Progress and problems of interhospital consulting by computer networking.
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Computer Communication Networks↗
Biomedical subjects
D F Parsons
Publications and source records attributed to D F Parsons.
Organelle rearrangement and cell volume changes during squeezing invasion of peritoneal elastic lamina by targeted murine breast carcinoma cells.
Murine breast cancer cell lines were developed to selectively invade the peritoneum while they proliferated in ascites form in the abdominal cavity. In a dominant form of invasion, tumor cells showed special affinity for elastin fibers and squeezed through narrow gaps in the elastic fiber meshwork of the stroma. Even in fixed tissue, such cells could be recognized as being in the process of invasive migration because of their dumbbell shape. This appearance was similar to that of diapedetic blood cells traversing bone marrow sinus endothelium. Three-dimensional STERECON graphics reconstruction from serial thick sections of 44 such cells was carried out. The reconstructions showed that, in mid-penetration, the cells spread extensively over the exterior surface of the elastic fiber meshwork. The cell surface contact of these forward projections was mainly with the elastic fiber outer coat of microfibrils, but small areas of the cell surface also fused directly to inner-core elastin. The morphological rearrangement of the cytoskeleton was minimal in both types of attachment areas. The location of these forward facing attachments is consistent with mechanisms for pulling the invasive cell through the gap. Lamellopodia formation and clustering of cytoplasmic organelles occurred more commonly at the forward-facing part of the cell. Morphometry of the reconstructions showed that a contraction of the whole cell occurred during the squeezing/migration process suggestive of an additional pushing process. However, our invasive cell lines showed marked differences in the degree of cell shrinkage. The process of adhesion and squeezing of tumor cells through elastin meshworks in vivo is clearly a complex phenomenon. Changes in cell surface activity appear to play a significant role in establishing the necessary 'foothold' component of invasion and, possibly, in the generation of tractive force as well.
Animals↗
The relative merits of direct morphometry of reconstructions of whole cells, and statistical morphometry by stereology of random sections of cells.
Stereology, or the derivation of quantitative, three-dimensional (3-D) data about cells by statistical analysis of the structures of random sections, is widely used in cytology and pathology. However, there are situations where this approach is inadequate, and only an analysis of a homogeneous population of whole cells will give the required results. This involved 3-D reconstruction from physical or optical sections, or tomography or photogrammetry of whole-cell mounts. Use of stereo views of individual sections or projections adds considerably to the information available for both contouring and reconstruction. Recent image-processing advances in clinical radiography have shown, for the first time, that rapid, high-resolution digitization and contrast enhancement enable nearly all structural details to be routinely extracted from the micrographs and adequately portrayed. Three-D whole-cell reconstructions provide the digital data for many kinds of morphometric measurements on both whole cells and their individual organelles and membranes. Rapid fixation or freezing allows improved quantitative structure/function correlations of organelles with disturbances in cell metabolism or gene expression.
Animals↗
Targeting of peritoneum by the small numbers of isogeneic and allogeneic ascites carcinoma cells that infiltrate or attach to peritoneum during ascites growth.
In the course of development of an in vivo invasion model, sublines of a series of allogenic and isogeneic carcinoma cell lines have been selected that show enhanced invasion of the peritoneum. It was found that, during the proliferation of tumor cell lines in ascitic form in the abdominal cavity, small numbers of cells infiltrated or firmly adhered to the peritoneum in at least 8/12 of the tumor-host combinations tried. After thorough washing of the peritoneum it was disaggregated by an enzyme mixture, and the resulting mixture of normal and tumor cells was inoculated intraperitoneally. Peritoneal isolations were made serially for 3 to 12 times. In 6 of 8 cases where the isolation produced a stable ascites, the cells showed enhanced peritoneal invasion compared with the parent cell line. The invasion of some of the cell lines was tested in another invasion model consisting of cultured mouse buccal mucosa (9/10 cell lines invaded the explant). In 3/3 cell lines showing enhanced peritoneum invasion in vivo, there was no enhanced invasion of the buccal mucosa. The enhanced peritoneum invasion appears to be tissue specific rather than a general increase in invasion potential. Pairs of high- and low-invasive cell lines were obtained that should be useful for screening for invasion modulating agents using the mouse ascites/peritoneum in vivo model. It is suggested that the method might be generalized to produce various tumor cell lines that target for the normal tissues that are adjacent to proliferating solid or circulating tumors.
