anti-Gal IgG Levels Research Articles

Antibodies against Pneumococcal Capsular Polysaccharides and Natural Anti-Galactosyl (Alpha-Gal) in Patients with Humoral Immunodeficiencies.

Humoral deficiencies represent a broad group of disorders. The aim of the study was to compare the levels of antibodies against pneumococcal capsular polysaccharides (anti-PCP) and natural anti-galactosyl (anti-Gal) antibodies in (1) patients with chronic lymphocytic leukaemia (CLL), (2) patients with common variable immunodeficiency (CVID), and (3) a healthy population and to explore their diagnostic and prognostic potential. Serum immunoglobulin levels and levels of anti-Gal IgG, IgA, and IgM and anti-PCP IgG and IgG2 were determined in 59 CLL patients, 30 CVID patients, and 67 healthy controls. Levels of IgG, IgA, IgM, anti-Gal IgA, anti-Gal IgM, and anti-PCP IgA were lower in CLL and CVID patients than in healthy controls (p value for all parameters < 0.0001). Decrease in the levels of IgA, IgM, anti-Gal IgA, and anti-PCP IgA was less pronounced in the CLL group than in the CVID group. IgA decline, anti-Gal IgA, anti-PCP IgA, and anti-PCP IgG2 were negatively correlated with CLL stage. We devise the evaluation of anti-Gal antibodies to be a routine test in humoral immunodeficiency diagnostics, even in cases of immunoglobulin substitution therapy. Significant reductions, mainly in anti-Gal IgA, IgM, and anti-PCP IgA levels, may have prognostic importance in CLL patients.

Levels of anti-Gal IgG2 negatively correlate with the survival of porcine islet graft in non-human primates under immunosuppression of anti-CD154

Abstract Background Anti-CD154, the most successful immunosupppression (IS) regimen in porcine islet transplantation (PITx) is prohibited for clinical use. Anti-CD40, an alternative to anti-CD154, is not as effective as anti-CD154. To understand the reason, the relationship between anti-Gal responses and the graft survival was investigated in rhesus monkeys of PITx. Methods Intraportal PITx was performed in 18 diabetic monkeys: 3 received various IS without CD40 pathway blockade (Tx-control); 12, IS with anti-CD154 (Tx-anti-CD154); and 3, IS with anti-CD40 (Tx-anti-CD40). Blood samples were weekly obtained from the monkeys after PITx until the graft ceased to function, and additionally from 3 monkeys without transplantation (non-Tx-control) for 6 months. Levels of anti-Gal IgG, IgG1, IgG2 and IgM in the samples were analyzed by in-house ELISA, and levels of D-dimer and high-sensitive CRP by turbidimetry. Results Intraportal PITx led to strong induction of anti-Gal IgG1 response, not IgG2, in Tx-control, which was suppressed in both Tx-anti-CD154 and Tx-anti-CD40. The level of anti-Gal IgG2 revealed negative association with the levels of CRP and D-dimer in Tx-anti-CD40 and non-Tx-control, not in Tx-anti-CD154. The time-weighted average levels of anti-Gal IgG2 in Tx-anti-CD154 were significantly lower than in non-Tx-control. Additionally, the levels of anti-Gal IgG2 and D-dimer negatively correlated with the graft survival in Tx-anti-CD154. Conclusions The strong correlation between the suppression of anti-Gal IgG2 response and the graft survival in Tx-anti-CD154 suggests that IgG2 response is implicated with the effector mechanism mediating rejection, possibly NK cell activation which is known to promote IgG2 response.

Increase in anti-Gal IgM level is associated with early graft failure in intraportal porcine islet xenotransplantation.

