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Summary Background Serum antinuclear antibodies giving the ‘multiple nuclear dots’ or the ‘rim-like/membranous’ patterns are frequently detected by indirect immunofluorescence on HEp-2 cells in patients with primary biliary cirrhosis. Aim To assess the accuracy of multiple nuclear dot and rim-like/membranous antinuclear antibodies for the diagnosis of primary biliary cirrhosis. Methods Sera from 4371 consecutive patients referred to our laboratory were analysed under code for antinuclear antibodies testing by indirect immunofluorescence on HEp-2 cells. Results Review of the clinical records of the 4371 patients allowed identification of 101 patients with antimitochondrial antibody-positive primary biliary cirrhosis and 22 with antimitochondrial antibody-negative variant. Multiple nuclear dot and/or rim-like/membranous patterns were found in 59 (1.3%) of the 4371 patients: 31 antimitochondrial antibody-positive primary biliary cirrhosis, 17 antimitochondrial antibody-negative primary biliary cirrhosis and 11 non-primary biliary cirrhosis. The specificity for primary biliary cirrhosis of both the antinuclear antibodies pattern was 99%. Positive predictive value and likelihood ratio for a positive test were 86% (95% CI: 72.7–94) and 221 (95% CI: 91.7–544) for multiple nuclear dot, 79% (95% CI: 62.2–90.1) and 132 (95% CI: 56.8–312.7) for rim-like/membranous, respectively. Conclusions Multiple nuclear dot and rim-like/membranous antinuclear antibodies are rare findings. Their positivity strongly suggests the diagnosis of primary biliary cirrhosis, irrespective of antimitochondrial antibody status. The high specificity for primary biliary cirrhosis makes them a useful diagnostic tool especially in antimitochondrial antibody-negative patients.

SUMMARY Serum IgA anti-gliadin antibodies (AGA) were positive in 25 (68%) of 37 untreated adults with coeliac disease belonging mostly to IgAl subclass (88%) and only in a few cases to IgA2(12%). Anti-secretory component IgA AGA were present in serum of seven patients (28%) by immunofluores-cence and in nine (36%) by ELISA. The search for IgA AGA in jejunal juice of eight untreated children with coeliac disease was positive in seven cases (88%), with consistent finding of anti-secretory component IgA AGA. These antibodies belonged with equal proportion to IgAl and IgA2 subclasses. This study shows that in intestinal secretions of untreated coeliac disease cases the IgA immune response to gliadin is confined to polymeric anti-secretory component IgA with the same prevalence of IgAl and IgA2 subclasses, while in serum IgA AGA are largely monomeric, more frequently of IgAl than of IgA2 subclass, and with a lower proportion of polymeric anti-secretory component IgA (20–36%). The finding of secretory IgA AGA in serum of patients with coeliac disease could result from a spill-over from the intestinal mucosal synthesis into the circulation.

Summary 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.

Editor,—Recent studies have demonstrated that coeliac disease can develop months or years after the clinical onset of insulin dependent diabetes mellitus (IDDM).1 Over the last 10 years, 200 consecutive children with IDDM at the onset have been tested for IgG and IgA antigliadin antibodies (AGA) by indirect immunofluorescence2 and for IgA antiendomysial antibodies (EmA) using monkey oesophagus and, more recently, human umbilical cord as substrate.3 An antibody follow up was also performed in 151 of these patients every 3–6 months. At initial testing six diabetic children (four girls and two boys, median age 9 years, range 2–13) were positive for both IgA EmA and IgA AGA (associated with IgG …