9,10-Dimethyl-1,2-benzanthracene↗
Telecommunication discussion groups for health services and medical research.
International university and research institute telecommunication links provide free or low-cost access to several computer networks (commercial and non-commercial). Previously used by science researchers, such networks are now used in clinical and social medicine. University affiliation, a personal computer, and a modern and software are needed. A host-computer, at a local computing centre, operates server software that automatically distributes information to subscribers. The server stores all messages in a monthly logbook that can be edited to produce an electronic magazine. The magazine can also be accessed by non-subscribers. The software gives convenient, fast, single-key processing of messages and files and also allows on-line conferences. Discussion groups on several medical specialties and data bases for AIDS and other communicable diseases are already operating.
Computer Communication Networks↗
Shape, size, and distribution of cell structures by 3-D graphics reconstruction and stereology. I. The regulatory volume decrease of astroglial cells.
This report discusses fundamental limitations in attempting to derive cell size, shape, or distribution from the two-dimensional images provided by conventional electron microscopy. Morphometric or stereologic measurement of random thin sections is a convenient way to obtain some information of this type. However, it cannot provide complete, objective information about real size, shape, or connectivity of cells containing irregular or unevenly distributed structures or nonuniform populations of cells. Anisotropic structures require analysis of a complete set of serial sections. The analysis may utilize either stereo, mono, or tilted optical slices, and subsequent integration of this information into a single 3-D computer data set. In this study, we analyze stereo pairs of high-voltage electron micrographs of serial thick sections (0.5 micron) and critical-point-dried whole-cell mounts of rat brain astroglial cell cultures. The Z-axis resolution is increased by digitizing contours at discrete levels within each stereo view. This is accomplished with a new type of stereoscopic contouring device. We calculated area and volume changes accompanying hypo-osmolar swelling and spontaneous reversal of the swelling. (Regulatory Volume Decrease-RVD). An understanding of the mechanism of swelling of astroglial cells is important for improving the treatment of brain injury. The total cell-volume results are comparable with results previously obtained using nonmetabolized, radioactively tagged compounds that diffuse into various cell compartments. Our serial-section and whole-cell data also provide new information about the relative swelling of nucleus, cytoplasm, and individual organelles such as mitochondria. The basic biological problem being approached is whether homeostasis of cell function is accompanied by surface area and volume regulation of enzyme-rich membranes and organelles. Conversely, it is proposed to explore the possibility that abnormal organelle areas and volumes are indicators of perturbations of cell division, metabolism, or gene expression.
Animals↗
Near-diploid tumor cell lines for cancer research.
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Animals↗
Establishment and characterization of two new squamous cell carcinoma cell lines derived from tumors of the head and neck.
Two human cell lines were established from untreated squamous cell carcinomas of the head and neck. Line 183 was derived from a head and neck squamous cell carcinoma of the tonsil and 1483 from a head and neck squamous cell carcinoma of the retromolar trigone. Both lines grow in a cobblestone pattern demonstrating their epithelial heritage. Immunofluorescence studies and one-dimensional polyacrylamide gel electrophoresis indicated that both lines contain cytokeratins. Line 1483 is more aggressive in nude mice, has a higher efficiency for anchorage-independent growth, expresses p21ras (product of the ras oncogene) at a higher level, and is more aneuploid than 183. 1483 also grows as a multicellular tumor spheroid. Line 1483, which was established from the primary tumor of a patient with nodal metastasis, thus displays more progressed characteristics than line 183, which was established from a patient with no clinically positive nodes.
Aged↗
Resolution as a function of accelerating voltage in electron microscopy of semithick biological specimens.
