Anti-HLA alloantibodies of the IgA isotype in re-transplant candidates part II: Correlation with graft survival.

We reported previously on the widespread occurrence of anti-HLA alloantibodies of the IgA isotype (anti-HLA IgA) in the sera of solid-organ re-transplantation (re-tx) candidates (Arnold et al., ). Specifically focussing on kidney re-tx patients, we now extended our earlier findings by examining the impact of the presence and donor specificity of anti-HLA IgA on graft survival. We observed frequent concurrence of anti-HLA IgA and anti-HLA IgG in 27% of our multicenter collective of 694 kidney re-tx patients. This subgroup displayed significantly reduced graft survival as evidenced by the median time to first dialysis after transplantation (TTD 77 months) compared to patients carrying either anti-HLA IgG or IgA (TTD 102 and 94 months, respectively). In addition, donor specificity of anti-HLA IgA had a significant negative impact on graft survival (TTD 74 months) in our study. Taken together, our data strongly indicate that presence of anti-HLA IgA, in particular in conjunction with anti-HLA-IgG, in sera of kidney re-tx patients is associated with negative transplantation outcome.

Functional Fc gamma receptor gene polymorphisms and donor-specific antibody-triggered microcirculation inflammation.

Fc-dependent effector mechanisms may contribute to antibody-mediated rejection (ABMR), and distinct gene polymorphisms modifying the function of Fc gamma receptors (FcγRs) may influence the capability of donor-specific antibodies (DSAs) to trigger inflammation. To evaluate the relevance of functional FcγR variants in late ABMR, 85 DSA-positive kidney allograft recipients, who were recruited upon antibody screening of 741 prevalent patients, were genotyped for polymorphisms in FcγRIIA (FCGR2A-H/R131 ; rs1801274), FcγRIIIA (FCGR3A-V/F158 ; rs396991), and FcγRIIIB (FCGR3B-neutrophil antigen 1 ([NA1]/NA2; rs35139848). Individuals with high-affinity FCGR3A-V158 alleles (V/V158 or V/F158 ) showed a higher rate (and extent) of peritubular capillaritis (ptc) in protocol biopsies than homozygous carriers of the lower-affinity allele (ptc score ≥1: 53.6% vs 25.9%; P = .018). Associations were independent of C1q-binding to DSA or capillary C4d. In parallel, there was a trend toward increased macrophage- and injury-repair response-associated transcript subsets. Kidney function over 24 months, however, was not different. In support of a functional role of FcγRIIIA polymorphism, NK92 cells expressing FCGR3A-V158 produced >2 times as much interferon gamma upon incubation with HLA antibody-coated cells as those expressing FCGR3A-F158 . FcγRIIA and FcγRIIIB polymorphisms were not associated with allograft morphology. Our data suggest that the presence of high-affinity FcγRIIIA variants may favor DSA-triggered microcirculation inflammation.

Effects of weak/non-complement-binding HLA antibodies on C1q-binding.

It is unknown under what conditions and to what extent weak/non-complement (C)-binding IgG subclasses (IgG2/IgG4) can block C1q-binding triggered by C-binding IgG subclasses (IgG1/IgG3). Therefore, we investigated in vitro C1q-binding induced by IgG subclass mixtures targeting the same HLA epitope. Various mixtures of HLA class II specific monoclonal antibodies of different IgG subclasses but identical V-region were incubated with HLA DRB1*07:01 beads and monitored for C1q-binding. The lowest concentration to achieve maximum C1q-binding was measured for IgG3, followed by IgG1, while IgG2 and IgG4 did not show appreciable C1q-binding. C1q-binding occurred only after a critical amount of IgG1/3 has bound and sharply increased thereafter. When both, C-binding and weak/non-C-binding IgG subclasses were mixed, C1q-binding was diminished proportionally to the fraction of IgG2/4. A 2- to 4-fold excess of IgG2/4 inhibited C1q-binding by 50%. Very high levels (10-fold excess) almost completely abrogated C1q-binding even in the presence of significant IgG1/3 levels that would usually lead to strong C1q-binding. In sensitized renal allograft recipients, IgG subclass constellations with ≥ 2-fold excess of IgG2/4 over IgG1/3 were present in 23/66 patients (34.8%) and overall revealed slightly decreased C1q signals. However, spiking of patient sera with IgG2 targeting a different epitope than the patient's IgG1/3 synergistically increased C1q-binding. In conclusion, if targeting the same epitope, an excess of IgG2/4 is repressing the extent of IgG1/3 triggered C1q-binding in vitro. Such IgG subclass constellations are present in about a third of sensitized patients and their net effect on C1q-binding is slightly inhibitory.

