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Kx:
Its
Relationship
to Chronic
Granulomatous
Linkage
By
The
Peter
Densen,
relationship
neutrophil
chronic
Kx.
was
Xg
Wilkinson-Kroovand,
between
antigen.
with
Xk,
Susan
neutrophil
as well
examined
granulomatous
function
as the
linkage
in a kindred
disease.
Four
and
of the
with
of
the
L. Mandell,
the
male
siblings
had chronic
granulomatous
disease
by clinical
history
and tests of neutrophil
function,
and all four had
Kx-negative
neutrophils.
The remaining
four were in good
C
as
HRONIC
granulocyte
an X-linked
granulomatous
function
recessive
disorder
in some
is transmitted
kindreds.
recurrent
infections
neutrophils
have
ingested
inability
disease
is a disorder
of
that is inherited
primarily
trait,
although
a similar
as an autosomal
Affected
individuals
because
their
defective
recessive
suffer
activity
This
defect
to generate
neutrophils
that
that
linked
a causal
relationship
surface
antigen
and
other erythrocyte
surface
antigen
an X-linked
mode of inheritance.
Evans
Department
Boston,
of
The
New
Supported
from
and
Va. and
York
Address
Room
5) 1981
the
Blood
in part
the National
Submitted
tal,
the Division
Medicine
lottesville,
June
reprint
E529,
by Grune
of
Pathology,
by Grants
Institutes
Diseases,
F. Kimball
New
York,
requests
to Dr.
siblings
been
reported.4
75 E. Newton
of
Red
& Stratton,
Inc.
data
on the
suggest
X chromosome
In addiand Xg,
and
HLO9OI
Boston,
University
Mass.
02118.
If ospi-
other
of the
ABO,
Rh,
groups
using
have
granulomatous
of recurrent
patients
of
an
four
previously
disease
infections
to reduce
parameters
details
of the
was
and
nitroblue
the
tetra-
oxidative
burst
of
phagocytosis.
Typing
from
MN,
standard
all family
members
Lutheran,
Kell,
were
Duffy,
tested
Kidd,
for antigens
and
Xga
bkod
techniques.5
Typing
Erythrocyte
indirect
Kx determinations
antiglobulin
Serum
from
a patient
been
inadvertently
Equal
cells
was
null
(K0)
with
Marsh.6
erythrocytes,
ability
on the
neutrophil
blood
The
neutrophil-rich
mm,
22#{176}C),and
hypotonic
(0.2%)
packed
neutrophils
standard
anti-Kx
test
used
were
the
system
performed
using
of Kx antigen,
to reduce
and
removed,
in
blood
twice
10%
suspended
in
an
for
I hr
Vol.
cells
equal
at
58,
dextran.
(200
in isotonic
incubated
Blood,
anti-Kx
1 5 ml of ACD-treated
centrifuged
red
washing
the
of Kx antigen
sedimentation
of contaminating
serum,
red
erythro-
polyspecific
as a measure
from
by
was
After
were
blood
the
final
using
an excess
Neutrophils
saline.
and
The
scored
red
at 37#{176}C
for
centrifuged
saline.
neutrophils
obtained
freed
subject’s
had
cells.2
Kx antigen.
was
plasma
red
of commercial
have
subject’s
surface.
were
Kx who
incubated
controls
lack
antiserum
test
for
obtained
Kx-positive
then
drops
which
that
of the
of a standard
two
scored
neutrophil
hemagglutination
Appropriate
erythrocytes
The
was
in isotonic
and
of 32 was
were
times
incubated
titer
of the
mixture
a standard
being
normal
serum
using
result
and
with
anti-Kx
three
globulin
by
an anti-Kx
suspension
and
incubation
washed
pellet
McLeod
saline
the
erythrocyte
of a 3%-5%
The
cells
with
absent
performed
with
transfused
volumes
30 mm.
Kell
with
in isotonic
blood
were
technique
hemagglutination.
1
with
studies
disease
of chronic
history
these
together
function
granulomatous
during
Cell
family
neutrophil
diagnosis
exhibit
Blood
described
2, 198/.
Densen.
to
cells
of
and
from
activity
blood
titer
AI16476,
METHODS
three-generation
chronic
The
Red
Kx
Char-
Institute
with
or
metabolic
of
Virginia,
Research
March
Street,
Kx-negative
X chromosome.
