Reproductive immunology in viviparous mammals: evolutionary paradox of interactions among immune mechanisms and autologous or allogeneic gametes and semiallogeneic foetuses.

Literally, reproductive immunology was born in bovine on-farm reproduction where seminal experiments intended for developing methods for embryo transfer in cattle were performed. Actually, these experiments led to two of major concepts and fundamental principles of reproductive immunology using the bovine species as a model for biomedical research, namely the concept of acquired immunological tolerance and the paradox of the semiallogeneic bovine foetus whereby such organism can develop within an immunologically competent host. Peter Medawar, a scientist who together with Frank Macfarlande Burnet shared the 1960 Nobel Prize in physiology or medicine for discovery of acquired immunological tolerance, while studying dizygotic cattle twins, thereby giving birth to reproductive immunology. Also, these findings significantly influenced development of organ transplants and showed that using farm animals as models for studying transplantation immunology had general relevance for mammalian biology and health including those of humans. However, the interest for further research of the fascinating maternal immune influences on pregnancy and perinatal outcomes and of the prevention and treatment of immunologically mediated reproductive disorders in viviparous mammals of veterinary relevance by veterinary immunologists and reproductive clinicians have been very scarce regarding the application of nonspecific immunomodulatory agents for prevention and treatment of subfertility and infertility in pigs and cattle, but still broadening knowledge in this area and hold great potential for improving such therapy in the future. The aim of the current overview is to provide up-to-date information and explaining/translating relevant immunology phenomena into veterinary practice for specialists and scientists/clinicians in reproduction of animals.

A new extended haplotype Cw*0602-B57-DRB1*0701-DQA1*0201-DQB1*0201 associated with psoriasis in the Croatian population.

In this study, we have analysed the distribution of HLA class II alleles and the extended haplotype HLA-Cw-B-DRB1-DQA1-DQB1 in Croatian patients with type I and type II psoriasis by hybridization with specific oligonucleotide probes. Type I psoriasis showed a significant association with the DRB1*0701 [P < 0.00001; relative risk (RR) = 5.83], DQA1*0201 (P < 0.00001; RR = 6.12), DQB1*0201 (P = 0.0006; RR = 3.29) and DQB1*0303 alleles (P = 0.0008; RR = 7.51). A negative correlation with type I disease was observed for the DQA1*0102 allele (P = 0.002; RR = 0.26). Type II psoriasis did not show any association with any class II alleles. The extended haplotype HLA-Cw*0602-B57-DRB1*0701-DQA1*0201-DQB1*0201 was present at a significantly higher frequency in type I patients (P < 0.00001; RR = 7.72). However, this haplotype was not detected at all in patients with type II psoriasis. In conclusion, the extended haplotype HLA-Cw*0602-B57-DRB1*0701-DQA1*0201-DQB1*0201 is a risk haplotype for type I disease in the Croatian population. This particular haplotype has not been reported previously in association with psoriasis in any other ethnic groups.

HLA class II haplotypic association and DQCAR microsatellite polymorphisms in Croatian patients with psoriasis.

The purpose of the present study was to investigate polymorphism of HLA class II haplotypic associations (HLA-DRB1, -DQA1, -DQB1) and DQCAR alleles in 78 Croatian patients with psoriasis. Patients were divided into two groups according to a family history of disease and age of onset: type I (positive family history and early onset) and type II (negative family history and late onset). The difference in frequency of HLA class II haplotypic associations between type I patients and controls was observed for the following combinations: HLA-DRB1*0701, -DQA1*0201, -DQB1*02 (23.6% vs. 7.2%; p < 0.001), HLA-DRB1*0701, -DQA1*0201, -DQB1*0303 (8.5% vs. 1.3%; p = 0.0018) and HLA-DRB1*1601, -DQA1*0102, -DQB1*0502 (2.8% vs. 9.3%; p = 0.06). The difference between type II psoriasis and controls for association: HLA-DRB1*1501, -DQA1*0102, -DQB1*0602 is not significant (20.0% vs. 8.9%; p = 0.06). The significantly higher frequency of DQCAR 113bp and 119bp alleles in patients with type Ipsoriasis is a result of linkage disequlibrium of these alleles with both HLA-DRB1*0701 haplotypic associations. Analysis ofDQCAR alleles in the HLA-DRB1*0701 haplotypic associations in patients with psoriasis vulgaris and matched controls did not reveal any difference in polymorphism of DQCAR alleles. These data suggest that HLA-DRB*0701 haplotypic combinations are associated with type I but not for type II psoriasis in the Croatian population. DQCAR polymorphism is not useful genetic marker to distinguish susceptible HLA class II haplotypic association.

