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Hyper-M-immunoglobulinemia in children with bronchial asthma
222
August 1977 TheJournalofPEDIATRICS
Hyper-M-immunoglobulinemia in children with bronchial asthma Radial immunodiffusion was used to measure the concentrations of lgG, IgA, and IgM in the sera of 224 asthmatic children, ages 6 months to 14 years. IgM was greater than two standard deviations above agematched normal values in 51% of these asthmatic subjects, lmmunoglobulin profiles were repeated up to four years later in 29 individuals with elevated l g M and increased IgM synthesis was sustained in 14 of these asthmatic subjects. Serum lgE concentrations were elevated in 23 o f 35 asthmatic patients. There was no statistical difference in mean IgE between asthmatic children with normal or increased IgM. The distribution of lgE in each group was similar. Only 10 of 224 subjects had an IgA concentration less than 2 SD below age-matched control subjects. The significance and possible mechanisms of hyperM-immunoglobulinemia are discussed.
Ming S. Lin, M.D., Ph.D., Bruce S. Rabin, M.D., Ph.D., Ralph LaNeve, M.D.,
and Philip Fireman, M.D.,* Pittsburgh, Pa.
ASTHMA is a syndrome of several etiologies characterized by bronchospasm, increased bronchial mucous secretions, and bronchial edema. The immunologic development of asthma is often mediated by reaginic (IgE) antibodies; however, in many patients reagin-mediated immediate hypersensitivity cannot be demonstrated. Even though several investigators have reported an elevation of mean serum IgE values in asthmatic individua!sl 1 it is not unusual for individual patients with severe bronchial asthma to have normal or low serum IgE, 2 whereas increased concentrations of serum IgE may be present in patients without any asthmatic symptoms? Thus, measurement of serum IgE concentrations cannot be used alone to assess the severity of asthma. In an attempt to uncover immunologic mechanisms which may be operative in severely asthmatic BRONCHIAL
From the Departments of Pediatrics and Pathology, University o f Pittsburgh School of Medicine, and Children's Hospital of Pittsburgh. Supported by United States Public Health Service Grants HD-02662 and HD-07850. Presented in part at the annual meeting of the American Academy of Allergy, February, 1975. *Reprint address: Children's Hospital, 125 Desoto St., Pittsburgh, PA 15213.
Vol. 91, No. 2, pp. 222-227
patients, several investigators have demonstrated inconsistent changes from normal in serum IgG, IgA, or IgM concentration in asthmatic patients. ~-7 Several factors could contribute to this: (1) the number of patients studied was rather small, (2) the data obtained were not analyzed by age and sex, and (3) the criteria used to select the subjects were different in each study. The present study was undertaken to evaluate serum immunogl0bulin concentrations and calculate the prevalence of immunoglobulin abnormalities in children with recurrent asthma. The results were analyzed by age and sex of the patients and then compared to those of normal individuals. The study demonstrated that many asthmatic children who have normal or elevated serum IgE concentrations may have another immunoglobulin abnormality, usually an elevation of serum tgM values.
MATERIALS AND METHODS Study population. The subjects evaluated in this study consisted of 224 consecutive asthmatic patients, aged 6 months to 14 years, referred to the allergy clinic of Children's Hospital of Pittsburgh and evaluated by one of the authors (M.S.L.) The diagnosis o f asthma was established on the basis of history and physical examination. The history and pfiysical examination elicited specific
Volume 91 Number 2
Hyper-M-immunoglobulinemia
223
Table I. Descriptive statistics for IgM for patients and normal subjects classified by age and sex
Normal
Asthmatic t-Statistic
Age
Sex
N*
3-6 mo
M &F
7
7-12 mo
M&F
6
13,24 mo
M &F
11
25-36 mo
M &F
10
3-4 yr
M
9
3-4 yr
F
8
5-6 yr
M
10
5-6 yr
F
9
7-8 yr
M
9
7-8 yr
F
7
9-11 yr
M
13
9-11 yr
F
9
12-14 yr
M
13
12-14 yr
F
6
GM'~
N
43.7 (28.8, 66.5) 53.6 (25.1, 114,7) 67.3 (35.8, 126.5) 54.5 (26.3, 113.4) 77.4 (30.5, 196.3) 64.1 (23.4, 175.8) 62.2 (25.8, 150.0) 94.8 (37.8, 190.2) 87.1 (34.2, 186.8) 65.3 (21.4, 180.1) 66.8 (30.8, 144.9) 69.0 (32.3, 147.6) 88.1 (36.4, 213.3) 64.0 (39.4, 104.7)
5 9 36 15 8 13 18 13 15 10 20 10 11 5
GM
t%
93.4 (25.2, 345.8) 100.5 (37.8, 266.7) 116.4 (49.2, 275.4) 133.4 (64.1, 277.3) 117.8 (46.7, 297.2) 157.8 (76.6, 325.1) 130.6 (47.2, 361.4) 143.5 (68.4, 301.3) 120.7 (45.7, 391.3) 172.2 (76.2, 389.0) 126.8 (57.7, 278.6) 156.3 (91.6, 266.6) 123.9 (76.2, 423.6) 152.1 (65.2, 354.8)
--3.17w --2.65 N --3.90w --5.99w --1.86
--4.79w --3.87w -2.34 82 -1.61 -4.15w --4.60w --5.47w
-2.5311 -4.23w
*N = number of observations. tGM = geometric mean = antilog x, where g is the mean of logarithm of IgM. The two values appearing below the geometric mean are, respectively,antilog (g 2 SD) and antilog (x+ 2 SD) where SD = standard deviation of logarithm of lgM. :~The t-statistics were obtained from a comparison of the mean logarithm of lgM for normal subjects and patients using comparable age-sex classes. w < 0.005. prP< 0.01.
