Tufting Enteropathy and Chronic Arthritis: A Newly Recognized Association With a Novel <i>EpCAM</i> Gene Mutation
AbstractIntractable diarrhea of infancy is a descriptive term rather than a specific diagnosis. Patients usually present in the neonatal period with severe intractable watery diarrhea, malabsorption, and failure to thrive. It is etiologically heterogeneous but is often infectious or inflammatory in origin (1). Tufting enteropathy, also known as intestinal epithelial dysplasia, is one of the causes of congenital intractable diarrhea and is characterized by persistent villous atrophy and a pathognomonic histological appearance (1–3). Recently, mutations in the epithelial cell adhesion molecule (EpCAM) gene were identified as the responsible genetic defect in patients with tufting enteropathy (4). The coexistence of intractable diarrhea of infancy and chronic arthritis is rare and has limited possible underlying causes (5,6). To the best of our knowledge, there are no previous reports of chronic arthritis in children with tufting enteropathy. We describe the clinical, biochemical, molecular, and radiological analysis as well as treatment and outcome of 4 children with tufting enteropathy seen at our hospital. METHODS We retrospectively reviewed data of children with tufting enteropathy and chronic arthritis obtained at King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia. Data included demographics, clinical presentation, laboratory parameters, radiological and histopathological features, limited molecular analysis, treatment, disease course, and outcome of these patients. DNA Extraction Genomic DNA was extracted from whole blood anticoagulated with ethylenediaminetetraacidic acid using the Purgene Gentra DNA Extraction Kit (Cat. # D-5000, Gentra Systems, Minneapolis, MN) according to the manufacturer's instructions. The DNA was quantified spectrophotometrically and stored in aliquots at −20°C until required. Polymerase Chain Reaction and Direct Sequencing Polymerase chain reaction amplification was performed on a thermocycler (DNA Engine Tetrad. MJ Research, Inc, Waltham, MA) in a total volume of 25 μL, containing 10 ng DNA, 50 mmol/L KCl, 10 mmol/L Tris-HCl (pH 9.0), 1.5 mmol/L MgCl2, 0.1% Triton X-100, 0.25 mmol/L of each deoxyribonucleotide triphosphate, 0.8 μmol/L of each primer and 0.5 U of Taq polymerase (QIAGEN, D-40724, Hilden, Germany). For polymerase chain reaction, an initial denaturation step at 95°C for 10 minutes was followed by 40 cycles of denaturation at 95°C for 30 seconds, annealing at 59°C for 30 seconds and extension at 72°C for 30 seconds followed by a final extension step of 72°C for 10 minutes. The primers used for amplification of the coding regions of EpCAM are available upon request. Genomic DNA from the patients were screened for mutations in the exons and splice sites of EpCAM by using the BigDye Terminator v3.1 Cycle Sequencing Kit from Applied Biosystems, and samples were loaded onto ABI PRISM 3730XL DNA analyzer system (Applied Biosystems, Foster City, CA), following the manufacturer's instructions. Sequence analysis was performed using the SeqManII module of the Lasergene (DNA Star Inc, Madison, WI) software package, then compared with a reference sequence. RESULTS Four patients (2 males and 2 females) from 2 unrelated families who were referred with infantile intractable diarrhea were found to have tufting enteropathy. The parents of these patients were first cousins and healthy. All patients who presented at the age of 1 to 2 weeks were diagnosed with intractable diarrhea, electrolyte disturbance, abdominal distension, and failure to thrive (growth parameters were significantly below the fifth percentiles). They were fed different formula but did not show any improvement and eventually all required total parenteral nutrition (TPN). There was no dysmorphism noted on examination. The initial laboratory findings showed normal complete blood count and electrolyte disturbance (mainly low sodium, chloride, magnesium, and calcium). Liver function tests were normal, but 1 patient had progressive liver function derangement probably because of TPN. Serum immunoglobulin levels, sweat chloride test, and thyroid function test were normal. Autoimmune screen including antimitochondrial, antismooth muscle, and celiac disease panel were negative. Although 2 patients were weakly positive for anti-nuclear antibody titer, they were negative for extractable nuclear antigens. All cultures including stool culture were negative. Small bowel and colonic biopsies showed subtotal villous atrophy associated with mild crypt hyperplasia. The lamina propria contained a relatively normal number of chronic inflammatory cells. There was no chronic crypt damage or active duodenitis. The intraepithelial lymphocytes were not increased. Interestingly, the luminal enterocytes were closely packed and showed characteristic tufting with rounding of their apical cytoplasm. These findings were consistent with tufting enteropathy (Fig. 1).FIG. 1: Subtotal villous atrophy with crypt hyperplasia and tufting of the epithelial surface.Unfortunately, 2 patients (1 from each family) died in the infancy period because of the intractable diarrhea and sepsis (1 additionally had progressive liver failure secondary to TPN). The other 2 patients are still alive but continue to have intractable diarrhea and failure to thrive. Interestingly, at around 4 years of age, both started to develop morning stiffness, pain, and swelling in multiple joints including the interphalangeal and metacarpophalangeal joints of hands, wrists, and knees. During follow-up it was noticed that both had progressive contractures associated with flexor and extensor tenosynovitis of the hands but no evidence of enthsitis or other associated systemic symptoms. The surviving patients had elevated erythrocyte sedimentation rate and