Irena Hrstić

NOD2/CARD15 mutations in Croatian patients with Crohn's disease : prevalence and genotype-phenotype relationship

Background Crohns disease (CD) is a chronic inflammatory disorder of the gastrointestinal tract with variations in localization and behaviour. Mutations in the NOD2/CARD15 gene on chromosome 16q have been implicated in the pathogenesis of the disease and three main sequence variants, all single nucleotide polymorphisms (SNPs), have been identified in North American and European populations. Aims and methods As no data exist in the Croatian population, we consecutively collected a cohort of 136 CD patients and 91 healthy controls to determine the prevalence of NOD2/CARD15 mutations and their association with phenotypic expression of the disease. All patients and controls were genotyped for Arg702Trp (Hugot SNP8), Gly908Arg (Hugot SNP12), and Leu1007fsinsC (Hugot SNP13) and allele frequencies were compared between the Crohns patients and controls. The correlation of NOD2/CARD15 genotypes with the phenotypic expression of Crohns disease was further assessed by logistic regression analysis. Results NOD2/CARD15 variants were found in 38/136 CD patients (27.9%) compared to 10/91 (10.9%) healthy controls (P=0.0022). Allele frequencies in patients with CD were 13.97%, 4.4% and 11.76%, respectively, for SNP8, 12 and 13, compared to 5.49%, 1.12% and 4.40% in controls (P=0.041, P=0.162, P=0.055). Six CD patients carried double mutations and, remarkably, we identified two homozygous mutants amongst the healthy control group. Surgery over the course of the disease and a younger age at onset of the disease were significantly more frequent in patients who were carriers of NOD2/CARD15 mutations. Conclusions This report on NOD2/CARD15 mutations in Croatian patients with CD demonstrates that this gene is also implicated in susceptibility to CD in the Croatian population. Phenotypic association showed a younger age at diagnosis and a higher need for surgery in patients carrying NOD2/CARD15 mutations. However, the prevalence is somewhat lower compared to other reports, likely due to a more prominent colonic inflammation.

Endoscopic ultrasound elastography as a method for screening the patients with suspected primary sclerosing cholangitis.

Objectives Currently, magnetic resonance cholangiography is being used for establishing the diagnosis of primary sclerosing cholangitis, whereas endoscopic retrograde cholangiography is reserved for therapeutic interventions. The aim of this study was to determine the role of endoscopic ultrasound elastography in the detection of inflammatory and fibrotic lesions of the common bile duct. Methods Linear endoscopic ultrasound elastography of the common bile duct was performed in 41 patients. The patients were divided into two groups: disease group (20 patients with both, primary sclerosing cholangitis and inflammatory bowel disease) and control group (21 patients). Main outcome measurements were diameter, wall thickness and wall qualitative Elasto Score of the common bile duct. Results The disease group consisted of nine females and 11 males, whereas the control group consisted of 13 females and eight males, with no sex differences observed (χ2u2009=u20090.6, d.f.u2009=u20091, Pu2009=u20090.443). There was no significant difference in the diameter of common bile duct between the studied groups: 4.67±1.83u2009mm in the disease group and 5.88±2.47u2009mm in controls (tu2009=u2009−1.77, d.f. u2009=u200939, Pu2009=u20090.085). Hard or mixed Elasto Score was found in 16 patients and four controls, being significantly different compared with the soft Elasto Score found in four patients and 17 controls (χ2u2009=u20091.8, d.f. u2009=u20091, P<0.001). A significant difference was observed in the common bile duct wall thickness: 0.89±0.59u2009mm in the disease group and 0.39±0.14u2009mm in controls (tu2009=u20093.75, d.f. u2009=u200939, P<0.001). Conclusion Endoscopic ultrasound elastography might be a useful noninvasive method in diagnosing primary sclerosing cholangitis.

Hypervariable Region 1 of Hepatitis C Virus Genome and Response to Interferon Therapy

Abstract The relationship between the complexity of the hypervariable region 1 (HVR1) quasispecies of hepatitis C virus (HCV) and responsiveness to interferon-α (IFN) therapy was studied in patients with chronic hepatitis C. Twelve HCV-RNA-positive patients were treated daily with high dose IFN and ribavirin for 4 weeks, and then with IFN 3 MIU (Million International Units) TIW (three times per week) and ribavirin for 6 months. The HVR1 quasispecies complexity was analyzed by nested polymerase chain reaction-mediated single-strand conformation polymorphism (SSCP). The baseline HCV-RNA levels in the study group ranged from 106 to 107 copies/ml. All patients exhibited HCV genotype 1 b. Initial SSCP analysis revealed four (33.3%) patients with a low complexity pattern (SSCP bands ≤4) and eight (66.6%) patients with high complexity pattern (SSCP bands >4). After 4 weeks of IFN therapy, one patient became HCV negative, and among those remaining positive, the HCV-RNA levels decreased by 2 to 3 logs and the number of SSCP decreased by 2 to 3 bands per sample. After 6 months of IFN therapy, five (41.7%) patients became HCV-RNA-negative. Seven (58.3%) patients did not respond to IFN therapy with sustained viral load from 103 to 105 copies/ml, and high complexity SSCP patterns. Our data support the HVR quasispecies complexity to be an independent predictive factor for IFN responsiveness in patients infected with HCV.