]1P < 0.05. information regarding recurrent asthmatic symptoms and signs, including more than six previous episodes of wheezing, cough, or exercise-induced bronchospasm which were relieved by theophylline or sympathomimetic medications. None of the patients studied was receiving corticosteroid therapy and none had received hyposensitization therapy at the time the first blood sample was drawn. Normal sera were obtained from venous blood of healthy infants and children attending the general pediatric practice of one of the authors (R.L.) The detailed medical background of each donor was evaluated and the serum was included in the study only if there was no history of frequent infections, allergy, or autoimmune diseases. The normal subject group consisted of 75 boys and 51 girls. Serum samples were collected by venipuncture and stored at - 2 0 ~ C until analyzed. No sodium
azide or other bactericidal agents were added to the serum. Informed consent was obtained from all subjects; the research was approved by the H u m a n Right Committee of the Children's Hospital of Pittsburgh. Q u a n t i t a t i o n of serum i m m u n o g l o b u l i n s . Serum concentrations o f IgG, IgA, and IgM were measured by radial immunodiffusion using quantitative i m m u n o g l o b u l i n plates (Meloy Laboratories, Springfield, V a . ) ? " Sera were collected from the normal subjects and from 189 patients who were evaluated between 1971 and 1974; o f these 112 were male and 77 were female. The same immunoglobulin reference standard was used throughout the study. Serum IgE concentrations, in addition to serum IgG, IgA, and IgM, were measured in 35 asthmatic patients who were seen between 1974 and 1975; of these 24 were male and 11 were female. IgE was quantified by
224
Lin et al.
400
The Journal of Pediatrics August 1977
1
1
1
1
1
1
(
1
1
1
1
[
== r ~- 3 0 0
=,
200
<[ z
z_
200
A--
-
~
IOO
9
H
bJ H
I00 4/12 8/12
I
2
3
4
5
6
7
8
9
I0
II
12
13
14
AGE (YEARS)
Fig. 2. Serum IgM concentrations for male asthmatic children, Similar graphic representation as in Fig. 1. 0 4 / 1 2 8/12
I
2
3
4
5
6
7
8
9
I0
II
12
13
14
AGE ( Y E A R S |
Fig. 1. Serum IgM concentrations for female asthmatic children. The shaded area represents the range of normal serum IgM concentration for that age, and the continuous line in the shaded area represents the geometric mean of the normal IgM. radioimmunoassay with kits purchased from Pharmacia AB, Uppsala, Sweden. Statistical analysis. The serum concentrations of various immunogl0bulins, IgG, IgA, IgM, and IgE were converted to their logarithms for statistical analysis/~ For the normal children and the asthmatic patients the mean (x) and the standard deviation of these transformed IgG, IgA, IgM, and IgE values were determined for the various age and sex groups. Detailed results for IgM are presented in Table I which shows the antilog (x), antilog (x - 2 SD), and antilog (x + 2 SD). Corresponding age and sex groups for normal subjects and patients were compared on the basis of the mean logarithm values using the tstatistic. These comparisons were preceded by assurance that the variances of the corresponding groups were similar. Such conditions were met in all instances. Limits for normal values were considered to be within 2 SD of the antilog of the geometric mean. RESULTS Quantitative serum immunoglobulins, IgG, IgA, and IgM were determined in 224 patients with bronchial asthma. Their ages ranged from 6 months to 14 years: 136 were boys and 88 were girls. Among the total 224 patients we have studied, only 8.8% had values of all three
immunoglobulin classes within 1 SD of the mean and 30.8% within 2 SD. Serum IgM concentrations were elevated in many asthmatic patients and no individual value was below 2 SD of the geometric mean (Figs. 1 and 2). Of the female patients, 40% had IgM levels within normal limits; the remainder were above 2 SD of the mean (Fig. 1). In male patients 54% h a d normal serum IgM, concentrations the rest (46%) had elevated IgM levels (Fig. 2). In the 224 patients we have studied, more than half (51%) of the patients had elevated serum IgM concentrations. In Table I statistical comparison of IgM levels in normal and asthmatic subjects by age and sex showed significant differences in 12 of the 14 groups studied. To determine whether increased IgM synthesis was sustained~ immunoglobulin profiles have been repeated in 29 patients with increase d IgM concentrations. They were divided into four groups on the basis of time interval after the initial immunog!obulin determinations. The results (Fig. 3) indicated that the serum IgM Concentrations decreased during subsequent follow-up examinations in all but four of the asthmatic patients who had hyper-Mimmunoglobulinemia, but 14 patients still had increased serum IgM values. To study the relationship between the IgE and IgM values, IgE was quantified in 35 patients seen between 1974 and 1975. Elevated IgE was found in 23 patients. These patients were divided into two populations; one with normal IgM and the other with increased IgM. The geometric mean values of IgE for the patients with elevated IgM values was 347.1 I U / m l and that of the
Volume 91 Number 2
Hyper-M-immunoglobulinernia
225
3.0
Au -~
2.5
2.01 a E
\
Z
E E
1.5
-~
1.0
5
0.5
i
!
