Both apoptosis and complement membrane attack complex deposition are major features of murine acute graft-vs.-host disease

The parent-into-F1 mouse model (P-->F1) of acute graft-vs.-host disease (GVHD) is a useful model of human acute GVHD because it allows the study of the T cell contribution to pathology without the complicating effects of conditioning regimens. To determine the similarity of this model to human GVHD, we assessed injury in organs typically involved in human acute GVHD (skin, liver) and less typically involved organs (spleen, kidney, lung). Mice were assessed histologically at early (2 weeks), intermediate (3 months) and late (6 month) time points. Based on the emerging roles of Fas ligand killing and complement deposition in allograft rejection, we correlated the amount of tissue specific TUNEL positive apoptosis and deposition of complement (C5b-9) with histopathologic changes. Our results indicate a striking similarity histologically between acute GVHD occurring in this model and in humans following bone marrow transplant. Moreover, C5b-9 deposition and apoptotic cell accumulation were found to parallel tissue injury in major organs of acute GVHD mice, although not all organs exhibited the same kinetic pattern. These results indicate a role for both adaptive immunity and innate immunity in this model of GVHD and support its use in modeling human acute GVHD in the nonmyeloablative setting.     

Genomic variation of adenovirus type 5 isolates recovered from bone marrow transplant recipients.

We characterized the genomic variation of adenovirus type 5 isolates recovered from bone marrow transplant recipients in Seattle between 1976 and 1982. By restriction endonuclease analysis, we identified three new adenovirus genomic variants, each associated with a single invasive adenovirus infection. In addition, we were able to obtain suggestive evidence for a nosocomial spread of a particular group of isolates within this population. This study demonstrates that the technique of restriction endonuclease analysis is an important epidemiological tool for investigating viral infections.     

Production of interleukin (IL)-10 and IL-12 by murine colonic dendritic cells in response to microbial stimuli

Intestinal dendritic cells (DC) are likely to regulate immunity to gut microflora, but little is known about their responses to bacterial antigens. Therefore, DC from normal murine colon were characterized and their cytokine responses to components of Gram-negative and/or Gram-positive bacteria assessed. Cells were obtained by digestion of colonic tissue and contained DC that were identified by flow cytometry as CD11c(+) major histocompatibility complex (MHC) class II(+) cells. Purified DC were obtained by immunomagnetic separation plus cell sorting. DC had the morphology of immature myeloid cells, were endocytically active, expressed low levels of co-stimulatory molecules and stimulated a weak allogeneic mixed leucocyte reaction. Analysis of flow cytometry data by a sensitive subtraction method allowed measurement of production of interleukin (IL)-12 and IL-10 by small numbers of gut DC by intracellular staining. Fewer than 5% of unstimulated DC produced either IL-10 or IL-12. IL-10 production was significantly up-regulated following stimulation with Bifidobacteria longum, but not after exposure to lipopolysaccharide (LPS) or Streptococcus faecium. In contrast, colonic DC produced IL-12 in response to both LPS and B.longum. Thus, colonic DC can produce both IL-12 and IL-10 following bacterial stimulation. Cell wall components from different bacteria stimulate distinct responses and may direct immune responses differentially in the gut.     

In vitro and in vivo inhibition of interleukin (IL)-5-mediated eosinopoiesis by murine IL-5Ralpha antisense oligonucleotide

The unique role of interleukin (IL)-5 in eosinophil production, activation, and localization makes this cytokine a prime target for therapeutic intervention in diseases characterized by a selective blood and tissue eosinophilia. In an attempt to block the effects of IL-5 on eosinophils, a strategy was developed to suppress the expression of the IL-5 receptor alpha chain (IL-5Ralpha) by antisense oligonucleotides (ASOs). IL-5Ralpha ASOs were identified which selectively and specifically suppress the expression of messenger RNA and proteins of both the membrane and the soluble form of the receptor in constitutively IL-5R-expressing murine BCL-1 cells in vitro. Moreover, these IL-5Ralpha-specific ASOs were able to selectively inhibit the IL-5-induced eosinopoesis from murine fetal liver and bone marrow cells in vitro, suggesting that these molecules may affect the development of IL-5-mediated eosinophilia in vivo. Indeed, intravenous administration of IL-5Ralpha-specific ASOs not only suppressed the bone-marrow and blood eosinophilia in mice after short-term treatment with recombinant murine IL-5 but also inhibited the development of blood and tissue eosinophilia in a ragweed-induced allergic peritonitis model. Thus, blocking the expression of IL-5Ralpha on eosinophil using ASOs may have therapeutic benefits in eosinophilic diseases such as asthma.     

