Killer Cell Immunoglobulin-Like Receptor Alleles Alter HIV Disease in Children

Background

HLA class I molecules are ligands for killer cell immunoglobin like receptors (KIR) that control the antiviral response of natural killer (NK) cells. However, the effects of KIR and HLA (KIR/HLA) alleles on HIV disease of children have not been studied.

Methods

993 antiretroviral naïve children with symptomatic HIV infection from PACTG protocols P152 and P300 were genotyped for KIR and HLA alleles using the Luminex platform. Linear regression was used to test the association between genotypes and baseline pre-ART HIV RNA, CD4⁺ lymphocyte count, and cognitive score, adjusting for age, race/ethnicity and study. The interaction between genetic markers and age was investigated. To account for multiple testing the false discovery rate (FDR) was controlled at 0.05.

Results

Children with the KIR2DS4*ALL FULL LENGTH (KIR2DS4*AFL) allele had higher CD4⁺ lymphocyte counts. Among children ≤2 years of age, the KIR2DS4*AFL was associated with lower plasma HIV RNA and higher cognitive index scores. KIR Cent2DS3/5_1 had lower CD4⁺ lymphocyte counts in children ≤2 years of age, while the presence of Tel1, Tel2DS4_2, Tel2DS4_4, Tel8, Tel2DS4_6 had higher CD4⁺ lymphocyte counts in all children. Presence of Cent2, Cent4 and Cent8 was associated with increased HIV RNA load in children ≤2 years. Presence of KIR3DL1+Bw4 was associated with higher CD4⁺ lymphocyte counts in all children. Among children >2 years old, KIR3DS1+Bw4-80I was associated with higher plasma HIV RNA, and Bw6/Bw6 was associated with lower plasma HIV RNA compared to children with KIR3DS1+Bw4-80I.

Conclusions

Presented data show for the first time that specific KIR alleles independently or combined with HLA ligands are associated with HIV RNA and CD4⁺ lymphocyte counts in infected, antiretroviral naive children; and many of these effect estimates appear to be age dependent. These data support a role for specific KIR alleles in HIV pathogenesis in children.     

Genetic Diversity of the KIR/HLA System and Outcome of Patients with Metastatic Colorectal Cancer Treated with Chemotherapy

Objective

To explore genes of the killer-cell immunoglobulin-like receptor (KIR) and of the HLA ligand and their relationship with the outcome of metastatic colorectal cancer (mCRC) patients treated with first-line 5-fluorouracil, leucovorin, and irinotecan (FOLFIRI).

Methods

A total of 224 mCRC patients were screened for KIR/HLA typing. The determination of the KIR/HLA combinations was based upon the gene content and variants. Genetic associations with complete response (CR), time to progression (TTP) and overall survival (OS) were evaluated by calculating odds and hazard ratios. Multivariate modeling with prognostic covariates was also performed.

Results

For CR, the presence of KIR2DL5A, 2DS5, 2DS1, 3DS1, and KIR3DS1/HLA-Bw4-I80 was associated with increased CR rates, with median ORs ranging from 2.1 to 4.3, while the absence of KIR2DS4 and 3DL1 was associated with increased CR rates (OR 3.1). After univariate analysis, patients that underwent resective surgery of tumor, absence of KIR2DS5, and presence of KIR3DL1/HLA-Bw4-I80 showed a significant better OS (HR 1.5 to 2.8). Multivariate analysis identified as parameters independently related to OS the type of treatment (surgery; HR 2.0) and KIR3DL1/HLA-Bw4-I80 genotype (HR for T-I80 2.7 and for no functional KIR/HLA interaction 1.8). For TTP, no association with KIR/HLA genes was observed.

Conclusion

This study, for the first time, evidences that the genotyping for KIR-HLA pairs are found predictive markers associated with complete response and improves overall survival prediction of FOLFIRI treatment response in metastatic colorectal cancer. These results suggest a role of the KIR/HLA system in patient outcome, and guide new research on the immunogenetics of mCRC through mechanistic studies and clinical validation.     

Does the KIR2DS5 Gene Protect from Some Human Diseases?

Background

KIR2DS5 gene encodes an activating natural killer cell receptor whose ligand is not known. It was recently reported to affect the outcome of hematopoietic stem cell transplantation.

