Ana Paula Barreiros

Improved Differentiation of Pancreatic Tumors Using Contrast-Enhanced Endoscopic Ultrasound

BACKGROUND & AIMS Endoscopic ultrasound is a widely accepted imaging method for staging of ductal adenocarcinoma and the localization of neuroendocrine tumors of the pancreas. We prospectively evaluated conventional color Doppler imaging and contrast-enhanced endoscopic Doppler ultrasound (CE-EDUS) as a new imaging technique for further characterization and differentiation of solid pancreatic tumors. METHODS From 300 patients with pancreatic lesions investigated using contrast-enhanced endoscopic ultrasound we could finally include 93 patients with an undetermined, solitary, predominantly solid, lesion 40 mm or less, and a definite histologically proven diagnosis. After bolus injection of the contrast agent SHU 508A 4 g (400 mg/dL) the vascular pattern of the lesion during the arterial phase was compared with the vascularity of the residual pancreatic parenchyma. RESULTS Color Doppler imaging did not reveal vascularity of the pancreatic parenchyma in any of the patients, and therefore tumor hypovascularity could not be determined in contrast to all CE-EDUS-examined patients revealing at least some degree of parenchymal vascularity. Fifty-seven of 62 patients with ductal adenocarcinoma of the pancreas showed a hypovascularity of the tumor using CE-EDUS. All other pancreatic lesions revealed an isovascular or hypervascular pattern using contrast-enhanced endoscopic ultrasound (20 neuroendocrine tumors, 10 serous microcystic adenomas, and 1 teratoma). Hypovascularity as a sign of malignancy in contrast-enhanced endoscopic ultrasound obtained 92% (82%-97%) sensitivity and 100% specificity (89%-100%). CONCLUSIONS Contrast-enhanced endoscopic ultrasound is effective in differentiating small solid pancreatic tumors of different origin in most cases. Hypovascularity indicates malignancy of pancreatic tumors.

Multicentric evaluation of model for end‐stage liver disease‐based allocation and survival after liver transplantation in Germany – limitations of the ‘sickest first’‐concept

Since the introduction of model for end‐stage liver disease (MELD) in 2006, post‐orthotopic liver transplantation (OLT) survival in Germany has declined. The aim of this study was to evaluate risk factors and prognostic scores for outcome. All adult OLT recipients in seven German transplant centers after MELD implementation (December 2006–December 2007) were included. Recipient data were analyzed for their influence on 1‐year outcome. A total of 462 patients (mean calculated MELD = 20.5, follow‐up: 1 year) were transplanted for alcoholic cirrhosis (33.1%), hepatocellular carcinoma (26.6%), Hepatitis‐C (17.1%), Hepatitis‐B (9.5%), primary sclerosing cholangitis (5.6%) and late graft‐failure after first OLT before December 2006 (8.7%). 1‐year patient survival was 75.8% (graft survival 71.2%) correlating with MELD parameters and serum choline esterase. MELD score >30 [odds ratio (OR) = 4.17, confidence interval: 2.57–6.78, 12‐month survival = 52.6%, c‐statistic = 0.669], hyponatremia (OR = 2.07), and pre‐OLT hemodialysis (OR = 2.35) were the main death risk factors. In alcoholic cirrhosis (n = 153, mean MELD = 21.1) and hepatocellular carcinoma (n = 123, mean MELD = 13.5), serum bilirubin and the survival after liver transplantation score were independent outcome parameters, respectively. MELD >30 currently represents a major risk factor for outcome. Risk factors differ in individual patient subgroups. In the current German practice of organ allocation to sicker patients, outcome prediction should be considered to prevent results below acceptable standards.

Indications and limitations of endoscopic ultrasound elastography for evaluation of focal pancreatic lesions

