Species: human
Tissue: Choriocarcinoma
Features: Gonadotropin, lactogen and steroid secreting
Culture Medium: 12K modification of Ham's medium, 85%; FBS, 15%
Freeze Medium: Culture medium, 95%; dimethyl sulfoxide (DMSO), 5%
ATCC:
ATCC CCL-98, BeWo (Choriocarcinoma, human) Gonadotropin, lactogen
and steroid secreting. Current medium for
propagation: F12K modification of Ham's medium, 85%; FBS, 15%.
The BeWo trophoblastic cell line, the first human endocrine cell
type to be maintained in continuous cultivation, was initiated in
June, 1966 by R.A. Pattillo and G.O. Gey from a malignant
gestational choriocarcinoma of the fetal placenta (Cancer Res.
28: 1231, 1968; Science (Washington, DC) 159: 1467, 1968). The
tumor was originally transplanted to the hamster cheek pouch by
R. Hertz where it was maintained for 304 serial transfers over a
period of 8 years before establishment in vitro (Proc. Soc. Exp.
Biol. Med. 102: 77, 1959). The original BeWo culture was
established from a parabiotic combination of the choriocarcinoma
and human decidual tissue embedded on a collagen matrix. The
initial medium consisted of Waymouth's MB 752/1 diluted with
Gey's BSS, 70%; placental-cord serum, 30%. Homogeneous
populations of trophoblast cells were developed after a period of
several months by means of microsurgically excising and
discarding the fibroblastic and other non-tumorous areas. Gradual
adaptation to growth on glass and plastic flasks in Waymouth's MB
752/1, 50%; Gey's BSS, 40%; newborn calf serum, 10% was
accomplished over a 5-month period. The BeWo cell line has been
shown to secrete the spectrum of placental hormones
including human chorionic gonadotropin (hCG), polypeptide
hormones, human placental lactogen (hPL), estrogenic and
progestational steroids, estrone, estradiol, estriol, and
progesterone (In Vitro (Rockville) 6: 205, 1970). The cells
synthesize hCG up to a rate of 1000 IU/10(8) cells/24 hours
(Ann.N.Y. Acad. Sci. 172: 288, 1971). Cellular differentiation of
this stem cell type occurs when hormonal precursors are added to
the incubation medium: 35-40% conversion of estrogenic precursors
to estrone and estradiol was observed (J. Clin. Endocrinol.
Metab. 34: 59-61, 1972). The predominant glycolytic metabolism
exhibited by these cells and a high carbon dioxide production and
rapid drop in pH, necessitates daily fluid renewals or addition
of sufficient culture medium to last 36-48 hours to prevent
glucose exhaustion. In culture, approximately 10 million cells
subcultured one to three at weekly intervals utilize 1 mg of
glucose per hour from the culture medium. Variations in the level
of glucose contained in the culture medium exert profound changes
on cell growth and glycogen content. A glucose concentration of
approximately 300-340 mg percent has been found to be optimal for
the cells. The cell line was submitted to the American Type
Culture Collection in the 187th passage. DESCRIPTION OF
REPOSITORY REFERENCE SEED STOCK: Number of Serial Subcultures
from Tissue of Origin: 191. Freeze Medium:
Culture medium, 95%; dimethyl sulfoxide (DMSO), 5%;
antibiotic-free. Viability: Approximately 80%
(dye exclusion). Culture Medium: Waymouth's MB
752/1, 50%; Gey's BSS, 40%; newborn calf serum, 10%;
antibiotic-free. Growth Characteristics of Thawed Cells:
An inoculum of 6-7 X 10(5) viable cells in 4 ml of the above
culture medium per 25 cm2 flask yields a 3- to 5-fold increase
within 7 days at 37C, provided the medium is renewed daily and
the total volume of medium per flask is increased by 1 ml each
day. Plating Efficiency: Not measurable in the
above culture medium. Morphology: Epithelial-like. Karyology:
Chromosome Frequency Distribution 75 Cells: 2n = 46 Cells: 1 1 2
3 3 4 3 8 5 17 1 3 4 5 4 1 10 Chromosomes: 71-73-76-79-81 82 83
84 85 86 87 88 89 90-92 93 97-178 Stemline number is
hypotetraploid. Karyotype relatively stable within stemline
number. Sterility: Tests for mycoplasma,
bacteria, and fungi were negative. Species:
Confirmed as human by the cytotoxic-antibody dye-exclusion test. Virus
Susceptibility: Susceptible to poliovirus type 3, and
vesicular stomatitis (Indiana Strain) virus. Reverse
Transcriptase: Not detected. Hormone Production:
At passage 195, an average of 64 + or - 3 (S.E.) IU of hCG were
secreted per 10(6) cells per 24 hours. At passage 198, an average
of 35 + or - 3 (S.E.) nanograms of progesterone were secreted per
mg of cell protein per hour. Both hCG and progesterone were
measured by radioimmunoassay. Isoenzymes: G6PD
type B. Submitted by: R.A. Pattillo, Department of Gynecology and
Obstetrics, The Medical College of Wisconsin, Milwaukee, WI.
Prepared and characterized by: ATCC, Rockville, MD. Price Code:
J