Immunohistology of skin and rectum biopsies in bone marrow transplant recipients.

We have studied histological and immunohistological specimens of 39 skin biopsies from 21, and 30 rectal biopsies from 17 bone marrow transplant recipients. The biopsies were taken before transplantation, during acute and chronic graft-versus-host disease (GVHD), and at times with no GVHD. In biopsies taken during cutaneous aGVHD grade I to III, epithelial changes were seen in 16/23 biopsies. The cutaneous infiltrates during aGVHD consisted of CD2-, CD4-, CD8- and FMC-33-positive cells both in the epithelium and in the dermis. CD57-positive NK cells were also detected in most biopsies. During chronic GVHD the cutaneous cellular infiltrates were similar to those seen in moderate aGVHD, i.e. both CD4- and CD8-positive lymphoid cells were present. When the biopsy was taken after the beginning of corticosteroid treatment for aGVHD, or at times when the patient did not have GVHD symptoms, the cellular infiltrates were considerably smaller in the dermis. During clinical intestinal aGVHD mucosal epithelial changes were relatively uncommon; instead, increased numbers of both CD4- and CD8-positive lymphocytes in the lamina propria (LP) were seen in 11/13 samples. During chronic GVHD the number of CD4-positive cells exceeded that of CD8-positive cells in the LP, and the large lymphoid infiltrates also reached the muscularis mucosae. In rectal biopsies the differences were not so prominent because most of the pretransplant biopsies showed CD2-, CD4-, CD8- and CD57-positive lymphocytes both in the lamina propria and epithelium.     

The secreted form of the p40 subunit of interleukin (IL)-12 inhibits IL-23 functions and abrogates IL-23-mediated antitumour effects

Interleukin (IL)-23 is a heterodimeric cytokine consisting of a novel p19 molecule and the p40 subunit of IL-12. Since secreted p40 can act as an antagonist for IL-12, we investigated whether p40 also inhibited IL-23-mediated immunological functions. p40 did not induce interferon (IFN)-gamma or IL-17 production from splenocytes but impaired IL-23-induced cytokine production by competitive binding to the IL-23 receptors. Furthermore, a mixed population of murine colon carcinoma Colon 26 cells transduced with the p40 gene and those transduced with the IL-23 gene developed tumours in syngenic mice, whereas the IL-23-expressing Colon 26 cells were completely rejected. p40 also suppressed IFN-gamma production of antigen-stimulated splenocytes and IL-23-mediated cytotoxic T-lymphocyte activities in the mice that rejected Colon 26 cells expressing IL-23. p40 can thereby antagonize IL-23 and is a possible therapeutic agent for suppression of IL-23 functions.     

The effects of interleukin (IL)-12 and IL-4 deficiency on worm development and granuloma formation in Schistosoma japonicum-infected mice

CD4⁺ T-helper (Th) cell is widely recognized to be capable of influencing worm development and egg granuloma formation after schistosome infection. Interleukin (IL)-12 and IL-4 play key roles in regulation of Th cell differentiation. In the present study, we subcutaneously inoculated mice with hybridoma cells secreting monoclonal antibodies to neutralize IL-12 and IL-4 and explored the effects of IL-12 and IL-4 deficiency on the worm development and granuloma formation in mice infected with cercariae of Schistosoma japonicum. It was found that deficiency of host IL-12 and IL-4 supported normal parasite survival and fecundity. However, worm development (length and female fecundity) was significantly enhanced in anti-IL-12-treated mice. Mean length of worms in anti-IL-12-treated group was significantly greater than that of intact controls on day 28 after infection (females, 11.84 ± 1.20 mm vs. 9.45 ± 1.34; males, 9.35 ± 1.21 mm vs. 8.10 ± 0.85 mm, p < 0.05). Liver egg load per pair of worms (1,770.12 ± 470.67 vs. 806.08 ± 232.37, p < 0.05) and uterine egg load of ovigerous females (93.08 ± 27.85 vs. 46.05 ± 34.24, p < 0.05) in anti-IL-12-treated mice were significantly higher than those in intact control 28 days postinfection. But these effects diminished 42 days postinfection (p > 0.05). Granuloma size in anti-IL-12-treated mice was significantly larger than that in intact mice 42 days postinfection (398.3 ± 80.7 μm vs. 294.4 ± 72.2 μm, p < 0.05). Granuloma fibrosis dramatically intensified in anti-IL-12-treated mice but diminished in anti-IL-4-treated mice. The results suggest that IL-12 may play an impeditive role in the development of S. japonicum and in granuloma formation as well as fibrosis. IL-4 may promote granuloma formation but have no effect on worm development.     

