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51.
Masahiro Miyazaki Yasunori Suzuki Munehiro Oda Akira Kawai Liyan Bai Jiro Sato 《In vitro cellular & developmental biology. Plant》1989,25(9):839-848
Summary For serum-free primary culture of adult rat hepatocytes, a synthetic medium DM-160 and rat-tail collagen were selected for
the basal medium and for the culture substratum, respectively. Barbiturates, such as phenobarbital and 1-ethyl-5-isobutylbarbiturate,
efficiently supported survival of hepatocytes and maintained their morphologic features at lower concentrations under the
serum-free conditions than under the serum-supplemented conditions. However, the hepatocyte survival rates under the serum-free
conditions were lower than those under the serum-supplemented conditions in the presence or absence of barbiturates. Supplementation
of the basal medium with a combination of five groups of factors (5Fs), such as eight amino acids (Ala, Arg, Gly, Ile, Met,
Phe, Pro, and Trp), two unsaturated fatty acids (linoleate and oleate), a protease inhibitor (aprotinin), three vitamins (A,
C, and E), and five trace elements (Mn, Fe, Cu, Zn, and Se), improved the hepatocyte survival under the serum-free conditions
in the presence or absence of barbiturates. In other words, the serum could be completely substituted by the 5Fs. Hepatocyte
cultures maintained in the 5Fs-suppelemented basal medium showed excellent induction of tyrosine aminotransferase activity
in response to dexamethasone in the presence or absence of barbiturates. The efficiency of the 5Fs-supplemented basal medium
for maintaining hepatocytes was not inferior to those of other media in common use with hepatocytes, such as Williams' medium
E and Waymouth's medium MB-752/1. In conclusion, maintenance of functional hepatocytes in serum-free primary culture could
be improved by use of the new medium preparation (the 5Fs-supplemented DM-160) in the presence of barbiturates.
This work was supported by a grant no. 61771923 from the Ministry of Education, Science and Culture of Japan. 相似文献
52.
Katrin Engelmann Peter Friedl 《In vitro cellular & developmental biology. Plant》1989,25(11):1065-1072
Summary Long-term cultivation of human corneal endothelial cells (HCEC) was optimized with respect to different components of the
culture system: 25 different nutrient media, different sera, 6 mitogens and various substrates were tested in their ability
to influence clonal growth and morphology of HCEC. F99, a 1∶1 mixture of the two media M199 and Ham’s F12, was the most effective
basal medium in promoting clonal growth of HCEC. Among various sera, human serum and fetal bovine serum showed optimal growth
promoting activities in combination with F99, whereas newborn bovine serum (NBS) was by far superior for the development of
a typically corneal endothelial morphology. Crude fibroblast growth factor (FGF), or alternatively endothelial cell growth
supplement, was absolutely essential for clonal growth of HCEC at low serum concentrations, for example 5% NBS. Formation
of a monolayer with a morphology similar to corneal endothelium in vivo was observed only on culture dishes coated with basal
membrane components such as collagen type IV, laminin, or fibronectin. The most pronounced effect on morphologic appearance
was obtained by culturing the cells on the extracellular matrix (ECM) produced by bovine corneal endothelial cells. Moreover,
ECM could substitute for crude FGF in clonal growth assays. 相似文献
53.
本文用正交实验设计法探讨了杂色曲霉(Aspergillus versicolr)在加有两类不同性质营养物的人胃液中产生杂色曲霉素(Sterigmatocystin,简称ST)的条件。发现在26℃斜面静置培养12天后可产生ST。加入半合成物质的最佳配伍是:蔗糖1,000.0mg;蛋白胨50.0mg;KH_2PO_4 7.5mg;MgSO_4·7H_2O_2.5mg;人胃液10.0ml,称之为SPKM人胃液培养基。加入天然物质的最佳配伍是:玉米粉0.5g;豆腐粉0.25g,人胃液10.0ml,称之为CS人胃液培养基。还进一步研究了pH值和培养时间对杂色曲霉产毒菌株在SPKM和CS人胃液培养基中生长及产生ST的影响。根据临床胃酸缺乏程度分级标准,分为pH 1.0,3.0,6.5,8.0四个组。发现在两种人胃液培养基中,无论是杂色曲霉生长,还是产生ST,pH3.0到6.5是发生质变的范围。在两种人胃液培养基pH为6.5时,37℃静置培养8天有痕量ST产生,10天后就明显增加。所以杂色曲霉产生的ST可能是慢性萎缩性胃炎易癌变的原因之一。 相似文献
54.
