Biomedical subjects

P Malý

Publications and source records attributed to P Malý.

16 recordsLinked to original sources

PubMed2001

The alpha(1,3)fucosyltransferases FucT-IV and FucT-VII exert collaborative control over selectin-dependent leukocyte recruitment and lymphocyte homing.

E-, P-, and L-selectin counterreceptor activities, leukocyte trafficking, and lymphocyte homing are controlled prominently but incompletely by alpha(1,3)fucosyltransferase FucT-VII-dependent fucosylation. Molecular determinants for FucT-VII-independent leukocyte trafficking are not defined, and evidence for contributions by or requirements for other FucTs in leukocyte recruitment is contradictory and incomplete. We show here that inflammation-dependent leukocyte recruitment retained in FucT-VII deficiency is extinguished in FucT-IV(-/-)/FucT-VII(-/-) mice. Double deficiency yields an extreme leukocytosis characterized by decreased neutrophil turnover and increased neutrophil production. FucT-IV also contributes to HEV-born L-selectin ligands, since lymphocyte homing retained in FucT-VII(-/-) mice is revoked in FucT-IV(-/-)/FucT-VII(-/-) mice. These observations reveal essential FucT-IV-dependent contributions to E-, P-, and L-selectin ligand synthesis and to the control of leukocyte recruitment and lymphocyte homing.

Animals↗

PubMed2001

Expression of human erythropoietin gene in the mammary gland of a transgenic mouse.

WAP is being recognized as the principal milk protein expressed in pregnant or lactating females of several mammalian species. Recently, it has been shown that the 6.3-kb 5' untranslated region of the rWAP gene is able to control, and almost completely restrict, the expression of the transgene into the mammary gland of the transgenic animal. We cloned the genomic fragment carrying the rWAP gene locus from the rabbit phage genomic library and used the 8.5-kb long 5' untranslated part of the rWAP gene to target the expression of hEPO, cloned from the human phage genomic library, into the mammary gland of the mouse. The vectors, carrying either the hEPO gene or the rWAP-hEPO hybrid gene, were injected into the mouse ova, and 12 transgenic animals were identified by PCR and Southern blot from the progeny of 168 tested littermates. Transgenic mice were viable, fertile and displayed a normal development. Recombinant human erythropoietin was produced in the milk of a transgenic mouse female at a secretion level of 5.3 mIU/ml, as detected by ELISA. Despite the low production of the transgenic glycoprotein in the milk we demonstrate that the hybrid gene can be expressed in the mammary gland of the host animal. Thus, WAP-based recombinant vectors, with additional optimizing modifications, can be useful for production of therapeutic proteins in the transgenic mammals.

Animals↗

PubMed2000

Detection of ICAM-1 in experimentally induced colitis of ICAM-1-deficient and wild-type mice: an immunohistochemical study.

Adhesion molecules (e.g. ICAM-1, CD 54) are known to be upregulated on activated vascular endothelial cells during inflammatory reactions. To study the role of ICAM-1 in intestinal inflammation in vivo, we induced acute experimental colitis in wild-type (C57BL/6) mice and ICAM-1-deficient mice, by feeding the animals with 3% dextran sodium sulphate (DSS) in drinking water for 7 days. In the control strain the immunohistochemical staining showed a very pronounced endothelial upregulation of ICAM-1 after the DSS treatment observed in areas of inflammatory infiltrate, especially in venules or arterioles of the propria and submucosa, and partly in the mesocolon. DSS-fed ICAM-1-deficient mice showed no endothelial enhancement and only faint staining of venules or capillaries approaching that encountered in the control ICAM-1-deficient animals. Our data indicate that ICAM-1 may play a crucial role in the development of acute intestinal inflammation, consistent with our finding that ICAM-1 deficiency can obviate severe forms of experimentally induced colitis in mice.

Animals↗

PubMed2000

Reduced phagocytic activity of polymorphonuclear leukocytes in alpha(1,3) fucosyltransferase VII-deficient mice.

