Biomedical subjects
Publications and source records attributed to A Leibovitz.
The authors present a case of a patient suffering from profuse diaphoresis as the sole manifestation of myocardial ischemia (syndrome x or coronary vasospasm). The diagnosis was established by positive exercise thallium 201 imaging and normal-appearing coronary arteriogram.
The authors describe the association of hypernatremic dehydration and peripheral gangrene in elderly patients. This association is known in infancy but hitherto has not been described in the elderly.
The authors present a typical case of Holt-Oram syndrome in an eighty-year-old woman. The patient had atrial-septal defect and phocomelia but was independent in the activities of daily living.
Two human cell lines (UACC-812 and 893), both containing significant amplification of the HER-2/neu gene, were established from biopsy specimens of breast carcinomas. One patient had Stage II breast carcinoma; the other had metastatic disease. Characterisation of these lines has revealed that both are highly aneuploid containing multiple clonal chromosome alterations, have doubling times near 100 h, and are oestrogen and progesterone receptor negative. Electron microscopy demonstrates that both lines contain numerous microvilli, cytoplasmic filaments, multivesicular bodies, and desmosomes. Immunoblot analysis for P-glycoprotein using the monoclonal antibody C219 was negative for both patient cell lines. These relatively rare cell lines may represent a useful model to investigate human breast carcinomas.
We have used an in vitro adhesion assay to study the interaction of tumor cells with lymphatic endothelium, a dynamic event that leads to tumor metastasis in vivo. 3H-thymidine-labeled human tumor cells from: one primary Ewing sarcoma, two established melanoma cell lines, two colon and two breast carcinomas (one established line and one primary culture of each) were added to 24-well culture dishes containing confluent monolayers of bovine lymphatic endothelium. Radioactivity associated with either the cells in suspension or the attached cells was assessed and compared at frequent intervals up to 360 minutes. Generally, tumor cell attachment increased as a function of time reaching a plateau between 180 and 360 minutes. the modular media system described here facilitates the primary and secondary culture (or co-culture) of a variety of normal and transformed cells. Primary cultures with a rounded morphology (one breast and one colon carcinoma) showed the lowest preferential attachment for lymphatic endothelium. All established cell lines and the primary Ewing sarcoma cell line displayed a more fibroblastic morphology and achieved the highest adhesion profiles. There was a correlation between the malignancy and attachment potential for the melanoma and breast carcinoma cell lines. Collectively, these data show that established tumor cell lines with fibroblastic-like morphology exhibit more rapid adhesion than primary tumor cell cultures with more rounded morphologies. While this property may reflect in vitro selection and/or adaptation, it does correlate with the metastatic propensity for some human tumor cells.
Doxorubicin, cisplatin, and cyclophosphamide are the three drugs most commonly used in the treatment of ovarian cancer, but no effect greater than additivity was observed for any combination of these drugs in the present study. Only a few studies have been reported concerning the degree of their additivity or their best order of sequencing. In our in vitro studies, cisplatin in combination with doxorubicin or 4-hydroperoxycyclophosphamide (4HC) was tested against seven human gynecologic tumor-cell lines in different sequences, using a double-agar layer tissue-culture system. Drug interactions with respect to inhibition of tumor clonogenicity were evaluated by isobologram and fractional survival methods. Doxorubicin and 4HC were sequenced simultaneously and at 1, 6 and 24 h after cisplatin, and cisplatin was sequenced at 1, 6 and 24 h after 4HC. The isobolograms constructed for doxorubicin or 4HC plus cisplatin revealed strict additivity between these agents against ovarian cancer clonogenicity. Both doxorubicin and 4HC showed the greatest additivity when used simultaneously and at 1 h vs 6 or 24 h after cisplatin. Although the mechanisms by which these sequencing effects occur are unknown, these studies provide new leads for the design of clinical trials with combinations of these three agents.
A number of techniques are now in use in attempting to establish cell lines from human solid tumors. Although short-term cultures are readily attained, relatively few become established as permanent cell lines, especially from primary carcinomas where metastasis has not already occurred and from such organs as the breast, prostate, and pancreas. With recent advances in techniques and in our knowledge of growth factors, however, these difficulties may be resolved in the near future.
Fibroxanthosarcoma is a tumor of soft tissues, mainly effecting the extremities, mediastinum and retroperitoneum. Primary fibroxanthosarcoma of the kidney is extremely rare and has been described in only 4 cases in the past. We present an additional patient, presenting with continuous spiking fever, with rapid deterioration of her condition due to widespread metastases and death within 4 months.