Depression and torture.

Torture represents an exceptionally traumatic experience in which horror, helplessness, and hopelessness are extreme. Therefore, it can be expected that depression, along with other trauma-related disorders is present in torture victims at higher rates than in other psychotraumatized individuals. To demonstrate this, we examined two groups of refugees, all suffering the post-traumatic stress disorder. The first group (N = 50) had combat experience but were imprisoned and tortured as well. Members of the second group (N = 29) had combat experience. A third group (N = 30) consisted of local people with no traumatic experience. Using the Hamilton scale, the Beck Depression Inventory and structured dedicated interviews, we tried to determine whether those groups differed in level of depression based on their different levels of traumatic experience. The results of our study indicate that torture victims showed a significantly higher level of depression that is clinically relevant.

[Variation in HLA-B27 gene subtypes and susceptibility of ankylosing spondylitis in the Croatian population]. / Raznolikost podtipova gena HLA-B27 i povezanost s ankilozantnim spondilitisom u hrvatskoj populaciji.

The differences in amino acid residues of HLA-B27 subtypes are minor, but may play role in pathogenesis of ankylosing spondylitis (AS). Aim of this study was to investigate of frequency of B27 subtypes in Croatian AS patients and B27 positive healthy controls. Group of 50 AS patients and 38 B27 positive controls were typed for B27 subtypes by PCR-SSP method. In the group of AS patients we found four subtypes: B*2705 (83.0%), B*2702 (13.2%), while remaining two alleles B*2701 and B*2704 had one individual each. In the group of B27+ controls we also observed B*2705 (76.3%) as most frequent allele while frequency of B*2702 was 21.1%. No significant evidence for association between AS and a particular HLA-B27 subtype in the Croatian population were found.

Distribution of alleles at DQCAR microsatellite locus in the Croatian population.

AIM: To investigate the polymorphism of DQCAR alleles and their association with HLA-DRB1, -DQA1, -DQB1 haplotypic associations in the Croatian population. METHODS: Blood samples were collected from 135 healthy unrelated donors from Zagreb area previously typed for HLA class II alleles (DRB1, DQA1, DQB1). The DQCAR samples were run on a standard denaturing sequencing gel in a DNA sequencer and the sequences were analyzed and compared. RESULTS: Among 10 different DQCAR alleles found in the population of Croatia, the most frequent were DQCAR 103 bp (41.5%), 121 bp (13.7%), 111 bp (11.9%), and 99 bp (10.7%). DQCAR alleles 101 bp, 115 bp, 123 bp, and 125 bp were not observed. Comparison of DQCAR allele frequencies between Croatians and other populations did not reveal any significant difference. The study proved a little diversity in DQ1 haplotypic associations. Among 141 examined DQ1 associations, 120 were DQCAR 103 bp, whereas the remaining 21 were DQCAR 107 bp. The DRB1*07 haplotypic association showed the highest diversity of DQCAR alleles (111 bp, 113 bp, 117 bp, 119 bp, and 121 bp). Three unusual haplotypic combinations were found: HLA-DRB1*0401, -DQA1*0301, -DQB1*0302, -DQCAR119bp; HLA-DRB1*0408, -DQA1*0301, -DQB1*0304, -DQCAR117bp; and HLA-DRB1*0701, -DQA1*0201, -DQB1*02, -DQCAR 105bp. CONCLUSION: Specific DQCAR alleles observed in association with common Caucasoid haplotypes are also found in the Croatian population, but in new and unusual associations. These associations have not been reported in other populations, which suggests that they might be a characteristic of Croatians.