Methodology/Principal Findings

In our studies on KIR2DS5 gene associations with human diseases, we compared the frequencies of this gene in patients and relevant controls. Typing for KIR2DS5 gene was performed by either individual or multiplex polymerase chain reactions which, when compared in the same samples, gave concordant results. We noted an apparently protective effect of KIR2DS5 gene presence in several clinical conditions, but not in others. Namely, this effect was observed in ankylosing spondylitis (p = 0.003, odds ratio [OR] = 0.47, confidence interval [CI] = 0.28–0.79), endometriosis (p = 0.03, OR = 0.25, CI = 0.07–0.82) and acute rejection of kidney graft (p = 0.0056, OR = 0.44, CI = 0.24–0.80), but not in non-small-cell lung carcinoma, rheumatoid arthritis, spontaneous abortion, or leukemia (all p>0.05). In addition, the simultaneous presence of KIR2DS5 gene and HLA-C C1 allotype exhibited an even stronger protective effect on ankylosing spondylitis (p = 0.0003, OR = 0.35, CI = 0.19–0.65), whereas a lack of KIR2DS5 and the presence of the HLA-C C2 allotype was associated with ankylosing spondylitis (p = 0.0017, OR = 1.92, CI = 1.28–2.89), whereas a lack of KIR2DS5 and presence of C1 allotype was associated with rheumatoid arthritis (p = 0.005, OR = 1.47, CI = 1.13–1.92). The presence of both KIR2DS5 and C1 seemed to protect from acute kidney graft rejection (p = 0.017, OR = 0.47, CI = 0.25–0.89), whereas lack of KIR2DS5 and presence of C2 seemed to favor rejection (p = 0.0015, OR = 2.13, CI = 1.34–3.37).

Conclusions/Significance

Our results suggest that KIR2DS5 may protect from endometriosis, ankylosing spondylitis, and acute rejection of kidney graft.     

Natural Killer KIR3DS1 Is Closely Associated with HCV Viral Clearance and Sustained Virological Response in HIV/HCV Patients

Aim

To evaluate the influence of the presence of the killer cell immunoglobulin-like receptor (KIR) 3DS1 on HCV treatment response in HIV/HCV genotype 1 co-infected patients

Methods

HIV/HCV co-infected patients were included. KIR3DS1, their specific HLA-B ligands and IL28B gene were genotyped. Reductions of plasma HCV RNA levels between baseline and week 1, week 2 and week 4 were analyzed for IL28B genotype and KIR3DS1 (HLA Bw4 or Bw6). Rapid and sustained virological response (RVR and SVR) rates were also analyzed.

Results

Sixty HIV/HCV genotype 1 co-infected patients were included. Patients with KIR3DS1 and Bw4 had higher rates of HCV viral decline than those who were not carriers of KIR3DS1 (week1: p = 0.01; week2: p = 0.038; week 4: p = 0.03). Patients carrying KIR3DS1/Bw4 had higher rates of RVR and SVR than those who did not carry KIR3DS1 (RVR: 46.15% versus 17.02%, p = 0.012; SVR: 63.6% versus 13 26.5%, p = 0.031). With respect to patients carrying the IL28B-CC genotype, those with KIR3DS1/Bw4 had greater rates of HCV viral clearance (week1: p<0.001; week2: p = 0.01; week 4: p = 0.02), RVR (p = 0.015) and SVR (p = 0.029) than those not carrying KIR3DS1.

Conclusion

Our results show that the KIR3DS1 genotype has a positive effect on HCV viral clearance during the first weeks of Peg-IFN/RBV treatment in HCV/HCV co-infected patients bearing genotype 1, and higher RVR and SVR rates.     

Exploring the Role of Killer Cell Immunoglobulin-Like Receptors and Their HLA Class I Ligands in Autoimmune Hepatitis

Background

Natural killer cells are involved in the complex mechanisms underlying autoimmune diseases but few studies have investigated their role in autoimmune hepatitis. Killer immunoglobulin-like receptors are key regulators of natural killer cell-mediated immune responses.

Methods and Findings

KIR gene frequencies, KIR haplotypes, KIR ligands and combinations of KIRs and their HLA Class I ligands were investigated in 114 patients diagnosed with type 1 autoimmune hepatitis and compared with a group of 221 healthy controls. HLA Class I and Class II antigen frequencies were compared to those of 551 healthy unrelated families representative of the Sardinian population. In our cohort, type 1 autoimmune hepatitis was strongly associated with the HLA-B18, Cw5, DR3 haplotype. The KIR2DS1 activating KIR gene and the high affinity HLA-C2 ligands were significantly higher in patients compared to controls. Patients also had a reduced frequency of HLA-Bw4 ligands for KIR3DL1 and HLA-C1 ligands for KIR2DL3. Age at onset was significantly associated with the KIR2DS1 activating gene but not with HLA-C1 or HLA-C2 ligand groups.

Conclusions

The activating KIR gene KIR2DS1 resulted to have an important predictive potential for early onset of type 1 autoimmune hepatitis. Additionally, the low frequency of the KIR-ligand combinations KIR3DL1/HLA-Bw4 and KIR2DL3/HLA-C1 coupled to the high frequency of the HLA-C2 high affinity ligands for KIR2DS1 could contribute to unwanted NK cell autoreactivity in AIH-1.     

