Protease inhibitors-based therapy induces acquired spherocytic-like anaemia and ineffective erythropoiesis in chronic hepatitis C virus patients.

BACKGROUND & AIMS: The addition of protease inhibitors, boceprevir (BOC) or telaprevir (TRV), to peg-interferon and ribavirin (PR) increases the incidence of anaemia in patients with chronic hepatitis C virus (HCV) infection. Although genetic variants in inosine triphosphatase (ITPA) gene have been linked to the haemolytic anaemia induced by PR, the mechanism sustaining severe anaemia during triple therapy is still unknown. This study aims to elucidate the molecular mechanisms underlying anaemia in chronic HCV patients with combined therapy. METHODS: We studied 59 patients with chronic HCV genotype-1: 29 treated with TRV/PR and 30 with BOC/PR. We evaluated biochemical and haematological parameters, red cell index at baseline, 4, 12, 16 and 24 weeks of treatment; in a subgroup, we performed functional studies: osmotic fragility, red cell membrane protein separation, mass spectrometry analysis, quantification of erythroid microparticles release. IL28B and ITPA polymorphisms were also evaluated. RESULTS: We found early acute normochromic normocytic haemolytic anaemia (4-8 weeks) followed by a late macrocytic hypo-regenerative anaemia with inappropriate low reticulocyte count (12-24 weeks). Studies on red cells revealed: (i) presence of spherocytes; (ii) increased osmotic fragility; (iii) abnormalities in red cell membrane protein composition; (iv) reduced membrane-cytoskeleton stability; (v) increased release of erythroid microparticles. ITPA polymorphisms impacted only the early phase of anaemia. CONCLUSIONS: The bimodal pattern of anaemia in chronic HCV patients on triple therapy might be because of acquired spherocytic-like anaemia in the early phase, followed by hyporegenerative anaemia, most likely related to the combined effects of PR and TRV or BOC on erythropoiesis.

There Is No Association between Coeliac Disease and Autoimmune Pancreatitis.

Autoimmune pancreatitis (AIP) is a rare disorder whose association with coeliac disease (CD) has never been investigated, although CD patients display a high prevalence of both endocrine and exocrine pancreatic affections. Therefore, we sought to evaluate the frequency of CD in patients with AIP and in further medical pancreatic disorders. The screening for CD was carried out through the detection of tissue transglutaminase (tTG) autoantibodies in sera of patients retrospectively enrolled and divided in four groups: AIP, chronic pancreatitis, chronic asymptomatic pancreatic hyperenzymemia (CAPH), and control subjects with functional dyspepsia. The search for anti-endomysium autoantibodies was performed in those cases with borderline or positive anti-tTG values. Duodenal biopsy was offered to all cases showing positive results. One patient out of 72 (1.4%) with AIP had already been diagnosed with CD and was following a gluten-free diet, while one case out of 71 (1.4%) with chronic pancreatitis and one out of 92 (1.1%) control subjects were diagnosed with de novo CD. No cases of CD were detected in the CAPH group. By contrast, a high prevalence of cases with ulcerative colitis was found in the AIP group (13.8%). Despite a mutual association between CD and several autoimmune disorders, our data do not support the serologic screening for CD in AIP. Further studies will clarify the usefulness of CD serologic screening in other pancreatic disorders.

Role of Amylase-α2A Autoantibodies in the Diagnosis of Autoimmune Pancreatitis.

