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1.
Summary During methotrexate-induced differentiation of cultured human choriocarcinoma (BeWo) cells, proliferation is inhibited, morphologic
and biochemical changes occur, and giant, often multinucleated, cells form. We have used the increase in cell volume as a
marker of the mature syncytiotrophoblastlike phenotype. Uninduced and differentiated BeWo cells are not spherical, and theoretical
considerations suggested that deviations in shape could result in significant errors in Coulter volume. To determine if the
values obtained by electrical pulse sizing reflected the actual mass of BeWo cells, we have evaluated the relationship between
Coulter volumes and intracellular water volumes obtained using a shape-independent estimate for eight cell types. A close
correlation (r
2=0.97) was found, indicating that cell volume changes in populations of irregularly shaped cells can be accurately measured
using a Coulter instrument.
Supported by an operating grant from the National Cancer Institute of Canada. N.S.B. was a recipient of a studentship award
from the Alberta Heritage Foundation for Medical Research. C.E.C. is a Senior Research Scientist of the National Cancer Institute
of Canada. The McEachern Laboratory is a research facility of the Faculty of Medicine, University of Alberta, and the Cross
Cancer Institute, Edmonton, Alberta. 相似文献
2.
Establishment and comparison of fibroblast cell lines from the medial collateral and anterior cruciate ligaments of the rabbit 总被引:3,自引:0,他引:3
Stephen M. Ross Rohit Joshi Cyril B. Frank 《In vitro cellular & developmental biology. Plant》1990,26(6):579-584
Summary We have developed a procedure to explant fibroblasts from the anterior cruciate ligament (ACL) and the medial collateral ligament
(MCL) of the rabbit knee, and have optimized conditions for maintaining them in culture. Maximal growth for both ACL and MCL
cells was obtained with Dulbecco's modified Eagle's medium supplemented with 15% fetal bovine serum and 250 μM ascorbate. ACL and MCL fibroblasts displayed intrinsic differences in their responses to changes in culture parameters. Specifically,
they displayed different growth responses when plated at different densities and responded to RPMI 1640 medium in very different
ways. There were also biochemical differences between the cell types. Both cell types produced similar amounts of collagen
in culture, but the ratio of type I to type III, the major collagen subtypes produced by these cells, were different. ACL
fibroblasts produced 86.7% type I and 13.3% type III, and MCL fibroblasts produced 71.1% type I and 28.9% type III. In addition,
total protein produced by ACL fibroblasts was higher than that produced by MCL cells. This confirms the suggestions of previous
researchers that such differences might exist.
This work was funded by a grant-in-aid from Medtronic of Canada, by an R&D Grant from the Alberta Ministry of Technology,
Research and Telecommunications, and by the Alberta Heritage Foundation for Medical Research. 相似文献
3.
Philip Skehan James E. Thomas Susan J. Friedman 《In vitro cellular & developmental biology. Plant》1986,22(11):632-636
Summary Sarcoma 180 monolayers spontaneously shed single cells and small multicellular aggregates into the surrounding medium to produce
a dual population of floating and substratum-attached cells. Shedding was a motility-associated event that occurred when cells
attempted to migrate over one another. It resulted from a combination of cell shape change and active motility, which increased
sensitivity to fluid shear dislodgement by reducing a cell's surface area of adhesive contact and increasing strain tension
at its adhesive contact points. Shedding occurred at all phases of the cell cycle. Extracellular matrix but not conditioned
medium enhanced the floating subpopulation by slowing the kinetics of rattachment to plastic and cellular substrata. Although
sarcoma 180 cells are anchorage independent in the sense that they grow readily in single cell suspension, they nevertheless
exhibited anchorage modulation of their cell cycle. Short periods in suspension produced a mild G1 accumulation, whereas longer periods of anchorage deprivation led to a mild G2 accumulation which appeared to result from an interference with cytokinesis.
This work was supported by grants from the Medical Research Council of Canada, The National Cancer Institute of Canada, the
Alberta Heritage Savings and Trust Fund for Applied Cancer Research, and the Alberta Heritage Fund for Medical Research. 相似文献
4.
