Distinct phases in recovery of reconstituted innate cellular-mediated immunity after murine syngeneic bone marrow transplantation.

Defects in immune reconstitution after hematopoietic stem cell transplantation confer extreme infection risk on to the transplant recipient. Perturbations in adaptive immune reconstitution have been well characterized, yet defects in reconstituted innate cellular-mediated immunity remain largely unstudied. Recovery in innate effector cells was defined by using an established murine model of autologous bone marrow transplantation. Cytokine induction after cell culture and systemic stimulation with pathogen-associated molecular patterns was also measured for control, transplant-recipient, and irradiated-only animals. Early reconstitution (7 to 14 days) of donor-derived macrophages, dendritic cells, and polymorphonuclear cells was associated with recovery in interleukin (IL)-12p70 and IL-6 production. Later reconstitution (21 days) of natural killer cells was associated with interferon (IFN)-gamma recovery. Hence, splenocyte innate cellular-mediated immunity recovered to normal levels in cellularity and IL-12p70, IFN-gamma, and IFN-alpha production by 21 days after transplantation. In contrast, levels of systemic cytokine production from transplant-recipient and irradiated-only animals were preserved despite incomplete or absent hematopoietic reconstitution. These results suggest that innate immune responses to systemic inflammatory challenges are largely intact after autologous bone marrow transplantation, whereas local innate cellular-mediated immunity within reconstituting lymphoid organs may be impaired. The disparate effects of autologous hematopoietic stem cell transplantation on host immune function may translate to differences in susceptibility to local versus systemic infectious challenges.     

Interleukin 12 is produced in vivo during endotoxemia and stimulates synthesis of gamma interferon.

Gamma interferon (IFN-gamma) is produced in response to circulating lipopolysaccharide (LPS) and contributes to the lethality of endotoxic shock. To address the cellular source of IFN-gamma production in vivo, T cells and B cells were magnetically purified from C57BL/6 mouse spleens 5 h following endotoxin injection. IFN-gamma RNA was abundant in splenic CD4+ and CD8+ T cells and in a T- and B-cell-depleted population of splenocytes containing 34% NK1.1+ natural killer (NK) cells. Because interleukin 12 (IL-12) is a known inducer of IFN-gamma synthesis by cultured T cells and NK cells, we examined whether IL-12 might be involved in IFN-gamma release during endotoxemia. mRNA encoding the p40 subunit of IL-12 increased markedly in the spleens of C57BL/6 mice at 2 h after LPS injection, whereas p35 IL-12 mRNA was constitutively expressed at all times. Bioactive IL-12 (p70 heterodimer) was detected in mouse serum at 2 to 4 h after LPS injection. Similar results were obtained using a p40 subunit-specific enzyme-linked immunosorbent assay. Endotoxin-insensitive C3H/HeJ mice generated threefold less IL-12 p70 and IFN-gamma at these times than endotoxin-sensitive C3H/HeOuJ mice. Pretreatment of mice with polyclonal anti-mouse IL-12 antibody reduced IFN-gamma levels present at 6 h post-LPS nearly sixfold in three separate experiments. These studies support a role for IL-12 as a proximal stimulator of IFN-gamma release during endotoxemia.     

Stoffwechseluntersuchungen bei tiefer Hypothermie unter 20° und langdauerndem artefiziellem Kreislaufstillstand

Ohne Zusammenfassung     

Mycobacterium tuberculosis-specific CD8+ T cells preferentially recognize heavily infected cells

Both CD4+ and CD8+ T cells are important for successful immunity to tuberculosis and have redundant effector functions, such as cytolysis and release of potent antimycobacterial cytokines such as interferon-gamma and tumor necrosis factor-alpha. We hypothesized that CD8+ T cells play a unique role in host defense to Mycobacterium tuberculosis infection as well. Possibilities include preferential and/or enhanced release of granular constituents and/or preferential recognition of heavily infected cells. Utilizing human, Mycobacterium tuberculosis-specific, CD4+ and CD8+ T cell clones, we demonstrate that, after recognition of antigen-presenting cells displaying peptide antigen, CD4+ T cells preferentially release interferon-gamma, whereas CD8+ T cells preferentially lyse antigen-presenting cells. Furthermore, utilizing dendritic cells infected with Mycobacterium tuberculosis expressing green fluorescent protein, we show that CD8+ T cells preferentially recognize heavily infected cells that constitute the minority of infected cells. These data support the hypothesis that the central role of CD8+ T cells in the control of infection with Mycobacterium tuberculosis may be that of surveillance; in essence, recognition of cells in which the containment of Mycobacterium tuberculosis is no longer effective.     

