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Biomedical subjects

C H June

Publications and source records attributed to C H June.

At least 181 records · Page 10Linked to original sources
PubMed1986

Antibody binding to CD5 (Tp67) and Tp44 T cell surface molecules: effects on cyclic nucleotides, cytoplasmic free calcium, and cAMP-mediated suppression.

T cells can be activated to proliferate by antibodies to the T cell antigen receptor or the molecularly associated CD3 complex if monocytes are present. We have shown previously that monoclonal antibodies to the human T cell differentiation antigens CD5 (Tp67) and Tp44 each augment and prolong proliferative responses of anti-CD3-activated T cells, even in the absence of monocytes. Here we show that the functional and biochemical mechanisms of CD5 and Tp44 signal transmission are distinct. T cell proliferation is suppressed by agents that increase the concentration of intracellular cAMP. We found that antibody binding to the Tp44 surface molecule overcomes this suppression, whereas antibody binding to CD5 does not, indicating that ligand-Tp44 interaction changes T cell sensitivity to cAMP-mediated growth inhibition. The ability of anti-CD3, anti-Tp44, and anti-CD5 monoclonal antibodies to directly alter cyclic nucleotide levels in the Jurkat T cell line was examined. Anti-CD3 alone caused a rapid four- to sixfold increase in cAMP levels, but did not affect cGMP levels. However, anti-Tp44 and anti-CD5 each caused a rapid three- to fourfold increase in cGMP levels without affecting cAMP levels. In other experiments, cytoplasmic free calcium levels were measured in resting T cells after CD5 or Tp44 stimulation by using the dye indo-1 and flow cytometry. This sensitive method showed that anti-CD5 alone caused an increase in cytoplasmic calcium free levels within 3 min of antibody addition, whereas anti-Tp44 had no effect. Finally, anti-Tp44 and IL 1 each augmented proliferation of phorbol ester-stimulated lymphocytes, whereas anti-CD5 did not. The effects of IL 1 and Tp44 could be further distinguished in that the effect of anti-Tp44 was resistant to inhibition by dBcAMP whereas IL 1 was not. These data suggest that the receptor function of both Tp44 and CD5 involves changes in cyclic nucleotides levels, and that the mechanism by which anti-Tp44 and anti-CD5 antibodies affect T cell proliferative responses may be related to their selective effects on cGMP levels and cytoplasmic calcium concentrations.

Antibodies, Monoclonal↗
PubMed1986

Heterogeneity among T cells in intracellular free calcium responses after mitogen stimulation with PHA or anti-CD3. Simultaneous use of indo-1 and immunofluorescence with flow cytometry.

Measurement of intracellular ionized calcium concentrations ([Ca2+]i) has been indispensable in elucidating the central role of [Ca2+]i as a trigger of cellular responses to activating stimuli. Such studies have employed the dye quin2, which has not been readily adapted to analysis of individual small cells. We show here that the calcium response of large numbers of single cells can be analyzed with the use of flow cytometry and the recently described dye, indo-1. Such analyses demonstrate for the first time the heterogeneous nature of the [Ca2+]i response to mitogenic stimuli within populations of peripheral blood lymphocytes (PBL). By simultaneous quantitation of one- or two-color surface immunofluorescence labels, some of this heterogeneity of [Ca2+]i response in PBL is shown to be related to cellular immunophenotype. Almost all T cells responded to anti-CD3 antibody; however, the response is greater among CD4+ than CD8+ cells, and within the CD4+ population the rate of response to stimulation by antibody to CD3 differed between subpopulations defined by expression of the common leukocyte marker p220. In contrast, not all T cells responded to phytohemagglutinin (PHA), even at very high doses. As with anti-CD3, after stimulation with PHA, CD4+ cells showed a larger proportion of responding cells than did CD8+ cells. In separate experiments, indo-1 was found not to impair reproductive viability of PBL, thereby providing the potential for analysis of functional activity after the separation of cells by sorting on the basis of the [Ca2+]i response to stimuli. Mixing experiments indicated that a response of a subpopulation representing as little as 0.3% of total cells could be readily detected. Thus, the flow cytometric assay with indo-1 is the first technique that allows the quantitative analysis of response differences of small subpopulations of cells and intercellular variation in [Ca2+]i.

