(AGAMMAGLOBULINEMIA):
REPORT
OF
3 CASES449
By ALVIN
B.
HAYLES,M.D.,#{176}f
AND BERNARD
F.
Rochester,
LTERATIONS in total serum proteins or in
various subdivisions of such proteins have been recognized for a long time. In recent years electrophoretic technics have replaced salting-out methods for
fractionat-ing proteins, partly because of precise
re-producibility of tile former procedure.
Changes in serum proteins in a variety of
clinical states were summarized by Stern
and Reiner,’ in 1946.
Of particular interest have been altera-tions in the gamma globulin of the serum proteins, for this fraction contains many of
the antibodies. Usually a depression of gamma globu!in is associated with
signifi-cant fluctuations in other components of proteins. Thus, in nephrosis the values for
serum albumin and gamma globulin are
decreased but those for the alpha and beta
globulins
are
increased.
It is a common
ob-servation
that
patients
who
have
nephrosis
are
prone
to
incur
infections,
especially
pneumococcic peritonitis.
Several reports have appeared concerning idiopathic hypoglobulinemia in infants and
children.27 The condition usually has been
characterized by disturbances in fluid bal-ance, edema, low blood pressure, lympho-penia, hypoproteinemia and, in some in-stances, increased susceptibility to infec-tions. Electrophoretic analyses of the serum proteins, when done, have shown virtual ab-sence of gamma globulin in addition to
From the Mayo Clinic and Mayo Foundation,
Rochester, Minnesota. The Mayo Foundation is a
part of the Graduate School of the University of
Minnesota.
(Received for publication May 25, 1954.)
#{176}Address: Mayo Clinic, Rochester, Minnesota.
+ Section of Pediatrics. #{176}#{176}Fellow in Pediatrics.
Section of Biochemistry.
GUNNAR
B.
STICKLER, M.D.,#{176}#{176}MCKENZIE, M.S4 Minnesota
rather pronounced reduction in albumin.5’6 Krebs,8 in 1946, noted the usual hypo-albuminemia in a patient with starvation but also found that gamma globulin was practically absent. After partaking of a wholesome diet, the patient returned to norma!; at this time, antibodies induced as a result of use of typhoid vaccine were demonstrable, whereas they were not be-fore.
In
the
past
two
years,
isolated
cases
of
idiopathic, probably congenital, deficiency in gamma globulin have been reported.
In May,
1952,
Bruton
and
associates9
pre-sented their observations on three boys, all approximately nine years of age, who had histories of multiple infections since
in-fancy
or
early
childhood.
Electrophoretic
analyses of the serum of these patients indi-cated a deficiency of gamma globulin. Bruton1#{176} treated one of these three patients
by means of monthly injections of 3.2 gm. of
gamma
globulin
and
reported
that
he
was
able to maintain the patient free of infec-tions. This was the first patient
recog-nized
to
have
isolated
absence
of
gamma
globulin. Bruton named the condition “agammaglobulinemia.”
Keidan
and
co-workers11
have
reported
on an 11-week-old girl who had generalized vaccinia and micrococcic (staphylococcic) septicemia after vaccination against
small-pox.
This
child
was
unable
to produce
anti-bodies
against
the
virus
of cowpox.
Electro-phoretic analysis of her serum proteins dis-closed an absence of gamma globulin.
Hyperimmune globulin was given but the
baby
died.
It
is the
purpose
of
this
publication
to
report
threecases
encountered
at the
Mayo
Clinic
in which
a decrease
in gamma
Normal
Aib
0(1 0(2
,8
-V
46.2%
14.3% 2.3.6 %
15.9%
0
Fic. 2 (Case 1). Distribution of serum proteins. Note absence of gamma globulin.
Aib is ‘y
Alb S9iL%
cx1 5.9%
cxZ 11.0%
/3 11.8%
-I 12.1%
Ftc. 1. Norma! distribution of serum proteins
as determined by the Tiselius method.
