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1.
Inhibition of p38 MAPK facilitates ex vivo expansion of skin epithelial progenitor cells 总被引:1,自引:0,他引:1
Juan Peng Wei Li Haibo Li Yanni Jia Zuguo Liu 《In vitro cellular & developmental biology. Animal》2009,45(9):558-565
Ex vivo expansion of skin epithelial stem cells has long attracted great interest because of the potential utilization in
transplantation and gene therapy. The use of cultured stem or progenitor cells was limited by the lack of applicable culturing
system with both satisfactory expansion efficacy and well suppressed differentiation ex vivo. The p38 mitogen-activated protein
kinase (MAPK) pathways are responsible for cell growth and differentiation process. We investigated the function of p38 inhibitor
SB203580 in the ex vivo expansion of skin epithelial progenitor cells by comparing media with or without addition of this
inhibitor. Our results showed that the culturing medium with murine 3T3 feeder layers added with 10 μM SB203580 was more effective
in promoting clonal growth of human skin epithelial progenitors or stem cells than the conventional medium without SB203580.
The clone initial day in cells treated with 10 μM SB203580 came 2 d earlier with higher colony formation efficiency. The skin
epithelial progenitor cells treated with 10 μM SB203580 formed clones that were uniformly smaller in size, longer in sustained
proliferation, shorter in clone doubling time, higher in S-phase cells percentage, and lower in levels of differentiation
markers such as K10 along with higher levels of stem-cell-associated markers such as p63, K15, and ABCG2 than those cultured
in the conventional medium. Collectively, these results indicate that the p38 MAPK pathways inhibitor SB203580 can be used
as a culture medium additive that helps to achieve more effective ex vivo expansion of skin epithelial progenitor cells. 相似文献
2.
Linda J. Loretz Catherine A. Reznikoff 《In vitro cellular & developmental biology. Plant》1988,24(4):333-342
Summary We report the development of culture conditions which routinely support clonal growth of normal human uroepithelial cells
(HUC). Secondary cultures seeded at clonal densities and grown under conditions described herein have a colony-forming efficiency
(CFE) and colony size that will be useful for in vitro experiments. Primary cultures were dispersed to single cells and seeded
in a supplemented Ham's F12 medium containing 1% fetal bovine serum together with 3×105 lethally irradiated Swiss 3T3 feeder cells on plastic substrates preequilibrated with F12 medium containing 5 or 10% serum.
Using these conditions, the average CFE was 16.1±2.5%. A cloning efficiency of 4.9±1.5% was obtained under the same conditions
in serum-free F12+ when supplemented with a mixture of trace elements or 0.1 mM ethanolamine. The epithelial nature of the cloned cells was confirmed by morphology and by positive immunofluorescent staining
for human epithelial keratin proteins. To make this system useful for mutagenesis experiments, a clone of Swiss 3T3 feeder
cells resistant to 5 μg/ml 6-thioguanine (6TG) was derived from the parental cell line. This 6-TG-resistant Swiss 3T3 clone
supports HUC clonal growth with a CFE of 17.9±2.0% CFE. We also report clonal growth of HUC without feeder cells using supplemented
MCDB 170 medium containing 70 μg/ml bovine pituitary extract. The average cloning efficiency using these conditions was 5.7±1.7%.
This work was supported by NIH grant 29525 to C. A. R. L. J. L. is a recipient of National Science Foundation predoctoral
fellowship. 相似文献
3.
Terramani TT Eton D Bui PA Wang Y Weaver FA Yu H 《In vitro cellular & developmental biology. Animal》2000,36(2):125-132
Summary The purpose of this study is to identify optimal culture conditions to support the proliferation of human macrovascular endothelial
cells. Two cell lines were employed: human saphenous vein endothelial cells (HSVEC) and human umbilical vein endothelial cells
(HUVEC). The influence of basal nutrient media (14 types), fetal bovine serum (FBS), and mitogens (three types) were investigated
in relation to cell proliferation. Additionally, a variety of extracellular matrix (ECM) substrate-coated culture dishes were
also tested. The most effective nutrient medium in augmenting cell proliferation was MCDB 131. Compared to the more commonly
used M199 medium, MCDB 131 resulted in a 2.3-fold increase in cell proliferation. Media containing 20% FBS increased cell
proliferation 7.5-fold compared to serum-free media. Among the mitogens tested, heparin (50 μg/ml) and endothelial cell growth
supplement (ECGS) (50μg/ml) significantly improved cell proliferation. Epithelial growth factor (EGF) provided no improvement
in cell proliferation. There were no statistical differences in cell proliferation or morphology when endothelial cells were
grown on uncoated culture plates compared to plates coated with ECM proteins: fibronectin, laminin, gelatin, or collagen types
I and IV. The culture environment yielding maximal HSVEC and HUVEC proliferation is MCDB 131 nutrient medium supplemented
with 2 mM glutamine, 20% FBS, 50 μg/ml heparin, and 50 μg/ml ECGS. The ECM substrate-coated culture dishes offer no advantage. 相似文献
4.
