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Original Studies |
Departments of Pediatrics (L.W., J.P., E.S., C.H., F.A., C.D.), Clinical Biochemistry (E.D.), Hôpital Sainte-Justine, Université de Montréal, Montréal, Québec, Canada H3T 1C5; Department of Endocrinology (P.C.), John Hunter Children’s Hospital, Newcastle, Australia 2310; Department of Medicine (O.K.), University Hospital, Uppsala, Sweden SE-75185
Address correspondence and requests for reprints to: Cheri L. Deal, Ph.D., M.D., Associate Professor of Research, Univ. of Montréal, Endocrinology Service, Hôpital Sainte-Justine, 3175 Côte Sainte-Catherine, Montréal, Québec Canada H3T 1C5. E-mail: dealc{at}ere.umontreal.ca
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Abstract |
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 Top Abstract IntroductionSubjects and MethodsResultsDiscussionReferences |
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Haplotype analysis suggested compound heterozygozity at the
AIRE locus. Direct sequencing of exon 8 revealed a
previously described mutation, a 13-bp deletion (1085–1097) of
maternal origin, found in the index patient, her affected sister, and
her unaffected sister. A novel missense mutation characterized by a
T
G transversion at nucleotide position 398, resulting in a leuarg
amino acid substitution (L93R), was found in exon 2. The mutation was
present in the father, the brother, the index patient, and the affected
sister. The presence of the mutation in the propositus was verified by
cloning of PCR products from genomic DNA. The mutation destroys a
PstI restriction enzyme site, as confirmed in the
aforementioned patients. Screening of 50 French-Canadian controls with
PstI digestion did not show destruction of the
restriction-enzyme site.
The index patient’s phenotype was severe, manifested by classic features of the illness (adrenal insufficiency, hypoparathyroidism, candidiasis, and keratoconjunctivitis with alopecia universalis), as well as by severe exocrine pancreatic insufficiency, diabetes mellitus, hepatic inflammation, growth hormone (GH) deficiency due to lymphocytic hypophysitis, and primary ovarian failure. Oral CyA (5 mg/kg/day) was initiated at 13 yr of age. After 8 months of therapy, stimulated pancreatic lipase increased 24-fold with normalization of stool fat (from 31.5 g/day to 2.5 g/day, normal(N) < 5). There was complete resolution of her photophobia, and considerable hair regrowth was diffusely apparent. Minimal side effects were noted. Our experience supports the use of oral CyA for the treatment of severe APECED-associated exocrine pancreatic failure and keratoconjunctivitis.
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Introduction |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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The phenotype of APECED is highly variable; however, conclusions regarding genotype-phenotype correlations have not been possible (1, 2, 3). Although recent reviews have discussed the natural history of APECED in depth (1, 3), little emphasis has been placed on therapy. With increasing experience in the use of immunosuppressive therapy (11, 12), more aggressive and comprehensive treatment of APECED is feasible.
We investigated a French-Canadian pedigree including the adolescent index case who presented rare features of the disease that posed diagnostic and therapeutic challenges, notably severe exocrine pancreatic insufficiency and isolated growth hormone (GH) deficiency secondary to lymphocytic hypophysitis. We further report the clinical investigations in the evaluation of this girl’s severe disease and the results of treatment with cyclosporine A (CyA).
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Subjects and Methods |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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The index case, a 13.6-yr-old girl, was the first of four children born to healthy, nonconsanguineous, French-Canadian parents. A 12-yr-old sister has suffered only mild episodes of vaginal candidiasis. An 11-yr-old sister also presented with features of APECED, including mucocutaneous candidiasis, hypoparathyroidism, and alopecia areata. An 8-yr-old brother has been treated for oral candidiasis, but is otherwise unaffected.
Clinical course of index patient (refer to Table 1
for clinical summary)
Candidiasis and ectodermal dystrophy. The patient presented in infancy with recurrent mucocutaneous candidiasis, unresponsive to topical antifungal agents, though controlled with systemic therapy. Immunological investigations showed a cell-mediated immune deficiency with a low thymocyte-helper to thymocyte-suppresser cell ratio. Photophobia and visual impairment occurred at 6 yr of age followed by alopecia universalis at age 9 yr. The keratoconjunctivitis was of such severity that home schooling became necessary. Systemic vitamin A was initially prescribed, with some improvement. This was eventually changed to topical vitamin A ointment (Dulcis, Allergan, Inc., Mougins, France) because of concern for potential hepatoxicity; the topical vitamin A treatment proved remarkably effective in preventing new corneal ulceration and allowing resumption of classroom studies, although photophobia persisted. Dental enamel hypoplasia was present without evidence of nail dystrophy.