REPORT OF CASES
CASE 1 : A 5-month-old white boy was
ad-mitted to the hospital in October, 1952. Fre-quent watery stools had appeared 2 mos. prior to admission; later, the stools had become bloody. He had been hospitalized on two
occa-sions for bronchopneumonia, bilateral otitis media, purulent conjunctivitis and diarrhea. Antibiotic therapy had given only temporary
relief. The baby had not received vaccination against smallpox or other immunizing pro-cedures. The family history was
noncontribu-tory. Two siblings were living and well. The infant did not appear acutely ill on ad-mission. His temperature was 101.6#{176} F. He weighed 7,080 gm. Pronounced chemosis and
bilateral purulent conjunctivitis were present. A seropurulent nasal discharge and mild pharyngitis were noted. A loose cough was
present. The remainder of the physical ex-amination showed nothing abnormal except for mild metatarsus varus of the right foot.
Laboratory studies disclosed a value for
hemoglobin of 12.6 gm. per 100 cc. of blood.
Leukocytes numbered 5,400 per cmm. of
blood; differential count showed 34% lympho-cytes, 2% monocytes, 1% eosinophiis, 41% seg-mented neutrophils and 22% young neutrophils. Results of urinalysis were normal. Cultures
made from the throat, conjunctivae and stool showed no pathogens. Cultures of the blood produced no growth of organisms. A roentgeno-gram of the thorax showed nothing abnormal.
The patient was given penicillin and
di-hydrostreptomycin. His condition appeared to improve temporarily but 2 days after admission the temperature had increased to 104#{176}F. and
recurrence of frequent loose stools was noted.
Electrophoretic
study
of
the serum proteinsdisclosed almost complete absence of gamma globulin (Figs. 1 and 2) . At the time this de-termination was carried out, the significance
of the absence of gamma globulin was not
ap-preciated. The patient received a variety of
antibiotics without evidence of improvement.
The septic temperature continued; pneumonia
developed and extended rapidly. The child died 42 days after admission.
Necropsy revealed chronic suppurative pneumonitis with formation of abscesses. Cul-ture of material from the abscesses showed
abundant growth of Micrococcus pyogenes.
Multiple, non-specific, granulomatous lesions of
the liver were present, in one of which a few cocci were present.
CASE 2: A 9-year-old white boy was ad-mitted to the hospital in April, 1953, with a his-tory of fever, irritability, stiff neck and pro-gressive weakness of the right arm and leg of 7 days’ duration. He had been studied at the clinic in 1948, 1949 and 1951 because of re-current infections. No cause for these infections had been found.
The past history was characterized by
in-numerable infections since the age of 2 yrs. These infections had included many colds, at-tacks of purulent otitis media, tonsillitis, bron-chitis, ethmoiditis, conjunctivitis, stomatitis, proctitis, inflammatory arthritis and pleuritis. Diarrhea had been present on occasions.
Measles,
in 1952,had
been
followed
by
deaf-ness.
The tonsils and adenoids had beenre-moved in 1952. The patient had been treated with chemotherapeutic or antibiotic agents for
the greater part of the past 7 yrs. The family history was noncontributory. Immunization for
diphtheria, tetanus, and pertussis had been done in infancy. Vaccination against smallpox
at the age of 18 mos. had been successful, with
formation of a good scar.
451
Examination on admission disclosed an acutely ill, malnourished, irritable boy who was deaf. The oral temperature was 102.6#{176} F. Nuchal rigidity was pronounced and there was weakness of the right arm and leg.
The value for hemoglobin was 10.4 gm.
Erythrocytes numbered 3,260,000 per cmm. of
blood. The leukocyte count was 9,100, with 30% lymphocytes, 4% monocytes and 66%
trophils. Results of urinalysis were normal. The erythrocytic sedimentation rate was 23 mm.
during the 1st hr. (Westergren method).