Summary We have recently examined the electrophysiology and ultrastructure of approximately 100 tactile spines from the metathoracic legs of adult cockroaches. In only one animal the single sensory neuron that innervates the spine was replaced with a pair of apparently identical neurons which we believe were both functional. As far as we are aware this is the first reported study of unprovoked duplication in a peripherally-located insect sensory neuron.Supported by the Canadian Medical Research Council and the Alberta Heritage Foundation for Medical Research 相似文献
5.
Summary The localization of urotensin I has been investigated in the caudal neurosecretory system of the white sucker (Catostomus commersoni). The peptide is present in all the cells of the system both large and small, in the large axons passing to the urophysis, and in fine beaded fibres not only within the urophysis but also in a fine plexus lateral to the large cells in the spinal cord proper. The possibility that the caudal neurosecretory system is not a functionally uniform system but rather a collection of dissimilar cells of different synaptic inputs with a common entity, urotensin I, is discussed. Moreover, the feasibility of a urotensin I feedback loop is described.Financial support for this investigation was provided in part by MRC (Canada). K.L. is MRC career investigator; K.L.W, was in receipt of an Alberta Heritage Foundation for Medical Research Fellowship. It is a pleasure to record the valuable technical assistance of Mrs. W. Ho and the dedicated assistance in the collection of the experimental animals by Mrs. Helen Wilson of Nanton, Alberta. 相似文献
6.
E. J. Sanders 《In vitro cellular & developmental biology. Plant》1984,20(7):521-527
Summary The cell-substratum adhesive characteristics of cultured chick embryo primary mesoderm cells have been examined by inteference
reflection microscopy and transmission electron microscoy under various conditions. Correlations were drawn between the type
of adhesion and the degree of motility shown by the cells. During the rapid spreading and motility of cells cultured on fibronectin-containing
substrate, focal contacts (10 to 15-nm gap) were rare and close contacts (about 30-nm gap) were pedominant. By contrast, when
the cells were immobile, after 5 d in cultue, extensive focal contacts were present, together with stress fibers. The results
indicate that tight cell-substratum contact is incompatible with rapid cell motility and that fibronectin acts by inducing
the formation of close contacts rather than focal contacts.
This work was supported by grants from the Medical Research Council of Canada and the Alberta Heritage Foundation for Medical
Research. 相似文献
7.
H. Muzik M. E. Shea C. C. Lin H. Jamro S. Cassol L. M. Jerry L. Bryant 《In vitro cellular & developmental biology. Plant》1982,18(6):515-524
Summary Attempts were made to adapt human long-term B lymphoblastoid cell lines to prolonged growth in serum-free, chemically defined
media. A newly described medium, which is an enriched modification of Dulbecco’s modified Eagle’s medium containing additional
amino acids and vitamins, was used. The serum is totally replaced by albumin, transferrin, and soybean lipid. The cell lines
were all adaptable from RPMI 1640 over a period of time during which the 10% fetal bovine serum (FBS) concentration was reduced
and then eliminated in successive steps. After 3 to 6 wk minor alterations in cell shape and adhesion were noted without significant
histological changes. Growth characteristics were comparable in the new medium provided a double initial inoculum was used.
A panel of cell surface markers, including surface immunoglobulins, Ia antigens, Fc and complement receptors, and T and B
erythrocyte rosettes, all showed no altered expression. Molecular genotyping of Ia antigens was carried out by 2-D gel electrophoresis.
The antigens showed their full polymorphism without change and were shed into the new culture medium without alteration. Chromosome
analysis was performed on Q-banded karyotypes from one of the lines and showed no alteration resulting from the change to
serum-free conditions. Thus long-term B lymphoblastoid cell lines can be adapted to prolonged growth in serum-free medium.
This will facilitate the assay and isolation of cell products regulating lymphocyte function and the identification and characterization
of cell surface molecules free of interference from undefined serum components.