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Gastrointestinal symptoms associated with growth failure occurred
between the ages of 7.5 and 9 yrs. These included severe abdominal
distension (Fig. 1A), cramping pain,
diarrhea, and constipation. The subject was normocalcemic at the time.
Upper gastrointestinal series and rectal manometry were normal. Upper
endoscopy with quantification of intestinal disaccharidases revealed
hypolactasia (1.2 µmol substrate/min/g, normal (N =
11–85); sucrase and maltase levels were normal (55 µmol
substrate/min/g, N = 27–145 and 174 µmol
substrate/min/g, N = 112–618, respectively). In the
face of normal serum immunoglobulin A, G, and M levels, anti-enterocyte
and anti-gliadin antibodies were negative. Bacterial overgrowth was
present: duodenal fluid cultures were positive for two strains of
aerobic bacteria (streptococcus viridans, 2.0 x
108 CFU/L) and coagulase negative
staphylococcus, 2.0 x 108 CFU/L). She was
treated with a lactose-free diet, lactase supplements, and oral
metronidazole, without recuperation of weight or
improvement in symptoms. Endoscopy-directed small bowel biopsies at
ages 8 and 10 yr confirmed normal villous architecture with mildly
increased lymphocytic infiltration of the lamina propria. Silver
staining of the duodenal biopsies obtained at 8 and 10 yr demonstrated
an absence of serotonin-secreting enterochromaffin cells.
Lymphangiecstasia was ruled out on these biopsies and by a normal
albumin 51CrCl3 test to quantify stool protein
loss (0.16%, N <0.7).
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). Nine
months after the initiation of pancreatic enzyme therapy, the stool fat
was still extremely elevated (31.7 g/day). Despite maximal enzyme
replacement (pancrelipase, Creon 25, Solvay Pharma, 22 capsules/day),
weight loss was evident. Nocturnal nasogastric gavages with a
fat-deficient elemental diet (Tolerex, Novartis Nutrition)
were thus added at 10.6 yr of age, resulting in satisfactory weight
gain. However, abdominal distension and cramps persisted. Endocrine dysfunction. At 4.6 yr of age, the proband presented with seizures and tetany secondary to hypocalcemia. Hypoparathyroidism was documented, and parathyroid antibodies were positive by indirect immunofluorescence. She was successfully treated with calcitriol. At 10 yr of age, first-phase insulin release during an intravenous glucose tolerance test was performed as described (15) and was compared to age-matched, female controls. A decrease in the first-phase insulin release from greater than the 95th to less than 10th percentile (140.0 µU/ml to 70.3 µU/ml) (1, 15) was observed over the ensuing year. In addition, very high serum anti-islet cell antibodies (ICAs) were detected (>80 JDF units). After complaints of polyuria and polydipsia at age 11, home blood glucose monitoring revealed an elevated post-prandial glucose (270 mg/dL) with preprandial glucose values between 126 and 162 mg/dL. The HbA1C was normal (0.052, N = 0.043–0.058), and urine ketones were negative. Long-acting subcutaneous insulin was initiated at 11.6 yrs of age (0.15 U/kg/day). At the time of this report, the proband was receiving 0.4 U/kg/day long-acting human insulin before supper as a once daily injection to maintain euglycemia.
