferative diseases. Solid tumors were found at later terms. Suppurative inflammation at the site of injection was observed in all mice after 3-month treatment with granulocyte colony-stimulating factor. Our results indicate that chronic administration of granulocyte colony-stimulating factor in low doses leads to the development of etiologically different tumors and sharply reduced animal life span. The use of granulocyte colony-stimulating factor during allogeneic transplantation of hemopoietic stem cells can be hazardous for donors. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 145, No. 5, pp. 568–573, May, 2008     

Development of allergy in children. I. Association with virus infections.

Children born into allergic families, with two allergic parents, are at high risk of developing allergy within the first 5 years of life. In order to observe possible external factors in the sensitization process, a prospective study of 13 such children was done, in which serial clinical and immunologic observations were made at 3- to 6-month intervals over a period of 1 to 4 yr. Eleven of these children are now clinically allergic; 5 have asthma. Immunologic evidence for allergic sensitization was observed in these 11 children by RAST, antigen-induced leukocyte histamine release, lymphoblastogenesis, and rise in serum IgE. Upper respiratory infections (URI) occurred in these 11 allergic children 1 to 2 months prior to the onset of allergic sensitization. In 10 of these 11 URI children, complement-fixing antibodies to viruses (parainfluenza, RSV, CMV) increased in the same blood samples in which immunologic allergic sensitization was first evidenced. This coincidence suggests that certain viruses may contribute to the allergic sensitization process.     

Lymphocyte Responses to an Aqueous Extract from Cow Hair and Dander

The proliferative responsiveness to cow dander antigens of lymphocytes from 23 asthmatic patients with allergy to cow hair and dander, from 10 patients with allergic asthma to other antigens and from 31 non-atopic subjects was investigated. Lymphocytes from patients with allergic asthma due to cow epithelium antigens showed a statistically significant decreased responsiveness to an extract from cow dander, compared to that seen with lymphocytes from newborn infants, lymphocytes from normal controls and from asthmatic patients without allergy to cow. The response to the cow dander extract seemed to be an accessory cell dependent T lymphocyte response.     

Standardization for image characteristics in telemammography using genetic and nonlinear algorithms

As the soft copy reading and computer assisted diagnosis (CAD) in mammography become more and more important, the standardization of digital images becomes paramount. Telemammography and telemedicine requires the standardization for image characteristics, such as image resolution, bit-depth and intensity response. Soft copy reading and CAD in mammography are both dependent on the characteristics of the source of the digital data, either direct digital mammography or digitized screen-film mammography. An algorithm developed on images from one database may not perform well as on images from another database (with a different digitization). In this paper, we describe two methods based on a genetic algorithm and a nonlinear algorithm for standardization of digitized and digital mammography. The proposed standardization techniques are based on geometric and intensity transformations that are discovered using a set of calibration images. A set of transformation algorithm is used to search for the best standardization.     

Biolistic transformation of conidia of Botryotinia fuckeliana

Botryotinia fuckeliana, the causal agent of grey mould, was biolistically transformed to hygromycin B resistance using a plasmid (pOHT) containing a bacterial hygromycin phosphotransferase gene fused to regulatory sequences from Aspergillus nidulans. Multiple copies of the plasmid, precipitated onto tungsten particles, were delivered into the conidia by a helium-driven gene gun. Southern analysis showed that the plasmid was integrated into the fungal genome at one single locus. After five subsequent transfers on selective medium, all transformants were mitotically stable. When propagated on non-selective medium, four out of eight transformants retained their resistance to hygromycin B. Southern analysis of the fifth generation of transformants showed that no genetic rearrangements occurred during vegetative growth of stable transformants.     

Transformation of Neurospora crassa with the trp-1 gene and the effect of host strain upon the fate of the transforming DNA

Summary Neurospora trp-1 ⁺ transformants, obtained by transforming a trp-1 inl strain with plasmid DNA containing the wild type trp1 ⁺ gene, were characterized by genetic and Southern blot analyses. The transforming trp-1 gene integrated at or near the resident site in all of the trp-1 ⁺ transformants obtained with circular DNA or DNA cut within the trp-1 coding region. The frequency of homologous integration decreased substantially when the donor DNA was cleaved outside the trp-1 coding region. The transformants were very stable mitotically and, in general, also showed meiotic stability. Analysis of trp-1 ⁺ transformants obtained with another recipient strain, trp-1 ⁺ ga-2 aro-9 inl, showed that homologous integration of donor DNA occurred in only 20% of the transformants, whether circular or linear DNA was used. Thus, the host strain employed for transformation appears to be a major factor in determining the fate of transforming DNA. Southern blot analysis of transformants showed that integration of the transforming DNA at the homologous site occurred by double crossover or gene conversion events rather than by insertion of the entire plasmid DNA. Multiple and apparently non functional integration events were observed in some transformants.     

Antigens targeted to the Leishmania phagolysosome are processed for CD4+ T cell recognition

Processing of antigen for recognition by class II-restricted CD4⁺ T cells occurs within acidic compartments of the antigen-presenting cell. The exact nature of this compartment has yet to be precisely defined, however, but may vary depending upon the cell type studied and the antigen used. The acidic compartments of macrophages are also responsible for the degradation of ingested micro-organisms and play host to others which are adapted to an intracellular existance. To determine whether the phagolysosome (PL) formed in activated macrophages after ingestion of Leishmania parasites is also a site for entry of antigen into the class II presentation pathway, we have used the approach of genetic transformation. Hence, Leishmania were transfected with the genes for the protein antigens ovalbumin (OVA) and β-galactosidase (β-gal) and after infection were able to deliver these antigens specifically into the PL. Delivery of antigen to this site resulted in the ability of infected macrophages to present these antigens to antigen-specific CD4⁺ T cells. After taking into account the absolute levels of antigen uptake by macrophages, a 4-h processing period for OVA delivered by this or a soluble route led to equivalent levels of T cell activation. Unlike macrophages pulsed with soluble OVA, those with PL-targeted OVA still retained the ability to stimulate T cells after a 24-h processing period. This enhanced lifespan of antigen in macrophages corresponded to the kinetics of degradation of the parasite, suggesting slow release of antigen into the processing pathway. β-gal presentation from the PL was tenfold less efficient under the same conditions. In addition to providing the first information on antigen processing in a protozoan PL, these studies highlight the usefulness of genetically transformed parasites for these types of studies.     

Critical role for SV40 small-t antigen in human cell transformation

Defining the ability of simian virus 40 (SV40) to transform human cells has become of even greater importance with the increased understanding that this virus may play a role in some human malignancies. This report documents the requirement for viral small-t (ST) antigen in large-T (LT)-driven transformation of primary fibroblasts, a requirement that cannot be met by a well-known oncogene, c-Ha-ras (EJ-ras), which can cooperate with LT in rodent systems. The cellular gene telomerase is not essential for transformation, although transformed clones are not immortal without it. Similarly, an immortal mesothelial cell line has been developed using LT and telomerase. Immortalized mesothelial cells are morphologically normal, but can be transformed by introduction of ST, or ST + ras, but not by ras alone. It is likely that ST will be required along with LT for transformation of most human cell types.