Knockout of apolipoprotein A-I decreases parenchymal and vascular ß-amyloid pathology in the Tg2576 mouse model of Alzheimer's disease.

AIMS: Apolipoprotein A-I (apoA-I), the principal apolipoprotein associated with high-density lipoproteins in the periphery, is also found at high concentrations in the cerebrospinal fluid. Previous studies have reported either no impact or vascular-specific effects of apoA-I knockout (KO) on ß-amyloid (Aß) pathology. However, the putative mechanism(s) by which apoA-I may influence Aß deposition is unknown. METHODS: We evaluated the effect of apoA-I deletion on Aß pathology, Aß production and clearance from the brain in the Tg2576 mouse model of Alzheimer's disease (AD). RESULTS: Contrary to previous reports, deletion of the APOA1 gene significantly reduced concentrations of insoluble Aß40 and Aß42 and reduced plaque load in both the parenchyma and blood vessels of apoA-I KO × Tg2576 mice compared to Tg2576 animals. This was not due to decreased Aß production or alterations in Aß species. Levels of soluble clusterin/apoJ were significantly higher in neurons of apoA-I KO mice compared to both wildtype (WT) and apoA-I KO × Tg2576 mice. In addition, clearance of Aß along intramural periarterial drainage pathways was significantly higher in apoA-I KO mice compared to WT animals. CONCLUSION: These data suggest that deletion of apoA-I is associated with increased clearance of Aß and reduced parenchymal and vascular Aß pathology in the Tg2576 model. These results suggest that peripheral dyslipidaemia can modulate the expression of apolipoproteins in the brain and may influence Aß clearance and aggregation in AD.

A missense mutation in the glucagon receptor gene is associated with non-insulin-dependent diabetes mellitus.

Non-insulin-dependent diabetes mellitus (NIDDM) affects about 5% of the world population. The disease presents a polygenic mode of inheritance, but mechanisms and genes involved in late-onset NIDDM are largely unknown. We report the association of a single heterozygous Gly to Ser missense mutation in the glucagon receptor gene with late-onset NIDDM. This mutation was highly associated with NIDDM in a pooled set of French and Sardinian patients (chi 2 = 14.4, P = 0.0001) and showed some evidence for linkage to diabetes in 18 sibships from 9 French pedigrees (chi 2 = 6.63, P < 0.01). Receptor binding studies using cultured cells expressing the Gly40Ser mutation demonstrate that this mutation results in a receptor which binds glucagon with a three-fold lower affinity compared to the wild type receptor.

Role of Ia-like products of the main histocompatibility complex in conditioning skin allograft survival in man.

This report correlates the survival time of 93 intrafamilial skin allografts performed under conditions of main histocompatibility complex (HLA) haploidentity with donor-recipient compatibility for products of the HLA-A, -B, -C, and -DR, as well as C3 proactivator, Glyoxalase I, and P loci located on the human 6th chromosome. Incompatibilities for HLA-A and -B (and to a lesser extent for HLA-C) and(or) for HLA-DR products exerted a strong influence upon the fate of skin allografts. When HLA-A and -B were considered alone, the most compatible group of grafts had a mean survival time of 15.8 d, as compared with 11.3 d for the most incompatible transplants. HLA-DR compatibility alone was associated with a mean survival time of 15.3 d, whereas HLA-DR-incompatible grafts had a mean survival time of 11.5 d. Incompatibilities for C3 proactivator, Glyoxalase I, and P did not have a significant effect upon graft survival. There was no evidence of an association between donor-recipient incompatibility at HLA-A, -B, or -C or at HLA-DR; such incompatibilities occurred independently of each other, in spite of the state of linkage disequilibrium known to exist between HLA-B and -DR. Incompatibilities for HLA-A, -B, and for HLA-DR exerted a potent additive effect upon graft survival. Skin grafts bearing one, two, or three incompatibilities had a mean survival time of 16.2, 13.7, and 10.7 d, respectively (P <0.0005).The results point to the important role played by the Ia-like products of the HLA complex (HLA-DR) in conditioning skin allograft survival in man. This consideration may be of direct relevance to the potential clinical usefulness of in vitro serological techniques for the detection of donor-recipient compatibility for HLA-DR.

