Experimental investigations in prolonged myocardial ischaemia. Note I. Biology of the myocardial fiber after transient myocardial fiber after transient myocardial ischaemia

The first ten days' evolution of post-ischaemic lesions of the premonitory or angina pectoris syndrome type was experimentally studied by the challenge of a short-term (10 and 15 min) ischaemia, of an adaptation to ischaemia and an adaptation followed by prolonged ischaemia (20 and 35 min). Worthy of note was the persistence of reversible lesions after short-term ischaemia and adaptation, and the progressive evolution towards cytolysis and cicatrization of some pancicellular foci after adaptation followed by prolonged ischaemia. The role of mitochondrial lesions, of lysosomal hydrolases, the inefficiency of renewed circulation, as well as problems of diagnosis are discussed.     

Activated macrophages and antibodies against the plant lectin, GSI-B4, recognize the same tumor-associated structure (TAS)

Activated macrophages that were stabilized with either formalin or glutaraldehyde absorbed two polypeptides (Mr 100,000 and 60,000) from detergent extracts of all of the tumor cell lines tested, but not from detergent extracts of normal human peripheral blood lymphocytes. A major polypeptide (Mr 95,000) was retained from spent culture media of tumor cell lines. Polypeptides with molecular sizes of 100,000 and 60,000 daltons were also adsorbed by activated macrophages from detergent extracts of chicken embryo cell membranes, suggesting an oncofetal nature for these proteins. The 100,000 dalton polypeptide, but not the 60,000 dalton component, was found to be available to lactoperoxidase-catalyzed cell surface iodination. Polypeptides with identical molecular sizes could be adsorbed to immobilized galactopyranoside, indicating that they are vertebrate lectins. Activated macrophages and affinity adsorbents prepared by the covalent coupling of galactopyranoside to agarose also bind the plant lectin isolectin B4 prepared from the seeds of Griffonia simplicifolia. On the basis of these findings, we put forth the hypothesis that macromolecules of the same specificity, that is affinity to galactopyranosyl residues, must show homologies in their binding sites. We have predicted therefore that antisera prepared against this plant lectin should cross-react with galactopyranosyl-binding vertebrate lectins present on the surface of tumor cells. In this communication, we also report the generation of hybridomas that produce antibodies reactive with both the plant and vertebrate lectins. Inhibition experiments that make use of various mono- and disaccharides suggest that the specificities of these antibodies are for determinants intimately associated with the galactosyl binding site on the lectin molecule. Two of the antibodies were found to have moderate selectivity for tumor cells when tested in an immunohistochemical procedure that made use of fresh-frozen or paraffin-embedded sections of human biopsy material. These two antibodies on immunoblots of tumor cell membrane extracts reacted with a polypeptide with an apparent molecular size of 100,000 daltons.     

The stoichiometry of the Ca2+ pump in human erythrocyte vesicles: modulation by Ca2+, Mg2+ and calmodulin

Active Ca2+ uptake and the associated (Ca2+ + Mg2+)-ATPase activity were studied under the same conditions in an inside-out vesicle preparation of human red blood cells made essentially by the procedure of Quist and Roufogalis (Journal of Supramolecular Structure 6, 375-381, 1977). Some preparations were treated with 1 mM EDTA at 30 degrees to further deplete them of endogenous levels of calmodulin. As the Ca2+ taken up by the EDTA-treated inside-out vesicles, as well as the non-EDTA treated vesicles, was maintained after addition of 4.1 mM EGTA, the vesicles were shown to be impermeable to the passive leak of Ca2+ over the time course of the experiments. In the absence of added calmodulin, both active Ca2+ uptake and (Ca2+ + Mg2+)-ATPase were sensitive to free Ca2+ over a four log unit concentration range (0.7 microM to 300 microM Ca2+) at 6.4 mM MgCl2. Below 24 microM Ca2+ the stoichiometry of calcium transported per phosphate liberated was close to 2:1, both in EDTA and non-EDTA treated vesicles. Above 50 microM Ca2+ the stoichiometry approached 1:1. When MgCl2 was reduced from 6.4 mM to 1.0 mM, the stoichiometry remained close to 2:1 over the whole range of Ca2+ concentrations examined. In contrast to the results at 6.4 mM MgCl2, the Ca2+ pump was maximally activated at about 2 microM free Ca2+ and significantly inhibited above this concentration at 1 mM MgCl2. Calmodulin (0.5-2.0 microgram/ml) had little effect on the stoichiometry in any of the conditions examined. The possible significance of a variable stoichiometry of the Ca2+ pump in the red blood cell is discussed.     

Possible involvement of DNA breaks in epigenetic regulation of cell differentiation

The review summarizes the authors’ and literature data on accumulation of DNA breaks in differentiating cells. Large 50-kb free DNA fragments were observed by several research teams in non-apoptotic insect, mammal, and plant cells. More intense DNA breakage was observed during maturation of spermatides, embryo development, and differentiation of myotubes, epidermal cells, lymphocytes, and neutrophils. In general, accumulation of DNA breaks in differentiating cells cannot be attributed to a decrease in the DNA repair efficiency. Poly(ADP)ribose synthesis often follows the DNA breakage in differentiating cells. We hypothesize that DNA fragmentation is an epigenetic tool for regulating the differentiation process. Scarce data on localization of the differentiation-associated DNA breaks indicate their preferable accumulation in specific DNA sequences including the nuclear matrix attachment sites. The same sites are degraded at early stages of apoptosis. Recent data on non-apoptotic function of caspases provide more evidence for possible existence of a DNA breakage mechanism in differentiating cells, resembling the initial stage of apoptosis. Excision of methylated cytosine and recombination are other possible explanations of the phenomenon. Elucidation of mechanisms of differentiation-induced DNA breaks appears to be a prospective research direction.