Effects of steroid hormones in fetal bovine serum on plating and cloning of human cells in vitro 总被引:1,自引:0,他引:1
George E. Milo William B. Malarkey John E. Powell James R. Blakeslee David S. Yohn 《In vitro cellular & developmental biology. Plant》1976,12(1):23-30
Summary Fetal bovine sera from each of three different commercial sources were tested for their ability to support cloning of human
fibroblastoid cells in vitro. Cloning efficiencies varied according to serum source. Serum (10 samples) from company A did
not support growth, while sera (10 samples) from companies B and C provided adequate to excellent conditions for cloning and
growth. Cells from neonatal foreskin or embryonic lung responded to each serum similarly. Bovine serum albumin type H7 from
company C supported cell growth in media without serum.
Sera containing 1.0 ng per ml or more of progesterone inhibited growth, whereas sera containing less than 1.0 ng per ml supported
cloning and growth. In the low progesterone sera, the concentration of 17-β-estradiol exceeded 100 pg per ml. Growth supporting
sera could be made non-supportive by adding 0.1 μg per ml of progesterone. The addition to non-supportive sera of 0.1 μg per
ml of 17-β-estradiol or hydrocortisone made these sera supportive of cell growth.
Addition of estrogen or hydrocortisone to a culture medium that inhibits growth, with subsequent reversal of the inhibitory
effect, implies that these hormones competitively regulate growth of responsive cells in vitro.
Supported in part by NIH-NCI-EC2074. 相似文献
5.
Tran Cong Toai Huynh Duy Thao Ciro Gargiulo Nguyen Phuong Thao Tran Thi Thanh Thuy Huynh Minh Tuan Nguyen Thanh Tung Luis Filgueira D. Micheal Strong 《Cell and tissue banking》2011,12(2):125-133
There have been many attempts to acquire and culture human keratinocytes for clinical purposes including from keratotome slices
in media with fetal calf serum (FCS) or pituitary extract (PE), from skin specimens in media with feeder layers, from suction
blister epidermal roofs’ in serum-free culture and from human umbilical cord blood (hUCB) mesenchymal stem cells (MSCs) in
media with skin feeder layers. Conversely this study was designed to investigate whether keratinocytes could be obtained directly
from hUCB MSCs in vitro. It is widely established that mesenchymal stem cells from human umbilical cord blood have multipotent
capacity and the ability to differentiate into disparate cell lineages hUCB MSCs were directly induced to differentiate into
keratinocytes by using a specific medium composed of primary culture medium (PCM) and serum free medium (SFM) in a ratio 1:9
for a period of 7 days and tested by immunostain p63 and K1-K10. Cells thus cultured were positive in both tests, confirming
the possibility to directly obtain keratinocytes from MSCs hUCB in vitro. 相似文献
6.
Summary L cells were grown in spinner cultures in a defined medium consisting of Waymouth medium MB752/1 (19) supplemented with 2
mg of fatty acid-free bovine serum albumin (BSA) per ml and 5 μg of oleate per ml (WO5 medium). Growth in WO5 medium was comparable to spinner L cell growth in two serum-containing media. The optimal concentration of oleate in the
WO medium was 5 to 10 μg per ml. The use of 20 to 80 μg of oleate per ml of medium resulted in lower peak populations and
earlier declines in viable cell counts. Cell death occurred rapidly in WO160 medium. Cell growth in WO medium containing 5 to 80 μg of oleate per ml was well above the level of growth observed when
no oleate was present in the medium. Since the total lipid and fatty acid compositions of the BSA used in this study have
been characterized by the authors, the WO medium may be considered a defined medium. L cells have been continuously maintained
in spinner cultures in WO5 medium for over 50 passages with no major variation in the growth pattern. A 1000-fold increase inChlamydia psittaci strain meningopneumonitis, with a peak titer of 9.3×107 plaque-forming units per ml, was observed when the chlamydial agents were grown in spinner L cells in WO5 medium.
