Anti-cyclic citrullinated peptide antibodies in type 1 autoimmune hepatitis.

BACKGROUND: Besides the autoantibodies included in the diagnostic criteria of type 1 autoimmune hepatitis, many other autoantibodies have been described in this condition. Recently, antibodies against cyclic citrullinated peptide have been validated as specific diagnostic and prognostic markers of rheumatoid arthritis. AIM: To assess whether these antibodies are part of the autoantibody repertoire of type 1 autoimmune hepatitis and correlate with rheumatological manifestations. METHODS: Antibodies against cyclic citrullinated peptide were tested by a commercially available enzyme-linked immunosorbent assay. RESULTS: The antibodies were found in 12 of 133 (9%) type 1 autoimmune hepatitis, two of 49 (4%) with primary biliary cirrhosis, one of 80 (1%) with hepatitis C virus-related chronic liver disease and 53 of 89 (60%) with rheumatoid arthritis serum samples. High titres were found only in rheumatoid arthritis and type 1 autoimmune hepatitis. No clinical (in particular rheumatological manifestations), biochemical or immunoserological differences were detectable between antibodies against cyclic citrullinated peptide positive and negative type 1 autoimmune hepatitis sera, with the exception of rheumatoid factor, always negative in the positive ones. CONCLUSIONS: Antibodies against cyclic citrullinated peptide can be detected in a subgroup of patients with type 1 autoimmune hepatitis. They might be part of the wide range of autoantibody production characteristic of this condition and/or, less probably, be predictive of future rheumatoid arthritis development.

Haplotype HLA-B8-DR3 confers susceptibility to hepatitis C virus-related mixed cryoglobulinemia.

Our aim was to investigate whether host genetic factors are involved in the onset of hepatitis C virus (HCV)-related mixed cryoglobulinemia (MC). We studied 25 consecutive patients presenting with a full-blown clinical picture of MC by physical examination, blood chemistry, assessment of cryoglobulins and their composition, nonorgan-specific autoantibodies, antibodies to HCV, serum HCV RNA, and HLA polymorphism. Biopsies of liver, bone marrow, and minor salivary glands were also performed in a number of patients. HLA results were compared with those of normal controls and patients with chronic HCV infection without MC and negative for autoimmune phenomena (pathological controls). Type II MC was found in 14 of 25 patients (56%), and type III MC was found in the remaining 11 (44%). All patients were positive for antibodies to HCV and/or serum HCV RNA. HLA-B8 was found in 40% (10 of 25) of patients compared with 10. 1% (38 of 377) of normal controls (P = .00003, Pcorrected = .0005, relative risk [RR] 5.9) and 6.7% (2 of 30) of pathological controls (P = .007, Pcorrected = not significant). As for class II HLA molecules, only DR3 was significantly more frequent in MC patients (40%, 10 of 25) than in normal controls (15.1%, 57 of 377; P = .003, Pcorrected = .03, RR 3.7). Odds ratio (OR) for the risk of developing MC was calculated in patients positive for B8 and/or DR3, and the highest OR (8.2) was observed in individuals possessing both. The results suggest that the development of HCV-related MC is associated with HLA-B8 and DR3 markers.

Wheat deficient in gliadins: promising tool for treatment of coeliac disease.

The toxicity of two varieties of bread wheat, one poor in alpha and beta gliadins and the other poor in alpha, beta, gamma, and omega gliadins, has been tested. The peptic-tryptic digest of these wheats was assessed using coeliac mucosa in an in vitro organ culture system. A significantly lower toxicity was found in respect of bread wheat containing all gliadin fractions. These results suggest new opportunities for the treatment of coeliac disease.

Asp57-negative HLA DQ beta chain and DQA1*0501 allele are essential for the onset of DQw2-positive and DQw2-negative coeliac disease.

The genetic predisposition to coeliac disease is associated with the HLA DQw2 allele. Coeliac patients lacking the DQw2 allele are very rare and always exhibit the DR4-DQw3 haplotype. We performed oligotyping of polymerase chain reaction (PCR)-amplified DQA1 and DQB1 genes in six DQw2-negative and 30 DQw2-positive coeliac patients. The DQB analysis showed that all six DQw2-negative patients possessed the DQB1*0302 allele. The other DQB alleles found in five of these patients were DQB1*0501, DQB1*0604 and DQB1*0302. The DQ beta chains encoded from all these alleles have the replacement of aspartic acid residue at position 57 (Asp57), as well as the DQB1*0201 allele which was found in all 30 DQw2-positive coeliac patients. The DQw2-negative proband who lacked the homozygous Asp57 replacement exhibited the DQA1*0501 allele in the DQA1 gene. The DQA1*0501 allele was also found in 27 of the 30 DQw2-positive coeliac patients. Among this group of coeliacs, the four cases lacking the DQA1*0501 allele exhibited the homozygous Asp57 replacement in the DQ beta chain. Our results indicate that Asp57-negative DQ beta alleles are involved in both DQw2-positive and -negative coeliac patients. Moreover, when the Asp57-negative DQ beta chain is encoded from only one of the two DQB1 genes the DQA1*0501 allele is always present.

