Full text of DOI:10.1007/BF03343975

J. Endocrinol. Invest. 25: 125-128, 2002
Lymphocyte subset pattern in acromegaly
A. Colao*, D. Ferone*, P. Marzullo*, N. Panza**, R. Pivonello*, F. Orio Jr.*,
G. Grande***, N. Bevilacqua***, and G. Lombardi*
Departments of *Molecular and Clinical Endocrinology and Oncology, “Federico II” University of
Naples; **Oncology and ***Clinical Chemistry, “A. Cardarelli” Hospital, Naples, Italy
ABSTRACT. Immune function in acromegalic pa-
tients has been poorly investigated. The aim of
this study was to evaluate the main surface anti-
gen clusters of circulating lymphocytes in acro-
megaly. One hundred patients with active acro-
megaly (55 women and 45 men, aged 20-70 yr)
and 200 healthy subjects sex- and age-matched
with the patients (110 women and 90 men, aged
20-70 yr) were enrolled in this study. All patients
and controls were born and live in Southern Italy.
No patient had received octreotide, bromocrip-
tine or corticosteroids for at least 3 months be-
fore entering the study. The analysis of lympho-
cyte subset pattern was performed by flow cy-
tometry and fluorescein isothiocyanate or phyco-
erythrin directly conjugated monoclonal antibod-
ies specific for the cell surface antigen clusters
(CD) representing T-cell population as a whole
(CD3), T helpers (CD4), T suppressors (CD8), nat-
ural killer cells (CD16) and B-cell population as a
whole (CD19). Acromegalics had significantly in-
creased levels of CD3 (67.1 7.2 vs 64.3 8.8%;
p=0.03) and CD4 (37.8 3.5 vs 36.4 4.3%;
p=0.004) and decreased levels of CD8 (31.4 3.3
vs 33.7 8.2%; p<0.01) and CD19 (12.1 3.1 vs
15.2 5.1; p=0.01) without age-difference. The re-
sults of the current study demonstrate an increase
in T-cell activity together with a decrease in B-cell
activity in a very large series of patients with active
acromegaly. These data further support the exis-
tence of abnormalities of the immune system in
patients with chronic GH/IGF-I excess.
(J. Endocrinol. Invest. 25: 125-128, 2002)
©2002, Editrice Kurtis
INTRODUCTION
other hand, GH-deficient humans are not immuno-
deficient (17) and immune function in GH-treated
children has been reported to be normal (18), in-
creased (19) or decreased (20). Despite this con-
flicting evidence on the association between GH
secretory status and modulation of immune re-
sponse, immune function in acromegalic patients
has been poorly investigated.
The aim of this study was to evaluate the lympho-
cyte subset pattern in a large series of consecutive
active acromegalic patients, and compare it to a con-
sistent control group of Caucasian healthy sex- and
age-matched individuals, living in Southern Italy.
In acromegaly, various tumors have been reported
to occur with a greater than expected incidence (1-
3). In particular, prevalence of tumors of the gas-
trointestinal tract was suggested to be higher in
acromegalics than in the healthy population (4-8),
although negative data have also been reported
recently (9). GH/IGF-I axis has long been supposed
to play a major role in immuno-modulation: in rats
hypothalamic damage leads to abrogation of nat-
ural killer cell activity (10), whereas fetal hypophy-
sectomy in mice results in thymic atrophy and im-
munodeficiency (11). Moreover, specific GH re-
ceptors have been found on peripheral mononu-
clear cells (12); GH modulates lymphoproliferation
in vitro (13-15) and is considered a minor growth
factor for normal lymphocyte in vivo (16). On the
PATIENTS AND METHODS
Subjects
One hundred acromegalic patients with active dis-
ease (55 women and 45 men, aged 20-70 yr) en-
tered this study. The diagnosis of acromegaly was
based on clinical, hormonal and radiological fea-
tures, as elsewhere reported (21, 22). GH was
31 8.5 μg/l not suppressed below 1 μg/l after oral
administration of 75 g glucose, IGF-I was 780.5 66
μg/l. No patient had received octreotide or bromo-
Key-words: Acromegaly, GH, IGF-I, immune system, lymphocyte subsets.
Correspondence: Dr. Annamaria Colao, Dipartimento di Oncologia e
Oncologia Molecolare, Università Federico II, Via S. Pansini 5, 80131
Napoli, Italia.
E-mail: colao@unina.it
Accepted August 1, 2001.