BACKGROUND AND STUDY AIM Endoscopic-ultrasound-guided elastography (EUS-elastography) is a recently introduced imaging procedure that distinguishes tissues on the basis of their specific consistency. The aim of this prospective study was to investigate the role of this new technique in the characterization and differentiation of focal pancreatic lesions. PATIENTS AND METHODS This prospective study enrolled 70 patients with unclassified solid lesions of the pancreas and 10 controls with a healthy pancreas. In all patients elastography recordings were compared with cytology/histology findings as the gold standard. RESULTS Adequate EUS-elastography of the pancreas was performed in all healthy controls but in only 56 % of patients with solid pancreatic lesions. The main limitation of elastographic image acquisition was incomplete delineation of the border of lesions greater than 35 mm in diameter (39 %) or of lesions at some distance from the transducer (10 %). Elastographic recordings were also hampered by the fact that the surrounding tissue, which is used as an internal reference standard for strain calculation, was insufficiently displayed in the case of larger lesions. The reduced ratio of target to surrounding tissue resulted in the formation of color artifacts and in impaired reproducibility. In contrast, the majority of lesions smaller than 35 mm in diameter were adequately and reproducibly evaluated by EUS-elastography (91 %). The clinical use for differential diagnosis, however, seems limited, since strain images from all kinds of pancreatic masses were found to be harder than the surrounding tissues, irrespective of the underlying nature of the lesion (i. e., malignant vs. benign). EUS-elastography predicted the nature of pancreatic lesions with poor diagnostic sensitivity (41 %), specificity (53 %), and accuracy (45 %). CONCLUSION EUS-elastography of the pancreas has the potential to obtain some complementary information that would improve tissue characterization. Its clinical utility, however, remains questionable, and it seems unlikely that the information provided will obviate the necessity of obtaining tissue samples for confirmation of a final pathologic diagnosis.

Contrast enhanced ultrasound for the diagnosis of hepatocellular carcinoma (HCC): Comments on AASLD guidelines

To the Editor: Contrast enhanced ultrasound (CEUS) has been introduced 10 years ago for liver imaging in many European and Asian countries, but FDA approval in the US is still lacking [1]. The excellent value of CEUS has been established by numerous prospective studies including the German DEGUM-Study with over 1000 patients [2] and the respective French multicentric study [3]. CEUS was proven able to detect and characterize liver tumours in clinical routine within at least the same accuracy range as contrast enhanced computed tomography (CECT) and contrast enhanced magnetic resonance imaging (CEMRI) [4,5]. Therefore, CEUS has been introduced into important guidelines and recommendations, like those from the American Association for the Study of Liver Diseases (AASLD) 2005 [6], the Asian Pacific Association for the Study of the Liver (APASL) [7], the Japanese Society of Hepatology [8] and the European Federation of Societies for Ultrasound in Medicine and Biology (EFSUMB) guidelines 2004 [9], 2008 [10], and WFUMB-EFSUMB guidelines 2012 (in preparation). However, CEUS has been eliminated from the diagnostic flow chart of nodules in cirrhosis in the updated AASLD guidelines 2011 [11]. This removal raised controversial discussion and was not well received in Europe and Asia. This is therefore the issue to which the present commentary refers. There are two reasons for which CEUS has been eliminated from the AASLD guidelines. 1. ‘‘Contrast-enhanced US may offer false positive HCC diagnosis in patients with cholangiocarcinoma and thus, has been dropped from the diagnostic techniques’’ Intrahepatic cholangiocellular carcinoma (ICC) is a rare tumour in liver cirrhosis (about 1–3% of newly developed tumors) [12,13] but the incidence appears raising [14]. In a retrospective series of 21 patients with histologically confirmed ICC on cirrhosis collected between 2003 and 2009, the Barcelona Clinic Liver Cancer (BCLC) group found that ten ICC had the same CEUS enhancement pattern considered diagnostic for HCC, consisting in homogeneous arterial hyperenhancement followed by washout [15]. At variance, these tumours showed hyperintense enhancement in the arterial phase but lacked wash-out at MRI, failing to show the typical HCC pattern, thus prompting biopsy. The difference in the enhancement patterns is probably related to the different pharmacokinetics of contrast agents used for US (strictly intravascular) and MRI (extravascular space diffusion of Gadolinium which transiently binds to fibrous tissue, explaining

Efficacy of an escalating dose regimen of pegylated interferon α-2a plus ribavirin in the early phase of HCV reinfection after liver transplantation

