PubMed Health⌕ Search

Biomedical subjects

I Yamane

Publications and source records attributed to I Yamane.

At least 55 records · Page 3Linked to original sources
PubMed1987

Role of iron chelators in growth-promoting effect on mouse hybridoma cells in a chemically defined medium.

The role of various iron chelators on the multiplication of mouse hybridoma cells in an albumin-free, transferrin-deficient defined medium was investigated. Fe(III)-dihydroxyethylglycine, Fe(III)-glycylglycine, Fe(III)-ethylenediamine-N,N'-dipropionic acid, or Fe(III)-iminodiacetic acid supported the excellent growth of the cells. In addition, the growth of the iron-starved cells, which had been preincubated in a protein-, iron- and chelator-free defined medium, restored rapidly when the medium was supplemented with holotransferrin, ferric iron, and chelator compared to that when supplemented with holotransferrin, but without iron and chelator. The results suggest that such chelators modulate a progression of transferrin cycle in the presence of transferrin and ferric iron. An alternative explanation is that there is a decrease in generation of iron-catalyzed free radicals.

Animals↗
PubMed1987

Increased tumorigenicity of polyploid Ehrlich variants during growth in vivo is associated with karyotypic changes.

In previous studies, treatment of Ehrlich ascites tumors (EAT) with either Sendai virus, Herpes virus or neuraminidase produced variants that differed significantly from EAT in both tumorigenicity and karyotype. In this study, less tumorigenicity variants of EAT were isolated by culturing in media containing low concentrations of serum or without serum. In contrast to the relative stability of tumorigenicity in such variants cultured in vivo, the tumorigenicity of these variant clones passaged in vivo was variable, depending on the variant clone. During growth in vivo, 3 variant clones independently isolated from serum-free medium, low serum-containing medium and after neuraminidase treatment produced markedly heterogeneous tumors, and after 4 passages the tumors were about 1000 times more tumorigenic than the initial variants. In generating new tumors, the tumors always segregated chromosomes. The banded chromosome analysis of a variant clone isolated from serum-free medium and its malignant tumor showed that most of the chromosomes segregated from the clone were normal types. On the other hand, the tumorigenicity of Herpes virus-induced variants was unchanged during growth in vivo.

Animals↗
PubMed1987

Effects of extracellular matrices on human keratinocyte adhesion and growth and on its secretion and deposition of fibronectin in culture.

Extracellular matrices (ECMs) play an important role as components of basement membrane of normal human skin and in migrating epidermal cells in wound healing. We investigated the effects of various ECMs on human keratinocyte adhesion and growth as well as on its secretion and deposition of fibronectin (FN) in vitro using a serum-free, low-calcium culture system. Since cell adhesion is the first step of cell growth, we performed cell adhesion assay for 14 h. Human keratinocytes adhered best on FN and less well on types I/III collagen, type IV collagen, and heparan sulfate proteoglycan (HSPG) as compared with bovine serum albumin (BSA) (control) or laminin (La). Cell growth assayed for 7-8 days on the dishes coated with various extracellular matrices revealed significantly increased keratinocyte growth on FN and on types I/III collagen in comparison with that on type IV collagen, HSPG, BSA (control), or La. Morphology of keratinocytes and of their colonies on FN and types I/III collagen was strikingly different from that of the control; the colonies were not so compact as in the control, but rather loose and larger; each keratinocyte was spread out more on these substrata. These morphologic features seemed to correlate with the increased keratinocyte growth on these extracellular matrices. Both immunofluorescence study for FN with keratinocytes in 8-day culture on various extracellular matrices and enzyme-linked immunosorbent assay for FN measurement on substratum or in conditioned medium with keratinocytes in 5-day culture demonstrated that extracellular matrices modulated the secretion and deposition of FN by human keratinocytes in culture; the keratinocyte growth correlated with the amount of FN detected on substratum but not with that in medium. Based on the results of the present investigation, we think that the growth of human keratinocytes depends on the amount of FN on substratum.

