Gina Mazzola

Familial clustering of IgA nephropathy: Further evidence in an Italian population

Several lines of evidence suggest that genetic factors have an important role in the pathogenesis of immunoglobulin A (IgA) nephropathy. We report the prevalence of familial IgA nephropathy in a referral center in northern Italy and present the data on HLA genotypes in the families identified. Twenty-six of 185 patients (14%) with IgA nephropathy investigated in Brescia, Italy, were related to at least one other patient with the disease. Restriction fragment length polymorphism (RFLP) analysis of HLA-DR beta and HLA-DQ alpha and beta genes, as well as polymerase chain reaction-based oligonucleotide typing, was performed in family members. The 26 patients with IgA nephropathy belonged to 10 families. Familial relationships between the patients varied greatly, ranging from parent-child to sib-pair to more distant familial relationships. No common nephrotoxic factor was identified in the families. The intervals separating the apparent onset of disease in relatives with IgA nephropathy varied from 8 months to 13 years. In patients with a family history of IgA nephropathy, there was an increased incidence of HLA-DRB1*08 compared with those with sporadic IgA nephropathy. The study shows that a significant number of the patients with IgA nephropathy followed up in Brescia had a family history of disease. The fact that the Italian population, an ethnic group not previously examined, also presents an increased familial susceptibility to IgA nephropathy suggests that familial predisposition is a very common finding for IgA nephropathy. Thus, clinicians should become aware that IgA nephropathy may aggregate within families in a substantial number of cases. In addition, this subgroup of patients with IgA nephropathy offers an ideal opportunity to elucidate the molecular genetics of this disease.

Molecular analysis of hyperoxaluria type 1 in Italian patients reveals eight new mutations in the alanine : glyoxylate aminotransferase gene

Abstract Systematic screening using the SSCP technique followed by sequencing of bands with abnormal mobility derived from the AGXT exons of 15 unrelated Italian patients with primary hyperoxaluria type 1 (PH1) allowed us to characterize both the mutant alleles in each individual. Eight new mutations were identified: C155del, C156ins, G244T, C252T, GAG408ins, G468A, G588A and G1098del. This study demonstrates both the effectiveness of the screening strategy chosen to identify all the mutant alleles and the high degree of allelic heterogeneity in PH1.

Role of non-HLA genetic polymorphisms in graft-versus-host disease after haematopoietic stem cell transplantation

Graft‐versus‐host disease (GvHD) is the main complication after haematopoietic stem cells transplantation (HSCT) and acute forms (aGvHD) occur in 20–40% of cases even after donor (D) and recipient (R) HLA matching, apparently because of D/R minor histocompatibility antigen (mHA) mismatches and cytokine polymorphisms. The genotype of cytokines and mHA of 77 haematological R following HSCT from HLA identical siblings were determined to detect genetic polymorphisms correlated with GvHD. We analysed TNFA (−863 C/A, −857 C/T and G/A at positions −574, −376, −308, −244, −238), IL‐10 (−1082 G/A, −819 C/A, −592 C/T), IL‐1B (T/C +3953), IL‐1RA (VNTR), HA‐1 (H/R allele) and CD‐31 (C/G at codon 125, A/G at codon 563). Allele frequencies were in Hardy–Weinberg equilibrium and similar to those of 77 healthy controls. We observed positive correlations between a lower risk of clinically significant aGvHD and both the presence of −1082G −819C −592C IL‐10 haplotype when both R and D are considered together and the absence of R IL‐1RA allele 2. Furthermore, we observed an association between the absence of TNF‐A −238 A allele and the risk of extensive chronic GvHD. mHA and cytokines genotyping would thus seem a valid source of information for the prior identification of recipients with a higher risk of aGvHD.

Angiotensin I-converting enzyme genotype significantly affects progression of IgA glomerulonephritis in an italian population.

