FTO rs9939609 Gene Polymorphism and Obesity: Lack of Association in Kidney Transplantation.

The fat mass and obesity-associated (FTO) gene is one of the most important obesity susceptibility genes. Some FTO gene polymorphisms have been associated with obesity, diabetes, and hypertension, all conditions for which, after transplant, there is increased susceptibility, due to effects of immunosuppressive regimens. To evaluate whether FTO could be a candidate for targeted preventive intervention in the transplant setting, we investigated whether the common genetic variation, FTO rs9939609T>A, could affect weight gain and risk of cardiovascular complications in kidney transplantation. METHODS: In 198 kidney transplant recipients, FTO rs9939609 was investigated in association with body mass index (BMI)/obesity and with other clinical markers of posttransplant risk, then monitored up to 5 years after transplantation. Genotyping was performed using an allelic discrimination method on a real-time polymerase chain (PCR) system. Associations were analyzed using the chi-square test; differences between genotypes were examined with analysis of variance or Kruskal-Wallis test; tests for repeated measures and a general linear model analysis controlling for age and gender were also utilized. RESULTS: Allele and genotype frequencies of FTO rs9939609 in recipients (T/T, 29.8%; T/A, 49.0%; A/A, 21.2%; A, 45.7%; T, 54.3%) reflect those present in healthy Caucasian populations. In the face of pre-/posttransplant differences in total cholesterol, triglycerides, or fasting glucose, results did not show significant changes in these factors among genotypes either before or after transplantation. CONCLUSION: This study highlights a lack of association of FTO rs9939609T>A genotypes and posttransplant weight gain, plasma lipids, and fasting blood glucose in kidney transplantation.

Kidney Transplantation, Polymorphisms of IL-18, and Other Pro-Inflammatory Genes and Late Post-Transplant Outcome.

BACKGROUND: Functional polymorphisms of molecules involved in immune-mediated mechanisms of allograft rejection could be predictive of increased risk for early and late post-transplant complications. In the past years, the challenge for long-term graft survival in kidney recipients is the implementation of personalized approaches. In this study, effects of interleukin (IL)-18-137G/C (rs187238), -607C/A (rs1946518), and other pro-inflammatory cytokine gene polymorphisms (tumor necrosis factor [TNF]-α-308G/A, rs1800629, IL-6-174G/C, rs1800795, and interferon [IFN]-γ+874A/T, rs2430561) on the main post-transplant risk parameters and diseases (metabolic, cardiovascular, infective, and chronic allograft rejection) were assessed in kidney-transplanted patients. METHODS: One hundred seventy-nine transplanted patients were retrospectively analyzed for clinical and biochemical parameters and onset of post-transplant complications. Taqman allelic discrimination and PCR-SSP (polymerase chain reaction-sequence specific primers) techniques were used for genotyping. RESULTS: No predictive effects of allele and genotypes of IL-18-607C/A, TNF-α-308G/A, IL-6-174G/C, and IFN-γ+874A/T gene polymorphisms and onset of risk factors and late complications were evidenced. However, Kaplan-Meier analysis evidenced a weak effect of IL-18-137G/C genotypes on graft survival. CONCLUSIONS: Analyzing associations between some pro-inflammatory cytokine gene polymorphisms and onset of the most relevant risk factors and late complications of kidney transplant, results suggested a possible impact of IL-18-137G/C genotypes on graft survival, which deserves further studies.

Increased CD1D polymorphism: identification of two novel alleles, CD1D*03 and *04, in individuals from Morocco.

Two novel CD1D alleles were identified in unrelated individuals from Morocco. They differ each from the common CD1D*01 allele by one nucleotide substitution in exon 2 resulting in one amino acid change in the G-ALPHA1-LIKE domain. According to the IMGT unique numbering for G domain, CD1D*03 has one nucleotide transition c136 > t in codon 46, with an arginine-to-cysteine amino acid change (R46 > C) in the D-STRAND, whereas CD1D*04 has one transition c98 > t in codon 33, with a threonine-to-methionine amino acid change (T33 > M) in the C-STRAND. This suggests that CD1D is more polymorphic than previously assumed.

