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Biomedical subjects

F Kristensen

Publications and source records attributed to F Kristensen.

At least 37 records · Page 2Linked to original sources
PubMed1984

Lymphokine regulation of human lymphocyte proliferation: formation of resting G0 cells by removal of interleukin 2 in cultures of proliferating T lymphocytes.

The question of whether lymphocytes which have once been activated and have completed one or several cell cycle(s) can return to the G0 phase and stay ready for a new activation (G0-G1 transition), rather than simply die, was investigated. To do so interleukin 2 (IL-2) was removed from cultures of continuously proliferating human T lymphocytes and the formation of resting (G0) cells was measured. Kinetic analyses in freshly prepared peripheral blood lymphocytes (PBL) revealed that the onset of detectable RNA synthesis and the appearance of structures binding the anti-Tac antibody occurred simultaneously. This allowed the expansion of the definition of G0 T lymphocytes as cells having a low RNA (and DNA) content, and no Tac antigen. When cultured human T cells proliferating continuously by means of IL-2 were characterized in terms of their distribution in the cell cycle, 7 days after the initial PHA stimulation, it could be demonstrated that very few cells were in the G0 phase, supporting the concept of direct S/G2/M-G1 transition. However, when IL-2 was removed from the cultures, the [3H]thymidine incorporation per 10(4) cells and correspondingly the number of cells in the S/G2/M and G1 phases were reduced drastically and during the following 72-hr period, the number of G0 cells increased markedly. Restimulation of such in vitro formed G0 cells, under conditions permitting observation of their shift from the G0 to G0 phase, demonstrated that most cells could respond normally. Based on these observations, it was concluded that IL-2 not only ensures T-lymphocyte survival and proliferation, but IL-2 starvation induces many continuously proliferating T lymphocytes to stop cycling and to return to the G0 phase of the cell cycle where they remain functional.

Antibodies, Monoclonal↗
PubMed1984

Interleukin 1- and 2-like activities in the dog.

Canine adherent and non-adherent peripheral blood leukocytes and spleen cells were examined for their ability to produce soluble factors with Interleukin 1- and 2- (IL-1 and IL-2) like activities. For this purpose, three conventional assay systems were used: (a) proliferation on an IL-2-dependent murine cytotoxic T-lymphocyte cell line, (b) enhancement of PHA-induced murine thymocyte proliferation and (c) proliferation of lectin-primed canine peripheral blood lymphocytes (PBL). Only the latter two types of cells respond to IL-1, whereas all three types respond to IL-2. Both types of factors were produced and the kinetics of their release/production were found to be identical to those of human PBL. Results suggested that species-related differences existed. Canine interleukin-containing supernatants had higher titers than murine interleukin-containing supernatants when analyzed on canine lymphocytes, and the reverse was found if murine target cells were used.

Animals↗
PubMed1984

Cell cycle-related changes in number of T-lymphocyte receptors for glucocorticoids and insulin.

Enriched human peripheral T-lymphocytes were stimulated with PHA and examined for variations in insulin and glucocorticoid (dexamethasone) receptor numbers during the early phases of the cell cycle. Cells in G0, G1a and G1b phases, where the G1a - G1b transition is an Interleukin 2 dependent event, were quantitated by flow cytometry. Few but significant numbers of glucocorticoid receptors (2700/cell) and no insulin receptors (-1/cell) were found in the resting (G0) phase. As cells entered the G1a phase the specific binding of dexamethasone increased and of insulin took place. Although the specific binding further increased as T-cells entered the G1b phase (as measured at 44 h of incubation and using hydroxyurea-treated cells), the major changes in the specific binding of dexamethasone took place during the period 16 - 20 h after stimulation. Based on these findings, it is concluded that both receptor types (cell membrane and cytoplasmic receptors) are being formed and increased at G1 phase prior to cell proliferation, indicating the importance of G1 phase in immunoregulation.

Adult↗
PubMed1984

Lymphokine regulation of activated (G1) lymphocytes. II. Glucocorticoid and anti-Tac-induced inhibition of human T lymphocyte proliferation.