Despite significant improvement in the patient’s nutritional status (height-to-weight ratios above the 50th percentile) after treatment for pancreatic insufficiency, growth velocity continued to decrease [3.0 cm/yr between 11 and 12 yr of age, < 3rd percentile for bone age (16)]. At 11.9 yr of age her growth parameters were as follows: height 129.6 cm (-2.9 SD), weight 28.1 kg (-1.2 SD), and bone age 8 yr. Plasma insulin-like growth factor (IGF)-I was low before nutritional supplementation (46 ng/mL), but demonstrated a modest rise after the sequential initiation of pancreatic enzyme supplementation, gavage feedings, and insulin (46, 64, and 93 ng/mL, respectively). Provocative GH testing after confirming euthyroidism revealed inadequate peak responses of 0.8 and 0.3 ng/mL to L-dopa-propranolol and to clonidine, respectively. The maximal nocturnal GH peak was 3.1 ng/mL. The peak GH response to clonidine with estrogen priming (20 µg ethinyl estradiol once daily for three days) was 4.6 ng/mL. Magnetic resonance imaging demonstrated a perihypophyseal "halo effect" with gadolinium enhancement, suggestive of hypophysitis. Immunoblotting of human pituitary membranes with consecutive samples of the patient’s sera detected antipituitary autoantibody reactivity to a 43–45 kDa protein (17). These findings were highly suggestive of autoimmune hypophysitis complicated by GH deficiency. GH replacement therapy was provided with recombinant human growth hormone (0.18 mg/kg/week, given as a daily injection sc) at age 12.0 yr, and over a 12 month period there was significant increase in growth velocity (9 cm/yr) and an increase in plasma IGF-I to 483 ng/mL.
At age 12.7 yr, four months before the initiation of immunosuppressive therapy as outlined below, the gonadotropin and corticotropin axes were assessed by dynamic testing. A baseline cortisol was 13.9 µg/dL at 0800 (N = 5–25), with a peak of 15.4 µg/dL (N > 20) after 250 µg ACTH, although 24-h urine cortisol was normal. An LHRH stimulation test (100 µg) demonstrated a prepubertal response (baseline LH 0.8 mIU/mL, FSH 5.47 mIU/mL; peak LH 2.3 mIU/mL, FSH 16 mIU/mL). Anti-ovarian antibodies were positive. To date, posterior pituitary function has been unaffected.
At 13 yr of age, immunosuppressive therapy was initiated with CyA, 5 mg/kg/day, after the informed consent of both the patient and her parents. Indications and treatment goals were: 1), to improve pancreatic function, given that the patient remained dependent upon nocturnal gavages to prevent weight loss despite maximal therapy with pancreatic enzyme supplements; 2), to prevent further progression of the severe keratoconjunctivitis and of the hepatic inflammation; and 3), to prolong residual pancreatic endocrine function.
Methods
Autoantibodies and hormonal assays. Antimicrosomal liver antibodies (13) and antipituitary membrane antibodies (17) were measured by Western immunoblots as previously described. Anti-adrenal antibodies were detected by indirect immunofluorescence using monkey adrenal tissue (Inova Diagnostic Inc., San Diego, California). Methods for antiparathyroid antibodies (18), anti-IA-2, and antiglutamate decarboxylase (GAD) antibodies (19), anti-aromatic-L-amino-acid decarboxylase (AADC) (2, 20), and antitryptophan hydroxylase antibodies (21) have been detailed elsewhere. Anti-ovarian and anti-ICAs were kindly performed by Dr. Noel MacLaren (Harahan, LA).
Plasma insulin was measured by a semi-automated microparticle enzyme immunoassay (Abbott Laboratories, Montréal, Québec). Plasma growth hormone, cortisol, and gonadotropins were measured by time-resolved immunofluorometric assay (AutoDELFIA, EG&G Wallac, Kirkland, Québec). Glycated hemoglobin was performed with an immunoturbidometric inhibition assay (Boehringer Mannheim, Montréal, Québec). Plasma aldosterone was measured by a one-step solid-phase radioimmunoassay (Diagnostic Product Corp., Los Angeles, CA), plasma renin with a solid-phase radioimmunometric assay (Sanofi Diagnostics Pasteur, Montréal, Québec) and ACTH by a two-antibody, equilibrium radioimmunoassay (INCSTAR Corp., Stillwater, MN). Plasma IGF-I was measured after acid-ethanol extraction using an immunoradiometric assay (Diagnostic Systems Laboratories, Webster, TX).