Polymorphism of HLA-A, -B, -DRB1, -DQA1 and -DQB1 haplotypes in a Croatian population.

We describe for the first time extended haplotypes in a Croatian population. The present study gives the HLA-A, -B, -DRB1, -DQA1 and -DQB1 allele and haplotype frequencies in 105 families with at least two offspring. All individuals were studied by conventional serology for HLA class I antigens (A and B), while class II alleles (DRB1, DQA1, DQB1) were typed using the PCR-SSOP method. HLA genotyping was performed by segregation in all 105 families. For extended haplotype analysis, 420 independent parental haplotypes were included. Fourteen HLA-A, 18 HLA-B, 28 DRB1, 9 DQA1 and 11 DQB1 alleles were found in the studied population. Most of the DRB1 alleles in our population had an exclusive association with one specific DQA1-DQB1 combination. This strong linkage disequilibrium within the HLA class II region is often extended to the HLA-B locus. A total of 10 HLA-A, -B, -DRB1, -DQA1, -DQB1 haplotypes were observed with a frequency

HLA class I polymorphism in the Albanian population.

The HLA class I polymorphism was studied in a sample of the Albanian population. Ninety-three unrelated healthy Albanians were typed for HLA-A, -B and -Cw antigens by standard microlyphocytotoxicity test. The antigens with the highest frequencies were: HLA-A2 (34.4%), A3 (14.5%) and A1 (12.4%); B51 (19.3%), B35 (12.4%) and B18 (10.2%); Cw4 (16.2%), Cw7 (16.2%) and Cw6 (10.8%). The HLA haplotypes with high frequency in Albanians included A2-B51 (4.3%), A2-B18 (2.4%), A2-B35 (2.4%), Cw4-B35 (7.6%), and Cw7-B18 (6.5%), which are not significantly different from the other neighboring populations. Low frequency of HLA-A1-B8 haplotype (1.1%) is noted in the Albanian population. The frequency of HLA-B27 antigen (1.1%) is one of the lowest frequencies observed in Caucasians. Such results are important in studies of HLA-A1-B8, HLA-B27 and disease associations. These findings should be also useful in understanding the origin of Albanians, representing a base for future studies about HLA polymorphism in the Albanian population.

Analysis of HLA antigens in Croatian patients with psoriasis.

In common with most autoimmune diseases, psoriasis is associated with some HLA antigens. We studied the distribution of HLA antigens in Croatian patients with psoriasis: 108 patients were divided into groups according to family history and age of disease onset. HLA antigens were analyzed serologically and HLA-C alleles were analyzed using polymerase chain reaction. We found significant increases in HLA-A2, -B17, -B37 and -B13 antigens and highly significant increases in HLA-Cw*0602 and DR7 antigens in psoriatic patients compared with controls. Patients with type I psoriasis (early onset, positive family history) showed highly significant associations with Cw*0602 [p < 0.00001; relative risk (RR) = 14.45] and DR7 (p < 0.00001; RR = 15.09) antigens. Patients with type II psoriasis (late onset, no family history) had a significant association with Cw*03 antigen (p = 0.008; RR = 0.17). In conclusion, HLA-B13, -B17, Cw*0602 and -DR7 antigens are associated with a significant risk of psoriasis in the Croatian population and the Cw*0602 allele has the strongest association, especially for type I psoriasis.

[Gluten enteropathy in Croatian children is primarily associated with the HLA-DR3-DQ2 haplotype]. / Glutenska enteropatija u hrvatske djece primarno je udruzena s haplotipom HLA-DR3-DQ2.

