Showing papers on "Tissue culture published in 1973"
TL;DR: It is demonstrated that it is possible to culture morphologically and immunologically identifiable human endothelial cells for periods up to 5 mo and ABH antigens appropriate to the tissue donor's blood type were not detectable on cultured smooth muscle cells or fibroblasts.
Abstract: Endothelial cells were isolated from freshly obtained human umbilical cords by collagenase digestion of the interior of the umbilical vein. The cells were grown in tissue culture as a homogeneous population for periods up to 5 mo and some lines were subcultured for 10 serial passages. During the logarithmic phase of cell growth, cell-doubling time was 92 h. Light, phase contrast, and scanning electron microscopy demonstrated that cultured human endothelial cells grew as monolayers of closely opposed, polygonal large cells whereas both cultured human fibroblasts and human smooth muscle cells grew as overlapping layers of parallel arrays of slender, spindle-shaped cells. By transmission electron microscopy, cultured endothelial cells were seen to contain cytoplasmic inclusions (Weibel-Palade bodies) characteristic of in situ endothelial cells. These inclusions were also found in endothelial cells lining umbilical veins but were not seen in smooth muscle cells or fibroblasts in culture or in situ. Cultured endothelial cells contained abundant quantities of smooth muscle actomyosin. Cultured endothelial cells also contained ABH antigens appropriate to the tissue donor's blood type; these antigens were not detectable on cultured smooth muscle cells or fibroblasts. These studies demonstrate that it is possible to culture morphologically and immunologically identifiable human endothelial cells for periods up to 5 mo.
6,874 citations
TL;DR: It is established that exogenous AHF procoagulant activity is not inactivated by the tissue culture system.
Abstract: Antihemophilic factor (AHF, Factor VIII) antigen has been demonstrated in cultured human endothelial cells by immunofluorescence studies using monospecific rabbit antibody to human AHF. Control studies with cultured human smooth muscle cells and human fibroblasts were negative. By radioimmunoassay it was demonstrated that cultured human endothelial cells contain AHF antigen which is released into the culture medium. Cultured smooth muscle cells and fibroblasts did not have this property. Cultured endothelial cells incorporated radioactive amino acids into high molecular weight, AHF antigen-rich protein fractions prepared from the culture media, 7% of the radioactive amino acid counts incorporated into this material were precipitated by globulin prepared from rabbit anti-AHF whereas normal rabbit globulin precipitated only 1.5% of the counts. Although cultured endothelial cells actively synthesize AHF antigen, AHF procoagulant activity was not detected in the culture medium. Studies seeking a basis for the lack of procoagulant activity have not clarified this deficiency, but they have established that exogenous AHF procoagulant activity is not inactivated by the tissue culture system.
1,242 citations
TL;DR: The permanent presence of tumour‐specific antigen (TSA) in this carcinoma cell line suggests that the TSA is a genetically determined characteristic of T24 cells.
Abstract: A cell line derived from human urinary bladder carcinoma, designated T24, was established in vitro. The growth of T24 cells in tissue culture was characterized by a disorderly pattern of growth in one or more layers and by mixed epithelioid-fibroblastoid morphology. The generation time of T24 cells was 19 h. The cells had a hypotetraploid stemline with marker chromosomes. The malignant character of the line was verified by inoculation of cell suspension into hamster cheek pouch. The presence of tumour-specific antigen (TSA) in T24 cells was demonstrated by a micromodification of the cytotoxicity test with autochthonous leukocytes and serum from the donor of the tumour. The TSA was also well detectable by the reaction with allogeneic leukocytes from tumour-bearing patients and persisted during a 30-month culture period. The permanent presence of TSA in this carcinoma cell line suggests that the TSA is a genetically determined characteristic of T24 cells.
522 citations
TL;DR: The findings indicate that hepatic parenchymal cells in this monolayer system are viable and behave in many respects like normal adult rat liver.
Abstract: Parenchymal cells from adult rat liver have been established in primary monolayer culture. Donor animals are subjected to a partial hepatectomy and, 4 days later, cells are prepared by collagenase perfusion of the regenerated liver. The hepatic parenchymal cells, separated from nonparenchymal material and suspended in serum-free medium, are placed in plastic tissue culture dishes, where they form a monolayer within 24 h. The monolayer cells exhibit minimal mitotic activity and demonstrate several major metabolic functions characteristic of liver in vivo; these include albumin synthesis and secretion, gluconeogenesis from 3-carbon precursors, responsiveness to insulin and glucagon, glycogen synthesis, and activity of two microsomal enzymes. These functions are present in the monolayer cells for several days at activities similar to those observed in the liver in vivo. The findings indicate that hepatic parenchymal cells in this monolayer system are viable and behave in many respects like normal adult rat liver.
