Modulation of Mycobacterium bovis-Specific Responses of Bovine Peripheral Blood Mononuclear Cells by 1,25-Dihydroxyvitamin D3

Historically, administration of vitamin D has been considered beneficial in the treatment of tuberculosis. The interaction of this vitamin {i.e., 1,25-dihdroxyvitamin D₃ [1,25(OH)₂D₃]} with the antitubercular immune response, however, is not clear. In the present study, in vitro recall responses of peripheral blood mononuclear cells (PBMC) from cattle infected with Mycobacterium bovis were used to study the immune-modulatory effects of 1,25(OH)₂D₃ on M. bovis-specific responses in vitro. Addition of 1 or 10 nM 1,25(OH)₂D₃ inhibited M. bovis-specific proliferative responses of PBMC from M. bovis-infected cattle, affecting predominately the CD4⁺ cell subset. In addition, 1,25(OH)₂D₃ inhibited M. bovis-specific gamma interferon (IFN-γ) production yet enhanced M. bovis-specific nitric oxide (NO) production. Lymphocyte apoptosis, measured by flow cytometry using annexin-V staining, was diminished by addition of 1,25(OH)₂D₃ to PBMC cultures. These findings support the current hypothesis that 1,25(OH)₂D₃ enhances mycobacterial killing by increasing NO production, a potent antimicrobial mechanism of activated macrophages, and suggest that 1,25(OH)₂D₃ limits host damage by decreasing M. bovis-induced IFN-γ production.     

C1-Inhibitor for treatment of acute vascular xenograft rejection in cynomolgus recipients of h-DAF transgenic porcine kidneys

Abstract: At present, the major barrier to successful discordant xenotransplantation of unmodified or complement regulator transgenic porcine xenografts is acute vascular xenograft rejection (AVR). AVR is associated with the intragraft deposition of induced recipient xenoreactive antibodies and subsequent complement activation. In a life-supporting pig to primate kidney xenotransplantation setting using h-DAF transgenic donor organs and postoperative immunosuppression, episodes of AVR were either treated with boluses of cyclophosphamide and steroids or with the same regimen supplemented by a three-day course of C1-Inhibitor, a multifunctional complement regulator. In 8 out of 10 animals stable initial graft function was achieved; in all animals one or more episodes of AVR were observed. When, in 4 animals, C1-Inhibitor was added to the standard anti-rejection treatment regimen, AVR was successfully reversed in 6 out of 7 episodes, while in another group of 4 animals receiving the standard anti-rejection treatment 0 out of 4 episodes of AVR responded to treatment. Response to anti-rejection treatment was associated with a significant increase in recipient survival time. We conclude that AVR of h-DAF transgenic porcine kidneys can be successfully treated by additional short-term fluid phase complement inhibition.     

Angiotensin-converting enzyme activity and the ACE Alu polymorphism in autosomal dominant polycystic kidney disease.

BACKGROUND: Previous studies concerning Alu I/D polymorphism in the ACE gene and ADPKD severity have used the Alu genotypes as a representative of the true biological variable, namely ACE activity. However, wide individual and ethnic differences in the proportion of variance in ACE activity explained by the I/D genotype may have confounded these studies. This investigation examines the association between ADPKD severity and ACE in terms of plasma enzyme activity and I/D genotypes in individuals from three different countries. METHODS: Blood samples were collected from 307 ADPKD patients (116 Australian, 124 Bulgarian and 67 Polish) for determination of ACE activity levels and I/D genotypes. Chronic renal failure (CRF) was present in 117 patients and end-stage renal failure (ESRF) in 68 patients. RESULTS: ACE activity was related to the I/D genotype, showing a dosage effect of the D allele (P=0.006). The proportion of variance due to the Alu polymorphism was 14%. No difference in ACE activity and I/D genotype distribution was found between patients with CRF versus normal renal function (P=0.494; P=0.576) or between those with ESRF versus those without ESRF (P=0.872; P=0.825). No effect of the I/D genotype on age at development and progression to renal failure (CRF; ESRF) was detected in the overall group, and in subgroups based on ethnic origin, linkage status and sex. CONCLUSION: ACE is not likely to play a role as a determinant of ADPKD phenotype severity.     

