From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
Evaluation
of Surface
Lymphocytes
Containing
A Quantitative
Markers
on Normal
Human
Parallel
Tubular
Arrays:
Ultrastructural
Study
By Claire M. Payne and Lewis
Normal
lymphocytes
sent
a
surface
tubular
and
arrays
whether
this
tutes
the
cell
B-lymphocyte
cytes
K cells.
of
subpopulations.
mine
if this
tutes
the
or
cell
was
using
fraction
used
Spontaneous
were
with
used
as a marker
R,R2
erythrocytes
T
of any
the
HAS
the
formed
used
as
for
three
with
complement
with
Fc receptors
from
for
T
rabbit
Human
Ripley
(EA6,-rosettes)
well
the
to
consti-
documented
peripheral
of
of
PTA.
healthy
1 1 .5%
of the
of
lymphocyte
1gM
major
group
anti-CD
were
that
circulation
0,
used
to
Fc
the
type
of 369
was
marker
populations
parallel
for
cytophilic
and
antithat
the
constitutes
the
of lymphocytes.
The
within
the
only
a population
assay
inclusions
within
EAC-
cross-
that
PTA-lymphocyte
and
1 51
that
for
only
cross-
of
concluded
population
the
that
lymphocyte
receptors
of
E-rosettes
revealed
EA,.,,-rosette
third
of
96%
cross-sections
examination
these
of the
inclusion
lymphocyte
tion
by
lympho-
subject,
revealed
subjects
It
subjects
lymphocyte
a morphological
containing
cell
In one
rosette-forming
bearing
identified
possible
type
serum
and
PTA.
are
lymphocytes
lymphocytes.
were
central
contained
body
PTA.
rosette-forming
four
five
rosette-forming
subjects
contained
arrays
Ultrastructural
examination
from
mean
E-rosettes.
the
lymphocyte
rosettes
erythrocytes
antibody
central
following
from
healthy
Light
obtained
79.2%
EA6,-rosettes.
central
sections
tubular
populations.
1 6%
of the
five
the
sections
assays
the
EA,-rosettes
In contrast,
from
3.4%
revealed
contained
PTA.
obtained
lymphocytes.
preparations
rosette-forming
contained
deter-
of
the
lymphocytes:
and
1 78
89%
central
lymphocyte-
major
that
of
subjects
of
the
lympho-
population
reacted
rosettes.
of
(EAC-rosettes)
B lymphocytes.
sensitized
revealed
marker
sheep
a marker
consti-
lymphocyte
a
EAC
examination
analysis
on
8.8%
cross-sections
was
surface
healthy
of
known
lymphocyte
three
five
cyte
lymphocyte
of these
with
sensitized
mouse
BEEN
lymphocytes
three
with
third
examination
percentage
or is unrelated
study
the
population.
ultrastructural
identify
erythrocytes
coated
this
performed
rosettes
(E-rosettes)
I
to
cells).
defined
A quantitative
commonly
null
of
from
these
parallel
identified
population
identifiable
type
enriched
identify
M cells.
purpose
PTA-lymphocytes
Sheep
third
of
It is not
T-Iymphocyte
for
microscopic
called
be
marker
different
lymphocyte
immunologically
morphologically
subpopulations.
and
of the
I cells,
The
major
only
defined
type
these
Some
microscope.
population.
(i.e..
any
can
electron
repre-
with
inclusions
ultrastructurally
major
cells
functions.
which
the
circulation
of
cytoplasmic
(PTA).
under
peripheral
different
contain
certainty
the
population
markers
lymphocytes
to
from
heterogenous
Glasser
monocyte
other
popula-
is discussed.
as a
normal
repre-
ies using scanning
T and B lymphocytes
electron
microscopy
do have subtle
indicated
that
but nevertheless
sent a heterogeneous
population
of cells with different
surface
markers
and different
functions.’