In the past, biological sections ranging in thickness from 0.10- to 0.50-micron have usually been examined with high-voltage (greater than 500 kV) electron microscopes (HVEM). Now investigators are increasingly using intermediate voltage (200-500 kV) electron microscopes (IVEM), which are more readily available and demand less maintenance. In a study of "typical" plastic-embedded, stained sections of mouse liver ranging from 0.10 to 1.0 micron thick, we determined the resolution obtainable at 100, 200, and 1000 kV. At all three accelerating voltages the resolution (2.7 nm) for 0.10-micron sections was limited only by the sections stain granularity. For 0.25-micron thickness the resolutions were 5.8, 3.1, and 3.1 nm at 100, 200, and 1000 kV, respectively. The maximum usable thickness at 200 kV with resolution sufficient to resolve membranes clearly was between 0.75 and 1.0 micron, depending on the magnification. Resolution at 100 kV was adequate for screening sections up to 1.0-micron thick for preparation defects prior to examination with an IVEM or HVEM.
Animals↗
Electron microscopy, electron diffraction, and element analysis of wet biological specimens.
Temperature controlled differentially pumped environmental chambers now allow more routine examination of wet specimens in the electron microscope. A sensitive test of their efficiency is the ability to provide high resolution electron diffraction patterns from wet, unfixed protein microcrystals. Fortunately, wet specimens can be prepared with only a few tens of nanometers thickness of remaining water, so extraneous electron scattering by liquid water can be kept to a minimum. It still remains to be determined whether microprobe analysis (X-ray or electron energy-loss spectroscopy) using wet specimens gives better element localization in cells than the current freezing methods. More extensive comparisons are also required of the ultrastructural preservation and visibility of macromolecules immersed in a thin layer of water vs immersion in a thin layer of amorphous ice. However, the recent introduction of commercial forms of the necessary equipment now make these comparisons more feasible.
Freeze Drying↗
Ultrastructural characterization of isolated human head and neck squamous carcinoma cells. Assessment of isopycnic centrifugation.
In an experiment to evaluate the merit of isopycnic centrifugation as a method of separating cell types in human head and neck squamous cell carcinomas, cells have been isolated from four specimens of these tumors and subjected to isopycnic centrifugation in continuous Percoll gradients. Cell types were identified by electron microscopy. The R- (ribosome-rich), T- (tonofilament-rich), and RT- (intermediate) cell types yielded broad bands overlapping extensively with one another, and partially with the bands of leukocytes. The pattern differed for each tumor studied, so that universal density levels separating given cell types could not be found. Isopycnic centrifugation proves less suitable in analyzing cells dispersed from solid tumors than for cells in suspended culture, blood, effusions, etc., probably because of heterogeneous growth conditions of cells in solid tumors.
Carcinoma, Squamous Cell↗
Detection and characterization of circulating rat mammary tumor cells in buffy coat and correlation with metastasis.
A new method for detecting bloodborne TMT-081 rat mammary tumor cells in buffy coat has revealed dose-dependent variations in the latency period after inoculation of tumor cells, the concentration of circulating tumor cells, and the incidence of metastases. Cells isolated from buffy coat of right ventricular blood were more tumorigenic than tryptically dispersed cells from solid tumors. With the new method circulating tumor cells can be detected at concentrations as low as 3 cells/microliter of buffy coat, or approximately 60 cells/ml of whole blood. The morphologic and ultrastructural features of the primary tumor were generally retained in both the circulating and tryptically dispersed cells, as shown by light and electron microscopy. A sparse distribution of intermediate filaments was revealed by high-voltage electron microscopy, although the filaments were not evident in conventional transmission electron micrographs. They were identified as keratin by immunofluorescence studies.
Animals↗
Patterns of blood-vessel invasion by mammary tumor cells.
Rat mammary tumor TMT-081 was employed as a model for blood vessel invasion because its mode of metastasis resembles that of human tumors. The invasive mechanism was studied with two methods of serial transplantation: transfer of enzymatically dispersed solid tumors, and transfer of buffy coat containing circulating tumor cells. The latter method produced greater invasion of blood vessels, including larger veins and occasionally arteries, perhaps by obviating damage to tumor cells during enzyme treatment. The course of migration was traced by three-dimensional examination in the high voltage electron microscope, as well as the light microscope. Two broad patterns were found for the course of invasion of small and large vessels respectively.
