CFU-c suppressor T cells were generated in vitro by culturing overnight peripheral blood T cells from healthy donors with pokeweed mitogen (PWM), or T cells from patients with severe aplastic anemia (SAA) in remission, with culture medium (RPMI). The supernatants were removed the next morning, the cells harvested and washed, and both tested for CFU-c suppression on normal marrow cells. Cyclosporin A (CyA) was added to this system to test whether it could abrogate or prevent the generation of suppressor cells. CyA was incubated with T cells, at a concentration of 0.1 microgram/ml, for 30 min at 37 degrees C and then washed away, in two different assays: (a) before T cells were incubated overnight in culture medium with or without PWM, or (b) after T cells had been kept overnight in culture. The results of this study indicate that CyA can prevent the generation of CFU-c suppressor T cells if preincubated with both normal or SAA T cells prior to in vitro priming, whereas it cannot abrogate the suppressor activity of primed T cells.

Anemia, Aplastic↗

PubMed1983

Mepartricin: a new antifungal agent for the treatment of disseminated Candida infections in the immunocompromised host.

Mepartricin, a new semisynthetic heptene, was given for systemic Candida infections to 11 immunocompromised patients. All patients were undergoing bone marrow transplantation or treatment with antilymphocytic globulin: 6 had acute leukemia, 4 aplastic anemia and 1 had Wiskott-Aldrich syndrome. In all patients systemic Candida infection was diagnosed on the basis of fungemia or positive cultures from at least three different sites, two of which were outside the gut. Mepartricin was given intravenously as a slow drip, at the dose of 100-700 U/kg/day for a total of 198 patient-days (range 9-36, mean +/- SD 18 +/- 8). Patients had to be premedicated with steroids and antihistamines to avoid reactions such as chills and fever. The treatment was well tolerated in all patients, and renal function tests were unchanged during therapy. There was a complete resolution of the fungal infection in 10 of the 11 patients.

Adolescent↗

PubMed1982

GM-CFC growth in chronic granulocytic leukaemia is not affected by a soluble inhibitor released by aplastic anaemia T-cells or mitogen-primed normal T-lymphocytes.

Peripheral blood (PB) and bone marrow (BM) cells were obtained from 12 patients with chronic granulocytic leukaemia (CGL) and cultured in agar for granulocyte macrophage colony formation (GM-CFC). In addition PB and BM CGL cells were co-cultured with T-lymphocytes from patients with immune severe aplastic anaemia (SAA), or with T-cells from healthy donors primed with pokeweed mitogen (PWM). The supernatants of SAA and PWM primed T-cells were also added to CGL cells grown in agar. Normal marrow cells obtained from eight healthy donors were used to set up control cultures and co-cultures. The results of this study indicate that, firstly, T-cells derived from SAA patients and their supernatants suppress GM-CFC growth of normal marrow cells but not of PB nor BM CGL cells, and, secondly, normal T-cells primed with PWM and their supernatants suppress normal marrow GM-CFC but not colony formation of BM nor PB CGL cells. These results provide further evidence that mediators which are effective in regulating normal myeloid progenitor cells fail to inhibit the in vitro growth of GM-CFC from CGL patients.

Adult↗

PubMed1982

Generation of CFU-C suppressor T cells in vitro. V. A multistep process.

Different cell fractions obtained from five patients with immune severe aplastic anaemia (SAA) in complete autologous haematologic reconstitution were tested for CFT-c suppression. Bone marrow mononuclear cells (BMMC), but not peripheral blood mononuclear cells (PBMC), showed definite CFU-c inhibitory activity. On the contrary, both peripheral blood and marrow E rosetting cells (E+) suppressed CFU-c growth. The suppressor activity of PBE+ cells could not be rescued by adding back PBE- cells and/or PB adherent cells (AC). In addition, unfractionated PBMC exposed to sheep red blood cells (SRBC) suppressed CFU-c growth. PBMC from normal donors exposed to SRBC had no suppressor activity. This study suggests that CFU-c suppressor T cells in the peripheral blood of SAA patients are in different activation state as compared to BM cells from the same patients, and also differ from normal PB cells. The identification of T cells with different requirements for in vitro activation in order to exhibit a suppressor activity, suggests that generation of suppressor cells is a multistep process, and this may have practical implications for in vitro assays designed to test for immune suppression of haematopoiesis.

Anemia, Aplastic↗

PubMed1982

Generation of CFU-C suppressor T cells in vitro. III. Failure of mitogen-primed T cells from patients with chronic granulocytic leukemia to inhibit the growth of normal CFU-C.

T lymphocytes were derived by E rosetting from the peripheral blood (PB) and bone marrow (BM) of 15 patients with chronic granulocytic leukemia (CGL) in the chronic phase of their disease. T cells were also obtained from 12 healthy individuals. T cells were incubated overnight either in culture medium (RPMI) or RPMI plus pokeweed mitogen (PWM). The supernatants were then recovered and the cells washed in fresh RPMI. T cells from normal donors and from CGL patients were then cocultured with normal allogeneic marrow cells grown in soft agar for CFU-C colony formation. Target marrow cells were also grown in agar in the presence of T-derived supernatants. The results of this study can be summarized as follows. (1) Normal PB and BM T cells efficiently suppressed autologous and allogeneic CFU-C growth after PWM stimulation. (2) T cells derived from peripheral blood or marrow of CGL patients failed to inhibit CFU-C growth, whether pretreated with PWM or not. (3) The supernatants of PWM-treated normal T cells strongly inhibited CFU-C colony formation, whereas the supernatants of PWM-treated CGL T cells had no CFU-C/suppressor activity. These data indicate that T cells from CGL patients cannot be primed to become CFU-C suppressor cells after PWM: stimulation in vitro and cannot release a soluble inhibitor of granulopoiesis produced by PWM-primed normal T cells.