Stroke in first-degree relatives of patients with cervical artery dissection.

BACKGROUND AND PURPOSE: Patients with ischaemic stroke (IS) caused by a spontaneous cervical artery dissection (CeAD) worry about an increased risk for stroke in their families. The occurrence of stroke in relatives of patients with CeAD and in those with ischaemic stroke attributable to other (non-CeAD) causes were compared. METHODS: The frequency of stroke in first-degree relatives (family history of stroke, FHS) was studied in IS patients (CeAD patients and age- and sex-matched non-CeAD patients) from the Cervical Artery Dissection and Ischemic Stroke Patients (CADISP) database. FHS ≤ 50 and FHS > 50 were defined as having relatives who suffered stroke at the age of ≤50 or >50 years. FHS ≤ 50 and FHS > 50 were studied in CeAD and non-CeAD IS patients and related to age, sex, number of siblings, hypertension, hypercholesterolemia, smoking and body mass index (BMI). RESULTS: In all, 1225 patients were analyzed. FHS ≤ 50 was less frequent in CeAD patients (15/598 = 2.5%) than in non-CeAD IS patients (38/627 = 6.1%) (P = 0.003; odds ratio 0.40, 95% confidence interval 0.22-0.73), also after adjustment for age, sex and number of siblings (P = 0.005; odds ratio 0.42, 95% confidence interval 0.23-0.77). The frequency of FHS > 50 was similar in both study groups. Vascular risk factors did not differ between patients with positive or negative FHS ≤ 50. However, patients with FHS > 50 were more likely to have hypertension and higher BMI. CONCLUSION: Relatives of CeAD patients had fewer strokes at a young age than relatives of non-CeAD IS stroke patients.

16(th) IHIW: anti-HLA alloantibodies of the of IgA isotype in re-transplant candidates.

In this multicentre study, sera from 803 retransplant candidates, including 775 kidney transplant recipients, were analysed with regard to the presence and specificity of anti-HLA alloantibodies of the IgA isotype using a modified microsphere-based platform. Of the kidney recipients, nearly one-third (n = 237, 31%) had IgA alloantibodies. Mostly, these antibodies were found in sera that also harboured IgG alloantibodies that could be found in a total of 572 (74%) of patients. Interestingly, IgA anti-HLA antibodies were preferentially targeting HLA class I antigens in contrast to those of the IgG isotype, which targeted mostly both HLA class I and II antigens. Donor specificity of the IgA alloantibodies could be established for over half of the 237 patients with IgA alloantibodies (n = 124, 52%). A further 58 patients had specificities against HLA-C or HLA-DP, for which no information regarding donor typing was available. In summary, these data showed in a large cohort of retransplant candidates that IgA alloantibodies occur in about one-third of patients, about half of these antibodies being donor specific.

Hybrid fitness, adaptation and evolutionary diversification: lessons learned from Louisiana Irises.