402 citations
TL;DR: Numerous lines of mouse lymphoid tumors showed, with increased sensitivity, a property of normal mouse splenic lymphocytes, the potential for growth promotion in vitro by specific thiols added to standard culture media, but the response varied with the mitogen preparation used and under some conditionsThiols-disulfides were inhibitory.
Abstract: Numerous lines of mouse lymphoid tumors (13 of 22 tested) showed, with increased sensitivity, a property of normal mouse splenic lymphocytes, the potential for growth promotion in vitro by specific thiols added to standard culture media. For lymphoma L1210 (V), structure activity relationships were examined; 9 of 30 thiols promoted growth; the most active was alpha-thioglycerol, effective at 0.2 microM. Thiols became oxidized under conditions of tissue culture and had half-lives of less than 8 h. Disulfides of active thiols promoted growth of lymphoma cells. The mitogenic response of splenic lymphocytes to lectins was increased by thiols-disulfides which promoted the growth of lymphoma cells, but the response varied with the mitogen preparation used and under some conditions thiols-disulfides were inhibitory.
217 citations
TL;DR: A quantitative assay for the migration of Balb/c3T3 cells into wounds is described, used in the partial purification of a migration factor released into serum-free medium by SV28 cells.
Abstract: When a monolayer culture of normal Balb/c3T3 cells is wounded by scraping away part of the cell sheet, the cells do not migrate into the cleared area unless there is serum in the culture medium. By contrast, SV40-transformed Balb/c3T3 cells do migrate into the wound area without serum. A quantitative assay for the migration of Balb/c3T3 cells into wounds is described. This assay is used in the partial purification of a migration factor released into serum-free medium by SV28 cells. SV28 is a line of BHK21/13 hamster cells transformed by SV40 chosen for its malignancy. The most purified fractions have about 1500 times the specific activity of whole calf serum. These fractions have an activity that promotes overgrowth of Balb/c3T3 cells to high density and an activity that prolongs cell survival without serum. The SV28 migration factor is not extractable from the medium of untransformed BHK21/13 cells or from serum. This migration factor might contribute to the malignancy of SV28 cells.
171 citations
TL;DR: Short-chain fatty acids (up to hexanoate) and parabens induce, at partially inhibitory concentrations, a jagged cell shape in continuous epithelial-like cell lines, such as HeLa, Chang liver, L-132, and Intestine 407.
Abstract: Human (HeLa, Chang liver, L-132, and Intestine 407) and other mammalian (XC, SV3T3, and chick-embryo) cells in tissue culture are at least as sensitive to inhibition by lipophilic acids and nitrite as bacteria. Some of these compounds are the most frequently used antimicrobial food additives. Short-chain fatty acids (up to hexanoate) and parabens induce, at partially inhibitory concentrations, a jagged cell shape in continuous epithelial-like cell lines, such as HeLa, Chang liver, L-132, and Intestine 407. This morphological effect is not mediated or enhanced by butyryl cyclic AMP, which specifically affects fibroblasts.
169 citations
Journal Article•
TL;DR: The results indicate that the acquisition of biological malignant potential, both in vitro and in vivo, is accompanied by increased thermosensitivity in mouse mesenchymal cells.
Abstract: Summary The effects of supranormal temperatures upon normal (derived from normal, embryonal tissues) and neoplastic (derived from 3-methylcholanthrene-induced sarcomas) mesenchymal cells from C57BL/6 mice have been quantitatively studied in tissue culture. It has been found that exposure to a temperature of 42.5° for 2 hr or more has a selective lethal effect upon tumor cells (defined as cells capable of developing into a malignant tumor when injected into syngenic mice at doses of 1 × 10 6 cells or less in adults or 1 × 10 5 or less in newborns). No other biological characteristic studied (aneuploidy, length of time in culture, and rate of growth) could be correlated significantly with thermosensitivity. All the cultures of tumor-derived and tumor-producing cells exhibited the same high degree of thermosensitivity, i.e. , death of 95% of the cells after 2 hr exposure at 42.5°. All the cultured normal, nontumor-producing cells exhibited a lower degree of heat sensitivity, i.e. , death of 43% of the cells after 2 hr exposure to 42.5°. When a cell subline having a high tumor-producing ability was derived from a nontumor-producing line, it acquired a greater thermosensitivity, although its rate of growth was the same as that of the original line. These results indicate that the acquisition of biological malignant potential, both in vitro and in vivo , is accompanied by increased thermosensitivity in mouse mesenchymal cells.