AND
by a compatible
zolium
Center,
Departments
of this
of neutrophils
peripheral
Peter
reduction
had
These
genes
on the
features
male
failure
N.Y.
AI09504,
accepted
normally.
distinct
linked
pedigree
clinical
anti-human
of Health.
2, 1980;
four
Patients
established
Research,
Medical
University
Lindsley
closely
The
might
exist
the defective
Clinical
University
of Infectious
Center,
0006-497l/81/5801-0005$01.00/0
34
Boston
but
tetrazolium
latter
functioned
L. Marsh
cate
that
the
absence
of Kx from
neutrophils
is
strongly
associated
with
chronic
granulomatous
disease,
but
does
not
preclude
normal
oxidative
neutrophil
function.
In addition,
the Xk and Xg loci
of the
that is known
to
The data mdi-
Department
Medicine,
Mass.;
Internal
Memorial
nitroblue
of these
W.
code for chronic
granulomatous
disease
and Kx.
tion, close linkage was demonstrated
between
Xk
a gene coding for an erythrocyte
surface antigen.
cyte
the
closely
#{216}yen, and
MATERIALS
the
of
We investigated
the association
of these two abnormalities
in a large
kindred
with
X-linked
chronic
granulomatous
disease.
We also examined
the possibility
of linkage
between
Xk and Xg loci. Xk alleles
code for the surface
antigen,
Kx, on both granulocytes
and erythrocytes,
while the Xg locus codes for Xga, the
From
normal
one
against
is due to
metabolites
to superoxide
ion, is believed
to account
for this defect
in oxidative
activity,
but
the
precise
biochemical
abnormality
has not been established.’
Recently,
granulocytes
from patients
with X-linked
chronic
granulomatous
disease
have
also been
reported
to lack
an
X-linked,
Kell-related
surface
antigen,
Kx. This antigen is present
on granulocytes
from normal
individu-
only
have
Ragnhild
had
However,
appear
oxygen
that
are
toxic
to ingested
bacteria.
The
dysfunction
or absence
of a membrane-bound
oxidase,
which
catalyzes
the one electron
reduction
of oxygen
als, suggesting
that
between
the absent
oxidase
activity.2’3
Genetic
polymorphonuclear
bactericidal
microorganisms.
of these
cells
trait
from
Sullivan,
and
tests.
X-Iinked
eight
Gail
health
gene.
and
Xg
With
Gerald
Disease
by
saline,
volume
37#{176}C,and
No.
g,
1 (July),
10
lysis
in
the
of
the
then
1981
From www.bloodjournal.org by guest on June 11, 2017. For personal use only.
Kx, CHRONIC
GRANULOMATOUS
DISEASE,
AND
Xg
35
Xg(a+)
Xg(ci+)
Fig. 1 .
Partial
family
pedigree
demonstrating
the relationship
between
chronic
granulomatous
disease
(CGD)
and neutrophil Kx antigen
as well as the genetic
linkage between
Xg and Xk.
The results
of
erythrocyte
Xg’ and neutrophil
Kx antigen
typing are shown for each individual
tested.
NT. not tested.
removed
titer
by
of
centrifugation
anti-Kx
twofold
dilutions
37#{176}C.The
globulin
red
inverse
was
taken
washed,
of the
examined
were
highest
titer
ofanti-Kx
mediated
previously
tous
disease
above.
hemagglutination
neutrophils
were
controls.
quantitative
described4
sibling
at
anti-human
Normal
four
male
and
with
all family
and
bactericidal
using
neutrophils
in this
Kx-negative
tetrazolium
tests’
nitroblue
shunt
siblings
Kx-negative
Xg(a-)
i(x+
isolated
kindred
neutrophils,
with
and
were
from
normal
chronic
the
In addition,
performed
assessed
Kx
volun-
healthy
qualitative
Typing
1 , all four male siblings
granulomatous
disease
However,
neutrophils
from
male
the
last
born
obtained
male
Neutrophil
Since
previously
from
RESULTS
Cell
healthy
globulin
tests on all erythrocyte
tive.
There
were
no unusual
samples
reactions
were negawhen
the
erythrocytes
were examined
for the blood group
antigens
described
in the methods,
and
none
of these
results
suggested
nonparentage.
The
crythrocytcs
from
type
all family
members
were
Kx-positive
K-k+,
Kp(a-b+).
