Analysis of Diagnosing Methods for Celiac Disease By: Lauren Rieves 3562W 007

Table of Contents
Abstract Introduction Methods Results Discussion Conclusion 3 3 7 8 12 12

Abstract: Celiac disease has dramatically increased worldwide over the past decade, primarily due to the advances in diagnosing methods. This research aims to find the most effective diagnosing method for celiac disease through comparing and contrasting a variety of methods. I looked at a variety of research studies that provided specificity and sensitivity values for five main blood tests that are used to determine celiac status. Currently, a definite celiac diagnosis can only come from an intestinal biopsy. Therefore, I also looked at the pros and cons of the biopsy method. As a result of my research I found that the EMA and tTG- IgA blood tests have the highest specificity and sensitivity values, and therefore are generally the best method for diagnosing celiac disease without a biopsy. However, I also concluded that other tests may be best for certain patients that are under 2 years of age, or those that are IgA deficient. The intestine biopsy method is the gold standard because it will positively diagnose everyone with celiac disease and negatively diagnose everyone that does not have celiac disease. However, I believe with more research and diagnostic developments , we can discover an accurate and noninvasive diagnosing method for celiac disease.

Introduction Celiac disease is one of the most common gastrointestinal disorders, with a prevalence of 1:100-1:200 in the general population(Brusca et al). There has been a large increase in the number of celiac diagnoses in the past decade because of improved

availability to diagnosing methods. This analytical paper aims to compare and contrast a variety of celiac disease diagnosing methods in order to determine the most effective procedure. I specifically compare several blood/serum assays to the intestinal biopsy procedure. This paper will outline general information on the disease and provide a detailed description of each diagnosis method along with its pros and cons. Overall effectiveness of the methods is compared using specificity and sensitivity. Specificity is the probability of obtaining a negative test result for someone that does not have celiac disease. Sensitivity is the probability of getting a positive test result for a patient with celiac disease. Lastly, I will analyze the methods specificity and sensitivity values to determine the most effective method for diagnosing. Simply put, celiac disease (CD) is both a food intolerance and an autoimmune disorder. It is intolerance to gluten, which is found in wheat, barley and rye. When an individual with CD consumes food with these components they experience decreased digestion, decreased absorption of all nutrients and increased secretion of water and solute (Rostom et al). CD patients can cause serious harm to their small intestine by consuming wheat, barley or rye. This damage to the villi and lining of the intestine is referred to as atrophy and is categorized into normal, partial atrophy I, II, III, sub atrophy and total atrophy. Intestinal damage can also be indicated using the Marsh scale, which is outlined in figure 3. Research has shown that CD is a result of inappropriate T cell-mediated immune responses to consuming gluten (Fasano, Catassi). Fortunately, it is the only treatable autoimmune disease. Treatment entails committing to a gluten-free diet for the remainder of life; I will discuss treatment more at the end of the introduction. Genetics

plays a large role in who develops CD. Individuals with immediate relatives that have been diagnosed should be tested. CD is one of the most frequent genetically based diseases (Fasano, Catassi). There is no genetic test to diagnose CD, but there are tests for the HLA DQ2 and HLA DQ8 genes. These genes are associated with celiac. However, 40% of North Americans have these genes but only .5-1% will develop CD (Blood Testing for Celiac Disease). Also, those that exhibit classic symptoms such as chronic diarrhea, malabsorption, failure to thrive (in children) and abdominal distention should be tested. Atypical symptoms include anemia, hyper/hypothyroidism, abdominal pain and osteopenia (Fasano, Catassi). A complete list of symptoms is located below in figure 1. Patients and relatives of patients with type-1 diabetes, thyroid or liver disorders and patients with connective tissue diseases, Down or Turner syndrome should all be tested and are considered to be at-risk (Fasano, Catassi). Celiac disease occurs because of the interactions involving by-products of dietary grains, immune factors, and an individuals genetic makeup (Rostom et al). Typical symptoms generally occur between 6 and 18 months of age. On the other hand, CD may be silent for years and not emerge until adulthood (Fasano, Catassi). As mentioned before, the only treatment of CD is eliminating all wheat, barley and rye from the diet. A gluten-free diet (GFD) will result in normal serologic test results after 6 to 12 months. Serological refers to the analysis of ones blood serum. All symptoms will improve on a GFD (Rostom et al). Some doctors will screen their patients for celiac. This is a progressive procedure because it identifies asymptomatic CD individuals. In Western countries, for each diagnosed case of CD, an average of 5-10 cases remain