Systemic lupus erythematosus in Europe at the change of the millennium: lessons from the "Euro-Lupus Project".

The "Euro-Lupus Cohort" is composed by 1000 patients with systemic lupus erythematosus (SLE) that have been followed prospectively since 1991. These patients have been gathered by a European consortium--the "Euro-Lupus Project Group". This consortium was originated as part of the network promoted by the "European Working Party on SLE", a working group created in 1990 in order to promote research in Europe on the different problems related to this disease. The "Euro-Lupus Cohort" provides an updated information on the SLE morbidity and mortality characteristics in the present decade as well as defines several clinical and immunological prognostic factors.

Unrelated donor stem cell transplantation in adult patients with thalassemia.

Allogeneic SCT remains the only potential cure for patients with thalassemia. However, most BMT candidates lack a suitable family donor and require an unrelated donor (UD). We evaluated whether BMT using UDs in high-risk adult thalassemia patients can offer a probability of cure comparable to that reported employing an HLA-compatible sibling as donor. A total of 27 adult thalassemia patients (15 males and 12 females, median age 22 years) underwent BMT from a UD selected by high-resolution HLA molecular typing. The conditioning regimen consisted of Busulphan (BU, 14 mg/kg) plus Cyclophosphamide (CY, 120 or 160 mg/kg) in 12 cases and BU (14 mg/kg), Thiotepa (10 mg/kg) and CY (120-160 mg/kg) in the remaining 15 cases. Cyclosporine-A and short-term Methotrexate were used for graft-versus-host disease (GVHD) prophylaxis. In all, 19 patients (70%) are alive and transfusion-independent after a median follow-up of 43 months (range 16-137). A total of 10 patients (37%) developed grade II-IV acute GVHD and six (27%) chronic GVHD. Eight patients (30%) died from transplant-related causes. UD-BMT can cure more than two-thirds of adult thalassemia patients, and is a particularly attractive option for patients who are not compliant with conventional treatment.

Association of HLA-DRB3*0202 and serum IgG antibodies to Chlamydia pneumoniae with essential hypertension in a highly homogeneous population from Majorca (Balearic Islands, Spain).

Separate studies investigating the relationship of essential hypertension (EH) with the HLA system and with Chlamydia pneumoniae (C. pneumoniae) infection have given conflicting results. Our aim was to clarify these relationships and determine whether the HLA system and C. pneumoniae infection interact with respect to the risk for EH. An association study (110 essential hypertensives and 107 controls) was conducted in a highly homogeneous population in the Balearic Island of Majorca (Spain). Molecular typing of HLA-B and HLA-DRB and quantification of serum levels of IgG antibodies to C. pneumoniae (sIgGa-Cp) were determined. Student's t-test, chi(2)-statistics, logistic regression analysis, and general linear model ANOVA were used for statistical analysis. The results showed that EH was related with HLA-DRB3*0202 in the whole study population, and with levels of sIgGa-Cp>63.5 BU/ml in the group of individuals with sIgGa-Cp>30 BU/ml (OR (95% CI) adjusted for obesity, familial history of EH and diabetes=2.06 (1.07-3.97), P=0.03, and =4.60 (1.06-19.90), P=0.04, respectively). The association between EH and sIgGa-Cp was observed in the DRB3*0202(+) individuals, but not in the DRB3*0202(-) subgroup (OR (95% CI)=11.14 (1.92-64.54), P=0.004, and =0.98 (0.22-4.43), P=0.64, respectively (P of the Mantel-Haenszel test for homogeneity of OR=0.06)). In our population, EH was positively associated with HLA-DRB3*0202 and with high levels of sIgGa-Cp. Moreover, a significant interaction of DRB3*0202 on the effect of sIgGa-Cp was observed, as the association of EH with these antibodies depended on the presence of DRB*0202.