Polymorphisms of killer immunoglobulin-like receptors (KIRs) and HLA ligands in northeastern Thais

Killer cell immunoglobulin-like receptors (KIRs) are cell surface receptors on natural killer (NK) cells and subsets of T cells. The functions of NK cells are partly regulated by interactions between KIRs and HLA ligands on target cells. In this study, the presence or absence of 17 KIR genes and their known HLA ligands have been investigated in 235 unrelated individuals living in northeastern Thailand (NET). Subtypes of KIR2DS4 including full length (KIR2DS4F) and deleted forms (KIR2DS4D) have also been determined. Framework genes (KIR2DL4, 3DL2, 3DL3, and 3DP1) were found in all individuals and KIR genes belonging to the A haplotype (KIR2DL1, 2DL3, 3DL1, and 2DS4) were present in more than 90 % of NET. KIR2DS4D (61.7 %) was more common than KIR2DS4F (52.8 %). A total of 33 different KIR genotypes were observed. Of these, three new genotypes were identified. The most common genotype (AA) was observed in 35.7 % of NET, and HLA-C alleles bearing the C1 epitope (HLA-C1) had the highest frequency (97 %). All individuals had at least one inhibitory KIR and its corresponding HLA ligand; 40.9 % of NET had three pairs of receptor–ligand combinations, and 18.3 % had all three receptor–ligand combinations of KIR2DL3+C1, 3DL1+Bw4, and 3DL2+A11. Surprisingly, the patterns of KIR gene frequencies in NET are more similar to those of Caucasians than Japanese, Korean, and Chinese. This is the first report on complete analysis of KIR and known HLA ligands in Thais. These data provide basic knowledge on KIR for further studies on disease associations and transplantation in northeastern Thais.     

Association of HLA Class-I and Inhibitory KIR Genotypes in Gabonese Patients Infected by Chikungunya or Dengue Type-2 Viruses

Background

Natural killer (NK) cells provide defense in the early stages of the immune response against viral infections. Killer cell immunoglobulin-like receptors (KIR) expressed on the surface of NK cells play an important role in regulating NK cell response through recognition of human leukocyte antigen (HLA) class I molecules on target cells. Previous studies have shown that specific KIR/ligand combinations are associated with the outcome of several viral infectious diseases.

Methods

We investigated the impact of inhibitory and activating KIR and their HLA-class I ligand genotype on the susceptibility to Chikungunya virus (CHIKV) and Dengue virus (DENV2) infections. From April to July 2010 in Gabon, a large outbreak of CHIKV and DENV2 concomitantly occurred in two provinces of Gabon (Ogooué-Lolo and Haut-Ogooué). We performed the genotypic analysis of KIR in the combination with their cognate HLA-class I ligands in 73 CHIKV and 55 DENV2 adult cases, compared with 54 healthy individuals.

Results

We found in CHIV-infected patients that KIR2DL1 and KIR2DS5 are significantly increased and decreased respectively, as compared to DENV2⁺ patients and healthy donors. The combination of KIR2DL1 and its cognate HLA-C2 ligand was significantly associated with the susceptibility to CHIKV infection. In contrast, no other inhibitory KIR-HLA pairs showed an association with the two mosquito-borne arboviruses.

Conclusion

These observations are strongly suggestive that the NK cell repertoire shaped by the KIR2DL1:HLA-C2 interaction facilitate specific infection by CHIKV.     

Low CD4+ T cell counts among African HIV-1 infected subjects with group B KIR haplotypes in the absence of specific inhibitory KIR ligands

Natural killer (NK) cells are regulated by interactions between polymorphic killer immunoglobulin-like receptors (KIR) and human leukocyte antigens (HLA). Genotypic combinations of KIR3DS1/L1 and HLA Bw4-80I were previously shown to influence HIV-1 disease progression, however other KIR genes have not been well studied. In this study, we analyzed the influence of all activating and inhibitory KIR, in association with the known HLA inhibitory KIR ligands, on markers of disease progression in a West African population of therapy-naïve HIV-1 infected subjects. We observed a significant association between carriage of a group B KIR haplotype and lower CD4+ T cell counts, with an additional effect for KIR3DS1 within the frame of this haplotype. In contrast, we found that individuals carrying genes for the inhibitory KIR ligands HLA-Bw4 as well as HLA-C1 showed significantly higher CD4+ T cell counts. These associations were independent from the viral load and from individual HIV-1 protective HLA alleles. Our data suggest that group B KIR haplotypes and lack of specific inhibitory KIR ligand genes, genotypes considered to favor NK cell activation, are predictive of HIV-1 disease progression.     

KIR and HLA loci are associated with hepatocellular carcinoma development in patients with hepatitis B virus infection: a case-control study

Background

Natural killer (NK) cells activation has been reported to contribute to inflammation and liver injury during hepatitis B virus (HBV) infection both in transgenic mice and in patients. However, the role of NK cells in the process of HBV-associated hepatocellular carcinoma (HCC) development has not been addressed. Killer cell immunoglobulin-like receptors (KIRs) are involved in regulating NK cell activation through recognition of specific human leukocyte antigen (HLA) class I allotypes.