We evaluated the safety and efficacy of an escalating dose regimen of pegylated interferon α‐2a (PEG‐IFNα‐2a) and ribavirin in the early phase of recurrent hepatitis C after orthotopic liver transplantation (OLT). In this prospective study, 26 patients transplanted for hepatitis C virus cirrhosis with recurrent hepatitis C were treated 3.4 ± 3.6 months after OLT and compared with an untreated historical control. PEG‐IFNα‐2a was initiated as monotherapy, following stepwise dose escalation up to 180 μg/week and the addition of ribavirin up to 1200 mg/day or maximally tolerated doses for 48 weeks. In the intent‐to‐treat analysis, 38% showed an early virological response (EVR), 35% an end of treatment response (ETR) and 19% a sustained virological response (SVR). SVR was associated with EVR (P = 0.0001) and cumulative PEG‐IFNα‐2a dose (P = 0.04). There was no significant histological improvement compared with untreated patients. There were no treatment‐related serious adverse events. Adverse events included leucopenia (77%) and thrombocytopenia (46%). Three patients discontinued therapy due to side effects, fourteen were nonresponders and four relapsers. Treatment with PEG‐IFNα‐2a and ribavirin in the acute phase of post‐transplant recurrent hepatitis C yielded an EVR of 38% and an SVR of 19%. The combination was safe and well tolerated.

Clinical symptomatic de novo systemic transthyretin amyloidosis 9 years after domino liver transplantation

Four years ago, Stangou et al. reported a patient who developed clinical symptoms of amyloidosis 8 years after domino liver transplantation (DLT). In order to alleviate the graft shortage, livers from patients with familial amyloidotic polyneuropathy (FAP) are used as domino grafts for other patients under the assumption that several decades will pass before clinical symptoms due to amyloid accumulation emerge. We now report on another case, the third reported case in the literature, of de novo systemic amyloidosis in a 75-year-old woman 9 years after DLT. The patient underwent DLT at 65 years of age for hepatocellular carcinoma exceeding the Milan criteria in hepatitis C cirrhosis. Transarterial chemoembolization was performed before liver transplantation. During treatment, progressive cachexia and increasing B symptoms occurred. Therefore, DLT was finally considered in this patient. The posttransplant course was uneventful. In 2006, the patient described progressive dysesthesia in the lower extremities. Immunosuppression was switched from a calcineurin inhibitor to a mammalian target of rapamycin inhibitor. Other reasons for neuropathy were excluded. A rectal biopsy specimen at this time was free of amyloid deposition. In 2007, neurological symptoms further deteriorated: burning pain and progressive sensory loss in the lower limbs occurred. In March 2008, amyloid deposition was detected in the gastrointestinal wall, reflecting de novo FAP in this patient. A literature review revealed 2 case reports of symptomatic patients: in the first patient, dysesthesia of the lower limb 8 years after DLT was described, and in the second patient, decreased temperature sensation and pain in the toes and fingertips 7 years after DLT were reported. Interestingly, in our patient and in both previously reported patients, neuropathic symptoms occurred after comparable intervals after DLT (7 and 8 versus 9 years), and the indication for DLT was hepatocellular carcinoma with underlying hepatitis C cirrhosis in the first described patient and our patient. All the patients had received an FAP graft with a transthyretin mutation; the former 2 received grafts with a Val30Met mutation, and our patient received a graft with a Gly47Glu mutation. In individuals with both inherited mutations, neurological symptoms usually occur within the age range of 20 to 35 years. After DLT, the onset of symptoms was thus significantly earlier than the onset in inherited FAP patients. Despite the risk of emerging amyloidosis in recipients, the graft shortage and increasing death rates on the waiting lists prompt alternative approaches to providing grafts. DLT is a feasible technique and may, therefore, be justified in selected patients. Nevertheless, our data reinforce the need for additional caution and meticulous follow-up programs.

Control of organ transplant-associated graft-versus-host disease by activated host lymphocyte infusions.

Background. Prolonged persistence of donor-derived T cells after organ transplantation has been proposed to improve long-term allograft survival. However, surviving transplant-derived T cells are also able to mediate devastating graft-versus-host disease (GvHD). Currently, GvHD after organ transplantation is usually refractory to conventional therapy and the disease outcome fatal. Methods. Graft-reactive host T cells were generated ex vivo from a patient suffering from a severe and refractory liver–transplant-associated GvHD. To control GvHD, activated alloreactive host T cells were repetitively retransferred into the patient (activated host lymphocyte infusion [aHLI]). Results. Adoptive transfer of ex vivo activated alloreactive host T cells (aHLI) led to the control and complete resolution of severe GvHD without inducing allograft rejection. Conclusions. aHLI opens a novel therapeutic window to control solid-organ transplant-associated GvHD while preserving allograft integrity.

Liver transplanted patients with preoperative autoimmune hepatitis and immunological disorders are at increased risk for Post-Transplant Lymphoproliferative Disease (PTLD).