162 citations
TL;DR: Two types of alterations in the enzyme hypoxanthine phosphoribosyl transferase are found, supporting the view that tissue culture cell variants can arise from structural gene mutations.
Abstract: Two selective procedures are compared in an effort to isolate variants of mouse L cells containing structural gene mutations. Among the resulting variant cloned cell lines are found two types of alterations in the enzyme hypoxanthine phosphoribosyl transferase (EC 2.4.2.8) (1): enzyme with altered kinetic constants causing in vivo and in vitro resistance to 8-azaguanine; and (2) enzyme with altered heat sensitivity in vitro. These results support the view that tissue culture cell variants can arise from structural gene mutations.
TL;DR: It is suggested that the direction and extent of nerve fiber growth may be related to the density of the normal potential sympathetic innervation of the tissues, and that a chemical substance diffusing from the explants is involved in this process.
TL;DR: While this material corrected the prolonged clotting time of factor VIII (AHF) deficient plasma, it is shown that no AHF antigen was present in the cell cultures, and it is demonstrated this material to be tissue factor by coagulation techniques and immunological cross-reactivity.
Abstract: The procoagulant material of lymphocytes has been characterized as tissue factor. Lymphocytes stimulated with phytohemagglutinin or the purified protein derivative of the tubercle bacillus developed procoagulant activity with incubation in tissue culture. While this material corrected the prolonged clotting time of factor VIII (AHF) deficient plasma, we have shown, utilizing a sensitive radioimmunoassay, that no AHF antigen was present in the cell cultures. Further, we have demonstrated this material to be tissue factor by coagulation techniques and immunological cross-reactivity. The published data regarding factor VIII synthesis is reviewed in light of these observations and comments are made regarding the role of the lymphocyte procoagulant.
TL;DR: Under the culture conditions employed, liver cells appear to have a limited replicative life span and growth potential showed that there was a negative correlation between age of the donor tissue andAge of the culture at the last passage, maximum number of doublings, number ofDoublings in phase II, and age ofThe culture atThe end of phase II.
Abstract: The morphological aspects of ageing and growth capacity were studied in 100 lines of human liver cells. Under the culture conditions employed, liver cells appear to have a limited replicative life span. The morphological effects of ageing were similar to those described in the tissue culture of other organs. Studies on growth potential showed that there was a negative correlation between age of the donor tissue and age of the culture at the last passage, maximum number of doublings, number of doublings in phase II, and age of the culture at the end of phase II. The growth indices were different for epithelial-like and fibroblast-like cells.
TL;DR: It is found that tyrosine hydroxylase activity in N1E-115 is related inversely to the rate of cell division and/or cell density in culture (ref. 2), and from early log to late stationary phase of growth, its activity increases more than 30-fold.
Abstract: Regulation of catecholamine biosynthesis can be studied in mouse neuroblastoma cells growing in tissue culture. We have isolated adrenergic clones1 and have studied in particular clone N1E-115 which has very high levels of tyrosine hydroxylase (EC 1.14.3a), the first enzyme within the adrenergic cell in the biosynthetic pathway to catecholamines. We found that tyrosine hydroxylase activity in N1E-115 is related inversely to the rate of cell division and/or cell density in culture (ref. 2 and E. R. and M. Nirenberg, manuscript in preparation) so that from early log to late stationary phase of growth, its activity increases more than 30-fold. The catecholamine precursors, phenylalanine and tyrosine, enter these cells by high affinity transport systems3 and are converted into catechols (E. R., unpublished data).
TL;DR: Findings combined with the finding of viral-specific RNA indicate that portions of the viral genome are being expressed with a much greater frequency than previously appreciated.
Abstract: A radioimmunoassay specific for a murine leukemia virus structural protein, the gs antigen, detects an antigenic reactivity in normal murine cells in culture and natural tissues. The assay was shown to measure an antigen that is highly related to the virion protein as shown by absorption tests, immunoadsorbent chromatography, and by analysis of linearized dose-response curves. These findings combined with the finding of viral-specific RNA indicate that portions of the viral genome are being expressed with a much greater frequency than previously appreciated.
