MEDICINAL PLANTS
THE IMMUNOMODULANT ACTION OF LECTINS ISOLATED FROM
CULTIVATED PEAVINE (Lathyrus sativus L.)
M. V. Kiselevskii
1
and S. G. Zaichikova
2
Translated from Khimiko-Farmatsevticheskii Zhurnal, Vol. 36, No. 6, pp. 30 – 31, June, 2002.
Original article submitted June 4, 2001.
As is known, lectins extracted from seeds of various
plants from Fabaceae family exhibit high biological activity
[1]. The most interesting lectins of plant origin are the kidney
bean phytohemagglutinins (PHAs), which can serve as
polyclonal activators of immunity effectors [2]. It was re
-
ported that lectins were also found in the seeds of cultivated
peavine (Lathyrus sativus L.) [3, 4]. Previously, we have iso-
lated and separated lectins from this material and determined
their hemagglutinating activity [5].
The aim of this study was to determine the ability of
lectins isolated from cultivated peavine to activate the killer
activity of mononuclears in donor blood and to induce the
production of inflammation mediators, including
interleukin-1b (IL-1b), interleukin-2 (IL-2), interleukin-6
(IL-6), interleukin-8 (IL-8), interferon-a (INF-a), and tumor
necrosis factor (TNF).
MATERIALS AND METHODS
The level of cytokines was determined in a supernatant
fraction of lymphocytes obtained from healthy donors. The
samples were treated with various lectin fractions (L
1
–L
4
)
isolated from peavine seeds. The reference preparation was
PHA from kidney bean taken, as well as lectins, in an opti
-
mum concentration (5 mg/ml). Lymphocytes were isolated
from a peripheral blood obtained from a hemotransfusion de
-
partment of the Blokhin Oncological Research Center (Mos
-
cow). The blood was diluted by equal amounts of a nutrient
medium (RPMI-1640), layered over ficoll – verografin solu
-
tion with a density of 1.077 g/cm
3
, and centrifuged for
35 min at 1500 rpm. The isolated lymphocytes were doubly
washed with the same medium (RPMI-1640) and resuspend
-
ed at a cell concentration of 4 ´ 10
6
ml
–1
in a complete me
-
dium (RPMI-1640 with 5% fetal serum). Lectins L
1
–L
4
were added to aliquots of the lymphocyte suspension placed
into wells of a Costar multiwell plate and the samples were
kept for 48 h in a CO
2
incubator (37°C, 4% CO
2
), after
which the lymphocyte supernatant fractions were separated
and used in the test.
The cytokine concentrations were determined by ELISA
using a commercial (ProCon) reagent kit. The optical densi-
ties of solutions were measured on an Labsystem MCC-340
multiscan unit. The concentrations were calculated on a com-
puter using a Biostat program package.
The ability of lectins to modulate the killer activity of
lymphocytes was studied on K-562 (transferred cell culture
of human erythroblastic leukemia) and AKL (transferred
monolayer cell culture of human lung adenocarcinoma) in a
1 : 5 ratio. The tests were performed on 96-well Costar plates
with 30 ´ 10
3
target cells per well. The cells in complete me
-
dium were incubated for 18 h at 37°C and 4% CO
2
and
stained with a vital dye MTT (Sigma). After a 4-h exposure,
actively matabolizing cells oxidize MTT to yield water-in
-
soluble formazan crystals, which were dissolved by adding
313
0091-150X/02/3606-0313$27.00 © 2002 Plenum Publishing Corporation
Pharmaceutical Chemistry Journal Vol. 36, No. 6, 2002
1
Blokhin Oncological Research Center, Russian Academy of Medical Sci
-
ences, Moscow, Russia.
2
Sechenov Medical Academy, Moscow, Russia.
TABLE 1. Modulant Action of Lectins from Culti
-
vated Peavine on the Spontaneous Cytotoxicity of
Lymphocytes from Healthy Donors
Lectin
fraction
Percentage cytotoxicity
with respect to tumor cells
AKL K-562
Control
29 ± 232± 3
L
1
32 ± 239± 5
L
2
42 ± 5* 49 ± 9*
L
3
45 ± 6* 48 ± 2*
L
4
47 ± 8* 49 ± 3*
*
Differences from control are reliable for p < 0.05.