Autoantibody and human leukocyte antigen profiles in children with autoimmune liver disease and their first-degree relatives.

OBJECTIVE: Familial clustering of juvenile autoimmune liver disease (AILD), including autoimmune hepatitis and autoimmune sclerosing cholangitis (ASC), is rare, despite a high prevalence of autoimmune disorders in AILD families. METHODS: To investigate this discrepancy, we measured autoantibodies diagnostic for AILD, anti-nuclear, anti-smooth muscle, anti-liver kidney microsomal type 1, anti-liver cytosol type 1, and anti-soluble liver antigen antibodies, and human leukocyte antigen profiles in 31 patients and 65 of their first-degree relatives (FDR). The autoantibody profile was compared with that of 42 healthy subjects (HS). RESULTS: Autoantibodies were detected in 71% (22/31) patients. Anti-nuclear antibody or anti-smooth muscle antibody were present in 4/65 FDR (6.2%). HS were negative for all autoantibodies. The frequencies of homozygous HLA DRB1*0301 (DR3) genes and haplotype A1-B8-DR3 were higher in the patients (25% and 43%) than in FDR (9% and 27%) and HS (0% and 16%). The frequencies of disease-protective genes DR4 and/or DR15 were lower in the patients (25%) than in FDR (42%) and HS (42%). Only 1 family contained 2 patients with AILD, 1 with ASC and 1 with primary sclerosing cholangitis. Both patients possessed A1-B8-DR3 genes, the ASC being homozygous and the primary sclerosing cholangitis heterozygous. Six FDR had nonhepatic autoimmune disorders, none being autoantibody positive. CONCLUSIONS: Homozygosity for DR3 plays a major role in the predisposition to juvenile AILD. Diagnostic autoantibodies for AILD are rare among patients' FDR and not linked to clinical manifestation of AILD.

Severity of the compensatory anti-inflammatory response determined by monocyte HLA-DR expression may assist outcome prediction in cirrhosis.

PURPOSE: To investigate variations in expression of the monocyte antigen presentation molecule HLA-DR in cirrhosis. METHODS: HLA-DR expression was measured by flow cytometry in 100 patients within 48 h of admission and repeated a week later in 21 patients admitted to ICU. IL-10, TNF-α and IFN-γ secretion in response to lipopolysaccharide and recall antigens were measured by enzyme-linked immunosorbent spot (ELISPOT) assay in 12 patients (7 with clinical immunoparesis, 5 stable). RESULTS: HLA-DR level was 71% in stable patients, 53% with single organ dysfunction and 34% with multiple organ failure (p < 0.02). Within these groups, no significant differences in admission HLA-DR were seen between survivors and non-survivors. HLA-DR expression less than 40% predicted 90-day mortality with a specificity of 80% and sensitivity of 59% [area under the receiver operator curve (AUROC) 0.76]. HLA-DR less than 40% was an independent predictor of prognosis in multivariate analysis with a relative risk of 2.35 (p = 0.04), although sequential organ failure assessment score (SOFA) score displaced HLA-DR when included. In those admitted to intensive care failure to increase HLA-DR expression was predictive of death within 30 days (risk ratio 6.9, p = 0.007). Follow-up values predicted outcome with similar accuracy to acute physiology and chronic health evaluation II (APACHE II)/SOFA scores (AUROC 0.88). Response to endotoxin and recall antigen was characterised by an anti-inflammatory cytokine secretion profile, and was associated with impairment in recall antigen presentation capacity. CONCLUSIONS: HLA-DR expression less than 40% and a failure of recovery predict poor outcome in decompensated cirrhosis, but overall prognostic power remains inferior to conventional markers. Ex vivo experiments demonstrate reduced Th1 response to antigenic stimulation and an exaggerated counter-inflammatory cytokine secretion profile.

The management of treatment-resistant biliary calculi using percutaneous endourologic techniques.

BACKGROUND: Complicated choledocholithiasis cannot always be managed by standard surgical, radiologic or endoscopic methods. One additional approach is to use percutaneous techniques developed by endourologists to treat renal calculi. In this report, we present our experience over the past 10 years with this novel approach. METHODS: We conducted a retrospective review of all patients who underwent percutaneous, endoscopic treatment of biliary calculi at our institution between January 1997 and August 2007. Primary outcomes of interest were symptom- and stone-free rates, length of stay in hospital and complications. RESULTS: Nineteen patients underwent 21 percutaneous treatments for biliary calculi. All were dependent on external drainage for symptom control. The primary indications for treatment were cholangitis, retained stone, biliary colic and jaundice. Seventeen patients (89.5%) had failed prior endoscopic retrograde cholangiopancreatography (ERCP) or open attempts at treatment. The 2 remaining patients (10.5%) were deemed unfit for a general anesthetic. Patients had experienced a mean of 1.8 (standard deviation [SD] 1.0) prior failed attempts at stone removal. We used several treatment modalities, including holmium:yttrium-aluminum-garnet laser (61.9%), electrohydraulic lithotripter (19.0%), ultrasound (9.5%), basket extraction (9.5%) and balloon dilatation of the ampulla (19.0%). Overall, treatment led to successful removal of the biliary drainage tube in 94.7% of patients and 76.2% were stone-free. We performed cholangiograms an average of 21.8 (SD 13.7) days after treatment. The average length of stay in hospital was 1.9 (SD 1.1) days. One patient experienced a perioperative acute coronary syndrome and another experienced prolonged biliary drainage. Both had successful endoscopic treatment of their calculi. There were no cases of treatment-related sepsis, and we observed no other complications. CONCLUSION: Biliary calculi may be successfully treated using standard endourologic methods with high stone-free rates. This technique is generally well-tolerated even among high-risk patients.

