Andreas Kreft

Five Cases of De Novo Inflammatory Bowel Disease After Orthotopic Liver Transplantation

Immunosuppression is currently the treatment of choice for severe inflammatory bowel disease (IBD). Thus, it was anticipated that the course of preexisting IBD should improve after orthotopic liver transplantation (OLT). Despite sufficient allograft immunosuppressive therapy, however, exacerbation of IBD or the development of de novo IBD after OLT were described in some cases, primarily in patients transplanted for end-stage primary sclerosing cholangitis (PSC). In addition, the development of de novo IBD in patients undergoing OLT for indications other than PSC was described. Evaluating our collective of 314 liver transplanted patients we found five patients transplanted for various indications other than PSC (autoimmune hepatitis [AIH], acute-on-chronic hepatitis B, Wilsons disease, and cryptogenic cirrhosis) who developed de novo IBD after OLT despite sufficient immunosuppressive therapy with tacrolimus or cyclosporine. PSC was widely excluded in these patients by clinical and histological examinations and there was no sign of an enteric infection. It was remarkable that all patients were suspected to have an autoimmune background. Four of our patients were women and almost all patients showed histologically typical signs of an ulcerative colitis (UC). To prevent allograft rejection, three of five patients were treated with cyclosporine and the other two with tacrolimus. After diagnosis, treatment with aminosalicylates and corticosteroids led to complete clinical and histological remission. However, relapses occurred frequently after termination of specific therapy. In combination with previous reports, our cases indicate an immune dysregulation leading to the development of de novo IBD after OLT under immunosuppressive therapy. Reviewing the literature, it should be considered that apart from the autoimmune background, immunosuppressive therapy may itself play a major role in the development of de novo IBD. From the clinical point of view, it is of critical importance to detect this phenomenon, since diarrhea is an important cause of morbidity and mortality in transplanted patients and therapy for this disorder completely differs from the treatment for other causes of diarrhea. Aminosalicylates and courses of corticosteroids offer an effective treatment.

The Incidence of Myelofibrosis in Essential Thrombocythaemia, Polycythaemia vera and Chronic Idiopathic Myelofibrosis: A Retrospective Evaluation of Sequential Bone Marrow Biopsies

The incidence of myelofibrosis (MF) among the three major Philadelphia chromosome-negative chronic myeloproliferative disorders, i.e. essential thrombocythaemia (ET), polycythaemia vera (PV) and chronic idiopathic myelofibrosis (CIMF), is not well documented since the diagnostic criteria have recently been redefined by the WHO. Therefore we performed a retrospective analysis of follow-up biopsies of 275 patients with ET, PV and CIMF according to the WHO classification of chronic myeloproliferative disorders. In the diagnostic bone marrow biopsies, MF was observed in 57 of the 136 CIMF patients (42%), 4 of the 73 PV patients (5%) and none of the 66 patients with ET. Within a median observation time of 2.9 years, 34 of the 79 patients with CIMF (43%), 13 of the 69 patients with PV (19%) and 1 of the 66 patients with ET (1.5%) – each initially without MF – developed MF regardless of myelosuppressive therapy.

Ablation of c-FLIP in hepatocytes enhances death-receptor mediated apoptosis and toxic liver injury in vivo.

BACKGROUND & AIMS Apoptosis is crucially involved in acute and chronic liver injury, including viral, cholestatic, toxic, and metabolic liver disease. Additionally, dysregulation of apoptosis signaling pathways has been implicated in hepatocarcinogenesis. The most prominent members of the apoptosis-mediating tumor necrosis factor receptor superfamily are the TNF-R1 (CD120a) and the CD95 (Apo-1/Fas) receptor. Although extensively studied, the intracellular signaling events in hepatocytes are only incompletely understood. METHODS To examine the role of the caspase-8 homolog cellular FLICE-inhibitory protein (c-FLIP) in liver injury, we generated mice with hepatocyte specific deletion of c-FLIP. Three models of acute liver injury were employed: the agonistic anti-CD95 antibody Jo2, d-galactosamine and LPS (GalN/LPS), and concanavalin A. RESULTS Conditional ablation of c-FLIP in hepatocytes augmented liver injury and cell death in all three models of liver injury. CD95- and GalN/LPS-induced liver injury was ameliorated by a pancaspase inhibitor, while ConA-induced injury was unaffected by caspase inhibition. Augmented activation of the MAPK JNK was observed in parallel to liver injury in c-FLIP knockout mice in all injury models; however, inhibition of JNK only affected TNF- and ConA-mediated injury. CONCLUSIONS In summary, c-FLIP is a central regulator of cell death in hepatocytes, involving increased activation of caspases and the MAPK JNK.