Expression of ATP7B in normal human liver.

ATP7B is a copper transporting P-type ATPase, also known as Wilson disease protein, which plays a key role in copper distribution inside cells. Recent experimental data in cell culture have shown that ATP7B putatively serves a dual function in hepatocytes: when localized to the Golgi apparatus, it has a biosynthetic role, delivering copper atoms to apoceruloplasmin; when the hepatocytes are under copper stress, ATP7B translocates to the biliary pole to transport excess copper out of the cell and into the bile canaliculus for subsequent excretion from the body via the bile. The above data on ATP7B localization have been mainly obtained in tumor cell systems in vitro. The aim of the present work was to assess the presence and localization of the Wilson disease protein in the human liver. We tested immunoreactivity for ATP7B in 10 human liver biopsies, in which no significant pathological lesion was found using a polyclonal antiserum specific for ATP7B. In the normal liver, immunoreactivity for ATP7B was observed in hepatocytes and in biliary cells. In the hepatocytes, immunoreactivity for ATP7B was observed close to the plasma membrane, both at the sinusoidal and at the biliary pole. In the biliary cells, ATP7B was localized close to the cell membrane, mainly concentrated at the basal pole of the cells. The data suggest that, in human liver, ATP7B is localized to the plasma membrane of both hepatocytes and biliary epithelial cells.

Association of monoamine oxidase B alleles with age at onset in amyotrophic lateral sclerosis.

An active role of monoamine oxidase B (MAO-B) in the pathogenesis of neurodegenerative disorders such as Parkinson's disease has been proposed as the enzyme is known to be a generator of free radicals which seem to be responsible for neuron oxidative damage. We evaluated the influence of MAO-B in the pathogenesis of the sporadic forms of Amyotrophic lateral sclerosis (ALS) by studying the MAO-B allele distribution in 51 patients and 71 healthy controls. MAO-B did not directly result in a risk factor for ALS but seemed to strongly influence age at onset. The mean ALS onset age was significantly higher in individuals carrying allele 5 compared to individuals without this allele (60.4 +/- 8.1 vs. 52.1 +/- 10.3 years; P = 0.004). These results, in agreement with findings in the literature, suggest an increased MAO-B expression in ALS and support the hypothesis that neuronal cell death in neurodegenerative diseases is triggered by astroglial reaction.

HLA-DM polymorphisms do not associate with multiple sclerosis: an association study with analysis of myelin basic protein T cell specificity.

The MHC region on 6p harbors at least one susceptibility gene for multiple sclerosis (MS). Within this region, HLA-DM loci are of interest being involved in class II antigen processing. We investigated the association of HLA-DM polymorphisms with MS. Sixty-three patients with MS and 46 healthy controls from continental Italy were typed for HLA-DM polymorphisms and HLA-DRB1 alleles. Besides, among the donors characterized for the DM polymorphisms, we considered 6 MS patients previously studied for the fine specificity of their MBP-specific T lymphocyte lines (TLL). The frequencies of allelic variants at the DMA and DMB loci were similar between MS patients and controls, even when HLA-DRB1*1501 positive and negative donors were analyzed separately. Patients with predominant responses to different MBP epitopes did not differ for their HLA-DM haplotype while patients with predominant responses to the same MBP epitope could present different HLA-DM haplotypes. HLA-DM polymorphisms do not associate with MS and may not affect specific patterns of T cell responses to MBP.

Is the MHC a general self-recognition system playing a major unifying role in an organism?