Estimates of hybrid fitness have been used as either a platform for testing the potential role of natural hybridization in the evolution of species and species complexes or, alternatively, as a rationale for dismissing hybridization events as being of any evolutionary significance. From the time of Darwin's publication of The Origin, through the neo-Darwinian synthesis, to the present day, the observation of variability in hybrid fitness has remained a challenge for some models of speciation. Yet, Darwin and others have reported the elevated fitness of hybrid genotypes under certain environmental conditions. In modern scientific terminology, this observation reflects the fact that hybrid genotypes can demonstrate genotype × environment interactions. In the current review, we illustrate the development of one plant species complex, namely the Louisiana Irises, into a 'model system' for investigating hybrid fitness and the role of genetic exchange in adaptive evolution and diversification. In particular, we will argue that a multitude of approaches, involving both experimental and natural environments, and incorporating both manipulative analyses and surveys of natural populations, are necessary to adequately test for the evolutionary significance of introgressive hybridization. An appreciation of the variability of hybrid fitness leads to the conclusion that certain genetic signatures reflect adaptive evolution. Furthermore, tests of the frequency of allopatric versus sympatric/parapatric divergence (that is, divergence with ongoing gene flow) support hybrid genotypes as a mechanism of evolutionary diversification in numerous species complexes.

Prevalence and specificity of immunoglobulin G and immunoglobulin A non-complement-binding anti-HLA alloantibodies in retransplant candidates.

The role of complement-binding donor-directed anti-human leukocyte antigen (HLA) antibodies in graft rejection is well established, whereas the prevalence and relevance of non-complement-binding (NCB) anti-HLA antibodies are less well defined. The aim of our study was to establish a sensitive and reliable test system for the detection and the specification of these NCB anti-HLA antibodies. Sera from 60 patients awaiting retransplantation were analysed for the presence of anti-HLA class I alloantibodies with complement-dependent cytotoxicity (CDC) tests. Immunoglobulin (Ig)G(all) anti-HLA class I and class II alloantibodies were differentiated on generic level by plate-based solid phase enzyme-linked immunosorbent assay. Subsequently, a modified bead-based (Luminex) assay was applied, allowing the investigation of IgG(2/4) NCB isotypes as well as IgA(1/2). The anti-HLA specificities of the NCB alloantibodies were determined and compared with known mismatches from previous transplants. Seventeen of the 60 sera (28%) were positive in the CDC increasing to 26 of 60 (43%) in the class I and 33 of 60 (55%) in the class II plate-based assay. Using the modified bead-based system 24 of 60 sera (40%) contained NCB IgG(2/4), which were mostly donor specific. In addition, a high prevalence of NCB IgA antibodies was detected (26 of 60 sera), which occurred independently of IgG(2/4) NCB, and half of which were donor specific. NCB anti-HLA alloantibodies, including the IgA isotype, can reliably be detected using the modified bead-based test system. These NCB alloantibodies had a high prevalence in retransplant candidates and were mostly donor specific.

MTHFR 677TT genotype increases the risk for cervical artery dissections.

The methylene tetrahydrofolate reductase (MTHFR) C677T polymorphism was studied in 174 German patients with cervical artery dissection (CAD). The results were compared with published data on 927 healthy German controls. In the series of patients, the frequency of T alleles and of TT carriers was slightly higher (13.8%) than among the healthy controls (10.6%). In patients with multiple dissections (n = 50), the proportion of TT carriers (18%) was found to be even higher and correlated with the number of events. The MTHFR C677T polymorphism was suggested to modify the risk for CAD.

Anti-HLA class II antibodies in kidney retransplant patients.

The relevance of anti-HLA class II antibodies for kidney graft survival is still controversial. In part, this can be attributed to difficulties to detect and differentiate anti-HLA class II antibodies. Anti-HLA class II IgG antibody screening was performed by enzyme-linked immunosorbent assay. Subsequently, all anti-HLA class II-positive sera were subjected to the determination and specification using color-coded microspheres coated with purified HLA antigens. In a cohort of 934 patients awaiting kidney transplantation, 41 sera (4.4%) were positive for IgG anti-HLA class II antibodies. The presence was confirmed in 90.2% sera by retesting. Subsequently, all anti-HLA class II-positive patients (n = 27) who in the past had undergone a kidney transplantation with an HLA-DR and/or -DQ-mismatched graft were selected. In 25 of 27 sera (92.6%), the alloantibody specificities corresponded to the known previous transplant mismatches on a broad antigen level. In 20 of 27 sera (74.1%), anticlass I antibodies were detected as well. Anti-HLA-DP antibodies were seen in 24 of the 27 sera of this cohort. In the majority of the cases, the reactivities with different DPB1 alleles could be explained by involvement of a single, specific DPB1 epitope. Donor-specific anti-HLA-DR and -DQ antibodies were seen in the majority of cases with graft failure following HLA class II alloantigen exposure in prior kidney transplantations. In addition, HLA-DP may serve as a transplantation antigen in kidney transplantation, leading to a humoral response.

