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M Boubelík

Publications and source records attributed to M Boubelík.

At least 19 recordsLinked to original sources
PubMed2001

Differential sensitivity to acute cholesterol lowering of activation mediated via the high-affinity IgE receptor and Thy-1 glycoprotein.

Lateral cross-linking of transmembrane high-affinity IgE receptors (FcepsilonRI) or glycosylphosphatidylinositol-anchored Thy-1 glycoproteins on the surface of rat mast cells and rat basophilic leukemia (RBL) cells triggers the signaling pathways that lead to the release of allergy mediators. Although both of these pathways are initiated by an increased activity of Lyn kinase, the exact mechanism by which Lyn kinase interacts with aggregated FcepsilonRI and Thy-1 is not completely understood. Here we demonstrate that pretreatment of RBL cells with methyl-beta-cyclodextrin (MBCD) resulted in a dose- and time-dependent decrease in cellular cholesterol, increased detergent solubilization of Thy-1 and Lyn kinase, and a transient increase in tyrosine phosphorylation of several proteins. Acute lowering of cholesterol suppressed the activation through Thy-1, as determined by tyrosine phosphorylation of Syk kinase and some other proteins, and modulation of free cytoplasmic calcium. In contrast, the FcepsilonRI-mediated activation events were more resistant. Thy-1 and FcepsilonRI in MBCD-pretreated cells also differed in the extent of aggregation after cross-linking: Thy-1 formed large caps, whereas FcepsilonRI accumulated in small patches. MBCD treatment induced an increased release of secretory components in both Thy-1- and FcepsilonRI-activated cells. The combined data indicate that cholesterol depletion does not merely block receptor signaling but has more complex consequences.

Animals↗
PubMed2001

Mixture of trypsin, chymotrypsin and papain reduces formation of metastases and extends survival time of C57Bl6 mice with syngeneic melanoma B16.

PURPOSE: The aim of the present study was to investigate the effect of a mixture of proteolytic enzymes (comprising trypsin, chymotrypsin and papain) on the metastatic model of syngeneic melanoma B16. METHODS: 140 C57B16 mice were divided into two control and two "treated" groups. Control groups received saline rectally, twice a day starting 24 h after intracutaneous transplantation (C1) or from the time point of the primary B16 melanoma extirpation (C2), respectively. "Treated" groups were rectally administered a mixture of 0.2 mg trypsin, 0.5 mg papain, and 0.2 mg chymotrypsin twice daily starting 24 h after transplantation (E1) or after extirpation of the tumor (E2), respectively. Survival of mice and B16 melanoma generalization were observed for a period of 100 days. Immunological evaluation of B16 melanoma cells in the ascites was accomplished. CD44, CD54 and CD106 cells were measured by flow cytometry. RESULTS: Administration of proteolytic enzymes to mice inhibited the growth of primary tumors, and tumor recurrences were less numerous. Importantly, metastasis was considerably curtailed both in the vicinity of the primary tumor and at distant locales. These findings correlated with a decreased expression of CD44 and CD54 molecules in tumors exposed to proteolytic enzymes in vivo. CONCLUSIONS: Our data suggest that serine and cysteine proteinases suppress B16 melanoma, and restrict its metastatic dissemination in C57B16 mice.

Administration, Rectal↗
PubMed2000

Soluble isoforms of CEACAM1 containing the A2 domain: increased serum levels in patients with obstructive jaundice and differences in 3-fucosyl-N-acetyl-lactosamine moiety.

CEACAM1 (biliary glycoprotein or CD66a) is a member of the carcinoembryonic antigen (CEA) subgroup of the CEA family. Eleven RNA isoforms derived from the splicing of a single CEACAM1 gene have been described. Some of the CEACAM1 isoforms have been recognized by the CD66 antibodies in T and B lymphocytes, natural killer cells, granulocytes and epithelial cells in several human tissues. Although it is also present in soluble form in bile and serum, and elevated levels have been found in the serum of patients with liver diseases, it is not known which isoforms are primarily involved. In order to learn more about the distribution and properties of particular CEACAM1 isoforms, we have prepared a monoclonal antibody specific for the A2 domain of CEACAM1, designated TEC-11. This antibody does not cross-react with other members of the CEA family. Immunoblotting analysis revealed that the TEC-11 epitope was present in all cell types expressing CEACAM1 containing the A2 domain [CEACAM1(A2)], including granulocytes (160 000 MW isoform) and sperm cells (140 000 MW isoform). A 115 000 MW isoform of CEACAM1(A2) was present in human serum, bile, saliva and seminal fluid. Human bile, saliva and seminal fluid also contained the 160 000 MW CEACAM1(A2) isoform. Significantly higher serum levels of the 115 000 MW CEACAM1(A2) isoform were detected in patients with obstructive jaundice. The 160 000 MW isoform of CEACAM1(A2) in bile, but not a 115 000 MW isoform in serum and bile, carried the 3-fucosyl-N-acetyl-lactosamine moiety. The combined data indicate that various isoforms of CEACAM1(A2) are present in different body fluids where they could take part in different CEACAM1-mediated functions.