Dynamics of serum hepatitis C virus load and quasispecies complexity during antiviral therapy in patients with chronic hepatitis C.

BACKGROUNDnHepatitis C virus (HCV) infection is a dynamic process during which viral genetic variants continuously develop as a result of the virus adaptation to the hosts immune system. The level of viremia and the complexity of the hypervariable region 1 (HVR 1) quasispecies of hepatitis C virus during antiviral therapy reflect the dynamic balance between the viral and host components in response to therapy.nnnOBJECTIVEnThe aim of the study was to evaluate the dynamics of HCV viremia and the complexity of the HVR 1 quasispecies during the induction phase of a triple combination therapy regimen in nonresponders to earlier anti-HCV treatment.nnnSTUDY DESIGNnTen patients with chronic hepatitis C undergoing antiviral combination therapy with interferon-alpha, ribavirin, and amantadine were studied. The serum HCV RNA level was monitored by a quantitative RT-PCR assay up to 3 months after start of treatment. The HVR 1 quasispecies complexity was analysed by an in house nested RT-PCR mediated single-strand conformation polymorphism (SSCP) assay.nnnRESULTSnBaseline serum HCV RNA levels ranged from 1.94x10(6) to 5.53x10(6) copies/ml. In all patients, HCV subtype 1b was found. At the start of therapy, the SSCP assay revealed a high complexity pattern (at least six SSCP bands) in all patients. None of the patients responded within 4 weeks of treatment, however, the serum HCV RNA level decreased by one to two logs in eight patients. At week 4 after start of treatment, there was a decrease of SSCP bands in five patients. In four patients, SSCP bands remained unchanged and in one patient SSCP bands increased. At month 3 after start of treatment, serum HCV RNA was not detectable in one patient.nnnCONCLUSIONnBecause of the low number of patients involved in this study, prediction of therapeutical success based on the quasispecies complexity was not possible. Larger studies are urgently needed.

Metabolic syndrome and non-alcoholic fatty liver disease after liver or kidney transplantation

Transplantation is a definitive treatment option for patients with end‐stage liver disease, and for some patients with acute liver failure, hepatocellular carcinoma or end‐stage renal disease. Long‐term post‐transplantation complications have become an important medical issue, and cardiovascular diseases (CVD) are now the leading cause of mortality in liver or kidney transplant recipients. The increased prevalence of metabolic syndrome (MS) likely plays a role in the high incidence of post‐transplantation CVD. MS and its hepatic manifestation, non‐alcoholic fatty liver disease (NAFLD), are prevalent among the general population and in pre‐ and post‐transplantation settings. MS components are associated with recurrent or de novo NAFLD in transplant recipients, potentially influencing post‐transplantation survival. Moreover, recent data reveal an important association between NAFLD and risk of incident of chronic kidney disease (CKD). Therefore, NAFLD identification could represent an additional clinical feature for improving the stratification of liver and kidney transplant recipients with regards to risks of CVD, CKD and renal allograft dysfunction. All MS components are potentially modifiable; therefore, it is crucial that hepatologists, nephrologists and primary care physicians become more engaged in managing post‐transplantation metabolic complications. The present review discusses the recent clinical evidence regarding the importance of MS and its components after liver and kidney transplantation, as well as the link between MS and NAFLD after liver and kidney transplantation.

Pegylated interferon for treatment of chronic hepatitis C in hemodialysis patients in Croatia

Background and Aims: Hepatitis C virus (HCV) infection is a frequent complication among long-term dialysis patients. The aim of the present study was to evaluate the efficacy and side effects of pegylated interferon-α2a (PEG-IFN-α2a) treatment in hemodialysis patients. Methods: We retrospectively reviewed charts of 16 HCV-RNA-positive hemodialysis patients. Results: There were 11 male and 5 female patients treated with dialysis for 6–28 years. Twelve patients had HCV genotype 1b, 2 patients had 3a, and 1 patient had genotype 2a. Although only 10 out of 16 patients completed 48 weeks of treatment, early virological response and end-of-treatment virological response were achieved in 9 and 13 patients, respectively. Sustained virological response was recorded in 9 patients. The most common side effect was anemia. A flu-like syndrome was documented in 6, myalgia in 4, and arthralgia in 5 patients. Rectorrhagia, endocarditis and severe cough were recorded in 1 patient each. Nine patients received a renal transplant, and all 6 responders remained HCV-RNA-negative. Conclusions: PEG-IFN-α2a has limited efficacy in dialysis patients. A significant proportion of patients discontinued treatment because of side effects. Additional studies with long-term follow-up are needed to determine the optimal treatment of HCV infection in the dialysis population.