Methodology/Principal Findings

To investigate whether KIR and HLA genes could influence the risk of HBV-associated HCC development, 144 HBV-infected patients with HCC and 189 well-matched HBV infectors with chronic hepatitis or cirrhosis as non-HCC controls were enrolled in this study. The presence of 12 loci of KIR was detected individually. HLA-A, -B, -C loci were genotyped with high-resolution. HLA-C group 1 homozygote (OR = 2.02; p = 0.005), HLA-Bw4-80I (OR = 2.67; p = 2.0E-04) and combination of full-length form and 22 bp-deleted form of KIR2DS4 (KIR2DS4/1D) (OR = 1.89; p = 0.017) were found associated with HCC incidence. When the combined effects of these three genetic factors were evaluated, more risk factors were observed correlating with higher odds ratios for HCC incidence (P trend = 7.4E-05). Because all the risk factors we found have been reported to result in high NK cell functional potential by previous studies, our observations suggest that NK cell activation may contribute to HBV-associated HCC development.

Conclusions/Significance

In conclusion, this study has identified significant associations that suggest an important role for NK cells in HCC incidence in HBV-infected patients. Our study is useful for HCC surveillance and has implications for novel personalized therapy strategy development aiming at HCC prevention in HBV-infected patients.     

Recipient-Related Risk Factors for Graft Failure and Death in Elderly Kidney Transplant Recipients

Background

Elderly patients with end-stage renal disease have become the fastest growing population of kidney transplant candidates in recent years. However, the risk factors associated with long-term outcomes in these patients remain unclear.

Methods

We retrospectively analyzed 166 recipients aged 60 years or older who underwent primary deceased kidney transplantation between 2002 and 2013 in our center. The main outcomes included 1-, 3- and 5-year patient survival as well as overall and death-censored graft survival. The independent risk factors affecting graft and patient survival were analyzed using Cox regression analysis.

Results

The 1-, 3-, 5-year death-censored graft survival rates were 93.6%, 89.4% and 83.6%, respectively. Based on the Cox multivariate analysis, panel reactive antibody (PRA)>5% [hazard ratio (HR) 4.295, 95% confidence interval (CI) 1.321–13.97], delayed graft function (HR 4.744, 95% CI 1.611–13.973) and acute rejection (HR 4.971, 95% CI 1.516–16.301) were independent risk factors for graft failure. The 1-, 3-, 5-year patient survival rates were 84.8%, 82.1% and 77.1%, respectively. Longer dialysis time (HR 1.011 for 1-month increase, 95% CI 1.002–1.020), graft loss (HR 3.501, 95% CI 1.559–7.865) and low-dose ganciclovir prophylaxis (1.5 g/d for 3 months) (HR 3.173, 95% CI 1.063–9.473) were risk factors associated with patient death.

Conclusions

The five-year results show an excellent graft and patient survival in elderly kidney transplant recipients aged ≥60 years. PRA>5%, delayed graft function, and acute rejection are risk factors for graft failure, while longer duration of dialysis, graft loss and low-dose ganciclovir prophylaxis are risk factors for mortality in elderly recipients. These factors represent potential targets for interventions aimed at improving graft and patient survival in elderly recipients.     

KIR-HLA genotypes in HIV-infected patients lacking immunological recovery despite effective antiretroviral therapy

Background

In HIV-infected individuals, mechanisms underlying unsatisfactory immune recovery during effective combination antiretroviral therapy (cART) have yet to be fully understood. We investigated whether polymorphism of genes encoding immune-regulating molecules, such as killer immunoglobulin-like receptors (KIR) and their ligands class I human leukocyte antigen (HLA), could influence immunological response to cART.

Methods

KIR and HLA frequencies were analyzed in 154 HIV-infected and cART-treated patients with undetectable viral load divided into two groups: ‘immunological non responders’ (INR, N = 50, CD4⁺ T-cell count <200/mm³) and full responders (FR, N = 104, CD4⁺ T-cell count >350/mm³). Molecular KIR were typed using polymerase chain reaction-based genotyping. Comparisons were adjusted for baseline patient characteristics.

Results

The frequency of KIR2DL3 allele was significantly higher in FR than in INR (83.7% vs. 62%, P = 0.005). The functional compound genotype HLA-C1⁺/KIR2DL3⁺, even at multivariable analysis, when adjusted for nadir CD4⁺ T-cell count, was associated with reduced risk of INR status: odds ratio (95% Confidence Intervals) 0.34 (0.13−0.88), P = 0.03.

Conclusions

Reduced presence of the inhibitory KIR2DL3 genotype detected in INR might provoke an imbalance in NK function, possibly leading to increased immune activation, impaired killing of latently infected cells, and higher proviral burden. These factors would hinder full immune recovery during therapy.     