From the review of several recent observations of cell-cell interactions, which occur preferentially in autologous or syngeneic situations such as rosettes, adhesion, homing, and contact inhibition, the existence of an active general process of cellular self-recognition, not limited to the immune system, is postulated. This process is MHC associated or dependent, and seems to require an identity of ubiquitous molecules of class I--or other linked gene products at the surface of interacting cells. In contrast, class II molecules are not apparently implicated in general self-recognition. The immune system is regarded as a late evolution from a self-recognition system. It retains the ability of self-evolution, but possesses the exclusive property of active discrimination against foreignness. The astonishing fact that identity of MHC products seems to be needed for recognition is discussed in the context of various possible mechanisms. From immunological and genetical consideration, it is proposed that the genuine biological role of the MHC would be that of a self-recognition and unifying system.

A new HLA-DR2 extended haplotype is involved in insulin-dependent diabetes mellitus susceptibility.

To ascertain why HLA-DR2 seems to confer only a moderate resistance to insulin-dependent diabetes mellitus (IDDM) in the high-incidence population of Sardinia, Italy, 32 families having one individual affected with IDDM (the proband) and 31 families without IDDM history were randomly selected from the same geographical area and serologically and molecularly HLA typed. The 64 haplotypes of the probands were then compared with the 122 haplotypes determined in the parents from the control families. Two haplotypes were found to have the highest percentage in the general population (12.3% and 7.3%, respectively). The first is the already described "Sardinian" extended haplotype A30, Cw5, B18, 3F130, DR3, DRw52, DQw2 (39.0% in IDDM patients). The second is an extended haplotype that has not been identified before (A2, Cw7, B17, 3F31, DR2, DQw1), and, due to the DR2 allele, we expected it to be decreased in IDDM. However, a stratified analysis performed by removing the DR3 and DR4 haplotypes showed that the frequency of this haplotype is significantly increased in IDDM patients. A peculiar feature of this haplotype is its DQw1 allele, which is DQB1*0502 and has serine in position 57 of the DQ beta chain. The absence of an aspartic acid in this position seems to confer susceptibility to IDDM and not resistance. The fact that DQB1*0502 was present in 75% of the Sardinian DR2 haplotypes may explain why, in Sardinia, DR2 is not providing the commonly recognized resistance to IDDM.

Defining the allelic variants of HLA-A30 in the Sardinian population using amplification refractory mutation system--polymerase chain reaction.

HLA-A30 is present in the Sardinian population at a frequency of 23%. We have designed a system using nested ARMS-PCR to determine the relative frequencies of the HLA-A*30 allelic variants (A*3001, A*3002, and A*3003) within this population. The use of a nested PCR approach, in which the first-round reaction provides HLA-A*30 specificity and template DNA for the subsequent nested reactions, is a powerful means of discriminating between alleles of very similar sequence. Using this method, we performed subtyping of 35 serologically defined HLA-A30 Sardinian individuals, and taking into account homozygotes, identified 38 A*30 alleles. Of these, 33 typed as A*3002, four typed as A*3001, and one sample did not conform to the patterns of reactivity of any of the published A*30 alleles. Haplotype information showed strong linkage disequilibrium between A*3002 and B18. This study underlines the potential of DNA-based methods for typing HLA class I in terms of adding further levels of definition to studies of population structure and also as a means of identifying new alleles.

A randomized trial comparing the introduction of ritonavir or indinavir in 1251 nucleoside-experienced patients with advanced HIV infection.