The regulation of the first cell cycle of human, activated (G1) PBL was analyzed by flow cytometry and [3H]thymidine incorporation. Endogenous IL 2 production was blocked in situ by pharmacologic concentration of DEX (100 to 1000 nM), resulting in an 80 to 90% reduction of thymidine uptake. Although T lymphocyte activation (G0-G1a transition) by PHA was unaltered, cells remained in the G1a phase of the cell cycle due to insufficient RNA synthesis for proliferation. The addition of IL 2-containing supernatants reversed this inhibitory effect of DEX by allowing the cells to synthesize more RNA (G1a-G1b transition). Such cells could enter the S phase and proliferate. Similar studies were performed on cells treated with a monoclonal antibody (anti-Tac) against the IL 2 receptor. In these studies, IL 2-induced RNA synthesis, and subsequent proliferation of DEX-treated and PHA-stimulated cells was inhibited by anti-Tac. Anti-Tac did not, however, inhibit the effect of endogenous IL 2 (PHA-stimulated PBL without DEX treatment), although it did bind equally well to such cells. Thus, IL 2 directly or indirectly regulates human T cell proliferation at the level of RNA synthesis. Furthermore, anti-Tac can inhibit the mitogenic signal given by endogenous IL 2, but not by in situ produced IL 2, an observation of importance to further investigations of the mechanisms by which IL 2 interacts with specific receptors to elicit proliferation.

Adult↗
PubMed1983

A simple and rapid flow cytometric method for routine assessment of baker's yeast uptake by human polymorphonuclear leukocytes.

A new method for measuring uptake of baker's yeast (BY) by human polymorphonuclear leukocytes (PMN) using flow cytometry is described. The method correlates excellently with the visual method, is reproducible and provides a means for investigating the early phases of the phagocytic process as well as the phagocytic capacity of PMN. This quick and accurate method allows the counting of large numbers of cells, and monitoring of the process of particle uptake and has a considerable potential in the routine assessment of polymorph function in various clinical situations.

Acridine Orange↗
PubMed1983

Biological and biochemical characterization of an interleukin 1-like factor from rat C6 glioma cells.

It has been previously reported that lipopolysaccharide-stimulated murine astrocytes produce a factor which enhances the proliferative response of thymocytes to lectins. The present report demonstrates that a rat astrocytoma cell (C6 cells)-derived factor appears to be similar to macrophage-derived interleukin 1 (IL 1) in its biological activities and biochemical characteristics. Upon injection into mice, supernatants of C6 cells induce the production of serum amyloid A. The C6 cell-derived factors enhance the response of thymocytes to phytohemagglutinin and the growth of fibroblasts whereas no effect on the growth of neuroblasts and of a strictly interleukin 2 (IL 2)-dependent T cell line was observed. On an AcA 54 column the C6-derived factors acting on thymocytes and fibroblasts coeluted as a single peak of Mr = 13 500 to 18 000; the semipurified factor was found to enhance the lymphocytes' production of IL 2. These observations demonstrate that cells not belonging to the mononuclear phagocyte lineage are able to produce factors identical or closely related to IL 1.

Animals↗
PubMed1983

Glial proteins in canine distemper virus-induced demyelination. A sequential immunocytochemical study.

A temporal series of demyelinating lesions in experimental canine distemper virus (CDV) infection was examined with immunohistological techniques demonstrating myelin basic protein (MBP), myelin-associated glycoprotein (MAG), and glial fibrillary acidic protein (GFAP) on serial sections. The earliest lesions were characterized by decreased MBP and MAG and increased GFAP. During the further progression of the disease, MBP and MAG losses continued to match each other. There was no indication of MAG loss preceding the disappearance of MBP. In the more advanced lesions there was a marked decrease of GFAP positive cells. Since these findings differed considerably from similar immunohistochemical studies in progressive multifocal leukoencephalopathy (PML) where demyelination results from oligodendroglial infection, it was concluded that the oligodendroglial cell body is not the primary target of CDV. The marked astroglial changes were also considered to contribute to demyelination in CDV infection but the mechanism by which this happens remains unknown.

Animals↗
PubMed1983

Lymphocyte responsiveness to lectin and myelin antigens in canine distemper infection in relation to the development of demyelinating lesions.