Influence of single amino acids on the development of hamster one-cell embryos in vitro 总被引:1,自引:0,他引:1
One-cell hamster embryos placed in culture have always shown a complete block to development at the two-cell stage. In a preliminary study using a chemically defined culture medium containing 20 amino acids (HECM-1), many one-cell embryos were able to escape the "two-cell block" and develop to the four-cell stage. Use of a simpler formulation containing only the amino acids hypotaurine and glutamine revealed marked inhibitory and stimulatory effects of adding the other amino acids. In the first experiment, 19 amino acids were separately examined for effects on one-cell embryo development. Six amino acids (phenylalanine, valine, isoleucine, tyrosine, tryptophan, and arginine) inhibited embryo development (reduced mean cell number; MCN), and three others (glycine, cystine, and lysine) stimulated development (increased MCN), compared with basic medium containing only glutamine and hypotaurine (low control). When the responses with the six inhibitory amino acids were totalled, only 3 of 185 (2%) one-cell embryos reached the six-or seven-cell stage compared to a total of 15 of 76 (20%) embryos that developed to these stages using the three stimulatory amino acids. When tested together in a second experiment, the six inhibitory amino acids significantly reduced the MCN, from 4.28 +/- 0.44 (low control) to 3.71 +/- 0.55. In this group, 17 of 117 (15%) of one-cell embryos reached more than four-cell and only 4 of 117 (3%) reached six- or 7-cell stages, compared with 39 of 117 (33%) and 12 of 117 (10%), respectively, for the basal medium group.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
55.
Charlotte L. Branchaud Cynthia G. Goodyer Harvey J. Guyda Yves Lefebvre 《In vitro cellular & developmental biology. Plant》1990,26(9):865-870
Summary We have compared hormone production by early gestation and term human placental trophoblasts cultured in Ham's F10 medium
containing 10% fetal bovine serum with that by cells cultured in serum-free HB102 medium. Mean daily production of progesterone
on Days 3 to 7 was approximately 25% less by both early gestation and term cells cultured in HB102 as compared to Ham's F10,
but production was maintained at a stable level for at least 7 d longer than the cells in Ham's. Estradiol production from
10−6
M dehydroepiandrosterone by both early gestation and term cells was comparable in both media. Human placental lactogen production
on Days 3 to 7 was 40% less by cells cultured in HB102. Human chorionic gonadotropin (hCG) output by early gestation cells
was also 50% less in HB102 but term cells in HB102 produced twice as much hCG as those in Ham's F10. 3B-Hydroxysteroid dehydrogenase
(3BHSD) activity in early gestation and term cells and 11B-hydroxysteroid dehydrogenase (11BHSD) activity of early gestation
cultures was comparable in the two media. 11BHSD activity was decreased in the term cultures, and this decrease was more marked
in Ham's than in HB102. Sulfatase and aromatase activities in the early gestation cultures were comparable in both media;
sulfatase activity was comparable and aromatase activity only 20% less in the term cells cultured in HB102. These results
indicate that serum-free HB102 supports differential function of human trophoblast cells and is useful for studies of placental
activity for as long as 14 d in culture. 相似文献
56.
Frederick J. Darfler 《In vitro cellular & developmental biology. Plant》1990,26(8):769-778
Summary A protein-free medium, termed ABC, has been developed which essentially eliminates the need for serum proteins. ABC supports
the long-term growth of murine hybridomas as well as other transformed cells of the immune system. The requirement of hybridoma
growth for transferrin has been met by substituting the soluble organo-iron compound, sodium nitroprusside. Substantial improvement
in the growth of hybridomas was afforded by the inclusion of 18 trace elements complexed to disodium ethylene diaminetetraacetate
(EDTA). The medium was further improved by the inclusion of components not found in Ham's F12 medium or by raising the concentrations
of existing low molecular weight components. Murine hybridomas can be cultured routinely in this protein-free medium in an
anchorage-independent manner with doubling times generally under 24 h. Visualized on electrophoretic gels, levels of monoclonal
antibody taken from those cultures often exceeded 80% of the total protein. The medium was also able to support the growth
of HuT 78 and H9 cells as well as certain other transformed cells of the immune system. In addition, normal human peripheral
blood lymphocytes, activated with phytohemagglutinin and cultured with 50 U/ml recombinant interleukin 2, could be grown for
2 wk with a 50-fold expansion over input cell number. 相似文献
57.