TL;DR: A high proportion of diluted serums of patients with amyotrophic lateral sclerosis were toxic to the anterior horn cells of the mouse in tissue culture, and Serums from other degenerative neurological diseases were inactive.
Abstract: A high proportion of diluted serums of patients with amyotrophic lateral sclerosis were toxic to the anterior horn cells of the mouse in tissue culture. This is not a general cytotoxicity, since apparently only the neurons were killed. Serums from other degenerative neurological diseases were inactive.
TL;DR: The advantage of diploid human skin flbroblastoid cell cultures is that they permit controlled studies of individual strain variation to an extent not possible with the better characterized human embryo lung lines.
Abstract: Publisher Summary This chapter focuses on human skin fibroblasts cultures. The advantage of diploid human skin flbroblastoid cell cultures is that they permit controlled studies of individual strain variation to an extent not possible with the better characterized human embryo lung lines. Except for a variable but significant degree of tetraploidy and occasional nondisjunctional progeny, the genotype of the cells established in culture is that of the donor. Many methods of skin biopsy provide material satisfactory for the establishment of predominantly diploid flbroblastoid cultures. The procedure discussed in the chapter employs a sterile skin prep of pHisoHex, sterile water rinse, and 70% ethanol. Local anesthetics are ordinarily not necessary for pinch, wedge, or punch biopsies; when used for surgical ellipse biopsies, the tissue which is to be used for culture should not be infiltrated. Autopsies provide an excellent source of culture material. In the laboratory, lines have been established from well over 100 refrigerated subjects. Except for rare microbial contamination, the success rate is virtually 100%. The biopsy tissues are best stored and transported in a standard sterile tissue culture medium with 10 to 20% newborn or fetal calf serum.
TL;DR: Inoculation of the Shope virus in tissue cultures of human fibroblasts from a patient with a deficiency of the enzyme arginase results in an induction of arginases activity, apparently virus coded.
Abstract: Inoculation of the Shope virus in tissue cultures of human fibroblasts from a patient with a deficiency of the enzyme arginase results in an induction of arginase activity, apparently virus coded.
TL;DR: It is reported that extracts from mesenchymal tissues will replace intact tissue in the normal development of rat pancreatic epithelia in vitro and recently shown that MF dramatically stimulates epithelial DNA synthesis and cell proliferation.
Abstract: The normal embryological development of many, if not all, organs depends on interactions between epithelial and mesenchymal tissues1. Some epithelial components require their own mesenchyme, while others, for example pancreatic epithelia, develop with any of a variety of mesenchymes1, 2. We have reported that extracts from mesenchymal tissues will replace intact tissue in the normal development of rat pancreatic epithelia in vitro2–5. The causal activity present in these extracts has been termed mesenchymal factor (MF). This activity appears to be a specific component of embryonic mesenchymal tissue; it has been detected only in extracts of tissues rich in mesenchyme and in tissue culture cells derived from mesenchyme5. No activity has been detected in extracts of epithelial cells, in a variety of adult tissues and in several embryonic organs such as liver, brain or heart. None of the common hormones or growth factors tested has MF activity2, 5. MF has been purified from chicken embryos and has the characteristics of a glycoprotein (ref. 5 and unpublished results). In addition to its reported effects on cytodifferentiation (refs 2–5 and unpublished results), we have recently shown that MF dramatically stimulates epithelial DNA synthesis and cell proliferation5.
TL;DR: It is shown that mouse peritoneal exudates contain cells that can give rise to colonies when grown in agar.
Abstract: Cells from the haematopoietic tissues of laboratory animals or humans can form colonies when grown in soft agar culture containing either a feeder layer or exogenous colony stimulating factors1–5. Although peritoneal exudate cells, consisting chiefly of macrophages and lymphocytes, proliferate in vitro6, 7 there is no information on whether these cells can form colonies. Here we show that mouse peritoneal exudates contain cells that can give rise to colonies when grown in agar.
TL;DR: For instance, this paper showed that heparin and Treburon (a synthetic polysaccharide-sulfuric ester) are structurally and chemically related to heparins, and showed that dextran sulfate was as effective in increasing the activity of the enzyme using collagen in the solid state as a substrate.