This investigation was supported by Public Health Service Research Grant HE 08214 from the Program Projects Branch, Extramural
Programs, National Heart and Lung Institute; The World Health Organization; and The Hormel Foundation. 相似文献
7.
Growth and differentiation of human keratinocytes without a feeder layer or conditioned medium 总被引:1,自引:0,他引:1
Summary An improved procedure has been developed for clonal growth of normal human epidermal keratinocytes (HK) without feeder cells
or conditioned medium. The use of medium 199, supplemented with 0.4 μg/ml hydrocortisone (HC) and 20% (v/v) whole fetal bovine
serum (wFBS) and conditioned overnight by 3T3 cells, eliminated the need for a feeder layer of lethally irradiated 3T3 cells
for HK growth. Several other media with equivalent conditioning and supplementation failed to support satisfactory multiplication
of HK, including Dulbecco's modified Eagle's medium, which is normally used for growth of HK with a feeder layer. Increasing
the concentration of HC to 10 μg/ml (2.8×10−5
M) made possible clonal growth of HK without any conditioning of the medium. The addition of 10−5
M putrescine, 10−5
M vitamin B12, or 3.7×10−6
M β-estradiol further enhanced growth in unconditioned medium. Substantially greater improvement was obtained by the addition
of pituitary extract or fractions prepared from pituitary extract. In medium 199 supplemented with 10 μg/ml HC, 20% (v/v)
wFBS, and 0.15 mg/ml each of two pituitary fractions, single HK attach with a colony-forming efficiency equal to that in conditioned
medium and form stratified, keratinized colonies that grow to confluency and can be subcultured. These results make it clear
that HK do not require special “conditioning factors” from fibroblasts for clonal growth and differentiation in culture. Thus,
factors directly involved in growth and the expression of differentiation can be analyzed without the interfering effects
of any other type of cell. Preliminary studies with epidermal growth factor (EGF), which stimulates growth and extends life
span of HK grown in the presence of fibroblasts, have shown that, in the absence of fibroblasts, EGF has no effect either
on clonal growth or on cumulative multiplication potential of HK.
This paper contains material from a thesis submitted to the Graduate School of the University of Colorado, Boulder, by Donna
M. Peehl in partial fulfillment of the requirements for the Ph.D. degree.
This work was supported by Grant CA 15305 from the National Cancer Institute and Grant AG 00310 from the National Institute
on Aging. 相似文献
8.
F. L. Vaughan L. L. Kass J. A. Uzman 《In vitro cellular & developmental biology. Plant》1981,17(11):941-946
Summary A procedure for the preparation and cultivation of rat epidermal basal cells from full thickness skin resulted in greater
than 99% viability and 90% plating efficiency. However, attempts to subculture monolayers of these epithelial cells grown
in medium with serum as the only supplement were totally unsuccessful. When hydrocortisone and insulin were added to the medium,
subcultivation of primary growth was obtained. It was demonstrated that hydrocortisone at concentrations as low as 0.1 μg/ml
was necessary for at least the initial attachment of the cells to the substrate—an essential step in subcultivation. Increasing
concentrations of insulin (0.1 to 50 μg/ml) caused the rate of proliferation and the cell density to increase, but insulin
alone did not support subcultivation.
This work was supported by Grants 1-R01-CA-19988 and 5-R01-AM-15206 from the National Institutes of Health, U.S. Public Health
Service. 相似文献
9.
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
10.