To obtain information on the mechanisms of hepatocellular damage and the determinants of response to interferon, hepatitis C virus (HCV) genotype, tissue HCV antigens, hepatocellular expression of HLA-A,B,C and intercellular adhesion-1 molecules, and the number of lobular T lymphocytes were studied in 38 anti-HCV-positive patients. 14 patients did not show a primary response to interferon treatment. HCV genotype 1b was detected in 11 of them. They displayed higher scores of HCV-positive hepatocytes, HLA-A,B,C, and ICAM-1 molecules expression than with the responders. HCV-infected hepatocytes maintained the capacity to express HLA-A,B,C and ICAM-1 molecules. CD8-positive T cells in contact with infected hepatocytes and Councilman-like bodies were observed. A significant correlation was found between the number of lobular CD8-positive T cells and alanine amino transferase levels. No differences were observed in clinical, biochemical, and histological features between patients with high and low number of hepatocytes containing HCV antigens. These data suggest a prominent role of T cell-mediated cytotoxicity in the genesis of hepatocellular damage. The high expression of interferon-inducible antigens like HLA-A,B,C molecules suggests the presence of strong activation of the interferon system possibly related to high HCV replication in nonresponder patients infected with genotype 1b.

SUMMARY The specificities of anti-nuclear antibodies (ANA) reacting with multiple nuclear dots (MND-ANA) present in about 15% primary biliary cirrhosis sera were studied by Western blot analysis with nuclear fractions from a human cell line. Reactivity with two broad bands of 78–92 kD and 96–00 kD of the insoluble fraction was present exclusively in MND-ANA-positive sera. Antibodies eluted from these proteins specifically retained the immunofluorescence reactivity of MND-ANA. Immunomorphological analysis by a pre-embedding technique revealed that the antibody specifically binds to nuclear regions resembling in size and number nuclear bodies. Since these structures are absent in immature rate endometrial cell and can be induced by diethylstilbestrol. we tested MND-ANA by immunofluorescence on cryostat sections of uteri from hormone-treated and untreated immature rats. A strong reaction of nuclear dots was observed predominantly in endometrial cells of hormone treated rats. We thus conclude that MND-ANA present in primary biliary cirrhosis sera are directed against 78–92-kD and 96–100-kD nuclear proteins located in nuclear bodies.

Serum samples from 66 seropositive subjects (56 with a history of intravenous drug abuse), including asymptomatic carriers and patients with persistent generalised lymphadenopathy (PGL), AIDS related complex (ARC), and AIDS, were tested by indirect immunofluorescence on rat tissue sections and HEp-2 cells for the presence of antibodies to nuclei, smooth muscle, intermediate filaments (anti-IMF) and microfilaments (anti-MF). Counterimmunoelectrophoresis was also used to detect antibodies to extractable nuclear antigens. Smooth muscle antibodies with the V pattern or antinuclear antibodies, mainly of the speckled type, or anti-IMF, occurred in 35 cases, being widely distributed in all groups. Such an autoantibody response resembles the "viral" autoimmunity described in various infectious diseases and in particular that of non-A, non-B post-transfusion hepatitis. Autoantibodies may be of some prognostic relevance, as the prevalence of smooth muscle antibodies V increased as the disease progressed (asymptomatic carriers 20%, those with PGL 29%, those with ARC 47%, and those with AIDS 63%. In the PGL group autoantibody positivity correlated with the presence of skin anergy. The fact that autoantibodies were more frequently detected in patients with circulating immune complexes suggests that these can contain autoantibodies and the corresponding autoantigens.

Recent evidence shows that chronic liver disease induced by hepatitis delta virus is closely associated with production of autoantibodies. To verify this sera from patients with hepatitis delta virus and from those with hepatitis B virus mediated chronic liver disease were tested for a panel of autoantibodies. Although the traditional non-organ specific autoantibodies were similarly distributed in the two groups, a considerably higher prevalence of IgG basal cell layer antibodies, IgM anti-intermediate filaments, and IgG antimicrotubule antibodies was found. Although these phenomena might be secondary to hepatitis delta virus infection, they might be the serological markers of underlying immune events.