Mutation analysis. Genomic DNA was isolated from peripheral
blood samples taken from the propositus, her three siblings, parents,
and extended family members after informed consent (Fig. 2). Haplotype analysis with the markers
D21S1912 (GDB Accession ID: 665559), D21S171 (GDB ID: 60660) and PFKL
(GDB ID: 60455) was performed using Human MapPairs and PCR conditions
from Research Genetics, Inc., Genome Services protocol
(Research Genetics, Inc., Huntsville, AL). The
AIRE gene (GenBank accession no. AB006684) is located just
centromeric (<10 kb) to the PFKL gene and approximately 150 kb from
the other two flanking markers noted above. Exons 2, 6, 8, and 10 of
the AIRE gene, together with adjacent intronic sequences
were amplified using primers and conditions specified by the Finnish
German APECED Consortium
(http://chr21.rz-berlin.mpg.de/APECED.html) and a Gene Amp
9600 PCR system (Perkin Elmer, Cetus Instruments, Norwalk,
CT). The PCR products were then purified with the Qia-quick PCR
purification columns kit (Qiagen Inc., Ontario, Canada)
according to the manufacturer’s instructions, directly sequenced by
the dideoxy nucleotide chain termination method (Thermo Sequenase
terminator cycle sequencing kit, Amersham Life Science),
and reaction products were run on a 6% denaturing polyacrylamide gel.
The exon 2 PCR product containing the mutant PstI
restriction site in the heterozygous form was subcloned into
pBluescript II KS (Stratagene, La Jolla, CA). Plasmids
(0.25 pmol) with either the mutant or the normal allele were then
directly sequenced in the manner described above.
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Results |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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Haplotype analysis suggested compound heterozygosity at the
AIRE locus (Fig. 2). Direct sequencing of exon 8 revealed a
previously described mutation, a deletion of nucleotides 1085–1097
(EMBL Accession no: mRNA Z97990) found in the index patient, her
affected sister, and her unaffected sister. This 13-bp deletion
mutation was of maternal origin, presumably inherited from the maternal
grandfather. It results in a frameshift and produces a truncated 371
amino-acid protein. The defect is predicted to lead to the loss of at
least one of the two AIRE PHD fingers (7).
Direct sequencing of exons 6 and 10 failed to show any of the
previously described or any novel mutations. However, a TG
transversion at nucleotide position 398, resulting in a leuarg amino
acid substitution (L93R) was detected in exon 2. The mutation was found
in the father, the brother, the index patient, and the affected sister.
The unaffected sister and mother were not carriers of the mutation. The
presence of the mutation in the propositus was confirmed by cloning of
PCR product from genomic DNA (Fig. 3).
The mutation destroys a PstI restriction-enzyme site. Using
PstI digestion, its presence was confirmed in the index
patient, the clinically affected sister, the father, the brother, one
paternal uncle, and the paternal grandfather. Screening of 50
French-Canadian controls with PstI digestion did not show
destruction of the restriction-enzyme site. Thus, none of the controls
were carriers of the novel mutation.
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An outline of the response to CyA is shown in Table 2. The pancreatic dysfunction and
ectodermal dysplasia responded dramatically to therapy. Stimulated
pancreatic lipase increased 24-fold, accompanied by a marked decrease
in the quantity of stool fat and requirements for pancreatic enzyme
supplements. Nocturnal gavage feedings were successfully discontinued 5
months after CyA. The patient also reported marked improvement in her
symptoms of abdominal bloating, diarrhea, and cramps. Eight months
after initiating CyA, ophthalmological re-evaluation showed no evidence
of new corneal ulcerations. The severe photophobia resolved completely,
obviating the need for artificial tears and vitamin A ointment. Hair
regrowth was apparent diffusely, most notably on the scalp (Fig. 1C).
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Hypomagnesemia (1.5 mg/dL, N = 1.7–2.4) and hyperuricemia (147 ng/mL, N = 55–110) developed as a result of the CyA therapy. The patient was treated with magnesium supplements and allopurinol (100 mg twice daily) with complete normalization of these parameters. Serum urea and creatinine were within normal limits both before and after CyA. The glomerular filtration rate (GFR) by 99Technetium DTPA scanning before initiation of immunosuppressant therapy was 150 cc/min/1.73 m2, followed by 100 cc/min/1.73 m2 after 8 months of treatment (N = 125 ± 13).