Adolescent↗

PubMed1982

Generation of CFU-c suppressor T cells in vitro. IV. effect of time on the inhibitory activity of mitogen-primed normal T lymphocytes.

Bone marrow and peripheral blood T cells were obtained from 15 normal individuals by E rosetting and cultured in round-bottomed microwells for 7 days in RPMI or in RPMI supplemented with mitogens (pokeweed mitogen, phytohemagglutinin or concanavalin A). Supernatants and cells were harvested on days 1, 2, 3, 4 and 7 and co-cultured with normal marrow cells in semi-solid agar to test their CFU-c suppressor activity. The results of this study indicate that (a) RPMI treated cells and their supernatants have no effect or an enhancing effect on CFU-c growth; (b) all 3 mitogens generate CFU-c suppressor T cells on day 1 of culture; (c) the inhibitory activity is detectable until day 4 of culture, though overall reduced, and is completely lost on day; 7 (d) the trend for supernatants of mitogen-treated T cells is quite similar with a tendency to complete loss of the inhibitory effect on day 7. We interpret these data as indicating that T cells release a soluble inhibitor of CFU-c growth within a few hours from polyclonal activation, the production of which is either controlled or lost with time in culture.

Anemia, Aplastic↗

PubMed1982

Generation of CFUC suppressor T cells in vitro. II. Effect of PHA, PWM, and Con-A on bone marrow and peripheral blood lymphocytes from healthy donors.

T lymphocytes were derived by E-rosetting techniques from the peripheral blood and bone marrow of 12 healthy donors. Following incubation of 18 h with PWM, PHA, Con-A or culture medium (RPMI) alone, the lymphocytes were harvested by centrifugation and washed. Both lymphocytes and culture supernatants were tested for CFUC suppressor activity in semisolid bone marrow cultures. The results of this study indicate that (a) T cells and supernatants from unstimulated cultures contained no CFUC suppressor activity, (b) T cells and supernatants from cultures stimulated with PHA, PWM or Con-A significantly suppressed autologous and allogeneic bone marrow CFUC, (c) there was no significant difference between the ability of bone marrow or peripheral blood T cells to inhibit CFUC, and (d) mitogens alone had either no effect or a moderate enhancing activity on marrow CFUC. These findings suggest that generation of CFUC suppressor T cells occurs normally within a few hours of polyclonal T cell stimulation.

Bone Marrow Cells↗

PubMed1981

Tobramycin versus gentamicin, in combination with cephalotin and carbenecillin, in patients undergoing bone marrow transplantation.

Thirty-six patients with severe aplastic anemia or acute leukemia undergoing bone marrow transplantation or intensive hematologic treatments were randomized to receive gentamicin (1 mg/kg/iv q8h) or tobramycin (1 mg/kg/iv q8h) in combination with carbenecillin (500 mg/kg/day i.v. in 4 doses) and cephalotin (200 mg/kg/day i.v. in 6 doses), at the onset of fever during granulocytopenia. Patients in the gentamicin group were treated for a total of 541 patient days (minimum 5, maximum 106), and patients in the tobramycin group for 426 patient days (minimum 9, maximum 48). All patients received oral decontamination, which included the amino glycoside (either gentamicin or tobramycin) given intravenously. This study showed that 1) no patient had an increase in creatinine level above 1.3 mg % and no patient developed renal failure, 2) there was no difference between gentamicin and tobramycin in the efficacy of treating febrile episodes and/or major infections, 3) prolonged intravenous administration (up to 48 consecutive days in the tobramycin group and 106 consecutive days in the gentamicin group) was well tolerated and effective in treating fever of unknown origin in granulocytopenic patients: 45 of 72 febrile episodes resolved while the patients had a granulocyte count below 500/mm3.

Adolescent↗

PubMed1981

Imbalance of T-cell subpopulations and defective pokeweed mitogen-induced B-cell differentiation after bone marrow transplantation in man.

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Adolescent↗

PubMed1981

A monoclonal antibody to antigens expressed on MLR-activated T cells inhibits granulocyte macrophage colony formation.

A monoclonal antibody specifically reactive with MLR-activated T cells (MLR2) was added to light density normal marrow cells, depleted of adherent cells and T lymphocytes, and plated in soft agar for granulocyte macrophage colony formation. Colonies from MLR2-treated marrow cells were reduced to less than 10% of expected growth. The inhibition was not complement dependent, did not require the continuous presence of MLR2 in culture, and could not be detected also when human placenta-conditioned medium was used in the place of leukocyte feeder layers as a source of colony-stimulating factor (CSF). Co-culture experiments with MLR2 treated and untreated marrow cells further excluded the possibility of an indirect effect of MLR2 on CFU-c via auxiliary cells. The results of this study suggest that myeloid progenitor cells express a lymphoid antigen that is absent on resting or activated B cells and on resting T cells, but is expressed on activated T cells.

Absorption↗

PubMed1981