Cell Adhesion↗
PubMed1987

[Analysis of interpersonal experience as a function of psychological distance and the level of acquaintanceship].

Psychological distance between self and others was assumed, semiologically, to be experienced in four aspects (active representation, active presentation, passive representation and passive presentation) which can assume different values from one another. It was further assumed that there is a behavioral tendency to reduce differences among these aspects by means of two types of behavioral principle: egoistic and contextual. With acquaintanceship classified into four levels (total acquaintance, peripheral acquaintance, known stranger including familiar stranger, and total stranger) in terms of psychological distance and behavioral norms, it was hypothesized that the 'other' person who embarrasses us most is an acquaintance by norm but a stranger psychically. The results of questionnaire surveys supported this hypothesis and confirmed that it is difficult to treat the peripheral acquaintance as an acquaintance.

Cognition↗
PubMed1987

[Primary cultures of various differentiated human cells and their transfer (3). Apparatus and instrument for primary culture].

Clean bench, water distilled, low speed centrifuge, CO2-incubator and invert microscope were described and discussed as those necessary for the primary culture of differentiated human cells. Also the instruments including knives, filter sterilization equipment, the reagents including culture media were described and discussed as ones necessary for those primary cultures.

Cell Differentiation↗
PubMed1987

[Primary cultures of various differentiated human cells and their transfer].

From our long experience with primary cultures of human differentiated cells, we have been able to come to the following conclusions. The culture media routinely employed have been developed using established cell lines but not primary cells as the growth marker. Therefore the culture media for primary cultures should be modified from those routinely employed. In addition the concentrated supplementation of serum to basal media dose not contribute to the growth of primary differentiated cells, and in fact on the contrary, is advantageous for the growth of non-target fibroblasts which would hamper the growth of the target cells. A hypoxic culture environment is more favorable for primary cultures especially when smaller cell numbers are used as the inoculum. The primary culture cells are more fragile in comparison with established cell lines. Therefore, the low-temperature cell diopersion procedure is recommended with the use of diluted crystalline trypsin saline. On the basis of the above findings, we have successfully carried out primary and transfer cultures of human esophageal and gall bladder epithelial cells, skin keratinocytes and endothelial cells. Their epithelial mature was proved by the presence of keratin in their cytoplasm, and the phenotype of endothelial cells was evident from the presence of Factor VIII-related antigen in their cytoplasm. Culture of endothelial cells requires the supplementation of a specific growth factor which we have utilized and isolated from conditioned medium of human diploid fibroblasts. Tiny amounts of the factor were shown to, induce the vascularization of rabbit eye cornea.

Animals↗
PubMed1987

[Primary cultures of various differentiated human cells and their transfer (2). The isolation of target cells from their specimen].

The specimen employed for the culture should be possibly rich of target cells in order to obtain the successful primary culture. When the target cells are located along the inner side of a duct forming tissue, they can be relatively easily isolated as major cell population in their whole dissegregated cells by means digesting the inner wall of ductive tissues. When the specimen supplied for the culture is so small and limited, the explant outgrowth culture method can be better applied for the culture. For the primary cultures of malignant cells, soft agar cell suspension method can be better available for the successful culture.

Agar↗
PubMed1986

Purification and characterization of an endothelial cell growth factor from serum-free culture medium of human diploid fibroblast cells.

Serum-free culture supernatants of human embryo fibroblast cells contain endothelial cell growth factor (f-ECGF) which supports the serial propagation of human umbilical vein endothelial cells in the serum-free culture medium (medium A). This growth-stimulating activity has been partially purified from serum-free culture-conditioned medium. The stability of the activity to acid (pH 4.0-4.5) was utilized for the first step in purification. f-ECGF had a high affinity to heparin-Sepharose CL-6B, and was isolated by the methods of heparin affinity, of ion-exchange and gel filtration chromatography from the serum-free culture-conditioned medium preparation. The purified f-ECGF had an isoelectric point in the pH range 4.5-6, and a molecular weight of approx. 30 kDa determined by either gel filtration or SDS-polyacrylamide gradient gel electrophoresis. The f-ECGF has high affinity for concanavalin A column, and the activity was partially eluted from the column with ethylene glycol and alpha-methylmannose. The results indicate that f-ECGF is an acidic-glyco-protein with heterogeneous sugar chain(s).