Summary Primary and passaged cultures of normal colon epithelial cells, derived from human fetuses (13 to 17 wk of conceptual age)
have been established. These cultures have been passaged 16 times thus far. The cultures have been initiated and maintained
in medium consisting of 50% Dulbecco's minimum essential medium and 50% Ham's F12 medium and supplemented with antibiotics
(penicillin, 100 U/ml; streptomycin, 100 μg/ml); ascorbic acid, 40 μg/ml;l-isoleucine, 50 μg/ml; epidermal growth factor, 20 ng/ml; insulin, 5 μg/ml; cholera toxin, 5 ng/ml; transferrin, 1 μg/ml;
fetal bovine serum (10%); and HEPES, 25 mM final concentration, and incubated at 37°C in humidified gas containing 5% CO2: 95% air.
The cellular and subcellular characteristics of primary and passaged cultures were defined using light microscopy and scanning
and transmission electron microscopy. The cells exhibited microvilli on cell surfaces and showed junctional complexes and
interdigitations between cells. Indented nuclei with dense chromatin and marginated heterochromatin, numerous mitochondria,
rough endoplasmic reticulum, polysomes, and extensive Golgi zones were conspicuous. Also, periodic acid Schiff's reagent-positive
staining of the cells suggests the active synthesis of complex mucopolysaccharides in the cytoplasm.
This study was supported by USPHS Grant CA-30185 from the National Large Bowel Cancer Project, National Cancer Institute. 相似文献
11.
Summary Rapid proliferation of mammary epithelial cells derived from biopsy specimens of human fibroadenomas was observed when medium
was supplemented with ten percent fetal bovine serum and hydrocortisone (5 μg per ml−1). Hydrocortisone in combination with FBS also led to a 2.5-fold increase in cell cluster attachment and subsequent colony
formation. A similar effect was not observed with human serum. In contrast to fibroblast cell systems, insulin did not significantly
alter cell growth. The results show that a mitogenic response to glucocorticoids by mammary epithelium may depend on the presence
of factors in sera.
Supported in part by NCI Contract CB-33898. 相似文献
12.
Gary D. Stoner Curtis C. Harris David G. Bostwick Raymond T. Jones Benjamin F. Trump Elizabeth W. Kingsbury Elliott Fineman Carnell Newkirk 《In vitro cellular & developmental biology. Plant》1978,14(7):581-590
Summary Epithelial cells derived from bovine pancreatic duct have been grown continuously in culture for 30 weeks (approximately 90
doublings of the cell population). The cells were grown in Eagle's minimal essential medium supplemented with 10% heat-inactivated
fetal bovine serum, 2 mM glutamine, 0.1 mM nonessential amino acids, and antibiotics. In confluent cultures, the cells are
multilayered and form circular structures. When tested at various passages, the cells neither formed colonies in soft agar
nor produced tumors after inoculation into athymic, nude mice. Hydrocortisone (1 and 5 μg per ml) and insulin (1,5 and 10
μg per ml) had no effect on the growth of the cells. β-Retinyl acetate inhibited growth rate and cell yield at a concentration
of 5 μg per ml but was not growth-inhibitory at lower concentrations. By electron microscopy the cells have numerous mitochondria,
Golgi and microvilli. Mucous droplets were observed in a small proportion of the cells. Desmosome-like structures and occluding
junctions were observed more frequently between cells that had been transferred as aggregates than between cells transferred
as single cells. Cytochemical studies indicated that some cells produce PAS positive granules that were not removed after
treatment of the cultures with diastase. Eleven cell clones were isolated from the mass culture. The growth rates of the clones
are different as well as the period of time in which the clones can be propagated in vitro.
This work was supported in part by Y01 CP 60204 and N01 CP 43237. 相似文献
13.
Karimullah A. Zirvi Darwin O. Chee George J. Hill 《In vitro cellular & developmental biology. Plant》1986,22(7):369-374
Summary Five human tumor cell lines were studied for growth factor requirements and for replication in serum-free media. Of the five
tumor lines HT-29 (colon carcinoma), TWI (melanoma), A-549 (lung carcinoma), Panc-1, (carcinoma of the pancreas) and EJ, (bladder
carcinoma) only HT-29 and TWI grew in the serum-free medium (SFM). In a series of additional experiments, a combination of
transferrin (5 μg/ml), insulin (5 μg/ml), triiodothyronine (2×10−10
M), epidermal growth factor (20 ng/ml), and selenium (5 ng/ml) was added to Chee’s essential medium (CEM) without serum (C-TITES
medium). The C-TITES modification of CEM was found to allow optimal replication of HT-29 and TWI cells. Both HT-29 and TWI
cells have replicated continuously in C-TITES medium for periods of more than 15 mo. These cells replicate with slightly lower
doubling times than in CEM supplemented with 10% fetal bovine serum. Deletion of insulin or transferrin from the C-TITES medium
resulted in cessation of cell growth of HT-29 and TWI. HT-29 assumed a somewhat rounded morphology, whereas TWI grew with
the characteristic fibroblastic morphology in C-TITES medium. Cell line EJ did not grow in C-TITES medium. The other two cell
lines, A-549 and Panc-1, grew in C-TITES medium but their growth rate was much slower than that in SSM. Availability of cell
lines that can be propagated in serum-free, hormone-supplemented medium may aid in the study of the mechanisms by which hormones
influence cell growth.