Antibodies, Monoclonal↗
PubMed1986

Valency of CD3 binding and internalization of the CD3 cell-surface complex control T cell responses to second signals: distinction between effects on protein kinase C, cytoplasmic free calcium, and proliferation.

We have studied the relationship of valency of CD3 stimulation and modulation of the CD3 receptor complex with biochemical and proliferative responses of T cells. Anti-CD3 Fab, as well as F(ab')2 and whole antibody caused rapid modulation of the CD3 antigen, whereas anti-CD3 conjugated to Sepharose did not. In the absence of monocytes, T cells stimulated with anti-CD3 Fab, F(ab')2, or F(ab')2-Sepharose showed differences in their ability to respond to second signals given by PMA, IL 1, IL 2, or antibodies to Tp67 and Tp44. None of the anti-CD3 signals alone caused resting T cells to produce IL 2, and only the Sepharose-bound anti-CD3 F(ab')2 caused T cells to express high levels of functional IL 2 receptors. Anti-CD3 F(ab')2-Sepharose-stimulated T cells produced IL 2 and proliferated in response to each of the second signals. Because anti-CD3-Sepharose did not cause modulation of the CD3 antigen, the ability of the Sepharose-bound antibody to induce T cells to express IL 2 receptors and to respond to individual second signals may be related to lack of modulation rather than valency of binding. Anti-CD3 Fab-stimulated T cells responded to PMA but required combinations of other second signals. T cells stimulated with unmodified anti-CD3 antibody or F(ab')2 fragments responded to PMA but did not respond to any other second signals alone or in combination. Stimulations that resulted in modulation (i.e., anti-CD3 whole antibody, anti-CD3 F(ab')2, or anti-CD3 Fab fragments) caused an increase in cytoplasmic calcium levels in resting T cells but blocked proliferation of T cells in response to mitogenic lectins or CD2 stimulation. Anti-CD3 F(ab')2 on Sepharose, however, did not block T cell proliferation. Whole bivalent anti-CD3 antibody or F(ab')2 fragments, but not monovalent Fab fragments, caused a rapid translation of protein kinase C activity from cytosol to membrane in the Jurkat T cell line. Because all of these modulate the receptor, these data indicate that the functional difference between monovalent and bivalent binding to CD3 is related to antibody valency and not to antigenic modulation. The use of Fab anti-CD3 stimulation that requires combinations of second signals for proliferation allowed an analysis of the functional relationships between IL 1, anti-Tp67, and anti-Tp44.

Animals↗
PubMed1986

A 44 kilodalton cell surface homodimer regulates interleukin 2 production by activated human T lymphocytes.

We previously described a cell surface antigen, termed Tp44, detected by monoclonal antibody 9.3 on approximately 80% of mature human T lymphocytes. Analysis by SDS-polyacrylamide gel electrophoresis and isoelectric focusing demonstrated that this antigen consists of two identical 44 kilodalton glycopeptides that form a disulfide-linked homodimer. Competitive binding experiments showed that antibody 9.3 and an anti-CD3 antibody (64.1) recognize distinct antigenic determinants; furthermore, the binding of antibody 9.3 was unaffected by prior modulation of CD3. Thus, Tp44 has no detectable cell surface association with CD3. By itself, antibody 9.3 had no detectable effect on either IL 2 receptor expression or IL 2 release, and did not cause T cell proliferation even when monocytes were present and exogenous IL 2 was provided, indicating that binding of antibody 9.3 does not provide a primary signal for T cell activation. However, the proliferative responses of T lymphocytes activated by phytohemagglutinin, concanavalin A, or an anti-CD3 monoclonal antibody were strikingly enhanced in the presence of antibody 9.3, an effect associated with increased IL 2 receptor expression and increased IL 2 secretion. Antibody 9.3 enabled anti-CD3-Sepharose-activated T cells and anti-CD3 antibody-activated Jurkat cells to release IL 2 in the absence of monocytes. Fab fragments of antibody 9.3 had no effect on anti-CD3-induced IL 2 release by Jurkat cells, whereas F(ab')2 fragments had activity comparable to that of unmodified antibody, indicating that bivalent binding of Tp44 molecules is required for IL 2 secretion. Together, these results suggest that TP44 may function as a receptor for accessory signals in the activation of T cells.

