Jörg Lindenmann

Extracellular concentrations of fosfomycin in lung tissue of septic patients

OBJECTIVES The present investigation explored the ability of fosfomycin to penetrate lung tissue of septic patients by utilizing the microdialysis technique. METHODS After microdialysis probe insertion into healthy and infected lung tissue, a single intravenous dose of 4 g of fosfomycin was administered. RESULTS The mean C(max), T(max), AUC(0-4) and AUC(0-infinity) for healthy lung were 131.6 +/- 110.6 mg/L, 1.1 +/- 0.4 h, 242.4 +/- 101.6 mgxh/L and 367.6 +/- 111.9 mgxh/L, respectively. The corresponding values for infected lung were 107.5 +/- 60.2 mg/L, 1.4 +/- 0.5 h, 203.5 +/- 118.4 mgxh/L and 315.1 +/- 151.2 mgxh/L. The half-life of fosfomycin ranged from 2.2 to 2.7 h between compartments. The magnitude of lung tissue penetration, as determined by the ratios of the AUC(0-infinity) for lung to the AUC(0-infinity) for plasma, was 0.63 +/- 0.31 and 0.53 +/- 0.31 for healthy and infected lung, respectively. CONCLUSIONS We conclude that fosfomycin achieves antimicrobially effective concentrations in infected lung tissue.

Docosahexaenoic acid-induced unfolded protein response, cell cycle arrest, and apoptosis in vascular smooth muscle cells are triggered by Ca2+-dependent induction of oxidative stress

Proliferation of vascular smooth muscle cells is a characteristic of pathological vascular remodeling and represents a significant therapeutic challenge in several cardiovascular diseases. Docosahexaenoic acid (DHA), a member of the n-3 polyunsaturated fatty acids, was shown to inhibit proliferation of numerous cell types, implicating several different mechanisms. In this study we examined the molecular events underlying the inhibitory effects of DHA on proliferation of primary human smooth muscle cells isolated from small pulmonary artery (hPASMCs). DHA concentration-dependently inhibited hPASMC proliferation, induced G1 cell cycle arrest, and decreased cyclin D1 protein expression. DHA activated the unfolded protein response (UPR), evidenced by increased mRNA expression of HSPA5, increased phosphorylation of eukaryotic initiation factor 2α, and splicing of X-box binding protein 1. DHA altered cellular lipid composition and led to increased reactive oxygen species (ROS) production. DHA-induced ROS were dependent on both intracellular Ca2+ release and entry of extracellular Ca2+. Overall cellular ROS and mitochondrial ROS were decreased by RU360, a specific inhibitor of mitochondrial Ca2+ uptake. DHA-induced mitochondrial dysfunction was evidenced by decreased mitochondrial membrane potential and decreased cellular ATP content. DHA triggered apoptosis as found by increased numbers of cleaved caspase-3- and TUNEL-positive cells. The free radical scavenger Tempol counteracted DHA-induced ROS, cell cycle arrest, induction of UPR, and apoptosis. We conclude that Ca2+-dependent oxidative stress is the central and initial event responsible for induction of UPR, cell cycle arrest, and apoptosis in DHA-treated hPASMCs.

Hypoxia increases membrane metallo-endopeptidase expression in a novel lung cancer ex vivo model – role of tumor stroma cells

BackgroundHypoxia-induced genes are potential targets in cancer therapy. Responses to hypoxia have been extensively studied in vitro, however, they may differ in vivo due to the specific tumor microenvironment. In this study gene expression profiles were obtained from fresh human lung cancer tissue fragments cultured ex vivo under different oxygen concentrations in order to study responses to hypoxia in a model that mimics human lung cancer in vivo.MethodsNon-small cell lung cancer (NSCLC) fragments from altogether 70 patients were maintained ex vivo in normoxia or hypoxia in short-term culture. Viability, apoptosis rates and tissue hypoxia were assessed. Gene expression profiles were studied using Affymetrix GeneChip 1.0 ST microarrays.ResultsApoptosis rates were comparable in normoxia and hypoxia despite different oxygenation levels, suggesting adaptation of tumor cells to hypoxia. Gene expression profiles in hypoxic compared to normoxic fragments largely overlapped with published hypoxia-signatures. While most of these genes were up-regulated by hypoxia also in NSCLC cell lines, membrane metallo-endopeptidase (MME, neprilysin, CD10) expression was not increased in hypoxia in NSCLC cell lines, but in carcinoma-associated fibroblasts isolated from non-small cell lung cancers. High MME expression was significantly associated with poor overall survival in 342 NSCLC patients in a meta-analysis of published microarray datasets.ConclusionsThe novel ex vivo model allowed for the first time to analyze hypoxia-regulated gene expression in preserved human lung cancer tissue. Gene expression profiles in human hypoxic lung cancer tissue overlapped with hypoxia-signatures from cancer cell lines, however, the elastase MME was identified as a novel hypoxia-induced gene in lung cancer. Due to the lack of hypoxia effects on MME expression in NSCLC cell lines in contrast to carcinoma-associated fibroblasts, a direct up-regulation of stroma fibroblast MME expression under hypoxia might contribute to enhanced aggressiveness of hypoxic cancers.