The combined effect of interleukin (IL)-10 and IL-12 polymorphisms on induced cytokine production

Interleukin (IL)-12 and IL-10 are immunoregulatory cytokines with an antagonistic effect of the T-helper (Th)1/Th2 cytokine balance and provide a functional link between innate and adaptive immune responses. The aim of the study was to investigate the combined effect of -1082A*G in IL10 and +16974A*C in IL12B single nucleotide polymorphisms (SNPs) on induced cytokine production by stimulated peripheral blood mononuclear cells (PBMCs) isolated from healthy donors. The presence of the high-producer IL-12p40 genotype led to diminished production of IL-10 as determined by the -1082*G allele of SNP in IL10. Significantly decreased IL-10 production was detected in AA+AG/GG in comparison with the low-producer IL-12p40 (AC/CC+AG/GG) genotype combination after stimulation with C3bgp (2 +/- 4 vs. 29 +/- 14.2 pg/ml; p = 0.0003) and LPS (33.4 +/- 13.5 vs. 93.3 +/- 59.6 pg/ml; p = 0.019). IL-12p40 production was independent of IL10 genotype. The present results demonstrated that the production of IL-10 from PBMC depended on both -1082A*G in IL10 and +16974A*C in IL12B polymorphisms.     

Inhibition of in vitroimmunoglobulin production by IL-12 in murine chronic graft- vs. -host disease: synergism with IL-18

We investigated the effects of IL-12 on immunoglobulin (Ig) production in vitro in murine chronic graft- vs. -host disease (cGVHD), a lupus-like model of overt B cell activation induced by allogeneic stimulation. Addition of IL-12 to cGVHD splenocytes strongly inhibited total Ig (Igκ), IgM and IgG1 production. Although IL-12 down-regulated IL-4, IL-5, IL-9 and IL-10 production, its inhibitory activity on Ig production could not be ascribed to down-regulation of these cytokines, as addition of saturating doses of IL-4, IL-5 and/or IL-9 did not reverse the inhibitory activity of IL-12. Interestingly, IL-12 was also found to suppress the stimulating effect of IL-4 and IL-5 on Ig synthesis by cGVHD splenocytes. Several lines of evidence indicated that the inhibitory activity exerted by IL-12 on Ig production was mediated by IFN-γ. First, IFN-γ was produced in large amounts upon IL-12 stimulation. Secondly, it displayed a potent inhibitory activity on Ig production. Thirdly, Ig production was also inhibited by IL-18, a recently cloned IFN-γ-inducing cytokine. Finally, the inhibitory activity of IL-12 was blocked by anti-IFN-γ monoclonal antibody. We also investigated whether IL-12 down-regulated Ig production by purified cGVHD B cells. We found that IL-12 had only a marginal inhibitory activity on highly purified B cell populations isolated from cGVHD splenocytes and stimulated with IL-4 and IL-5, and that IL-18 was inactive in this respect. However, when the two cytokines were combined, a striking synergy was unmasked not only for IgG1 inhibition but also for IFN-γ production by these B cell populations. Taken together, our results demonstrate that IL-12 inhibits in vitro Ig production by activated splenocytes through IFN-γ production and that it synergizes with IL-18 on activated B cells to inhibit Ig production, through up-regulation of IFN-γ production by B cells.     

A retrospective comparison of allogeneic peripheral blood stem cell and bone marrow transplantation results from a single center: a focus on the incidence of graft-vs.-host disease and relapse.