Cell Line↗
PubMed1986

Enhanced formation of mouse hybridomas without hat treatment in a serum-free medium.

A newly developed, serum-free medium (NYSF-404) selects for antibody-producing hybridomas after fusion of antigen-sensitized mouse spleen cells with myeloma cell lines P3-X63-Ag8-U1 (P3-U1), P3-X63-Ag8-6.5.3 (Ag8.653), or P3-NSI/1-Ag4-1 (NS-1). Without the need for hypoxanthine-aminopterin-thymidine (HAT) selection of hybrid cells, frequency of hybridoma formation in medium NYSF-404 is higher (twice) than that in serum- and HAT-containing medium. Colonies developed upon limiting dilution in the presence of the mortal parent myeloma cells in medium NYSF-404 and pure culture of antibody-secreting cells could be subsequently established. The results suggest that fusions can be done in serum-free medium and that the clonal growth of hybridomas is dependent on factors produced by parent myeloma cells under serum-free culture conditions. Such factors seem deficient in serum- and HAT-containing medium or are masked by serum.

Aminopterin↗
PubMed1986

Isolation and characterization of lectin-resistant mouse cell lines defective in their ability to yield hybrid cells.

Lectin-resistant variants of the mouse mammary tumor cell line FM3A in suspension were isolated after multiple-step selection using toxic concentrations of phytohemagglutinin (PHA) and concanavalin A (Con A). In fusion experiments, both PHA- and Con A-resistant lines were found to have lost the ability to yield hybrids with suspension-grown cells, although they were capable of yielding hybrids with monolayer-grown cells. This phenotype of PHA-resistant lines was stable during a long period of culture in the absence of PHA.

Animals↗
PubMed1986

Population-dependent requirements of vitamin B12 and metabolically related substances of several mouse cell types in serum-free, albumin-fortified medium.

Mouse FM3A cells propagated well in serum-free medium containing 0.5% serum albumin and 1 microgram of insulin/ml. The vitamin B12 (B12) requirement of the cells depended on the population density. This requirement disappeared when a sufficiently large cell population was present. A combination of 1-100 ng of B12/ml and 4 micrograms of hypoxanthine/ml resulted in a synergistic increase in cell growth at low cell densities. A similar growth response was obtained when the B12 plus hypoxanthine was replaced by 4 micrograms of hypoxanthine/ml in combination with 100 ng of thymidine/ml, 1 microgram of folic acid/ml or 1 microgram of folinic acid/ml, even though 1 microgram of folic acid/ml already was present in the medium. Experiments on single cell inoculation showed that colony size and the yield of cells grown in B12-supplemented medium were much larger than those for cells grown in B12-free medium. A more critical population-dependent B12 requirement was demonstrated in mouse Ehrlich and L cells and their hybrids. At less than 100 cells there was no propagation in serum-free medium lacking B12, folinic acid and thymidine; whereas, a satisfactory growth response was obtained in medium supplemented with these substances.

Animals↗
PubMed1985

Fibroblast products adsorb to the culture substrate and stimulate growth of human umbilical vein endothelial cells.

Human umbilical vein endothelial cells grew with a doubling time of approximately 20 hr in medium conditioned by human diploid fibroblasts and supplemented with 10% fetal bovine serum, whereas the cells did not grow substantially in the non-conditioned serum supplemented medium. Production of the fibroblast-derived activity required the presence of insulin, EGF, or PDGF. The fibroblast derived-factor adsorbed to native culture dishes or dishes coated with gelatin and collagen. The adsorbed activity was resistant to treatment with 1% Triton X-100, and was abolished by treatment with serine proteases. Further, the extracellular matrix produced by the fibroblasts also showed growth-stimulating activity. Fibroblast-derived factors may play a role in vascularization processes during wound healing, inflammation and normal development.