Photodynamic therapy enhanced by hyperbaric oxygenation in palliation of malignant pleural mesothelioma: clinical experience

INTRODUCTION Surgical debulking followed by radiotherapy/chemotherapy are the standards in the palliative treatment schedule of malignant pleural mesothelioma. The aim of this study was to evaluate the additional effect of intraoperative photodynamic therapy (PDT) under hyperbaric oxygenation (HBO) if compared to decortication alone. PATIENTS AND METHODS From January 1993 to August 2003, decortication was done in 34 patients (28 males, 6 females; mean age: 65 years) suffering from advanced malignant pleural mesothelioma. Twenty-two patients received additional intraoperative PDT under HBO. The surgery and PDT/HBO was done 48h after photosensitization with a polyhematoporphyrin, 2mg/kg BW using a diode laser delivering red light at 630nm through a microlens. The light dose was calculated for 300J at a distance of 1cm from the tumour surface. RESULTS At 6-month follow-up the Karnofsky performance status showed no significant difference (P≥0.05) between both groups. CT scans documented focal regrowth of the tumour after 6 months in 10/12 cases of the non-PDT group. However, in the PDT group tumour regrowth was detected in only 9/22 cases at 6-month follow-up. Survival analysis showed a significant advantage for the group with PDT (log-rank test: P=0.0179). CONCLUSION Although the study includes only a small number of patients, it indicates that additional PDT/HBO represents a safe and technically feasible approach in the palliative setting of advanced malignant mesothelioma of the pleura.

Method development and validation for the analysis of a new anti-cancer infusion solution via HPLC

A fast and simple HPLC method has been developed and validated for the quantification of a completely new anti-cancer drug during the manufacturing process. The combination of four compounds including α-ketoglutaric acid, hydroxymethylfurfural, N-acetyl-L-methionine and N-acetyl-L-selenomethionine, administered intravenously, is still in test phase but has already shown promising results in cancer therapy. HPLC separation was achieved on an RP-18 column with a gradient system. However, the highly different concentrations of the compounds required a variation in the detection wavelength within one run. In order to produce a chromatogram where peaks were comparable on a similar range scale, detection at absorption maxima for the two most concentrated components was avoided. After optimization of the gradient program it was possible to detect all four substances within 14 min in spite of their strongly different chemical structure. The method developed was validated for accuracy, repeatability, reproducibility and robustness in relation to temperature and pH of buffer. Linearity as well as the limit of detection and quantification were determined. This HPLC method was found to be precise, accurate and reproducible and can be easily used for in-line process control during the manufacture of the anti-tumour infusion solution.

TASK-1 Regulates Apoptosis and Proliferation in a Subset of Non-Small Cell Lung Cancers.

Lung cancer is the leading cause of cancer deaths worldwide; survival times are poor despite therapy. The role of the two-pore domain K+ (K2P) channel TASK-1 (KCNK3) in lung cancer is at present unknown. We found that TASK-1 is expressed in non-small cell lung cancer (NSCLC) cell lines at variable levels. In a highly TASK-1 expressing NSCLC cell line, A549, a characteristic pH- and hypoxia-sensitive non-inactivating K+ current was measured, indicating the presence of functional TASK-1 channels. Inhibition of TASK-1 led to significant depolarization in these cells. Knockdown of TASK-1 by siRNA significantly enhanced apoptosis and reduced proliferation in A549 cells, but not in weakly TASK-1 expressing NCI-H358 cells. Na+-coupled nutrient transport across the cell membrane is functionally coupled to the efflux of K+ via K+ channels, thus TASK-1 may potentially influence Na+-coupled nutrient transport. In contrast to TASK-1, which was not differentially expressed in lung cancer vs. normal lung tissue, we found the Na+-coupled nutrient transporters, SLC5A3, SLC5A6, and SLC38A1, transporters for myo-inositol, biotin and glutamine, respectively, to be significantly overexpressed in lung adenocarcinomas. In summary, we show for the first time that the TASK-1 channel regulates apoptosis and proliferation in a subset of NSCLC.

