Dominik Rüttinger

In vivo assessment of hepatic alterations following gadolinium chloride-induced Kupffer cell blockade

BACKGROUND/AIMS In recent years, Gadolinium chloride (GdCl3), a rare earth metal, has frequently been used to study the role and function of Kupffer cells under physiological and pathological conditions. This study was performed to elucidate the consequences of GdCl3-induced Kupffer cell blockade for hepatic microcirculation, hepatocellular function and integrity. METHODS/RESULTS Using intravital fluorescence microscopy, we studied the hepatic microcirculation of rats pretreated with either GdCl3 (n = 12; 10 mg/kg; 1 ml i.v. for 2 d) or saline (n = 9; 1 ml). The GdCl3-treated animals revealed a significantly lower phagocytic activity of Kupffer cells when compared to controls. Concomitantly, GdCl3-treatment resulted in a pronounced rise of serum cytokine activity (tumor necrosis factor-alpha; interleukin-6). The hepatic microvascular perfusion was characterized by a moderate increase in the number of non-perfused sinusoids accompanied by a reduction of bile flow. In addition, GdCl3-treatment caused a slight increase in liver enzyme activity (< 200 U/l) (aspartate aminotransferase and alanine aminotransferase) with no substantial parenchymal tissue injury (light microscopy). The groups did not differ in concentrations of circulating endotoxin (GdCl3-treatment: 0.044 +/- 0.042 ng/ml; controls: 0.052 +/- 0.014 ng/ml). CONCLUSIONS We conclude that hepatic alterations following Kupffer cell blockade with GdCl3 may possibly be the consequence of cytokine release as a response to the phagocytic challenge of GdCl3-aggregates. If used for Kupffer cell blockade, the hepatic alterations following GdCl3-treatment described in the present study should be taken into consideration.

Evaluation of a new prognostic score (Munich score) to predict long-term survival after resection of pulmonary renal cell carcinoma metastases

BACKGROUND The aim of this single-center study was to analyze factors predicting long-term outcomes following surgical resection of pulmonary metastases in patients with renal cell carcinoma. METHODS Two hundred two consecutive patients entered the study. Overall survival was analyzed by the Kaplan-Meier method. Multivariate analysis was performed using Cox regression models. RESULTS In 175 cases (87%), curative resection of the pulmonary metastases was achievable, with median survival of 43 months. Multivariate analysis revealed complete metastasectomy (R0), metastasis size >3 cm, positive nodal status of the primary tumor, synchronous metastases, pleural infiltration, and tumor-infiltrated hilar or mediastinal lymph nodes as independent prognostic factors for survival. On the basis of these findings, a new scoring system (the Munich score) was established to predict survival, which discriminates 3 groups with low, intermediate, and high risk for poor outcomes (median survival, 90, 31, and 14 months, respectively, P < .001). CONCLUSIONS The aim of the Munich score is to define patients with low, intermediate, and high risk for poor survival and will help identify patients who may benefit from further adjuvant therapy.

Hypertonic saline dextran attenuates leukocyte accumulation in the liver after hemorrhagic shock and resuscitation

BACKGROUND Hemorrhagic shock and resuscitation triggers a global ischemia/reperfusion phenomenon, in which activated leukocytes are considered strong contributors to the ensuing tissue damage. METHODS The aim of the study was to investigate the effects of hypertonic saline dextran (HSD) on the early leukocyte/endothelial interactions (intravital fluorescence microscopy) in a rat model of hemorrhagic shock (1 hour at mean arterial pressure of 40 mm Hg). The resuscitation was performed with lactated Ringers solution (RL, four times shed blood/20 minutes, n = 6), 6% dextran 60 (DEX, 100% shed blood/5 minutes, n = 8), and 7.2% NaCl/10% dextran 60 (HSD, 10% shed blood/2 minutes, n = 8). RESULTS After 1 hour of resuscitation, shock-induced stasis/adherence of leukocytes was further enhanced with RL (sinusoids 17.6+/-6.9%; venules 33.9+/-8.5%), whereas DEX and HSD attenuated leukocyte stagnation in sinusoids (DEX -7.4+/-6,1%; HSD -14.7+/-2.9%, p<0.01 vs. RL) and leukocyte adherence in postsinusoidal venules (DEX -12.2+/-8.6%, p<0.05 vs. RL; HSD -27+/-7.4%, p<0.01 vs. RL). CONCLUSION HSD reduced significantly the number of leukocytes accumulated in the liver after resuscitation of hemorrhagic shock, probably due to a combination of mechanisms of both components.