|
A
Initial
1-3 mos, later
Initial
4-12 mos.
later
,I Initial
|
|
|
1-2 yrs. later
Initial
2-4 yrs. later
,
Fig. 3. Serum IgM levels up to four years after initial determination and grouped on the basis of the time interval as shown on the abscissa. The ordinate represents the ratio between the patients' serum IgM and the highest normal serum IgM for that age and sex. Ratio greater than 1.0 indicates hyper-M-immunoglobulinemia; ratio smaller than 1.0 means normal serum IgM. Patients with sustained increased IgM levels are indicated by the open circle. The n value for this study was 29. patients with normal IgM was 219.0 lU/ml. The differonce between these two groups was not statistically significant (Fig. 4). When these 35 asthmatic patients were divided into four groups (Table IX) according to the frequency of asthmatic attacks that were observed during the next 12 months and without prior knowledge of the serum concentrations of IgE and IgM, the following was observed: the probability of frequent asthmatic attacks in a patient with increased serum IgE and IgM concentrations was 0.86 (6 out of 7 patients), in patients with elevated IgM values alone was 0.57 (12 out of 21 patients), and in patients with increased IgE concentrations was 0.21 (3 out of 14 patients). Serum IgG concentration was normal in the majority of the patients. In the female asthmatic patients, 14.0% had IgG higher than 2 SD and none had IgG deficiency. Similarly in the male patients, 15.0% had high and 3.5% had low IgG. Of the entire population studied, 14.8% of the asthmatic patients had elevated IgG and 2.1% had IgG deficiency. Statistical comparison of IgG levels by age and sex showed significant differences in only one of the 14 groups studied.
Serum IgA concentrations were normal in most asthmatic patients. In the female asthmatic patients, 20.7% had elevated IgA concentrations and 6.5% had serum IgA deficiency. In the male asthmatic patients, 21.4% had elevated serum IgA concentrations and 3.5% had IgA deficiency. As a group, 21% of the asthmatic patients had IgA values higher than 2 SD and 4.7% had low IgA concentrations. Statistical comparison of IgA levels by age and sex showed significant differences in only 3 of the 14 groups studied. DISCUSSION An immune response which depends on the presence of IgE and the balance between IgE and other specific antibodies has been postulated to play a central role in the pathogenesis of certain varieties of bronchial asthma?, ~A quantitative evaluation of serum immunoglobulins in bronchial asthma could, therefore, provide a better understanding of the humoral antibody system in this disease. In the first stage of our studies, serum concentrations of IgG, IgA, and IgM were measured in 189 asthmatic children. These data differ from those reported by other
226
Lin et al.
The Journal of Pediatrics August 1977
~1000 .
E
IlO00 .
00
.tO0.
9
i
i 12
9
AGE OF PATIENTSWITH HYPER IgM ( in years)
AGE OF PATIENTSWITH NORMAL IgM
( in years ]
F~. 4. Comparison of serum IgE levels at different ages in asthmatic subjects with hyper-IgM on left and in asthmatic subjects with normal IgM on right. Table U. Comparison of serum IgM and IgE levels with the frequency of acute asthmatic episodes in asthmatic children-~176
Frequency of attacks At least 1/wk:~ One to 3/mo Less than 1/mo Asymptomatic~
lgM( ~,)* IgE( ~ )
I
lgM(+ ) IgE(Nff
lgM(N) IgE( +)
12 5 1 3
2 0 1 5
6 0 0 1
I
IgM(N)lgE(N) 3 2 1 8
*(+) Indicatesincreasedserumlevel. t(N) Indicatesnormalserum level. :~Thereis no statisticalevidencebased on the chi square statisticsfor associationbetweenchangesin IgM and changesin IgE in those two groupsof patients. investigators who reported no deficiency or increase in immunoglobulins A, G, and M in patients with bronchial asthma?. 6. 7.11 Since the number of patients studied in these previous investigations was rather small and the results for each immunoglobulin class were not analyzed by age and sex, our data cannot be compared to data reported in the studies. Defective immunologic surveillance as a consequence of immune deficiency has been suggested to play an essential role in the pathogenesis and development of asthma and it has been reported that many forms of allergy are more common in patients with immunodeficiency than in healthy populations. 12 ~3 In the present study, data on individual serum immunoglobulin concentrations in 224 asthmatic children were examined for
the prevalence of selective immunoglobulin deficiencies and, more importantly, for interrelationships between immunoglobulin classes. The prevalence of selective immunoglobulin deficiency was found much less frequently than that of selective hyperimmunoglobulinemia. The prevalence of selective IgG, IgA, and IgM deficiency was 2.1%, 4.7%, and 0%, respectively. On the contrary, hypergammaglobulinemia, especially an increase in the IgM values was a rather common phenomenon ~n the children with bronchial asthma. The prevalences of increased serum IgG, IgA, and IgM concentrations were 15%, 21%, and 51%, respectively. This observation seriously challenged the previous speculation that a selective immunoglobulin deficiency, particularly of IgA, would result in a defect in eliminating the offending
Volume 91 Number 2
