Preparation of Neurospora crassa mitochondria

The fungus Neurospora crassa represents a eukaryotic cell with high biosynthetic activities. Cell mass doubles in 2-4 hr during expone ntial growth , even in simple salt media with sucrose as the sole carbon source. The microorgani sm forms a mycelium of long hyphae durlng vegetative growth . The mitochondria can be isolated under relatively gentle condi tions since a few breaks in the threadlike hyphae are sufficient to cause the outflow of the organelles. This article describes two methods for the physical disruption of the hyphae : (I) The cell s are opened in a grind mill between two rotating corundum di sks. This is a continuous and fast procedure and allows large- and small-scale preparations of mitochondria. (2) Hyphae are ground with sand in a mortar and pestle. This procedure can be applied to microscale preparations of mitochondria starting with minute amounts of cells. Other procedures for the isolation of Neurospora mitochondria after the physical di sruption or the enzymatic degradation of the cell wall have been described elsewher

Virtual 3D tumor marking-exact intraoperative coordinate mapping improve post-operative radiotherapy

The quality of the interdisciplinary interface in oncological treatment between surgery, pathology and radiotherapy is mainly dependent on reliable anatomical three-dimensional (3D) allocation of specimen and their context sensitive interpretation which defines further treatment protocols. Computer-assisted preoperative planning (CAPP) allows for outlining macroscopical tumor size and margins. A new technique facilitates the 3D virtual marking and mapping of frozen sections and resection margins or important surgical intraoperative information. These data could be stored in DICOM format (Digital Imaging and Communication in Medicine) in terms of augmented reality and transferred to communicate patient's specific tumor information (invasion to vessels and nerves, non-resectable tumor) to oncologists, radiotherapists and pathologists

Telomerase-pulsed dendritic cells: preclinical results and outcome of a clinical phase I/II trial in patients with metastatic renal cell carcinoma

Objective: Therapeutic vaccination with dendritic cells (DC) showed promising results in first clinical trials in cases of metastatic renal cell carcinoma (RCC). Human telomerase reverse transcriptase (hTERT) could be a potential target because it is detectable in more than 85% of human tumors including RCC

Bladder cancer cells acquire competent mechanisms to escape Fas-mediated apoptosis and immune surveillance in the course of malignant transformation

Mechanisms of resistance against Fas-mediated cell killing have been reported in different malignancies. However, the biological response of immune escape mechanisms might depend on malignant transformation of cancer cells. In this study we investigated different mechanisms of immune escape in 2 well-differentiated low-grade (RT4 and RT112) and 2 poorly differentiated high-grade (T24 and TCCSUP) bladder cancer cell lines. Fas, the receptor of Fas-ligand, is expressed and shedded by human transitional bladder carcinoma cell lines RT4, RT112, T24 and TCCSUP. Cytotoxicity and apoptosis assays demonstrate that in spite of the Fas expression, poorly differentiated T24 and TCCSUP cells are insensitive towards either recombinant Fas-ligand or agonistic apoptosis-inducing monoclonal antibody against Fas. In poorly differentiated T24 and TCCSUP cell lines we were able to detect marked Fas-ligand protein by flow cytometry and Western blot analysis. In grade 1 RT4 and RT112 cells only minor expression of Fas-ligand possibly because of proteinase action. Fas-ligand mRNA translation or post-translational processing seems to be regulated differentially in the cancer cell lines depending on malignant transformation. In co-culture experiments we show that poorly differentiated cells can induce apoptosis and cell death in Jurkat cells and activated peripheral blood mononuclear cells. This in vitro study suggests that bladder cancer cells can take advantage of different mechanisms of immune evasion and become more competent in avoiding immune surveillance during transformation to higher-grade malignant disease. © 2001 Cancer Research Campaign www.bjcancer.co

Designing the ideal model for assessment of wound contamination after gunshot injuries: a comparative experimental study