Animals↗
PubMed1986

Correlation of hypomagnesemia with the onset of cyclosporine-associated hypertension in marrow transplant patients.

Cyclosporine is known to cause hypertension, and we have recently reported that it causes hypomagnesemia and renal magnesium wasting in marrow transplant recipients. We performed a case-control study to ask whether hypomagnesemia might be related to this form of drug-induced hypertension. The charts of 188 patients treated with cyclosporine were evaluated for the development of hypertension. The 32 patients who became hypertensive were age, sex, and disease-matched with 32 cyclosporine-treated controls. Baseline serum Mg levels were normal in both groups. However at the time of development of hypertension, the hypertensive patients had a mean (+/- SD) Mg of 1.22 +/- 0.20 mEq/L versus controls 1.40 +/- 0.33 mEq/L (P less than 0.01). Serum calcium, albumin, creatinine, potassium, and cyclosporine concentrations were not different between the two groups. This study may indicate that hypertension and hypomagnesemia are coincident toxicities in cyclosporine-treated patients. Alternatively, our data support the hypothesis that acquired derangements in magnesium metabolism may contribute to the development of hypertension. Magnesium replacement may prove beneficial in the treatment and/or prevention of cyclosporine-associated hypertension.

Adolescent↗
PubMed1986

Distinct patterns of transmembrane calcium flux and intracellular calcium mobilization after differentiation antigen cluster 2 (E rosette receptor) or 3 (T3) stimulation of human lymphocytes.

We evaluated CD2 (E rosette) and CD3 (T3)-triggered activation of resting lymphocytes by measuring the intracellular free calcium concentration ([Ca2+]i) of individual cells. The [Ca2+]i of indo-1-loaded cells was measured by flow cytometry and responses were correlated with cell surface phenotype. Stimulation with anti-CD3 antibody caused an increase in [Ca2+]i in greater than 90% of CD3+ cells within 1 min, and furthermore, the response was restricted to cells bearing the CD3 marker. In contrast, stimulation of cells with anti-CD2 antibodies produced a biphasic response pattern with an early component in CD3- cells and a late component in CD3+ cells. Thus, the CD2 response does not require cell surface expression of CD3. In addition, stimulation of a single CD2 epitope was sufficient for activation of CD3- cells, whereas stimulation of two CD2 epitopes was required for activation of CD3+ cells. Both the CD2 and CD3 responses were diminished in magnitude and duration by EGTA. However, approximately 50% of T cells still had a brief response in the presence of EGTA, indicating that the increased [Ca2+]i results in part from intracellular calcium mobilization, and furthermore demonstrates that extracellular calcium is required for a full and sustained response. Our results support the concept that CD2 represents the trigger for a distinct pathway of activation both for T cells that express the CD3 molecular complex and for large granular lymphocytes that do not.

Antibodies, Monoclonal↗
PubMed1986

Monoclonal antibody-defined functional epitopes on the adhesion-promoting glycoprotein complex (CDw18) of human neutrophils.