Ex vivo generation of human alloantigen-specific regulatory T cells from CD4(pos)CD25(high) T cells for immunotherapy

Background

Regulatory T cell (Treg) based immunotherapy is a potential treatment for several immune disorders. By now, this approach proved successful in preclinical animal transplantation and auto-immunity models. In these models the success of Treg based immunotherapy crucially depends on the antigen-specificity of the infused Treg population. For the human setting, information is lacking on how to generate Treg with direct antigen-specificity ex vivo to be used for immunotherapy.

Methodology/Principal Findings

Here, we demonstrate that in as little as two stimulation cycles with HLA mismatched allogeneic stimulator cells and T cell growth factors a very high degree of alloantigen-specificity was reached in magnetic bead isolated human CD4^posCD25^high Treg. Efficient increases in cell numbers were obtained. Primary allogeneic stimulation appeared a prerequisite in the generation of alloantigen-specific Treg, while secondary allogeneic or polyclonal stimulation with anti-CD3 plus anti-CD28 monoclonal antibodies enriched alloantigen-specificity and cell yield to a similar extent.

Conclusions/Significance

The ex vivo expansion protocol that we describe will very likely increase the success of clinical Treg-based immunotherapy, and will help to induce tolerance to selected antigens, while minimizing general immune suppression. This approach is of particular interest for recipients of HLA mismatched transplants.     

KIR and HLA-C interactions promote differential dendritic cell maturation and is a major determinant of graft failure following kidney transplantation

Background

HLA-C is an important ligand for killer immunoglobulin like receptors (KIR) that regulate natural killer (NK) cell function. Based on KIR specificity HLA-C molecules are allocated into two groups, HLA-C1 or HLA-C2; HLA-C2 is more inhibiting to NK cell function than HLA-C1. We studied the clinical importance of HLA-C genotypes on the long-term graft survival of 760 kidney transplants performed at our centre utilising a population based genetic study and cell culture model to define putative mechanisms.

Methods and Findings

Genotyping was performed using conventional DNA PCR techniques and correlations made to clinical outcomes. We found that transplant recipients with HLA-C2 had significantly better long-term graft survival than transplant recipients with HLA-C1 (66% versus 44% at 10 years, log-rank p = 0.002, HR = 1.51, 95%CI = 1.16–1.97). In in-vitro NK and dendritic cell (DC) co-culture model we made several key observations that correlated with the population based genetic study. We observed that donor derived NK cells, on activation with IL-15, promoted differential HLA-C genotype dependent DC maturation. In NK-DC co-culture, the possession of HLA-C2 by DC was associated with anti-inflammatory cytokine production (IL-1RA/IL-6), diminished DC maturation (CD86, HLA-DR), and absent CCR7 expression. Conversely, possession of HLA-C1 by DC was associated with pro-inflammatory cytokine synthesis (TNF-α, IL-12p40/p70), enhanced DC maturation and up-regulation of CCR7 expression. By immunohistochemistry the presence of donor NK cells was confirmed in pre-transplant kidneys.

Conclusions

We propose that after kidney transplantation IL-15 activated donor derived NK cells interact with recipient DC with less activation of indirect allo-reactivity in HLA-C2 positive recipients than HLA-C1 positive recipients; this has implications for long-term graft survival. Early events following kidney transplantation involving NK-DC interaction via KIR and HLA-C immune synapse may have a central role in long-term kidney transplant outcomes.     

Conjunctival Scarring in Trachoma Is Associated with the HLA-C Ligand of KIR and Is Exacerbated by Heterozygosity at KIR2DL2/KIR2DL3

Background

Chlamydia trachomatis is globally the predominant infectious cause of blindness and one of the most common bacterial causes of sexually transmitted infection. Infections of the conjunctiva cause the blinding disease trachoma, an immuno-pathological disease that is characterised by chronic conjunctival inflammation and fibrosis. The polymorphic Killer-cell Immunoglobulin-like Receptors (KIR) are found on Natural Killer cells and have co-evolved with the Human Leucocyte Antigen (HLA) class I system. Certain genetic constellations of KIR and HLA class I polymorphisms are associated with a number of diseases in which modulation of the innate responses to viral and intracellular bacterial pathogens is central.

Methodology

A sample of 134 Gambian pedigrees selected to contain at least one individual with conjunctival scarring in the F₁ generation was used. Individuals (n = 830) were genotyped for HLA class I and KIR gene families. Family Based Association Tests and Case Pseudo-control tests were used to extend tests for transmission disequilibrium to take full advantage of the family design, genetic model and phenotype.

Principle findings

We found that the odds of trachomatous scarring increased with the number of genome copies of HLA-C2 (C1/C2 OR = 2.29 _BHP-value = 0.006; C2/C2 OR = 3.97 _BHP-value = 0.0004) and further increased when both KIR2DL2 and KIR2DL3 (C2/C2 OR = 5.95 _BHP-value = 0.006) were present.