Roent-genograms showed thickening of the mucous membranes of both antrums but nothing ab-normal in the skull, mastoids or thorax. Ex-amination of cerebrospinal fluid revealed 160
lymphocytes and 526 polymorphonuclear leuko-cytes per cmm. It contained 200 mg. of protein per 100 cc.; the value for sugar was 28 mg and that for chlorides was 637 mg. Smears and cultures of the cerebrospinal fluid showed no organisms. Serum proteins measured 4.6 gm. per 100 cc., with values for albumin and globu-lin being 3.7 and 0.9 gm. respectively (Kings-ley method) . Six wks. later, the total serum
protein was 5.6 gm., with 4.2 gm. of albumin and 1.4 gm. of globulin. Electrophoretic analy-sis disclosed virtual absence of gamma globulin and slight decrease in the other fractions of protein (Fig. 3).
On admission to the hospital, the patient began to receive chlortetracycline (aureo-mycin) and dihydrostreptomycin. Four days
later, 2 cc. of human pooled gamma globulin,
containing 0.3 gm. of gamma globulin, as pro-vided by the American Red Cross was given intramuscularly. Immediately after this injec-tion the oral temperature increased from 102#{176} to 105#{176}F. but within 4 hrs. the temperature
decreased to normal and remained so
through-out the period of observation. Injection of 2
cc. of gamma globulin was repeated on the 5th, 6th and 10th hospital days. Use of all medications was discontinued on the 20th day. Recovery of the patient was rapid and was
marked by improvement in appetite, decrease in irritability and return of strength in the arm
and leg.
Serum proteins were analyzed by paper
elec-trophoresis and by the method of Tiselius after the administration of gamma globulin. A small
but definite increase in gamma globulin could be detected on the paper electrophoretic pat-tern but the increase was not enough to be
Case 2
Alb 56.4% 01 11.2% 02 15.2%
13 17.2%
-V 0
FIG. 3 (Case 2). Distribution of serum proteins. Note absence of glmma globulin.
detected by the Tiselius technic.
The patient was dismissed 4 wks. after
ad-mission. Arrangements were made for ad-ministration of 5 cc. of gamma globulin every 2 wks. In spite of this, he continued to have frequent infections. An increase in the amount of gamma globulin to 10 cc. every 2 wks. did not alter the frequency or severity of the infec-tions nor did it appreciably change the elec-trophoretic pattern of the serum as deter-mined 2 wks. after an injection. It was duff!-cult to ascribe benefit to the patient as a result of the use of gamma globulin, although its ad-ministration was continued at a dosage of 5 cc.
every 2 wks. He gained weight and strength and attended a school for the deaf, but he con-tinued to have almost constant infection.
Information forwarded through the courtesy
of Drs. Kenneth Fijon and Frank Tanner, of Lincoln, Nebraska, indicates that generalized edema, high fever and progressive dyspnea appeared in this patient, and he died. Necropsy disclosed generalized anasarca, diffuse myo-carditis, cerebral edema, chronic hemorrhagic pachymeningitis and meningitis, diffuse myo-sitis, and fatty degeneration and hypertrophy of the liver.
CASE 3: A 9-year-old white boy was admitted to the hospital in May, 1953, because of
meningitis of 5 days’ duration. The past history
was characterized by an unusual number of illnesses, beginning with osteomyelitis when the patient was 18 mos. old. The episode of osteo-myelitis was followed by many febrile illnesses associated with pain, limitation of motion and swelling of the joints. Relief was obtained by treatment with antibiotics. Poliomyelitis at the
age of 7 yrs. was followed by a “lung
A1E
#{176}1
/3
7
Case 3
i 02 13
65.3% 7.3% 13.7% 13.7%
0
FIG. 4 (Case 3). Distribution of seruni proteins. Note absence of gamma globulin.
Measles at the age of 8 and chickenpox when he was 9 were uncomplicated. In addition, the patient had been subject to infections of the
upper part of the respiratory tract and
pyo-derma. The family history was noncontributory. The patient had received no immunizations or
vaccination against smallpox.
Physical examination on admission revealed a semicomatose, dehydrated boy. The oral tern-perature was 98.8#{176}F. Nuchal rigidity was
pro-nounced.