To detect the effect of the stem cell source, allogeneic peripheral blood stem cell transplantations (alloPBSCTs) performed between 1995 and 1997 from human leukocyte antigen (HLA)-identical siblings in 40 patients with acute and chronic hematological disorders were compared with a historical group of 40 patients with similar variables who had received allogeneic bone marrow transplants (alloBMTs) between 1993 and 1995. Patients in both groups were identical except that both the recipient and the donor ages were, on average, higher in the alloPBSCT group (26 vs. 36 [p = 0.005] and 27 vs. 32 [p = 0.024], respectively). Patients received similar therapy excluding posttransplant granulocyte colony-stimulating factor administration (97% in alloBMT vs. 12.5% in alloPBSCT). The median time to reach neutrophil counts >0.5 x 10(9)/L and platelet counts >20 x 10(9)/L was 13 and 14 days, respectively, in patients receiving alloPBSCTs compared with 19 and 27 days in patients receiving alloBMTs (p = 0.0014 and p = 0.0002). The alloPBSCT group required similar transfusions of red blood cells or platelets. The incidence of grade II-IV acute graft-vs.-host disease (aGVHD) was similar in both groups. However, chronic GVHD (cGVHD) of all grades developed in 78.1% of patients in the alloPBSCT group after a median follow-up period of 12.5 (range 0.5-34) months. In alloBMT recipients, cGVHD of all grades developed in 21.4% after a median follow-up period of 38 (range 0.5-62) months (p = 0.00001). Day 100 transplant-related mortality was also similar: 20% (8 of 40) in the alloBMT patients and 17.5% (7 of 40) in the alloPBSCT group. Although not statistically significant, a relatively higher relapse rate occurred in the alloBMT group (21.4 vs. 10.7%). The estimated disease-free survival in month 24 was 51.3% for alloBMT and 54.6% for alloPBSCT, and the estimated overall survival in month 24 was 56.1% for alloBMT and 64.6% for alloPBSCT. In conclusion, this retrospective comparison suggests that alloPBSCT from HLA-identical donors is associated with faster engraftment, fewer transfusions, and no greater incidence of aGVHD, but a high incidence of cGVHD.     

Impaired T and NK cell response of bone marrow and peripheral blood stem cell products to interleukin (IL)-2.

The function of steady-state and interleukin (IL)-2-co-cultured mononuclear cells differs significantly between bone marrow (BM) products, growth factor-mobilized peripheral blood stem cell (PSC) products and normal peripheral blood mononuclear cells (PBMC). The natural killer (NK) cell activity and T cell proliferative response of PSC products from non-Hodgkin's lymphoma (NHL) patients are significantly higher than that of BM products and similar to normal PBMC. However, following a five-day co-culture with IL-2 (100 IU/ml), the NK activity of PSC, PBMC, and BM products (lytic units) was increased 176-, 40-, and 14-fold, respectively, compared to that observed prior to IL-2 culture. In contrast, lymphokine activated killer (LAK) cytotoxicity prior to IL-2 culture was low in PSC and BM products and normal PBMC, but was significantly increased in PSC products and PBMC following IL-2 co-culture. The proliferative response of PSC and BM products to the T cell mitogen phytohemagglutinin (PHA) was significantly lower than that observed with normal PBMC; however, PSC had a significantly higher response than cells from BM products. Similar patterns of T cell PHA mitogenic response were observed after IL-2 co-culture. In addition, the IL-2 mitogenic responses of IL-2-co-cultured PSC and BM products were also significantly lower than that observed with PBMC co-cultured with IL-2. The IL-2 mitogenic response of PBMC was also significantly increased compared to prior to IL-2 co-culture; whereas, the IL-2 mitogenic responses from PSC and BM cells were not. In summary, co-culture with IL-2 can increase the NK and LAK cell cytotoxicity of PSC and BM products from NHL patients, but IL-2 co-culture does not improve T cell function within either BM or PSC products.     

Graft-versus-host disease in murine bone marrow transplantation. I. Modification of GVHD by preimmunization of recipients with spleen cells of primary recipients undergoing GVHD

Lethally irradiated (1000 rad) CBA/J mice were transplanted with anti-Thy 1 treated BALB/c bone marrow. Under these conditions, we uniformly observed the development of pathology suggestive of acute graft-vs.-host disease (GVHD), i.e. weight loss, diarrhoea, hypogammaglobulinemia and thymic hypoplasia. If, 2 wk before irradiation, the recipients were preimmunized with spleen cells taken from mice undergoing acute GVHD, these symptoms were avoided. Instead, such animals seemed to show long term survival either with or without signs of chronic GVHD (hypergammaglobulinemia, splenomegaly, lymphoid hyperplasia). The ability to show long term survival with bone marrow allografts was dependent upon successful immunization of the recipient mice. Long term survivors contain a splenic population not bearing detectable host MHC antigens, which can elicit a memory anti-host cytotoxic response from a population of quiescent donor lymphocytes previously immunized in vitro against host MHC antigens.     