Animals↗
PubMed2000

Genetic variation in in-vitro cytokine-induced production of nitric oxide by murine peritoneal macrophages.

Quantitative aspects of the in-vitro interferon (IFN)-gamma-induced nitric oxide (NO) production by peritoneal macrophages of eight inbred strains of mice were investigated. Animals employed in the study can be assorted into three phenotype categories: high, moderate, and low NO-responders. Concentration of nitrites in the 24-h supernatants of cells stimulated with recombinant murine IFN-gamma (25 U/ml) reached the following values (mean +/- SEM; in microM): C57BL/10 (33.7+/-1.88) = C57BL/6 (32.1+/-2.10) > SIL (24.0+/-1.55) > CBA/J (18.1+/-1.79) = C3H/HeN (18.0+/-1.10) > DBA/2 (11.4+/-1.16) = DBA/1 (11.0+/-1.20) = Balb/c (11.0+/-1.16). Approximately 80% of the total variation was found to be controlled by genetic factors. No association between the extent of NO formation and variation in the constitutive expression of macrophage IFN-gamma receptor was observed. Similar magnitude of inter-strain differences was sustained after enhanced NO-stimulation of the cells with IFN-gamma + tumour necrosis factor (TNF)-alpha, but only high (strains BL/10, BL/6, SJL, CBA/J, C3H/HeN) and low (DBA/1, DBA/2, Balb/c) NO-responder phenotypes were detected after the triple cytokine cocktail composed of IFN-gamma + TNF-alpha + interleukin (IL)-10. The strain differences remained unchanged after the supplementation of culture medium with L-arginine or tetrahydrobipopterin. Genetically governed differences in IFN-gamma-induced NO production have been found to be tightly associated with differential expression of inducible nitric oxide synthase mRNA. Possible implications of the findings for various fields of NO biomedical research are discussed.

Animals↗
PubMed1998

Lex glycosphingolipids-mediated cell aggregation.

Glycoconjugates bearing oligosaccharide Lex, Galbeta1-->4(Fucalpha1-->3)GlcNAcbeta1-->3R, are found on the surface of several cell types. Although recent studies have indicated that Lexon both glycosphingolipids (GSL) and polylactosaminoglycans can mediate under certain experimental conditions Lex-Lexinteractions, cell-cell interactions based exclusively on LexGSLs have not been demonstrated. In this study we show that preincubation of nonaggregating rat basophilic leukemia (RBL) cells with purified LexGSLs resulted in incorporation of the GSLs into plasma membrane, as determined by immunostaining, and formation of aggregates in the presence of Ca2+; no aggregates were formed after preincubation of the cells with globoside or sphingomyelin. Lex-mediated aggregation was inhibited by removal of Ca2+or by addition of lactofucopentaose III but not by lactose or lacto-N-fucopentaose II. In a mixture of Lex-positive and Lex-negative RBL cells most of the aggregates were composed exclusively of Lex-positive cells. The combined data suggest that interactions between LexGSL on opposite cell surfaces are strong enough to allow formation of stable cell-cell contacts.

Animals↗
PubMed1997

Inhibition of murine macrophage nitric oxide synthase expression by a pivoxil prodrug of antiviral acyclic nucleotide analogue 9-(2-phosphonomethoxyethyl)adenine.