We have coined the term 'helioid inclusion' to describe a new type of single-membrane-bound intranuclear inclusion which contains one or more rounded bodies with radiating filaments (helioid bodies). Helioid bodies were not found in the rough endoplasmic reticulum in the cytoplasm, or in the cytoplasmic matrix, but a helioid body was seen in dilated rough endoplasmic reticulum included in a double-membrane-bound intranuclear pseudoinclusion. No firm conclusion regarding the genesis of helioid inclusions has emerged from our studies, but two possibilities exist: 1) that helioid bodies derive from modification of serous secretory granules sequestrated in an intranuclear inclusion; or 2) that they develop by condensation of the contents of the perinuclear cistern or rough endoplasmic reticulum subsequent to its sequestration in an intranuclear inclusion.
Two new human myeloma cell lines have been established from a 36-year-old woman with refractory IgG kappa multiple myeloma in whom bilateral malignant pleural effusions developed. The malignant plasma cells from each effusion were set up in a liquid culture using an L-15 medium containing catalase, glutathione, selenous acid, ascorbic acid, insulin, transferrin, additional glutamine hydrocortisone, and 2-mercaptoethanol and designated as M-3 medium. Two IgG kappa cell lines, LB -831 and LB-832, were established and proved to be Epstein-Barr virus negative using the internal repeat sequence DNA probe. Characteristic plasma cell morphology was evident by light and electron microscopy. Immunotyping revealed an IgG kappa , B1+, B2-, Ia (HLA-DR)+, CALLA+ phenotype for each cell line as well as for the original pleural fluid and bone marrow myeloma cells. The supernatants also contained IgG kappa, beta 2 microglobulin, and large amounts of osteoclast-activating factor (indicating bone-resorbing activity). Cytogenetic analysis of the LB-831 cell line revealed a nearly triploid highly abnormal karyotype with numerous clonal chromosomal abnormalities involving chromosomes 1, 3, 5, 7, 13, and 15; several structurally abnormal marker chromosomes; and a putative homogeneously staining region on chromosome 7p at band p22. Analysis of the LB-832 cell line revealed several additional clonal abnormalities. These additional cytogenetic changes suggest that in vivo sequential clonal evolution occurred in this patient. Therefore, two new but related cell lines have been established, which should prove useful for further biological studies.
We categorized established human colon carcinoma cell lines into three biological groups. Our studies were performed on six colorectal cancer lines representing the proposed three groups. Group I consisted of two lines designated LoVo and SW 48; Group II comprised two lines called SW 480 and SW 620; and Group III was represented by lines SW 403 and SW 1116. Group I consisted of the most differentiated cells. This differentiation encompassed morphological markers, gland and signet ring formation, and ciliary development. The outstanding morphological characteristic of Group III was the development of numerous multinucleated giant cells. The range and modal chromosome number increased from Group I to Group III, a change reflected by the higher DNA content of these cells as measured by flow cytometry. Carcinoembryonic antigen synthesis was maximal for Group III and virtually absent for Group II. The number of clonogenic cells decreased from Group I to Group III, while the proportion of nonproliferating cells calculated both by experiments using continuous labeling with tritiated thymidine, and by the primer-available alpha-DNA polymerase index, increased from Group I through Group III. Another important cytokinetic difference was that Group I had an exponential cell cycle stage distribution not seen for the other groups. Cells in Group I were easily propagated in athymic (nude) rats by s.c. injection; cells in Group II injected s.c. grew for about 30 days and then regressed spontaneously. Cells in Group III could only be grown when inoculated intracerebrally. Thus, our studies have now confirmed and extended the hypothesis that cultured human colorectal carcinomas can be separated into at least three groups on the basis of morphological differentiation, chromatin distribution, carcinoembryonic antigen production, cytokinetic properties, and xenograft propagation. Perhaps this classification is just the tip of the iceberg, and future studies will determine the existence of additional groups or subgroups on the basis of other markers. However, at present it appears established that malignant cells with a common histological origin in the gut express their phenotypic potential in a sufficiently discrete manner as to permit their classification into distinct biological groups. Thus, the stage is set for extrapolating this in vitro classification for an in vivo segregation of human colorectal tumors into categories with specific properties and diverse prognosis.
A hypo-osmolar medium and tissue processing technique is described which is useful for disaggregation of residual human tumor cell clumps persisting after mechanical or enzymatic treatment of solid tumors and malignant effusions. The addition of the hypo-osmolar procedure to the standard methods for disaggregation increased the viable single cell yield in solid tumors by 47% and in malignant effusions by 67%. In 5 of the 26 solid tumor specimens tested in the human tumor stem cell assay, clonogenic single cells were obtained with the hypo-osmolar procedure, whereas no growth was observed using standard methods. Overall, the success rate for clonogenicity increased from 46% to 65% for the 26 solid tumors, with the major improvement occurring in ovarian cancer. Clonogenicity was obtained in 80% of malignant effusions both by standard methods and the hypo-osmolar techniques. The increased total yield of clonogenic cells obtained with this procedure enhances the opportunity for experimental versatility and in vitro drug testing.