The twocomponent
concept
or duality
of the immune
system5
is gradually
being
replaced
by a three-component
concept
after a third major
population
of lymphocytes
was identified
in mice69
and later in humans.’#{176}’3 This
discernable
differences
in surface
morphology.26
It is
possible
to demonstrate
an abundance
of rough
endoplasmic
reticulum
in some B cells.27’28 This represents,
however,
a stage
of differentiation
following
B-cell
third
population
of cells
is characterized
presence
of Fe receptors
that bind cytophilic
and
have
been
variously
described
as K
contain
unique
cytoplasmic
inclusions
called
parallel
tubular
arrays
that
may
be visible
as nonspecific
azurophilic
granules
on blood
smears
stained
with
Romanowsky
dyes. These
inclusions
can only be identified
with certainty
under
the electron
microscope.
Normal
M cells,’8 and
lymphocyte
null cells.’9
subpopulations
present
by the
antibody
L
in the
peripheral
circulation
cannot
be identified
morphologically
by light microscopy.
Although
some
investigators report
successful
separation
of populations
using
cytochemistry,
there are conflicting
reports
regarding
the
specificity
of these
findings.20
Ultrastructural
differences
have been
noted
using
transmission
electron microscopy,2123
however,
these
subtle and do not define homogeneous
of cells.
microvilli
Scanning
electron
on B lymphocytes
microscopy
initially
showed
and a smooth surface
on T
lymphocytes,24
but subsequent
difference
to be a preparative
Blood,
Vol. 57, No. 3 (March).
differences
are
subpopulations
1981
studies have shown
this
artifact.25
Further
stud-
and is not seen
B lymphocytes.
activation
circulating
The
purpose
morphologically
the
major
From
Health
of
cell
the
Submitted
type
August
Pathology,
University
is to determine
if this
lymphocyte
constitutes
of the
of
Center.
reprint
Address
this article
identifiable
Department
Sciences
in the majority
of normal
Some human
lymphocytes
T lymphocyte,
Pathology,
Tucson.
28. 1980;
requests
University
B lympho-
of
Arizona
Ariz.
accepted
to Dr.
of Arizona
November
Claire
M.
Payne.
Health
Sciences
1 1, 1980.
Department
Center,
of
Tucson.
Ariz. 85724.
© 198!
by Grune
& Stratton,
inc.
0006-4971/81/5703--0027$01.00/o
567
From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
PAYNE
568
cyte,
or third
tive
ultrastructural
was
performed
assays
population
using
used to identify
these three
conclude
that parallel
tubular
commonly
populations.
Preparation
of lymphocytes.
A quantitaof lymphocyte
rosettes
three different
surface
marker
analysis
We
major
arrays
lymphocyte
within
The possible
inclusion
of the
other
lymphocyte
populations
population
is discussed.
within
monocyte
the
MATERIALS
Lymphocyte
blood
adults
evacuated
gradient
fraction
mm
(3
were
2 male)
Whole
The
from
which
96%
to
99%
recoveries
lymphocyte
purities
I .077).
and
(mean
ranging
incubated
spun
in large
thin
acetate
and
Hitachi
HU-I2electron
prepared
blood
rosette
according
cells
Following
wrapped
serum
(FCS)
Microscopy
A
excess
as measured
to 83%
by
(mean
(mean
In the EA66-rosette
examined
rang-
37#{176}C
and
balanced
lymphocytes
at
eosin
dye
that
and
the surrounding
96.6%).
200
g, and
(0.25
described.29#{176} Sheep
sheep
ml),
were
incubated
range
with
erythrocytes
FCS
incubated
62%
at
ml),
15 mm
with
a mean
uranyl
examination
with
a rosette
a
as a lympho-
One-hundred
by light
in each
was
erythrocytes.
was defined
examined
scored
a rosette
attached
crythrocytes.
rosettes
microscopy.
