Moritz Schmelzle

Making sense of regulatory T cell suppressive function.

Several types of regulatory T cells maintain self-tolerance and control excessive immune responses to foreign antigens. The major regulatory T subsets described over the past decade and novel function in transplantation will be covered in this review with a focus on CD4(+)CD25(+)Foxp3(+) regulatory T (Treg) cells. Multiple mechanisms have been proposed to explain how Treg cells inhibit effector cells but none can completely explain the observed effects in toto. Proposed mechanisms to explain suppressive activity of Treg cells include the generation of inhibitory cytokines, induced death of effector cells by cytokine deprivation or cytolysis, local metabolic perturbation of target cells mediated by changes in extracellular nucleotide/nucleoside fluxes with alterations in intracellular signaling molecules such as cyclic AMP, and finally inhibition of dendritic cell functions. A better understanding of how Treg cells operate at the molecular level could result in novel and safer therapeutic approaches in transplantation and immune-mediated diseases.

Infusion of CD133+ bone marrow-derived stem cells after selective portal vein embolization enhances functional hepatic reserves after extended right hepatectomy: a retrospective single-center study.

Objective:This study was designed to evaluate the clinical outcome of patients undergoing portal vein embolization (PVE) and autologous CD133+ bone marrow–derived stem cell (CD133+ BMSC) application before extended right hepatectomy. Background:We have previously shown that portal venous infusion of CD133+ BMSCs substantially increases hepatic proliferation, when compared with PVE alone. Methods:Among 40 consecutive patients with a median follow-up of 28 months (7.4–57.2) scheduled for extended right hepatectomy, we compared a preconditioned group with PVE and CD133+ BMSC cotreatment (PVE+SC group, n = 11) and a group pretreated only with PVE (PVE group, n = 11). Functional and overall outcomes after extended right hepatectomy were evaluated. Patients without presurgical treatment served as controls (n = 18). Results:In preconditioned patients, mean hepatic growth of segments II/III 14 days after PVE in the PVE+SC group was significantly higher (138.66 mL ± 66.29) when compared with that of PVE group patients (62.95 mL ± 40.03; P = 0.004). There were no significant differences among all 3 groups regarding general and oncological characteristics and functional parameters on postoperative day (POD) 7. Lack of hepatic preconditioning, extrahepatic extension of resection, and postoperative complications were of negative prognostic value, using univariate analysis (P < 0.05). In multivariate analysis, freedom from postoperative major complications (P = 0.012), coagulation status on POD 7 (international normalized ratio < 1.4; P = 0.027), and presurgical expansion of the future liver remnant volume (P = 0.048) were positively associated with overall survival. Post hoc analysis revealed a better survival for the PVE+SC group (P = 0.028) compared with the PVE group (P = 0.094) and compared with controls. conclusion:Promising data from this survival analysis suggest that PVE, together with CD133+ BMSC pretreatment, could positively impact overall outcomes after extended right hepatectomy.

Sublethal heat treatment promotes epithelial‐mesenchymal transition and enhances the malignant potential of hepatocellular carcinoma

Radiofrequency ablation (RFA) is a potentially curative therapy for hepatocellular carcinoma (HCC). However, incomplete RFA can induce accelerated invasive growth at the periphery. The mechanisms underlying the RFA‐induced tumor promotion remain largely unexplored. Three human HCC cell lines were exposed to 45°C‐55°C for 10 minutes, simulating the marginal zone of RFA treatment. At 5‐12 days post‐treatment cell proliferation, parameters of epithelial‐mesenchymal transition (EMT), and activation of mitogen‐activated protein kinases were analyzed. Livers from patients with viral hepatitis without and with HCC (n = 114) were examined to confirm the relevance of altered kinase patterns. In vivo tumorigenic potential of heat‐treated versus untreated HCC cells was studied in nude mice. Heating to 55°C killed all HCC cells, whereas 65%‐85% of cells survived 48°C‐50°C, developing spindle‐like morphology and expressing CD133, cytokeratin (CK)7, CK19, procollagen‐α1(I), and Snail at day 5 after heat exposure, which returned to baseline at day 12. Heat‐exposed HCC cells showed enhanced proliferation and prominent activation of p46‐Shc (Src homology and collagen) and downstream extracellular signal‐related kinase (Erk)1/2. In patients, Shc expression correlated with malignant potential and overall survival. Blocking Erk1/2 reduced proliferation and EMT‐like changes of heat‐treated HCC cells. Implantation of heat‐exposed HEPG2 cells into nude mice induced significantly larger, more aggressive tumors than untreated cells. Conclusions: Sublethal heat treatment skews HCC cells toward EMT and transforms them to a progenitor‐like, highly proliferative cellular phenotype in vitro and in vivo, which is driven significantly by p46Shc‐Erk1/2. Suboptimal RFA accelerates HCC growth and spread by transiently inducing an EMT‐like, more aggressive cellular phenotype. (Hepatology 2013;58:1667–1680)