Marrow ablative doses of gamma-irradiation and protracted changes in peripheral lymph node microvasculature of murine and human bone marrow transplant recipients

Gamma-irradiation has been extensively utilized as a bone marrow ablative agent during human bone marrow transplantation. Although the effects of ionizing radiation on lymphoid and hematopoietic cells are well documented, little is currently known about its effect on the nonhematopoietic tissues which are important for the restoration of normal immune function. The vast majority of lymphocyte movement into peripheral lymph nodes takes place via the bloodstream and requires a specific receptor-ligand interaction between the lymphocyte and anatomically distinct postcapillary venules. Due to the importance of lymphocyte recirculation in the initiation and amplification of immune responses, an understanding of the radiosensitivity of the postcapillary venules may provide insight into the pathogenesis of the immune deficiencies commonly seen after bone marrow transplantation. Our studies disclosed that the ability of normal blood-borne lymphocytes to enter peripheral lymph nodes was markedly depressed (less than 50% of normal) in mice which had been exposed to 7.5 Gy of gamma-irradiation. This radiation-induced effect lasted longer than 6 months after irradiation and syngeneic reconstitution, and its magnitude was radiation-dose dependent. Immunochemical staining of the lymph node microvasculature with the monoclonal antibody MECA-325 established that the radiation protocol induced persistent anatomic changes in the lymphocyte-receptive areas of endothelium (high endothelial venules). These vessels developed the appearance of endothelial cell proliferation. Electron microscopy demonstrated significant intracellular edema, with virtual occlusion of many microvascular lumens by edematous endothelial cells. Lymph nodes from human bone marrow transplant recipients were found to exhibit similar ultrastructural changes. These studies for the first time demonstrate that doses of irradiation similar to those used to prepare bone marrow transplant recipients can have significant anatomic and functional sequellae on host endothelial cells.     

Cytomegalovirus diagnosis in renal and bone marrow transplant recipients: the impact of molecular assays.

BACKGROUND: Cytomegalovirus (CMV) infections are a major threat in transplant recipients. In recent years, new assays for routine CMV diagnosis, based on molecular techniques, have become available. OBJECTIVE: The impact of molecular assays for CMV diagnosis in transplant recipient was evaluated. STUDY DESIGN: A total of 51 transplant recipients were screened for CMV infection. Serological (AxSYM CMV IgG and recombinant CMV IgM assays), antigenemia, CMV DNA (qualitative in house PCR and the quantitative COBAS AMPLICOR CMV MONITOR Test), and CMV mRNA (NucliSens CMV pp67 Test) tests were compared. RESULTS: In 11/20 bone marrow transplant (BMT) recipients and 10/31 renal transplant (RTX) recipients there was no evidence of active CMV infection. Ten RTX recipients and one BMT recipient were antigenemia positive, 21 RTX and seven BMT recipients were PCR positive (qualitative CMV PCR). There were more BMT recipients CMV DNA positive in serum (7/21) than antigenemia positive (1/21). CMV mRNA was found positive in two BMT recipients (one case with no other evidence of CMV infection, the other one CMV DNA positive and antigenemia negative). The only antigenemia positive BMT recipient was found negative for CMV mRNA, but positive in all other tests. Eight RTX recipients were found positive for CMV mRNA. Six of them were also antigenemia positive and five of those were also found positive for CMV IgM. One CMV mRNA positive RTX recipient was CMV IgM positive but antigenemia negative and the other one CMV mRNA positive RTX recipient was found negative in all other tests. Two antigenemia positive RTX recipients were found negative for mRNA and CMV IgM. CONCLUSION: Antigenemia was found to be a good screening test for CMV infection in RTX recipients. In BMT recipients, tests based on molecular techniques appeared to be superior compared to antigenemia.     

Distinct genotypic distributions of cytomegalovirus (CMV) envelope glycoprotein in bone marrow and renal transplant recipients with CMV disease.