Eleven dogs were experimentally infected with canine distemper virus and studied for periods of up to 63 days post-inoculation. The responsiveness of lymphocytes in vitro toward phytohemagglutinin, myelin basic protein and galactocerebroside was tested at regular intervals during the course of infection by means of [3H]thymidine incorporation and flow cytometry. All dogs developed a marked decrease of lymphocyte responsiveness toward phytohemagglutinin. Four dogs recovered rapidly from the immunosuppression and did not develop demyelination or had only mild lesions, while two others failed to recover at all and developed severe non-inflammatory demyelinating lesions. The remaining dogs exhibited a slow or partial immune recovery and had various degrees of inflammatory demyelination. Lymphocytes from 2 dogs with demyelination and 2 dogs without myelin lesions responded to myelin antigens. The findings indicate that the degree of immunosuppression in canine distemper virus infection may determine the type of demyelination and autoimmune reactions that occur during the inflammatory stage of demyelination may be epiphenomena.

Animals↗
PubMed1983

Lymphokine regulation of activated (G1) lymphocytes. I. Prostaglandin E2-induced inhibition of interleukin 2 production.

PGE2-induced inhibition of the proliferatory response of PHA-stimulated human PBL and Con A-stimulated murine thymocytes was analyzed by flow cytometry. It was found that the activation process (G0-G1a transition) was not influenced by PGE2 over a wide range of concentrations (10(-10) to 10(-6) M), nor was the formation of IL 2 receptors inhibited. Similarly, the viability of human lymphocytes was practically unaltered. In contrast, the IL 2-dependent cell cycle event (G1a-G1b transition), which is required for proliferation, was inhibited in a dose-dependent fashion. The addition of IL 2-containing supernatants to such cultures prevented the PGE2-mediated block in the G1a phase and reconstituted a normal lymphocyte proliferation. Furthermore, lower IL 2 titers were measured in supernatants from PHA-stimulated human PBL treated with PGE2. These findings strongly suggest that PGE2 primarily exerts its inhibitory effect on lymphocyte proliferation through an inhibition of IL 2 production.

Adult↗
PubMed1983

Analysis of 3H-histamine interaction with lymphocytes: receptor binding or uptake?

The immunoregulatory role of histamine is presumably mediated by specific receptors on the plasma membrane of lymphocytes. However, using murine spleen cells and a whole cell assay commonly applied in hormone receptor studies, specific histamine receptors with an affinity higher than that of non-specific binding could not be identified. Nevertheless, approximately 30% of the totally bound histamine was undissociable over a range of added histamine concentrations (9 X 10(-6)-1 X 10(-2) M). Lectin stimulation of spleen cells caused an additional two-fold increase of undissociable histamine. The H1 receptor antagonist, diphenhydramine, blocked histamine uptake, whereas the H2 receptor antagonist, cimetidine, had no effect. Binding experiments carried out at 4 degrees C demonstrated that the amount of undissociable histamine was much reduced. Even at 4 degrees C, evidence for specific membrane associated histamine receptor could not be obtained. It was therefore concluded that lymphocytes take up histamine by an energy-dependent mechanism inhibitable by diphenhydramine but not cimetidine, and that the usual hormone receptor methodology did not allow the identification of specific membrane associated histamine receptors.

Animals↗
PubMed1982

Human lymphocyte proliferation. II. Formation of activated (G1) cells.

The kinetics of phytohemagglutinin (PHA)-stimulated human peripheral blood lymphocytes in the early phases (G0, G1a, G1b) of the first cell cycle have been analyzed in vitro by cytofluorometry and [3H]thymidine incorporation. Cells were detected in the G1a, G1b and S phases 12-14, 20-22 and 24-26 h after stimulation, respectively. The total number of PHA-induced G1 cells reached a plateau after 32 h of incubation, upon which a second increase followed, 10 h later. The latter increase was considered to be a result of cells initiating their second cell cycle, since it could be abolished by hydroxyurea. By examination of supernatants, interleukin 2 (IL 2) activities could be recovered 6-10 h after PHA stimulation. In contrast, the monokine IL 1 was already detected after 2 h. Both interleukins were present in the culture medium before the major G1a cell formation took place. The titer of free IL 2 increased to a maximum after 18-22 h, whereafter a decline was observed. This decline was less pronounced if cultures were treated with hydroxyurea. Finally, among individual donors, the total number of lymphocytes entering the G1 phase and the time at which the G1a-G1b transition took place varied. Lower numbers of G1 cells and delayed G1a-G1b transition coincided with lower IL 2 titers and delayed occurrence of maximal titers in the supernatants.