Subclinical celiac sprue. Increasing occurrence and clues to its diagnosis.

We have reviewed the clinical records of 226 consecutive adult patients with celiac sprue diagnosed in our department from 1972 to 1989. The study period has been divided into three subperiods of 6 years each (1972-1977, 1978-1983, 1984-1989). From the first to the third subperiod a significant increase in the number of new diagnoses and of the proportion of patients recognized by minor symptoms has been observed. These patients represent 50% of our series in the last 3 years of the study and 70% in 1989. Diagnosing subclinical forms of celiac sprue has significantly lowered the mean age at diagnosis and the female/male ratio. First-degree relatives of celiac patients; subjects who had a gluten-free diet for a period during childhood; patients with short stature, anemia, or amenorrhea of no obvious cause; or patients with unexplained immunological abnormalities are the groups in which most patients with subclinical celiac sprue have been found and in which potential patients should be sought. Helpful diagnostic tools include antigliadin antibody testing and the observation of absent or reduced Kerckring folds in the descending duodenum in the course of upper gastrointestinal endoscopy.

Gliadin immune reactivity is associated with overt and latent enteropathy in relatives of celiac patients.

Antigliadin antibodies have been widely used in the screening of celiac disease. Using this test, candidates for jejunal biopsy were selected from 328 first-degree relatives of 128 adult celiac patients. All relatives geographically available were willing to participate in the study. Twenty-one turned out to be positive for antigliadin antibodies, and in 13 jejunal histology was consistent with celiac disease. In the remaining 8, the morphometric evaluation of jejunal biopsy specimens showed a mean surface to volume ratio that, although in the normal range, was significantly lower than that of other 10 relatives negative for antigliadin antibodies and 16 biopsied controls. It was concluded that antigliadin antibody testing is a valuable method for the screening of celiac disease among family members and that relatives with genetically predisposed gliadin sensitivity, without gross histological lesions but with minor morphometric abnormalities of the jejunal mucosa, may be regarded as subjects with latent celiac disease.

HLA-DP polymorphism in northern Italian celiac patients.

The contribution of HLA-DP genes to celiac disease susceptibility has been investigated in 95 Italian patients, 41 with childhood and 54 with adult disease onset. Polymerase chain reaction amplification, sequence-specific oligonucleotide probe hybridization and restriction fragment length polymorphism analyses have been carried out. All celiac patients and 56 out of 128 random healthy controls were DQw2-positive. The frequency of the DPB1*0101 allele was significantly increased (pc = 0.002, relative risk 5.21) in patients with celiac disease (23.2%) compared to the whole panel of controls (5.5%), but not to the 56 controls bearing DQw2 (10.7%). No significant difference in the frequency of DPB1*0101 was found between celiac patients with pediatric (24.4%) or adult (22.2%) onset. The DPB1*0101 allele was associated with both the DR3-DQw2 and DR7-DQw2 haplotypes. Moreover, our study has not confirmed the association with DPB1*0402 and DPB1*0301 previously reported in celiac children from southern Italy. The linkage of the DPB1*0101 allele with the DQ locus and the observation that the DP but not the DQ association appears to be ethnically dependent strongly support a secondary role of DP molecules in celiac disease susceptibility.

Molecular analysis of HLA-DQ A alleles in coeliac disease lack of a unique disease-associated sequence.

Susceptibility to coeliac disease is strongly associated with some HLA class II antigens, encoded by the HLA-D region. Since the HLA-DQ locus seems to be primarily involved, we have analysed by polymerase chain reaction amplification and allele-specific oligonucleotide hybridization the most polymorphic region of the HLA-DQ A1 gene. No difference was observed between the 20 coeliac patients and 20 HLA-D-matched healthy controls who took part in the study. Furthermore, in patients and controls, the restriction fragment length polymorphism analysis of the HLA-DQ A gene using the restriction enzyme BglII did not disclose any specific disease-associated fragment. Our results are not consistent with a unique DQ A coeliac disease-associated sequence, but rather with the hypothesis that some polymorphic residues or allelic hypervariable regions, although found also in the normal population, can predispose to coeliac disease due to their higher frequency in this condition.