Distinct tumour necrosis factor alpha, interferon gamma, interleukin 10, and cytotoxic T cell antigen 4 gene polymorphisms in disease occurrence and end stage renal disease in Wegener's granulomatosis.

BACKGROUND: Cytokines and T cell regulatory proteins play an important role in the pathogenesis of Wegener's granulomatosis (WG). OBJECTIVE: To investigate cytokine and cytotoxic T cell antigen-4 (CTLA4) gene polymorphisms and HLA class II alleles in generalised WG. METHODS: The distribution of cytokine and cytotoxic T cell antigen 4 (CTLA4) gene polymorphisms and HLA class II alleles was analysed in 32 patients with generalised WG and 91 healthy controls. Genotyping was carried out for HLA-DRB1 and HLA-DQB1 and for polymorphism of the genes encoding TNF alpha (-238, -308, -376), TGF beta (codon 10 and 25), IFN gamma (+874), IL6 (-174), IL10 (-592, -819, -1082), CTLA4 (-318, +49), and the (AT)(n) repeats of the CTLA4 gene. In addition, stratification analysis was carried out according to the presence (n = 15) or absence (n = 17) of end stage renal disease. RESULTS: An increase in the IFN gamma +874 T/T (odds ratio (OR) = 3.14) and TNF alpha -238 G/A (OR = 5.01) genotypes was found in WG patients. When ESRD positive and negative patients were compared, the IFN gamma +874 A/A and the CTLA4 -318 C/C genotypes were found more often in the ESRD subgroup (OR = 10.6 and OR = 2.25). WG patients without ESRD had a higher frequency of the IL10 GCC/ACC promotor genotype (OR = 0.13) and long CTLA4 (AT)(n) repeats (OR = 0.4). No effect was seen for HLA-DR and -DQ markers. CONCLUSIONS: Disease susceptibility and clinical course in WG may be associated with distinct polymorphisms of cytokine and CTLA4 genes.

An accelerator based system for in vivo neutron activation analysis measurements of manganese in human hand bones.

Manganese (Mn) is an essential nutrient for growth and development. Unfortunately, overexposure can lead to neurological damage, which is manifested as a movement disorder marked by tremors. Preclinical symptoms have been found in populations occupationally exposed to the element, and it is suggested that in late stages of the disorder, removing the Mn exposure will not prevent symptoms from progressing. Hence, it is desirable to have a means of monitoring Mn body burden. In vivo neutron activation analysis (IVNAA) is a technique which allows the concentration of some elements to be determined within sites of the body without invasive procedures. Data in the literature suggests that the Mn concentration in bone is greater than other tissues, and that it may be a long term storage site following exposure. Therefore, using the McMaster KN-accelerator to produce neutrons through the 7Li(p,n)7Be reaction, the feasibility of IVNAA for measuring Mn levels in the human hand bone was investigated. Mn is activated through the 55Mn(n,gamma)56Mn reaction, and the 847 keV gamma rays emitted when 56Mn decays are measured outside the body using NaI(Tl) detectors. An optimal incident proton energy of 2.00 MeV was determined from indium foil and microdosimetry measurements. Hand phantom data suggest a minimum detectable limit of approximately 1.8 ppm could be achieved with a reasonably low dose of 50 mSv to the hand (normal manganese levels in the human hand are approximately 1 ppm). It is recommended the technique be developed further to make human in vivo measurements.

Genetics and the fitness of hybrids.

