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Leukemia cells from 32 instances of intense leukemia were refined
in vitro with the tumor advertiser 12-O-tetradecanoylphorbol-13-
acetic acid derivation (TPA) to examine their separate potential. Three cases
of intense undifferentiated leukemia (AUL) were contemplated seriously.
We found that refined of leukemic cells with TPA can incite
changes in cell surface antigens. Specifically, MCS-2, a "skillet"
granulocyte/monocyte marker, was inducible in vitro in AUL and
in intense myelogenous leukemia, while it was not inducible in
intense lymphoblastic leukemia. BA-2 (perceiving the M, 24,000
protein) and TA-1 (perceiving the M, 170,000 and M, 95,000
proteins) were likewise inducible in instances of AUL, intense myelocytic
leukemia, and intense monoblastic leukemia, despite the fact that these enemy of
gens are not constrained distinctly to leukemias of the myelomonocytic
heredity. Our examinations additionally demonstrate that undifferentiated cells could
be incited to vague esterase and once in a while to chloroac-
etate esterase reactivity while losing terminal deoxynucleotidyl
transferase. Morphological examinations in these cases uncovered cy-
tological development following TPA treatment. By and large,
these progressions were likewise somewhat inducible by refined cells in
medium alone or with the expansion of dimethyl sulfoxide however not
to the degree that was exhibited by TPA. Our examinations
demonstrated that MCS-2 is an awesome, explicit marker of intense
nonlymphocytic leukemia. A potential use for TPA to help in the
subclassification of patients with AUL is additionally recommended.
Presentation
TPA4 has been appeared to advance the separation of numerous
cell types in vitro (1). Its impacts have been exhibited on
human lymphocytes (14), hematopoietic cells (2, 38), leukemic
cells (5, 9,19, 41), and cell lines (11, 22, 27, 28, 35, 37, 42). The
promyelocytic cell line HL-60 has been utilized widely to
exhibit both utilitarian and morphological changes asso-
1Presented to some degree at the 67th Federation of American Societies for Experimental
Science meeting in Chicago (10). This work was bolstered by Grants CA-17034
what's more, CA-31685 from the National Cancer Institute and by the Coleman Leukemia
Research Fund.
2To whom demands for reprints ought to be tended to, at Department of Labo
ratory Medicine and Pathology, Box 609 Mayo, 420 Delaware Street S.E., University
of Minnesota, Minneapolis, MN 55455.
'Present location: Department of Immunology, Kurume University, School of
Drug, Kurume. Fukuoka 830, Japan. 4The shortenings utilized are: TPA, 12-O-tetradecanoylphorbol-13-acetic acid derivation; AUL,
intense undifferentiated leukemia; ALL, intense lymphocytic leukemia; AML, intense
myelocytic leukemia; AMOL, intense monoblastic leukemia; AMML, intense myelo
monocytic leukemia; AMEL, intense megakaryocytic leukemia; AEL, intense erythroid
leukemia; ANLL, intense nonlymphocytic leukemia; NSE, vague esterase; CAE,
chloroacetate esterase; TdT, terminal deoxynucleotidyl transferase; DMSO, di
methyl sulfoxide; FBS, fetal ox-like serum; PWM, pokeweed mitogen; PHA, phy-
tohemagglutinin; Con A. concanavalm A; Slg. surface immunoglobu
ciated with TPA acceptance. A portion of these modifications incorporate the
acceptance of NSE (35, 37), a marker of monocytic heredity (45),
joined by lost CAE (35), a marker of myelocytic
heredity, expanded phagocytosis (13, 35), and morphological
changes, for example, an indented core (36, 37), the entirety of which
show development towards a macrophage-like cell. Other cell
lines have likewise been utilized to show this differentiative
potential. Koeffler ef a/. (20) announced that myeloblastic-promye-
locytic cell lines (HL-60, ML-3, KG-1, and KG-1 clones 1, 2, and
3) could be incited to macrophage attributes, while early
myeloid impact cells (KG-1 an and K562) proved unable. Papayannopou-
lou ef a/. (31) found that TPA restrains globin combination in the
human erythroleukemia cell line while prompting morphological,
useful and biochemical changes trademark for large scale
phage-like cells. Pegoraro ef a/. (32) indicated that crisp cells from
AML and AMML uncovered changes after TPA treatment comparative
to those revealed for HL-60 and KG-1 lines. Likewise, Onta ef al.
(30) and Polliack ef al. (33) found that crisp leukemic cells from
patients with AML, however not ALL, have gotten follower after
momentary hatching with TPA. Since TPA has been found
to conquer the separation square average of leukemias, we
have utilized this compound to incite leukemic impacts to express
a myelomonocytic phenotype as characterized by monoclonal antibod
ies and cytochemical markers in 3 instances of AUL.
MATERIALS AND METHODS
Readiness and Culturing of Patient Cells. Mononuclear cells were
disengaged from the fringe blood or bone marrow of patients with
leukemia by Ficoll-Hypaque centrifugaron. Just examples containing at
least half leukemic cells before Ficoll-Hypaque enhancement were stud
ied. Cells were refined promptly or solidified in 10% DMSO (J. T. Bread cook Chemicals, Phillipsburg, NJ) and put away at - 170â°. At the point when required, solidified
cells were immediately defrosted in a 37â°water shower; gradually weakened with RPMI
1640 (Associated Biomedics Systems, Buffalo, NY) enhanced with
penicillin, streptomycin, and gentamicin (Grand Island Biological Co.,
Excellent Island, NY); and washed multiple times. To improve endurance and suitability, the cells were permitted to recover at 4â°for 3 to 4 hr in RPMI 1640
with 10% fetal ox-like serum (FBS; Grand Island Biological Co.) and were
at that point checked for practicality by trypan blue avoidance. All cells were refined
at a convergence of 106 cells/ml in RPMI 1640 with 10% FBS at 37â°in
5% CO2. In certain trials, 10% human AB-positive serum or 10%
autologous serum was fill in for FBS.
TPA, Mitogens, and Other Chemicals. TPA (CCR, Inc., Eden Prairie, MN) was broken down in DMSO, put away as a stock arrangement at 4â°for up to
multi week, and utilized at a grouping of 5 to 100 ng/ml in RPM11640.
Control societies containing the fitting weakenings (0.005 to 0.01%) of
DMSO were run in parallel with TPA-containing societies. PWM (Chemical
Elements, Inc., Lot 119051), PHA (Burroughs-Wellcome Co., Durham,
NC), and Con A (3 times solidified; Miles-Yeda, Ltd., Elkhart, IN) were
all utilized at 5 ¿ig/ml,and cholera poison (Sigma, St. Louis, MO; Lot 3129C-
0455) was utilized at 1 ng/ml. In certain investigations, DMSO was us