Click to increase image sizeClick to decrease image sizeРЕЗЮМЕРассмотрены возможности применения моноклональных антител в экспериментальных исследованиях и клинической медицине. Вкратце проанализированы преимущества и недостатки моноклональных антител мышей при использовании их в клинической практике, описаны экспериментальные основы получения гибридомных клеточных линий, секретирующих человеческие иммуноглобулины. Изложены и проанализированы опыты по конструированию гетерогибридом — клеточных линий, полученных вследствие гибридизации миеломных клеток мышей с человеческими лимфоцитами. Приводятся наиболее существенные литературные источники об условиях и трудностях получения гетерогибридомных линий и показаны перспектипы этого нового метода в биотехнологии.SUMMARYThe paper considers the possibilities for application of monoclonal antibodies in experimental research and clinical medicine. Following a brief analysis of advantages and shortcomings of the use of mouse monoclonal antibodies in clinical practice, the experimental principles for obtaining hybridoma cell lines secreting human immunoglobulins have been revealed. Experiments for construction of heterohybridomas (cell lines obtained after hybridization of mouse myeloma cells with human lymphocytes) have been described and analyzed. A review of the most important scientific information on the conditions and the difficulties in developing heterohybridoma lines has been made and the prospects of this new biotechnological method have been outlined.
The induction of an acute inflammatory response followed by the release of polypeptide cytokines and growth factors from peripheral blood monocytes has been implicated in mediating the response to vascular injury. Because the Cu2+-binding proteins IL-1alpha and fibroblast growth factor 1 are exported into the extracellular compartment in a stress-dependent manner by using intracellular Cu2+ to facilitate the formation of S100A13 heterotetrameric complexes and these signal peptideless polypeptides have been implicated as regulators of vascular injury in vivo, we examined the ability of Cu2+ chelation to repress neointimal thickening in response to injury. We observed that the oral administration of the Cu2+ chelator tetrathiomolybdate was able to reduce neointimal thickening after balloon injury in the rat. Interestingly, although immunohistochemical analysis of control neointimal sections exhibited prominent staining for MAC1, IL-1alpha, S100A13, and the acidic phospholipid phosphatidylserine, similar sections obtained from tetrathiomolybdate-treated animals did not. Further, adenoviral gene transfer of the IL-1 receptor antagonist during vascular injury also significantly reduced the area of neointimal thickening. Our data suggest that intracellular copper may be involved in mediating the response to injury in vivo by its ability to regulate the stress-induced release of IL-1alpha by using the nonclassical export mechanism employed by human peripheral blood mononuclear cells in vitro.
Murine hybridomas were generated to DNA/tight binding proteins complex isolated from the residual nuclear structure following a procedure analogous to that yielding "empty" shells of nuclear envelope. A monoclonal antibody designated 2A8 was selected because of its differential immunostaining of mitotic cells of a synchronized mouse fibroblast cell culture L-929. The target antigen was rendered insoluble by a sequence of extractions of isolated nuclei of diverse cell types with detergents, urea, DNase I and alkali thus reproducing some solubility properties of proteins constituting an operationally defined residual nuclear matrix. The cognate polypeptide was localized on a subset of proteins of M(r) 58-65 kDa, 70 kDa in isolated fibroblast nuclear matrices. The functional implication of the antigen in mitosis-related disassembly-assembly process of the nuclear matrix/envelope was detected. At prophase the antibody decorated the nuclear periphery and nuclear envelope fixed inward filaments. A fibrous network of cytoplasmic localization was stained in metaphase. At anaphase the antigen was dispositioned into peripheral fibrogranular clusters of polar orientation predominantly on one side of the nucleus. Proceeding to telophase a spreading fluorescence was manifested over the entire contour of the nuclear periphery to delineate the reforming nucleus. By immunogold electron microscopy of interphase cells the antigen was identified as evenly distributed in chromatin and interchromatin regions. At initiation of chromosome condensation in mitosis the label was detected predominantly in the chromosomal area.
Imbalance of activation and inhibition of matrix metalloproteinases (MMPs) lead to an increase in their activity and the occurrence of pathological changes in the vascular wall. The purpose of this paper is to determine the role of MMP-2 and MMP-9 in vascular and ventricular remodelling in patients with heart failure with preserved ejection fraction. The patients were divided into three groups: 15 patients with heart failure with preserved ejection fraction (HFpEF), 72.73 ± 10.44 years old; 15 patients with arterial hypertension (AH), 63.73±7.06 years old; 10 healthy controls, 58.7± 5.87 years old. Arterial stiffness was assessed by pulse wave velocity (PWV). Diastolic dysfunction was assessed by the ratio of early diastolic mitral flow velocity and early diastolic myocardial velocity (E/Em ratio). Left ventricular mass was calculated by area-length method and indexed to body surface area (LVMI). MMP-2, MMP-9 and Brain Natriuretic Peptide (BNP) were measured by ELISA technique. MMP-2 was higher in patients with HFpEF and AH versus controls (13987±4464 ng/ml and 13040±5060 ng/ml vs 9260±4135 ng/ml, p = 0.047). MMP-9 was similar across the groups. BNP was higher in HFpEF versus AH and controls (775.33±443.59pg/ml vs 370.00±158.29 pg/ml and 345.00±94.39 pg/ml, p = 0.002). In HFpEF patients, PWV (12.04±2.46 m/s vs 10.06±1.94 m/s vs 7.22±1.19 m/s, p < 0.0001), LVMI (134.11±29.40g/m 2 vs 122.45±23.73 Elisa kits for MMP-2, MMP-9 and BNP were purchased with funds from project “Young researcher” No 4D/2011 of the Medical University of Sofia.