High extracellular levels of cefpirome in unaffected and infected lung tissue of patients

OBJECTIVES the objective of the present investigation was to measure the extracellular concentrations of cefpirome in unaffected and infected lung tissue of septic patients. METHODS a single intravenous dose of 30 mg/kg total body weight of cefpirome was administered to eight patients every 12 h prior to insertion of microdialysis probes into lung tissue. RESULTS the median (minimum, maximum) peak concentration (C(max)), time to C(max) (T(max)), area under the concentration-time curve from 0 to 4 h (AUC(0-4)) and AUC(0-∞) of unbound cefpirome for unaffected lung were 48 (32, 107) mg/L, 0.83 (0.17, 3.17) h, 117 (60, 177) mg · h/L and 182 (80, 382) mg · h/L, respectively. The corresponding values for infected lung tissue were 45 (6, 122) mg/L, 1.17 (0.83, 2.83) h, 92 (17, 253) mg · h/L and 206 (49, 379) mg · h/L, respectively. The median apparent terminal elimination half-lives (t(½z)) of cefpirome were 2.61, 3.05 and 3.39 h for plasma, unaffected lung and infected lung, respectively. The median ratios of the AUC(0)(-∞) for lung to the AUC(0)(-∞) for plasma were 0.63 (0.19, 1.55) and 0.46 (0.32, 0.98) for unaffected and infected lung, respectively. CONCLUSIONS we provide strong evidence that cefpirome penetrates effectively into the extracellular space fluid of lung tissue. Under steady-state conditions, the median concentrations of cefpirome in plasma, unaffected lung and infected lung exceeded the MICs of the majority of relevant bacteria over the entire dosing interval of up to 12 h after intravenous administration of a dose of 30 mg/kg total body weight.

Incidental finding of a splenic hamartoma with tumour-like extramedullary erythropoiesis.

ZusammenfassungHamartome der Milz sind sehr selten vorkommende gutartige tumoröse, meist asymptomatische Läsionen, wobei solche mit tumoröser extramedullärer Blutbildung extrem selten auftreten. Ein 70-jähriger männlicher Patient mit Dyspnoe und einem CT-verifizierten pulmonalen Rundherd unterzog sich einer Bronchoskopie, bei der ein Fremdkörper in Form einer Erbse erfolgreich entfernt werden konnte. Als Zufallsbefund zeigte sich in der CT eine Läsion im Bereich der Milz mit einem Durchmesser von ungefähr 8 cm. Eine MRT bestätigte einen expansiven Prozess im Milzzentrum. Der Patient war zu diesem Zeitpunkt völlig asymptomatisch. Aufgrund der unklaren Genese des Tumors und dem hohen Risiko einer spontanen Milzruptur wurde eine offene Splenektomie durchgeführt. Bei der histologischen Untersuchung des Operationspräparates wurde ein Hamartom der Milz mit tumoröser extramedullärer Blutbildung diagnostiziert. Der Patient konnte schließlich am zehnten postoperativen Tag in gutem Allgemeinzustand entlassen werden. Ein Hamartom der Milz mit extramedullärer tumoröser Erythropoese sollte als Differentialdiagnose einer unklaren Milzläsionen bedacht werden.SummaryHamartomas of the spleen are rare benign tumour-like lesions that tend to be asymptomatic incidental findings. Splenic hamartomas with tumour-like extramedullary erythropoiesis are extremely rare. A 70-year-old male patient with dyspnoea and a nodular intrapulmonary lesion morphologically verified by CT underwent fibre bronchoscopy, which showed a pea-shaped foreign body that was retrieved without complication. As a secondary finding, the CT showed a splenic lesion of about 8 cm in size. An MR study showed an expansive process in the center of the spleen. The patient was completely free of clinical symptoms. Because of the unknown nature and the high risk for spontaneous rupture of the lesion, a splenectomy was performed. The histological analyses revealed tumour-like extramedullary erythropoiesis of the spleen. The patient was discharged in good condition ten days postoperatively. A hamartoma of the spleen with extramedullary tumor-like erythropoiesis should be considered in the differential diagnosis of a splenic lesion.

5-Aminolaevulinic acid compared to polyhematoporphyrin photosensitization for photodynamic therapy of malignant bronchial and esophageal stenosis: clinical experience

BACKGROUND AND OBJECTIVE Polyhematoporphyrins (PhP) as sensitizers for photodynamic therapy (PDT) in malignant bronchial and esophageal stenosis carry the risk of prolonged photosensitivity of the skin. New line sensitizers such as 5-aminolaevulinic acid (ALA) with low rates of skin phototoxicity appear to be promising alternatives. The aim of this study was to evaluate the efficacy of ALA compared to PhP for PDT regarding phototoxicity of the skin, reduction of tumour stenosis and tumour length and Karnofsky performance status. PATIENTS AND METHODS After diagnostic work-up, photosensitization was done in 38 patients with ALA (60mg/kg body weight, oral, 6-8h prior to PDT) and in 51 patients with PhP (2mg/kg body weight, i.v., 48h before PDT). The light dose was calculated as 300J/cm fibre tip. Light at 630nm was applied using a pumped dye laser. In both groups, additional hyperbaric oxygenation was applied at a level of 2 bar absolute pressure. RESULTS Improvement regarding stenosis diameter, tumour length and Karnofsky performance status could be obtained in both treatment arms with a significant difference in favour of the PhP-group, P = 0.00073; 0.000014, and 0.00015, respectively. No sunburn or other major treatment related complications occurred in either treatment arms. CONCLUSION Photosensitization with PhP compared to ALA seems to be more effective in PDT of malignant bronchial and esophageal stenosis.