Conclusions

To explain the observations in the context of chlamydial infection and trachoma we propose a two-stage model of response and disease that balances the cytolytic response of KIR expressing NK cells with the ability to secrete interferon gamma, a combination that may cause pathology. The data presented indicate that HLA-C genotypes are important determinants of conjunctival scarring in trachoma and that KIR2DL2/KIR2DL3 heterozygosity further increases risk of conjunctival scarring in individuals carrying HLA-C2.     

Adiponectin Mediated MHC Class II Mismatched Cardiac Graft Rejection in Mice Is IL-4 Dependent

Background

Adiponectin regulates glucose and fatty-acid metabolism but its role in chronic graft rejection mediated by Th2 cytokines remains ill-defined.

Methodology/Principal Findings

Wild type and adiponectin-null mice were used as graft recipients in mouse MHC class II disparate cardiac transplantation (bm12 toB6) and the graft rejection was monitored. In adiponectin-null mice we observed that the cellular infiltrate of eosinophils, CD4⁺ and CD8⁺ T cells was reduced in grafts compared to the controls as was collagen deposition and vessel occlusion. A similar outcome was observed for skin transplants except that neutrophil infiltration was increased. Low levels of IL-4 were detected in the grafts and serum. The effect of adiponectin signaling on IL-4 expression was further investigated. Treatment with AMPK and p38 MAPK inhibitors blocked adiponectin enhanced T cell proliferation in mixed lymphocyte reactions. Inhibition of AMPK reduced eosinophil infiltration in skin grafts in wild type recipients and in contrast AMPK activation increased eosinophils in adiponectin-null recipients. The addition of adiponectin increased IL-4 production by the T cell line EL4 with augmented nuclear GATA-3 and phospho-STAT6 expression which were suppressed by knockdown of adiponectin receptor 1 and 2.

Conclusions

Our results demonstrate a direct effect of adiponectin on IL-4 expression which contributes to Th2 cytokine mediated rejection in mouse MHC class II histoincompatible transplants. These results add to our understanding of the interrelationship of metabolism and immune regulation and raise the possibility that AMPK inhibitors may be beneficial in selected types of rejection.     

KIR3DL1/S1 genotypes and KIR2DS4 allelic variants in the AB KIR genotypes are associated with Plasmodium-positive individuals in malaria infection

The importance of innate immunity in malaria has been suggested for early protection from maturation and multiplication of Plasmodium parasites injected via infected mosquitoes. In this study, the killer cell immunoglobulin-like receptor (KIR) genes in innate immunity were investigated for an association with malaria in the comparison between Plasmodium-positive and Plasmodium-negative Melanesian individuals in the Solomon Islands, one of the most hyperendemic malaria regions in the world. The higher frequency of a pair of KIR3DL1 and KIR2DS4 was observed in the Plasmodium-positive individuals, which led to the investigation of KIR3DL1/S1 genotypes in concert with KIR2DS4 allelic variants. The positive individuals showed the highest frequency of KIR3DL1/KIR3DS1 heterozygosity, which might suggest the masking of activating KIR3DS1 by inhibitory KIR3DL1 at allelic levels to maintain the KIR3DS1-driven activation of natural killer cells diminished in controlling Plasmodium proliferation. The extended analysis with A/B genotypes further revealed the trend of parasitic positive individuals to be KIR3DL1/KIR3DS1 heterozygous in pair with KIR2DS4 nondeleted variants in a set of KIR genes inheritable as the AB genotypes. To the best of our knowledge, this study is the first KIR investigation of the malaria-infected population, which strengthened the potential associations of KIR with malaria pathogenesis. The balance of inhibitory and activating KIR3D genes (KIR3DL1/S1) and membrane-bound or secreted status of KIR2DS4 alleles in the interaction with the other KIR genes in the AB genotypes might constitute a part of KIR characteristics to determine resistance or susceptibility to Plasmodium parasitic infection.     

Co-Graft of Allogeneic Immune Regulatory Neural Stem Cells (NPC) and Pancreatic Islets Mediates Tolerance,while Inducing NPC-Derived Tumors in Mice

Background

Data available on the immunomodulatory properties of neural stem/precursor cells (NPC) support their possible use as modulators for immune-mediated process. The aim of this study was to define whether NPC administered in combination with pancreatic islets prevents rejection in a fully mismatched allograft model.