A
rosette preparation
as a lymphoid
that
was
at a scope
found
could
cell
The
of each
through
magnification
the
io,000x.
of
parallel
central
attached
This
tubular
a similar
at
as a parallel
cytoplasm
examined
can have
containing
be identified
necessary-since
cytoplasm-and
percentages
electron
close
arrays
density
as
be easily missed at a low magnifi-
section
fraction
taken
for
of the total
Preparation
Electron
0.50
ml of
values
of
PTA-lymphocytes
are an underestimate
4#{176}C
and
The
scored
in
value
since
of the true
electron
microscopy
cell volume.
constitutes
each
rosette
an ultrathin
only
a small
in our
of Unfractionated
removing
Whole
Blood
subjects
were
for
Microscopy
buffy
electron
at 37#{176}C,
pelleted
at 4#{176}C.
Normal
to 86%
grids.
and
red
in Hanks’
and
ultraMT2-B
copper
citrate
preparations,
magnification.
are not membrane-bound
calf
Fetal
suspension
(0.25
for
overnight
from
was
solution.
and
cation.
erythrocytcs
in Alsever’s
at 56#{176}C.A 2% SRBC
solution
(106/ml)
laboratory
sheep
previously
suspended
absorbed
inactivated
salt
system
preparation
using
at high
79%),
200-mesh
or more
was defined
was
selected
lead
cut and
microscope.
on a Sorvail
before
three
inclusion
lymphocyte
ocular
were
microscopy.
one cytoplasmic
lox
were
with
EAC-rosette
attached
by electron
rosette-forming
of ethanol,
the light
knife
in 0.1
tetroxide
series
sections
with
later
by Light
of 26 rosettes was
examination
a graded
rosettes
in each preparation
array
1% osmium
One-micron
preparations,
five or more
tubular
in
stained
with
PTA-lymphocyte
least
iron
viabilities
and
as a lymphocyte
with
described.32
carbon-coated
of Rosettes
Electron
as previously
microscope.
Quanlitalion
Instrument
and
had
to 98%
formation
were
was
lightly
and
tubes
on uncoated,
were
and
minimum
incubation,
the
around
a magnet
phagocytes
95i
to methods
(SRBC)
then
for stabilization
a diamond
mounted
sections
I .5% giutaralde-
7.2)
and examined
with
GLASSER
Microscopy
with
through
numerous
cut
and
were scored
magnetic
epoxy.33
blue
were
ultrathin
cyte
of E-Rosettes
Spontaneous
were postfixed
containing
ultramicrotome
The
(pH
plastic
in Spurr’s
blocks
Electron
in suspension
dehydrated
toluidine
sections
The
Preparation
buffer
pellets
with
Those
phosphate
buffer,
and embedded
stained
fixed
round-bottomed
M phosphate
at 37#{176}C
for 30
micron-sized
76%
5
mononuclear
contains
97.2%)
from
The
(Technicon
from
from
heparinized
by Ficoll-Hypaque
preparations
ranging
into
Reagent
iron-laden
lymphocyte-rich
exclusion,
gr.
poly L-lysine.
tygon
tubing
removed
final
drawn
was separated
was removed
with
through
by venipuncture
and
(sp.
N.Y.)
effectively
filings.
blood
Separator
Tarrytown,
that
METHODS
collected
centrifugation
particles
sensitized
cells were passed
ing
samples
Lymphocyte
Corp.,
in 0.1 M
for
initially
in the E-rosette
at the interface
with
hyde
defIned
female,
test tubes.
density
PTAand
Isolation
Peripheral
healthy
AND
were
The resulting
are a morphological
marker
for a population
of
lymphocytes
bearing
Fc receptors
for cytophilic
antibodies
identified
by the
EAhU-rosette
assay.
The
lymphocyte
containing
these
inclusions
by definition
constitutes
the major
cell type of the third population
of lymphocytes.
of Rosettes
Rosettes
AND
coats
microscopy
from
I 6 normal
by centrifuging
the plasma,
and
whole
immediately
blood
fixing
prepared
in Wintrobe
the cells
for
tubes,
as previously
described.34
of 78%.