Adsorption↗
PubMed1985

Cell fusion and cell agglutination: enhancing effect by a combined use of lectin and polycation.

The technique efficient for hybridization of mammalian cells was improved by combining a procedure of cell agglutination of cells pretreated with a combination of lectin and polycation and the procedure of conventional polyethylene glycol (PEG) -induced cell fusion. The agglutinability of cells treated with lectin, polycation, and both of them was tested. The effects of these agglutinogens on the hybridization frequencies of cells were also compared. The appearance rate of hybrid colonies was found to be correlated to the extent of cell agglutination. The pretreatment with a combination of lectin and polycation induced the highest degree of cell agglutination and the highest frequency of resulting hybridization. The enhancing effect by the pretreatment with a combination of these agglutinogens on the hybridization frequency was confirmed through experiments on the crosses of several cell types including nuclear fraction (karyoplast).

Agglutination↗
PubMed1985

Long-term serial cultivation and growth requirements for human umbilical vein endothelial cells.

Human umbilical vein endothelial cells (HUV-EC) grew rapidly in vitro in medium supplemented with epidermal growth factor, fetal bovine serum (FBS) and human diploid fibroblast-conditioned medium. The effect of FBS could be replaced partially by bovine serum albumin, cholesterol, and vitamin E, and completely by further addition of serum dialysate or refeeding every other day. Among these components, fibroblast-conditioned medium is essential for HUV-EC growth. The HUV-EC were cultured serially for over 50 population doublings in the 10% FBS containing fibroblast-conditioned medium and for over 40 population doublings in the serum-free medium. Mitogenic factor(s) present in the medium conditioned by fibroblasts may be related to endothelial cell growth factor and play an important role in angiogenesis and regeneration of vascular endothelium in vitro.

Cell Division↗
PubMed1985

Cell hybridization and cell agglutination. I. Enhancement of cell hybridization by lectins.

A great increase in hybridization frequency of cultured rodent cells was obtained when conventional cell fusion using 50% polyethylene glycol (PEG) was combined with a cell agglutination produced by plant lectins. The rate of appearance of hybrid colonies was found to be correlated with the extent of cell agglutination by lectin, as well as with cell fusion induced by subsequent PEG treatment. Phytohemagglutinin (PHA), wheat germ agglutinin, Wistaria floribunda agglutinin and concanavalin A were all active; the most effective was PHA. When parental cells in a monolayer were treated with PHA followed by PEG, the resulting hybridization frequency was very low because of markedly decreased viability, whereas the same cells in suspension yielded hybrid colonies at a higher rate. These results suggest that the enhancement of hybridization by PHA/PEG treatment was brought about by the ability of lectin to agglutinate cells.

Agglutination↗
PubMed1985

Cell hybridization and cell agglutination. II. Enhancement of cell hybridization by polycations.

An efficient technique for hybridization of mammalian cells was developed by combining agglutination by pretreatment with polycations, such as polyarginine, and conventional polyethylene glycol(PEG)-mediated cell fusion. Polyarginine and subsequent PEG treatment resulted in markedly decreased viability in the treated cells, but addition of polyvinyl pyrrolidone or glycerol to the polyarginine prevented this cytotoxicity. Polyarginine was much more effective than polylysine or polyornithine in inducing hybridization. Other polycations, including polybrene and protamine but not DEAE-dextran, were also active in inducing hybridization. The condition of the cells at the time of polycation treatment was an important factor in the enhancement of hybridization. The condition of the cells at the time of polycation treatment was an important factor in the enhancement of hybridization. The enhancement of hybridization of cells in monolayer incubated for 2 h was much higher than that of cells incubated for 24 h. These findings suggest that polycations do not necessarily operate by agglutinating cells. The mechanism of polycation-enhanced cell hybridization is discussed.

Agglutination↗