Adult↗
PubMed1982

Immunological and pathological findings in demyelinating encephalitis associated with canine distemper virus infection.

Nine dogs with canine distemper encephalitis (CDE) were examined with immunological techniques including demonstration of antibodies against canine distemper virus (CDV) in the serum and against myelin basic protein (MBP) in serum and in CSF. Mitogen stimulation tests of lymphocytes were also done. The brains were examined pathologically and immunoglobulin and C3 were demonstrated in lesions by means of immunohistological techniques. Six dogs with acute CDE had none or low antibody levels against CDV or MBP, and there was no immunoglobulin in demyelinating lesions. Some of these dogs had depressed lymphocyte mitogen responses. Two dogs with chronic CDE showed recovery of lymphocyte mitogen responses. One of these had a significant antibody response against CDV and MBP in the serum. Both dogs with chronic CDE had very high antibody titers against MBP in the CSF and demyelinating lesions contained immunoglobulin. These results suggest that acute demyelination in CDE is probably due to some direct viral activity and that the progression of demyelination in chronic CDE is associated with a local immune response.

Animals↗
PubMed1982

Canine lymphocyte cultures in vitro: evaluation of peripheral blood lymphocyte response to mitogens.

Lymphocytes from dog peripheral blood have been stimulated in vitro with 3 different mitogens (Con A, PHA and PWM). Culture medium was RPMI 1640 enriched with either autologous plasma, fetal calf serum of a newly described defined serum substitute. In such cultures the number of surviving and activated cells was measured by cytofluorometry and the proliferation was assessed by thymidine incorporation. In unstimulated cultures, up to 70% of all cells had disappeared (died) during the first 42 hours of incubation, whereas the number of viable cells was reduced to 50-60% in mitogen stimulated cultures. Of the surviving lymphocytes, between 25-40% of the cells appeared to have an elevated RNA-content (activated or G1 cells). By comparison between thymidine incorporation and number of mitogen induced G1 cells, a very high correlation was found (r=0.92). However, the Slope of the regression line was much lower than expected. The low thymidine incorporation per activated cell was primarily related to the high cell death and a resulting dilution of tritiated thymidine. Indeed, preliminary results suggested that the same thymidine incorporation per G1b cells could be obtained if peripheral blood lymphocytes were washed immediately before pulsing as could be obtained with lymph node cells without washing.

Animals↗
PubMed1982

Technical aspects of low 3H-thymidine incorporation by mitogen stimulated canine peripheral blood lymphocytes in vitro.

The value of [3H]-thymidine incorporation as a measurement for mitogen induced proliferation of dog peripheral blood lymphocytes (PBL) has been examined. The cells were cultured in RPMI 1640, enriched with 10% autologous plasma for 48 hours at 37 degrees C, 5% CO2 and 95% relative humidity. Under these conditions a great variability in [3H]-thymidine incorporation was observed. By analysis of CPM and number of activated cells (G1), it was found that comparable number of G1 cells were generated in human and dog PBL. Also, the membrane transport of thymidine was very similar for lymphocytes of the two species. Nevertheless, a low [3H]-thymidine incorporation by dog PBL was frequently seen, and this phenomenon could be related to a release of soluble substance(s) within the cultures. When the cultured cells were washed and resuspended in fresh medium immediately before pulsing, the expected CPM per G1 cell could be obtained. Since it has been described in the literature that macrophages can produce cold thymidine in macrophage enriched lymphocyte cultures, the in vitro response of non-adherent dog PBL was analyzed. Mitogen stimulation of such non-adherent cells resulted in CPM per G1 cells very similar to those obtained with washed cells. Based on these data, it is suggested that the production of cold thymidine might be one of the technical problems related to cultures of lectin stimulated dog PBL in vitro and it should be taken into consideration, if [3H]-thymidine incorporation is used as the only measure of lymphocyte proliferation.

Adult↗