<p>Abstract</p> <p>Background</p> <p>Modern high-velocity projectiles produce temporary cavities and can thus cause extensive tissue destruction along the bullet path. It is still unclear whether gelatin blocks, which are used as a well-accepted tissue simulant, allow the effects of projectiles to be adequately investigated and how these effects are influenced by caliber size.</p> <p>Method</p> <p>Barium titanate particles were distributed throughout a test chamber for an assessment of wound contamination. We fired .22-caliber Magnum bullets first into gelatin blocks and then into porcine hind limbs placed behind the chamber. Two other types of bullets (.222-caliber bullets and 6.5 × 57 mm cartridges) were then shot into porcine hind limbs. Permanent and temporary wound cavities as well as the spatial distribution of barium titanate particles in relation to the bullet path were evaluated radiologically.</p> <p>Results</p> <p>A comparison of the gelatin blocks and hind limbs showed significant differences (<it>p </it>< 0.05) in the mean results for all parameters. There were significant differences between the bullets of different calibers in the depth to which barium titanate particles penetrated the porcine hind limbs. Almost no particles, however, were found at a penetration depth of 10 cm or more. By contrast, gas cavities were detected along the entire bullet path.</p> <p>Conclusion</p> <p>Gelatin is only of limited value for evaluating the path of high-velocity projectiles and the contamination of wounds by exogenous particles. There is a direct relationship between the presence of gas cavities in the tissue along the bullet path and caliber size. These cavities, however, are only mildly contaminated by exogenous particles.</p

Assessment of therapeutic responses to gametocytocidal drugs in Plasmodium falciparum malaria.

Indirect clinical measures assessing anti-malarial drug transmission-blocking activity in falciparum malaria include measurement of the duration of gametocytaemia, the rate of gametocyte clearance or the area under the gametocytaemia-time curve (AUC). These may provide useful comparative information, but they underestimate dose-response relationships for transmission-blocking activity. Following 8-aminoquinoline administration P. falciparum gametocytes are sterilized within hours, whereas clearance from blood takes days. Gametocytaemia AUC and clearance times are determined predominantly by the more numerous female gametocytes, which are generally less drug sensitive than the minority male gametocytes, whereas transmission-blocking activity and thus infectivity is determined by the more sensitive male forms. In choosing doses of transmission-blocking drugs there is no substitute yet for mosquito-feeding studies

Epigenetic therapy in urologic cancers: an update on clinical trials

Epigenetic dysregulation is one of many factors that contribute to cancer development and progression. Numerous epigenetic alterations have been identified in urologic cancers including histone modifications, DNA methylation changes, and microRNA expression. Since these changes are reversible, efforts are being made to develop epigenetic drugs that restore the normal epigenetic patterns of cells, and many clinical trials are already underway to test their clinical potential. In this review we analyze multiple clinical trials (n=51) that test the efficacy of these drugs in patients with urologic cancers. The most frequently used epigenetic drugs were histone deacetylase inhibitors followed by antisense oligonucleotides, DNA methyltransferase inhibitors and histone demethylase inhibitors, the last of which are only being tested in prostate cancer. In more than 50% of the clinical trials considered, epigenetic drugs were used as part of combination therapy, which achieved the best results. The epigenetic regulation of some cancers is still matter of research but will undoubtedly open a window to new therapeutic approaches in the era of personalized medicine. The future of therapy for urological malignancies is likely to include multidrug regimens in which epigenetic modifying drugs will play an important role

MicroRNAs in Renal Cell Carcinoma: Diagnostic Implications of Serum miR-1233 Levels

BACKGROUND: MicroRNA expression is altered in cancer cells, and microRNAs could serve as diagnostic/prognostic biomarker for cancer patients. Our study was designed to analyze circulating serum microRNAs in patients with renal cell carcinoma (RCC). METHODOLOGY/PRINCIPAL FINDINGS: We first explored microrna expression profiles in tissue and serum using taqman low density arrays in each six malignant and benign samples: Although 109 microRNAs were circulating at higher levels in cancer patients' serum, we identified only 36 microRNAs with up-regulation in RCC tissue and serum of RCC patients. Seven candidate microRNAs were selected for verification based on the finding of up-regulation in serum and tissue of RCC patients: miR-7-1*, miR-93, miR-106b*, miR-210, miR-320b, miR-1233 and miR-1290 levels in serum of healthy controls (n = 30) and RCC (n = 33) patients were determined using quantitative real-time PCR (TaqMan MicroRNA Assays). miR-1233 was increased in RCC patients, and thus validated in a multicentre cohort of 84 RCC patients and 93 healthy controls using quantitative real-time PCR (sensitivity 77.4%, specificity 37.6%, AUC 0.588). We also studied 13 samples of patients with angiomyolipoma or oncocytoma, whose serum miR-1233 levels were similar to RCC patients. Circulating microRNAs were not correlated with clinical-pathological parameters. CONCLUSIONS/SIGNIFICANCE: MicroRNA levels are distinctly increased in cancer patients, although only a small subset of circulating microRNAs has a tumor-specific origin. We identify circulating miR-1233 as a potential biomarker for RCC patients. Larger-scaled studies are warranted to fully explore the role of circulating microRNAs in RCC