Fas expression on peripheral blood lymphocytes in systemic lupus erythematosus (SLE): relation to lymphocyte activation and disease activity.

Levels of apoptotic lymphocytes have been found to be increased in SLE and persistence of apoptotic cells has been associated with autoantibody production. Increased lymphocyte Fas (CD95) expression due to lymphocyte activation may account for increased susceptibility to Fas-mediated apoptosis in SLE. Flowcytometry was performed to evaluate membrane expression of Fas in combination with the activation markers CD25, HLA-DR and CD38 on, respectively, CD4+, CD8+ and CD19+ lymphocytes of SLE patients with inactive (n = 20) and with active disease (n = 13). SLEDAI-scores were calculated. Healthy volunteers (n = 14) served as controls. Percentages of CD4+ T-cells expressing CD25 and CD19+ B-cells expressing CD38 were increased in patients with active disease compared to controls (P = 0.03, P = 0.04, respectively). In contrast to CD4+ and CD8+ cells, percentages of CD19+ cells expressing Fas were increased in SLE patients with active disease (P = 0.0002 vs controls). In these patients percentages of cells double positive for both CD38 and Fas were increased compared to patients with inactive disease (P = 0.006) and controls (P = 0.0007). Percentages of CD19+ cells expressing Fas correlated with SLEDAI-scores. In SLE patients, percentages of Fas-expressing B-lymphocytes are increased, are related to the state of lymphocyte activation, and correlate to disease activity. Increased Fas expression results in a higher susceptibility for Fas-mediated apoptosis, which might contribute to the increased levels of apoptotic lymphocytes in SLE patients.     

Patient Position Reproducibility in Fractionated Stereotactically Guided Conformal Radiotherapy Using the BrainLab? Mask System

Purpose: Dedicated mask systems nowadays allow the use of stereotactic radiotherapy in fractionated regimes, therefore combining the advantages of high precision radiotherapy with the biological benefit of fractionation. Therefore the knowledge of institution specific isocenter accuracy is essential for decision-making about margins to be allowed to form the planning target volume. Patients and Methods: Measurements of isocenter deviations during fractionated treatments were performed in 33 patients using the simulator Simulix-xy (Oldelft) in connection with the BrainLab^® angiographic localizer-box as well as port-films. In both cases repeated images were overlaid by use of anatomical landmarks with a methodical accuracy in the order of 0.5 mm. Results: Both methods yield random isocenter deviations of less then 2 mm (standard deviation) in all three directions and no significant systematic deviations. These values are in the order of the accuracy of the method, obtained by comparison of two independent investigators, as well as they are comparable with the literature. Conclusions: The accuracy of less than 2 mm indicates safety margins of 3-4 mm as sufficient for clinical routine to cover the target in 95.5% of all set-ups (2 SD). Ziel: Spezielle Maskensysteme erlauben heutzutage die Anwendung der stereotaktischen Strahlentherapie in fraktionierten Regimes und damit die Kombination der Vorteile der Hochpräzisionsbestrahlung mit dem biologischen Nutzen der Fraktionierung. Deshalb ist die Kenntnis der institutsspezifischen Genauigkeit der Isozentrumseinstellung eine notwendige Voraussetzung für die Entscheidung über die erforderlichen Sicherheitsabstände. Patienten und Methode: Die Messung der Isozentrumsgenauigkeit erfolgte bei 33 Patienten sowohl durch Simulatorkontrollen in Verbindung mit der BrainLab^®-Localizer-Box oder durch Verifikation mit Portfilmen. In beiden Fällen wurden die Filme anhand anatomischer Lankdmarken mit einer methodischen Sicherheit von unter 0,5 mm überlagert. Ergebnisse: Beide Methoden zeigten zufällige Isozentrumsabweichungen von weniger als 2 mm (Standardabweichung) in allen drei Raumebenen (Abbildung 1, Tabelle 2) und keine signifikanten systematischen Abweichungen. Damit liegen die Ergebnisse im Bereich der methodischen Genauigkeit, wie durch den Vergleich der Befunde zweier unabhängiger Untersucher gezeigt wird (Tabelle 1), und sind mit Literaturdaten gut vergleichbar. Schlussfolgerung: Die Genauigkeit von unter 2 mm zeigt, dass ein Sicherheitsabstand von 3-4 mm für die klinische Routine ausreichend ist, um bei 95,5% der Einstellung die sichere Erfassung des Targets zu garantieren.     