Animals↗
The usefulness of the high voltage electron microscope in biomedical ultrastructure analysis.
We now have the tools to quantitate and correctly interpret the size, shape, and distribution of organelles in cells. It appears likely that the results will require some changes in some basic assumptions of ultrastructural cytology. This should also result in some new biochemical hypotheses about control of cell metabolism.
Cells↗
Immediate ascites conversion of mammary tumors induced in NYLR/Nya mice by 7,12-dimethylbenz-[a] anthracene and urethane feeding and by forced breeding.
Intragastric feeding of dimethylbenz-[a]anthracene in corn oil together with urethane in the drinking water and forced breeding were successful in rapidly inducing (2-4 months) mammary tumors (adenoacanthomas) in inbred NYLR/Nya mice, which have a low incidence of spontaneous breast tumors and of other tumors. The tumors could be quickly and permanently transformed to an ascites form by intraperitoneal inoculation of enzyme-dissociated cells and subsequent serial passage of free cells. Only 10 (3 in some cases) serial passages were required, thus conveniently providing multiple isogeneic carcinoma cell lines in this strain of mice. Some tumor cell lines proliferated strongly in the abdominal cavity even on the first passage.
9,10-Dimethyl-1,2-benzanthracene↗
In vitro modulation of differentiation by calcium in organ cultures of human and murine epithelial tissue.
The differentiation of epithelial tissue in organ cultures of murine buccal mucosa, various human oral mucosa, and human newborn foreskin was found to be dependent on the calcium concentration of the culture media. In low calcium medium (less than or equal to 0.07 mM) epithelial differentiation was inhibited. The original stratifying layers separate and can be removed, producing a destratified explant. Histologically such an explant consists of a dorsal epithelial layer of basal keratinocytes resting on an intact basal lamina with subjacent stroma. At 0.01 mM calcium, the epithelial layer was one to two cells thick whereas at 0.07 mM it could be three or more layers in thickness with the most superficial cells being spread over the underlying cells. In addition to differentiation, keratinocyte migration over the sides of the explant (epiboly) and epithelial proliferation as determined by [3H]thymidine autoradiography were reduced by culture in low calcium medium. Redifferentiation occurs upon return to normal calcium levels (1.8 mM); addition of hydrocortisone to low calcium media was found to facilitate this redifferentiation.
Adult↗
Shape changes and polarization of cells migrating through tissue. A high-voltage electron microscope and computer graphics study of serial thick sections.
Structural changes of carcinoma cells and fibroblasts migrating through small spaces in the elastic-collagen reticulum of mouse peritoneum have been studied by high-voltage electron microscopy of serial thick sections and by computer graphics reconstruction of cell profiles. The change of shape profile of an individual cell, between serial sections is large and the distribution of organelles is very non-uniform and changes markedly between sections. Conclusions about adhesion, intercell contact, cell shape and polarization of cytoplasmic organelles could only be reached by assessing a complete set of serial sections. Our preliminary results suggest that interesting structural changes occur in both carcinoma cells and fibroblasts when migrating through this tissue.
Adenocarcinoma↗
Inflammation with restricted lysosomal proteolysis during early ascites carcinoma invasion of mouse parietal peritoneum. A medium and high-voltage electron microscopic and cytochemical study.
A carcinoma invasion system (Krebs-2 and Ehrlich tetraploid ascites tumors invading mouse peritoneum) was studied by high-voltage electron microscope (HVEM) stereoscopy, conventional (medium voltage) electron microscopy (MVEM), and cytochemistry. Tumor cells entered areas of peritoneum (mainly parietal) only where mesothelial cells were damaged and where there was inflammation of the underlying stroma. The initial invasion was different from that of most other invading carcinomas in that there was minimal breakdown of basal lamina and collagen. Neither tumor cells, inflammatory leukocytes nor peritoneal fibroblasts showed significant secondary lysosome production or release of intracellular or extracellular acid phosphatase. Morphological and cytochemical criteria suggest that in some invading carcinomas, as with non-tumor migrating cells such as leukocytes, widespread proteolysis due to diffusion of proteases is not a prerequisite for invasion of stromal connective tissue.
Acid Phosphatase↗