A prospective study of the spectrum of glycoprotein B (gB) and glycoprotein H (gH) genotypes of cytomegalovirus (CMV) was conducted with five categories of patients: viremic bone marrow-transplant (BMT) recipients who developed CMV disease after BMT (n = 22), viremic BMT recipients without CMV disease (n = 11), viremic renal-transplant recipients who developed CMV disease after transplantation (n = 14), viremic renal-transplant recipients without CMV disease (n = 13), and premature babies with asymptomatic congenital CMV infections (n = 13). Genotypic stability was observed because the gB and gH genotypes of multiple isolates obtained from a single patient were identical. The distribution of gH genotypes in patients of all groups studied were similar. However, there was a unique distribution of the gB genotype in the first category of patients, i.e., BMT recipients with CMV disease, which was distinct from those of all other categories (P < 0.05). CMV isolates from 54% of BMT recipients with CMV disease exhibited gB type 2, while isolates from 46, 50, 69, and 77% of the BMT recipients without CMV disease, renal-transplant recipients with and those without CMV disease, and premature babies with congenital CMV infection, respectively, were of gB type 1. An analysis of the clinical characteristics of BMT recipients with CMV disease indicated that all underwent either an allogeneic or matched, unrelated donor transplant, and half had severe acute graft-versus-host disease (grades 2 to 4). The statistically significant genotypic difference between CMV isolates from BMT recipients with and without CMV disease was not observed between isolates from renal-transplant recipients with and without CMV disease. We speculate that differences in pathogenesis in different patient groups might account for these observations. These findings would also facilitate decision making about the choice of recombinant CMV glycoprotein vaccine required to immunize transplant donors and the subsequent adoptive transfer of immunity to BMT recipients. When the source of CMV DNA required for genotyping was investigated among renal-transplant recipients, direct use of peripheral blood leukocytes was 95% effective compared to the effectiveness of cells obtained from conventional culture of peripheral blood specimens.     

Sphingosine-1-phosphate differently regulates the cytokine production of IL-12, IL-23 and IL-27 in activated murine bone marrow derived dendritic cells

Sphingosine-1-phosphate (S1P) modulates many cell functions such as lymphocyte trafficking and signaling as well as keratinocyte proliferation. However, less is known about the specific effects of S1P on cytokine production, particularly on the interaction between dendritic cells (DCs) and keratinocytes, cell types which are crucial for the initiation and maintenance of chronic inflammatory skin diseases like atopic dermatitis or psoriasis. Especially the cytokines of the IL-12 family play a dominant role in many inflammatory diseases as they have a significant impact on T-helper cell function. In the present study we show that S1P decreased the production of the pro-inflammatory cytokines IL-12 and IL-23 in LPS-stimulated DCs via the common subunit p40 as well as in the crosstalk with activated keratinocytes. By using specific S1P receptor agonists (SEW2871, FTY720-P) and antagonist (JTE013) we identified an important role for S1P receptor 1 in the modulation of the cytokine profile. While diminishing IL-12 and IL-23 secretion, S1P enhanced IL-27 production in DCs. To elucidate the mechanism of the different impact on the IL-12 family cytokine production, we investigated the mitogen-activated protein kinase (MAPK) and phosphatidylinositide 3-kinase (PI3K) pathways in DCs. By using specific MAPK-Inhibitors (U0126, SB202190, SP600125) we demonstrated that ERK, p38 and JNK differently regulate each pathway of each cytokine. While p38 and JNK did not seem to play a role in the modulation properties of S1P on cytokine production, ERK is at least partially involved in the S1P mediated modulation of IL-12 and IL-27. The PI3K-Inhibitor abrogated the S1P-induced decrease of IL-12 and IL-23 secretion, while it had no influence on the S1P-induced increase of IL-27 production. These data implicate, that S1P has an anti-inflammatory impact on the production of IL-12 family cytokines, indicating therapeutic potential for S1P treatment of several inflammatory diseases like psoriasis.     

Selective and differential binding of interleukin (IL)-1 alpha, IL-1 beta, IL-2 and IL-6 to glycosaminoglycans.

The binding of interleukin (IL)-1 alpha, IL-1 beta, IL-2 and IL-6 to acidic polysaccharides was investigated by affinity chromatography of the recombinant, radioiodinated interleukins on columns of immobilized polysaccharide. Each interleukin showed selective binding retention. Overall heparin bound all four interleukins significantly, whereas chondroitin sulfate provided little retention. IL-1 alpha and IL-1 beta showed differential binding, with only the latter binding to hyaluronic acid. IL-2 was virtually completely retained on fucoidan. Noniodinated recombinant IL-2 bound similarly to fucoidan, and fucoidan was found to sequester IL-2 activity in a bioassay employing IL-2-dependent CTLL cells. In all other cases tested, interleukin retention was partial, implying that interleukin binding sites are sparsely distributed along the polysaccharide chains. These findings suggest that during the immune response, interleukins will tend to be retained at sites of secretion by interaction with glycosaminoglycans in the extracellular matrix and on cell surfaces.     