Rectal Cancer: Mucinous Carcinoma on Magnetic Resonance Imaging Indicates Poor Response to Neoadjuvant Chemoradiation

PURPOSE To assess response of locally advanced rectal carcinoma to chemoradiation with regard to mucinous status and local tumor invasion found at pretherapeutic magnetic resonance imaging (MRI). METHODS AND MATERIALS A total of 88 patients were included in this prospective study of patients with advanced mrT3 and mrT4 carcinomas. Carcinomas were categorized by MRI as mucinous (mucin proportion >50% within the tumor volume), and as nonmucinous. Patients received neoadjuvant chemoradiation consisting of 50.4 Gy (1.8 Gy/fraction) and 5-fluorouracil on Days 1 to 5 and Days 29 to 33. Therapy response was assessed by comparing pretherapeutic MRI with histopathology of surgical specimens (minimum distance between outer tumor edge and circumferential resection margin = CRM, T, and N category). RESULTS A mucinous carcinoma was found in 21 of 88 patients. Pretherapeutic mrCRM was 0 mm (median) in the mucinous and nonmucinous group. Of the 88 patients, 83 underwent surgery with tumor resection. The ypCRM (mm) at histopathology was significantly lower in mucinous carcinomas than in nonmucinous carcinomas (p ≤ 0.001). Positive resection margins (ypCRM ≤ 1 mm) were found more frequently in mucinous carcinomas than in nonmucinous ones (p ≤ 0.001). Treatment had less effect on local tumor stage in mucinous carcinomas than in nonmucinous carcinomas (for T downsizing, p = 0.012; for N downstaging, p = 0.007). Disease progression was observed only in patients with mucinous carcinomas (n = 5). CONCLUSION Mucinous status at pretherapeutic MRI was associated with a noticeably worse response to chemoradiation and should be assessed by MRI in addition to local tumor staging to estimate response to treatment before it is initiated.

Rectal cancer: assessment of response to neoadjuvant chemoradiation by dynamic contrast-enhanced MRI.

To assess pretreatment functional and morphological tumor characteristics with magnetic resonance imaging (MRI) in advanced rectal carcinoma and to identify factors predicting response to neoadjuvant chemoradiation.

Donor and host B cell-derived IL-10 contributes to suppression of graft-versus-host disease.

Graft‐versus‐host disease (GvHD) is a frequent life‐threatening complication following allogeneic HSC transplantation (HSCT). IL‐10 is a regulatory cytokine with important roles during GvHD, yet its relevant sources, and mode of action, remain incompletely defined in this disease. Using IL‐10‐deficient donor or host mice (BALB/c or C57BL/6, respectively) in a MHC‐mismatched model for acute GvHD, we found a strongly aggravated course of the disease with increased mortality when either donor or host cells could not produce this cytokine. A lack of IL‐10 resulted in increased allogeneic T‐cell responses and enhanced activation of host DCs in spleen and MLNs. Remarkably, IL‐10 was prominently produced by host‐ and donor‐derived CD5intCD1dintTIM‐1int B cells in this disease, and consistent with this, allogeneic HSCT resulted in exacerbated GvHD when mice lacking IL‐10 expression in B cells were used as donor or host, compared with controls. Taken together, this study demonstrates that host and donor B cell‐derived IL‐10 provides a unique mechanism of suppression of acute GvHD, and suggests that DCs are the targets of this B cell‐mediated suppressive effect. These findings open novel therapeutic possibilities based on the use of B cells to increase the feasibility of allogeneic HSCT.

Bone marrow findings in multicentric Castleman disease in HIV-negative patients.

Because bone marrow histology in multicentric Castleman disease in human immunodeficiency virus-negative patients is not well reported, we investigated sequential bone marrow biopsies of 3 affected human immunodeficiency virus-negative patients, of which one was human herpes virus 8 (HHV8)-positive. The histologic evaluation of the bone marrow revealed lymphoid follicles with regressed germinal centers in 1 patient. Another patient showed tumorlike but bland polyclonal plasmacytosis with large perivascular plasma cell clusters. The HHV8-positive patient revealed interstitial HHV8-positive cells accompanied by a mild plasmacytosis. The atypical lymphoid follicles could be regarded as a bone marrow manifestation of multicentric Castleman disease, whereas the plasmacytosis most likely is the result of excess interleukin 6 production. The presence of HHV8-positive cells within the bone marrow may indicate the dissemination of the virus in a compromised immune system.