The distribution of KIR-HLA functional blocks is different from north to south of Italy.

The killer cell immunoglobulin-like receptor (KIR)-human leukocyte antigen (HLA) interaction represents an example of genetic epistasis, where the concomitant presence of specific genes or alleles encoding receptor-ligand units is necessary for the activity of natural killer (NK) cells. Although KIR and HLA genes segregate independently, they co-evolved under environmental pressures to maintain particular KIR-HLA functional blocks for species survival. We investigated, in 270 Italian healthy individuals, the distribution of KIR and HLA polymorphisms in three climatic areas (from cold north to warm south), to verify their possible geographical stratification. We analyzed the presence of 13 KIR genes and genotyped KIR ligands belonging to HLA class I: HLA-C, HLA-B and HLA-A. We did not observe any genetic stratification for KIR genes and HLA-C ligands in Italy. By contrast, in a north-to-south direction, we found a decreasing trend for the HLA-A3 and HLA-A11 ligands (P = 0.012) and an increasing trend for the HLA-B ligands carrying the Bw4 epitope (P = 0.0003) and the Bw4 Ile80 epitope (P = 0.0005). The HLA-A and HLA-B KIR ligands were in negative linkage disequilibrium (correlation coefficient -0.1211), possibly as a consequence of their similar function in inhibiting NK cells. The distribution of the KIR-HLA functional blocks was different along Italy, as we observed a north-to-south ascending trend for KIR3DL1, when coupled with HLA-B Bw4 ligands (P = 0.0067) and with HLA-B Bw4 Ile80 (P = 0.0027), and a descending trend for KIR3DL2 when coupled with HLA-A3 and HLA-A11 ligands (P = 0.0044). Overall, people from South Italy preferentially use the KIR3DL1-HLA-B Bw4 functional unit, while those from the North Italy equally use both the KIR3DL2-HLA-A3/A11 and the KIR3DL1-HLA-B Bw4 functional units to fight infections. Thus, only KIR3DL receptors, which exert the unique role of microbial sensors through the specific D0 domain, and their cognate HLA-A and HLA-B ligands are selectively pressured in Italy according to geographical north-to-south distribution.

Correlation between genetic HLA class I and II polymorphisms and anthropological aspects in the Chaouya population from Morocco (Arabic speaking).

The aim of this study was to provide genetic and anthropological information on the Chaouya (CH), an Arabic-speaking population living in West Morocco, Atlantic coast (Settat). In 98 unrelated healthy CH volunteers, we first investigated the human leukocyte antigen (HLA) class I and II allele polymorphisms using a sequence-based typing method and examined haplotypes and relatedness of this group to other African and Mediterranean populations. The study showed the close relatedness with Tunisian population and other North Africans, together with a strong influence of various immigrations, mainly Spaniards, French, and Portuguese, as expected. Nevertheless, analysis of class II allele frequencies (afs) showed that Oromo and Amhara Ethiopian groups cluster together with the Berbers and other North Africans, confirming the relationship between these populations (Afro-Asiatic linguistic group, Hamites). South and sub-Saharan Africans cluster separately at a great distance from CH, except the sub-Saharan Bantu population from Congo Kinshasa, which shows a relatively close genetic relationship ascribable to the effect of a diversifying selection. On the other hand, considering HLA class I afs analyses, it was noteworthy that CH grouped together with sub-Saharans, showing a close genetic distance mainly with Ugandas and Kenians Luo.

Analysis of the HLA population data (AHPD) submitted to the 15th International Histocompatibility/Immunogenetics Workshop by using the Gene[rate] computer tools accommodating ambiguous data (AHPD project report).