No weak or unusual
with
Kell
system
antisera
were
observed.
who
lacked
on two
male
did
Kx when
occasions.
siblings
Therefore,
the
at the Xk locus.
not
have
a
all
mother
had
separate
The three
Kx-positive
must
also
be het-
Function
the
Kx-negative
history
of
Typing
None of the family
members’
sera contained
atypical blood
group
antibodies
and
direct
anti-human
in generation
five of the
sibling
in generalacked
Kx
obtained
history
suggestive
of chronic
granulomatous
and whose
qualitative
nitroblue
tetrazolium
neutrophils.
erozygous
nitro-
members.
Blood
CARRIER
As shown
in Fig.
tion II with chronic
on their
neutrophils.
clinical
disease
Xq(a.)
Ks-
granuloma-
one
on neutrophils
as
Xg(o+)
Ks-
CGD
remaining
reduc-
myeloperoxidase-
activity
neutrophils.
were
tetrazolium
activity,
Xg(o.)
Ks-
#{149} ‘c,
NORMAL
absence
only
been
chronic
granulomatous
phil function
of the
Red
Xg(o+)
Ks-
test
was
normal,
also
samples
were examined
monophosphate
iodination,
the
for 30 mm
Xq(o-)
i(+
Neutrophil
serial
as described
producing
antibody.
as positive
consumption.
hexose
teers,
still
residual
incubating
polyspecific
scored
dilution
Xg(o-)
Kx+
Function
Oxygen
tion,
then
22#{176}C).The
by
K, erythrocytes
hemagglutination
concurrently
Neutrophil
blue
cells
Xg(o+)
Xx-
LII
10 mm,
determined
with
and
as the
g,
was
of the supernate
added,
The
(1000
antibody
Xg(os)
NT.
and Kell
reactions
Both
I had Xg(a+)
red blood cells,
eight
male
siblings
in generation
parents
but only
II had
quantitative
nitroblue
of
single
neutrophil
recognized
disease,
male
neutrophils,
but
repeated
infection
defect,
tetrazolium
Kx
in
antigen
patients
has
with
we studied
the neutroin this family
who had
who
did not have
(Table
I ). Neither
as measured
by
test,
hexose
a
a
the quantitative
monophosphate
shunt,
or iodination,
nor a kinetic
defect,
as measured
by continuously
monitored
oxygen
consumption
and
bactericidal
activity,
could
be demonstrated.
DISCUSSION
The
KelI
determined
blood
allelic
white,
blood
cells.
precursor
substance
group
is a system
antigens
present
of autosomally
on red, but not
It has been proposed
that Kx is a
that is converted
to the appropri-
she is genotypically
XgaXg.
The only female
sibling
in
the second
generation
had Xg(a + ) erythrocytes,
as
would be expected
since her father’s
erythrocytes
were
ate Kell antigens
by an enzyme
coded
for on the
autosome
determining
Kell type.2
In contrast
to the
Kell antigens,
Kx is normally
present
on both red cells
and granulocytic
white
cells, although
lesser
amounts
are detected
on red cells,
presumably
due
to the
conversion
of Kx to Kell
system
antigens.
It has
also
further
Xg(a+)
that the
Xg(a
on their
red blood
cells
mother
was heterozygous
+).
(Fig.
I), indicating
at the Xg locus, i.e.,
been
suggested
that
Kx
results
from
the
From www.bloodjournal.org by guest on June 11, 2017. For personal use only.
36
DENSEN
Table 1 . Neutrophil
Function
Qualitative
NBT
NBT
CGDt
92±2.7
91
±4.1
2.8±
1.0
reduction
monophosphate
(% increase
Pat,entt
reduction
(%increasenormalcontrol)
Hexose
AL.
Studies
Normal
(%NBT-positivePMN)
Quantitative
Fun ction
El
100
82.5
±
18.5
29.8
±
13.1
100
80.0
±
3.0
26.3
±
2.0
±
1,658
±
226
±
1.20
±
0.15
shunt
normal
control)
Iodination
(cpm/1O6PMN)
Oxygen
(MIO2/hr/1O6PMN)
Bactericidal
956
9.04
±
1.92
96.6
±
2.8
killed)
§Patient
7.27
0.43
89.6
for bactericidal
activity,
each test was performed
on the patient
chronic granulomatous
disease; NBT. nitroblue tetrazolium;
PMN.
CGD,
8,546
12
SEM.