Long-term Survival After Surgery for Primary Hepatic Sarcoma in Adults

HYPOTHESIS Patients with primary hepatic sarcomas benefit from resection, with possible long-term cure. DESIGN Retrospective and prospective cohort study. SETTING University hospitals of Hamburg-Eppendorf and Düsseldorf, Germany. PATIENTS Between 1985 and 2006, 22 patients (8 men and 14 women; median age at initial diagnosis, 54 years [range, 19-80 years]) were surgically treated for primary hepatic sarcomas. INTERVENTION Tumor resection with curative intent ranging from nonanatomical resection to liver transplant. MAIN OUTCOME MEASURES Effects on overall survival were analyzed using the log-rank test. RESULTS The majority of tumors were more than 5 cm (n = 19), with a median tumor size of 7 cm (range, 4-14 cm); of intermediate differentiation (G2; n = 15); and classified as leiomyosarcoma (n = 7). Ten patients received a hemihepatectomy. In 4 patients, a bisegmentectomy was performed and in 2 patients, a segmentectomy, while 4 patients received a nonanatomical resection. Liver transplant was performed in 2 patients. In 18 patients, complete tumor resection (R0) was achieved. Perioperative mortality was 0%. Median follow-up was 88 months (range, 6-246 months). Local recurrence occurred in 6 patients. Distant metastases were diagnosed in 10 patients, predominantly in the lung (n = 6). The 5-year survival after surgery was 65%, with 41% of the patients living more than 10 years without disease. Patients with angiosarcoma had a poor prognosis (P = .03). CONCLUSIONS Although primary hepatic sarcoma is a rare malignant tumor, no standard treatment is established. A long-term survival is possible after complete tumor resection in a preselected population with early-stage disease.

Disordered purinergic signaling and abnormal cellular metabolism are associated with development of liver cancer in Cd39/ENTPD1 null mice.

Liver cancer is associated with chronic inflammation, which is linked to immune dysregulation, disordered metabolism, and aberrant cell proliferation. Nucleoside triphosphate diphosphohydrolase‐1; (CD39/ENTPD1) is an ectonucleotidase that regulates extracellular nucleotide/nucleoside concentrations by scavenging nucleotides to ultimately generate adenosine. These properties inhibit antitumor immune responses and promote angiogenesis, being permissive for the growth of transplanted tumors. Here we show that Cd39 deletion promotes development of both induced and spontaneous autochthonous liver cancer in mice. Loss of Cd39 results in higher concentrations of extracellular nucleotides, which stimulate proliferation of hepatocytes, abrogate autophagy, and disrupt glycolytic metabolism. Constitutive activation of Ras‐mitogen‐activated protein kinase (MAPK) and mammalian target of rapamycin (mTOR)‐S6K1 pathways occurs in both quiescent Cd39 null hepatocytes in vitro and liver tissues in vivo. Exogenous adenosine 5′‐triphosphate (ATP) boosts these signaling pathways, whereas rapamycin inhibits such aberrant responses in hepatocytes. Conclusion: Deletion of Cd39 and resulting changes in disordered purinergic signaling perturb hepatocellular metabolic/proliferative responses, paradoxically resulting in malignant transformation. These findings might impact adjunctive therapies for cancer. Our studies indicate that the biology of autochthonous and transplanted tumors is quite distinct. (HEPATOLOGY 2013)

Coagulation, platelet activation and thrombosis in xenotransplantation.

Purpose of reviewXenotransplantation may become clinically feasible once the mechanisms of graft loss and rejection are better understood. Inflammatory reactions to vasculature of grafted pig organs and pancreatic islets have been linked to procoagulant activation and consumption with resulting thrombosis that precludes long-term function. Although development of α-1,3-galactosyltransferase gene-knockout swine with removal of a dominant xenoantigen has been an important advance, major problems still persist. Recent findingsConsumptive coagulopathy and platelet sequestration are initiated by immune responses associated with xenograft rejection and are exacerbated by putative intrinsic functional incompatibilities. Thrombotic processes together with progressive xenograft microangiopathy and infarction are intertwined with humoral immune reactions that may be secondary, at least in part, to ‘natural’ or elicited nongalactosyl antibodies. These immune responses are further exacerbated by the documented intrinsic molecular incompatibilities in the vascular regulation of blood clotting and extracellular nucleotide homeostasis between discordant species. Hence, limited benefits have been achieved with currently available pharmacological antithrombotics and anticoagulants. SummaryProposed strategies to tackle this problem will include optimal immunosuppressive interventions, attempts to induce tolerance, judicious and more effective use of anti-thrombotics with development of mutant swine either transgenic for human anticoagulants and thromboregulatory factors or null for defined porcine procoagulants.