Deficiencies in adhesion molecules or their counter-receptors in humans may have severe consequences as exemplified by leukocyte adhesion deficiency (LAD) I or II syndromes. Because such diseases occur with great rarity, animal models are valuable for studying the role of particular adhesion molecules and their natural ligands in immunity. We studied selected immune parameters and general health in mice with a defect in the sialyl-Lewis X antigen (selectin ligand) caused by disruption of the gene encoding alpha(1,3)fucosyltransferase VII (Fuc-TVII). Leukocytes from Fuc-TVII -/- and control mice were tested for adherence to cellophane membranes or polymer particles in vivo and phagocytic activity in vitro. While no difference in adherence was found, the number of neutrophil granulocytes in exudate induced by intraperitoneal injection of polymer beads was reduced in knock-out mice. Moreover, the phagocytic activity in Fuc-TVII -/- mice was significantly reduced. These animals have splenomegaly due to increased hematopoiesis and reduced weight but do not exhibit clinical signs of immunodeficiency. In conclusion, the lack of Fuc-TVII activity leads to several morphological and functional abnormalities without an impact on survival rate.

Animals↗

PubMed2000

Intercellular adhesion molecule-1 (ICAM-1) deficiency protects mice against severe forms of experimentally induced colitis.

ICAM-1 (CD54), the ligand for LFA-1 and Mac-1, is up-regulated during inflammatory reaction on the activated vascular endothelium. To determine its role in intestinal inflammation, we induced acute experimental colitis in mice with a deleted ICAM-1 gene, by feeding them with 3% dextran sodium sulphate (DSS) in drinking water for 7 days. Chronic colitis was elicited by DSS similarly, followed by 2 weeks with water. In the acute phase of inflammation, ICAM-1-deficient mice exhibited a significantly lower mortality rate (5%) than control C57Bl/6J mice (35%). Control animals, but not the ICAM-1-deficient mice, exhibited diarrhoea and rectal bleeding. Histological examination of large-bowel samples evaluated the intensity of inflammatory changes, and type and extent of mucosal lesions. In the acute phase, 33.3% of samples from ICAM-1-deficient mice exhibited mucosal defects (flat and fissural ulcers), predominantly mild to moderate inflammatory infiltrate within the lamina propria mucosae and lower grades of mucosal lesions. Much stronger inflammatory changes were present in control animals, flat ulcers (sometimes multiple) and fissural ulcers being observed in 62.5% of samples. Mucosal inflammatory infiltrate was moderate to severe, typically with higher grades of mucosal lesions. In chronic colitis, smaller inflammatory changes were found in the large bowel. The two mouse strains differed, the chronic colitis being accompanied by an increased serum level of anti-epithelial IgA autoantibodies in C57Bl/6 control mice but not in ICAM-1-deficient mice. These findings provide direct evidence of the participation of ICAM-1 molecule in the development of experimentally induced intestinal inflammation.

Animals↗

PubMed1996

The alpha(1,3)fucosyltransferase Fuc-TVII controls leukocyte trafficking through an essential role in L-, E-, and P-selectin ligand biosynthesis.

alpha(1,3)Fucosylated oligosaccharides represent components of leukocyte counterreceptors for E- and P-selectins and of L-selectin ligands expressed by lymph node high endothelial venules (HEV). The identity of the alpha(1,3)fucosyltransferase(s) required for their expression has been uncertain, as has a requirement for alpha(1,3)fucosylation in HEV L-selectin ligand activity. We demonstrate here that mice deficient in alpha(1,3) fucosyltransferase Fuc-TVII exhibit a leukocyte adhesion deficiency characterized by absent leukocyte E- and P-selectin ligand activity and deficient HEV L-selectin ligand activity. Selectin ligand deficiency is distinguished by blood leukocytosis, impaired leukocyte extravasation in inflammation, and faulty lymphocyte homing. These observations demonstrate an essential role for Fuc-TVII in E-, P-, and L-selectin ligand biosynthesis and imply that this locus can control leukocyte trafficking in health and disease.

Animals↗

PubMed1995

Oocyte Gal alpha 1,3Gal epitopes implicated in sperm adhesion to the zona pellucida glycoprotein ZP3 are not required for fertilization in the mouse.