The effect of the acyclic nucleotide analogue, 9-(2-phosphonomethoxyethyl)adenine (PMEA, Adefovir), and its (bis)pivaloyloxymethyl ester (bis-POM-PMEA, Adefovir Dipivoxil) on in vitro nitric oxide (NO) production by murine peritoneal macrophages was investigated. Bis-POM-PMEA inhibited in a concentration-dependent manner the formation of NO generated by interferon-gamma and lipopolysaccharide, the IC50 being 15 microM. Suppressed transcription of mRNA for inducible NO synthase (EC 1.14.13.39) resulting in decreased synthesis of NO synthase protein was found. Parent compound PMEA was virtually ineffective.

Adenine↗
PubMed1996

Carbohydrate-mediated sorting in aggregating embryonal carcinoma cells.

The oligosaccharide structure carried by embryoglycan, Lex hapten, as well as E-cadherin, has been described to mediate Ca(2+)-dependent cell-cell adhesions in embryonal carcinoma cells. To examine the contribution of these two systems to intercellular adhesion, we analyzed aggregation properties of previously isolated embryoglycan-defective mutants of P19 embryonal carcinoma cells. Our data indicate that the absence of Lex and embryoglycan has no effect on homotypic cell aggregation. Pretreatment of the cells with E-cadherin-specific antibody reduced homotypic aggregation of both parental and mutant cells, suggesting that E-cadherin plays a major role in this system. When parental cells were mixed with mutant cells, the aggregates contained either parental or mutant cells; no heterotypic aggregation was observed. The absence of mixed aggregates formed between parental and Lex-or embryoglycan-negative mutant P19 cells suggests that carbohydrates are involved in cadherin-mediated cell sorting.

Animals↗
PubMed1994

Immunological properties of heterozygous nu/+ mice: changes in antibody response and inducibility of tolerance to protein antigens.

Heterozygous nu/+ mice are not fully identical in their immunological properties with the mice of wild +/+ genotype. A colony of nu/nu, nu/+ and +/+ mice from the same breeding nucleus was established and their immune reactivity to human serum albumin, inducibility of adult immune tolerance to hen egg lysozyme (HEL), sensitivity of their lymphoid cells to stimulation by mitogens and ratio of CD3, CD4 and CD8 positive cell populations was studied. Both the numbers of antibody-forming cells in regional lymph nodes and the antibody titres in sera of nu/+ mice were highly variable, between undetectable values of nu/nu and high values of +/+ homozygotes. Intravenous pretreatment with soluble HEL, leading in +/+ mice to a deep hyporeactivity to subsequent immunization with the same antigen, did not decrease the response of nu/+ mice significantly. These results indicate that the immunological alteration of nu/+ mice is not only quantitative and that T cell subpopulations might be differentially modified by the presence of nu allele. The finding of decreased CD4:CD8 ratio in nu/+ mice also supports this idea.

Animals↗
PubMed1993

A modified two-step method for cryopreservation of mouse embryos for purposes of embryo banking.

So far, more than 17,000 embryos have been frozen in the repository of the cryobank in the Institute of Molecular Genetics. The freezing of embryos from more than 20 inbred, congenic and mutant strains has been completed. For purposes of embryo banking, a modified two-step method was used. The first step includes slow freezing, after seeding, to -25 degrees C, rate 0.3 degrees C/min, and is followed by a second step involving direct transfer of the embryos into liquid nitrogen. A relatively high percentage of survival is found (85-95%) after thawing, depending on the strain used. If thawed embryos are cultivated briefly and transferred into recipients, 20-35% living offspring are obtained. Mice born after embryo transfer are subjected to genetic control in order to check the correct procedure of embryo banking.

Animals↗
PubMed1986

Mouse sera contain "natural" antibodies directed exclusively against polymorphic--non-histocompatibility--antigens of the chicken Ea- A blood group system.

Natural agglutinating antibodies in normal sera from A and B10 mice react exclusively with antigenic determinants of the polymorphic Ea-A blood group system (a non-histocompatibility system) of the chicken. Agglutinating activity of these "natural" antibodies, which was demonstrated in the IgM fraction of the serum, did not correlate with the cytotoxic activity against chicken cells in A mice compared to B10 mice. "Natural" agglutinating antibodies against antigens A13 and A14 present in chickens of congenic lines were demonstrated in NMS from A mice. On the other hand, B10 mice showed no antibody reactivity against A13 antigen. The antibody reactions against antigen A13 could be induced in B10 mice by immunization with the respective erythrocytes. In this case, the antibody activity was concentrated in the IgG fraction of the serum. After immunization of B10 mice, agglutinating antibodies against antigenic determinants of the Ea-B (MHC) system appeared, which were not detected in NMS.