We have evaluated the functional and immunochemical activities of three monoclonal antibodies (MoAbs) minimally reactive with adherence-defective neutrophils (PMN) from a patient with recurrent bacterial infections. In studies with normal PMN, MoAbs OKM1 and 60.1 both precipitate the same 165kd alpha-subunit (alpha M) within an alpha-beta heterodimer complex (CD11). The CD11 complex is part of a larger complex composed of four glycoproteins (CDw18) precipitated by MoAb 60.3, with properties suggesting that the CDw18 complex is equivalent to the Mac-1, LFA-1, p150, 95 glycoprotein family implicated in adherence-dependent leukocyte functions. PMN adherence to endothelium, spreading on surfaces, aggregation, and phagocytosis of zymosan particles were all inhibited in a dose-dependent fashion by MoAb 60.1 (analogous to previous studies with MoAb 60.3) while MoAb OKM1 had no effect. These findings unify previously disparate observations and suggest that a functionally active site on the adherence promoting glycoprotein complexes CD11 and CDw18 is distant from the alpha M epitope recognized by MoAb OKM1 but closely associated with the alpha M epitope recognized by MoAb 60.1 and the beta-epitope (or epitope created by alpha-beta quaternary structure) recognized by MoAb 60.3.

Antibodies, Monoclonal↗
PubMed1985

Profound hypomagnesemia and renal magnesium wasting associated with the use of cyclosporine for marrow transplantation.

We prospectively studied 41 marrow transplant patients to evaluate the possible association of hypomagnesemia with cyclosporine immunosuppressive therapy. During the 3 months posttransplant the mean nadir serum magnesium level was 1.06 +/- 0.16 mEq/L in 24 patients treated with cyclosporine and 1.33 +/- 0.13 in 14 patients treated with methotrexate (P less than 0.0001). Eleven of 24 patients receiving cyclosporine versus 1 of 14 patients receiving methotrexate had magnesium levels less than 1 mEq/L, or were begun on replacement therapy for presumed symptomatic hypomagnesemia (P less than 0.02). Agents known to be associated with hypomagnesemia, aminoglycosides and amphotericin B, were given in both lesser quantity and frequency to cyclosporine-treated patients than to methotrexate-treated patients. Hypomagnesemic patients treated with cyclosporine had inappropriately elevated urine magnesium excretion. Renal magnesium wasting may be added to the spectrum of nephrotoxicity resulting from cyclosporine. Several adverse reactions previously attributed to cyclosporine may be secondary to magnesium deficiency.

Adolescent↗
PubMed1985

Ultrasonography and computed tomography in severe urinary tract infection.

A prospective study evaluated the utility of renal computed tomography (CT) and ultrasonography in 35 patients hospitalized for treatment of urinary tract infection. Renal computed tomograms were abnormal in 18 of 28 patients with acute pyelonephritis and three of four patients with urosepsis, showing findings consistent with pyelonephritis in 17 patients and intrarenal abscess or focal bacterial nephritis in four patients. Renal sonograms were abnormal in only eight patients, showing findings compatible with pyelonephritis in four and intrarenal abscess or focal bacterial nephritis in the other four. Flank tenderness was absent in only four patients with CT findings of pyelonephritis, of whom three were diabetic. We therefore found that (1) renal CT is a sensitive test for acute upper urinary tract infection, (2) ultrasonography detects focal bacterial nephritis and abscesses but is insensitive to uncomplicated upper urinary tract infection, and (3) painless pyelonephritis may be more common in patients with diabetes mellitus.

Abscess↗
PubMed1984

Association between cyclosporin neurotoxicity and hypomagnesaemia.

The serum magnesium levels of all 12 allogeneic bone-marrow transplant recipients who experienced the neurotoxic effects of cyclosporin (CyA) were more than two standard deviations below the normal range. The neurological events seemed to segregate into three separate syndromes. 7 patients had grand-mal seizures, which occurred within the first several weeks of CyA therapy (median onset 12 days). At the time of their first seizure all 7 patients had hypomagnesaemia, which had developed rapidly over the preceding 1-3 weeks. 3 patients had four episodes of cerebellar ataxia, tremor, and depression. These subacute episodes developed after prolonged CyA therapy (median onset 67 days). Each episode was associated with hypomagnesaemia. 2 patients had a transient episode of expressive aphasia following a long period of hypomagnesaemia. In all cases symptoms resolved or did not recur with adequate magnesium replacement. These data suggest that CyA neurotoxicity is associated with hypomagnesaemia and may be treated or prevented with magnesium replacement.