OBJECTIVES: Coeliac disease (CD) has one of the strongest class II HLA association of any human disease. The goal of this study was to establish the frequency of HLA-DR and DQ antigens, as well as common HLA-DR-DQ haplotypes among CD patients. No similar study has previously been done in Croatian patients. PATIENTS AND CONTROLS: There were 58 biopsy proven patients with CD. The control groups comprised 502 and 118 healthy person for HLA-DR and HLA-DQ typing, respectively. METHODS: The class II HLA-DR and DQ antigens typing were carried out by standard microcytotoxicity assay for B cells. Only antisera to three specificities (DQ1, 2 and 3) were available to us at the time of the study. RESULTS: We confirmed the high frequency of HLA-DR3 (84.48%; RR = 23.7; p < 0.00001) in our patients with CD. As reported in other populations, most of the patients negative for DR3, were heterozygous for DR7 and DR5 (10.34%) or DR4/DR5, DR4/DR6 (3.44%). CD was significantly correlated with the presence of HLA-DQ2 (96.55%, RR = 75.9; p < 0.00001). Correlation with CD was strongly dependent on homozygosity, 15 out of 58 (25.86%) of the patients and 4 (2.87%) of the controls being homozygotes for DQ2 (RR = 11.9; p < 0.00001). The remaining two patients were DR4-DQ3 positive. Among extended haplotypes only DR2-DQ1 was under-represented (RR = 0.3; p < 0.0033). CONCLUSION: The results suggest that in Croatia CD is primarily associated with HLA-DQ2 specificities on the DR3-DQ2 haplotype.

[Immunogenetic study of the HLA system in families of patients with ankylosing spondylitis]. / Imunogenetska istrazivanja sustava HLA u obiteljima bolesnika s ankilozantnim spondilitisom.

In this study the immunogenetic relationships among 141 unrelated HLA-B27+ patients with ankylosing spondylitis (AS) and 792 members of their families were studied. Two control groups, with at least one B27+ parent were used (families undergoing transplantation program and triplet families undergoing paternity testing). All subjects were typed for HLA-A and -B antigens by microlyphocytotoxity test (MLCT) on local typing trays. The frequency of HLA-A and -B alleles was equal in the all tested groups. The segregation of all tested genes was regular regarding to the total number of positive and negative siblings, while regarding to the sex of sibs was irregular for HLA-B27 and -B5 gene. The statistical significance (p < 0.05) was found when ratio between B27+ and B27- sons in AS group was compared with the same ration in control families. In AS group was detected statistical significant (p < 0.01) high number of B5+ than B5- daughters and statistical significant (p < 0.05) less number of B5+ sons. HLA-B21 was shown to be decreased among B27+ AS patients. A synergistic effect between additional HLA-B alleles and B27 was not observed. The distribution of B27 haplotypes in AS and control families was similar except for haplotype HLA-A10, B27 which was significant (p < 0.001) less present in AS families.

Polymorphism of DR52-associated haplotypes in a Croatian population.

We investigated DR52 haplotype polymorphism in a population of 78 Croatian families with at least one parent and one offspring positive for a DR52-associated allele, using the PCR-SSOP method. The haplotypes DRB1*0301-DQA1*0501-DQB1*0201, DRB1*11-DQA1*0501-DQB1*0301 and DRB1*1201-DQA1*0501-DQB1*0301 seem to be conserved haplotypes in this Croatian population, while DRB1*13 haplotypes showed high diversity. Among 10 different DRB1*13 haplotypes, four consist of common alleles, while six have an unusual combination of DRB1-DQA1-DQB1 alleles. Three haplotypes (DRB1*1301-DQA1*0103-DQB1*0503, DRB1*1302-DQA1*0102-DQB1*0502 and DRB1*1303-DQA1*0102-*DQB1*0502) have not been reported. These results on DR52-associated haplotype polymorphisms in a Croatian population must be taken into consideration in organ transplantation, especially when searching for unrelated bone marrow donors.

Distribution of alleles at two microsatellite loci (D6S273 and TNFa) in Croatian population.