125

A. Colao, D. Ferone, P. Marzullo, et al.
Controls
Patients
0.03
criptine for at least 3 months before entering the
study. L-thyroxine, given at the dose of 75-125
μg/day for goiter to 63 patients, was withdrawn 15-
30 days before entering the study. Two-hundred
healthy subjects of the medical and paramedical
staff of the Departments of Molecular and Clinical
Endocrinology and Oncology of Federico II Uni-
versity and of the Unit of Oncology and of Labo-
ratory of the Cardarelli Hospital in Naples, sex- and
age-matched with patients, served as controls (110
women and 90 men, aged 20-70 yr). All patients
and controls were born and lived in Southern Italy.
The exclusion criteria for acromegalics and controls
were: age over 70 yr, previous benign and malig-
nant tumors, anemia, previous radiation and corti-
costeroid therapy performed 3 months before en-
tering the study. All patients and controls gave their
informed consent orally in the presence of a third
person.
75
50
25
0
0.004
0.01
0.01
CD3
CD4
CD8
CD19
Fig. 1 - Lymphocyte whole population (CD3), T helpers cells (CD4),
T suppressor cells (CD8), and B-cell population as a whole (CD19)
in 100 patients with acromegaly and 200 controls.
RESULTS
Methods
In acromegalic patients, the expression of the whole
lymphocyte population (CD3) and of T helpers cells
(CD4) was significantly increased, when compared to
control subjects (Fig. 1). By contrast, the expression of
T suppressor cells (CD8), and B- cell population as a
whole (CD19) was significantly decreased (Fig. 1). No
significant difference was found between acrome-
galics and controls in the expression of natural killer
cells, represented by the natural killers (CD16) ex-
pressing cells (9.9 4.9 vs 10.2 3.3%). In addition, a
significant decrease of the ratio between CD4 and
CD8 was observed in acromegalic patients compared
to controls (1.2 0.5 vs 1.5 0.5; p<0.001). No differ-
ence in the expression of CD3, CD4, CD8, CD16 and
CD19 was found among young (age below 40 yr),
middle-aged (40-60 yr) or elderly (>60 yr) patients
with acromegaly (Fig. 2).
The analysis of lymphocyte subset pattern was per-
formed by flow cytometry using aliquots of whole
fresh blood collected from an antecubital vein and
fluorescein isothiocyanate (FITC) or phycoerythrin (PE)
directly conjugated monoclonal antibodies (MoAbs)
specific for the cell surface antigen clusters (CD) 3, 4,
8, 16 and 19 (Becton Dickinson Monoclonal Center
Inc., Mountain View, CA, U.S.A.). Lymphocyte gated
on light scatter were analyzed for fluorescence in-
tensity using FACScan (Becton Dickinson) acquiring
5000 events for at least each sample.
Assays
Serum GH levels were measured by IRMA (HGH-
CTK-IRMA Sorin, Saluggia, Italy). The sensitivity of
the assay was 0.2 μg/l; 1 μg/l corresponds to 2.5
mU/l. The intra- and inter-assay coefficients of varia-
tion (CV) were 4.5 and 7.9%, respectively. Plasma
IGF-I was measured by IRMA after ethanol extrac-
tion using DSL kits (Webster, Tx, U.S.A.). The sensi-
tivity of the assay was 0.8 μg/l. The intra-assay CV
were 3.4, 3.0 and 1.5% for the low, medium and high
points on the standard curve, respectively. The in-
ter-assay CV were 8.2, 1.5 and 3.7% for the low, me-
dium and high points on the standard curve. Fasting
GH levels were considered above the normal range
when >2.5 μg/l. In our laboratories the normal IGF-I
range was 110-450 μg/l in ꢀ40, 100-300 μg/l in 41-
59, and 78-258 μg/l in ꢁ60 yr old subjects.
100
75
50
25
0
<40 yr
>40 yr
CD3
CD4
CD8
CD19
Statistical analysis
Fig. 2 - Lymphocyte whole population (CD3), T helpers cells
(CD4), T suppressor cells (CD8), and B-cell population as a
whole (CD19) in 100 patients with acromegaly grouped on the
basis of age below (no.=38) or above 40 yr (no.=62).
Statistical analysis was performed by ANOVA. Si-
gnificance was set at the 5%. Data are reported as
Mean SD.