RESULTS
Preparation
Sheep
erythrocytes
(Cordis
the
of EAC-Rosettes
were
Laboratories,
minimum
prepared
The
sensitized
Miami,
agglutinating
absorbed
37#{176}C.3’A 0.5-mi
whole
suspension
of 1% EAC-SRBC
ous mixing
for 30 mm.
of EAh6-Rosettes
Ripley
anti-CD
Equal
volumes
and
lymphocytes
type
serum.”
placed
upright
in our
laboratory
coated
using
with
(EAC-SRBC)
(l06/ml)
incubated
at
and
at 37#{176}C
with
in our
half
freshly
laboratory
0.5-mI
continu-
range
from
were
at
room
range
five volunteers
chosen
for
this
study
had
normal
hematologic
values. The white
blood cell counts
had a
mean of 6300 with a range of 3500-9900.
The mean
percentage
of lymphocytes
was 29.8 with a range
of
23-38.
The
mean
absolute
lymphocyte
count
was
I 762/tl
with a range of I 2 1 6-2574/s1.
Light
microscopic
analysis
of the three
different
rosette preparations
(Table
I gave results comparable
to those previously
described.”293’
The mean percent-
)
R1R2
The
(l06/mi)
tubes
antibody
temperature
then
complement
values
1gM
of 8%.
of suspensions
values
rabbit
lymphocytes
were
Preparation
0,
and
Normal
a mean
group
at room
of
4% to I 2% with
Human
Fla.)
titer
mouse
suspension
with
erythrocytes
were
of the
antiserum
titer
of 1% sensitized
centrifuged
temperature
from
sensitized
was
with
1:80,000.
R,R2 erythrocytes
for
5 mm
for
8% to 25% with
30
at 200 g and
mm.
a mean
Normal
of I 5%.
age of E-rosettes
was 79 with a range of 72-86.
The
mean absolute
number
of E-rosetting
cells was 1415/
l with a range
of 876-2162/.zl
(Table
2). The mean
percentage
of EAC-rosettes
was 8.8 with a range of
5-12.
The mean
absolute
number
of EAC-rosetting
cells was 140/tl with a range of6I-237/sl.
The mean
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SURFACE
MARKERS
Table
ON PTA-
1 . Percentage
569
LYMPHOCYTES
of Cen tral Rosette-Forming
Cells
Containing
P arallel
Tubular
Percent
PTA-Lymphocytest
Percent
Volunteer
as CRFC
Lym phocyte
Percent
as CRFC
EAC-Rosettes
Populations
Percent
PTA-Lymphocytest
in EAC-
Percent
as CRFC*
‘
Rosettes
EA,-Rosettes
Rosettes
13(31)
18
93(29)
25
74
0(38)
12
2
72
2(44)
5
12(34)
3
86
2(180)
6
12(51)
8
4
80
7(41)
12
9(35)
13
92(48)
84
6 (66)
ND
ND
16
96 (26)
5
§ Numbers
-
cellular
cell.
in parentheses
of
percentage
cross-sections
indicate
contained
the total
EAh-rosettes
number
was
16
parallel
of rosette
with
a
tubular
Ultrastructural
rosette-forming
analysis
of the percentage
cells
containing
parallel
(PTA-lymphocyte-rosettes)
of the EAh5-rosettes
majority
the
central
mean
The
cross-sections
range
of
of central
tubular
revealed
that
PTA-lymphocytes
had
mean of I 1 .5 ± I .5 SD and a range of 9-1 3. The
absolute
number
of PTA-lymphocyte-E-rosettes
58/tl
with a range
of 0-130/.tl.
number
of PTA-lymphocyte-EAC
with a range of 7-2 1 /l. The
PTA-lymphocyte-EA5-rosettes
range of I 2 1-396/l.
Except
appeared
arrays
at
I).
ultrastructurally
Table
2.