BackgroundAnti-Gal is a major antibody induced in non-human primates (NHPs) after xenotransplantation. To understand the mechanism of graft rejection, we investigated the association between anti-Gal responses and graft failure in NHP recipients of porcine islet transplantation (PITx).MethodsIntraportal PITx was performed in 35 diabetic NHPs, and graft function was monitored. Early graft failure (EGF) was defined as loss of graft function within a month after PITx. Seven, 19, nine NHPs received immunosuppression (IS) without CD40 pathway blockade (Group I), with anti-CD154 (Group II), and with anti-CD40 (Group III), respectively. The anti-Gal levels on day 0 and day 7 of PITx were measured by ELISA.ResultsThe frequency of EGF was significantly lower in Group II (26.3%) than in Group I (100%, P=0.0012) and Group III (77.8%, P=0.0166). While levels of anti-Gal IgG in Group I and anti-Gal IgM in Group III increased on day 7 compared with day 0 (P=0.0156 and 0.0273), there was no increase in either on day 7 in Group II. The ratio of anti-Gal IgM or IgG level on day 7 to that on day 0 (Ratio7/0) was significantly higher in recipients with EGF than without EGF (P=0.0009 and 0.0027). ROC curve analysis of anti-Gal IgM Ratio7/0 revealed an area under the curve of 0.789 (P=0.0003).ConclusionsIS with anti-CD154 suppressed anti-Gal responses and prevented EGF in PITx. Anti-Gal IgM Ratio7/0, being associated with EGF, is a predictive marker for EGF.

An increase in the level of anti-Gal IgM is associated with an early graft failure in intraportal porcine islet xenotransplantation (TRAN1P.935)

Abstract In order to develop laboratory tests to predict graft outcome in porcine islet transplantation (ITx), we investigated the association between anti-Gal antibody responses and the graft failure in NHP recipients after porcine ITx. Total 35 naïve diabetic NHP recipients were intraportally administered with adult porcine islets and then graft function was monitored. Early graft failure (EGF) was defined as returning to diabetic state within a month after ITx. Among the recipients, 7 and 28 received induction immunosuppression (IS) without and with CD40 pathway blockade, respectively: 19, anti-CD154 and 9, anti-CD40. The levels of anti-Gal IgG and IgM in the recipients on the day 0 (0 d) and on the day 7 (7 d) of ITx were quantitatively measured by in-house ELISA. Whereas an increase in the level of anti-Gal IgG after ITx was suppressed by the use of either anti-CD154 or anti-CD40, a response of anti-Gal IgM was suppressed by anti-CD154, not by anti-CD40. The anti-Gal antibody ratio of 7 d/0 d was significantly higher in the recipients with EGF than in those without EGF (IgG ratio, 1.54 vs 0.75, P=0.0025; and IgM ratio, 1.48 vs 0.87 P=0.0035). An increase in the level of anti-Gal IgM on the day 7 of ITx is significantly associated with EGF in the recipients with porcine ITx regardless of use of CD40 pathway blockade, suggesting that an early response of anti-Gal IgM may reflect the innate immune responses which account for killing of porcine islets in early period of porcine ITx.

288 Non-HLA Antibodies and Risk of Rejection after Heart Transplantation: Profiling with Antigen Microarrays

antibody levels decreased in both WT and GTKO recipients after implantation. WT recipients, however, retained elevated levels of anti-Gal IgG for at least the first year post implant. Conversely anti-Gal IgG levels in all GTKO recipients fell within one month. The area under the curve showed a significant increase of anti-Gal IgG in the WT BHV group compared to GTKO BHV recipients (p 0.01). Conclusions: Persistently and significantly (p 0.01) elevated levels of anti-Gal IgG were observed in WT but not GTKO BHV recipients. This indicates a BHV-specific continuing immune stimulation to the Gal antigen which is known to be present in current commercially available BHVs, and other bioprosthetic materials of animal origin. These data support the hypothesis that clinical use of current xenogeneic bioprosthetic materials can induce an anti-Gal antibody response which increasing evidence suggests promotes subsequent calcification. BHVs made from GTKO pigs would have reduced antigenicity, may have greater durability and be potentially used in younger patients with more active immune systems.

Importance of the Gal alpha1-3 Gal antigen in discordant islet xenotransplantation: immunosuppression, which inhibits porcine islet xenograft rejection in ordinary mice, is equally effective in Gal-knockout mice.

