Leg ulcers are a diagnostic and therapeutic challenge

Leg ulcers are common. They cause a substantial burden to the patient and society. However, there is no need for therapeutic nihilism. The target of leg ulcer therapy is the individual patient. To be treated in a rational and successful way, exact diagnosis of the underlying cause(s) and associated diseases is necessary. This can be done in the most effective way with an interdisciplinary approach. The collection of cases demonstrates the need for careful clinical investigation substantiated and supported by vascular, histopathologic, and microbiologic techniques whenever needed. It is difficult to heal every ulcer completely, but improvement of the medical situation as well as the quality of life of the patient is possible in most cases.     

The SlideReactor--a simple hollow fiber based bioreactor suitable for light microscopy

Most bioartificial liver support systems are based on hollow fiber capillaries within modified dialysis cartridges or more sophisticated bioreactor constructions. Due to their design microscopic follow-up of reorganization and growth of tissue between the hollow fibers is not possible. The SlideReactor is a simple hollow fiber based bioreactor construction suitable for light microscopy and time-lapse video observation. The SlideReactor offers a cell compartment separated from a medium inflow and outflow compartment. Cell compartment access ports enable easy filling of the cell compartment with cell suspension, as well as fixation of the tissue. For more complex procedures or full access to all the cells, the bioreactor can be opened easily by cutting the silicone seal with a scalpel. Due to its simple design and the utilization of standard materials, it could serve as a suitable, cost-efficient tool to evaluate the behavior of cells cultured between hollow fiber capillaries. The paper describes the production process: similar to open source projects in software engineering, we would like to propose the concept as an open platform to anyone interested in hollow fiber based cell culture.     

"Blogs" and "wikis" are valuable software tools for communication within research groups

Appropriate software tools may improve communication and ease access to knowledge for research groups. A weblog is a website which contains periodic, chronologically ordered posts on a common webpage, whereas a wiki is hypertext-based collaborative software that enables documents to be authored collectively using a web browser. Although not primarily intended for use as an intranet-based collaborative knowledge warehouse, both blogs and wikis have the potential to offer all the features of complex and expensive IT solutions. These tools enable the team members to share knowledge simply and quickly-the collective knowledge base of the group can be efficiently managed and navigated.     

Functional and molecular characterization of interleukin-2 transgenic Ewing tumor cells for in vivo immunotherapy

BACKGROUND: Interleukin-2 (IL-2) is a potent cytokine with potential activity against several tumors including Ewing tumors (ET). Side effects of systemic IL-2 can be circumvented by the use of transgenic tumor cells. However, in vitro manipulation may change the overall gene expression profile of tumor cells unfavorably. Therefore, we assessed gene expression profiles, safety, and immunomodulatory efficacy of IL-2 transgenic (IL-2-tg) ET cells in vitro and in NOD/scid mice. PROCEDURE: Viable wild type A673 tumor cells were co-cultured together with irradiated IL-2-tg or mock-transfected cells and HLA matched peripheral blood mononuclear cells. Activation of T and NK cells was assessed by FACS analysis. The effect of irradiated IL-2-tg cells on tumor growth in vivo was investigated by using NOD/scid mice. Gene expression profiles of wild type and transfected cells were analyzed with Affymetrix HG-U95A microarrays. RESULTS: IL-2-tg cells activated and increased the number of T cells and NK cells in vitro. Co-culture with IL-2-tg but not with mock-transfected cells almost completely suppressed wild type tumor cell growth in vitro. Cell depletion experiments indicated a major contribution of NK cells to this tumor cell suppression. Co-transfer of irradiated IL-2-tg cells significantly reduced wild type tumor growth in NOD/scid mice. Side effects in the treated animals were not observed and no tumor growth was observed after injection of irradiated IL-2-tg cells alone. Gene expression profiling revealed a substantial degree of homogeneity of gene expression in transfected and wild type cells and suggests that transfection and selection procedures had no major impact on the gene expression profile. CONCLUSIONS: Next to a high degree of homogeneity between transgenic and wild type cells, our data suggest that irradiated IL-2-tg ET cells can activate cytolytic effector cells. These cells may have therapeutic potential for ET patients.     