During the 15th International Histocompatibility and Immunogenetics Workshop (IHIWS), 14 human leukocyte antigen (HLA) laboratories participated in the Analysis of HLA Population Data (AHPD) project where 18 new population samples were analyzed statistically and compared with data available from previous workshops. To that aim, an original methodology was developed and used (i) to estimate frequencies by taking into account ambiguous genotypic data, (ii) to test for Hardy-Weinberg equilibrium (HWE) by using a nested likelihood ratio test involving a parameter accounting for HWE deviations, (iii) to test for selective neutrality by using a resampling algorithm, and (iv) to provide explicit graphical representations including allele frequencies and basic statistics for each series of data. A total of 66 data series (1-7 loci per population) were analyzed with this standard approach. Frequency estimates were compliant with HWE in all but one population of mixed stem cell donors. Neutrality testing confirmed the observation of heterozygote excess at all HLA loci, although a significant deviation was established in only a few cases. Population comparisons showed that HLA genetic patterns were mostly shaped by geographic and/or linguistic differentiations in Africa and Europe, but not in America where both genetic drift in isolated populations and gene flow in admixed populations led to a more complex genetic structure. Overall, a fruitful collaboration between HLA typing laboratories and population geneticists allowed finding useful solutions to the problem of estimating gene frequencies and testing basic population diversity statistics on highly complex HLA data (high numbers of alleles and ambiguities), with promising applications in either anthropological, epidemiological, or transplantation studies.

HLA-G 14-bp insertion/deletion polymorphism in kidney transplant patients with metabolic complications.

HLA-G, a nonclassical HLA molecule with limited polymorphism has immunomodulating/tolerogenic properties. The most common polymorphism of HLA-G is a deletion/insertion of 14 bp, located at the 3'UTR region of the gene (exon 8). This polymorphism is associated with modifications of mRNA stability that can lead to variations of membrane versus soluble HLA-G expression. HLA-G may be involved in the clinical outcomes of transplantation, as evidenced by studies in hematopoietic cell transplantation. We evaluated the possible prognostic importance of 14-bp polymorphisms of HLA-G among kidney transplantation patients. Using polymerase chain reaction amplification we genotyped 124 patients (mean organ survival: 878.95 +/- 595.12 days; range = 1-2565) and 98 control individuals representative of the Italian population. Products were visualized by electrophoresis on agarose gels. The results showed no differences between patients and controls. Twenty-nine patients with acute or chronic rejection or in whom clinical conditions required the use of steroid bolus treatments also showed no association with HLA-G 14-bp genotypes or alleles. The subset of patients with dyslipidemia during follow-up showed a significant decrease among the HLA-G-14/-14 genotype, compared with heterozygous (+14/-14) and nondeleted homozygous (+14/+14) genotype patients (P(c) = .03). These preliminary data showed that HLA-G 14-bp genotypes, although not predictive of rejection, may be useful to identify individuals at risk for the development of posttransplant complications.

Total IL-12 levels are increased in paediatric atopic dermatitis: correlations with age and disease severity.

Alterations in the immune response play an important role in the pathogenesis of atopic dermatitis (AD). We evaluated the role of Th1/Th2 cytokine imbalance in paediatric AD and its progression towards other allergic manifestations. The levels of total interleukin (IL)-12 (p70+p40), IL-12 p70 and soluble CD30 (sCD30) were measured in paediatric patients affected by AD and in age-matched, non-atopic subjects (controls). The serum levels of total IL-12 and sCD30 were higher in patients than in controls (p=0.003 and p=0.053, respectively). Total IL-12 and sCD30 were also particularly increased in patients with severe disease. Serum levels of total IL-12 were negatively correlated with patient age (p=0.001): they were significantly higher in patients younger than 30 months, as compared to age-matched controls and to older patients and controls. Total IL-12 and sCD30 production were not significantly correlated with disease outcomes (atopic march) or with a family history of atopy. Our data show that total IL-12 levels are strongly associated with Th2 activation and severe disease in children with AD. The increased production of total IL-12 at an early age might indicate a different immune modulation and suggests the possibility of new therapeutic approaches for allergic diseases, in particular AD, early in childhood.

A new HLA-A allele identified in a leukemic patient attending hematopoietic cell transplantation: A*2318.