antigens, which in turn would stimulate the host to produce various i m m u n e substances such as IgE. Although h y p e r - M - i m m u n o g l o b u l i n e m i a was a common p h e n o m e n o n in children with bronchial asthma, the increased IgM synthesis was not sustained at the same level in most asthmatic patients and in fact returned to normal serum concentration in 15 of 29 asthmatic patients re-evaluated one month to four years later (Fig. 3). Nevertheless, four of these 29 patients sustained increased IgM synthesis; these four patients had more severe symptoms in spite of appropriate m a n a g e m e n t and required more medication in comparison to those patients in whom IgM synthesis had decreased. There were few significant differences in the serum levels of IgG and IgA between asthmatic and normal children when the various age and sex groups were compared; these data were similar to those observed by several investigators, 1' * 7 but different from those reported by Collins-William and associates? The m e a n IgE values in asthmatic children with normal IgM concentration were similar to those with increased IgM values and the distribution of IgE in each group was also quite similar (Fig. 4). Furthermore, the probability tha t an asthmatic patient with normal serum IgM concentration would have an elevated serum IgE value was 0.70, whereas the probability that those with increased IgM concentrations would have an elevated IgE value was 0.68. These observations suggest the probability that a patient with h y p e r - M - i m m u n o g l o b u l i n e m i a would have an elevated serum IgE value was not greater than for those with normal IgM concentrations. It may be important to point out that the sum of the probability of frequent attacks in patients with elevated IgM alone (0.57) and elevated IgE alone (0.25) was 0.82, which was close to, but lower than, that of the patients with elevated serum IgM and IgE concentrations (0.86). This suggested that the presence of at least two distinct pathologic processes affecting the respiratory tract might result in a clinical situation worse than either condition alone. Since hypergammaglobulinemia has been previously described in patients with frequent infections, "-1'~ and since viral infections have been reported as precipitants of asthmatic attacks in children, ~7-1~'this study may suggest that elevated IgM levels were due to repeated infection with different strains of viruses. As a result, specific clones of lymphocytes in response to various antigenic stimulation began to proliferate and accordingly more IgM was synthesized. The offending antigens could be either the antigens which initiated the allergic response or microorganisms which had infected the patients. The latter possibility is currently undergoing investigation in our laboratory and clinic.
Hyper-M-immunoglobulinemia
227
REFERENCES
1. Johansson SGO: Raised levels of a new immunoglobulin class (IgND) in asthma, Lancet 2:951, 1967. 2. Berg T, and Johansson SGO: IgE concentration in children with atopic diseases, Int Arch Allergy 36:219, 1969. 3. Aas K" The biochemical and immunological basis of bronchial asthma, Springfield, IL, 1972, Charles C Thomas, Publisher Chapter 13. 4. Collins-Williams C, Lamenza C, and Kokubu H: Deficiency of IgA in serum and respiratory secretions, Can Med Assoc J 99:1069, 1968. 5. Collins-Williams C, Tkachyk SJ, Toft B, Generoso L, and Moscarello M: Quantitative immunoglobulin levels (IgG, IgA and IgM) in children with intractable asthma, Ann Allergy 25:177, 1967. 6. Palma-Carlos AG, and Palma-Carlos ML: Immunoglobulins IgG, IgA and IgM in allergic diseases, Acta Allergol 26:161, 1971. 7. Frouchtman R, Vinas J, Rodriguez JL, and Garcia YP: Immunoglobulins in bronchial asthma, Rev Clin Esp 122:323, 1971. 8. Mancini G, Carbonara AO, and Heremans JF: Immunochemical quantitation of antigens by single radial immunodiffusion, Immunochemistry 2:235, 1965. 9. Leonardy JG, and Peacock LB: An evaluation of quantitative serum immunoglobulin determinations in clinical practice, Ann Allergy 30:378, 1972. 10. Sokol RR, and Rohlf F J, editors: Single classification analysis of variance, in: Biometry, San Francisco, 1969, W H Freeman & Co, p 220. 11. Radermecker M, and Rose B: Bronchial asthma, in Samter M, editor: Immunological Diseases II, Boston, 1971, Little, Brown & Company, p 878. 12. Taylor B, Norman AP, Orgel HA, Stokes CR, Turner MW, and Soothill JF: Transient IgA deficiency and pathogenesis of infantile atopy, Lancet 2:111, 1973. 13. Strober W, Blaese RM, and Waldman TA: Immunologic deficiency diseases, Bull Rheum Dis 22:686, 1972. 14. Rachelefsky GS, Osher A, Dooley RE, and Stiehm ER: Coexistent respiratory allergy and cystic fibrosis, Am J Dis Child 128:355, 1974. 15. Davis AT, and Quie PG: Phagocytes and phagocytosis, in Stiehm and Fulginiti, editors: Immunologic disorders in infants and children, WB Saunders Company, p 85. 16. Janeway CA, Craig J, Davidson M, Doroney W, Gitlin D, and Sullivan JC: Hypergammaglobulinemia associated with severe recurrent and chronic non-specific infections, Am J Dis Child 88:388, 1954. 17. Minor TE, Dick EC, DeMeo AN, Ouellette JJ, Cohen MC, and Reed CE: Viruses as precipitants of asthmatic attacks in children, JAMA 227:292, 1974. 18. McIntosh K, Ellis EF, Hoffman LS, Lybass TG, Eller JJ, and Fulginiti VA: The association of viral and bacterial respiratory infections with exacerbations of wheezing in young asthmatic children, J PEDIATR82:578, 1973. 19. Berkovich S, Millian SJ, and Snyder RD: The association of viral and mycoplasma infections with recurrence of wheezing in the asthmatic child, Ann Allergy 28:43, 1970. 20. Sokoi RR, and Rohlf FJ, editors: Analysis of frequencies, in: Biometry, San Francisco, I969, W. H. Freeman & Co, p 589.