This work is supported by grants from the Medical Research Council of Canada, the National Cancer Institute of Cancer, and
the Alberta Heritage Fund. 相似文献
8.
Human chorion cells respond to growth factors but lose steroidogenic capacity in primary monolayer cell culture 总被引:1,自引:0,他引:1
K. A. Ferguson B. F. Mitchell A. K. Tanswell 《In vitro cellular & developmental biology. Plant》1986,22(6):320-324
Summary This study has defined a method for preparation and monolayer culture of cells fromchorion laeve. Cell number and cell protein content are table over 7 d in culture. The cells will divide in response to epidermal growth
factor in the presence of a supplemented, enriched medium and a collagen matrix, but they lose steroidogenic activity over
time in culture. This culture system can be used as the starting point for the development of a chemically defined hormone-supplemented,
serum-free culture system for studies of chorion cell differentiation and fetal membrane cell interactions.
This work was supported by the Medical Research Council of Canada through Grant MT-7867 (to A. K. T.), Grant MA-7731 (to B.
F. M.), and a Summer Studentship (to K. A. F.). 相似文献
9.
Enterochromaffin (EC) cells regulate gut motility and secretion in response to luminal stimuli, via the release of serotonin
(5-HT). Inflammatory bowel disease and other gastrointestinal disorders are associated with increased numbers of EC cells
and 5-HT availability. Our aim was to determine whether proliferation of EC cells contributed to the hyperplasia associated
with intestinal inflammation. Ileitis was induced in guinea-pigs by intraluminal injection of 2,4,6-trinitrobenzene sulphonic
acid (TNBS). A single pulse of 5-bromo-2′-deoxyuridine (BrdU) was injected 1 or 24 h before the collection of tissue, 6 or
7 days after TNBS treatment. In the controls, the labelling index (percentage of BrdU-labelled EC cells) was less than 1%.
Despite a significant increase in EC cells in the inflamed ileum, the labelling index was similar in the TNBS-treated animals
to that of controls. An increased occurrence of EC cells in the BrdU-labelled zone accounted for the increase in EC cells
in the inflamed ileum. Goblet cell numbers were also significantly increased in the inflamed ileum, indicating that cell hyperplasia
was not limited to the enteroendocrine cell lineage. This study demonstrates that a small portion of EC cells retain some
proliferative capacity; however, hyperplasia associated with ileitis is not attributable to the increased proliferation of
EC cells and is not limited to one cell lineage. Therefore, EC cell hyperplasia most probably occurs at the level of the stem
cell or recruitment from the progenitor pool.
This work was supported by an operating grant from the Crohn’s and Colitis Foundation of Canada (CCFC; to Keith Sharkey and
Dr. Gary Mawe, University of Vermont, USA). Keith Sharkey is an Alberta Heritage Foundation for Medical Research (AHFMR) Medical
Scientist and the CCFC Chair in Inflammatory Bowel Disease Research. Jennifer O’Hara is an AHFMR graduate student. 相似文献
10.
