Three-Dimensional Imaging of Rhodamine 123 Fluorescence Distribution in Human Melanoma Cells by Means of Confocal Laser Scanning Microscopy

Porwol, T.; Merten, E.; Opitz, N.; Acker, H.

doi:10.1159/000147872

Cited by 13 publications

(14 citation statements)

References 1 publication

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Two new technical developments might improve the impact of the proposed action: 1. cultivation of individual tumor biopsy material in multicellular spheroid culture (Fjellbirkeland et al 1995) and 2. the detection of fluorescent probes specific for various tumor cells activities non invasively in different depths of the spheroids by confocal laser microscopy combined with multiphoton excitation for mapping with a subsequent threedimensional reconstruction the heterogeneity of the tumor cell response to the various therapeutic modalities (Porwol et al 1996).…”

Section: Resultsmentioning

confidence: 99%

The Use of Human Tumor Cells Grown in Multicellular Spheroid Culture for Designing and Improving Therapeutic Strategies

Acker

1997

Computational and Mathematical Methods in Medicine

Self Cite

View full text Add to dashboard Cite

Human tumor cells grown in threedimensional multicellular spheroid culture represent an ideal system to study the heterogeneous interaction between microenvironment and different therapeutic modalities in patients. Overcoming this heterogeneity is the most challeging task for the improvement of cancer therapy. Examples of this interaction are given with the meaning of p 0 2 and pH for the treatment of tumor cells with radiation, cytostatics, cytokines and gene therapy. The presented results might form a base for the use of multicellular spheroids in a concerted action between clinicians, biologists and mathematicians to improve tumor therapy individually for each patient.

show abstract

Section: Resultsmentioning

confidence: 99%

The Use of Human Tumor Cells Grown in Multicellular Spheroid Culture for Designing and Improving Therapeutic Strategies

Acker

1997

Computational and Mathematical Methods in Medicine

Self Cite

View full text Add to dashboard Cite

show abstract

“…For the 3D reconstitution work, we used the software package AVS (Porwol et al 1996;Sheehan et al 1996). This approach is convenient because of the high flexibility provided by the programming environment.…”

Section: Discussionmentioning

confidence: 99%

“…The command line driven program allows linear and nonlinear filtering as well as application of several edge detection algorithms (Porwol et al 1996). Because filtering of large data sets (e.g., 3 ϫ 512 ϫ 512 ϫ 60 pixels) is a rather time-consuming task, the initial data processing was done on a SPARC 10 workstation (SUN Microsystems; Mountain View, CA) equipped with 96 megabytes of RAM.…”

Section: Three-dimensional Reconstructionmentioning

confidence: 99%

“…A high number (60 or 79) of optical sections was obtained from the double-stained translucent assemblies. For the 3D reconstitution of the sectioned space, we used the software package "Application Visualization System" (AVS) (Foley et al 1994;Sheehan et al 1996) that was modified to meet the requirements of confocal laser scanning microscopy (Porwol et al 1996). The reconstructed images obtained show the gel matrix, the cell surface, and the spatial relationship of the stained intracellular compartments containing cathepsin B.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Tomography of Cells by Confocal Laser Scanning Microscopy and Computer-assisted Three-dimensional Image Reconstruction: Localization of Cathepsin B in Tumor Cells Penetrating Collagen Gels In Vitro

Strohmaier

Porwol

Acker

et al. 1997

J Histochem Cytochem.

View full text Add to dashboard Cite

SUMMARYWe used the nondestructive procedures of confocal laser scanning microscopy in combination with computer-assisted methods to visualize tumor cells in the process of penetrating collagen gels. Three independent sets of images were collected. The image information of all data sets was combined into one image, giving a three-dimensional (3D) impression at high light microscopic resolution and sensitivity. We collected information about the extracellular matrix using the reflection mode, the cell surface/morphology by staining with the fluorescent dye DiOC 6 (3), and the distribution of cathepsin B by Cy-3-labeled immunolocalization. The specific aim of our study was visualization of the spatial relationship of cell organelles as far as they contain the enzyme cathepsin B to cell morphology and motility in a 3D model of extracellular matrix. The majority of the enzyme was localized pericellularly, with no visible relationship to the direction of movement. However, substantial amounts also appeared in intramatrix pseudopodia and associated with the extracellular face of the plasma membane, which may be indicative either of secretion and/or epicellular activity. Our approach has general applicability to study of the spatial relationships of Biological structures have a dynamic three-dimensional (3D) organization. However, our understanding of structures and functions and their interrelations in cells is almost exclusively based on two-dimensional images and is therefore incomplete. In medical sciences, 3D imagery of whole organs or large anatomic regions of the body is possible by a number of noninvasive methods. Moreover, at the molecular level, spatial reconstitution of molecules or molecular assemblies is a common, although complicated procedure. At the cellular level, knowledge of spatial extension or interaction of structures at the light and electron microscopic level was obtained by destructive sectioning methods. Confocal laser scanning microscopy is now a well-established method to obtain optical sections of spatially extended biological objects (Brakenhoff et al. 1989;Masters and Farmer 1993;Wright et al. 1993). This precise and nondestructive procedure can render series of a sufficient number of equally thick sections to allow accurate 3D reconstitution of the imaged volume. The invention of appropriate fluorescent dyes and the development of specific antibodies to cell constituents are the last links in a chain leading to realistic 3D images of cells and their constituents.Such a methodology is an ideal tool to study cell locomotion (movement). In situ, this is a 3D process. It involves complex rearrangements of the intracellular compartments, intercellular interactions, and interactions of cells with the extracellular matrix. Movement of cells through natural barriers (invasion) is a basic phenomenon in living organisms. In higher organisms, such invasive processes occur during all stages of deCorrespondence to: Dr. Eberhard Spiess, Biomedizinische Strukturforschung 0195,