Patient Position Reproducibility in Fractionated Stereotactically Guided Conformal Radiotherapy Using the BrainLab® Mask System

PURPOSE: Dedicated mask systems nowadays allow the use of stereotactic radiotherapy in fractionated regimes, therefore combining the advantages of high precision radiotherapy with the biological benefit of fractionation. Therefore the knowledge of institution specific isocenter accuracy is essential for decision-making about margins to be allowed to form the planning target volume. PATIENTS AND METHOD: Measurements of isocenter deviations during fractionated treatments were performed in 33 patients using the simulator Simulix-xy (Oldelft) in connection with the BrainLab angiographic localizer-box as well as port-films. In both cases repeated images were overlaid by use of anatomical landmarks with a methodical accuracy in the order of 0.5 mm. RESULTS: Both methods yield random isocenter deviations of less then 2 mm (standard deviation) in all three directions and no significant systematic deviations. These values are in the order of the accuracy of the method, obtained by comparison of two independent investigators, as well as they are comparable with the literature. CONCLUSIONS: The accuracy of less than 2 mm indicates safety margins of 3-4 mm as sufficient for clinical routine to cover the target in 95.5% of all set-ups (2 SD).     

Lokale Tumorkontrolle und kosmetisches Ergebnis nach brusterhaltender Operation und Strahlentherapie bis 56 Gy Gesamtdosis ohne Boost beim Mammakarzinom

PURPOSE: To evaluate overall survival, local tumor control and cosmetic outcome after breast-conserving surgery followed by radiotherapy without boost irradiation. PATIENTS AND METHODS: In a retrospective study 270 breast cancer patients were treated with breast conserving surgery combined with a homogenous radiation of the tumor bearing breast up to a total dose of 56 Gy without local boost irradiation. Mean follow-up was 48 months. Local tumor control, side effects, cosmetic results and contentment with treatment were assessed using physical examinations and interviews based on a standardized questionnaire. RESULTS: Cause-specific survival at 5 years after treatment was 88.3%, actuarial disease-free survival at 5 years was 76.1%. Within 23 to 78 months after treatment 12 patients suffered from ipsilateral breast recurrence. The actuarial freedom from local recurrence (single tumor manifestation) was 96.8% at 5 years after treatment, 89% at 10 years. The occurrence of local failures was not significantly correlated to tumor size, margins, grading, nodal status, age or lymphangiosis. 15.6% of the patients developed distant metastases. In all patients treatment was performed without interruption. Side effects were predominantly of mild degree, no severe side effects were detected. 73% of physicians and 81% of patients scored their cosmetic outcome as excellent or good. 93% of patients would again decide in favor of this procedure. Whereas use of adjuvant chemotherapy as well as subcutaneous reconstruction of breast tissue did not significantly affect breast cosmesis, analysis demonstrated impaired cosmetic results related to a larger breast size. CONCLUSION: The data of this study show that tumor control achieved by breast conserving surgery in combination with a radiation technique up to a total dose of 56 Gy which omits boost irradiation is within the range of literature data. Side effects of the therapy were tolerable. The treatment displayed a good compatibility with tolerable side effects and good cosmetic results.     