Absolute
of section,
EAh-rosettes
in any
Numbers
given
E-rosettes/zl’
of Central
mean
lymphocyte
cally
EAhU-,
and
rosettes
EAC
was
257
±
there were less
and
rosetted
was not statisti-
tubular
arrays
inclusions
found
These
(PTA)
in
blood
distinct
lymphocytes
population
and
is to what
ultrastructurally
inclusions
Rosette-Forming
Lymphocyte
Cells
Containing
population
Parallel
ultrastructurally
of some
observed
or nonhematopoietic
cell.
that
normal
peripheral
containing
of cells
A question
distinct
are
the cytoplasm
have not been
PTA
behave
as a
when
challenged
with
we now
propose
to
of lymphocytes
cells
belong:
Tubular
Arrays
the
do these
B-lympho-
Populations
PTA-Lymphocyte
EAC-Rosettes/I
EA,-rosettes/pl
984
0
239
222
876
18
61
7
304
231
3
1,471
29
103
12
137
121
4
1,582
‘Determined
from
light
tDetermined
from
electron
microscopic
microscopic
21
PTA-Lymphocyte
EA-Rosettes/I
1
2.162
111
237
21
257
236
130
ND
ND
412
396
results.
results.
±
of PTA-lymphobuffy coat prepa-
2
5
the
significant.
glucocorticoids.34
159
induced
from
unfractionated
bully
absolute
number
of PTAafter
adding
up the PTA-
in any other
hematopoietic
We have
previously
shown
the
Human
assay,
I 18.7/tl.
Although
in the
purified
fractions,
this decrease
328
was
PTA-lymphocytes
lymphocytes.
EAC-Rosettes/Ml
E-Rosettes/lt
rations
Parallel
PTA-Lymphocyte
Volunteer
E-,
answer
in Normal
EAhU-rosette
84.7/,l.
The mean
absolute
number
cytes determined
from unseparated
distinct
subject
procedure,
microscope.
DISCUSSION
absolute
number
of
was
241/tl
with
a
for the absence of paral-
similar
the
mean
was
mean
from subject
to subject.
In order to determine
if the rosetting
the electron
particular
lymphocyte
The mean
absolute
rosettes
was 15/jtl
in some of the planes
the
center
of the
in
under
lymphocytes
determined
coat preparations.
The
lymphocytes
determined
the
cell (Table
I , Fig.
PTA-lymphocyte-EA-
rosette-forming
percentage
of
examined
formation
of parallel
tubular
arrays,
the
absolute
number
of PTA-lymphocytes
determined
after
rosetting was compared
to the absolute
number
of PTA-
rosettes
was 89.0 ± 7.0 (SD)
with a range
of 76-96.
The mean
percentage
of PTA-lymphocyte-E-rosettes
was only 3.4 ± 2.6 (SD)
with a range
of 0-7.
The
mean
percentage
of PTA-lymphocyte-EAC-rosettes
was slightly
higher
than
that of the E-rosettes
with a
lel tubular
lymphocytes
76(42)
88(33)
arrays.
8-25.
The mean absolute
number
of EA5-rosetting
cells was 270/gil
with a range of 137-412/.tl.
arrays
EA.-
by light microscopy.
tLymphocytes
whose
CentraI
rosette-forming
and
Human
1
‘Determined
as
i n Normal
PTA-Lymphocytest
in
E-Rosettes
E-rosettes
Arrays
Percent
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570
PAYNE
AND
GLASSER
Fig. 1 .
Composite
electron
micrograph showing
a typical
EA,,,-rosette.
(A) Low-power
electron
micrograph
of an EA6,,-rosette.
The
central
rosette-forming
lymphocyte
is surrounded
by 8-1 1 erythrocytes.
The
lymphocyte
has a fairly large cytoplasmic area containing
numerous
dense
granules
(arrow)
and
an irregular
surface
membrane.
There
is much
interdigitation
evident
between
the
lymphocyte
and erythrocyte
surfaces.
The area of the cytoplasm
indicated
by the arrow
is shown
at higher
magnification
in the bottom
electron
micrograph.
(Uranyl
acetate.
lead
citrate;
x 5900.)
(B)
Higher
power
electron
micrograph
of the cytoplasm
of the central
rosette-forming
lymphocyte
shown
in A. Numerous
paralId tubular
arrays
having
a typical
microtubular-like
appearance
(arrows) are present
in the ‘hof” region
of the nucleus
near the centriole
(Ce)
and the Golgi apparatus
(Go). (Uranyl
acetate.
lead citrate;
x46.700.)
cyte population,
population
of
T-lymphocyte
lymphocytes
population,
(i.e.,
K cells,
or the third
L cells,
M
cells,
null
cells)?
