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Biomedical subjects

T Maciag

Publications and source records attributed to T Maciag.

122 records · Page 7Linked to original sources
PubMed1982

Organizational behavior of human umbilical vein endothelial cells.

Culture conditions that favor rapid multiplication of human umbilical vein endothelial cells (HUV-EC) also support long-term serial propagation of the cells. This is routinely achieved when HUV-EC are grown in Medium 199 (M-199) supplemented with fetal bovine serum (FBS) and endothelial cell growth factor (ECGF), on a human fibronectin (HFN) matrix. The HUV-EC can shift from a proliferative to an organized state when the in vitro conditions are changed from those favoring low density proliferation to those supporting high density survival. When ECGF and HFN are omitted, cultures fail to achieve confluence beyond the first or second passage: the preconfluent cultures organize into tubular structures after 4-6 wk. Some tubes become grossly visible and float in the culture medium, remaining tethered to the plastic dish at either end of the tube. On an ultrastructural level, the tubes consist of cells, held together by junctional complexes, arranged so as to form a lumen. The smallest lumens are formed by one cell folding over to form a junction with itself. The cells contain Weibel-Palade bodies and factor VIII-related antigen. The lumens contain granular, fibrillar and amorphous debris. Predigesting the HFN matrix with trypsin (10 min, 37 degrees C) or plasmin significantly accelerates tube formation. Thrombin and plasminogen activator had no apparent effect. Disruption of the largest tubes with trypsin/EDTA permits the cells to revert to a proliferative state if plated on HFN, in M-199, FBS, and ECGF. These observations indicate that culture conditions that do not favor proliferation permit attainment of a state of nonterminal differentiation (organization) by the endothelial cell. Furthermore, proteolytic modification of the HFN matrix may play an important role in endothelial organization.

Cell Differentiation↗
PubMed1982

Bovine fibroblast growth factor: comparison of brain and pituitary preparations.

Bovine brain and pituitary fibroblast growth factors (FGF) have been compared with regard to their chemical and biological properties. Pituitary and one preparation of brain FGF (Prep A) contain a basic mitogenic activity, which migrates to the same position on electrophoresis in acid pH gels as detected by incorporation of [methyl-3H]-thymidine into BALB/c 3T3 cells. In contrast, another preparation of brain FGF (Prep B) contains two mitogens, one (20-30%) indistinguishable from the basic components in pituitary and brain (Prep A) FGF preparations and an acidic activity (70-80%), pl 5-6, that migrates more slowly on acid gels, corresponding to the acidic component of brain FGF described previously (Thomas, K. A., M. C. Riley, S. K. Lemmon, N. C. Baglan, and R. A. Bradshaw. 1980. J. Biol. Chem. 255:5517-5520.) In agreement with that report, none of the mitogens comigrates with fragments of myelin basic protein. Pituitary FGF was virtually inactive, brain (Prep A) FGF had a small amount of activity, and brain (Prep B) FGF was highly potent (50% maximal stimulation at 15-30 ng/ml) in stimulating the growth of human umbilical vein endothelial (HUVE) cells. The acidic component of brain FGF, which is much more unstable at pH 8.5 than the basic one, can be protected by reducing agents, whereas the basic constituent of brain FGF as well as pituitary FGF is unaffected by reducing conditions. Thus, brain FGF preparations may contain two distinct mitogenic activities, one that is acidic and contains HUVE cell activity, and a basic mitogen that is similar to and may be identical with pituitary FGF.

Animals↗
PubMed1981

Hormonal requirements for growth of arterial smooth muscle cells in vitro: and endocrine approach to atherosclerosis.

In this study the hormonal requirements for the growth of arterial smooth muscle cells in vitro were determined. A serum-free, biochemically defined medium, supplemented with the relevant hormones, permitted proliferation and propagation of normal diploid mammalian arterial smooth muscle cells. Serum-free, hormone-supplemented cultures spontaneously formed atherosclerotic plaque-like nodules. Thus atherosclerosis may be mediated by a complex endocrine system.

Animals↗
PubMed1981

An endocrine approach to the control of epidermal growth: serum-free cultivation of human keratinocytes.

Human keratinocytes, derived from the skin of newborns and of adults, were grown in the complete absence of serum, in a hormone-supplemented medium on fibronectin-coated cell culture dishes at low seed density. The cell culture medium consisted of Medium 199 containing epidermal growth factor, triiodothyronine, hydrocortisone, Cohn fraction IV, insulin, transferrin, bovine brain extract, and trace elements. Removal of the brain extract from the hormone supplement had a greater negative impact on proliferation of the keratinocyte cultures than did the removal of epidermal growth factor, hydrocortisone, and triiodothyronine or Cohn fraction IV. The growth of keratinocytes in this hormone-supplemented medium suggests that control of keratinocyte growth depends in large part on endocrine stimulation by other body organs, including the brain.

Brain Chemistry↗
PubMed1981

Serial propagation of human endothelial cells in vitro.

Human umbilical vein (HUV) endothelial cells were grown for 15 to 21 passages at a split ratio of 1:5 (at least 27 population doublings) on a human fibronectin (HFN) matrix in Medium 199 supplemented with fetal bovine serum (FBS) and endothelial-cell growth factor (ECGF). This system also permitted the growth of HUV endothelial cells at cell densities as low as 1.25 cells/cm2. In addition to delaying the premature senescence of HUV endothelial cells, ECGF also reduced the serum requirement for low-density HUV endothelial-cell growth; 2.5% serum and ECGF yields half-maximum growth as compared to high serum controls. Significant HUV endothelial-cell growth was also observed in medium supplemented with either ovine hypophysectomized (HYPOX) serum, plasma-derived serum (PDS), or HYPOX-PDS in the presence of ECGF, suggesting that neither the pituitary nor the platelet contributes to HUV endothelial-cell growth.