IgA antigliadin antibodies and persistence of jejunal lesions in adult coeliac disease.

In 46 adult patients with coeliac disease, 41 (89%) of whom were positive for IgA and/or IgG antigliadin antibodies (AGA) when untreated, we investigated after a gluten-free diet the relationship between the persistence of AGA, the persistence of jejunal lesions, and the duration and compliance with the diet. IgG AGA were positive in 21 coeliac patients (46%) after variable periods of gluten-free diet and were associated with IgA positivity only in 4 cases (9%). Both IgA and IgG AGA positivity appeared to be more related to the lack of improvement of the jejunal lesions than to the strictness and duration of gluten withdrawal. Nine coeliacs showed no improvement of jejunal lesions after the gluten-free diet. Of these 9, 4 showed persistent IgA AGA, while the remaining 5 resulted IgAAGA-negative as before when untreated. Though intestinal biopsy remains the best means of determining the positive effect of gluten withdrawal, the persistence of IgA AGA in treated coeliacs is always predictive of the persistence of severe jejunal lesions. The persistence of IgG AGA, on the contrary, should be regarded as an immunological memory.

Howell-Jolly body counting as a measure of splenic function. A reassessment.

Non-surgical and surgical asplenia predisposes to fatal infections; therefore, simple, non-invasive and repeatable tests for assessing splenic function are required, even in non-specialized medical institutions. Howell-Jolly bodies are the most characteristic peripheral blood abnormality after splenectomy, but their counting is not considered a reliable measure of splenic function. In this study, in a group of splenectomized subjects and of patients with non-surgical hyposplenism, we have compared counting of Howell-Jolly bodies, stained by both the May-Grünwald/Giemsa method and the Feulgen reaction, with pitted cell counting which is considered a reliable technique for the assessment of splenic hypofunction. A significant correlation has been found between Howell-Jolly body counts, stained by either technique, and pitted cell counts (P less than 0.0001). Through Howell-Jolly bodies were never detectable when pitted cell counts fell between 4 and 8%, values consistent with a very mild splenic hypofunction, for pitted cell counts above 8% their increase was always associated with increasing Howell-Jolly body counts. These data suggest that, although pitted cell counting represents a more sensitive method for evaluating splenic function, Howell-Jolly body counting may still be regarded as a simple and reliable technique for identifying and monitoring those cases associated with a real risk of overwhelming infections.

Cytophilic antibodies cause leucocyte migration inhibition in coeliac disease.

Leucocyte migration inhibition in response to challenge with gluten fraction III was prevented by pretreatment of coeliac leucocytes with pronase, an enzyme that removes cytophilic antibodies. Leucocytes from healthy volunteers preincubated with serum from untreated and treated coeliac patients showed marked migration inhibition when incubated with gluten fraction III. No migration inhibition was observed with leucocytes from healthy volunteers preincubated with serum from patients with small bowel diseases other than coeliac disease and from other healthy volunteers. Untreated and treated coeliac patients who were leucocyte migration inhibition negative in the direct assay had migration indices in the sensitized range in the serum induced assay. This work is in keeping with a previous suggestion that in coeliac disease leucocyte migration inhibition induced by gluten fraction III is caused by cytophilic antibodies.

Gluten specific suppressor T cell dysfunction in coeliac disease.

A T lymphocyte direct migration inhibition factor test has been used to investigate the function of the specific suppressor T cell population controlling the immune response to gluten in coeliac disease. The test has been carried out in 21 adult coeliac patients, 22 Mantoux- healthy controls and eight Mantoux+ donors using gluten fraction III and purified protein derivative, as antigens. All coeliacs, but two, were Mantoux-. When gluten fraction III was used a significant migration inhibition was observed in coeliac patients compared to controls; such migration inhibition was abrogated by coculturing in a 1:1 ratio coeliac T cells with T cells from controls or Mantoux+ donors. On the contrary, the addition to coeliac T cells of T lymphocytes from other coeliacs did not abolish migration inhibition to gluten. Pretreatment of normal T cells with mitomycin C prevented their abrogating activity on migration inhibition of coeliac T lymphocytes. When purified protein derivative was used as antigen a significant migration inhibition was observed in Mantoux+ donors compared with healthy subjects and such migration inhibition was abolished by co-culturing T cells from Mantoux+ donors with those from Mantoux- controls and coeliac patients. Our results show that coeliac T cells, while retaining their ability to suppress the immune response to purified protein derivative, cannot suppress the immune response to gluten and are consistent with the hypothesis that a gluten specific suppressor T cell dysfunction, rather than a generalised T lymphocyte defect, may play a role in the pathogenesis of coeliac disease.