Cyclosporine as prophylaxis for graft-versus-host disease: a randomized study in patients undergoing marrow transplantation for acute nonlymphoblastic leukemia

Seventy-five patients, 13 to 49 years of age, with acute nonlymphoblastic leukemia in first remission were treated with cyclophosphamide, fractionated total body irradiation, and marrow transplantation from an HLA-identical sibling and randomized to receive either cyclosporine (CSP) (n = 36) or methotrexate (MTX) (n = 39) as prophylaxis for graft-v-host disease (GVHD). All patients engrafted, and 22 who were given CSP and 21 who were given MTX, are alive at 20 to 47 (median, 35) months (P = .5). Engraftment as assessed by granulocyte recovery (P less than .0005) and platelet transfusion requirement (P = .01) was faster in patients on CSP. Twelve patients (33%) on CSP and 22 (56%) on MTX developed acute GVHD of grades II through IV (P = .07) and 15 of 30 on CSP and 14 of 32 on MTX that were at risk developed chronic GVHD. The most frequent causes of death were interstitial pneumonitis and marrow relapse of leukemia, which occurred with similar frequency in both groups. Beneficial effects observed in patients on CSP included less severe mucositis and shorter duration of hospitalization; adverse effects included renal function impairment and hypertension. These data confirm that CSP is a useful immunosuppressant in patients undergoing marrow transplantation but fail to show a significant improvement in survival as compared with the standard regimen of MTX.     

Phase I trial of interleukin-2 after unmodified HLA-matched sibling bone marrow transplantation for children with acute leukemia

Allogeneic bone marrow transplantation (BMT) for advanced acute leukemia is associated with a high risk of relapse. It is postulated that interleukin-2 (IL-2) administered after BMT might induce or amplify a graft-versus-leukemia effect and thereby reduce the relapse rate. To identify an IL-2 regimen for testing this hypothesis, a phase I trial of IL-2 (Roche) was performed in children in complete remission (CR) without active graft-versus-host disease (GVHD) off immunosuppressive agents after unmodified allogeneic matched-sibling BMT for acute leukemia beyond first remission. Beginning a median of 68 days after BMT, 17 patients received escalating doses of induction IL-2 (0.9, 3.0, or 6.0 x 10(6) IU/m2/d representing levels I, II, and III) for 5 days by continuous intravenous infusion (CIV). After 6 days of rest, they received maintenance IL-2 (0.9 x 10(6) IU/m2/d) for 10 days by CIV infusion. Levels I and II were well-tolerated, but, of 6 patients at level III, 1 developed pulmonary infiltrates, 1 developed hypotension (both resolved), and 1 died of bacterial sepsis and acute respiratory distress syndrome. Grade II acute GVHD developed in 1 patient at level I and 1 at level III. The maximum tolerated dose of induction IL-2 was level II. IL-2 induced lymphocytosis, with an increase in CD56+ and CD8+ cells. Ten patients remain in CR at 5+ to 67+ months. Thus, a regimen of IL-2 has been identified that did not induce a high incidence of acute GVHD when administered to children after unmodified allogeneic BMT. Its clinical activity will be assessed in a phase II trial.     

Transplantation of allogeneic peripheral blood stem cells mobilized by recombinant human granulocyte colony-stimulating factor [see comments]

Peripheral blood stem cells (PBSCs) are widely used in autologous transplantation because of ease of collection and rapid hematopoietic reconstitution. However, PBSCs have rarely been used for allogeneic transplantation because of concerns about donor toxicities from cytokine administration and the theoretical increased risk of graft- versus-host-disease (GVHD) from the large number of T cells infused. Eight patients with advanced malignancies received allogeneic PBSC transplants from genotypically HLA-identical sibling donors. All donors received 5 days of recombinant human granulocyte colony-stimulating factor (rhG-CSF; 16 micrograms/kg/day) subcutaneously and were leukapheresed for 2 days. After treatment of the patient with total body irradiation and cyclophosphamide (n = 7) or etoposide, thiotepa, and cyclophosphamide (n = 1), PBSCs were infused immediately after collection and without modification. All patients received cyclosporine and either methotrexate (n = 6) or prednisone (n = 2) for GVHD prophylaxis, rhG-CSF was well tolerated with mild bone pain requiring acetaminophen occurring in two donors. All patients engrafted and in seven hematopoietic recovery was rapid, with 500 neutrophils/microL achieved by day 18 and 20,000 platelets/microL by day 12. Complete donor engraftment was documented by Y chromosome analysis in all four sex-mismatched donor-recipient pairs tested and by DNA analysis in two sex-matched pairs. One patient died on day 18 of veno-occlusive disease of the liver with engraftment but before chromosome analysis could be performed (results are pending in 1 patient). A second patient died of fungal infection 78 days after transplant. Grade 2 acute GVHD occurred in two patients and grade 3 GVHD occurred in one patient. One patient is 301 days from transplant in remission with chronic GVHD; the remaining five patients are alive and disease free 67 to 112 days after transplantation. Preliminary results indicate that allogeneic PBSCs mobilized by rhG-CSF can provide rapid hematologic recovery without an appreciably greater incidence of acute GVHD than would be expected with marrow. Further follow-up is required to determine the incidence of chronic GVHD and any potential beneficial effects on relapse after transplant.