Methodology/Principal Finding

Diabetic Balb/c mice were co-transplanted under the kidney capsule with pancreatic islets and GFP⁺ NPC from fully mismatched C57BL/6 mice. The following 4 groups of recipients were used: mice receiving islets alone; mice receiving islets alone and treated with standard immunosuppression (IL-2Rα chain mAbs + FK506 + Rapamycin); mice receiving a mixed islet/NPC graft under the same kidney capsule (Co-NPC-Tx); mice receiving the islet graft under the left kidney capsule and the NPC graft under the right kidney capsule (NPC-Tx). Our results demonstrate that only the co-transplantation and co-localization of NPC and islets (Co-NPC-Tx) induce stable long-term graft function in the absence of immunosuppression. This condition is associated with an expansion of CD4⁺CD25⁺FoxP3⁺ T regulatory cells in the spleen. Unfortunately, stable graft function was accompanied by constant and reproducible development of NPC-derived cancer mainly sustained by insulin secretion.

Conclusion

These data demonstrate that the use of NPC in combination with islets prevents graft rejection in a fully mismatched model. However, the development of NPC-derived cancer raises serious doubts about the safety of using adult stem cells in combination with insulin-producing cells outside the original microenvironment.     

Conferral of enhanced natural killer cell function by KIR3DS1 in early human immunodeficiency virus type 1 infection

A flurry of recent reports on the role of activating and inhibitory forms of the killer cell immunoglobulin-like receptors (KIR) in natural killer (NK) cell activity against human immunodeficiency virus type 1 (HIV-1) have yielded widely divergent results. The role of the activating NK receptor encoded by the KIR3DS1 allele and its putative ligands, members of the HLA class I Bw4Ile80 cluster, in early HIV-1 disease is controversial. We selected 60 treatment-naïve adults for study from the OPTIONS cohort of individuals with early HIV-1 infection in San Francisco. We performed NK cell functional assays measuring gamma interferon (IFN-γ) and CD107a expression by NK cells in the unstimulated state and after stimulation by the major histocompatibility complex class I-deficient 721.221 B-lymphoblastoid cell line. In addition, we measured CD38 expression (a T-cell activation marker) on T and NK cells. Persons who have at least one copy of the KIR3DS1 gene had higher IFN-γ and CD107a expression in the unstimulated state compared to those who do not possess this gene. After stimulation, both groups experienced a large induction of IFN-γ and CD107a, with KIR3DS1 carriers achieving a greater amount of IFN-γ expression. Differences in effector activity correlating with KIR3DS1 were not attributable to joint carriage of HLA Bw4Ile80 and KIR3DS1. We detected a partial but not complete dependence of KIR3DS1 on the members of B*58 supertype (B*57 and B*58) leading to higher NK cell function. Possessing KIR3DS1 was associated with lower expression of CD38 on both CD8⁺ T and NK cells and with a loss or weakening of the known strong associations between CD8⁺ T-cell expression of CD38 mean fluorescence intensity and the HIV-1 viral load. We observed that possessing KIR3DS1 was associated with higher NK cell effector functions in early HIV-1 disease, despite the absence of HLA Bw4Ile80, a putative ligand of KIR3DS1. Carriage of KIR3DS1 was associated with diminished CD8⁺ T-cell activation, as determined by expression of CD38, and a disruption of the traditional relationship between viral load and activation in HIV-1 disease, which may lead to better clinical outcomes for these individuals.NK cell function is regulated by a family of receptors encoded by the killer cell immunoglobulin-like receptor (KIR) genes (18, 33). Within the KIR family, certain genes encode inhibitory receptors that recognize HLA class I ligands (i.e., HLA-Bw4 or HLA-C), whereas other KIR genes encode activating receptors which are not completely known. Studies on the role of KIRs in human immunodeficiency virus (HIV) disease have focused on the activating receptor encoded by the KIR3DS1 allele. However, recent genetic association and functional studies of KIR and HIV disease have yielded widely disparate results on the role of KIR3DS1 and its putative ligands, a subset of HLA class I-B alleles referred to as Bw4Ile80. The Bw4Ile80 cluster is a subset of HLA-B alleles that bear an isoleucine at position 80 in the α-1 helix, on the rim of the peptide-binding cleft. The inhibitory receptors encoded by KIR3DL1 alleles, which are highly related in the extracellular domains to the activating receptor encoded by KIR3DS1, specifically recognize HLA-Bw4 ligands (5). Because of this similarity, KIR3DS1 has been assumed to also recognize Bw4Ile80 ligands. In 2002, Martin et al. reported that HIV-infected individuals in the Multicenter AIDS Cohort Study possessing the KIR3DS1 allele demonstrated significantly delayed progression to AIDS, provided that the individuals also expressed a Bw4Ile80 allele (20).In 2005, Gaudieri et al. reported on the association of the entire KIR gene cluster and HLA class I in HIV disease progression in an Australian HIV cohort (8, 9). These authors observed a trend toward slowed CD4⁺ T-cell percent loss among those who carried both Bw4Ile80 and KIR3DS1 (8). However, this trend was not statistically significant, and Gaudieri et al. simultaneously observed an acceleration of time to AIDS (1987 definition) among joint KIR3DS1 and Bw4Ile80 carriers. In 2006, Qi et al. published a follow-up report from the Multicenter AIDS Cohort Study cohort documenting an association between the coexpression of KIR3DS1 and Bw4Ile80 and enhanced protection against certain opportunistic infections in HIV-infected individuals (26), an effect partially attributed to very modest differences in viral load. In 2007, our group observed that KIR3DS1 gene carriage was associated with higher CD4⁺ T-cell counts and hence protection against HIV type 1 (HIV-1) progression in early disease (4); however, we observed that this effect was not attributable to differences in the viral load and further was independent of Bw4Ile80. In other words, our analyses suggested that the KIR3DS1 and Bw4Ile80 genes were each associated with protection against HIV disease but via different mechanisms.Until recently, it was not clear if KIR3DS1 was expressed on the surface of NK cells; however, two recent reports have conclusively established that KIR3DS1 is expressed on NK cells (6, 24) and that expression is dose dependent, with higher expression for homozygotes. These studies also demonstrated that KIR3DS1 recognizes neither HLA-Bw4 nor HLA-Bw6 ligands, at least when these major histocompatibility complex (MHC) class I molecules are expressed on Epstein-Barr virus-transformed B-lymphoblastoid cell lines. Similarly, an independent study by another group reported that KIR3DS1 fails to bind to soluble Bw4Ile80 tetrameric complexes (10). In contrast, Alter et al. have presented results from in vitro cytotoxicity assays suggesting that target cells possessing HLA-Bw4Ile80 are better targets for NK cells possessing KIR3DS1 (1); however, no evidence was provided to confirm a physical interaction between the KIR3DS1 and HLA-Bw4 proteins.Here, we present a study of the NK cell phenotype and function in 60 treatment-naïve, recently HIV-1-infected persons with defined HLA-B and KIR3DS1/KIR3DL1 allotypes. We also measured the expression of CD38 on NK cells and CD8⁺ T cells, a widely used marker of disease progression and virulence in HIV research and a marker of immune activation. The expression of CD38, as measured by flow cytometry, is known to be elevated on CD8⁺ T cells in HIV disease, reaching steady-state levels in early HIV-1 infection (7), and predicts disease progression independently of the viral load (19). The individuals studied were selected from our recent genetic association study of KIR and HLA among 255 recently HIV-1-infected persons (4), in which KIR3DS1 carriage alone was associated with higher CD4⁺ T-cell counts, despite the absence of a difference in the viral loads. On the basis of these clinical findings, we performed this study to determine whether persons who carried the KIR3DS1 gene had enhanced NK cell phenotypic and functional profiles and if these profiles were further enhanced by carriage of the putative KIR3DS1 ligands encoded by HLA-Bw4Ile80 alleles. Flow cytometry-based detection of KIR3DS1 has been hampered by the absence of a monoclonal antibody that can bind to KIR3DS1 specifically and not cross-react with the related KIR3DL1 proteins (25). Hence, we used genotypic KIR assignments for our analyses rather than flow cytometry-based methods.     