Sequence-based typing procedure (SBT) procedure permitted us to identify a new human leukocyte antigen-A allele in a patient attending hematopoietic stem cell transplantation.

Identification of a novel HLA-B allele, HLA-B*3580, with possible implication in transplantation and CTL response.

A new human leukocyte antigen (HLA)-B allele, named B*3580, with an amino acid substitution at residue 156, has been identified during the sequence-based typing of a patient waiting for a hematopoietic cell transplantation.

Reduced number and function of peripheral dendritic cells in coeliac disease.

Dendritic cells (DC) play a pivotal role in shaping the immune response in both physiological and pathological conditions. In peripheral blood at least two subsets, the myeloid and plasmacytoid, have been described as having different T stimulatory functions and a variable degree of maturation. Certainly, antigen presentation plays a crucial role in the pathogenesis of coeliac disease and circulating immune cells are thought to reflect the state of immune response within the gut. Therefore, we aimed to investigate the quantitative and phenotypical modifications of peripheral blood DC, together with their functional properties, in this pathological condition. Blood samples from 11 untreated patients before and after a course of gluten-free diet, 27 treated patients and 14 controls underwent flow-cytometric analysis, while immunomagnetically sorted DC from the CD patients and eight human leucocyte antigen (HLA)-DQ2/8(+) bone marrow donors were used to evaluate maturation status through the CD83 expression, cytokine profile for interleukin (IL)-6, IL-10, IL-12 and interferon (IFN)-alpha by enzyme-linked immunosorbent assay (ELISA), and functional properties by mixed leucocyte reaction before and after pulsing with digested gliadin. We found that in both untreated and treated patients, a significant reduction of the entire DC population, mainly the plasmacytoid subset, in comparison to healthy controls was observed. In active disease, an impaired allogenic lymphocyte reaction and a significant reduction of IFN-alpha production, paralleled by the presence of a more immature status, were also demonstrated. All the latter modifications have been reverted by pulsing DC with digested gliadin.

Distribution of killer cell immunoglobulin-like receptors genes in the Italian Caucasian population.

BACKGROUND: Killer cell immunoglobulin-like receptors (KIRs) are a family of inhibitory and activatory receptors that are expressed by most natural killer (NK) cells. The KIR gene family is polymorphic: genomic diversity is achieved through differences in gene content and allelic polymorphism. The number of KIR loci has been reported to vary among individuals, resulting in different KIR haplotypes. In this study we report the genotypic structure of KIRs in 217 unrelated healthy Italian individuals from 22 immunogenetics laboratories, located in the northern, central and southern regions of Italy. METHODS: Two hundred and seventeen DNA samples were studied by a low resolution PCR-SSP kit designed to identify all KIR genes. RESULTS: All 17 KIR genes were observed in the population with different frequencies than other Caucasian and non-Caucasian populations; framework genes KIR3DL3, KIR3DP1, KIR2DL4 and KIR3DL2 were present in all individuals. Sixty-five different profiles were found in this Italian population study. Haplotype A remains the most prevalent and genotype 1, with a frequency of 28.5%, is the most commonly observed in the Italian population. CONCLUSION: The Italian Caucasian population shows polymorphism of the KIR gene family like other Caucasian and non-Caucasian populations. Although 64 genotypes have been observed, genotype 1 remains the most frequent as already observed in other populations. Such knowledge of the KIR gene distribution in populations is very useful in the study of associations with diseases and in selection of donors for haploidentical bone marrow transplantation.

Cytokine levels in amniotic fluid: a marker of preterm labor?