Immunotherapy of Peritoneal Carcinomatosis with the Antibody Catumaxomab in Colon, Gastric, or Pancreatic Cancer: An Open-Label, Multicenter, Phase I/II Trial

Background: Peritoneal carcinomatosis (PC) is common in gastrointestinal (GI) cancer and there is no effective standard treatment. We investigated the tolerability and maximum tolerated dose (MTD) of the trifunctional antibody catumaxomab in patients with PC. Methods: In this openlabel, phase I/II clinical trial, patients with epithelial cell adhesion molecule (EpCAM)-positive PC from GI cancer received 4 sequential intraperitoneal catumaxomab infusions: day 0: 10 µg; day 3: 10 or 20 µg; day 7: 30, 50, or 100 µg; and day 10: 50, 100, or 200 µg. Dose escalation was guided by dose-limiting toxicities. Results: The MTD was 10, 20, 50, and 200 µg on days 0, 3, 7, and 10, respectively. Catumaxomab had an acceptable safety profile: Most common treatment-related adverse events (at the MTD) were fever, vomiting, and abdominal pain. At final examination, 11/17 evaluable patients (65%) were progression free: 1 patient had a complete and 3 a partial response. Median overall survival from the time of diagnosis of PC was 502 days. Conclusions: Intraperitoneal catumaxomab is a promising option for the treatment of PC from GI cancer.

Use of gastric residual volume to guide enteral nutrition in critically ill patients: a brief systematic review of clinical studies.

OBJECTIVE In critically ill patients, the optimal procedure to monitor upper gastrointestinal function is controversial. Several authors have proposed gastric residual volume (GRV) as a tool to guide enteral nutrition. The aim of this contribution is to briefly discuss corresponding studies. METHODS We electronically searched MEDLINE, EMBASE, and CINAHL for studies relevant to the subject. RESULTS Six randomized controlled trials (RCTs) and six prospective observational studies were identified. Each analyzed different thresholds of GRV to guide enteral nutrition and to avoid complications (e.g., vomiting, aspiration, nosocomial pneumonia) in artificially ventilated patients. Due to heterogeneity in outcome measures, patient populations, type and diameter of feeding tubes, and randomization procedures, combination of the results of the six RCTs into a meta-analysis was not appropriate. High-quality RCTs studying medical patients could not demonstrate an association between complication rate and the magnitude of GRV. The only observational study that adjusted results to potential confounders and that studied surgical patients found, however, that the frequency of aspiration increased significantly if a GRV > 200 mL was registered more than once. CONCLUSION For mechanically ventilated patients with a medical diagnosis at admission to the intensive care unit, monitoring of GRV appears unnecessary to guide nutrition. Surgical patients might profit, however, from a low GRV threshold (200 mL).

Prolonged Overall Survival After Pulmonary Metastasectomy in Patients With Breast Cancer

BACKGROUND We investigated whether overall survival (OS) in patients with primary breast cancer (BC) is prolonged by pulmonary metastasectomy and which prognostic criteria may facilitate the decision in favor of thoracic surgical intervention. METHODS We assessed the median OS of 81 women after resection of pulmonary primary BC metastases by means of Kaplan-Meier estimators. Statistical interferences regarding prognostic factors were based on univariate log-rank tests and multivariate Cox proportional hazards regression. Matched patients who had not undergone resection from the Munich Tumor Registry served as controls. RESULTS Between 1982 and 2007, 81 patients were recruited prospectively. In 81.5% of the patients R0 resection was achieved, which was associated with significantly longer median OS than occurred after R1 or R2 resection (103.4 months versus 23.6 months versus 20.2 months, respectively; p<0.001). Multivariate analysis revealed R0 resection, number (n≥2), size (≥3 cm), and estrogen receptor (ER) and/or progesterone receptor (PR) positivity of metastases as independent prognostic factors for long-term survival. Presence of metastases in mediastinal and hilar lymph nodes correlated with decreased survival only in the univariate analysis (32.1 versus 103.4 months; p=0.095). Matched pair analysis confirmed that pulmonary metastasectomy significantly improved survival. CONCLUSIONS OS in patients with isolated pulmonary primary BC metastasis is prolonged by metastasectomy. Patients with multiple pulmonary lesions or metastases with negative hormone receptor (HR) status are at greater risk of disease relapse and should be followed closely. Moreover, additive treatment tailored to the biological subtype defined by HR expression should be considered for this group.