Oxidative burst and neutrophil elastase contribute to clearance of Aspergillus fumigatus pneumonia in mice.

Polymorphonuclear neutrophils (PMN) are important for the control of invasive aspergillosis (IA), a major threat to immunocompromised individuals. For clearance of Aspergillus fumigatus infections, PMN employ their potent oxidative and non-oxidative mechanisms. To clarify the relative contribution of these mechanisms, we analyzed p47(phox-/-), gp91(phox-/-) and elastase (ELA) deficient mice (ELANE) after intratracheal infection with A. fumigatus. Infected p47(phox-/-) and gp91(phox-/-) mice died within 4 days and had a significant higher fungal burden in the lungs compared to wild-type controls. Interestingly, the survival of ELANE mice after infection was unimpaired suggesting that ELA is not essential here. Nevertheless, A. fumigatus clearance was delayed in ELANE mice indicating a partial contribution of ELA to fungal immunity. Comparing p47(phox-/-), gp91(phox-/-) or ELANE mice for PMN activation and recruitment to the lungs, we were unable to detect significant differences in vitro or in vivo among mutant or wild-type strains suggesting intact PMN functionality of basic effector mechanisms. Fungal killing in vitro by ELA deficient PMN was comparably reduced as in p47(phox-/-) and gp91(phox-/-) deficient PMN corroborating the importance of oxidative and non-oxidative PMN mechanisms for the control of fungal outgrowth. Taken together, this suggests that intact oxidative as well as non-oxidative PMN effector functions are highly relevant for the control of A. fumigatus infections in vitro and in vivo. While ELA contributes to clearance of A. fumigatus, the oxidative functions are essential for survival.

Polycythaemia vera: bone marrow histopathology under treatment with interferon, hydroxyurea and busulphan

Abstract: Little is known about long‐term effects of myelosuppressive therapy on bone marrow of patients with polycythaemia vera, since histopathology from follow‐up biopsies has not been frequently reported. Thus we conducted a retrospective morphometrical analysis of diagnostic and follow‐up biopsies of 62 patients, evaluating fibre content, megakaryocytes and bone marrow cellularity. 8/62 patients were treated with interferon‐α (INF), 11/62 with hydroxyurea (HU) and 11/62 with busulphan (BU). 32/62 served as controls; they were not treated with myelosuppressive drugs but with phlebotomy only. The median observation time was 2.3 yr. Results were compared on the basis of change per time. The bone marrow of the patients with phlebotomies only was characterised by increasing cellularity of haematopoesis, number and volume ratio of megakaryocytes and fibre content. In BU‐ and HU‐treated patients, the haematopoesis was significantly reduced. The IFN patients revealed a reduction of cellularity, which was not significant. The fibre content was reduced by BU only, but not significantly. No correlation between megakaryocytes and fibres was found. – It could be concluded therefore that: 1) fibre proliferation within the bone marrow was not significantly altered by IFN, HU or BU. 2) Cellularity of haematopoesis was reduced significantly by HU and BU but only partly by IFN, corresponding with haematological remission.

STAT5 Is Crucial to Maintain Leukemic Stem Cells in Acute Myelogenous Leukemias Induced by MOZ-TIF2

MOZ-TIF2 is a leukemogenic fusion oncoprotein that confers self-renewal capability to hematopoietic progenitor cells and induces acute myelogenous leukemia (AML) with long latency in bone marrow transplantation assays. Here, we report that FLT3-ITD transforms hematopoietic cells in cooperation with MOZ-TIF2 in vitro and in vivo. Coexpression of FLT3-ITD confers growth factor independent survival/proliferation, shortens disease latency, and results in an increase in the number of leukemic stem cells (LSC). We show that STAT5, a major effector of aberrant FLT3-ITD signal transduction, is both necessary and sufficient for this cooperative effect. In addition, STAT5 signaling is essential for MOZ-TIF2-induced leukemic transformation itself. Lack of STAT5 in fetal liver cells caused rapid differentiation and loss of replating capacity of MOZ-TIF2-transduced cells enriched for LSCs. Furthermore, mice serially transplanted with Stat5(-/-) MOZ-TIF2 leukemic cells develop AML with longer disease latency and finally incomplete penetrance when compared with mice transplanted with Stat5(+/+) MOZ-TIF2 leukemic cells. These data suggest that STAT5AB is required for the self-renewal of LSCs and represents a combined signaling node of FLT3-ITD and MOZ-TIF2 driven leukemogenesis. Therefore, targeting aberrantly activated STAT5 or rewired downstream signaling pathways may be a promising therapeutic option.