We have examined the distribution and extent of phosphorylation of the tight junction-associated protein ZO-1 in the epithelial MDCK cell line, and in three cell types that do not form tight junctions: S180 (sarcoma) cells, S180 cells transfected with E-cadherin (S180L), and primary cultures of astrocytes. In shortterm calcium chelation experiments on MDCK cells, removal of extracellular calcium caused cells to pull apart. However, ZO-1 remained concentrated at the plasma membrane and no change in ZO-1 phosphorylation was observed. Maintenance of MDCK cells in low calcium medium, conditions where no tight junctions are found, resulted in altered ZO-1 distribution and lower total phosphorylation of the protein. In S180 cells, ZO-1 was diffusely distributed along the entire cell surface, with concentration of the antigen in motile regions of the cell. Cell-cell contact was not a prerequisite for ZO-1 localization at the plasma membrane in this cell type, and the phosphate content of ZO-1 was found to be lower in S180 cells relative to MDCK cells. Expression of Ecadherin in S180L cells did not alter either the distribution or phosphorylation of ZO-1. In contrast to S180 cells, ZO-1 in primary cultures of astrocytes was concentrated at sites of cell-cell contact, and the phosphorylation state was the same as that in control MDCK cells. Comparison of one-dimensional proteolytic digests of 32P-labeled ZO-1 revealed the phosphorylation of two peptides in control MDCK cells that was absent in both MDCK cells grown in low calcium and in S180 cells.We would like to thank Cheryl Richards for her help with the cell culture and immunohistochemistry; David Begg, Gary Firestone, Vik Maraj, Manijeh Pasdar and Colin Rasmussen for helpful discussions; Jaclyn Peebles and Greg Morrison for help with graphics and photography; and Grace Martin and Bob Campenot for rat tail collagen. We are grateful to all the members of our laboratories for their friendship, advice and support. This work was supported by an Establishment Award to B.R.S. from the Alberta Heritage Foundation for Medical Research and grants to B.R.S. from the Kidney Foundation of Canada and the Medical Research Council of Canada. A.H. is funded by a Studentship from the AHFMR. K.L.S. was supported by a grant from the National Institutes of Health (DK-42799) to Gary L. Firestone. B.R.S. is a Medical Research Council of Canada and AHFMR Scholar. 相似文献
11.
David G. Chilton Betty H. Johnson Laurence Danel-Moore Simon Kawa E. Brad Thompson 《In vitro cellular & developmental biology. Plant》1990,26(6):561-570
Summary CEM-C7, a human leukemic CD4+ T-lymphocyte cell line and three of its subclones, CEM-4R4, CEM-3R43, and ICR-27, previously
cultured in a medium supplemented with 5 to 10% fetal bovine serum, have been adapted to serum-free media. The best medium
of those tested was RPMI 1640 supplemented with 5 μg/ml each transferrin and insulin + 5 ng/ml sodium selinite ± 0.1% bovine
serum albumin. While growing either with or without albumin, the several clonal lines of CEM cells displayed growth similar
to serum-supplemented cultures. Cell proliferation of CEM-C7 cells cultured in both serum-free media has been sustained for
3 mo, with culture doubling times of about 25 h for both serum-supplemented and serum-free cultures (viability ≥ 90%). Cell
morphology remained essentially the same in serum-free or serum containing media. The expression of CD4, a marker for T-derived
lymphoid cells, was not significantly different in serum-free medium. When grown in serum-free medium, CEM-C7 cells exhibited
increased steroid responsiveness as evidenced by increased glucocorticoid receptor binding sites, increased induction of glutamine
synthetase, and cell lysis at lower concentrations of steroid. Receptor mutant subclones of CEM-C7, which are proven to be
completely unresponsive to micromolar concentrations of dexamethasone when grown in serum-supplemented medium, become partially
sensitive to the hormone after growth in defined medium. The increased sensitivity of CEM-C7 cells and its subclones to dexamethasone
in serum-free medium returned to previous levels when these cells were recultured in serum-containing medium. Our results
suggest that substances in serum influence steroid effects on these cells and that the molecular details of glucocorticoid
hormone action may be pursued more precisely in a clearly defined culture medium.
This work was conducted in conjunction with the Walls Medical Research Foundation. 相似文献
12.
Model of diffusion of oxygen to spheroids grown in stationary medium —I. Complete spherical symmetry
The use of spheroids as a tumor model has become commonplace since it was discovered that many cell lines can form spheroids
when grown on a surface to which the cells cannot attach. This culture system complicates experiments which depend on oxygen
supply because the oxygen concentration in the vicinity of a stationary spheroid has not been well defined. We present in
this paper solutions to the oxygen diffusion equation for simple geometries: a spheroid in an infinite stationary medium and
in a finite spherical stationary medium. Comparison of these solutions provides an estimate of the oxygen supply to a spheroid
in a Petri dish. We show that typical spheroids can be expected to cause a substantial depletion of the oxygen in the nearby
medium. Any disturbance of the medium or the spheroids will temporarily increase the oxygen supply. We provide a method for
estimating the rate of return to equilibrium in the finite cases. These results indicate that the oxygen supply to stationary
spheroids can be altered temporarily by small movements or changes in temperature which cause convection currents, or permanently
by changes in the depth of the medium.