Predictors of general complications after video-assisted thoracoscopic surgical procedures

BackgroundThe video-assisted thoracoscopic approach has become the preferred method for many procedures due to the reduced trauma, complication rate and morbidity. The aim of this study was a risk evaluation of patients undergoing video-assisted thoracoscopic surgery (VATS) procedures.MethodsBetween 1991 and 2004, 1,008 patients were included in this single-center retrospective analysis. Risk assessment was performed using univariate and multivariate analysis.ResultsMultivariate analysis revealed that patient age (p = 0.003), the duration of the VATS procedure (p = 0.008), redo-VATS (p < 0.001) and conversion to open thoracotomy (p < 0.001) correlated significantly with the incidence of complications. Patients with immune deficiency following organ transplantation had the highest complication rate at 31.7%, which was significantly higher than for patients with either benign disease (p = 0.010) or malignant disease (p = 0.019).ConclusionsVATS is a safe procedure, but extra caution is recommended for patients with a higher risk profile (age, redo-VATS, immune deficiency).

Therapeutic T cells induce tumor-directed chemotaxis of innate immune cells through tumor-specific secretion of chemokines and stimulation of B16BL6 melanoma to secrete chemokines

BackgroundThe mechanisms by which tumor-specific T cells induce regression of established metastases are not fully characterized. In using the poorly immunogenic B16BL6-D5 (D5) melanoma model we reported that T cell-mediated tumor regression can occur independently of perforin, IFN-γ or the combination of both. Characterization of regressing pulmonary metastases identified macrophages as a major component of the cells infiltrating the tumor after adoptive transfer of effector T cells. This led us to hypothesize that macrophages played a central role in tumor regression following T-cell transfer. Here, we sought to determine the factors responsible for the infiltration of macrophages at the tumor site.MethodsThese studies used the poorly immunogenic D5 melanoma model. Tumor-specific effector T cells, generated from tumor vaccine-draining lymph nodes (TVDLN), were used for adoptive immunotherapy and in vitro analysis of chemokine expression. Cellular infiltrates into pulmonary metastases were determined by immunohistochemistry. Chemokine expression by the D5 melanoma following co-culture with T cells, IFN-γ or TNF-α was determined by RT-PCR and ELISA. Functional activity of chemokines was confirmed using a macrophage migration assay. T cell activation of macrophages to release nitric oxide (NO) was determined using GRIES reagent.ResultsWe observed that tumor-specific T cells with a type 1 cytokine profile also expressed message for and secreted RANTES, MIP-1α and MIP-1β following stimulation with specific tumor. Unexpectedly, D5 melanoma cells cultured with IFN-γ or TNF-α, two type 1 cytokines expressed by therapeutic T cells, secreted Keratinocyte Chemoattractant (KC), MCP-1, IP-10 and RANTES and expressed mRNA for MIG. The chemokines released by T cells and cytokine-stimulated tumor cells were functional and induced migration of the DJ2PM macrophage cell line. Additionally, tumor-specific stimulation of wt or perforin-deficient (PKO) effector T cells induced macrophages to secrete nitric oxide (NO), providing an additional effector mechanism for T cell-mediated tumor regression.ConclusionThese data suggest two possible sources for chemokine secretion that stimulates macrophage recruitment to the site of tumor metastases. Both appear to be initiated by T cell recognition of specific antigen, but one is dependent on the tumor cells to produce the chemokines that recruit macrophages.