To evaluate the role of angiotensin-converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism in the progression of immunoglobulin A glomerulonephritis (IgA-GN), genotype distribution in 81 biopsy-proven cases of IgA-GN was studied. A logistic regression model showed that the risk for homozygous DD was not significantly elevated in patients with IgA-GN compared with healthy subjects (odds ratio = 1.16; confidence interval [CI], 0.4 to 3.3). However, the 5-year (78% v 90%) and 10-year (52% v 82%) renal survival rates for 47 patients with serum creatine (Cr) levels of 1.5 mg/dL or less at biopsy was significantly less in DD patients (n = 18; chi2 = 5.41; P = 0.02). The hazard ratio (HR) for DD (multivariate analysis from Cox proportional model after adjustment for known factors of progression, such as hypertension [HPT] and proteinuria [PTO]) was 3.07 (CI, 1.1 to 9.4). The HR for heavy PTO was 6.1 (CI, 1.9 to 19). The association of DD genotype with progression was even more striking when patients with other risk factors (heavy proteinuria) were excluded, as shown by DD-related risk in the absence (HR = 3.6; CI, 1.1 to 12) and presence (HR = 2; CI, 0.4 to 10) of PTO. The risk ratio was further increased by the coexistence of DD + PTO (HR = 9.16; CI, 1.8 to 15.7). Furthermore, in a cross-sectional study among patients with IgA-GN, a logistic regression model showed that the risk for homozygous DD was greater, although not at a statistically significant level in the end-stage renal failure subgroup compared with the normal renal function subgroup (odds ratio = 3.16; CI, 0.7 to 13.7) after adjustment by sex, age at biopsy, HPT, PTO, and therapy. Last, DD was significantly more frequent in those patients who started hemodialysis at an earlier age (chi2 for trend = 6.81; P = 0.009). Our study further supports that ACE genotype is a risk factor not for the development, but for the worsening of IgA-GN clinical course. However, on the basis of current knowledge, we cannot exclude that I/D polymorphism may simply serve as a prognostic marker, eventually linked with other discrete loci involved in the progression of renal damage.

Are HLA class II and immunoglobulin constant region genes involved in the pathogenesis of mixed cryoglobulinemia type II after hepatitis C virus infection

BACKGROUND/AIMS Hepatitis C virus infection is known to play an important role in the pathogenesis of essential mixed cryoglobulinemia type II. Progression of hepatitis C virus infection to mixed cryoglobulinemia may be influenced by host immune response. To analyze the immunogenetic background of mixed cryoglobulinemia, we studied HLA-DR, DQ loci and the switch regions of immunoglobulin heavy chain gamma1 and gamma4 constant genes. METHODS HLA typing was performed in 84 hepatitis C virus-infected patients (46 with cryoglobulins and 38 without), and 109 healthy controls, through analysis of restriction fragment length polymorphisms, supplemented with other techniques. Immunoglobulin heavy chain gamma1 and gamma4 polymorphisms, detected by restriction fragment length polymorphisms, were studied in 41 patients with mixed cryoglobulinemia and 51 controls. RESULTS The gene frequency of DRB1*11 was significantly higher in patients with mixed cryoglobulinemia than in controls (0.36 and 0.20, respectively; p= 0.0035). However, DRB1*11 was also increased in the subgroup of patients without mixed cryoglobulinemia who did not develop severe liver disease, while it was decreased in those with severe liver damage (0.50 and 0.13; p=0.0035). The frequency of 5.4 kb allele of the immunoglobulin heavy chain gamma1 switch region was higher in patients with mixed cryoglobulinemia than in controls (0.47 and 0.22; pc=0.002), while the frequency of 5.5 kb allele was lower (0.51 and 0.78; pc= 0.001). CONCLUSIONS Susceptibility to develop cryoglobulins after hepatitis C virus infection was not associated with HLA-DR or DQ. HLA-DRB1*11-positive individuals were protected from serious chronic liver disease after hepatitis C virus infection. Immunoglobulin heavy chain constant gamma1 switch region restriction fragment length polymorphisms were associated with mixed cryoglobulinemia.

The HLA-DRβ16 allogenotype constitutes a risk factor for hypertrophic scarring

Nineteen patients that had developed hypertrophic scars subsequent to thermal injury were typed for HLA class II allogenotypes with the restriction fragment length polymorphism technique. A significant association was found with DR beta 16 (pc = 1.45 x 10(-4); relative risk = 12.25). This finding adds evidence to other data suggesting that immunologic phenomena are involved in pathologic scarring. Moreover, the results presented here have allowed an identification of a genetically determined risk factor for hypertrophic scar formation located in the HLA region.