ISS-IP1, a multicenter, randomized, 48-week open trial, was designed to compare the introduction of ritonavir or indinavir in patients with previous nucleoside experience and CD4+ cell counts below 50/mm3. Concomitant antiretroviral treatment with nucleoside analogs was allowed. Primary efficacy measures were survival and time to a new AIDS-defining event or death, analyzed through the whole period of observation by the intention-to-treat approach. Primary toxicity measures were time to treatment discontinuation and adverse events, grade at least 3/serious, analyzed by an on-treatment approach. Evaluation-of efficacy also included CD4+ cell and RNA response. The trial enrolled 1251 patients in 5 months. At baseline, mean CD4+ cell count was about 20 cells/mm3 and mean HIV RNA copy number was 4.9 log10/ml in both groups. Overall, 402 patients in the ritonavir group and 250 patients in the indinavir group permanently discontinued the assigned treatment (relative risk, 1.96; 95% CI, 1.68-2.30; p = 0.0001), with most of this difference dependent on a higher number of discontinuation for adverse events in the ritonavir group. After a mean follow-up of 307 days (ritonavir, 304; indinavir, 309), 124 deaths (ritonavir, 61; indinavir, 63; relative risk, 0.96; 95% CI, 0.67-1.36; p = 0.80) and 330 new AIDS-defining events (ritonavir, 170; indinavir, 160; relative risk, 1.05; 95% CI, 0.85-1.31; p = 0.60) were observed. CD4+ cell counts increased in both groups in patients still receiving treatment, with about 100 cells gained by week 24 and 150 cells gained by week 48. Body weight also increased over time in both groups. Analysis of RNA response showed a decrease of 1.5 log10 or higher in both treatment groups. Overall, 400 patients in the ritonavir group and 338 patients in the indinavir group developed at least one grade 3/serious new adverse event during follow-up (relative risk, 1.48; 95% CI, 1.28-1.72; p = 0.0001). Favorable CD4+ cell and RNA responses at 24 and 48 weeks were observed in both groups of patients remaining on treatment. Indinavir showed slightly better effects in sustaining RNA, CD4+ cell, and body weight responses. Ritonavir and indinavir results were comparable in terms of clinical outcome (survival and AIDS-defining events).

Familial hypercholesterolemia study in Sardinia using 6 LDLR polymorphic markers based on PCR.

Twenty-two Sardinian families with multiple cases of hypercholesterolemia were investigated with six polymorphic markers of the low-density lipoprotein receptor (LDLR) gene that could be quickly analyzed by PCR-based methods. Five single nucleotide polymorphisms (SNP) in exons 8, 10, 13, 15, and 18 and a microsatellite marker flanking the 3' end of the LDLR gene were used to define the haplotypes at the LDLR locus for familial hypercholesterolemia (FH) diagnosis within families. No significant differences were observed between the allele frequencies of the normal and mutant chromosomes. In two families, hypercholesterolemia did not cosegregate with the LDLR locus. In the remaining 20 FH chromosomes, seven different haplotypes were identified. The same haplotypes were found with a similar frequency among the 61 normal chromosomes. Other five haplotypes were characteristic only of normal chromosomes. These data provide no evidence for a gene founder effect in the Sardinian population and, instead, highlight a pattern of genetic heterogeneity comparable with that found in mainland European populations. The replacement of the restriction fragment length polymorphisms currently used in the genetic analysis of FH with PCR-based markers proved to be a simple and less time-consuming method and did not reduce informativity in the molecular analysis of FH families.

Association of the HLA-A2, CW2, B27, S31, DR2 haplotype with ankylosing spondylitis. A possible role of non-B27 factors in the disease.

With the aim of searching for HLA haplotypes and non-B27 allele frequency variations in Sardinian AS patients, HLA-A, B, Cw, DR, DQ and Bf, C4A and C4B typing and haplotype assignment was carried out in the families of 25 AS patients and in 44 healthy individuals, all B27 heterozygotes. In the AS patients a significant increase of the A2, Cw2, B27, DR2, DQ1 haplotype was found. This depends only partially on the linkage disequilibrium existing in the Sardinian population between B27 and the other alleles of this haplotype, and rather seems to be due to a primary association of Cw2 and DR2 alleles with AS. Preliminary data seem to show that this haplotype bears the S31 complotype and the DRB1*1601 allele both in the AS patients and in the healthy controls. The pathogenetic implications of these findings are discussed.

Implication of HLA-DMA alleles in corsican IDDM.