Non-alcoholic fatty liver disease and colorectal cancer

As a significant cause of cancer death worldwide, colorectal cancer (CRC) is still one of the most common cancers in the world. The most efficient strategies to reduce CRC incidence include identifying risk factors for CRC and performing a preventive colonoscopy in high-risk populations. Some well-established risk factors for CRC development include hereditary syndromes and inflammatory bowel disease. Of note, in recent years, attention has been given to new evidence indicating that more than 75%–95% of CRC occurs in individuals with little or no genetic risk. For these individuals, the risk for CRC is associated with their lifestyle and dietary factors, including central obesity, overweight and physical inactivity. Recently, evidence demonstrated a connection between non-alcoholic fatty liver disease (NAFLD) and CRC. Insulin resistance and metabolic syndrome (MetS) are common risks that NAFLD and colorectal neoplasms share. The incidence of NAFLD is increasing in parallel with an increasing prevalence of MetS and obesity. Consequently, the question arises: will the incidence of CRC increase together with this dramatic increase in obesity, MetS and ultimately NAFLD prevalence? Recent studies of adenomatous polyps, CRC and NAFLD are discussed in this manuscript.

Chronic Budd-Chiari syndrome as a rare complication of Crohn's disease: a case report.

The Budd–Chiari syndrome (BCS) is characterized by hepatic venous outflow obstruction involving the hepatic veins, inferior vena cava, or both. BCS has occasionally been reported in the literature as a very rare complication of ulcerative colitis. However, association of Crohns disease (CD) and BCS is extremely rare with only a single case reported in the world literature to date. We report a case of a young woman with chronically active, therapy-resistant CD who developed massive ascites, elevation of liver enzymes, and coagulopathy in the course of her disease. She was subsequently diagnosed with BCS for which a successful liver transplantation was performed. Chronically active therapy resistant CD and methylenetetrahydrofolate reductase gene mutation have been identified as possible risk factors for development of BCS in this patient.

Nonalcoholic fatty liver disease and liver transplantation - Where do we stand?

Nonalcoholic fatty liver disease/nonalcoholic steatohepatitis (NAFLD/NASH) is a challenging and multisystem disease that has a high socioeconomic impact. NAFLD/NASH is a main cause of macrovesicular steatosis and has multiple impacts on liver transplantation (LT), on patients on the waiting list for transplant, on post-transplant setting as well as on organ donors. Current data indicate new trends in the area of chronic liver disease. Due to the increased incidence of metabolic syndrome (MetS) and its components, NASH cirrhosis and hepatocellular carcinoma caused by NASH will soon become a major indication for LT. Furthermore, due to an increasing incidence of MetS and, consequently, NAFLD, there will be more steatotic donor livers and less high quality organs available for LT, in addition to a lack of available liver allografts. Patients who have NASH and are candidates for LT have multiple comorbidities and are unique LT candidates. Finally, we discuss long-term grafts and patient survival after LT, the recurrence of NASH and NASH appearing de novo after transplantation. In addition, we suggest topics and areas that require more research for improving the health care of this increasing patient population.

Endoscopic Ultrasound Elastography in Inflammatory Bowel Disease

The diagnosis of inflammatory bowel disease (IBD) is based on clinical, endoscopic, radiologic and histologic criteria1. There are two main IBD phenotypes – Crohns disease (CD) and ulcerative colitis (UC). In some circumstances, especially when disease extension is restricted to the colon or in cases of acute severe pancolitis, recognition of specific IBD phenotype is very difficult. Recognition of the exact IBD phenotype is essential for guiding therapeutic decisions and detection of complications that warrant treatment. Endoscopic examination is the mainstay in the diagnosis of IBD. Endoscopic appearance (distribution and shape of lesions) helps to differentiate CD from UC in most cases. Pathohistologic analysis confirms the elements of chronic inflammation but it is frequently not diagnostic. Patients with UC may have atypical histological features such as microscopic inflammation of the ileum, patchiness of inflammation and rectal sparing at the time of diagnosis prompting physicians to make the diagnosis of CD in UC cases. Other endoscopic findings such as cobble stoning, segmental colitis, ileal stenosis and ulceration, perianal disease and pathologically confirmed multiple granulomas in the small bowel or colon strongly suggest a diagnosis of CD. The progress in genetics, serological markers and imaging studies will lead to more reliable determination of exact IBD phenotype in the future2. In the meantime, it is reasonable to explore other diagnostic options for better differentiation between different IBD phenotypes. We think that endoscopic ultrasound (EUS) elastography is a promising method to achieve this goal, picture 1 and 2. It is a new endoscopic procedure which can differentiate the stiffness of normal and pathological tissue by ultrasound. This finding is based on B-mode scanning during compressions3. There are some data on elastography applied on the GI tract, biliary tract, kidney, muscle, breast and the heart3-5. Primary sclerosing cholangitis (PSC) is a chronic liver disease of unknown etiology, characterized by cholestasis, inflammation, fibrosis and stricture formation of the biliary ducts. The pathophysiology of PSC is a complex multistep process included unclear immunological mechanisms, genetic susceptibility and various defects of the biliary epithelial cells. The disease is rare in the general population but is strongly associated with inflammatory bowel disease. The prevalence of IBD, predominantly ulcerative colitis, among PSC patients is approximately 70-90% while only 5% of patients with UC develop PSC. The percentage of