Research supported by the Alberta Heritage Savings and Trust Fund-Applied Cancer Research.
Research supported by the Natural Science and Engineering Research Council of Canada, Grant No. NSERC A 4823. 相似文献
13.
We have characterized biochemical effects of Idh
GB1
in Drosophila melanogaster. This is a null-activity allele for NADP+-dependent isocitrate dehydrogenase (NADP-IDH) isolated from a natural population. The homozygous mutant strain has 5% of the NADP-IDH specific activity found in controls and less than 24% of the immunologically cross-reacting material (CRM). This mutation maps to 27.2 on the third chromosome, to the right of h. The biochemical phenotype of this mutant strain includes a coordinate reduction in malic enzyme (ME) specific activity and CRM and an increase in specific activity for the pentose-phosphate shunt enzymes, 6-phosphogluconate dehydrogenase and glucose-6-phosphate dehydrogenase. The K
m
values for purified NADP-IDH are not different from those found for the purified control enzyme for NADP+ or isocitrate. It is suggested that this allele may represent a cis-acting control mutation for one of at least two loci involved in the production of NADP-IDH in D. melanogaster.Research supported by an Alberta Heritage Foundation for Medical Research Establishment Grant to MMB and a Natural Sciences and Engineering Research Council Operating Grant to JHW. 相似文献
14.
New mutant alleles of theadenosine2 locus (ade2; 2–17.7) have been isolated using the eye-color phenotype exhibited by the prototype auxotrophic alleleade2
1 as the screening criterion. The new mutants form a single complementation group, suggesting that they all exhibit purine auxotrophy and defective formylglycineamide ribotide amidotransferase enzyme, likeade2
1. Tests carried out on particular new alleles confirm these suggestions. The new mutants all exhibit more extreme physical defects than the prototype. They have wing abnormalities like mutants defective in pyrimidine biosynthesis and reduced bristles like those defective in protein synthesis; thus they exhibit the combined visible phenotype ofrudimentary wings,rosy eyes, andbobbed bristles. Cytogenetic analysis places the locus in the interband proximal to26B1-2.This work was supported by NSERC Operating Grant A3269 to D.N., an Alberta Heritage Foundation for Medical Research Postdoctoral Fellowship to S.Y.K.T., and National Institute on Aging Grant AG00029 to D.P. 相似文献
15.
Wouter H. Lamers Marian van Roon Piet G. Mooren André De Graaf Robert Charles 《In vitro cellular & developmental biology. Plant》1985,21(11):606-611
Summary A completely defined medium (EHM-1), which reflects the amino acid composition of fetal rat serum and contains albumin as
the sole proteinaceous compound, allows the accumulation of carbamoylphosphate synthetase and phosphoenolpyruvate carboxykinase
in the presence of dexamethasone, dibutyryl cyclic AMP, and triiodothyronine to approximately twice the level attained in
a standard culture medium (RPMI 1640) supplemented with 10% fetal bovine serum (and hormones). Using the EHM-1 medium we could
show that the capacity of hepatocytes to synthesize phosphoenolpyruvate carboxykinase in the presence of hormones is manifest
as soon as the cells differentiate from the embryonic foregut (embryonic Day 11). Furthermore we could show that embryonic
hepatocytes can become binuclear or polyploid when cultured in the presence of thyroid hormone.
These investigations were supported in part by the Dutch Foundation for Medical Research FUNGO (grant 13-50-38). 相似文献
16.
17.