OBJECTIVES: Several serological markers have been reported in autoimmune pancreatitis (AIP) patients. However, only serum IgG4 (sIgG4) is available in clinical practice for AIP diagnosis. Antiamylase α antibodies (AMY-α Abs) have been proposed to diagnose AIP. This study evaluates the utility of AMY-α Abs and sIgG4 for AIP diagnosis. METHODS: Twenty-five AIP patients, 84 disease control groups (31 chronic pancreatitis, 30 acute pancreatitis, 23 pancreatic adenocarcinoma), and 59 healthy donors were prospectively studied. The AMY-α Abs were determined by homemade enzyme-linked immunosorbent assay and sIgG4 by nephelometry. RESULTS: Increased sIgG4 were detected to be present in 52% of AIP, 5% in control groups, and 0% in healthy donors, and AMY-α Abs, respectively, in 76%, 36%, and 2%. sIgG4 was elevated in 92% of the 13 patients with type 1 AIP, but in none of 3 with type 2 and of 8 with not otherwise specified AIP. The AMY-α Abs were present in 79%, 67%, and 75% of type 1, type 2, and not otherwise specified AIP, respectively. Sensitivity and specificity of AMY-α Abs were 76% and 78%, and of sIgG4 50% and 94%. By combining the 2 serological markers, sensitivity was 41%, and specificity was 99%. CONCLUSIONS: The AMY-α Abs may help to diagnosis of AIP and to differentiate AIP subtypes.

Functional role of p38 mitogen activated protein kinase in platelet activation induced by a thromboxane A2 analogue and by 8-iso-prostaglandin F2alpha.

We investigated similarities in the signaling pathways elicited by the F2 isoprostane 8-iso-PGF2alpha and by low doses of U46619 to induce platelet activation. Both 0.01-0.1 micromol/L U46619 and 0.01-1 micromol/L 8-iso-PGF2alpha triggered shape change and filopodia extension, as well as adhesion to immobilized fibrinogen of washed platelets. At these doses the two platelet agonists failed to trigger secretion and aggregation, which were however induced by higher doses of U46619 (0.1-1 micromol/L). SB203580 (1-10 micromol/L), a specific inhibitor of the p38 mitogen activated protein (MAP) kinase blunted platelet shape change and adhesion induced by 0.05-1 micromol/L 8-iso-PGF2alpha and by 0.01 micromol/L U46619. These platelet responses were also inhibited by 20 micromol/L cytochalasin D, an inhibitor of actin polymerization, and 50 micromol/L piceatannol, an inhibitor of the Syk tyrosine kinases. Both 8-iso-PGF2alpha and U46619-induced p38 MAP kinase phosphorylation in suspended platelets and this was inhibited by piceatannol, indicating that Syk activation occurs upstream of p38 MAP kinase phosphorylation. These findings suggest that the signaling pathway triggered by both 8-iso-PGFalpha and low concentrations of U46619 to induce platelet adhesion and shape change implicates Syk, the p38 MAP kinase, and actin polymerization.

Altered intestinal function precedes the appearance of bacterial DNA in serum and ascites in patients with cirrhosis: a pilot study.

OBJECTIVES: Bacterial translocation seems to precede the occurrence of overt bacterial infection in patients with cirrhosis. The presence of bacterial DNA in blood and ascites correlates with bacterial translocation and is frequent in patients with advanced cirrhosis without overt infection. Our aim was to search for bacterial DNA in patients with cirrhosis both with and without ascites, and to study its correlation with abnormal intestinal motility or permeability and the presence of bacterial overgrowth. METHODS: Blood and ascites samples were obtained on day 1, and blood samples were taken twice a day for the following 3 days. Bacterial DNA was assayed by polymerase chain reaction using universal primers for rRNA 16 s. Oro-caecal transit time and bacterial overgrowth were assessed with Lactulose H(2) breath testing. Intestinal permeability was assessed by determining urinary lactulose and mannitol excretion with high performance liquid chromatography. RESULTS: We studied seven patients (six were male, age range was 42-78 years). Aetiology was alcohol in four, HCV in two, HBV in one; ascites was present in four and Child-Pugh grade was A in four and B in three. All patients had increased intestinal permeability, six had decreased transit time and one had bacterial overgrowth. In only one patient (with ascites), polymerase chain reaction was positive for bacterial DNA both in ascites and serum for all 4 days on which samples were taken. CONCLUSION: Increased intestinal permeability and abnormal motility were frequent without evidence of bacterial translocation in cirrhosis even without ascites. They are likely to be facilitators for bacterial translocation and thus precede it.