August 1977 TheJournalofPEDIATRICS
Hyper-M-immunoglobulinemia in children with bronchial asthma Radial immunodiffusion was used to measure the concentrations of lgG, IgA, and IgM in the sera of 224 asthmatic children, ages 6 months to 14 years. IgM was greater than two standard deviations above agematched normal values in 51% of these asthmatic subjects, lmmunoglobulin profiles were repeated up to four years later in 29 individuals with elevated l g M and increased IgM synthesis was sustained in 14 of these asthmatic subjects. Serum lgE concentrations were elevated in 23 o f 35 asthmatic patients. There was no statistical difference in mean IgE between asthmatic children with normal or increased IgM. The distribution of lgE in each group was similar. Only 10 of 224 subjects had an IgA concentration less than 2 SD below age-matched control subjects. The significance and possible mechanisms of hyperM-immunoglobulinemia are discussed.
Ming S. Lin, M.D., Ph.D., Bruce S. Rabin, M.D., Ph.D., Ralph LaNeve, M.D.,
and Philip Fireman, M.D.,* Pittsburgh, Pa.
ASTHMA is a syndrome of several etiologies characterized by bronchospasm, increased bronchial mucous secretions, and bronchial edema. The immunologic development of asthma is often mediated by reaginic (IgE) antibodies; however, in many patients reagin-mediated immediate hypersensitivity cannot be demonstrated. Even though several investigators have reported an elevation of mean serum IgE values in asthmatic individua!sl 1 it is not unusual for individual patients with severe bronchial asthma to have normal or low serum IgE, 2 whereas increased concentrations of serum IgE may be present in patients without any asthmatic symptoms? Thus, measurement of serum IgE concentrations cannot be used alone to assess the severity of asthma. In an attempt to uncover immunologic mechanisms which may be operative in severely asthmatic BRONCHIAL
From the Departments of Pediatrics and Pathology, University o f Pittsburgh School of Medicine, and Children's Hospital of Pittsburgh. Supported by United States Public Health Service Grants HD-02662 and HD-07850. Presented in part at the annual meeting of the American Academy of Allergy, February, 1975. *Reprint address: Children's Hospital, 125 Desoto St., Pittsburgh, PA 15213.
Vol. 91, No. 2, pp. 222-227
patients, several investigators have demonstrated inconsistent changes from normal in serum IgG, IgA, or IgM concentration in asthmatic patients. ~-7 Several factors could contribute to this: (1) the number of patients studied was rather small, (2) the data obtained were not analyzed by age and sex, and (3) the criteria used to select the subjects were different in each study. The present study was undertaken to evaluate serum immunogl0bulin concentrations and calculate the prevalence of immunoglobulin abnormalities in children with recurrent asthma. The results were analyzed by age and sex of the patients and then compared to those of normal individuals. The study demonstrated that many asthmatic children who have normal or elevated serum IgE concentrations may have another immunoglobulin abnormality, usually an elevation of serum tgM values.
MATERIALS AND METHODS Study population. The subjects evaluated in this study consisted of 224 consecutive asthmatic patients, aged 6 months to 14 years, referred to the allergy clinic of Children's Hospital of Pittsburgh and evaluated by one of the authors (M.S.L.) The diagnosis o f asthma was established on the basis of history and physical examination. The history and pfiysical examination elicited specific
Volume 91 Number 2
Hyper-M-immunoglobulinemia
223
Table I. Descriptive statistics for IgM for patients and normal subjects classified by age and sex
Normal
Asthmatic t-Statistic
Age
Sex
N*
3-6 mo
M &F
7
7-12 mo
M&F
6
13,24 mo
M &F
11
25-36 mo
M &F
10
3-4 yr
M
9
3-4 yr
F
8
5-6 yr
M
10
5-6 yr
F
9
7-8 yr
M
9
7-8 yr
F
7
9-11 yr
M
13
9-11 yr
F
9
12-14 yr
M
13
12-14 yr
F
6
GM'~
N
43.7 (28.8, 66.5) 53.6 (25.1, 114,7) 67.3 (35.8, 126.5) 54.5 (26.3, 113.4) 77.4 (30.5, 196.3) 64.1 (23.4, 175.8) 62.2 (25.8, 150.0) 94.8 (37.8, 190.2) 87.1 (34.2, 186.8) 65.3 (21.4, 180.1) 66.8 (30.8, 144.9) 69.0 (32.3, 147.6) 88.1 (36.4, 213.3) 64.0 (39.4, 104.7)
5 9 36 15 8 13 18 13 15 10 20 10 11 5
GM
t%
93.4 (25.2, 345.8) 100.5 (37.8, 266.7) 116.4 (49.2, 275.4) 133.4 (64.1, 277.3) 117.8 (46.7, 297.2) 157.8 (76.6, 325.1) 130.6 (47.2, 361.4) 143.5 (68.4, 301.3) 120.7 (45.7, 391.3) 172.2 (76.2, 389.0) 126.8 (57.7, 278.6) 156.3 (91.6, 266.6) 123.9 (76.2, 423.6) 152.1 (65.2, 354.8)
--3.17w --2.65 N --3.90w --5.99w --1.86
--4.79w --3.87w -2.34 82 -1.61 -4.15w --4.60w --5.47w
-2.5311 -4.23w
*N = number of observations. tGM = geometric mean = antilog x, where g is the mean of logarithm of IgM. The two values appearing below the geometric mean are, respectively,antilog (g 2 SD) and antilog (x+ 2 SD) where SD = standard deviation of logarithm of lgM. :~The t-statistics were obtained from a comparison of the mean logarithm of lgM for normal subjects and patients using comparable age-sex classes. w < 0.005. prP< 0.01.