An ex vivo culture system was developed for assessing the activity of cancer chemotherapeutic agents against tumor cells. The system utilizes artificial capillary culture units and the technique of hemodialysis to expose tumor cells to a chemotherapeutic drug and its metabolites following injection of the drug into an experimental animal. This ex vivo culture system was used to test the activity of 5-fluorouracil (5-FU) against four human colorectal adenocarcinoma cell lines (SW 403, SW 480, SW 620, and SW 707). Cell killing by 5-FU or its metabolites in blood dialysate following intravenous injection was measured by determining colony formation of cells attached to plastic and suspended in 0.3% agar after short-term exposures of 1 to 2 h. The technique was shown to discriminate between the sensitivities of these cell lines and the respective sensitivities to the drug were reproducible. Kinetics of drug clearance from the host animal's blood were shown to be similar to that in humans. The results suggest the system may be useful for testing diverse drugs, including those requiring metabolic activation, against a variety of types of tumor cells.
An established cell line, SW756, derived from a primary squamous carcinoma of the uterine cervix is described by its morphology, ultrastructure, karyotype, genetic signature analysis, HLA typing, and tumorigenesis in the nude mouse. Cultured cells obtained from the SW756 derived nude mouse tumor also were studied for chromosome and isozyme markers. The original tumor was poorly differentiated carcinoma with minimal keratinization and is compared with that occurring in the nude mouse after the cultured cells were inoculated. The nude mouse tumor showed similar histological features, but better differentiation than the original tumor. Karyotype analysis of SW756 demonstrated a hyperdiploid stem line number and several marker chromosomes (MI-M6). No HeLa marker chromosomes were identified. The isozyme pattern for SW756 reported by others has been confirmed. The unique chromosome and isozyme features have been identified repeatedly in the cultured cells and, most importantly, in the post nude mouse culture. We recommend SW756 as a defined human tumorigenic cell line derived from a primary squamous carcinoma of the uterine cervix.
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Four human colorectal adenocarcinoma tumor cell lines, previously established and characterized in monolayer culture were grown in a matrix-perfusion culture system to determine the suitability of this technique for synthesis of carcinoembryonic antigen (CEA). Production of CEA in excess of 100,000 ng was attained from one cell line, SW 403, during 15-day growth trials. In growth trials and cell-free diffusion studies, CEA passed through membranes of 100,000-dalton molecular weight porosity but not 10,000 porosity. Using cell cultures of high, moderate, or low producers, CEA synthesis tended to reach a plateau after several days of culture and remained nearly constant as the cells attained a maintenance condition. Basic biologic characteristics of the cell lines, expressed as growth rates and CEA produced per 10(6) cells, were comparable in monolayer and perfusion culture. The high cell densities, (10(8) to 10(9) cells per ml) achieved in matrix perfusion made it possible to routinely obtain continuous high yields of CEA over an extended time period.
Thirty-one different long-term established human carcinoma culture cell lines were examined for the presence or absence of receptors for the Fc portion of immunoglobulin G (IgG) ("Fc receptors"), the third component of complement (C3 receptors), sheep erythrocytes and mouse erythrocytes. Included in the catalogue of lines tested were adenocarcinoma of the adrenal cortex, colon, rectum, lung, liver, breast and kidney, and carcinomas of the uterine cervix, bladder and vulva. All lines were found to be consistently Fc receptor-negative, as assayed by rosette formation with sheep erythrocytes coated with subhemagglutinating amounts of anti-sheep erythrocyte antibodies, or bovine erythrocytes heavily coated with non-hemagglutinating anti-bovine erythrocyte antibodies. This is similar to results obtained with murine tumors: carcinomas are consistently Fc receptor-negative, while those lines which are Fc receptor-positive are invariably lymphoreticular in nature. The human carcinoma cell lines were also negative for complement (C3) receptors and spontaneous T and B rosette markers. It is proposed that, in most cases, rosette-forming cells found in primary human carcinomas are, as in the mouse or rat, representative of infiltrating non-malignant lymphoreticular cells.
A veterinarian contracted rabies in the course of laboratory work with homogenates of rabid goat brain. Epidemiologic study determined a respiratory mode of transmission. After a fulminant encephalitic illness, formed rabies virions were identified in the synaptic zones of the olfactory glomeruli. Identification, isolation, experimental disease production, and tissue cytopathic effects of virus recovered from the brain fulfilled Koch's postulates in this unusual instance of virus disease of the nervous system.