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Discussion |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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APECED complicated by steatorrhea is well-documented, as it occurs in up to 24% of cases in the Finnish population (1). However, the etiology of fat malabsorption in this setting is poorly defined. Malabsorption was initially attributed to hypoparathyroid-induced hypocalcemia (22); more recently it has been proposed that malabsorption is the primary event that subsequently leads to poor absorption of calcium and vitamin D and exacerbation of diarrhea (23, 24). Steatorrhea in APECED has also been linked to intestinal infection (23). In the present case, bacterial overgrowth of the small bowel was a contributing, though not the only, factor. Bereket et al. (24) report a patient with APECED-associated steatorrhea who had histological evidence of dilated lacteals in the presence of normal villous architecture. However, other clinical features of intestinal lymphangiectasia, such as edema, hypoproteinemia, and lymphocytopenia were absent, thus weakening the possibility that lymphangiectasia was the sole source of the patient’s malabsorption. Steatorrhea has also been associated with autoimmune enteropathy, recently described in a patient with APECED, intractable diarrhea and hypocalcemia unresponsive to supplementation (25). Villous atrophy and mild subacute inflammation were present on intestinal biopsy, with positive serum autoantibodies to the brush border of normal gut enterocytes. This patient was successfully treated with iv high-dose methylprednisolone and maintenance oral methotrexate (25). Interestingly, pancreatic function tests were consistent with mild pancreatic insufficiency, which was unresponsive to pancreatic enzyme replacement. Evidence for autoimmune destruction of the exocrine pancreas has been suggested by others: Scirè et al. (23) reported a 15-yr-old girl with APECED and exocrine pancreatic insufficiency who was successfully treated with enzyme replacement and in whom pancreatic, smooth muscle, and mitochondrial auto-antibodies were present.
In the present report, exocrine pancreatic insufficiency and duodenal inflammation heralded the onset of islet-cell failure in the presence of anti-ICAs. This temporal association suggests that the pathogenesis of the exocrine and endocrine failure, as well as the enteritis, may have a common mechanism. Recently, antibodies to the autoantigen tryptophan hydroxylase have been identified in the sera of patients with APECED (21). Tryptophan hydroxylase is a 230 kDa tetramer that is expressed in serotonin-producing cells of the central nervous system and of the intestine. In the gut, tryptophan hydroxylase and serotonin are present in enterochromaffin cells of the mucosa, the neuronal cells of the submucosa, and in the myenteric plexus. Ekwall et al. (21) reported that patients with APECED-associated gastrointestinal dysfunction and positive tryptophan hydroxylase antibodies had complete absence of enterochromaffin cells on duodenal biopsy, as was noted in our patient. Antitryptophan hydroxylase antibodies have been found to almost completely inhibit the enzyme (21), which converts L-tryptophan to 5-hydroxy-L-tryptophan, the precursor of serotonin. Intestinal serotonin depletion has been shown in mice to result in diarrhea (26), and serotonin receptors have been found in ganglia and acinar nerves of the pancreas (26). Communication between the gut and the pancreas via serotonin is suggested by the finding that serotoninergic enteropancreatic axons terminate near the pancreatic serotonin receptors (27). The absence of enterochromaffin cells in the gut of patients with APECED-related intestinal dysfunction and the occurrence of pancreatic exocrine as well as endocrine insufficiency in these patients raises the possibility that abnormal regulation of serotonin plays a role in the development of both the gastrointestinal and pancreatic disease.
The risk of developing type 1 diabetes has clearly been associated with
the occurrence of high antibody titers, including anti-ICAs, anti-GAD
antibodies, anti-IA-2 and anti-insulin antibodies (28). Within the
context of APECED, however, their significance is harder to evaluate,
as anti-ICA and anti-GAD antibodies can be present for many years
without progression to overt diabetes (29). High anti-ICAs were
identified in our patient at the age of 9.6 yr, and progression to
diabetes mellitus, as recently defined (30), was evident by the age of
11 yr. Anti-AADC antibodies have also been implicated in the
pathogenesis of diabetes mellitus (20) and were positive in our
patient. Like GAD, AADC is a pyridoxal phosphate-dependent enzyme that
catalyzes the decarboxylation of amino acids in the production of
neurotransmitters (the former leads to 
-aminobutyric acid
production, while the latter produces serotonin). Thus the AADC
antigen, located in the pancreas, liver, and enterochromaffin cells of
the intestine, may also be involved in the pathogenesis of the
gastrointestinal and the pancreatic dysfunction (20).