Previous
stt’dies
have
failed
to
unequivocally
determine
the
true
identity
of PTAlymphocytes.
Brunning
and Parkin35
and McKenna
et
al.36 in a study of a chronic
lymphoproliferative
disorder and
McKenna
et al.37 in a study
of infectious
mononucleosis,
rosettes
and
described
concluded
PTA-lymphocytes
that
the
PTA-lymphocyte
in
E-
is
most
probably
PTA-lymphocyte
was
challenged
a T lymphocyte.
as simply
when
Payne
The
a subset
et
identity
of the
of T lymphocytes
al.,38
in
a study
of
severe combined
immunodeficiency
disease,
identified
PTA-lymphocytes
in both E- and EAC-rosettes.
Since
the EAC-rosette
has been criticized
as measuring
both
Fc receptors
and complement
receptors,
Payne
and
Nagle39 identified
true complement
receptors
on PTAlymphocytes
from
the normal
peripheral
circulation
From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
SURFACE
MARKERS
ON PTA-
LYMPHOCYTES
using
an assay system
antibody
and activates
571
that circumvents
complement
by
the
the use of
alternate
pathway.
Appay
et al.,4#{176}
in a study
of six normal
subjects,
could
not detect
surface
immunoglobulin
on
PTA-lymphocytes
using anti-human
1gM peroxidaselabeled
Fab
lymphocytes
fragments.
containing
inclusions”
(BST)
could
not
to the
third
are
if
determine
consistent
lymphocyte
with
neoplastic
reported
of
that
the
tubular
not B lymphocytes,
but they
they belonged
to T cells and/or
population.
the
cells
They
concluded
these “bundle-shaped
This
observation
absence
proven
of
PTA
is
the
in
B-cell-derived
neo-
plasms.35’36
In this
study,
a quantitative
cell
rosette-forming
using
to distinguish
the three
that
the PTA-lymphocytes
type
of the third
evidenced
used
by
by
major
fact
analysis
of the
assays
commonly
populations
constitute
population
the
as a marker
three
the
89%
for the third
of the
used
has revealed
major
cell
of lymphocytes.
that
central
This
was
EAh-rosettes,
population,
were
formed
in contrast
to only
3% of the
E-rosettes
and I 2% of the EAC-rosettes.
It is noteworthy
that
in one subject,
96%
of the
EAhU-rosette
cross-sections
contained
parallel
tubular
arrays (Table
I ). In order
to determine
if the rosetting
procedure,
and
PTA-lymphocytes
in particular
formation
number
the
of
preparations
of
PTA-lymphocytes
coat
determined
determined
in the
from
Although
there
significant.
The
loss
in PTA-lymphocytes
the
absolute
to the absolute
was compared
preparations.
induced
the
arrays,
roset-
number
unseparated
was
in the number
of PTA-lymphocytes
preparations,
this decrease
was not
decrease
rosetted
cally
assay,
tubular
of PTA-lymphocytes
ted
buffy
EAhU-rosette
parallel
a 22%
in the
statistiin the
purified
and rosetted
fractions
may be a reflection
of
the 2 1 % loss in total lymphocytes
resulting
from the
isolation
procedure.
The parallel
tubular
array
can
therefore
be used as a morphological
marker
for the
third
population
for cytophilic
assay.
The
lymphocytes
of lymphocytes
antibodies
finding
in E-
that
identified
bear
by the
Fc receptors
EAhU-rosette
of a small
percentage
of PTAand
EAC-rosetting
preparations
two different
types of lymphocytes
with strong
PAPlabeling
of their membranes
were recognized.
The first
subgroup
of strongly
PAP-positive
lymphocytes
showed
a low degree
of morphological
differentiation,
and the second
subgroup
contained
parallel
tubular
arrays.
