Clinical and pathological features associated with HSP70, ENO1 and RNH1 autoantibodies in patients with CCA.

<p>*When the sum of subset numbers does not match patient totals, data were missing or unavailable.</p

Two-dimensional (2D) gel electrophoresis and western blotting.

<p>MMNK1 and CCA cell lines (M055, M214 and M139) lysates were separated by 2D electrophoresis and stained with coomassie brilliant blue (panels A, C, E and G). Parallel gels were transferred to PVDF and incubated with pooled plasma of patients with CCA (n = 10) and incubated with secondary anti-human IgG (panels B, D, F and H). Matching IgG autoantibodies spots were numbered 1–14 in both CBB gels and western blot membranes. Duplicate experiments were performed. Spot numbers 1, 11 and 13 are RNH1, ENO1 and HSP70, respectively.</p

Western blotting analysis of HSP70 and RNH1 autoantibodies.

<p>Panel A) Recombinant human HSP70 was incubated with plasma of healthy individuals (lane 1–5) and patients with CCA (lanes 6–12). Immunoreactive bands were detected in 5/7 patients and none of the healthy controls. Panel B) Recombinant human RNH1 was incubated with plasma of healthy individuals (lane 1–5) and patients with CCA (lanes 6–12). Immunoreactive bands were detected in 6/7 patients and was weak in controls. Recombinant HSP70 and RNH1 proteins were probed with pooled healthy plasma served as negative controls (−) and pooled plasma from patients with CCA served as positive controls (+). HSP70 and RNH1 proteins were also demonstrated by staining with coomassie brilliant blue (CBB). PH  =  healthy individuals, CCA  =  cholangiocarcinoma.</p

Diagnostic performance (sensitivity, specificity, predictive values) of the possible combination of autoantibodies against HSP70, RNH1 and ENO1 in patients with CCA <i>vs</i> healthy controls.

<p>PPV  =  positive predictive value, NPV  =  negative predictive value.</p

Identification of autoantigens reacting against plasma samples from patients with CCA using M139 cell line-derived proteins.

<p>Identification of autoantigens reacting against plasma samples from patients with CCA using M139 cell line-derived proteins.</p

Plasma levels of HSP70, ENO1 and RNH1 autoantibodies by ELISA.

<p>Plasma levels of autoantibodies against HSP70, ENO1 and RNH1 were determined in clinical groups including cholangitis (n = 12), CCA (n = 31) and healthy controls (n = 23). Panel A) HSP70 autoantibody level is significantly higher in CCA compared to cholangitis and healthy controls. Panel B) Receiver operating characteristic (ROC) curves of HSP70 autoantibody in patients with CCA <i>vs</i> healthy controls. Panel C) ENO1 and Panel E) RNH1 autoantibodies are significantly higher in CCA than healthy controls. Panels D and F) ROC curves of ENO1 and RNH1 autoantibodies in patients with CCA <i>vs</i> healthy controls. Data are illustrated as mean ± SD using student’s <i>t</i> test or non-parametric Mann-Whitney test; *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001.</p

Identification of autoantigens reacting against plasma samples from patients with CCA using MMNK1 cell line-derived proteins.

<p>Identification of autoantigens reacting against plasma samples from patients with CCA using MMNK1 cell line-derived proteins.</p

Diagnostic performance of autoantibodies against HSP70, RNH1 and ENO1 in pairwise comparisons between CCA, cholangitis, or healthy controls.

<p>AUC  =  area under the curve, 95%CI = 95% confidence interval (lower-upper).</p

Pancreatic function in control and MSG-treated groups as measured by (A) insulin levels at 1, 3, 6, and 9 months (mean ± SEM) and (B) oral glucose tolerance test (OGTT) at 9 months(mean ± SD).

<p>Pancreatic function in control and MSG-treated groups as measured by (A) insulin levels at 1, 3, 6, and 9 months (mean ± SEM) and (B) oral glucose tolerance test (OGTT) at 9 months(mean ± SD).</p

Islet size (um²) distribution in control and MSG-treated groups at 1, 3, 6, and 9 months.

<p>Islet size (um²) distribution in control and MSG-treated groups at 1, 3, 6, and 9 months.</p