Polymorphism at the level of two microsatellite loci (D6S273 and TNFa) was studied in Croatian population. The most frequent alleles at D6S273 locus are D6S273 134 bp and 136 bp, while at TNFa locus two most frequent alleles are TNFa 117 bp and 99 bp. This study confirms the irregularity in distribution of microsatellite alleles in different populations with the predominance of two or three alleles on these two investigated microsatellite loci.

High resolution molecular typing of HLA class II region in the population of the island of Krk, Croatia.

The HLA class II alleles (DRB1, DRB3, DRB5, DQA1, and DQB1) and haplotypic associations were studied in the population of the island of Krk using the PCR-SSOP method and the 12th International Histocompatibility Workshop primers and probes. Allele and haplotypic frequencies were compared with the general Croatian population. Significant differences were observed between the population of the island of Krk and Croatians for: a) three broad specificities at DRB1 locus (DRB1*01, *15, and *07), b) one allele at DRB3 locus (DRB3*0301), c) one allele at DQA1 locus (DQA1*0201), d) one allele at DQB1 locus (DQB1*0303). Four unusual haplotypic associations, which have not yet been described in the Croatian population, DRB1*1301-DQA1*0103-DQB1*0607, DRB1*1302-DQA1*0102-DQB1*0605, DRB1*1305-DQA1*0102-DQB1*0605 and DRB1*1305-DQA1*0103-DQB1*0603 were observed in the population from the island of Krk.

HLA class II gene and haplotype diversity in the population of the island of Hvar, Croatia.

The DRB1, DRB3, DRB5, DQA1 and DQB1 allele polymorphisms were analysed in 3 western and 3 eastern villages of the island of Hvar using PCR-SSOP method and 12th International Workshop primers and probes. Three DQB1 alleles (*0304, *0305, *0607) detected in the population of the island of Hvar (HP) have not yet been observed in general Croatian population (GCP). Significant differences were observed between two regions of Hvar for: a) DRB1*0701 allele (p < 0.001), b) DQA1*0201 allele (p < 0.01), and c) DRB1*0101-DQA1*0101-DQB1*0501 haplotypic association (p < 0.05). Two unusual haplotypic associations, which have not yet been described in general Croatian population (GCP), DRB1*0101-DQA1*0102-DQB1*0501 and DRB1*1501-DQA1 *0102-DQB1*0604 were observed in the population from the island of Hvar (HP). Measures of genetic kinship and genetic distances revealed isolation and clusterization which coincides with the known ethnohistorical, as well as biological and biocultural data obtained from a series of previous investigations. The five studied village subpopulations formed two clusters (East-West) to which the far eastern village (with the highest rii of 0.0407) joined later, thus indicating possible impact of historical immigrations from the mainland.

Implication of molecular analysis of HLA-A*02 subtyping for unrelated bone-marrow donor selection.

The HLA-A*02 allele is the most heterogeneous allele at HLA-A locus with 22 different subtypes so far identified. All of these subtype polymorphisms are located in alpha 1 and alpha 2 domains which are responsible for peptide biding and HLA restricted recognition by T-cell receptor. The aim of the present study was to determine the frequency of different HLA-A*02 alleles in 33 healthy unrelated Croatians. HLA-A*02 subtyping has also been retrospectively performed in 2 recipient-unrelated donor pairs and in 4 recipient-HLA phenotypically identical parent pairs. All subjects, previously typed as HLA-A2 by serology were tested using HLA-A*02 ARMS-PCR kit which discriminates 17 different A*02 alleles. Among 17 A*02 alleles we have found 4 different A*02 subtypes in healthy unrelated Croatians. The most frequent A*02 allele was A*0201 (84%). The frequency of the remaining A*02 alleles were as follows: A*0205 (3%), A*0207 (6%) and A*0213 (6%). Among 6 tested bone-marrow transplantation (BMT) pairs, only one has been found to be A*02 subtype incompatible (A*0201/A*0205). Four different A*02 alleles are found in Croatian population with the predominance of A*0201. However these results suggest that A*02 subtyping is also necessary for optimal matching of HLA-A*02 positive donor-recipient pairs in HLA incompatible BMT.