Antimitochondrial antibodies of immunoglobulin G3 subclass are associated with a more severe disease course in primary biliary cirrhosis.

BACKGROUND/AIMS: Primary biliary cirrhosis (PBC) is characterised by the presence of immunoglobulin (Ig) G antimitochondrial antibodies (AMA), which are routinely detected by indirect immunofluorescence (IFL) using composite rodent tissue substrate. The IgG subclass distribution and clinical significance of IFL-detected AMA in patients with PBC have not been previously studied in detail. METHODS: We have examined IgG subclass-specific AMA detected by IFL on rodent liver, kidney and stomach tissue substrate using affinity-purified IgG subclass monospecific antisera as revealing reagents in 95 AMA-positive PBC patients from Greece. RESULTS: AMA of any of the IgG1, IgG2 or IgG3 subclasses were present in 89/95 (93.7%) patients. Among those 89, 55 (61.8%) had IgG1, 2, 3 AMA positivity; eight (9%) had IgG1, 2; seven (7.9%) had IgG2, 3; eight (9%) had IgG1, 3; nine (10.1%) had IgG1 subclass and two (2.2%) single IgG3 AMA reactivity. IgG4 AMA was absent. IgG3 titres were higher than IgG2 and IgG1 (P<0.001) and IgG1 higher than IgG2 (P<0.001). IgG3 AMA-positive patients had a histologically more advanced disease (P<0.01) and were more frequently cirrhotic compared with those who were negative (P<0.01). There was a positive correlation between AMA IgG3 titre and Mayo risk score (r=0.55, P=0.009, Spearman's correlation). CONCLUSIONS: Our findings suggest that AMA are not restricted to a specific IgG subclass. AMA of the IgG3 subclass are associated with a more severe disease course, possibly reflecting the peculiar ability of this isotype to engage mediators of damage.

Reduced monocyte HLA-DR expression: a novel biomarker of disease severity and outcome in acetaminophen-induced acute liver failure.

Acute liver failure (ALF) shares striking similarities with septic shock where a decrease in HLA-DR expression on monocytes is associated with disease severity and predicts outcome. We investigated monocyte HLA-DR expression in ALF in relation to inflammatory mediator levels and clinical outcome. Monocyte HLA-DR expression was determined in 50 patients with acetaminophen-induced ALF (AALF) and 20 non-acetaminophen-induced ALF (NAALF). AALF patients were divided into dead/transplanted (AALF-NS, n = 26) and spontaneous survivors (AALF-S, n = 24). Fifty patients with chronic liver disease (CLD) and 50 healthy volunteers served as controls. Monocyte HLA-DR expression was determined by double-color flow-cytometry with monoclonal antibodies detecting HLA-DR and monocyte specific CD14. Serum levels of interleukin (IL) -4, -6, -10, tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma were concomitantly measured by ELISA. Compared to healthy volunteers (75%) and CLD (67%) monocyte HLA-DR percentage expression was lower in AALF (15%, P < .001) and NAALF (22 %, P < .001). Compared to AALF-S, AALF-NS had lower monocyte HLA-DR % (11% vs. 36%, P < .001) and higher levels of IL-4, IL-6, IL-10 and TNF-alpha (P < .001). HLA-DR percentage negatively correlated with INR, blood lactate, pH and levels of encephalopathy (r = -0.8 to -0.5, P < .01), IL-10 (r = -0.8, P < .0001), TNF-alpha (r = -0.4, P = .02). HLA-DR percentage level

Visual-spatial ability, learning modality and surgical knot tying.