]1P < 0.05. information regarding recurrent asthmatic symptoms and signs, including more than six previous episodes of wheezing, cough, or exercise-induced bronchospasm which were relieved by theophylline or sympathomimetic medications. None of the patients studied was receiving corticosteroid therapy and none had received hyposensitization therapy at the time the first blood sample was drawn. Normal sera were obtained from venous blood of healthy infants and children attending the general pediatric practice of one of the authors (R.L.) The detailed medical background of each donor was evaluated and the serum was included in the study only if there was no history of frequent infections, allergy, or autoimmune diseases. The normal subject group consisted of 75 boys and 51 girls. Serum samples were collected by venipuncture and stored at - 2 0 ~ C until analyzed. No sodium
azide or other bactericidal agents were added to the serum. Informed consent was obtained from all subjects; the research was approved by the H u m a n Right Committee of the Children's Hospital of Pittsburgh. Q u a n t i t a t i o n of serum i m m u n o g l o b u l i n s . Serum concentrations o f IgG, IgA, and IgM were measured by radial immunodiffusion using quantitative i m m u n o g l o b u l i n plates (Meloy Laboratories, Springfield, V a . ) ? " Sera were collected from the normal subjects and from 189 patients who were evaluated between 1971 and 1974; o f these 112 were male and 77 were female. The same immunoglobulin reference standard was used throughout the study. Serum IgE concentrations, in addition to serum IgG, IgA, and IgM, were measured in 35 asthmatic patients who were seen between 1974 and 1975; of these 24 were male and 11 were female. IgE was quantified by
224
Lin et al.
400
The Journal of Pediatrics August 1977
1
1
1
1
1
1
(
1
1
1
1
[
== r ~- 3 0 0
=,
200
<[ z
z_
200
A--
-
~
IOO
9
H
bJ H
I00 4/12 8/12
I
2
3
4
5
6
7
8
9
I0
II
12
13
14
AGE (YEARS)
Fig. 2. Serum IgM concentrations for male asthmatic children, Similar graphic representation as in Fig. 1. 0 4 / 1 2 8/12
I
2
3
4
5
6
7
8
9
I0
II
12
13
14
AGE ( Y E A R S |
Fig. 1. Serum IgM concentrations for female asthmatic children. The shaded area represents the range of normal serum IgM concentration for that age, and the continuous line in the shaded area represents the geometric mean of the normal IgM. radioimmunoassay with kits purchased from Pharmacia AB, Uppsala, Sweden. Statistical analysis. The serum concentrations of various immunogl0bulins, IgG, IgA, IgM, and IgE were converted to their logarithms for statistical analysis/~ For the normal children and the asthmatic patients the mean (x) and the standard deviation of these transformed IgG, IgA, IgM, and IgE values were determined for the various age and sex groups. Detailed results for IgM are presented in Table I which shows the antilog (x), antilog (x - 2 SD), and antilog (x + 2 SD). Corresponding age and sex groups for normal subjects and patients were compared on the basis of the mean logarithm values using the tstatistic. These comparisons were preceded by assurance that the variances of the corresponding groups were similar. Such conditions were met in all instances. Limits for normal values were considered to be within 2 SD of the antilog of the geometric mean. RESULTS Quantitative serum immunoglobulins, IgG, IgA, and IgM were determined in 224 patients with bronchial asthma. Their ages ranged from 6 months to 14 years: 136 were boys and 88 were girls. Among the total 224 patients we have studied, only 8.8% had values of all three
immunoglobulin classes within 1 SD of the mean and 30.8% within 2 SD. Serum IgM concentrations were elevated in many asthmatic patients and no individual value was below 2 SD of the geometric mean (Figs. 1 and 2). Of the female patients, 40% had IgM levels within normal limits; the remainder were above 2 SD of the mean (Fig. 1). In male patients 54% h a d normal serum IgM, concentrations the rest (46%) had elevated IgM levels (Fig. 2). In the 224 patients we have studied, more than half (51%) of the patients had elevated serum IgM concentrations. In Table I statistical comparison of IgM levels in normal and asthmatic subjects by age and sex showed significant differences in 12 of the 14 groups studied. To determine whether increased IgM synthesis was sustained~ immunoglobulin profiles have been repeated in 29 patients with increase d IgM concentrations. They were divided into four groups on the basis of time interval after the initial immunog!obulin determinations. The results (Fig. 3) indicated that the serum IgM Concentrations decreased during subsequent follow-up examinations in all but four of the asthmatic patients who had hyper-Mimmunoglobulinemia, but 14 patients still had increased serum IgM values. To study the relationship between the IgE and IgM values, IgE was quantified in 35 patients seen between 1974 and 1975. Elevated IgE was found in 23 patients. These patients were divided into two populations; one with normal IgM and the other with increased IgM. The geometric mean values of IgE for the patients with elevated IgM values was 347.1 I U / m l and that of the
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Hyper-M-immunoglobulinernia
225
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Z
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1-3 mos, later
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4-12 mos.