This is in contrast
to our finding
in which
the
Fc receptor-bearing
lymphocytes
identified
by means
of the EAhU-rosette
assay
appeared
to represent
a
morphologically
homogenous
cell population,
most of
which
could be determined
to contain
parallel
tubular
arrays.
the
Perhaps
EA-rosette-forming
to the
(ADCL),
correspond
lymphocytes
lymphocytes
Other
support
having
Fc receptors
work
from
the concept
antibody-dependent
shown
that after
thasone,
the
our
that
percentage
tion
include
third
population
EAC-rosettes
third
popula-
cells”2’44 as well.
The finding
of Fc receptors
on PTA-lymphocytes
is
consistent
with
the
finding
of Huhn
et al.45 who
demonstrated
Fc-receptors
using
soluble
peroxidaseanti-peroxidase
(PAP)
complexes.
They
stated
that
We have
of dexame-
of PTA-lymphocytes34
the
and
percentage
of EAhU-rosettes46
increased
significantly
in
the
peripheral
circulation,
whereas
the
absolute
number
of each remained
essentially
unchanged.
This
to steroids
glucocorticoid
populations.
insensitivity
have
fewer
lymphocyte
phocyte
Parrillo
population
and Fauci.47
significant
may
indicate
that
receptor
sites
The postdexamethasone
was
They
increase
in
analyzed
showed
these
cells
than
functionally
that
there
other
lymwas
antibody-dependent
by
a
cell-
mediated
cytotoxicity
by the poststeroid
population.
The finding
of a small
percentage
of E- and EACrosettes
sistent
containing
with
the
parallel
concept
that
tubular
arrays
the
PTA-lymphocytes
is not inconare
“killer”
cells exhibiting
antibody-dependent
cytotoxicity. West
et al.4’ found
that
low affinity
E-rosetteforming
cells were highly
enriched
for cytotoxic
reactivity
against
antibody-sensitized
Chang
liver
cells,
whereas
tamed
cells).
high-affinity
E-rosette-forming
cells
confew antibody-dependent
cytotoxic
cells
(K
It is also known
that in addition
to B lympho-
found on the same PTA-lymphocyte.
absolute
PTA-lymphocyte-rosette
has shown
that if these surface
percentage
ADCL).
cytotoxic
lymphocytes.
the in vivo administration
origin,30
Lymphocytes
known
to form
not only B lymphocytes42’43
but
and
laboratory
and elsewhere
the PTA-lymphocytes
are
cytes,42’43
complement
antibody-dependent
a small
cytotoxic
lym-
phocytes
identified
by the use of PAP
complexes
represent
two or more functionally
distinct
lymphocytes (i.e., a subpopulation
of T lymphocytes
and/or
B
can be explained
by the lack of specificity
of these
surface
marker
assays
or the inclusion
of parallel
tubular
arrays
in other
minor
subpopulations.
Although
most
E-rosetting
lymphocytes
are of T-cell
cells.”2’4’
include
lymphocytes
antibody-dependent
and the agg-Ig-positive
receptors
can be detected
on
cytotoxic
lymphocytes.’
It is not
possible
in the experimental
study
to determine
if all
three
four
cells
mutually
exclusive
design
used
three
surface
Determination
of
counts
(Table
2)
markers
represent
subpopulations,
times as many
PTA-lymphocyte
as PTA-lymphocyte
E-rosetting
peripheral
lymphocyte
circulation
EA5-rosetting
and
in the present
markers
are
there
EA-rosetting
cells
in
are
the
16 times
as many
PTAcells
as PTA-lymphocyte
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572
PAYNE
EAC-rosetting-cells.
although
This
all
cytophilic
Ripley
cytes,
seems
lymphocytes
to
antibody
indicate
that
receptors
to be PTA-lympho-
bearing
appear
shown,
for
Fc
PTA-lymphocytes
seem
to form
Erosettes
even
less have
complement
receptors.