Over the years, the evolutionary importance of natural hybridization has been a contentious issue. At one extreme is the relatively common view of hybridization as an evolutionarily unimportant process. A less common perspective, but one that has gained support over the past decade, is that of hybridization as a relatively widespread and potentially creative evolutionary process. Indeed, studies documenting the production of hybrid genotypes exhibiting a wide range of fitnesses have become increasingly common. In this review, we examine the genetic basis of such variation in hybrid fitness. In particular, we assess the genetic architecture of hybrid inferiority (both sterility and inviability). We then extend our discussion to the genetic basis of increased fitness in certain hybrid genotypes. The available evidence argues that hybrid inferiority is the result of widespread negative epistasis in a hybrid genetic background. In contrast, increased hybrid fitness can be most readily explained through the segregation of additive genetic factors, with epistasis playing a more limited role.

Differential inhibitory effects of intravenous immunoglobulin preparations on HLA-alloantibodies in vitro.

BACKGROUND: Treatment of allosensitized patients with intravenously administered pooled immunoglobulin preparations (IVIG) may lead to a long-lasting reduction of anti-HLA alloantibody titers. An inhibitory response of IVIG preparations on lymphocytotoxicity is suggested to depend on IgG and to predict a successful reduction of anti-HLA alloantibodies upon the administration of high-dose IVIG in vivo. METHODS: In this study, we evaluated different IVIG preparations for their in vitro inhibitory capacity on lymphocytotoxicity and binding of anti-HLA alloantibodies to purified HLA antigens. For that purpose sera from 24 highly sensitized patients awaiting kidney transplantation and serological HLA testing reagents were used. Panel-reactive antibody (PRA) determinations using standard complement-dependent cytotoxicity testing and anti-HLA alloantibody determination by ELISA were carried out in the presence and absence of 50% (v/v) IVIG. RESULTS: The addition of IgG-containing IVIG preparations gave only a moderate inhibitory response judging from the average decrease of PRA levels (absolute DeltaPRA range: -2% to 16%), whereas the largest inhibition of lymphocytotoxicity was seen after the addition of IgM/IgA-containing IVIG preparations (absolute DeltaPRA range: 19% to 44%). For both IgG and IgM/IgA-containing IVIG preparations, the reduction of lymphocytotoxicity occurred in a dose-dependent fashion without a preference for particular anti-HLA class I antibody specificities. Significantly lower inhibitory effects on anti-HLA antibody reactivity were observed when the effects of IVIG preparations were monitored by ELISA (absolute DeltaPRA range: 7% to 22%). CONCLUSIONS: Our data suggest that the immunomodulatory capacity is largely caused by the IgM/IgA fraction of IVIG when analyzed by lymphocytotoxicity. The different effect on ELISA versus complement-dependent cytotoxicity testing suggests that interactions of IVIG with complement rather than anti-idiotypic antibodies may contribute to the inhibitory effects of IVIG in vitro.

Environment-dependent performance and fitness of Iris brevicaulis, I. fulva (Iridaceae), and hybrids.

We tested the relative fitness of two Louisiana Iris species (Iris brevicaulis and I. fulva) and their first-generation backcross hybrids in three experimental watering treatments: dry, field capacity, and flooded. Leaf area expansion rate, gas exchange (A(max), g(s), c(i)), and biomass at final harvest were measured for each species and hybrid class in all three environmental treatments. Fitness (based on total biomass) of the four genotypic classes differed significantly with environment. All genotypic classes performed most poorly in the dry treatment. The fitness ranking of genotypic class also changed across environments (significant genotypic class by treatment interaction) with hybrid genotype fitness shifting relative to parental genotypes. Integrating over all treatments, backcrosses to I. fulva showed the lowest fitness, whereas backcrosses to I. brevicaulis outperformed I. fulva. The differences in fitness were apparently achieved by a combination of differences in photosynthesis and allocation. In this system, hybrids are not necessarily less fit than their parents, and the relationship between hybrid and parental fitness is influenced by environmental conditions, lending support to the Hybrid Novelty model of hybrid zone evolution.

Age-specific fitness components in hybrid females of Drosophila pseudoobscura and D. persimilis.