T cell neoepitope discovery in colorectal cancer by high throughput profiling of somatic mutations in expressed genes

Objective Patient-specific (unique) tumour antigens, encoded by somatically mutated cancer genes, generate neoepitopes that are implicated in the induction of tumour-controlling T cell responses. Recent advancements in massive DNA sequencing combined with robust T cell epitope predictions have allowed their systematic identification in several malignancies. Design We undertook the identification of unique neoepitopes in colorectal cancers (CRCs) by using high-throughput sequencing of cDNAs expressed by standard cancer cell cultures, and by related cancer stem/initiating cells (CSCs) cultures, coupled with a reverse immunology approach not requiring human leukocyte antigen (HLA) allele-specific epitope predictions. Results Several unique mutated antigens of CRC, shared by standard cancer and related CSC cultures, were identified by this strategy. CD8+ and CD4+ T cells, either autologous to the patient or derived from HLA-matched healthy donors, were readily expanded in vitro by peptides spanning different cancer mutations and specifically recognised differentiated cancer cells and CSC cultures, expressing the mutations. Neoepitope-specific CD8+ T cell frequency was also increased in a patient, compared with healthy donors, supporting the occurrence of clonal expansion in vivo. Conclusions These results provide a proof-of-concept approach for the identification of unique neoepitopes that are immunogenic in patients with CRC and can also target T cells against the most aggressive CSC component.

Prognostic values of soluble CD30 and CD30 gene polymorphisms in heart transplantation.

Pretransplant soluble CD30 (sCD30) is a predictor of kidney graft outcome. Its status as a predictor of heart transplant (HT) outcome has not been established. We have studied this question by assessing sCD30 levels and the number of (CCAT)n repeats of the microsatellite in the CD30 promoter region, which is able alone to repress gene transcription, in the sera of 83 HT patients and 77 of their donors. sCD30 was non-significantly increased in the patients, whereas there were no differences in the CD30 microsatellite allele frequencies. A negative correlation between the number of (CCAT)n and sCD30 levels was evident in the donors. Patients with pretransplant sCD30≤30 U/ml displayed a significantly better survival. In conclusion, sCD30 levels are predictive of HT outcome.

HLA supratypes in an Italian population

A supratype analysis of a North Italian population was performed, using 16 polymorphisms in the HLA region spanning the HLA-A-DP segment. Fourteen supratypes were identified, mostly corresponding to those found in other Caucasiod populations. The degree of their conservation both within the B-DR/DQ region and in the regions telomeric and centromeric from HLA-A and DP was evaluated and linkage disequilibria among several DR and DP alleles were identified. Notably, the degree of association with DP increased when the DR marker was part of a conserved B-DR/DQ supratype. These data are relevant to the definition of the genetic structure of the population and to the prediction of probabilities of histocompatibility matching between unrelated individuals.

Human Leukocyte Antigen Class II Polymorphisms and Genetic Susceptibility of IDDM in Egyptian Children

OBJECTIVE To analyze the association between human leukocyte antigen (HLA) and insulin-dependent diabetes mellitus (IDDM) in the Egyptian population for the first time and, thus, to determine the frequency of risk-associated alleles identified by a genomic HLA class II typing. Egyptians are genetically classified as North Africans and considered to be between Caucasoids and Africans (closer to Caucasoids). RESEARCH DESIGN AND METHODS HLA class II typing was performed for 50 IDDM patients and 50 healthy control subjects by a restriction fragment-length polymorphism (RFLP) technique. The analysis of position 57 of the DQB1 molecules was conducted by polymerase chain reaction and specific sequence oligonucleotide hybridization. RESULTS The frequency of DRB1*0301-DRB3*0201-DQA1*0501-DQB1*0201 haplotype was 43.9% in the IDDM patients and 7.1% in the control subjects (P < 0.00001), reflecting the increased prevalence of DQA1*0501 susceptibility allele coding for arginine (Arg) in position 52 and DQB1*0201 susceptibility allele non-coding aspartic acid (Asp) at position 57. Alleles DQB1*0601 and 0603, both carrying Asp at position 57 of the β-chain, and DQA1*0103, encoding a non-Arg 52 alpha-chain, were significantly decreased among the IDDM patients. The presence of four susceptibility residues (two DQA1 Arg 52+ and two DQB1 Asp 57−) conferred the highest relative risk at 20.2. On the other hand, homozygous genotypes for DQA1 non-Arg 52 and DQB1 Asp 57 were found only in the control group. CONCLUSIONS IDDM susceptibility and resistance in the Egyptian population is strongly associated with the expressed DQ α- and β-heterodimers in a dose-effective manner, as already defined in many different ethnic groups.