The value for hemoglobin was 11.9 gm. Erythrocytes numbered 4,010,000 and leuko-cytes 20,700. Differential count showed 12% lymphocytes and 88% neutrophils (44% band forms). The cerebrospina! fluid contained 17,760 polymorphonuclear leukocytes and 171 lymphocytes per crnm. Protein in the cerebro-spinal fluid measured 100 mg., while chloride measured 646 mg. and sugar 15 mg. Cultures of the fluid disclosed the presence of Hernoph-ilus influenzae. Total serum protein was 4.6 grn., with values for albumin and globulin being 3.9 and 0.7 grn. respectively. Electro-phoretic analysis of the serum showed almost complete absence of gamma globulin but nearly normal proportions of other protein frac-tions (Fig. 4).
A sulfonamide compound (gantrisin), chlor-tetracycline (aureornycin) and dihydrostrepto-mycin were given. On the 3rd hospital day, 4 cc. of gamma globulin (American Red Cross) was given intramuscularly. During the next few days he was given 12 cc. of gamma globulin in divided doses. On the 5th hospital day the patient was alert and on the 7th day was asymptomatic. Administration of all
medica-tions
was
discontinued
on the
8th
day
and
he
was dismissed 15 days after admission.
He was next seen 6 wks. later, at which time
he was well except for a few infected insect bites. Serum proteins measured 5.62 grn. per 100 cc., with 4.37 gm. of albumin and 1.25 gm. of globulin. Gamma globulin, as deter-mined by the Tiselius method, could not be detected. Therefore, he has been given 4 cc. of gamma globulin each month. He has remained
ill good health.
COMMENT
The recognition of agammaglobulinemia
in three patients at the clinic during a pe-nod of seven months suggests that the con-dition has escaped detection in the past
because electrophoretic analysis of serum proteins was not done. It may be that be-fore the advent of chemotherapeutic and antibiotic agents, patients who had this
condition did not live long enough to
estab-lish a history of repeated infections. Wide-spread use of these agents and increased
awareness of the condition undoubtedly will result in the diagnosis of many more cases in the future.
We have found paper electrophoretic
analysis of the serum proteins, as described by Durrum,12 to be a satisfactory method of study. This procedure has the advantage
of requiring only a small amount of blood; the technic is relatively simple and little
equipment is necessary.
The method used at the clinic consists of the application of two microliters of serum transversely at the middle of a strip of Whatman No. 2 filter paper measuring 1 by
29 cm. The ends of the paper are then immersed in electrode chambers, each con-taining a veronal buffer solution of 0.0416 ionic strength at pH 8.6. The paper is
moistened over its entire length and a full-wave rectified current of 0.5 milliamperes is passed through the paper. Frequent adjust-ments of the voltage by means of a variable
voltage transformer are necessary to main-tain the current at a constant rate. After 120 minutes, the paper is removed, dried at
1100 C. for 10 minutes and stained by im-mersion for five minutes in a 0.1% solution
which has been added 10 gm. of mercuric chloride per 100 cc.
The
excess
of
dye
is
removed by washing with 2% acetic acid.The paper is washed finally with 0.5%
sodium acetate, which maintains the dye at
a sufficiently high pH to provide a deep
blue color.
By this technic, differences in the color bands representative of the various serum proteins are easily recognized. Deficiency of gamma globulin in two of our patients
was detected by this method and con-firmed by the technic of Tiselius. Experi-ence has shown that it is possible to demon-sb-ate small amounts of gamma globulin by paper electrophoresis while simultaneous determinations by the Tiselius method show no evidence of gamma globulin.
We
were
unable
to reproduce
the
changes
in the
Tiselius
pattern
of
the
serum
after
the administration of gamma globulin such
as Bruton
did
in
his patient. We attemptedthis study in only one instance (Case 2).