Engraftment by recipient's (R) cells has been already demonstrated in gender mismatched liver grafts using fluorescence in situ hybridization (FISH), with contrasting results concerning epithelial cells. Mismatch for human leukocyte antigen (HLA) class I (HLA-I) is quite common in patients with orthotopic liver transplantation (OLT). We thus aimed to assess whether monoclonal antibodies (MoAbs), currently employed in the HLA typing process, could be used to study the dynamics of R cells in liver grafts. A total of 50 frozen liver biopsies from 37 patients receiving a HLA mismatch liver were tested. Biopsies were obtained from 3 days to more than 360 days after OLT. Frozen sections of graft biopsies were stained using an immunoperoxidase technique with the proper MoAbs. In selected cases, a double immunofluorescence was also performed. Circulating R blood cells and sinusoidal cells were occasionally observed in liver biopsies obtained within 10 days after OLT and were commonly detected after 1 month. The number of sinusoidal cells continued to increase up to 6 months, as shown on serial biopsies. On the whole, R blood cells and R sinusoidal cells were detected in 86% and 82% of the biopsies, respectively. R hepatocytes and biliary cells were detected after 40 and 60 days after OLT, respectively, in 14% (hepatocytes), 8% (bile ducts), and 12% (proliferating bile ducts) of the biopsies. R hepatocytes presented as single cells or groups of few cells; their number was lower than 1% and apparently did not increase with time after OLT. In conclusion, it is possible to detect R cells in liver graft using MoAbs to specific mismatched HLA-I alleles. R sinusoidal cells start to appear after 10 days and are commonly observed after 1 month; bile duct cells and hepatocytes appear later and their number does not increase with time. Engraftment by R epithelial cells seems to be less important than previously reported.

Liver Kidney Microsomal autoantibody type 1(LKM1) directed to cytochrome P4502D6 (CYP2D6) characterises autoimmune hepatitis type-2 (AIH-2), but is also found in a proportion of chronic hepatitis C virus (HCV) infected patients, CYP2D6252 – 271 being a major B- cell autoepitope. Molecular mimicry and immunological cross-reactivity between CYP2D6252 – 271, HCV polyprotein and the infected cell protein 4 (ICP4) of herpes simplex virus type 1 (HSV-1) have been suggested as triggers for the induction of LKM1, but reactivity and cross-reactivity to the relevant sequences have not been investigated experimentally. CYP2D6252 – 271 and its viral homologues were constructed and tested by ELISA in the sera of 46 chronically infected HCV patients, 23 of whom were LKM1 positive. Reactivity to the E1 HCV and ICP4 HSV1 mimics was frequently found in HCV infected patients irrespectively of their LKM1 status; viral/self cross-reactivity (as indicated by inhibition studies), however, was present in the only 2 of the 23 LKM1 seropositive HCV patients, who possessed the HLA allotype B51. Our results indicate that in HCV infected patients virus/self cross-reactivity is dependent on a specific immunogenetic background, a finding awaiting confirmation by studies in larger series of patients.

In the past five years 956 patients (306 male and 650 female) aged 60 and older (median 76, range 60-93) were referred to our allergy clinic for suspected atopic conditions. All of them underwent skin prick testing with the most common inhalant allergens, and 318 (33%) were found positive with a wheal of at least 3 mm, men being more often positive than women: 40% (123/306) v 30% (195/650), P < 0.005. The median age of these subjects was 65 (range 60-86). The allergen profile of skin positivity was similar in men and women and was characterised by a clear prevalence of pollens (78% v 75% ) over dust mites (41% v 36%), animal danders (18% v 16%) and moulds (6% v 8% ). Rhinitis (hay fever) was the most common complaint (74%), whereas asthma was present in 33% (105/318). In 22% (70/318) of the subjects the clinical onset of atopy was at or after their 60th birthday. We conclude that atopy is spreading to old age. We need to be aware of this phenomenon so as not to underestimate the role of atopy in the respiratory conditions affecting elderly people. We will also need to draw specific guidelines for the management (allergen avoidance, pharmacological treatment, immunotherapy) of atopy in such patients, who often have important comorbidities.