Evaluation of primary human liver cells in bioreactor cultures for extracorporeal liver support on the basis of urea production

Primary human liver cells from donor organs unsuitable for transplantation were cultivated in bioreactors developed for extracorporeal liver support. Because each system contains cells originating from an individual organ, each bioreactor culture must be individually characterized. The objective of this study was to identify suitable decisive parameters for the evaluation of cell culture performance. We analyzed the data from 47 bioreactor cultures containing 437 +/- 110 g of cells. Choosing urea production as the decisive parameter, the bioreactor cultures were divided into high-performance (daily urea production > or = 110 mg per bioreactor between culture days 3 and 14) and low-performance cultures. Comparing the mean courses of the groups revealed a significant distinction in most other investigated biochemical parameters. In conclusion, urea production seems to be an appropriate parameter for evaluating the performance of liver cell cultures in bioreactors because it corresponds to all other evaluated parameters of cell function.     

CpG island methylation and expression of tumour-associated genes in lung carcinoma

In this study, microarray analysis was used to identify tumour-related genes that were down regulated in lung carcinoma. The promoter sequences of the identified genes were analysed for methylation patterns. In lung cancer cell lines, CpG island methylation was frequently detected for TIMP4 (64%), SOX18 (73%), EGF-like domain 7 (56%), CD105 (71%), SEMA2 (55%), RASSF1A (71%), p16 (56%) SLIT2 (100%) and TIMP3 (29%). Methylation was however rarely observed in cell lines for SLIT3 (18%) and DLC1 (18%). In primary lung tumours, methylation of TIMP4 (94%), SOX18 (100%), EGF-like domain 7 (100%), CD105 (69%), SEMA2 (93%), DLC1 (61%), RASSF1A (44%), p16 (47%), SLIT2 (100%) and TIMP3 (13%) was also detected. Methylation of several CpG islands was frequently found in normal lung tissue of cancer patients and this may have been attributed to epigenetic field defect and/or infiltrating tumour cells. Interestingly, inactivation of RASSF1A and p16 correlated well with an extended smoking habit (P=0.02), and exposure to asbestos (P=0.017) or squamous cell carcinoma (P=0.011), respectively. These results have identified genes whose aberrant promoter methylation could play a crucial role in the malignancy of lung carcinoma.     

Effects of combined therapy with clopidogrel and acetylsalicylic acid on platelet glycoprotein expression and aggregation

SUMMARY: This study aimed to compare the effects of clopidogrel, acetylsalicylic acid (ASA), and the combination of both substances on platelet aggregation and expression of platelet membrane glycoproteins in patients with chronic coronary artery disease. We investigated platelet activation by flow cytometry and by platelet aggregation and disaggregation in 60 patients randomly assigned to 3 treatment groups: ASA, clopidogrel, combination of clopidogrel and ASA, treated for 14 days. Adenosine diphosphate (ADP)-induced expression of P-selectin and of PAC-1 was significantly reduced after 2 wk of clopidogrel but not of ASA treatment. Treatment with clopidogrel reduced the ADP-induced platelet aggregation. The combination of clopidogrel and ASA did not increase the inhibition of platelet activation compared with clopidogrel alone. A significant increase in platelet disaggregation was observed with clopidogrel alone and was more pronounced with the combination of clopidogrel and ASA. ADP-induced platelet degranulation, activation of GPIIb/IIIa receptor, and aggregation in vivo are effectively inhibited by clopidogrel. The significantly increased disaggregation under clopidogrel and ASA suggests that the combined therapy may be superior to the monotherapy in patients with coronary artery disease and a high risk for vascular events.     

Nuclei of chicken neurons in tissues and three-dimensional cell cultures are organized into distinct radial zones

We used chicken retinospheroids (RS) to study the nuclear architecture of vertebrate cells in a three-dimensional (3D) cell culture system. The results showed that the different neuronal cell types of RS displayed an extreme form of radial nuclear organization. Chromatin was arranged into distinct radial zones which became already visible after DAPI staining. The distinct zones were enriched in different chromatin modifications and in different types of chromosomes. Active isoforms of RNA polymerase II were depleted in the outermost zone. Also chromocenters and nucleoli were radially aligned in the nuclear interior. The splicing factor SC35 was enriched at the central zone and did not show the typical speckled pattern of distribution. Evaluation of neuronal and non-neuronal chicken tissues showed that the highly ordered form of radial nuclear organization was also present in neuronal chicken tissues. Furthermore, the data revealed that the neuron-specific nuclear organization was remodeled when cells spread on a flat substrate. Monolayer cultures of a chicken cell line did not show this extreme form of radial organization. Rather, such monolayer cultures displayed features of nuclear organization which have been described before for many different types of monolayer cells. The finding that an extreme form radial nuclear organization, which has not been described before, is present in RS and tissues, but not in cells spread on a flat substrate, suggests that it would be important to complement studies on nuclear architecture performed with monolayer cells by studies on 3D cell culture systems and tissues.