This work was supported by Veterans Administration Research Awards to two of the authors (Karimullah A. Zirvi and George J.
Hill) and grant no. CA-37138 from the National Cancer Institute. 相似文献
14.
Summary Serial passage cultures of colonic epithelial cells from young rats have been maintained for more than 6 months in Eagle's
minimum essential medium buffered with HEPES (25 mM) and supplemented with 2.5% fetal bovine serum, 0.5 μg/ml insulin, 5.0 μg/ml transferrin, and antibiotics. The cells proliferated
in this medium with a population doubling time of approximately 53 h. The cells retained differentiated morphology as evidenced
by secretory activity and the presence of secretory granules, microvilli, tonofilaments, and desmosomal junctions. Further
cells at the fourth passage had normal karyotypes with 42 chromosomes and exhibited anchorage dependent growth. High concentrations
of fetal bovine serum (10 to 15%) exerted toxic effects on the colonic epithelial cell cultures.
Supported by National Cancer Institute Contract N01-CP-75914. 相似文献
15.
Robert W. Pumper Peter Fagan Dr. William G. Taylor 《In vitro cellular & developmental biology. Plant》1971,6(4):266-268
Summary Two mammalian cell lines which multiply in vitro in culture medium devoid of serum were investigated for sensitivity to twice
crystallized trypsin. The minimum trypsin concentration which showed a concomitant increase in cell numbers and detachment
from the surface of the culture vessel in one cell line was 0.01 μg per ml or approximately 10−8 μg per cell. These effects could be neutralized by normal human or calf serum at a dilution of 1∶500. The second cell line,
which normally grows in suspension, was unaffected by these concentrations of trypsin. 相似文献
16.
Yasuhiro Tomooka Stephen E. Harris John A. McLachlan 《In vitro cellular & developmental biology. Plant》1985,21(4):237-244
Summary Epithelial cells from mouse seminal vesicles were enzymatically dissociated enriched by gradient centrifugation, and maintained
in collagen gel cultures with defined (serum-free) media. The epithelial origin of the cells was determined morhologically,
immunocytochemically, and biochemically. Cells formed three-dimensional colonies with a lumen in collagen gels. Cell number
was increased eight-fold within a 8 to 12-d culture period in a medium supplemented with epidermal growth factor (EGF) (10
ng/ml), insulin (10 μg/ml), transferrin (10 μg/ml), cholera toxin (10 ng/ml), and hydrocortisone (0.1 μg/ml). The cells required
eGF and insulin; the growth-promoting effects of these two peptide hormones were optimized by transferrin, cholera toxin,
and hydrocortisone. Fetal bovine serum did not support growth; rather, it suppressed the stimulated growth observed in serum-free
media. A time-course study revealed that a lag period preceded rapi growth. The collagen gel, serum-free culture provides
a powerful tool to study the effects of hormones on proliferation and differentiation of androgen sensitive cells. 相似文献
17.
Susumu Yoshida Shigeo Kasuga Yuzo Hirao Tohru Fuwa Shizutoshi Nakagawa 《In vitro cellular & developmental biology. Plant》1987,23(7):460-464
Summary Biosynthetic human epidermal growth factor (Bh-EGF) induced dose-dependent synthesis and secretion of neutral mucin glycoprotein
when the fundal cells isolated from rabbit stomach were cultured in serum-free medium containing Bh-EGF at concentrations
as high as 10 to 100 ng/ml. At these high concentrations, Bh-EGF had no effect on the cell growth. In marked contrast, much
lower concentrations from 0.1 to 1.0 ng/ml of Bh-EGF failed to stimulate mucin synthesis, but enhanced proliferation of the
cells. Electrophoretic pattern of the mucin secreted from the cultured mucosal cells was very similar to that of the authentic
mucin obtained from rabbit stomach. Maximal secretion of the mucin from the cells was observed at Hour 96 of the culture.