Islet xenotransplantation will most likely be performed in diabetic patients treated with immunosuppressive drugs. The importance of the galactosyl alpha(1-3) galactose (Galalpha1-3Gal) antigen in immunosuppressed islet xenograft recipients has not been studied. Fetal porcine islet-like cell clusters (ICCs) were transplanted into the renal subcapsular space of both Gal-knockout mice and ordinary mice. Transplantations were performed in untreated mice and mice immunosuppressed with cyclosporine A (CsA) plus 15-deoxyspergualin (DSG). Studies were also performed in immunosuppressed Gal-knockout mice that had been actively immunized against Galalpha1-3Gal. Evaluation was performed 12 days after transplantation using morphologic techniques. The levels of serum immunoglobulin (Ig)G and IgM to the Galalpha1-3Gal antigen or to the ICCs were determined. No difference in the morphologic appearance could be seen between ordinary mice and Gal-knockout mice. No deposits of IgG, IgM, or C3 could be detected. Almost no difference could be seen between immunosuppressed Gal-knockout mice and immunosuppressed ordinary mice. In immunosuppressed, immunized Gal-knockout mice, the results were similar. In ordinary mice treated with CsA+DSG, the levels of anti-Gal IgM were lower than they were in untreated mice, whereas the levels of anti-Gal IgG were similar. In Gal-knockout mice (including immunized animals) treated with CsA+DSG, the levels of anti-Gal IgG and IgM were lower than they were in untreated Gal-knockout mice. After renal subcapsular transplantation, antibodies against Galalpha1-3Gal have no major influence on islet xenograft rejection in the pig-to-mouse model. Immunosuppression, which inhibits rejection in the pig-to-mouse model, is equally effective when transplantation is performed across the Galalpha1-3Gal barrier.

Anti-Gal alpha 1-3Gal IgM and IgG antibody levels in sera of humans and old world non-human primates.

Organs transplanted from pig to primate are rejected within minutes or hours by an antibody-dependent, complement-mediated mechanism [hyperacute rejection (HAR)]. Even after depletion of anti-Gal alpha 1-3Gal (Gal) antibody (Ab), for example by extracorporeal immunoadsorption, return of natural Ab is believed to be a major factor in the initiation of acute humoral xenograft rejection. Various non-human primates are used as recipients of pig organs in experimental discordant xenotransplantation (XTx) models. However, anti-Gal IgM and IgG levels in non-human primates may differ from those in humans. Serum levels of anti-Gal IgM and IgG were measured by enzyme-linked immunosorbent assay (ELISA) in humans (n=14), chimpanzees (n=8), baboons (n=214), cynomolgus monkeys (n=29), rhesus monkeys (n=23) and Japanese monkeys (n=6). The mean level of anti-Gal IgM was significantly higher in chimpanzees than in other groups, while in rhesus monkeys it was significantly lower than in other groups, except baboons and Japanese monkeys. The mean human anti-Gal IgG level was higher than in other groups and this difference reached statistical significance except with regard to chimpanzees. The mean anti-Gal IgG level in baboons was significantly lower than that in humans, chimpanzees and cynomolgus monkeys. The measured differences in anti-Gal IgM and IgG levels may affect the kinetics of Ab removal and rate of return in different species, and thus may have relevance for translating work in non-human primate models to the clinical setting.

PIG KIDNEY TRANSPLANTATION IN BABOONS: Anti-Gal??1-3Gal IgM Alone Is Associated with Acute Humoral Xenograft Rejection and Disseminated Intravascular Coagulation1