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2-4 yrs. later
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Fig. 3. Serum IgM levels up to four years after initial determination and grouped on the basis of the time interval as shown on the abscissa. The ordinate represents the ratio between the patients' serum IgM and the highest normal serum IgM for that age and sex. Ratio greater than 1.0 indicates hyper-M-immunoglobulinemia; ratio smaller than 1.0 means normal serum IgM. Patients with sustained increased IgM levels are indicated by the open circle. The n value for this study was 29. patients with normal IgM was 219.0 lU/ml. The differonce between these two groups was not statistically significant (Fig. 4). When these 35 asthmatic patients were divided into four groups (Table IX) according to the frequency of asthmatic attacks that were observed during the next 12 months and without prior knowledge of the serum concentrations of IgE and IgM, the following was observed: the probability of frequent asthmatic attacks in a patient with increased serum IgE and IgM concentrations was 0.86 (6 out of 7 patients), in patients with elevated IgM values alone was 0.57 (12 out of 21 patients), and in patients with increased IgE concentrations was 0.21 (3 out of 14 patients). Serum IgG concentration was normal in the majority of the patients. In the female asthmatic patients, 14.0% had IgG higher than 2 SD and none had IgG deficiency. Similarly in the male patients, 15.0% had high and 3.5% had low IgG. Of the entire population studied, 14.8% of the asthmatic patients had elevated IgG and 2.1% had IgG deficiency. Statistical comparison of IgG levels by age and sex showed significant differences in only one of the 14 groups studied.
Serum IgA concentrations were normal in most asthmatic patients. In the female asthmatic patients, 20.7% had elevated IgA concentrations and 6.5% had serum IgA deficiency. In the male asthmatic patients, 21.4% had elevated serum IgA concentrations and 3.5% had IgA deficiency. As a group, 21% of the asthmatic patients had IgA values higher than 2 SD and 4.7% had low IgA concentrations. Statistical comparison of IgA levels by age and sex showed significant differences in only 3 of the 14 groups studied. DISCUSSION An immune response which depends on the presence of IgE and the balance between IgE and other specific antibodies has been postulated to play a central role in the pathogenesis of certain varieties of bronchial asthma?, ~A quantitative evaluation of serum immunoglobulins in bronchial asthma could, therefore, provide a better understanding of the humoral antibody system in this disease. In the first stage of our studies, serum concentrations of IgG, IgA, and IgM were measured in 189 asthmatic children. These data differ from those reported by other
226
Lin et al.
The Journal of Pediatrics August 1977
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i 12
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AGE OF PATIENTSWITH HYPER IgM ( in years)
AGE OF PATIENTSWITH NORMAL IgM
( in years ]
F~. 4. Comparison of serum IgE levels at different ages in asthmatic subjects with hyper-IgM on left and in asthmatic subjects with normal IgM on right. Table U. Comparison of serum IgM and IgE levels with the frequency of acute asthmatic episodes in asthmatic children-~176
Frequency of attacks At least 1/wk:~ One to 3/mo Less than 1/mo Asymptomatic~
lgM( ~,)* IgE( ~ )
I
lgM(+ ) IgE(Nff
lgM(N) IgE( +)
12 5 1 3
2 0 1 5
6 0 0 1
I
IgM(N)lgE(N) 3 2 1 8
*(+) Indicatesincreasedserumlevel. t(N) Indicatesnormalserum level. :~Thereis no statisticalevidencebased on the chi square statisticsfor associationbetweenchangesin IgM and changesin IgE in those two groupsof patients. investigators who reported no deficiency or increase in immunoglobulins A, G, and M in patients with bronchial asthma?. 6. 7.11 Since the number of patients studied in these previous investigations was rather small and the results for each immunoglobulin class were not analyzed by age and sex, our data cannot be compared to data reported in the studies. Defective immunologic surveillance as a consequence of immune deficiency has been suggested to play an essential role in the pathogenesis and development of asthma and it has been reported that many forms of allergy are more common in patients with immunodeficiency than in healthy populations. 12 ~3 In the present study, data on individual serum immunoglobulin concentrations in 224 asthmatic children were examined for
the prevalence of selective immunoglobulin deficiencies and, more importantly, for interrelationships between immunoglobulin classes. The prevalence of selective immunoglobulin deficiency was found much less frequently than that of selective hyperimmunoglobulinemia. The prevalence of selective IgG, IgA, and IgM deficiency was 2.1%, 4.7%, and 0%, respectively. On the contrary, hypergammaglobulinemia, especially an increase in the IgM values was a rather common phenomenon ~n the children with bronchial asthma. The prevalences of increased serum IgG, IgA, and IgM concentrations were 15%, 21%, and 51%, respectively. This observation seriously challenged the previous speculation that a selective immunoglobulin deficiency, particularly of IgA, would result in a defect in eliminating the offending