As already indicated, injection of 10 cc. of
gamma globulin every two weeks produced
no demonstrable gamma globulin in the
serum of this patient as indicated by the
Tiselius pattern but did produce evidence of a slight increase in gamma globulin in
the paper electrophoretic pattern. Since this
patient
did
not
respond
clinicafly
to the
use
of gamma globulin in the same manner as did Bruton’s patient, there must be some as yet unexplained abnormality in the utiliza-tion, destruction
or excretion
of protein
in
our patient.The possible relationship between gamma
globulin and lymphocytes has been the sub-ject of some discussion.13 Two of our pa-tients had relatively low lymphocyte counts at some time during observation. In both instances (Cases 1 and 3) the counts were
made
when
the
patients
were
acutely
ill
and
itis difficult
to attach
any
significance
to
them.The
cause
of
agammaglobulinemia
re-mains
obscure
but
we
are
in
agreement
with
Bruton
and
associates
that
the
con-dition
may
well
represent
a congenital
de-fect in formation of gamma globulin
analogous to the congenital lack of specific
proteins seen in hemophilia and
afibrino-genemia. It is interesting that all patients reported thus far have been males except for the infant described by Keidan and co-workers.
The term “agammaglobulinemia”
im-plies complete absence of gamma globulin in the serum. This absence is not actually complete but no particular advantage is to
be gained by changing the name to the
more accurate but even more cumbersome term “hypogammaglobulinemia.”
SUMMARY
A study
has
been
made
regarding
three
boys who had agammaglobulinemia and
who were seen at the Mayo Clinic during a period of seven months. In each there was a
history of frequent infections since infancy.
All three patients demonstrated relatively normal serum proteins except for greatly decreased amounts of gamma globulin. The deficiency in gamma globulin was demon-strated by use of paper electrophoretic
analysis
of
serum proteins.One of the patients, an infant, died while
under treatment at the clinic. The second
patient
continued
to
have
frequent
infec-tions in spite of the administration of
gamma
globulin
at
frequent
intervals;
he
died
later
at
home.
The
third
patient
has
been maintained in good health by monthly injection of gamma globulin.
REFERENCES
1. Stern, K. G. and Reiner, Miriam:
Electro-phoresis in medicine. Yale
J.
Biol. &Med., 19:67-99, 1946.
2. Thompson, W. H., McQuarrie, Irvine and Bell, E. T.: Edema associated with hy-pogenesis of serum proteins and atrophic changes in the liver, with studies of water and mineral exchanges.
J.
Pediat., 9:604-619, 1936.3. Schick, Bela and Greenbaum,
J.
W.: Ede-ma with hypoproteinemia due to454
4. Hertzog, F. V. and Faust, 0. A.: Edema
associated with temporary idiopathic hypoproteinemia.
J.
Pediat.,36:641-645, 1950.
5. Wyngaarden,
J.
B., Crawford,J.
D.,Chamberlin, H. R. and Lever, W. F.: Idiopathic hypoproteinemia: Report of a case of transient edema, depression of plasma albumin and gamma globulin with eosinophilia. PEDIATRICS, 9:729-734, 1952.
6. Bound,
J.
P. and Hackett, W. R.: Idio-pathic hypoproteinemia, oedema and amino-aciduria in an infant. Arch. Dis. Childhood, 28: 104-109, 1953.7. Fanconi, G. and Waligren, A.: Lehrbuch der P#{228}diatrie. Basel, Switzerland, Benno Schvabe & Co., 1950, p. 173.
8. Krebs, E. G.: Depression of gamma glob-ulin in hypoproteinemia due to
ma!-nutrition.
J.
Lab. & Clin. Med., 31:85-89, 1946.9. Bruton, 0. C., Apt, Leonard, Gitlin, David
and Janeway, C. A.: Absence of serum gamma globulins. (Abstr.) Am.
J.
Dis. Child., 84:632, 1952.10. Bruton, 0. C.: Agammaglobulinemia.
PE-DIATRICS, 9:722-727, 1952.
11. Keidan, S. E., McCarthy, K. and Haworth,
J.
C.: Fatal generalized vaccinia with failure of antibody production and ab-sence of serum gamma globulin. Arch. Dis. Childhood, 28:110-116, 1953. 12. Durrum, E. L.: A microelectrophoreticand microionophoretic technique. Am. Chem. Soc., 72:2943-2948, 1950. 13. Huth, F.: Lymphocyten system, wachstum