Management of allogeneic bone marrow transplant recipients at risk for cytomegalovirus disease using a surveillance bronchoscopy and prolonged pre-emptive ganciclovir therapy.

BACKGROUND: Patients undergoing allogeneic bone marrow transplant (BMT) are considered to be at increased risk of cytomegalovirus (CMV) disease if they and/or their donor are CMV seropositive pre-transplant. Although several pre-emptive strategies have been shown to be effective in preventing early CMV disease, the ability of pre-emptive strategies using prolonged ganciclovir therapy to reduce the incidence of late-onset CMV infection, disease and mortality has not been fully evaluated. OBJECTIVE: To assess the efficacy of 18 weeks of pre-emptive ganciclovir therapy in preventing late-onset (> 100 days post-transplant) CMV disease when administered to asymptomatic BMT patients found to have CMV in bronchoalveolar lavage (BAL) fluid obtained during a surveillance bronchoscopy approximately 35 days post-transplant. To determine whether or not survival of BMT recipients is influenced by pre-transplant donor and recipient CMV serostatus in the context of this pre-emptive ganciclovir strategy. STUDY DESIGN: Consecutive patients undergoing allogeneic BMT were assessed for their risk of developing CMV disease based on their pre-transplant CMV serostatus and that of their donor. Patients who were CMV seropositive and/or received marrow from a CMV seropositive donor underwent a surveillance bronchoscopy and BAL approximately 35 days post-transplant. Patients with positive BAL fluid for CMV received pre-emptive ganciclovir therapy for 18 weeks at decreasing dose levels. Patients considered to be at low risk for the development of CMV disease (donor and recipient CMV seronegative) were followed without intervention. RESULTS: Of 98 consecutive patients, 55 were considered to be at risk for CMV disease and underwent a surveillance bronchoscopy. Sixteen (29%) patients had a positive BAL fluid for CMV and were started on pre-emptive ganciclovir therapy. Two patients progressed and died with CMV-related pneumonia. One additional patient developed CMV-related enteritis on day 42 post-transplant and recovered with continuing ganciclovir treatment. Of the 39 patients with a negative BAL fluid for CMV, one developed a fatal CMV pneumonia 150 days post-transplant and two additional patients developed gastrointestinal CMV disease 28 and 57 days post-BMT, respectively. None of the patients in the low risk group developed CMV disease. CONCLUSIONS: The strategy utilizing a surveillance bronchoscopy for CMV and initiating prolonged (18 weeks) pre-emptive ganciclovir therapy for patients with a positive BAL fluid for CMV resulted in a low incidence of CMV-related post-transplant complications. After a minimum follow-up of 16 months, late CMV reactivations (occurring > 100 days post-transplant) were not observed in the group of individuals pre-emptively treated with ganciclovir. This observation suggests that prolonged therapy with a reduced dose of ganciclovir may be important in the prevention of CMV reactivation. The CMV serostatus of donors and recipients prior to BMT did not correlate with survival.     

Dose response and timing effects in the therapy of the LP-BM5 murine retrovirus-induced lymphoproliferative immunodeficiency disease with diethyldithiocarbamate

Diethyldithiocarbamate (DTC) was used to treat the murine, retrovirus-induced, immunodeficiency disease (MAIDS). Once-weekly treatment was not effective and 800 mg/kg was toxic. When 200, 400 and 600 mg/kg were given i.p., 5 days per week, starting either on the day of virus inoculation or 14 days later, a dose- response and time- response relationship was noted. Higher doses and a 2-week delayed onset of treatment were generally more effective in reducing the development of lymphadenopathy, hypergammaglobulinemia and in prolonging survival than treatment started on the day of virus inoculation. When treatment was delayed until 10 weeks after virus inoculation existing lymphadenopathy was abrogated (treated node area 0 mm² compared to control 175 mm², P<0.0001) and survival was improved (treated 100% compared to control 12.5%, P<0.0001). However, when therapy was stopped animals died at the same rate as the untreated controls. These data indicate that DTC is active in MAIDS in a dose-responsive and time-dependent manner.