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antigens, which in turn would stimulate the host to produce various i m m u n e substances such as IgE. Although h y p e r - M - i m m u n o g l o b u l i n e m i a was a common p h e n o m e n o n in children with bronchial asthma, the increased IgM synthesis was not sustained at the same level in most asthmatic patients and in fact returned to normal serum concentration in 15 of 29 asthmatic patients re-evaluated one month to four years later (Fig. 3). Nevertheless, four of these 29 patients sustained increased IgM synthesis; these four patients had more severe symptoms in spite of appropriate m a n a g e m e n t and required more medication in comparison to those patients in whom IgM synthesis had decreased. There were few significant differences in the serum levels of IgG and IgA between asthmatic and normal children when the various age and sex groups were compared; these data were similar to those observed by several investigators, 1' * 7 but different from those reported by Collins-William and associates? The m e a n IgE values in asthmatic children with normal IgM concentration were similar to those with increased IgM values and the distribution of IgE in each group was also quite similar (Fig. 4). Furthermore, the probability tha t an asthmatic patient with normal serum IgM concentration would have an elevated serum IgE value was 0.70, whereas the probability that those with increased IgM concentrations would have an elevated IgE value was 0.68. These observations suggest the probability that a patient with h y p e r - M - i m m u n o g l o b u l i n e m i a would have an elevated serum IgE value was not greater than for those with normal IgM concentrations. It may be important to point out that the sum of the probability of frequent attacks in patients with elevated IgM alone (0.57) and elevated IgE alone (0.25) was 0.82, which was close to, but lower than, that of the patients with elevated serum IgM and IgE concentrations (0.86). This suggested that the presence of at least two distinct pathologic processes affecting the respiratory tract might result in a clinical situation worse than either condition alone. Since hypergammaglobulinemia has been previously described in patients with frequent infections, "-1'~ and since viral infections have been reported as precipitants of asthmatic attacks in children, ~7-1~'this study may suggest that elevated IgM levels were due to repeated infection with different strains of viruses. As a result, specific clones of lymphocytes in response to various antigenic stimulation began to proliferate and accordingly more IgM was synthesized. The offending antigens could be either the antigens which initiated the allergic response or microorganisms which had infected the patients. The latter possibility is currently undergoing investigation in our laboratory and clinic.
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REFERENCES
1. Johansson SGO: Raised levels of a new immunoglobulin class (IgND) in asthma, Lancet 2:951, 1967. 2. Berg T, and Johansson SGO: IgE concentration in children with atopic diseases, Int Arch Allergy 36:219, 1969. 3. Aas K" The biochemical and immunological basis of bronchial asthma, Springfield, IL, 1972, Charles C Thomas, Publisher Chapter 13. 4. Collins-Williams C, Lamenza C, and Kokubu H: Deficiency of IgA in serum and respiratory secretions, Can Med Assoc J 99:1069, 1968. 5. Collins-Williams C, Tkachyk SJ, Toft B, Generoso L, and Moscarello M: Quantitative immunoglobulin levels (IgG, IgA and IgM) in children with intractable asthma, Ann Allergy 25:177, 1967. 6. Palma-Carlos AG, and Palma-Carlos ML: Immunoglobulins IgG, IgA and IgM in allergic diseases, Acta Allergol 26:161, 1971. 7. Frouchtman R, Vinas J, Rodriguez JL, and Garcia YP: Immunoglobulins in bronchial asthma, Rev Clin Esp 122:323, 1971. 8. Mancini G, Carbonara AO, and Heremans JF: Immunochemical quantitation of antigens by single radial immunodiffusion, Immunochemistry 2:235, 1965. 9. Leonardy JG, and Peacock LB: An evaluation of quantitative serum immunoglobulin determinations in clinical practice, Ann Allergy 30:378, 1972. 10. Sokol RR, and Rohlf F J, editors: Single classification analysis of variance, in: Biometry, San Francisco, 1969, W H Freeman & Co, p 220. 11. Radermecker M, and Rose B: Bronchial asthma, in Samter M, editor: Immunological Diseases II, Boston, 1971, Little, Brown & Company, p 878. 12. Taylor B, Norman AP, Orgel HA, Stokes CR, Turner MW, and Soothill JF: Transient IgA deficiency and pathogenesis of infantile atopy, Lancet 2:111, 1973. 13. Strober W, Blaese RM, and Waldman TA: Immunologic deficiency diseases, Bull Rheum Dis 22:686, 1972. 14. Rachelefsky GS, Osher A, Dooley RE, and Stiehm ER: Coexistent respiratory allergy and cystic fibrosis, Am J Dis Child 128:355, 1974. 15. Davis AT, and Quie PG: Phagocytes and phagocytosis, in Stiehm and Fulginiti, editors: Immunologic disorders in infants and children, WB Saunders Company, p 85. 16. Janeway CA, Craig J, Davidson M, Doroney W, Gitlin D, and Sullivan JC: Hypergammaglobulinemia associated with severe recurrent and chronic non-specific infections, Am J Dis Child 88:388, 1954. 17. Minor TE, Dick EC, DeMeo AN, Ouellette JJ, Cohen MC, and Reed CE: Viruses as precipitants of asthmatic attacks in children, JAMA 227:292, 1974. 18. McIntosh K, Ellis EF, Hoffman LS, Lybass TG, Eller JJ, and Fulginiti VA: The association of viral and bacterial respiratory infections with exacerbations of wheezing in young asthmatic children, J PEDIATR82:578, 1973. 19. Berkovich S, Millian SJ, and Snyder RD: The association of viral and mycoplasma infections with recurrence of wheezing in the asthmatic child, Ann Allergy 28:43, 1970. 20. Sokoi RR, and Rohlf FJ, editors: Analysis of frequencies, in: Biometry, San Francisco, I969, W. H. Freeman & Co, p 589.