It is noteworthy that insulin-dependent diabetes mellitus is seen in up to 18% of APECED patients, in contrast to the higher prevalence of most of the other disease components (3). In contrast to isolated diabetes mellitus type 1 and diabetes in the context of types 2 and 3 autoimmune polyglandular syndromes, no linkage to the histocompatibility leukocyte antigen (HLA)-DR/DQ alleles has been demon- strated (29, 31, 32), although questionable linkage of APECED to the HLA-A region has been described in the Finnish population (33), and linkage to the HLA-B region has been seen among North Americans (31). We have therefore explored the possible contribution of other loci implicated in the genetic susceptibility for development of type 1 diabetes. The genotyping of the insulin-dependent diabetes mellitus type 2 locus as previously described (34, 35) revealed that our index patient was homozygous for the class I susceptibility alleles (814 bp, 828 bp), whereas her siblings were heterozygous for the class I and III alleles (data not shown). Class III alleles are dominantly protective in the development of type 1 diabetes mellitus. While our intention is not to imply causality, it is of interest that none of the siblings manifest type 1 diabetes to date, including the sister with APECED, thus future studies should seek to define the IDDM2 locus in APECED.
Autoimmune hypophysitis is another rare feature of this patient’s disease. Lymphocytic hypophysitis classically presents as a pituitary mass in young women in late pregnancy or early post-partum. It preferentially causes ACTH deficiency, sometimes in isolation or with TSH deficiency, in contrast to this child with isolated GH-deficiency (36). Antipituitary antibodies in the present case to a 43–45 kDa membrane protein were more in keeping with previous findings in GH-deficient children (17) than were autoantibodies to a 49 kDa cytosolic protein in women with hypophysitis (37). The ring-enhancement on magnetic resonance imaging is highly suggestive of autoimmune hypophysitis (38), but the immunological mechanism may be different between patients with APECED and those with polyglandular autoimmune disease type II. It is interesting to note that immunoblotting in the index case showed reactivity to a protein in the microsomal fraction of human liver of 45–47 kDa and to a pituitary membrane protein of 43–45 kDa, suggesting the possibility of a common antigen in both tissues. The migration of this antigen appears to be distinct from that of P450 1A2 (52 kDa), another hepatic autoantigen recently reported in at least two patients with APECED and autoimmune hepatitis (39).
Hepatitis, although a rare feature of APECED, has been associated with fulminant hepatic failure, accounting for 25% of deaths in the Finnish group of patients (1). Our patient’s aspartate aminotransferase and alanine aminotransferase levels have returned to normal on CyA. Immunosuppressive therapy has been used to halt the liver disease in APECED, but transplantation has ultimately been necessary in some cases (40). Similarly, while oral CyA has been used for isolated autoimmune enteropathy (12) and hepatitis (41), as well as for a number of other autoimmune diseases (42) including Behcet’s syndrome, uveitis, psoriasis, atopic dermatitis, rheumatoid arthritis, Crohn’s disease, and nephrotic syndrome, its use for APECED-associated pancreatic exocrine insufficiency has not, to our knowledge, been reported.
Our experience suggests that CyA is useful in the treatment of APECED, although we would only recommend its use for specific components of the illness. The gastrointestinal dysfunction, alopecia universalis, and keratoconjunctivitis responded dramatically to the immunosuppressive therapy: these aspects of the disease are associated with organ systems in which there is a high cell turnover. It is possible that the regenerative capacity of the gut, the hair follicle, and the corneal epithelium is more amenable to therapy for this reason. In this report, it does not appear that CyA altered the course of at least two of the endocrinopathies, as progression to primary ovarian and adrenal failure ensued over the 8 months of therapy.
Although it has been shown that the remission period in patients with
non-APECED-associated diabetes mellitus type 1 is prolonged when oral
CyA is administered early in the course of the disease, it has been
clearly demonstrated that this is effective only temporarily in
patients with both preclinical and established diabetes mellitus (43),
and clinical trials in this instance have been abandoned. Our patient
developed hyperinsulinemia after the initiation of subcutaneous,
long-acting insulin and then CyA. This may, in part, be explained by
GH-replacement therapy, which was initiated after the pre-CyA
intravenous glucose tolerance test presented in Table 2 and was
maintained in conjunction with CyA treatment. GH replacement in
GH-deficient children is known to increase insulin secretion and
hepatic insulin resistance, although peripheral insulin sensitivity
appears to be unchanged (44, 45). Anti-insulin receptor and
anti-insulin antibodies in the context of multisystem autoimmune
disorders are associated with hyperinsulinemia and insulin resistance
(46), a possible explanation in this case which remains to be verified.