Evaluation of Angiogenesis Using Micro-Computed Tomography in a Xenograft Mouse Model of Lung Cancer

Quantitative evaluation of lung tumor angiogenesis using immunohistochemical techniques has been limited by difficulties in generating reproducible data. To analyze intrapulmonary tumor angiogenesis, we used high-resolution micro-computed tomography (micro-CT) of lung tumors of mice inoculated with mouse Lewis lung carcinoma (LLC1) or human adenocarcinoma (A549) cell lines. The lung vasculature was filled with the radiopaque silicone rubber, Microfil, through the jugular vein (in vivo application) or pulmonary artery (ex vivo application). In addition, human adenocarcinoma lung tumor-bearing mice treated site-specifically with humanized monoclonal antibody (bevacizumab) against vascular endothelial growth factor. Quantitative analysis of lung tumor microvessels imaged with micro-CT showed that more vessels (mainly small, <0.02 mm²) were filled using the in vivo (5.4%) compared with the ex vivo (2.1%) method. Furthermore, bevacizumab-treated lung tumor-bearing mice showed significantly reduced lung tumor volume and lung tumor angiogenesis compared with untreated mice as assessed by micro-CT. Interestingly, microvascularization of mainly the smaller vessels (<0.02 mm²) was reduced after bevacizumab treatment. This observation with micro-CT was nicely correlated with immunohistochemical measurement of microvessels. Therefore, micro-CT is a novel method for investigating lung tumor angiogenesis, and this might be considered as an additional complementary tool for precise quantification of angiogenesis.     

Imaging of integrin αvβ3 expression in patients with malignant glioma by [18F] Galacto-RGD positron emission tomography

Inhibitors targeting the integrin α_vβ₃ are promising new agents currently tested in clinical trials for supplemental therapy of glioblastoma multiforme (GBM). The aim of our study was to evaluate ¹⁸F-labeled glycosylated Arg-Gly-Asp peptide ([¹⁸F]Galacto-RGD) PET for noninvasive imaging of α_vβ₃ expression in patients with GBM, suggesting eligibility for this kind of additional treatment. Patients with suspected or recurrent GBM were examined with [¹⁸F]Galacto-RGD PET. Standardized uptake values (SUVs) of tumor hotspots, galea, and blood pool were derived by region-of-interest analysis. [¹⁸F]Galacto-RGD PET images were fused with cranial MR images for image-guided surgery. Tumor samples taken from areas with intense tracer accumulation in the [¹⁸F]Galacto-RGD PET images and were analyzed histologically and immunohistochemically for α_vβ₃ integrin expression. While normal brain tissue did not show significant tracer accumulation (mean SUV, 0.09 ± 0.04), GBMs demonstrated significant but heterogeneous tracer uptake, with a maximum in the highly proliferating and infiltrating areas of tumors (mean SUV, 1.6 ± 0.5). Immunohistochemical staining was prominent in tumor microvessels as well as glial tumor cells. In areas of highly proliferating glial tumor cells, tracer uptake (SUVs) in the [¹⁸F]Galacto-RGD PET images correlated with immunohistochemical α_vβ₃ integrin expression of corresponding tumor samples. These data suggest that [¹⁸F] Galacto-RGD PET successfully identifies α_vβ₃ expression in patients with GBM and might be a promising tool for planning and monitoring individualized cancer therapies targeting this integrin.     

Mortality among Thorotrast-exposed patients and an unexposed comparison group in the German Thorotrast study

Thorotrast was the brand name of a stabilised colloidal solution of thorium dioxide which was used preferentially as an X-ray contrast medium for arteriography between 1930 and 1950. The administration of the medium led to lifelong chronic α-particle irradiation by thorium decay products, mainly in the organs of deposition. Several epidemiological follow-up studies were set up after recognition of these side-effects among which the German study was the largest. After an extended follow-up, by 2004 only nine out of 2326 originally exposed subjects were still alive (while 151 of the comparison group, which originally numbered 1890 subjects, survived) and partially more than 70 years observation and chronic exposure time could be studied allowing for further observations to be made about long-term mortality effects of Thorotrast exposure. Median life-expectancy was shortened by 14 years and mortality increased, affecting total mortality SMR = 287 for males, SMR = 387 for females) as well as cause-specific, especially liver cancer (SMR = 16,695 and SMR = 12,680, respectively), and the haematopoietic system (SMR = 556 and SMR = 504, respectively), but not lung cancer. Mortality (total and selected cause-specific) increased with cumulative time since first exposure.