The Gal alpha 1-->3Gal structure is displayed on the zona pellucida glycoprotein ZP3 on murine oocytes. This trisaccharide has been implicated in sperm-zona pellucida adhesive events thought to be essential to fertilization in the mouse. To determine directly if this molecule is required for fertilization, we have generated mice that are deficient in a gene (alpha 1,3GT) encoding the UDP-Gal:beta-D-Gal-alpha 1-->3Gal-galactosyltransferase enzyme responsible for Gal alpha 1-->3Gal synthesis and expression. These mice develop normally and exhibit no gross phenotypic abnormalities. The Gal alpha 1-->3Gal epitope is absent from the vascular endothelium and other tissues in alpha 1,3GT (-/-) adult mice. By contrast, alpha 1,3GT (-/-) mice, like humans, develop naturally occurring anti-alpha-galactoside antibodies normally absent in wild type mice. Female alpha 1,3GT (-/-) mice yield oocytes that are devoid of the Gal alpha 1-->3Gal epitope; however, these mice are fully fertile. These observations indicate that the Gal alpha 1-->3Gal moiety is not essential to sperm-oocyte interactions leading to fertilization or to essentially normal development. They further suggest that alpha 1,3GT (-/-) mice will find utility for exploring approaches to diminish anti-Gal-dependent hyperacute xenograft rejection, which presents a major barrier to the use of porcine and other non-primate organs for xenotransplantation in humans.

Animals↗

PubMed1995

[A method of using intraluminal splints in the treatment and prevention of recurrent ileus of the small intestine].

The authors present a group of 23 patients where splints were used in the loops of the small intestine. The group is divided into two sub-groups. The first one comprises patients where the splint was inserted because of relapsing ileus of the small intestine after previous surgical operations. The second sub-group comprises patients where this operation was performed either preventively or as part of a primary operation or as part of a reoperation called for by complications immediately preceding operation. The internal splint is inserted via jejunostomy and through the loops of the small intestine it is guided by means of a sutured Foley catheter which after proper insertion of the splint is left in the caecum. The authors emphasize the importance of effective decompression of the small intestine during the early postoperative period which is made possible by suction of the ileous contents through lateral openings of the inserted probe. After three years' work, using this method, the authors confirm that it gives good results.

Adult↗

PubMed1995

[Subtotal colectomy in emergency situations].

One-stage subtotal colectomy is the most radical solution of ileous conditions caused by an obturating tumour of the left half of the colon. The authors report on their experience with this procedure in 10 patients operated in the course of three years. They emphasize the advantages such as oncological radicality, immediate detoxication of the organism, a favourable postoperative course with a low morbidity (10% dehiscences) and lethality (10%), shorter hospitalization period, life of the patients without a stoma, lower costs and satisfactory functional results. For an experienced surgeon, if perfect intensive postoperative care is available, this operation is the method of choice even in very old patients.

Aged↗

PubMed1993

[Peroperative glove damage as a possible risk for the surgical team].

With regard to the relative absence of data in our as well as the world literature on possible peroperative injuries of the surgical team, the authors investigated damage of surgical gloves during operation. They divided operations by types and the structure of the surgical team. It was found that on average 45.5% of the gloves used during operations were damaged.

General Surgery↗

PubMed1993

[Fasciocutaneous flaps].

Fasciocutaneous flaps proved useful for covering soft tissue defects in the region of the leg. After experience with the use of these flaps in six patients the authors recommend this method for covering these defects. In all patients very satisfactory results were achieved.

Humans↗

PubMed1992

Retinoic acid-induced changes in differentiation-defective embryonal carcinoma RAC65 cells.

RAC65 is a mutant clone of mouse embryonal carcinoma cells, P19, which does not undergo terminal differentiation upon treatment with retinoic acid (RA). RAC65 cells express a truncated RA receptor alpha (RAR alpha) which, however, does not fully explain their defect. Here we show that RAC65 cells exhibit an additional defect in RAR alpha mRNA which may reflect a defect in RNA splicing. The parental and mutant cells also differ in their capacities to bind [3H]RA into nuclear fractions and in expression of cellular RA binding protein (CRABP) mRNA after treatment with RA. The combined data suggest that the defect in RAC65 RAR alpha results in reduced expression of the CRABP gene after RA treatment and, therefore, increased flow of RA into the nucleus.

Animals↗

PubMed1992

Differential expression of the mouse and human Thy-1 gene in embryonal carcinoma cells.

Mouse P19 embryonal carcinoma (EC) cells express on their surfaces a Thy-1 glycoprotein. The expression of Thy-1 at the mRNA and protein levels is down-regulated during differentiation induced by retinoic acid (RA). Thy-1 is also expressed in human NTERA-2 EC cells, but its expression is not down-regulated during RA-induced differentiation. As a first step towards understanding differential regulation of the mouse and human Thy-1 gene in EC cells, we have introduced genomic DNA fragments encompassing the mouse or human Thy-1 gene into NTERA-2 and P19-derived cells and analyzed surface properties of the transfectants. In the transient transfection assay, both mouse and human Thy-1 genes were expressed on cell surfaces at comparable levels. P19-derived stable transfectants exhibited great clonal variations in the expressions of the transfected Thy-1 gene products, which in part reflected copy numbers. There was no simple correlation between the expression of the transfected Thy-1 gene and two stem cell surface markers, TEC-1 and TEC-4. In the course of differentiation induced by RA several clones with a surface phenotype of EC cells exhibited a significant decrease in the expression of the transfected mouse Thy-1, whereas expression of the human Thy-1 was less efficiently down-regulated. The results suggest the presence of multiple cis- and trans-acting elements controlling expression of the mouse and human Thy-1 genes in P19 EC cells and their differentiated derivatives.