The HLA-DM molecule catalyses the CLIP/antigen peptide exchange in the classical class II peptide-binding groove. As such, DM is an antigen presentation regulator and may be linked to autoimmune diseases. Using PCR derived methods, a relationship was revealed between DM gene polymorphism and IDDM, in a Corsican population. The DMA*0101 allele was observed to confer a significant predisposition to this autoimmune disease while the DMA*0102 allele protected significantly. Experiments examining polymorphism of the HLA-DRB1 gene established that these relationships are not a consequence of linkage disequilibrium with HLA-DRB1 alleles implicated in this pathology. The study of the DMA gene could therefore be an additional tool for early IDDM diagnosis in the Corsican population.

Allogeneic bone marrow transplantation combined with multiple anti-HIV-1 treatment in a case of AIDS.

A 25-year-old woman with AIDS was submitted to HLA-identical allogeneic BMT after cytoablation with busulphan and cyclophosphamide and combined anti-HIV-1 therapy with zidovudine, IFN-alpha 2 and anti-HIV-1-specific T cell clones. Marrow engraftment occurred after 18 days and tests for HIV-1 were negative after 30 days but the hematologic reconstitution of the patient was poor. A second BM infusion from the same donor was ineffective and treatment with GM-CSF only induced a transient increase of the blood cell count, suggesting iatrogenic damage to the BM microenvironment. The development of ARDS led to the death of the patient 10 months after transplantation. Post-mortem investigation did not reveal any active infections and PCR on autopsy tissues was negative for HIV-1.

Successful unrelated bone marrow transplantation in beta-thalassaemia.

A 16-year-old Sardinian girl affected by homozygous beta-thalassaemia was submitted to allogeneic BMT using an HLA-identical, MLC-negative, unrelated donor. The donor and the patient were homozygous for the entire extended haplotype A30, Cw5, B18, F130, DRB1*0301, DRB3*0202, DQA1*0501, DQB1*0201 and heterozygous for DPB1*0301/DPB1*0202. The conditioning regimen consisted of 14 mg/kg busulphan and 160 mg/kg cyclophosphamide. Engraftment was achieved 14 days from BMT and the haematological reconstitution was complete without any signs of acute or chronic GVHD. Seven months after the transplant the patient was in excellent general condition. The hypothesis is advanced that when two HLA extended haplotypes are shared by donors and recipients, particularly in homozygosity, this is a very favourable immunogenetic condition in unrelated BMT.

alpha 2 Antiplasmin and disseminated intravascular coagulation in liver cirrhosis.

Subnormal concentrations of alpha 2 Antiplasmin (alpha 2 AP) in liver cirrhosis may be due to an impaired hepatic synthesis and/or to a fibrinolysis activation in disseminated intravascular coagulation (DIC). In order to clarify this problem, in 26 cirrhotic patients (15 compensated and 11 decompensated) alpha 2 AP plasma activity and plasma Fibrinopeptide A (FPA) were measured. Serum albumin, p-Cholinesterase (p-CHE), Fibrinogen and Fibrinogen Degradation Products (FDP) were also carried out. Our data show that alpha 2 AP and FPA were equally abnormal in compensated and decompensated cirrhosis. The significant negative correlation obtained between alpha 2 AP and FPA as well as the lack of correlation between alpha 2 AP and albumin, alpha 2 AP and p-CHE in both groups suggests that, in our patients, alpha 2 AP decrease may be due to a fibrinolysis activation induced by a DIC which appears chronic since Fibrinogen and FDP were normal. These findings are in agreement with the results obtained in the four subgroups a posteriori selected on the basis of FPA levels: alpha 2 AP in subgroups with high FPA was significantly different from controls while it did not differ in subgroups with normal FPA.

The role of HLA-DR antigens in transplantation--survival of skin allografts in HLA-haploidentical donor-recipient combinations.

The results of 79 skin grafts performed in haploidentical donor-recipient pairs are correlated with HLA-A, -B, -C, and -DR compatibility. A strong detrimental effect of DR incompatibilities has been demonstrated. This effect is independent from that exerted by products of the HLA-A, -B, and -C loci. An additive effect of HLA-A, -B, and -DR incompatibilities on allograft survival time has been observed.