±
tExcept
26,035
activity
(% bacteria
Mean
±
24,063
consumption
but not CGD is within
expression
Erythrocytes
of
an
and
Kx independent
4 siblings
with
polymorphonuclear
CGD
2-3
structural
white cells
another,
gene,
Xk.8
may possess
depending
on which
of
lytic
anemia.9”
Individuals
chronic
Kx from
both
cell
types
(chronic
granulomatous
disease,
McLeod
phenotype).
The X3k allele codes for
Kx on red cells but not granulocytes
(chronic
granulomatous
disease
phenotype),
while
the reverse
is true
for X4k (McLeod
phenotype).3
In the family
reported
here, all members
had Kx-positive
red cells. However,
five of the males
in the second
generation
had Kxnegative
granulocytes
and three
had Kx-positive
gran-
abnormal
ulocytes.
These
data
heterozygous
at the
lomatous
disease.
The
granulomatous
disease
genotype.
expression
The
on
demonstrated
affected
males
indicate
Xk locus
consistent
both
that
and
the mother
must
have
pattern
erythrocytes
must be
the X’k
of Kx antigenic
and
in these eight male siblings,
and
in other
families
having
either
in other
the X3k
or X4k allele,
indicates
that the independent
sion of Kx on these
two cell types
is not
due
expresto the
effect
that
modi-
of an independently
segregating
fies the expression
of Kx.
The results
of Kx and
Xga
gene
typing
were
analyzed
granulomatous
in
disease,3
result
from
the dysfunction
The rare patients
with
the
red and
are unaware
granulomatous
Kx positive.
phils
have
appear
to
X2k
or X3k
white
they
have
a disorder
At the
neutrophils
evidence
expression
of chronic
kindred
appears
to be
mild in that the four
recurrent
infections
affected
males
did not
until after the age of 5,
have all
overcome
survived
into their
their susceptibility
tion.4 Given
the
affected
brothers
relatively
with
30s, and appear
to
to recurrent
infec-
mild clinical
course
Kx-negative
neutrophils,
neutrophils
evidence
Xg(a-).
linkage
absence
of Kx from
neutrophils
is closely
with chronic
granulomatous
disease,
the
for the
family
is
this
Xk and
between
2. 1 04
score of
+
at
+
Xg.
the
The lod score calculated
Xk and Xg loci in this
a recombination
fraction
2.0 or greater
is customarily
of zero.
accepted
level of significance
for linkage
of X-borne
Individuals
of the X2k or X4k genotype
their
ically
surface
red blood
abnormal.
cells,
is associated
A lod
as the
characters.
lack Kx on
and their red cells are morphologThis
abnormality
of the red cell
with
a mild
compensated
hemo-
in the
we
investigated
the unaffected
male
sibling
with
Kxnegative
neutrophils
for the possibility
of a partial
defect
in the function
of his neutrophils.
We were
three
These
between
linkage
we
clinical
in this
unable
function.
were
for genetic
time,
but does not have
of chronic
granu-
All five males
with
KxXg(a+)
red cells, while
all
Kx-positive
provide
strong
to
oxidase.
have
both
present
the eight
male
siblings.
negative
neutrophils
had
with
data
believed
of a membrane
X2k
genotype
cells.3
geno-
of Kx has
X-linked
of any individual
with X-linked
chronic
disease
whose
neutrophils
have
been
That all persons
with Kx-negative
neutrochronic
granulomatous
disease
does
not
be true,
since
one
male
sibling
in this
kindred
has Kx-negative
either
clinical
or laboratory
relatively
experience
granulocytes
of the
type lack Kx on their granulocytes.
This lack
shown
a high
degree
of association
with
four
recognized
Xk alleles
occupies
the Xk
locus.
Thus,
the X’k
allele
codes
for Kx on both cell types
(normal
phenotype),
but X2k results
in the absence
of
x3k
times.
neutrophils.
2 SD of the mean for normal.
X-linked
granulocytic
of one
or on the
to identify
These
antigen
any
results
is not
defect
in neutrophil
demonstrate
that
required
function.
Our data are
tion that separate
genes
chronic
granulomatous
case,
then
neutrophils
suggesting
that
associated
presence
normal
of
neutrophil
consistent
with the interpretaon the X chromosome
code for
disease
and Kx. If this is the
the
normal
reflects
a
chronic
granulomatous
the recombination
did
for
oxidative
while
the
brother
recombination
with
Kx-negative
between
the
disease
and Xk loci. However,
not separate
the Xk and Xg loci,
the order
of these
three
genes
on the
X
From www.bloodjournal.org by guest on June 11, 2017. For personal use only.