Animals↗

PubMed1989

The Thy-1 glycoprotein is expressed in mouse embryonal carcinoma cells P19.

Thy-1 is a well characterized glycoprotein known to be variably expressed on the surface of different cell types. Serological analysis of a limited number of teratocarcinoma-derived cell lines suggested that mouse embryonal carcinoma cells do not express Thy-1 and that its expression is associated with the appearance of differentiated cells. In this report we show that monoclonal antibody 1aG4, recognizing Thy-1.2 epitope, binds specifically to P19 embryonal carcinoma cells and their undifferentiated subclones. A number of control experiments confirmed that 1aG4 antibody binds to the Thy-1.2 glycoprotein expressed on the surface of P19 embryonal carcinoma cells and not to the antigen expressed on differentiated derivatives of these cells or to a cross-reactive epitope. Transcriptional activity of the Thy-1 gene in undifferentiated P19 cells was shown by transfection experiments in which transfer of the Thy-1.1 gene into P19 cells resulted in stable expression of the Thy-1.1 antigen on the surface of recipient cells. Direct evidence for the presence of Thy 1 mRNA in P19 cells was obtained by Northern blot analysis with a Thy-1-specific cDNA probe. Treatment of P19 cells with retinoic acid resulted in a decrease in the expression of Thy-1 antigen which preceded changes in morphology of the cells. These data indicate that Thy-1 is a developmentally regulated surface marker of P19 embryonal carcinoma cells which is amenable to direct genetic analysis.

Animals↗

PubMed1987

Mutants of embryonal carcinoma cells defective in the expression of embryoglycan.

Embryonal carcinoma cells defective in the expression of developmentally regulated carbohydrate epitope of teratocarcinoma cells (TEC-1) were isolated from mutagenized P19X1 and P19S1801A1 cells by a single-step selection technique using monoclonal antibody TEC-01 conjugated to plant toxin ricin. Three independently isolated mutant cell lines were characterized in detail. Analysis of the expression of the TEC-1 epitope in somatic cell hybrids constructed between wild-type and mutant cells and between two mutant cell types revealed that the mutant phenotypes are recessive and that the mutants belong to, at least, two complementation groups. Each mutant cell line exhibited a unique binding pattern of four monoclonal antibodies and five lectins, and different properties of large glycopeptides were distinguished by Sephadex G-50 column chromatography. The combined data suggest that our mutants identify three genes involved in the synthesis of embryoglycan, one of which appears to be the regulatory or structural gene for fucosyltransferase. One mutant cell line was completely deprived of embryoglycan and several carbohydrate structures typical of early embryonic and embryonal carcinoma cells; however, the cells were similar to parental cells in their morphology, their ability to form aggregates when cultured in suspension, their ability to differentiate into neuron-like cells after treatment with retinoic acid, and their ability to form tumors composed of embryonal carcinoma cells. Thus, embryoglycan is not required for the expression of a number of properties of the embryonal carcinoma phenotype.

Animals↗

PubMed1985

Studies on lectins. LVIII. Sugar-binding properties, as determined by affinity electrophoresis, of alpha-D-galactosidases from Vicia faba seeds possessing erythroagglutinating activity.

The interaction of alpha-D-galactosidases from Vicia faba seeds with saccharides was studied by means of affinity electrophoresis on polyacrylamide gel in an acidic buffer system. For the preparation of affinity gels, water-soluble O-glycosyl polyacrylamide copolymers and polysaccharides were used. alpha-D-Galactosidases interact with immobilized O-alpha-D-galactosyl residues and glycogen, but no interaction was observed with immobilized O-alpha-D-mannosyl residues. On the basis of the results of affinity electrophoresis performed in the presence of various free sugars, dissociation constants of the various alpha-D-galactosidase-free sugar complexes were calculated.

Agglutination↗