Although fetal bovine serum (5%) and insulin (0.5 μg/ml) also stimulated the mucosal cells, both in growth and in mucin synthesis
and release, the enhancing activity of the mucin synthesized and released by Bh-EGF at a concentration of 100 ng/ml per microgram
DNA of cultured cells was far superior to that of 5% fetal bovine serum and 0.5 μg/ml insulin. 相似文献
18.
Summary Fetal bovine serum has been reported to delay or inhibit “spontaneous” neoplastic transformation in vitro as compared with
all other sera tested. The present results indicate that fetal bovine serum is also unique in containing high levels of protein-glutathione
mixed disulfides (3 to 7 μg glutathione as mixed disulfide per ml serum). The level of mixed disulfide appears to vary in
accordance with the period of gestation of the fetal calves used to prepare the serum, decreasing below detectable levels
(less than 0.2 μg per ml) with nearterm fetal calves. Calf, adult bovine, fetal horse, and swine sera did not contain detectable
levels of this type of mixed disulfide.
This work was supported by a grant from the National Institutes of Health (CA 08348). 相似文献
19.
Pamela W. Roy Gail E. Ryan Edwin D. Bransome 《In vitro cellular & developmental biology. Plant》1976,12(2):115-119
Summary A simple method is described for primary culture and for maintenance of hormone-producing cells from normal human placenta.
A consistent yield of cells was obtained and an average survival of 3 to 4 months in culture using 1 mm3 explants from the most vascular area of the placentas. These explants were placed in a variety of culture media in 30 ml
flasks and incubated at 37°C in an atmosphere of 5% CO2 and 95% air. The best yields in terms of cell growth were observed with Eagle’s MEM (minimum essential medium) with supplements
of horse serum and fetal calf serum or human cord serum. (Ham’s F-10 with supplement of horse serum and fetal calf serum supports
growth for the longest period and media containing human cord serum had the best yield of steroids.)
Presented in part at the 25th meeting of the Tissue Culture Association, June 5, 1974, Miami Beach, Florida. Supported by
a contract from the Agency for International Development (2491/csd) and a grant from the National Institutes of Health (CA-12455). 相似文献
20.
Calvin D. Roskelley Nelly Auersperg 《In vitro cellular & developmental biology. Plant》1990,26(5):493-501
Summary We have developed a method that separates rat adrenocortical cells by density into populations which retain zone specific
properties in primary culture. Two different parenchymal populations were obtained and designated 2FASC (1.034 g/ml, 18.0
μm cell diameter) and 7GLOM (1.069 g/ml, 11.7 μm cell diameter). In freshly isolated cell suspensions the physical characteristics
and differential steroidogenic responses to adrenocorticotropin and angiotensin II suggested that 2FASC cells originated predominantly
from the zona fasciculata and 7GLOM cells from the zona glomerulosa. In primary culture (Dulbecco's Modified Eagle's Medium-F12
medium with 15% horse serum and 2.5% fetal bovine serum) the two populations exhibited different morphologies. 2FASC cells
retained lipid and formed cohesive epithelial monolayers that remained stationary for 3 wk. 7GLOM cells were initially epithelial
but rapidly lost lipid, spread, and assumed fibroblastic shapes. Both cell types were ositive for the cholesterol side-chain
cleavage cytochrome P-450 by immunofluorescence. Therefore, the morphologic changes seen in 7GLOM cultures were due to modulation,
not fibroblastic overgrowth. This phenotypic plasticity may reflect the mesodermal origin of the adrenal cortex, and the subcapsular
location of 7GLOM cells in vivo. In contrast, cells such as 2FASC which are located deeper in the cortex seem to have a more
restricted, fully committed parenchymal phenotype.
This work was supported by a studentship to C. D. R., and by a grant and research associateship to N. A., from the National
Cancer Institute of Canada. 相似文献