Given that the pathogenesis of diabetes mellitus in APECED may be
different than in isolated diabetes mellitus or diabetes associated
with types 2 and 3 polyglandular syndromes, it is possible that the
effect of CyA may also be different in APECED; that is, the positive
effect of immunosuppression may persist longer. However, this remains
to be confirmed in our patient and in others with APECED-associated
diabetes mellitus and will be difficult to prove given the paucity of
patients.
Our patient experienced minimal short-term side effects of the therapy, including hypomagnesemia and hyperuricemia. Both were easily remedied with supplementation of magnesium and allopurinol. Although the GFR fell during the 8 months of treatment, the pre-CyA GFR was supranormal, possibly secondary to renal hyperfiltration seen in the early course of diabetes mellitus. The assessment one year later on CyA was in the normal range. Renal dysfunction is relatively common during CyA therapy, though serious and irreversible damage is rare (42). No other adverse effects of the therapy were noted during the 8-month period. Common side effects such as neurological and gastrointestinal symptoms, when they do occur, are usually mild to moderate and resolve on dosage reduction. In the present report, trough CyA levels were maintained between 200–300 ng/mL on 5 mg/kg/day for the first 8 months; we now aim for a nadir (pre-dose) level of 100 ng/mL, with a lowering of the CyA dose to 3.5 mg/kg/day. From the patient’s and family’s perspective, the marked improvement in vision and the ameliorated body image are worth the risks of low dose CyA therapy.
The recent cloning of the AIRE gene has permitted investigation of the molecular basis of APECED in a number of patients from various geo-ethnic groups. The AIRE gene encodes a nuclear protein containing two zinc-finger (PHD-finger) motifs, suggestive of a transcription factor. Eight mutations have been reported (7, 8, 9, 10), and here we report a novel missense mutation in exon 2, the same exon in which another missense mutation (K83E) has been described in the Finnish population (8). The mutation we describe also predicts a change in the protein conformation as a nonpolar amino acid (leucine) is replaced by a polar (arginine) amino acid. We also report the common 13-bp deletion mutation (1085–1097) that results in a frameshift producing a truncated 371 amino-acid protein. This mutation has been previously identified in a number of populations (British, Dutch, German, Northern Italian, Finnish, New Zealand, and American) (8, 9, 10). Haplotype analysis suggests that the 13-bp deletion in this French-Canadian family is not due to the founder effect; rather, it appears to represent an independent mutational event (8, 9). It has been proposed that the AIRE gene contains hypermutable sites, accounting for the recurrence of the 13-bp deletion mutation and the Finnish major mutation (R257X, producing a truncated protein) in patients from a variety of geographical locations and ethnic groups (9).
Genotype-phenotype correlations have been difficult to ascertain to date, as there is tremendous variability in phenotypes among family members with APECED (1, 47). Furthermore, Scott et al. (9) noted that although the X546C mutation, which is expected to produce a functional protein, was found in compound heterozygozity with the R257X mutation in two Finnish patients, the clinical features of patients with this mutation were indistinguishable from the phenotype of the Finnish patients who were homozygous for the R257X mutation, which is expected to lead to complete loss of function of the zinc-finger protein. Phenotype-genotype correlations may not be possible until long-term follow-up establishes the full manifestations of the disease, as it tends to evolve over years. Our index patient was severely affected at a young age, suggesting that compound heterozygozity for the novel missense mutation and the common 13-bp deletion may be associated with a particularly difficult clinical course.
In summary, we describe a French-Canadian adolescent with severe failure to thrive in the context of unique features of APECED, including isolated GH deficiency secondary to autoimmune hypophysitis and exocrine pancreatic insufficiency. It is apparent that immunosuppressive therapy is generally underused in severe APECED, and we recommend a trial of CyA in the treatment of APECED-associated exocrine pancreatic failure and keratoconjuctivitis. The genetic analyses of this French-Canadian pedigree revealed a novel missense mutation in exon 2 as well as the common 13-bp deletion mutation in exon 8 of the AIRE gene. Haplotype analysis suggests the 13-bp deletion represents an independent mutational event.
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Acknowledgments |
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Footnotes |
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Received November 4, 1998.
Revised December 17, 1998.
Accepted January 5, 1999.