HGF and SDF-1-mediated mobilization of CD133+BMSC for hepatic regeneration following extensive liver resection

The molecular mechanisms of haematopoietic stem cells (HSC) mobilization and homing to the liver after partial hepatectomy (PH) remain largely unexplored.

Increased plasma levels of microparticles expressing CD39 and CD133 in acute liver injury

Background We have previously demonstrated that CD133 and CD39 are expressed by hematopoietic stem cells (HSC), which are mobilized after liver injury and target sites of injury, limit vascular inflammation, and boost hepatic regeneration. Plasma microparticles (MP) expressing CD39 can block endothelial activation. Here, we tested whether CD133+ MP might be shed in a CD39-dependent manner in a model of liver injury and could potentially serve as biomarkers of liver failure in the clinic. Methods Wild-type and Cd39-null mice were subjected to acetaminophen-induced liver injury. Mice were sacrificed and plasma MP were isolated by ultracentrifugation. HSC and CD133+ MP levels were analyzed by fluorescence-activated cell sorting. Patients were enrolled with acute (n=5) and acute on chronic (n=5) liver injury with matched controls (n=7). Blood was collected at admission and plasma CD133+ and CD39+ MP subsets were analyzed by fluorescence-activated cell sorting. Results HSC and CD133+ MP levels were significantly increased only in the plasma of wild-type mice with acetaminophen hepatotoxicity (P<0.05). No increases in CD133+ MP were noted in Cd39-null mice. Plasma MP increases were observed in patients with liver injury. These MP were characterized by significantly higher levels of CD39 (P<0.05). Conclusions HSC and plasma CD133+ MP levels increase in a CD39-dependent manner during experimental acute liver injury. Increased levels of CD39+ MP are differentially noted in patients with liver injury. Further research is needed to determine whether MP fluxes are secondary to pathophysiologic insults to the liver or might reflect compensatory responses.

Long-Term Vacuum-Assisted Closure in Open Abdomen due to Secondary Peritonitis: A Retrospective Evaluation of a Selected Group of Patients

Background/Aims: Vacuum-assisted closure (VAC) leads to a high fascial closure rate in open abdomen within the first week of treatment. However, little data exist on the role of long-term VAC treatment in patients with peritonitis, where fascial closure cannot be accomplished within the first days. Methods: We reviewed the medical records of 49 patients with open abdomen for more than 7 days due to secondary peritonitis, who underwent a VAC-treatment. Nonparametric analysis was performed using χ2 test or Fisher’s exact test. Results: Fascial closure could be accomplished in only 11 patients (22%), whereas complications occurred in 43 patients (88%). Re-explorations after starting VAC were associated with the occurrence of enterocutaneous fistula (p < 0.001) and were also of prognostic value regarding the rate of fascial closure (p = 0.033). Conclusions: If fascial closure cannot be accomplished within the first days, patients show a dramatically lower fascial closure and an increased complication rate with VAC. Further studies are needed to evaluate whether this subgroup really benefits from VAC.

CD39 modulates hematopoietic stem cell recruitment and promotes liver regeneration in mice and humans after partial hepatectomy.

Objective:To study molecular mechanisms involved in hematopoietic stem cell (HSC) mobilization after liver resection and determine impacts on liver regeneration. Background:Extracellular nucleotide-mediated cell signaling has been shown to boost liver regeneration. Ectonucleotidases of the CD39 family are expressed by bone marrow–derived cells, and purinergic mechanisms might also impact mobilization and functions of HSC after liver injury. Methods:Partial hepatectomy was performed in C57BL/6 wild-type, Cd39 ectonucleotidase-null mice and in chimeric mice after transplantation of wild-type or Cd39-null bone marrow. Bone marrow–derived HSCs were purified by fluorescence-activated cell sorting and administered after hepatectomy. Chemotactic studies were performed to examine effects of purinergic receptor agonists and antagonists in vitro. Mobilization of human HSCs and expression of CD39 were examined and linked to the extent of resection and liver tests. Results:Subsets of HSCs expressing Cd39 are preferentially mobilized after partial hepatectomy. Chemotactic responses of HSCs are increased by CD39-dependent adenosine triphosphate hydrolysis and adenosine signaling via A2A receptors in vitro. Mobilized Cd39high HSCs boost liver regeneration, potentially limiting interleukin 1&bgr; signaling. In clinical studies, mobilized human HSCs also express CD39 at high levels. Mobilization of HSCs correlates directly with the restoration of liver volume and function after partial hepatectomy. Conclusions:We demonstrate CD39 to be a novel HSC marker that defines a functionally distinct stem cell subset in mice and humans. HSCs are mobilized after liver resection, limit inflammation, and boost regeneration in a CD39-dependent manner. These observations have implications for monitoring and indicate future therapeutic avenues.