Adjuvant therapeutic vaccination in patients with non-small cell lung cancer made lymphopenic and reconstituted with autologous PBMC: first clinical experience and evidence of an immune response

BackgroundGiven the considerable toxicity and modest benefit of adjuvant chemotherapy for non-small cell lung cancer (NSCLC), there is clearly a need for new treatment modalities in the adjuvant setting. Active specific immunotherapy may represent such an option. However, clinical responses have been rare so far. Manipulating the host by inducing lymphopenia before vaccination resulted in a magnification of the immune response in the preclinical setting. To evaluate feasibility and safety of an irradiated, autologous tumor cell vaccine given following induction of lymphopenia by chemotherapy and reinfusion of autologous peripheral blood mononuclear cells (PBMC), we are currently conducting a pilot-phase I clinical trial in patients with NSCLC following surgical resection. This paper reports on the first clinical experience and evidence of an immune response in patients suffering from NSCLC.MethodsNSCLC patients stages I-IIIA are recruited. Vaccines are generated from their resected lung specimens. Patients undergo leukapheresis to harvest their PBMC prior to or following the surgical procedure. Furthermore, patients receive preparative chemotherapy (cyclophosphamide 350 mg/m2 and fludarabine 20 mg/m2 on 3 consecutive days) for induction of lymphopenia followed by reconstitution with their autologous PBMC. Vaccines are administered intradermally on day 1 following reconstitution and every two weeks for a total of up to five vaccinations. Granulocyte-macrophage-colony-stimulating-factor (GM-CSF) is given continuously (at a rate of 50 μg/24 h) at the site of vaccination via minipump for six consecutive days after each vaccination.ResultsTo date, vaccines were successfully manufactured for 4 of 4 patients. The most common toxicities were local injection-site reactions and mild constitutional symptoms. Immune responses to chemotherapy, reconstitution and vaccination are measured by vaccine site and delayed type hypersensitivity (DTH) skin reactions. One patient developed positive DTH skin tests so far. Immunohistochemical assessment of punch biopsies taken at the local vaccine site reaction revealed a dense lymphocyte infiltrate. Further immunohistochemical differentiation showed that CD1a+ cells had been attracted to the vaccine site as well as predominantly CD4+ lymphocytes. The 3-day combination chemotherapy consisting of cyclophosphamide and fludarabine induced a profound lymphopenia in all patients. Sequential FACS analysis revealed that different T cell subsets (CD4, CD8, CD4CD25) as well as granulocytes, B cells and NK cells were significantly reduced. Here, we report on clinical safety and feasibility of this vaccination approach during lymphoid recovery and demonstrate a patient example.ConclusionThus far, all vaccines were well tolerated. The overall trial design seems safe and feasible. Vaccine site reactions associated with infusion of GM-CSF via mini-pump are consistent with the postulated mechanism of action. More detailed immune-monitoring is required to evaluate a potential systemic immune response. Further studies to exploit homeostasis-driven T cell proliferation for the induction of a specific anti-tumor immune response in this clinical setting are warranted.

Failure of Kupffer cell blockade to prevent disseminated intravascular coagulation in endotoxemic rats despite improved survival

Objective: Studies were conducted to evaluate the impact of gadolinium chloride (GdCl3), an agent which blocks the phagocytosis of liver macrophages (Kupffer cells, KC), on the coagulation system and on mortality in a model of rats subjected to a lethal dose of Escherichia coli lipopolysaccharide (LPS) (10 mg/kg body weight, intravenously). Methods: Rats were either pretreated with GdCl3 (10 mg/kg, i.v., 48 h and 24 h prior to LPS exposure) or saline vehicle. A variety of coagulation parameters such as activated partial prothrombin time (aPTT), fibrinogen, systemic platelet count, antithrombin III (AT III), and activities of factors V, VII, and XII were monitored in the early (1 h) and late time course (16 h) following administration of E.coli LPS. Results: The administration of LPS resulted in the development of disseminated intravascular coagulation (DIC) and was associated with a mortality rate of 47% within 16 h. Blockade of KC by GdCl3 completely abolished LPS-related mortality (0%). However, despite improved survival, GdCl3 failed to prevent laboratory and clinical signs of DIC. GdCl3 per se even contributed to coagulatory and fibrinolytic disorders. Conclusion: These results confirm reports on the protective potential of GdCl3 pretreatment in experimental endotoxemia. However, the present study does not support the concept of DIC as a strong prognostic criterion for the outcome of sepsis and septic shock. Furthermore, the results presented suggest a minor role for KC in LPS-mediated activation of coagulation and indicate an involvement of KC in LPS-associated lethality independent of the coagulation system.