E-rosetting
PTA-lymphocytes
may include
T-y cells,
since
Fc receptors
for Ripley-equivalent
antibodies
have been demonstrated
on their
surface.48
The relationship
between
third population
lymphocytes
and Ty
fewer
and
and
numerous
cytoplasmic
not have the
parallel
tubular
granules
microscopy
granules,
but
these
typical
ultrastructural
arrays.5#{176}The relationship
arrays
seen
in T”y cells
and
seen in EA5-rosette-forming
further
study.
did
granules
appearance
between
the
cells
receptors
since these
ultrastructural
cytochemical
in some
and
but are atypical
by Kay and
tion
the
Horwitz52
OKMI
origin
of
lymphocytic
monocytes.
We
peroxidase
activity
have
unable
been
PTA-lymphocytes
granulocytic
that
have
the
with
are
iden-
been
unable
conclusion,
to demonstrate
cell
type
of the
or
B-lymphocyte
however,
population.
represent
the
major
antiFuture
to determine
the ontogand the functional
signifiarrays
in the physiology
experimentation
is necessary
eny of the PTA-lymphocyte
cance
of parallel
tubular
immunology
popu-
ultrastructurally
of these
distinct
cells
and
either
and
as a morphological
marker
for this lymphocyte
lation,
which
contains
Fc receptors
for cytophilic
body
identified
by the
EAhU-rosette
assay.
ACK
to demonstrate
thus
wish
Pathology)
for
excellent
or
We
The authors
of
far
monocytic
differentiation.
immunodeficiency,38
mononucleosis,37
cell type of the third
population
of lymphocytes.
It is
our opinion
that the parallel
tubular
array can be used
and/or
cells.
ques-
T-
do,
a
in
shown
that
lymphocytes
arrays
do not represent
the
have
tubular
PTA-lymphocytes
a
lymphocytes
combined
we
parallel
cytotoxicity.
were probably
in PTA-lymphocytes39
enzymatic
severe
recircu-
From
constitute
circulating
increase
in numbers
in infectious
may become
neoplastic.3653
In
effector
cellular
the cells
the
patients
containing
antibodies
of
portion
of
the
features
of lymphorespects.
Recent studies
using
involved
in antibody-dependent
They concluded,
however,
not
standpoint,
the
determined
not
glucocorticoids.
did
tified on third population
cells.5’ It is unlikely
that C3d
receptors
are found
on PTA-lymphocytes,
since
these
receptors
are restricted
to B lymphocytes,5’
and Appay
et al.4#{176}
have shown
that
PTA-lymphocytes
are not B
lymphocytes.
The cells containing
parallel
tubular
arrays
have
cytes
do
with
tubular
on PTA-lymphocytes
receptors
have been
been
challenged
major
The complement
most likely C3b,
have
late
have
of
some
PTAcomplement
apparently
and
when
activity,39
least
and
have
E-rosettes
PTA-
majority
At
to be steroid-resistant34
requires
parallel
form
that
immunoglobu-
the
PTA-lymphocytes
pathologic
show
indicates
peroxidase
cells.
by
phagocy-
surface
constitute
EA-rosette-forming
major
Electron
possess
no endogenous
lymphocytes
differentiation
to date
not
GLASSER
PTA-lymphocytes
bacteria.39
evidence
do
Fe-receptors,45
light
microscopy.5#{176}
elucidated.
functional
some
lins,4#{176}
have
receptors.39
has
some
Experimental
lymphocytes
Functionally,
both
third
population
lymphocytes
and
Ty cells
express
killer-cell
activity.’4’949
Of interest
is the finding of cytoplasmic
azurophilic
granules
in Ty cells by
cells
not been completely
however,
demonstrating
that
tize complement-coated
AND
College
have
anti-CD
NOWLEDGMENT
to thank
his
Dr.
Jack
invaluable
Layton
support,
(Head,
Roger
Marion
Department
Fiederlein
technical
assistance,
and
Dr.
Wailer
of Virginia,
Richmond,
Va.)
for her generous
for
(Medical
gift
of Ripley
serum.
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From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
1981 57: 567-573
Evaluation of surface markers on normal human lymphocytes containing
parallel tubular arrays: a quantitative ultrastructural study
CM Payne and L Glasser
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