INTRODUCTION: The ability to mentally rotate an object in 3 dimensions has been shown with an individual's score on the Vandenberg and Kuse Mental Rotations Test. The was to determine whether this Mental Rotations Test could be used to predict performance complex surgical skill - the tying of a 1-handed surgical reef knot. In addition, we learning a spatially complex surgical skill could be achieved more effectively via a computer-based selfdirected learning approach than with a didactic lecture-based teaching method. METHODS: preclerkship medical students at the University of Western Ontario were randomized into computer-based self-directed learning group and a didactic lecture-style learning group. administration of the Mental Rotations Test, the students were taught how to tie a reef knot via the learning modality assigned to their respective group. RESULTS: Students Mental Rotations Test scores were able to tie more surgical knots in the allocated time Students learning how to tie the surgical knot via the computer-based self-directed showed improvement on their knot tying abilities more rapidly than their didactically trained colleagues. CONCLUSION: The ability to mentally rotate an object in 3 dimensions played an important initial learning of a spatially complex surgical technical skill. Our data demonstrated learning was as effective and more practical than traditional lecture-based learning.

The effect of anti-lymphocyte serum on subpopulations of blood and tissue leucocytes: possible supplementary mechanisms for suppression of rejection and the development of opportunistic infections.

Xenogeneic anti-lymphocyte serum (ALS) remains a major reagent for immunosuppression in clinical practice, but mechanisms of action and risks of opportunistic infection have not been considered in the context of innate immunity and its role in immune responsiveness. Rabbit anti rat ALS was administered intraperitoneally. Blood was taken for flow cytometry to establish absolute counts of leucocyte subsets. Tissues were harvested for immunohistology to evaluate interstitial dendritic cells and tissue macrophages. At day 2 of ALS therapy, T cells are completely depleted, as anticipated. B cells are undiminished and form approximately 90% of blood leucocytes. Monocytes and natural killer (NK) cells are substantially (approximately 80%), but not completely, depleted, and there is a trend for diminished numbers of putative dendritic cells. Neither interstitial dendritic cells nor tissue macrophages in heart are affected. The results at day 7 were very similar to day 2. Substantial depletion of blood monocytes and NK cells might attenuate the innate immune system, and represent a possible supplementary mechanism (in addition to T cell depletion) for suppression of rejection. It might be of particular importance in reducing defences against infections. Monitoring these parameters could be of clinical value.

Microbial mimics are major targets of crossreactivity with human pyruvate dehydrogenase in primary biliary cirrhosis.

BACKGROUND/AIMS: Previous studies on patients with primary biliary cirrhosis (PBC) have shown extensive cross-reactivity between the dominant B- and T-cell epitopes of human pyruvate dehydrogenase complex-E2 (PDC-E2), and microbial mimics. Such observations have suggested microbial infection as having a role in the induction of anti-mitochondrial antibodies, through a mechanism of molecular mimicry. However the biological significance of these cross-reactivities is questionable, because PDC-E2 is so highly conserved among various species. METHODS: Interrogating protein databases, ten non-PDC-E2 microbial sequences with high degree of similarity to PDC-E2(212-226) were found in Escherichia coli (6), Helicobacter pylori, Pseudomonas aeruginosa, Cytomegalovirus, and Haemophilus influenzae. We report on a study testing reactivity and competitive cross-reactivity against these respective peptides, and in some cases the parent protein, using sera from 55 patients with PBC, compared to reactivity of 190 pathological and 28 healthy controls. RESULTS: Cross-reactivity to E. coli mimics was commonly seen in PBC, and in a subset of pathological controls except where there was no evidence of urinary tract infection and correlated with anti-mitochondrial reactivity. CONCLUSIONS: E. coli/PDC-E2 cross-reactive immunity characterizes primary biliary cirrhosis; the large number of E. coli immunogenic mimics may account for the dominance of the major PDC-E2 autoepitope.

Neonatal liver disease associated with placental transfer of anti-mitochondrial antibodies.

BACKGROUND: Anti-mitochondrial antibody is the diagnostic hallmark of primary biliary cirrhosis. Its role in the aetiology of primary biliary cirrhosis is controversial. METHODS: Two cases of neonatal hepatitis seropositive for anti-mitochondrial antibody are described. Anti-mitochondrial antibody Ig isotype and epitopic specificity were investigated by immunofluorescence and enzyme immunoassays. RESULTS: In both infants anti-mitochondrial antibody was of the G class, mainly G1 and G3 subclasses, and reacted with two synthetic peptides reproducing major M2 epitopicregions: innerlipoyl domain pyruvate dehydrogenase complex (PDC)-E2(162-176) and PDC-E3 binding protein (PDC-E3BP)86-100. One infant also reacted with outer lipoyl domain PDC-E2(35-49), and 2-oxoglutarate dehydrogenase complex (OGDC)-E2(99-113). An identical pattern of reactivity was present in their mothers, indicating the maternal origin of the antibodies. Anti-mitochondrial antibody disappeared in the infants with the disappearance of the liver pathology. CONCLUSIONS: The simultaneous disappearance of hepatitis and anti-mitochondrial antibody in the infants suggests a possible causal link between the two.