Mitsuo Satoh Shinji Hosoi Seiji Sato 《In vitro cellular & developmental biology. Plant》1990,26(11):1101-1104
Summary The protease activity in serum-free conditioned medium of chinese hamster ovary (CHO) cells was measured using peptidyl (or
aminoacyl)-4-methylcoumaryl-7-amides (MCAs) as the substrates. Aminopeptidase increased in level as amounts of nonviable cells
increased during cultivation in serum-free medium, indicating that the activity seems to be originated from intracellular
proteases. The activity toward Boc-Leu-Arg-Arg-MCA, which was strongly inhibited by p-chloromercuribenzonate and N-ethylmaleimide,
was the strongest among those toward peptidyl-MCAs in the conditioned medium within 48 h-cultivation in serum-free medium.
In contrast to the case of aminopeptidase activity, the endopeptidase activity decreased in level after 48 h-cultivation although
amounts of nonviable cells increases. Thus, CHO cells continuously secrete the cysteine proteases.
This work was supported by the management of the Research Association for Biotechnology as a part of the R&D of Basic Technology
for Future Industries sponsored by NEDO (New Energy and Industrial Technology Development Organization). 相似文献
58.
Koichi Rikimaru Hitomi Toda Noriko Tachikawa Nobuyuki Kamata Shoji Enomoto 《In vitro cellular & developmental biology. Plant》1990,26(9):849-856
Summary A novel protein-free synthetic medium has been developed for the culture of human squamous cell carcinoma cells. This medium,
designated PF86-1, supports the serial subcultivation of six out of nine human squamous cell carcinoma cell lines in a protein-free,
chemically defined condition without the adapting culture from serum-containing conditions. These cell lines growing in PF86-1
exhibited nearly equal potency to grow in massive culture without noticeable changes in morphology but presented a significantly
decreased level of colony forming efficiency when compared with the cells cultured in serum-containing media, suggesting the
implication of some autocrine mechanism. Interestingly, this medium supported the growth of normal human squamous cells of
oral mucosa and skin for more than 2 mo. in the primary explant culture in spite of high levels of calcium ion concentration,
where the overgrowth of fibroblasts as contaminant was not observed. These results suggest that PF86-1 supports the growth
of cells derived from epidermal tissues selectively and provides the same defined condition for growth of malignant and nonmalignant
human squamous cells. It seems, therefore, that PF86-1 allows investigations on the products of squamous cell carcinoma cells
or on the differences of growth mechanisms between normal and neoplastic human squamous cells. 相似文献
59.
In vitro development of one-cell embryos from outbred mice: Influence of culture medium composition 总被引:2,自引:0,他引:2
Akiko Spindle 《In vitro cellular & developmental biology. Plant》1990,26(2):151-156
Summary One-cell embryos from outbred mice (CF1, CD-1, and Dub:ICR) were cultured in various modifications of egg culture medium (ECM).
The best development was observed in medium in which inorganic salts of modified T6 medium (mT6) replaced those of ECM. In
this modification (TE), 66% of one-cell CF1 embryos developed into blastocysts, comared to 46 and 43% for ECM and mT6, respectively.
Moreover, the cell numbers of blastocysts developing in TE (74.9±3.3) were higher than the cell numbers of those developing
in ECM (55.1±2.4). The culture requirements of embryos varied between different stocks of mice: Fewer CF1 embryos developed
to the blastocyst stage than either Dub:ICR embryos (90%) or CD-1 embryos (84%). Lowering the osmolarity of the medium from
300 to 280 mOsm, increasing the concentration of KC1 from 1.42 to 25 mM, or omitting lactate from the medium during Day 1 of culture did not further improve development of embryos, in contrast
to previous reports. However, the time at which embryos were transferred to outgrowth medium influenced their postblastocyst
development. The best development was observed when embryos were transferred on Day 4 of culture at the late morula-early
blastocyst stage.
This work was supported by the Office of Health and Environmental Research, U.S. Department of Energy, Washington, DC, contract
DE-AC03-76-SF01012. 相似文献
60.
JOÃO BATISTA TAVARES DA SILVA ISAAC ROITMAN 《The Journal of eukaryotic microbiology》1990,37(6):521-523
ABSTRACT. Three strains of Phytomonas serpens two from tomatoes, Lycopersicon esculentum one from the insect Phtia picta (Hemiptera, Coreidae), were cultivated in a chemically defined medium developed from a defined medium for cultivating insect flagellates. Besides organic growth factors required by other insect trypanosomatids this flagellate requires, serine and inositol. Glutamine stimulates growth, and, surprisingly, does not require heme. 相似文献