Adolescent↗
PubMed1980

Immunopathological aspects of Plasmodium berghei infection in five strains of mice. I. Immune complexes and other serological features during the infection.

The development of circulating immune complexes was studied in mice of the BALB/c, A/J, OF1, CBA and C57B1 strains infected with P. berghei. Complexes were evaluated in relation to levels of parasitaemia, soluble antigen, specific antibody and C3. Susceptibility to infection was greatest in BALB/c, A/J and OF/a mice. The maximum parasitaemia was 30% in CBA and 70% in all other strains. Levels of soluble antigen paralleled those of parasitaemia. Specific antibody was detected in all strains, but the titre continued to rise throughout the infection only in CBA mice. Circulating immune complexes occurred in mice of all strains from day 6; the level fell after day 9 in C57B1 whereas it was maintained in CBA mice. The development of immune complexes was associated with marked depression of C3 levels in all except CBA mice, in which a transient reduction was followed by recovery. Partial characterization of the complexes showed that IgM-containing complexes appeared earliest and reached highest levels in BALB/c mice while in CBA mice, IgM complexes were found in lesser amounts and the level fell in late infection. IgG complexes rose throughout infection in CBA and fell in later stages in BALB/c and C57B1 mice. In nude BALB/c mice, immune complexes were usually not detectable and only low levels of antibody of IgM class were produced. Differences in mortality pattern could not be related to any single serological factor.

Animals↗
PubMed1980

Immunopathological aspects of Plasmodium berghei infection in five strains of mice. II. Immunopathology of cerebral and other tissue lesions during the infection.

Histological changes during the course of P. berghei infection were investigated in A/J, BALB/c, OF1, CBA and C57B1 mice. The findings were studied in relation to serological aspects (Contreras et al., 1980). High mortality and acute deaths occurred in A/J, BALB/c and OF1 mice and marked cerebral lesions were found in these strains from day 15, including congestion of meningeal and cerebral veins and capillaries, blocking of these vessels by heavily parasitized RBC, cerebral oedema and haemorrhages. Such lesions were minimal in CBA and C57B1 mice, and absent in mice examined 21 and 24 days after infection. Small deposits of IgG and traces of C3 were detected by immunofluorescence in the choroid plexus of most mice from day 9. Renal lesions included congestion, plugging of veins and capillaries, low-grade mononuclear infiltration and mesangial thickening; these changes were most marked in CBA, C57B1 and A/J mice. Glomerular deposits of IgM were present in all strains in the first week of infection. IgG and C3 were detected in the second week, but only traces were found in CBA mice. The livers showed congestion, accumulation of pigment in swollen Kupffer cells and mononuclear portal infiltration; these were most pronounced in A/J mice. In the spleen, there was a great increase in the reticuloendothelial cell population, white pulp proliferation, congestion and accumulation of pigment and plasma cell reaction; the pattern of white pulp expansion varied in the different strains. The results suggest that cerebral lesions play a significant role in the aetiology of acute deaths in this malaria model.

Animals↗
PubMed1979

Circulating and tissue-bound immune complex formation in murine malaria.

Immune complex formation during Plasmodium berghei infection of OF1 mice was investigated. Circulating immune complexes (CIC) were detected by the Clg-binding assay and the conglutinin-binding solid-phase assay in lethal or drug-limited infections. CIC appeared on day 9 of infection, peaked on day 11, and disappeared only after complete cure of the infection. Analysis of the immune complexes detected by the Clq-binding assay revealed the following characteristics: sedimentation coefficients of 13S to 21S, resistance to DNAse, and selective removal by filtration through protein A bound to Sepharose. Glomerular deposits of IgM preceded the appearance of CIC, whereas deposits of IgG and C3 were concomitant with the appearance of CIC. Tissue-bound immunoglobulins were also found in the choroid plexus. The appearance of anti-malarial antibodies and malarial antigens in the serum was closely associated with a depression of C3 levels and the presence of CIC. Drug treatment was followed by normalization of C3 levels, and clearance of both CIC and malarial antigens.

Animals↗