126

Immune system in acromegaly
DISCUSSION
polyp itself (7). However, patients with and without
polyps had a similar lymphocyte subset pattern, thus
excluding a pathogenetic role for colonic polyps in
altering the lymphocyte subset pattern (7). The exis-
tence of an altered immune function in acromegaly
was also reported in a small series of acromegalic pa-
tients and controls: while natural killer cell activity,
serum concentrations of immunoglobulins (IgG, IgM,
IgA) and metabolic burst activity were within the nor-
mal range in both groups, a significantly enhanced
phagocytic activity was observed in patients with
acromegaly. Kotzmann et al. (35) reported also that
surface markers on T lymphocytes (CD3, CD4, CD8),
B lymphocytes (CD19) and natural killer cells
(CD16/56) were normal in both groups, but CD4+ and
CD8+ cells showed a significantly higher expression
of transferrin receptors in acromegalic patients, indi-
cating enhanced T-cell activity. In our current series
of patients with acromegaly, T-cell activity, measured
as CD3 and CD4, was also shown to be enhanced.
Furthermore, aging has been shown to be accompa-
nied by various changes in the lymphocyte subset
distribution. In fact, a significant increase in both the
absolute counts and the proportions of CD3 and
CD57 was found with age, whereas the cytolytic T-
cell population showed less change with age (36). In
addition, soluble interleukin-2 receptor levels were
found to increase significantly with age and correlat-
ed with certain natural killer cell subsets (36). The func-
tions of these changes are still unclear but the ex-
pansion of some lymphocyte subsets in the elderly
was suggested to represent a remodeling of the im-
mune system with ageing, with an increase in non-
MHC (major histocompatibility complex)-restricted
cells likely to compensate the previously reported de-
cline in T- and B-cells (36). This decline in T- and B-
cells was also found in elderly acromegalics as com-
pared to young and middle-aged patients, suggest-
ing the persistence of this remodeling mechanism in
patients with chronic GH/IGF-I hypersecretion.
The results of the current study demonstrate a de-
crease of T-and increase of B-cell lines in patients with
active acromegaly as compared to sex- and age-
matched controls. The changes in the lymphocyte sub-
set pattern were independent from the patients’ age.
GH has been proposed to be an essential hormone
for the development, maintenance and regulation of
immune function (23, 24): it is necessary to maintain
lymphatic tissues populated with lymphocytes since
the removal of GH results in thymus atrophy and sec-
ondary lymphoid tissues (25-28). GH has been shown
to induce differentiation both of thymocyte precur-
sors into helper T-cells (29) and of stem cells into
granulocytes (30). Moreover, GH can increase in vi-
tro development of cytotoxic T lymphocytes in mixed
lymphocyte cultures (24) and activate macrophages
(31). On the other hand, GH may enhance the in vi-
tro proliferation of virus-injected erythroleukemia
cells (14) and of certain types of transformed cells
(13-15). In addition, circulating GH and IGF-I have
been reported to promote leukemia blast cell repli-
cation in vivo (32). Over-expression of heterologous
or homologous GH in transgenic mice was shown to
induce significant stimulation of some parameters of
immune function (32). In fact, in metallothionein I
(MT)-bGH transgenic mice with high peripheral lev-
els of bovine GH, the absolute weight of the thymus
and the spleen was significantly increased and the
mitogenic responses of splenocytes to concanavalin
A (ConA), lipopolysaccharide (LPS) and phyto-
hemagglutinin (PHA) were enhanced, as compared
to age-matched normal animals (32).
Although the neoplastic risk has been claimed to be
increased in acromegaly (1-9), data regarding immune
functions in patients with acromegaly are currently
limited. In a study reported by others (7), the lym-
phocyte subset pattern in the colonic lamina propria
of 34 acromegalics showed a significant decrease of
B-cells (CD19, CD20), CD16, ꢂ/ꢃ ratio, considered as
index of intraepitelial cytotoxic activity (33), and an in-
crease of T-cells as a whole (CD3), when compared
to controls. Interestingly, the changes of the lympho-
cyte subset pattern were found in patients with active
disease while those with inactive disease had a pattern
similar to healthy subjects. Since the study was per-
formed at the mucosal level of the colonic lamina pro-
pria close to the polyp lesion, the decrease of the
helper-inducer (CD4+/leu8-) lymphocytes suggested
an impairment of the immunoglobulin synthesis me-
diated by the above-mentioned T-cells at mucosal
level (34). In contrast, the increase of the helper-sup-
pressor (CD4+/leu8+) lymphocytes, supposed to reg-
ulate the cell-mediated immune response at mucos-
al locoregional site, could be caused by the colonic
The results of the current study demonstrate an in-
crease in T-cell activity together with a decrease in
B-cell activity in a very large series of patients with
active acromegaly. These data further support the
existence of abnormalities of the immune system
in patients with chronic GH/IGF-I excess.
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