Kx, CHRONIC
chromosome
Xk-Xg.
GRANULOMATOUS
DISEASE.
AND
Xg
37
may be chronic
granulomatous
It should
be noted that this interpretation
that
the neutrophils
amount
of Kx antigen
diseasedoes
not explain
the high degree
of association
(linkage
disequilibrium)
of chronic
granulomatous
disease
with
Kx-negative
granulocytes.
Other
possible
explanations
for the separation
of
Kx-negative
neutrophils
and chronic
granulomatous
disease
ing
must
Kx
be considered.
on
The
granulocytes
measures
the
standard
anti-Kx
procedure
is an
residual
for detect-
indirect
method
hemagglutination
antiserum
after
by the test granulocytes.
results,
the method
has
that
titer
it has
been
Although
giving
low sensitivity.
in
absorbed
level
on the
neutrophil
membrane
have
a reduced
for functional
results
full protection
from the disease.
Kx antigen
on neutrophils
from
as from
disease
are
ities.
reproducible
It is possible
this sibling
that is sufficient
activity
but which
is below the level of detection
by the
present
technique.
Another
possibility
is that
some
compensatory
process
operating
at the
phenotypic
well
a
of
individuals
needed
Kx-negative
rewarding.
with
of
the
neutrophils
granulomatous
among
these
descendants
may
of
prove
or
studies
of the
individuals
as
chronic
to distinguish
Evaluation
in partial
Further
normal
possibil-
males
with
to be particularly
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cytes.
BM:
N EngI
2.
Marsh
lomatous
Oxygen
I Med
WL,
dependent
298:659,
721,
microbial
#{248}yenR, Nichols
disease
and
the
Kell
killing
by
phago-
1978
ME,
blood
Allen
groups.
FH:
Chronic
Br I Haematol
method
for
carriers.
I Pediatr
granu-
8.
29:247,
syndrome,
3.
Marsh
WL,
phenotype,
and
Sang3l:356,
4.
tous
ME:
Kx
disease:
antigen,
Further
the
McLeod
studies.
Vox
Mandell
GL:
of “childhood.”
Am
Methods
Association
6.
Marsh
WL:
12:352,
1972
HD,
WL:
and
Wimer
Dail
the
BM,
WG:
chronic
83:77,
granulomatous
Chronic
Kell
granulomatous
blood
Marsh
groups.
WI,
Haematological
phenotype
of the
Kell
disease
and
female
1973
disease,
Birth
Taswell
changes
blood
HF,
Galey
associated
group
system.
the
Defects
McLeod
14:9,
1978
WR,
Evan
with
the
AP,
McLeod
Br I Haematol
36:2
19,
I 977
IA,
American
7. Ochs
Nichols
granulomatous
1976
Dilworth
disease
#{216}yenR,
chronic
5. Technical
sion
Marsh
9.
I975
detecting
and
of Blood
Scoring
Adults
with
I Med
63:233,
Procedures
(ed
Banks,
chronic
granuloma-
1977
7).
10.
Galey
Cooper
RA:
cyte.
Washington
DC,
reactions.
Iransfu-
SB,
Iinehan
McLeod
Igo
RP:
The
NBT
slide
test:
A simple
screening
42:575,
Van
and
Vox
WA,
BI:
phenotype
1979
AP,
electron
properties.
1 1 . Symmans
of hemagglutination
Evan
Morphology
I. Scanning
transport
1977
WR,
PS,
Shepherd
34: 1 52,
CS.
blood
WG,
and
Wimer
Mcleod
BM,
erythro-
electrolyte
and
water
WI,
R, Shohet
1978
Marsh
acanthocytosis
Kell
Dail
of the
microscopy
Sang
Hereditary
of the
Nice
physiology
group
Oyen
associated
system.
with
Br I Haematol
the
From www.bloodjournal.org by guest on June 11, 2017. For personal use only.
1981 58: 34-37
Kx: its relationship to chronic granulomatous disease and genetic linkage
with Xg
P Densen, S Wilkinson-Kroovand, GL Mandell, G Sullivan, R Oyen and WL Marsh
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