Most models of hybridization assume that hybrids are less fit than their parental taxa. In contrast, some researchers have explored the possibility that hybrid individuals may actually have higher fitness and so play an important role in the generation of new species or adaptations. By estimating age-specific fitness components, we can determine not only how hybrid fitness differs from parental taxa, but also whether the fitness of hybrids relative to parental taxa changes with age. Here we describe an analysis of age-specific fitness traits in two species of Drosophila, D. pseudoobscura and D. persimilis, and their F1 hybrids. At early ages, hybrid females lay as many eggs as parental individuals, on average, but produce far fewer offspring. By late ages, in contrast, parental taxa show a steep decline in production not seen in hybrids, such that hybrids produce more offspring, on average, than parental taxa. Furthermore, egg-adult survival in hybrids is negatively correlated with egg density, whereas these traits are only weakly correlated in parental taxa. The results are limited somewhat by the fact that we analyze only two strains, and that these may be partially inbred. Nonetheless, the results are certainly illustrative, pointing out not only that at least some hybrid individuals may be as fit or fitter than parental taxa, but also that the difference between hybrids and parental taxa varies with age.

Adult generativity and the socialization of adolescents: relations to mothers' and fathers' parenting beliefs, styles, and practices.

Mothers, fathers, and their adolescent children participated in two studies investigating the relations between Erikson's concept of generativityin adulthood and patterns of parenting. Study 1 involved 77 mothers and 48 fathers of 1st-year university students; Study 2 was part of an investigation of socialization processes in 35 families with an adolescent, aged 14-18. Parental generative concern was assessed by the Loyola Generativity Scale (LGS) of McAdams and de St. Aubin (1992) in each study. In both studies, mothers demonstrated positive relations between scores on the LGS and an authoritative style of parenting, as well as between generativity and more positive, optimistic views of adolescent development. In Study 2, these more positive views in turn mediated some aspects of autonomy-fostering practices used with the adolescent. Variations in fathers' levels of generative concern were less consistently related to these indices of parenting, however.

Intimately linked or hardly speaking? The relationship between genotype and environmental gradients in a Louisiana Iris hybrid population.

Several models of hybrid zone evolution predict the same spatial patterns of genotypic distribution whether or not structuring is due to environment-dependent or -independent selection. In this study, we tested for evidence of environment-dependent selection in an Iris fulva x Iris brevicaulis hybrid population by examining the distribution of genotypes in relation to environmental gradients. We selected 201 Louisiana Iris plants from within a known hybrid population (80 m x 80 m) and placed them in four different genotypic classes (I. fulva, I. fulva-like hybrid, I. brevicaulis-like hybrid and I. brevicaulis) based on seven species-specific random amplified polymorphic DNA (RAPD) markers and two chloroplast DNA haplotypes. Environmental variables were then measured. These variables included percentage cover by tree canopy, elevation from the high water mark, soil pH and percentage soil organic matter. Each variable was sampled for all 201 plants. Canonical discriminant analysis (CDA) was used to infer the environmental factors most strongly associated with the different genotypic groups. Slight differences in elevation (-0.5 m to +0.4 m) were important for distinguishing habitat distributions described by CDA, even though there were no statistical differences between mean elevations alone. I. brevicaulis occurred in a broad range of habitats, while I. fulva had a narrower distribution. Of all the possible combinations, I. fulva-like hybrids and I. brevicaulis-like hybrids occurred in the most distinct habitat types relative to one another. Each hybrid class was not significantly different from its closest parent with regard to habitat occupied, but was statistically unique from its more distant parental species. Within the hybrid genotypes, most, but not all, RAPD loci were individually correlated with environmental variables. This study suggests that, at a very fine spatial scale, environment-dependent selection contributed to the genetic structuring of this hybrid zone.

System design for in vivo neutron activation analysis measurements of manganese in the human brain: based on Monte Carlo modeling.

Manganese is an essential nutrient required by the human body, but conversely, over exposure to the element may cause central nervous system damage. The technique of in vivo neutron activation analysis, using the McMaster KN-accelerator, is being investigated as a possible method of noninvasively determining manganese concentrations within the human body. Since the brain is the primary target of damage from exposure it would be the ideal site for measurements. Thus, Monte Carlo simulations have been undertaken to define the optimum experimental parameters for such a measurement, examining the use of possible moderator, reflector and collimator materials.