The aim of this study was to determine levels of Interleukin 6 (IL-6) in amniotic fluid at the beginning of the second trimester and to establish whether IL-6 can be used as a marker for premature birth as it would appear to be an important prenatal marker of chorionic inflammation. Thirty-three patients, between 16 and 19 weeks of gestation, who were undergoing amniocentesis to establish the presence or not of fetal genetic pathologies were enrolled into the study. Amniotic fluid (3 ml) was taken from each patient and used to perform enzyme-linked immunosorbent assays (ELISAs). The results were analyzed using the Mann-Whitney test and Pearson and Spearman coefficient. The patients were divided into three groups on the basis of the levels of IL-6 found: a) up to 450 pg/ml; b) between 450 and 900 pg/ml; c) over 900 pg/ml; These data were then evaluated alongside the date of parturition and the presence of any maternal or fetal pathologies. The results of our analyses, however, were inconclusive: levels of IL-6 were normal in patients presenting pathologies while obstetric pathologies were absent in patients with high levels of IL-6. In conclusion, this data would indicate that a different method or approach is required for the identification of a marker for premature birth.

Identification of a novel HLA-A*02 allele, A*027401*.

A novel HLA-A*02 allele was detected in a Caucasian patient from central Italy, requiring a hematopoietic cell transplantation. Direct sequencing identified a variation in one nucleotide position, which was confirmed by cloning. The name A*027401 was officially assigned by the WHO Nomenclature Committee in November 2004. A*027401 differs from A*02010101 by a single G to A substitution at nucleotide position 595 in exon 3. The new variant would lead to a nonsynonymous nucleotide change (GGG to AGG) at codon 175, resulting in a basic Arg in the alpha-helix of the alpha2-domain, in place of a non-polar Gly. The presence of an uncommon variation at a highly conserved nucleotide position could have implications in unrelated hematopoietic cell transplantation.

Characterization of a novel HLA-Cw*02 variant, Cw*0208, in a Caucasian individual.

We describe an additional HLA-Cw*02 variant, HLA-Cw*0208, which has been identified in a renal transplant recipient of Caucasian origin (Italy). After performing preliminary serological typing, we analyzed exons 2 and 3 of the HLA-C locus polymorphism by cloning the amplified DNA and using a sequence-based typing method. The new allele differs from Cw*020202 by one nucleotide substitution at nucleotide 61 (G-->A) of exon 2, which translates to a difference of one amino acid at residue 21 (His-->Arg) of the HLA-C heavy chain. We propose that Cw*0208 was generated by a random point mutation in codon 21 from the Cw*020202 allele, or through gene conversion of Cw*020202 with another allele, probably the Cw*1205 and Cw*1602 alleles.

Human leukocyte antigen-A, -B, and -Cw polymorphism in a Berber population from North Morocco using sequence-based typing.

Class I human leukocyte antigen (HLA) polymorphism was examined in a Berber population from North Morocco, named Metalsa (ME). All data were obtained at high-resolution level, using sequence-based typing. The most frequent alleles were: HLA-A*0201 and A*0101; HLA-B*44 (B*4403 and B*4402); B*0801 and the B*50 allele group (B*5001 and B*5002); HLA-Cw*0602; and Cw*07 group (Cw*070101, Cw*070102, Cw*0702, Cw*0704, and Cw*0706), and Cw*040101. The novel HLA-B*570302 allele was identified. It differs at position 486 and position 855 from B*570301, resulting in synonymous Thr and Val. The analysis also evidenced some alleles common in Africans (A*3402, A*6802, A*7401, B*1503, B*4102, B*4202, B*7801, B*5802, Cw*1701, and Cw*1703) and some uncommon alleles (A*3004, B*2702, B*2703, B*5001,02, B*3503, and Cw*0706). The predominant HLA-A-Cw-B-DRB1-extended haplotypes in ME population were A*0101-Cw*0501-B*4402-DRB1*0402, A*240201-Cw*0701-B*0801-DRB1*030101, A*2301-Cw*040101-B*4403-DRB1*040501, A*0201-Cw*040101-B*4403-DRB1*1302, and A*3002-Cw*0602-B*5002-DRB1*0406. This study demonstrates a strong relatedness of ME to other Moroccan and North African populations, some characteristics of sub-Saharan Africans and evidenced the influence of various immigrations during centuries. Nevertheless, this study highlights some unique genetic traits of the ME population compared to other ethnic groups within Morocco, which could be of great interest for clinical aims, transplantation, and diseases.