Serum Bilirubin Affects Graft Outcomes through UDP-Glucuronosyltransferase Sequence Variation in Kidney Transplantation

Background

Oxidative stress is a major mediator of adverse outcome after kidney transplantation. Bilirubin is produced by heme oxygenase-1 (HO-1), catalyzed by UDP-glucuronosyltransferase (UGT1A1), and has potential as an antioxidant. In this study, we investigated the effects of HO-1 and UGT1A1 sequence variations on kidney allograft outcomes.

Methods

Clinical data were collected from 429 Korean recipients who underwent kidney transplantation from 1990–2008. Genotyping for UGT1A1*28 and HO-1 (A−413T) was performed. Acute rejection and graft survival were monitored as end-points.

Results

Serum levels of total bilirubin were significantly increased after transplantation (0.41±0.19 mg/dL to 0.80±0.33 mg/dL, P<0.001). Post-transplant 1-year bilirubin level was higher in 6/7 or 7/7 carriers compared with 6/6 homozygotes in terms of the UGT1A1*28 polymorphism (6/6 vs. 6/7 vs. 7/7: 0.71±0.27 vs. 1.06±0.36 vs. 1.10±0.45 mg/dL, P<0.001). According to an additive model of genotype analysis, the 7-allele genotype had a protective effect on the development of acute rejection compared with the 6-allele (odds ratio 0.43, 95% CI 0.25–0.73, P for trend = 0.006). Multivariate Cox regression analysis revealed that individuals carrying the 7-allele had a decreased risk of graft loss, by a factor of 0.36 (95% CI 0.15–0.85, P = 0.019). The HO-1 (A−413T) polymorphism had no effect on serum bilirubin levels or graft outcomes.

Conclusions

The UGT1A1*28 polymorphism is associated with changes in serum bilirubin and with graft outcome after kidney transplantation.