Animals↗
PubMed1983

Opposite effects of the synthetic immunomodulator, muramyl dipeptide, on rejection of mouse skin allografts.

The influence of muramyl dipeptide (N-acetylmuramyl-L-alanyl-D-isoglutamine) on the rejection of mouse skin allografts was investigated. While the 0.1-mg dose administered on days 7, 6, 5 prior to transplantation caused significant prolongation of the graft survival, the 0.5-mg dose administered on days 3, 2, 1 prior to transplantation resulted in remarkable augmentation of the graft rejection. The present results support the view that muramyl dipeptide can induce both stimulatory and suppressive immune mechanisms, depending on the treatment regimen.

Acetylmuramyl-Alanyl-Isoglutamine↗
PubMed1983

Alloreactive cytotoxic anti-H-2 antibodies in sera of the syngeneically immunized C57BL/10ScSnPh, BALB/cJPh, and B10.A/SnPh mice.

Mice of the inbred strains B10.A (H-2a), B10 (H-2b), and BALB/c (H-2d) were immunized with the syngeneic cells, and the sera obtained from individual animals on day 10 after the 8th immunizing injection were assayed for the presence of alloreactive cytotoxic antibodies. Anti-H-2Kk antibodies were present in the sera from 50% of the syngeneically immunized BALB/c mice. The syngeneic-immune B10 sera contained weak anti-H-2Dd antibodies in 14% of the immunized animals. B10.A mice produced no antibodies after syngeneic immunization.

Animals↗
PubMed1982

Interstrain differences in the reactivity of normal mouse sera against erythrocyte surface antigens of inbred chicken lines.

Normal mouse sera from non-immunized individuals of different strains gave high titres of "natural" antibodies which agglutinated RBC's of different inbred lines of chickens. Normal mouse sera of A strain and its congenic lines agglutinated RBC's of all chicken lines whereas normal sera of B10 strain and its congenic lines did not react with RBC's of Iowa A line. Normal sera of other mouse strains behaved similarly. The different reactivity of the congenic chicken lines IA and IC, which are identical in the B complex (MHC) and differ only in erythrocyte antigens of the A blood group system, suggests that natural antibodies may also react with antigens other than the chicken MHC antigens. On the other hand, normal sera from chickens of inbred lines reacted with all erythrocytes of the different mouse strains tested.

Alleles↗
PubMed1981

A comparison of the macrophage electrophoretic mobility test and the leukocyte migration inhibition test in the detection of histocompatibility antigens on tumor cells.

Lymph node cells from mice sensitized by rejection of skin allografts were incubated in vitro with tumor cells carrying the sensitizing alloantigens. Products of the lymphocyte--antigen interaction released into the culture medium were simultaneously assessed by the macrophage electrophoretic mobility test and the leukocyte migration inhibition assay. Both macrophage slowing factor (MSF) and a factor which inhibited leukocyte migration (LIF) were produced. The release of both factors was detectable after comparable periods of incubation; their production was transient and ceased within 24 hours after it started.

Animals↗
PubMed1981

Further testing of several strains of mice with respect to detection of H - 2 antigens on neonatal erythrocytes.

In previous experiments the existence of two types of neonatal erythrocytes was confirmed in different inbred and congenic strains of mice. Erythrocytes of the "early" type were agglutinable by the appropriate antisera on the day of birth; the "late" type erythrocytes were not agglutinable until the third day of postnatal life. Results of earlier studies on segregating generations supported the proposed "remote-cis-effect" model for genetic control of H-2 antigen detectability in newborns. We report on tests that bear further upon this phenomenon.

Animals↗
PubMed1979

Macrophage electrophoretic mobility test as sensitive probe of transplantation immunity in mice.

Transplantation immunity was examined in mouse H-2- and non-H-2-disparate strain combinations by the macrophage electrophoretic mobility (MEM) assay. Lymph node cells from the recipients rejecting histoincompatible skin grafts were incubated in the presence of solubilized histocompatibility antigens prepared from spleens of the grafts donors. Product(s) of lymphocyte-antigen interaction in appropriate combinations (sensitized lymphocytes with the sensitizing antigen) significantly diminished the electrophoretic mobility of guinea pig macrophages in all of the histoincompatible strain combinations examined. The data indicate that transplantation immunity in mice can be detected by the MEM assay.

Animals↗