Kidneys harvested from miniature swine or pigs transgenic for human decay-accelerating factor (hDAF) were transplanted into baboons receiving an anti-CD154 monoclonal antibody (mAb) and either a whole body irradiation (WBI)- or cyclophosphamide (CPP)-based immunosuppressive regimen. Group 1 baboons (n=3) underwent induction therapy with WBI and thymic irradiation, pretransplantation antithymocyte globulin, and immunoadsorption of anti-Gal(alpha)1-3Gal (Gal) antibody (Ab). After transplantation of a miniature swine kidney, maintenance therapy comprised cobra venom factor, mycophenolate mofetil, and an anti-CD154 mAb (for 14-28 days). In group 2 (n=2), WBI was replaced by CPP in the induction protocol. Group 3 (n=3) animals received the group 2 regimen, but underwent transplantation with hDAF pig kidneys. Group 1 and 2 animals developed features of disseminated intravascular coagulation (DIC), with reductions of fibrinogen and platelets and increases of prothrombin time, partial thromboplastin time, and fibrin split products. Graft survival was for 6-13 days. Histology showed mild acute humoral xenograft rejection (AHXR) of the kidneys, but severe rejection of the ureters. Group 3 animals developed features of DIC in two of three cases during the fourth week, with AHXR in the third case. Graft survival was for 28 (n=1) or 29 (n=2) days. Histology of day 15 biopsy specimens showed minimal focal mononuclear cellular infiltrates, with predominantly CD3+ cells. By days 28 and 29, kidneys showed mild-to-moderate features of AHXR. In all groups, the humoral response was manifest by reappearance of anti-Gal IgM below baseline level, with no or low return of anti-Gal IgG. All excised kidneys showed IgM deposition, but no complement and no or minimal IgG deposition. No baboon showed a rebound of anti-Gal Ab immediately after excision of the graft, and anti-Gal Ab increased over pretransplantation levels only when anti-CD154 mAb was discontinued. DIC was observed with WBI- or CPP-based therapy, and after miniature swine or hDAF kidney transplantation. AHXR+/-DIC was observed in all recipients even in the absence of complement and no or low levels of anti-Gal IgG, but was significantly delayed in the hDAF recipients. These results confirm our earlier observation that CD154 blockade prevents T cell-dependent sensitization in baboons to pig antigens, but that baseline natural anti-Gal Ab production is not inhibited. We suggest that IgM deposition, even in the absence of IgG and complement, leads to endothelial cell activation with the development of DIC, even when there are only minimal histologic changes of AHXR.

Tolerization of Gal alpha 1,3Gal-reactive B cells in pre-sensitized alpha 1,3-galactosyltransferase-deficient mice by nonmyeloablative induction of mixed chimerism.

Using a alpha 1,3-galactosyltransferase wild-type (GalT(+/+)) to deficient (GalT(-/-)) mouse bone marrow transplantation model, we have previously demonstrated that a non-myeloablative conditioning regimen is capable of permitting induction of allogeneic and xenogeneic mixed chimerism. Chimerism is associated with the rapid and lasting tolerization of anti-Gal alpha 1,3Gal (Gal) natural antibody (Ab)-producing B cells. However, one limitation of this model is that anti-Gal natural Ab levels are lower in GalT(-/-) mice than in humans and other primates. To overcome this limitation, we have now investigated the possibility of inducing such tolerance in GalT(-/-) mice that produce much higher levels of anti-Gal Abs due to presensitization with Gal-bearing xenogeneic cells. B6 GalT(-/-) mice that were pre-sensitized with rabbit red blood cells received non-myeloablative conditioning with depleting anti-CD4 and CD8 mAbs, 3Gy whole body and 7Gy thymic irradiation, and infusion of BALB/c GalT(+/+) bone marrow cells (BMC). Although engraftment of standard marrow doses was inhibited by the presensitization, long-lasting mixed chimerism could be induced in recipients of a high dose [160 x 10(6)] of allogeneic wild-type BMC. Achievement of persistent chimerism was associated with high levels of anti-Gal IgG(1) pretransplant, suggesting an inhibitory effect of non-complement-fixing IgG(1) Ab on anti-Gal-mediated marrow rejection. Induction of mixed chimerism was associated with a rapid disappearance of serum anti-Gal and tolerization of anti-Gal Ab-producing cells. B cells with anti-Gal receptors became undetectable in mixed chimeras. Mixed chimeras accepted subsequently transplanted donor-type GalT(+/+) hearts (> 140 days), whereas rapid (within 2 days) rejection of GalT(+/+) hearts occurred in conditioned control GalT(-/-) mice. In conclusion, when a high dose of GalT(+/+) BMC was administered to pre-sensitized GalT(-/-) mice, chimerism and tolerance were achieved. The absence of B cells with receptors recognizing Gal in mixed chimeras suggests a role for clonal deletion/receptor editing in the maintenance of B cell tolerance.