S. Arch T. Smock R. Gurvis C. McCarthy 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》1978,128(1):67-70
Summary The atrial gland of the large hermaphroditic duct of the reproductive tract ofAplysia californica contains a substance that will cause egg laying when injected into sexually mature recipient animals. Preliminary chemical characterization of the atrial gland factor indicates that it is quite similar to the egg-laying hormone synthesized and secreted by the bag cell neuroendocrine organs of the central nervous system. In view of the apparent identity of its effects with those of the egg-laying hormone on presumed neural and reproductive system targets, it is possible that the atrial gland factor plays an hitherto unsuspected role in reproductive regulation. Moreover, the molecular similarity between the atrial gland factor and the egg-laying hormone suggests the possibility that the same humorally active species may be elaborated by two distinct tissue types.This work was supported by grants from the Medical Research Foundation of Oregon and USPHS (NS 11149). C. McC. was recipient of a Zlinkoff Research Fellowship. Dr. R. Beeman kindly confirmed our identification of the location and general morphology of the atrial gland. 相似文献
18.
Di Lorenzo Teresa P. De Maro Joseph A. Pumo Dorothy E. 《In vitro cellular & developmental biology. Plant》1989,25(10):909-913
Summary A serum-free culture system was used to compare the nutritional requirements of mouse mammary cells transformed by bovine
papillomavirus type 1 (ID13 cells) and the uninfected parent line (C127 cells). The serum-free, chemically defined medium
used for this study was an MCDB 151-based medium (MCDB 151+S+I), supplemented with epidermal growth factor, transferrin, hydrocortisone,
ethanolamine, phosphoethanolamine, retinoic acid, trace metals, and insulin. Proliferation of either cell type in serum-free
culture required the addition of 250 μg/ml of insulin. ID13 cells have a doubling time of greater than 96 h in MCDB 151+S+I,
whereas C127 cells have a doubling time of 60 h. This is in sharp contrast to the growth characteristics of the two cell types
in 10% fetal bovine serum, where doubling times for the ID13 and C127 cells are 24 and 30 h, respectively. Culture of the
cells in a serum-free medium has therefore revealed that the papillomavirus-transformed cells have more stringent growth requirements
than the uninfected parent line.
This work was supported in part by grant #1-P01 NS19214 from the National Institutes of Health, Bethesda, MD, NSF grant #R11-8217798
from the National Science Foundation, Washington, DC, and by a grant from the Otolaryngology Foundation. 相似文献
19.
James T. Cooper Samuel Goldstein 《In vitro cellular & developmental biology. Plant》1977,13(8):473-476
Summary Three strains of human skin fibroblasts were cultivated in nutrient medium supplemented either with human serum or fetal bovine
serum, and growth and lipid synthesis were compared. Rates of cellular growth were similar in both kinds of medium, but the
replicative life spans of all three strains were curtailed significantly in human-serum medium. Incorporation of label into
the major classes of neutral lipids from [14C]acetate and3H2O was increased also in human-serum medium. Since human serum contained higher concentrations of cholesterol known to reduce
endogenous cholesterol synthesis, these results were unexpected. Nonlipid factors in human serum may account for the shortened
cellular life spans and increased lipogenesis and perhaps for the potential to develop atherosclerosis.
Supported by grants from the Ontario Heart Foundation and Medical Research Council of Canada during the tenure of a Senior
Research Fellowship from the Ontario Heart Foundation (J.T.C.) and a Scholarship from the Medical Research Council of Canada
(S.G.). 相似文献
20.
Lucian J. Cuprak Carol J. Lammi Janet I. Crane 《In vitro cellular & developmental biology. Plant》1979,15(11):900-909
Summary An improved basal medium is presented that requires only minimal supplementation with dialyzed fetal bovine serum or bovine
serum albumin and fetuin to be comparable to Ham's F-10, which requires 15% horse serum (HS) and 2.5% fetal bovine serum (FBS)
for the growth and function of Y-1, mouse adrenal cortex tumor, cells. Cell monolayers maintained for up to 2 weeks without
any protein supplementation have retained their steroid response to ACTH. The medium differs from Ham's F-10 in its buffer
composition and higher calcium-ion concentration. This medium should be a useful adjunct to studies pertaining to steroid
and lipid intermediary metabolism, the retention of a specialized physiological function in a chemically defined medium, and
the mechanism of hormonal response.
Supported by the Medical Research Service of the Veterans Administration. 相似文献