C-reactive protein. Urine and stool cultures were negative. X-ray of affected joints showed generalized osteopenia with increased trabeculation especially at the epiphysis of the metacarpophalangeal joints and carpal bones with synovial hypertrophy but no erosive changes. A novel homozygous single base pair insertion was identified in exon 5 of EpCAM in 1 of the 2 surviving patients (c.498insC). The resulting frameshift introduces 21 novel amino acids followed by premature truncation of the protein (Q167Pfsx21). Nonsteroidal anti-inflammatory drugs, prednisone, sulfasalazine followed by methotrexate were tried but the response was poor. However, the addition of antitumor necrosis factor (Etanercept) resulted in resolution of the joint pain, swelling, and tenosynovitis as well as significant improvement in the range of motion of the affected joints. Fortunately, we were able to wean off prednisone, methotrexate, and etanercept. During follow-up it was noticed that both had inactive arthritis and low inflammatory markers. There has been some improvement in their bowel habit; it became less frequent than before and both were tolerating and continued to be TPN free. DISCUSSION Tufting enteropathy is a rare autosomal recessive form of intractable diarrhea of infancy. Typically, patients develop watery diarrhea within the first days after birth and have the characteristic appearance of subtotal villous atrophy with crypt hyperplasia and tufting of the epithelial surface (7). It is more frequent in patients of Arabic origin probably because of a higher degree of consanguinity (7). Sivagnanam et al (4) identified EpCAM responsible gene for tufting enteropathy. They speculated that EpCAM plays an important role in the normal development of the crypt villus axis, where epithelial cells originate from stem cells in the crypt and migrate distally to the tip of the villous before shedding, so that perturbation of this developmental process may underlie the pathogenesis of tufting enteropathy in patients with EpCAM mutations. Several cases of tufting enteropathy have been reported as being associated with phenotypic abnormalities (8). The association between tufting enteropathy and autoimmune disorders has only been documented in a recent case report that described autoimmune anemia, thrombocytopenia, and skeletal dysplasia but no associated arthritis (9). The 4 patients described in this report have the typical clinical and histological features of tufting enteropathy; 2 of them later developed chronic inflammatory arthritis in multiple joints at the age of 4 years. This previously undescribed observation was not seen in their siblings, but that could be because of their early death in infancy. Both had active arthritis in small and large joints as well as tenosynovitis associated with elevated inflammatory markers, and radiological changes are consistent with chronic arthritic changes. Fortunately, their musculoskeletal symptoms responded well to the intensive therapy. Our patients did not have evidence of systemic disease despite extensive workup, although their autoimmune workup was inconclusive. Of note, bowel biopsy did not show any inflammatory cell infiltrate in the lamina propria, which makes the possibility of autoimmune enteropathy unlikely. The factors responsible for the late initiation and perpetuation of arthritis in our patients are unclear. Although no enteric pathogens were isolated, the possible roles of enteric infection or allergic reactions to food absorbed across an inflamed mucosa remain speculative. Rheumatic manifestations of diseases primarily of the gastrointestinal tract are frequent and variable. These manifestations include arthralgia, transient or chronic arthritis, and osteoporosis. The association of chronic diarrhea and arthritis in early childhood may lead to suspect limited differential diagnoses including inflammatory bowel disease, which is a familial disease that can affect all age groups, although it has not been well described in infants. Patients with inflammatory bowel disease may present with isolated chronic diarrhea but go on to develop arthritis later in life (10,11). Celiac disease is a primary gastrointestinal disease with a wide clinical spectrum that sometimes includes polyarthritis (12). There are rare conditions characterized clinically by chronic diarrhea and nondestructive arthritis. Biopsies from the colorectal mucosa showed a thickened subepithelial collagen layer consistent with collagenous colitis (13). Infantile systemic hyalinosis is a rare inherited disease of connective tissue characterized by widespread deposition of hyaline material in skin, musculoskeletal, and deep organs including the gut in addition to nodule formation and joint contractures. Most of the patients failed to thrive, with severe muscle wasting and intractable diarrhea (6). Juvenile idiopathic arthritis is the most frequent form of chronic arthritis in children. Juvenile idiopathic arthritis rarely occurs as a familial disease, although detailed family studies of patients and first-degree relatives have been published (14). Systemic onset subtype of juvenile idiopathic arthritis may associate with abdominal pain because of peritonitis but never associate with chronic diarrhea. Our clinical observation indicates that patients with tufting enteropathy are prone to develop chronic arthritis, and this report should increase the awareness of this association, especially in older children, and help in the identification of similar cases. Our ability to speculate on a possible genotype-phenotype correlation is limited by the fact that we have molecular data on only 1 of the 2 patients with this apparently novel association. Future clinical reports on tufting enteropathy with mutation information will be necessary to test for such correlations.
https://doi.org/10.1097/mpg.0b013e3181acaeae