Abstract: The amount of mouse bone collagenase recovered in the tissue culture medium of bone culturedin vitro was increased by the addition of heparin at an optimal concentration of approximately 50 units/ml of tissue culture medium. Dextran sulfate and Treburon (a synthetic polysaccharide-sulfuric ester) which are structurally and chemically related to heparin were as effective as heparin in increasing the amount of mouse bone collagenase recovered in the tissue culture medium. In addition to stimulating the synthesis and/or release of mouse bone collagenase, heparin was also found to increase the specific activity of both crude and purified preparations of the enzyme when assayed using collagen in the solid state as the substrate, but showed no enhancement of enzyme activity when assayed using collagen in solution as the substrate. Dextran sulfate was as effective as heparin in increasing the activity of the enzyme using collagen in the solid state as a substrate. Neither heparin or dextran sulfate enhanced the activity ofClostridium histolyticum collagenase. For the first time, a purified tissue collagenase has been shown to both degrade and solubilize undenatured, insoluble tissue collagen at 37°. Moreover, since this action was markedly enhanced by the addition of heparin, it suggests that heparin and similar substances may play an important role in the regulation of collagen degradation during the remodeling of collagenous tissuesin vivo.
Journal Article•
TL;DR: Adult human retinal pigment epithelium has been enzymatically isolated and established In tissue culture andMelanin granules decreased with the age of the culture, became less pigmented but retained some characterstics of melanin-producing cells.
Abstract: Adult human retinal pigment epithelium has been enzymatically isolated and established In tissue culture. Primary cultures survived three to six months. Occasionally, cell lines spontaneously evolved from primary cultures. Melanin granules decreased with the age of the culture, became less pigmented but retained some characterstics of melanin-producing cells. Lipofuscin granules are present, but differ from lipofuscin of testicular origin. Mitotic activity is present throughout the life of the cultures.
TL;DR: Tumor cell lines exposed to immunoglobulins specific for cell surface antigens developed increased cellular incorporation of [125I]iododeoxyuridine and [3H]thymidine, which may have significance in regard to enhancing effects of antibodies on tumor cell growth in vivo.
Abstract: Tumor cell lines exposed to immunoglobulins specific for cell surface antigens developed increased cellular incorporation of [125I]iododeoxyuridine and [3H]thymidine (up to 200-fold increases over cells treated with normal rabbit immunoglobulins). Antibody-stimulated cells multiplied more rapidly and lived longer than control cells in tissue culture. These observations were made both with cells substituted with 2,4,6-trinitrophenol and purified antibody against 2,4,6-trinitrophenol, and with several cell lines and their respective whole-cell antibodies. Antibodies that were stimulatory at low concentrations were cytotoxic at high concentrations. These observations may have significance in regard to enhancing effects of antibodies on tumor cell growth in vivo.
Journal Article•
TL;DR: The results indicate that polyunsaturated fatty acids are probably not indispensible to the normal functioning of the individual cell and the possibility that they perform additional essential functions in intact tissues is still an open question.
Journal Article•
TL;DR: In this article, the free fatty acid (FFA) pool of corticosteroid-sensitive mouse lymphosarcoma P1798S cells was investigated in vitro.
Abstract: When cells of the thymus or corticosteroid-sensitive mouse lymphosarcoma P1798S are treated in vitro with 0.27 µm cortisol in tissue culture medium with 10% serum or 0.5% human albumin, nuclear damage occurs that ends with karyorrhexis. The steroid-resistant subline P1798R does not show these changes.
Corticosteroid-sensitive lymphocytes that undergo lysis differ from the resistant subline in several respects with regard to changes in fatty acid metabolism. Corticosteroid treatment in vivo for 2 hr raised the free fatty acid (FFA) pool in thymus and P1798S cells, while decreasing it in P1798R cells. In vitro , cortisol had no effect on the uptake of 14C-labeled palmitic acid but decreased oxidation of this acid by 46 and 17% in thymus and P1798S, respectively, while increasing it 9% in P1798R.
After sensitive P1798S cells were incubated in a medium that contained FFA calculated to be equivalent to that accumulated after steroid treatment, electron microscopy revealed that certain effects of corticosteroids could be reproduced by fatty acids of chain length C-9 and higher, namely, nuclear edema, focal dissolution and disintegration of the nuclear membrane and, ultimately, karyolysis. Steroid-resistant cells show cytological changes only at 10-fold higher concentrations of FFA.
On the basis of these results, the following scheme is proposed as the mechanism by which cytolysis occurs in corticosteroid-sensitive lymphoid tissues:
![Figure][1] [1]: pending:yes