show abstract

“…Furthermore, the blood vessel diameters, as well as volumes were determined from the 3-D reconstructions of serial confocal sections. Details on image processing and data visualization in confocal laser scanning microscopy have been published elsewhere (Ehleben et al, 1997;Lorensen et al, 1987;Porwol et al, 1996;Strohmayer et al, 1997).…”

Section: Scanning Confocal Microscopymentioning

confidence: 99%

Endostatin reduces vascularization, blood flow, and growth in a rat gliosarcoma

et al. 2002

Self Cite

View full text Add to dashboard Cite

Endostatin, the 20-kDa C-terminal fragment of collagen XVIII, has previously been shown to inhibit growth and induce regression of different experimental tumors in rodents. In this study, we show that recombinant murine and human endostatin, produced in 293 EBNA cells and yeast, respectively, inhibit ectotopic as well as orthotopic growing BT 4 C n gliosarcomas in BD-IX rats. In rats in which s.c. gliomas were grown for a total of 29 days, systemic treatment with recombinant murine endostatin induced about 50% reduction of intratumoral blood ow and tumor size after only 10 days of therapy. In contrast, the blood ow to irrelevant organs was unaffected by endostatin, indicating its speci city of action. Tumors were not observed to increase in size or regrow after cessation of therapy. Furthermore, endostatin-treated rats with i.c. tumors had signi cantly longer survival time than did untreated controls. In the treated rats, endostatin therapy resulted in a reduced tumor blood vessel volume and an increased tumor cell density with an increased apoptotic index within a given tumor volume, as veri ed by ow cytometry and by staining with deoxynucleotidyltransferase-mediated dUTP nick-end labeling. This work veri es the general anti-angiogenic and antitumor effects of endostatin and indicates that the protein may also be considered as a treatment strategy for malignant brain tumors. Neuro-Oncology 4, 1-8, 2002 (Posted to Neuro-Oncology [serial online], Doc. 01-015, September 27, 2001 T he recruitment of new blood vessels is regarded as essential for continuous tumor growth and metastasis (Folkman, 1990). Recently, several inhibitors of angiogenesis have been identi ed, and much attention has been paid to endostatin, which is a 20-kDa C-terminal proteolytic fragment of collagen XVIII (O'Reilly et al., 1997). This heparin-binding fragment has been shown to inhibit tumor growth in different experimental model systems Chen et al., 1999;Read et al., 2001;Yamaguchi et al., 1999). The exact mechanism of action is still not understood, but the molecule may induce apoptosis of bovine pulmonary endothelial cells (Dhanabal et al., 1999). However, whether this applies to endothelial cells from the tumor vasculature is unknown.Murine endostatin, produced as a recombinant protein in Escherichia coli, has been shown to inhibit tumor growth when administered daily to mice bearing ectopic Lewis lung carcinoma, T241 brosarcoma, or B16F10 melanoma. Importantly, the tumors apparently do not develop resistance to endostatin therapy, but tumor Neuro-Oncologyregrowth may be observed upon termination of endostatin treatment . However, one study suggests this may be avoided by giving cycled endostatin therapy, which appears to induce a dormant tumor state that persists even after discontinuation of therapy .Most in vivo tumor inhibition experiments with endostatin have been performed in mice with s.c. tumors using precipitated endostatin produced in E. coli and given s.c. in doses of 3 to 20 mg/kg body weight (O'Reilly et al., 1997). Because...

show abstract

Three-Dimensional Imaging of Rhodamine 123 Fluorescence Distribution in Human Melanoma Cells by Means of Confocal Laser Scanning Microscopy

Cited by 13 publications

References 1 publication

The Use of Human Tumor Cells Grown in Multicellular Spheroid Culture for Designing and Improving Therapeutic Strategies

The Use of Human Tumor Cells Grown in Multicellular Spheroid Culture for Designing and Improving Therapeutic Strategies

Tomography of Cells by Confocal Laser Scanning Microscopy and Computer-assisted Three-dimensional Image Reconstruction: Localization of Cathepsin B in Tumor Cells Penetrating Collagen Gels In Vitro

Endostatin reduces vascularization, blood flow, and growth in a rat gliosarcoma

Contact Info

Product

Resources

About