BACKGROUND Long term immunosuppression and therapy of acute rejections result in a 20-120-fold increased risk to develop Non Hodgkin lymphoma (NHL). Since immunosuppressive therapy and immunological disorders are major risk factors for the development of NHL in the non-transplant population we aimed to analyze risk factors for PTLD in our cohort of liver transplanted (LT) patients. METHODS We analyzed retrospectively 431 patients liver transplanted between 1998 and 2008. RESULTS PTLD was diagnosed in eleven of 431 patients (2.6%). PTLD, especially late PTLD, was significantly more frequent in patients who received steroids before LT (Kaplan-Meier: p<0.001). Moreover PTLD in immunocompromised patients with preoperative steroid treatment occurred at a significantly younger age (49.5+/-4.7 years) compared to patients without steroids (60.6+/-5.1 years; p=0.006). Multivariate analysis revealed pretransplant steroid treatment and liver transplantation for autoimmune hepatitis as main risk factors for the development of PTLD after liver transplantation (p<0.001). CONCLUSION Liver transplanted patients who received steroids before LT due to immunological disorders and patients with autoimmune hepatitis seem to be at particular high risk to develop PTLD. Prospective cohort studies including immunoepidemiologic investigations of abnormalities of cellular, humoral and innate immunity should be carried out to identify predictive factors and patients at risk.

GM-CSF Restores Innate, But Not Adaptive, Immune Responses in Glucocorticoid-Immunosuppressed Human Blood In Vitro

Infection remains the major complication of immunosuppressive therapy in organ transplantation. Therefore, reconstitution of the innate immunity against infections, without activation of the adaptive immune responses, to prevent graft rejection is a clinically desirable status in transplant recipients. We found that GM-CSF restored TNF mRNA and protein expression without inducing IL-2 production and T cell proliferation in glucocorticoid-immunosuppressed blood from either healthy donors or liver transplant patients. Gene array experiments indicated that GM-CSF selectively restored a variety of dexamethasone-suppressed, LPS-inducible genes relevant for innate immunity. A possible explanation for the lack of GM-CSF to restore T cell proliferation is its enhancement of the release of IL-1βR antagonist, rather than of IL-1β itself, since exogenously added IL-1β induced an IL-2-independent Con A-stimulated proliferation of glucocorticoid-immunosuppressed lymphocytes. Finally, to test the in vivo relevance of our findings, we showed that GM-CSF restored the survival of dexamethasone- or cyclosporine A-immunosuppressed mice from an otherwise lethal infection with Salmonella typhimurium. In addition to this increased resistance to infection, GM-CSF did not induce graft rejection of a skin allotransplant in cyclosporine A-immunosuppressed mice. The selective restoration potential of GM-CSF suggests its therapeutic use in improving the resistance against infections upon organ transplantation.

EGF and HGF levels are increased during active HBV infection and enhance survival signaling through extracellular matrix interactions in primary human hepatocytes

The hepatitis B virus (HBV) is a major causative agent of chronic liver disease and subsequent liver cirrhosis worldwide. The reduced sensitivity of virus‐infected liver cells to apoptosis may play a role in the failure to remove virus‐infected cells and eventually promote viral chronicity. The purpose of our study was to investigate whether survival factors induced during compensatory liver regeneration may protect hepatocytes against apoptosis. We evaluated the serum levels of hepatocyte growth factor (HGF) and epidermal growth factor (EGF) in HBV‐infected patients and found significant increases in HGF and EGF in patients with active virus infection. In primary human hepatocytes we show that HGF and EGF have a protective effect against CD95‐mediated apoptosis and cytotoxic T‐cell killing. Simultaneous treatment with both regeneration factors enhanced the cytoprotective effect. The PI 3‐K/Akt kinase inhibitor, wortmannin, and the STAT3 pathway inhibitor, Tyrphostin AG490, both effectively attenuated the cytoprotective effect of HGF and EGF. Furthermore, we show an EGF/HGF‐dependent upregulation of β1‐integrin chains, increased adhesion to extracellular matrix and an increase in focal adhesions, suggesting outside‐in signaling from the extracellular matrix as an additional cytoprotective mechanism. Our study demonstrates that HGF and EGF can interfere with CD95‐mediated apoptosis and the action of cytotoxic T‐cells through multiple mechanisms in human hepatocytes. Together our results argue that a survival mileau generated by activation of liver regeneration factors may be a risk factor for establishing viral persistence.