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References |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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J. Perheentupa Autoimmune Polyendocrinopathy-Candidiasis-Ectodermal Dystrophy J. Clin. Endocrinol. Metab., August 1, 2006; 91(8): 2843 - 2850. [Abstract] [Full Text] [PDF]
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T. Ulinski, L. Perrin, M. Morris, M. Houang, S. Cabrol, C. Grapin, N. Chabbert-Buffet, A. Bensman, G. Deschenes, and I. Giurgea Autoimmune Polyendocrinopathy-Candidiasis-Ectodermal Dystrophy Syndrome with Renal Failure: Impact of Posttransplant Immunosuppression on Disease Activity J. Clin. Endocrinol. Metab., January 1, 2006; 91(1): 192 - 195. [Abstract] [Full Text] [PDF]
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 W. Jiang, M. S. Anderson, R. Bronson, D. Mathis, and C. Benoist Modifier loci condition autoimmunity provoked by Aire deficiency J. Exp. Med., September 19, 2005; 202(6): 805 - 815. [Abstract] [Full Text] [PDF]
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C. Cocco, A. Meloni, F. Boi, G. Pinna, R. Possenti, S. Mariotti, and G.-L. Ferri Median Eminence Dopaminergic Nerve Terminals: A Novel Target in Autoimmune Polyendocrine Syndrome? J. Clin. Endocrinol. Metab., July 1, 2005; 90(7): 4108 - 4111. [Abstract] [Full Text] [PDF]
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R. Perry, O. Kecha, J. Paquette, C. Huot, G. Van Vliet, and C. Deal Primary Adrenal Insufficiency in Children: Twenty Years Experience at the Sainte-Justine Hospital, Montreal J. Clin. Endocrinol. Metab., June 1, 2005; 90(6): 3243 - 3250. [Abstract] [Full Text] [PDF]
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F. Skoldberg, G. M. Portela-Gomes, L. Grimelius, G. Nilsson, J. Perheentupa, C. Betterle, E. S. Husebye, J. Gustafsson, A. Ronnblom, F. Rorsman, et al. Histidine Decarboxylase, a Pyridoxal Phosphate-Dependent Enzyme, Is an Autoantigen of Gastric Enterochromaffin-Like Cells J. Clin. Endocrinol. Metab., April 1, 2003; 88(4): 1445 - 1452. [Abstract] [Full Text] [PDF]
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 C. Betterle, C. Dal Pra, F. Mantero, and R. Zanchetta Autoimmune Adrenal Insufficiency and Autoimmune Polyendocrine Syndromes: Autoantibodies, Autoantigens, and Their Applicability in Diagnosis and Disease Prediction Endocr. Rev., June 1, 2002; 23(3): 327 - 364. [Abstract] [Full Text] [PDF]
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A. Meloni, R. Perniola, V. Faa, E. Corvaglia, A. Cao, and M. C. Rosatelli Delineation of the Molecular Defects in the AIRE Gene in Autoimmune Polyendocrinopathy-Candidiasis-Ectodermal Dystrophy Patients from Southern Italy J. Clin. Endocrinol. Metab., February 1, 2002; 87(2): 841 - 846. [Abstract] [Full Text] [PDF]
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 W. Creutzfeldt, O. Ekwall, F. Rorsman, O. Kampe, C. Hogenauer, G. J. Netto, and J. S. Fordtran Malabsorption Due to Cholecystokinin Deficiency in a Patient with Autoimmune Polyglandular Syndrome Type I N. Engl. J. Med., July 5, 2001; 345(1): 64 - 66. [Full Text] [PDF]
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 M. Halonen, M. Pelto–Huikko, P. Eskelin, L. Peltonen, I. Ulmanen, and M. Kolmer Subcellular Location and Expression Pattern of Autoimmune Regulator (Aire), the Mouse Orthologue for Human Gene Defective in Autoimmune Polyendocrinopathy Candidiasis Ectodermal Dystrophy (APECED) J. Histochem. Cytochem., February 1, 2001; 49(2): 197 - 208. [Abstract] [Full Text]
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O. Ekwall, J. Haavik, J. Perheentupa, C. Betterle, J. Gustafsson, E. Husebye, F. Rorsman, and O. Kämpe Pteridin-Dependent Hydroxylases as Autoantigens in Autoimmune Polyendocrine Syndrome Type I J. Clin. Endocrinol. Metab., August 1, 2000; 85(8): 2944 - 2950. [Abstract] [Full Text]
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