Animals↗
PubMed1981

The morphological and biochemical characterization of a line of rat promegakaryoblasts.

Biochemical and morphological evidence is presented to support the characterization of a rat bone-marrow-derived cell line (RPM) as an analog of the promegakaryoblast. The conditions for in vitro growth and maturation of the RPM line are described. Rapid proliferation of the RPM line is readily achieved when cultures are supplemented with moderate levels of fetal bovine serum (FBS). The proliferative compartment is a small blast-like cell. Immunofluorescent staining demonstrates that the RPM cells contain factor VIII:antigen and fibrinogen in their cytoplasm. The cells secrete, into their conditioned medium, a potent mitogenic activity for rat aortic smooth muscle cells. When incubated under conditions of relative serum deprivation, the cells stop proliferating and undergo a process of maturation. The sequence of maturation is described as stage I (promegakaryoblast--the proliferative compartment); stage II (immature megakaryocyte or promegakaryocyte); stage III (mature megakaryocyte). The stage III cells release, from their cytoplasm, small membrane-bound vesicular bodies containing lavender granules and cytoplasmic organelles. These have been designated stage IV. The RPM line may provide a useful model for the in vitro study of megakaryocyte maturation and the synthesis of megakaryocyte-specific proteins.

Animals↗
PubMed1980

Hormonal requirements of baby hamster kidney cells in culture.

Baby hamster kidney cells (BHK) were able to proliferate in the complete absence of serum in synthetic medium supplemented with insulin, transferrin, fibroblast growth factor and epidermal growth factor on cell culture dishes coated with fibronectin. Although the addition of individual supplements had little or no effect on cell growth, the combination of the supplements resulted in a significant synergistic effect. The defined medium was also capable of supporting the clonal growth of BHK cells in the absence of serum. Studies on the effect of hormonal supplementation in the presence of low concentrations of fetal calf serum suggest that the function of serum is to supply hormones and growth factors for cell growth.

Animals↗
PubMed1980

Growth of human keratinocytes on fibronectin -coated plates.

Keratinocytes derived from the skin of newborns and of adults aged 19 to 57 years were grown on plates coated with human fibronectin (HFN) in the absence of a 3T3 monolayer. The cells grew well, attained confluence and could be sub-cultivated at densities approximately 10% of those necessary for successful cultivation of human keratinocytes on collagen coated dishes. Growth was excellent at concentrations of fetal bovine serum (FBS) as low as 5%, and appreciable growth occurred over a six day period even in the complete absence of serum. Growth was enhanced by addition of cholera toxin to the medium. Fibroblast overgrowth of the keratinocyte colonies was not observed. The observation that keratinocytes grow well on fibronectin in the absence of a fibroblast feeder-layer should simplify further study of this fastidious cell type and increases our understanding of keratinocyte growth requirements in vitro.

Adult↗
PubMed1979

An endothelial cell growth factor from bovine hypothalamus: identification and partial characterization.

Extracts of bovine hypothalamus were found to contain a significant level of mitogenic activity when tested in a Swiss 3T3 cell [3H]dThd incorporation assay and in a human umbilical vein endothelial cell growth assay. The mitogenic activity responsible for 3T3 cell activity was purified and characterized as a fibroblast growth factor (FGF)-like mitogen. Neither the biologically active FGF-like mitogen purified from the hypothalamus extracts nor FGF purified from bovine pituitary glands was mitogenic when added to human endothelial cells in vitro, suggesting the presence of more than one mitogen in the hypothalamic extracts. The 3T3 and endothelial cell biological activities of hypothalamic extracts were both found to be inactivated by trypsin, subtilisin, and heat treatment, but were stable to dialysis. The endothelial cell growth factor activity could be efficiently separated from the FGF activity by gel exclusion chromatography. The endothelial cell mitogen possessed a molecular weight of approximately 75,000, whereas that of FGF was approximately 15,000. The endothelial cell growth factor activity was found to be inactivated with reducing agents whereas the 3T3 cell mitogenic activity was stable after incubation with 2-mercaptoethanol. Significant levels of endothelial cell mitogenic activity were also found in extracts of bovine brain and pituitary glands.

Animals↗
PubMed1979

The biosynthetic precursor of epidermal growth factor and the mechanism of its processing.

The biosynthesis of epidermal growth factor (EGF) was studied in mouse submaxillary glands incubated with L-[35S]cystine. EGF and EGF-like proteins were isolated from the gland homogenates by immunoprecipitation with anti-EGF antiserum. The major species appearing after short labeling periods is significantly larger (Mr, 9000) than EGF. The label in the Mr 9000 species plateaus after 1 hr whereas tha in EGF continuously increases. When glands are chased with unlabeled L-cystine after a brief period of labeling, the Mr 9000 peak decreases and a corresponding amount of label appears in EGF. The Mr 9000 species was isolated from boiled homogenates in which it accounts for approximately 1% of the total EGF content. It contains five of the six chymotryptic peptides of EGF and a sixth peptide which is a modified form of the COOH-terminal chymotryptic peptide of EGF. Of the arginyl esteropeptidases, gamma subunit of 7S nerve growth factor, beta-endopeptidase, trypsin, and EGF-binding protein, only the latter converts the isolated Mr 9000 species to EGF. The extrapeptide material released in the conversion comes from the COOH terminus of the Mr 9000 species. These results suggest that the Mr 9000 species is a biosynthetic precursor of EGF and that the EGF-binding protein is the specific intracellular cleaving enzyme that converts the precursor to EGF. In the process, the stable high molecular weight complex of EGF is formed.

Animals↗