undiagnosed. Clearly, accurate diagnosis of celiac disease is imperative. Described next is the process I used to determine the best method of diagnosis. Figure 1- Symptoms and Conditions related to Celiac Disease (Fasano, Catassi) Typical symptoms Chronic diarrhea Failure to thrive Abdominal distention Atypical symptoms Secondary to malabsorption Sideropenic anemia Short stature Osteopenia Recurrent abortions Hepatic steatosis Recurrent abdominal pain Gaseousness Independent of malabsorption Dermatitis herpetiformis Dental enamel hypoplasia Ataxia Alopecia Primary biliary cirrhosis Isolated hypertranaminasemia Recurrent aphthous stomatitits Myasthenia gravis Recurrent pericarditis Psoriasis Polyneuropathy Epilepsy Vasculitis Dilatative cardiomyopathy Hyper/hypothyroidism Associated conditions Possibly gluten dependent IDDM Autoimmune thyroiditis Autoimmune hepatitits Sjogren syndrome Addison disease Autoimmune atrophic gastritis Autoimmune emocytopenic diseases Gluten Independent Down syndrome Turner syndrome Williams syndrome Congenital heart defects IgA deficiency

Methods: Because CD is a rapidly growing health issue, there is an abundance of research on the diagnosing methods. The goal of my research was to compare different diagnosing

methods for celiac disease to determine the most effective method. For this paper, most effective and best refer to the sensitivity and specificity levels that are closest to 100. In the beginning of my research it became clear that specificity and sensitivity would be the most effective means of comparing different methods. As a reminder, specificity is the probability of obtaining a negative test result for someone that does not have celiac disease. Sensitivity is the probability of getting a positive test result for a patient with celiac disease. An ideal test would have 100 percent specificity and 100 percent sensitivity. I primarily used the PubMed database through the U of M library website and searched for titles including the words celiac and diagnosis. After I had a good knowledge base of the topic, I refined the search results by searching for titles with specific assays, such as AGA. Most of the sources I used were empirical and provided their findings in a specificity and sensitivity fashion. However, it was a priority to triangulate my materials so I also incorporated some non-empirical web based sources. These proved useful for providing background information in the introduction. I found empirical resources to be the most useful for comparing methods because,unfortunately, I dont have the resources to conduct a study myself. To determine which method of diagnosing CD is the most effective, I looked for the method with the highest levels of specificity and sensitivity. However, research shows that certain blood tests are not well suited for certain groups of people, this will be better explained in the results section. Results: A very common way to test for CD is through blood and serologic assays, which analyze blood and serum for certain enzymes or antibodies. There are a variety of tests that

can be preformed which collectively make up the celiac panel. Blood tests are relatively inexpensive and easy to conduct. However, they are not always accurate in children under 3 years of age. Blood/ serum tests have a 10 percent possibility of resulting in a false positive diagnosis (Blood Testing for Celiac Disease). Prior to having any test done for celiac diagnosis, the patient should consume at least the equivalent of 1-4 slices of bread per day until the test (Blood Testing for Celiac Disease). Blood tests can be divided into two catergories; the immunoglobin A (IgA) class and the immunoglobin G (IgG) class (Provider points:Testing for Celiac Disease) . I specifically looked at 5 main blood tests that can be preformed to diagnose CD and the intestinal biopsy method. Values for the blood tests are found below in figure 2.

EMA First is the EMA serologic test. EMA status is measured using an immunofluorescence technique. This method uses monkey esophagus or human umbilical cord as the tissue substrate. The tissue is stained and observed under a microscope to determine a positive or negative pattern. This test requires more time, money and human labor than some of the other methods (Blood Testing for Celiac Disease). Not long ago the EMA assay was suggested to be the confirmatory diagnosis procedure if the levels are above 100u/mL. This would save patients from going through a biopsy (Ada Aita et al). EMA is highly specific for celiac disease and is considered to be the best method for diagnosing without a biopsy by many professionals.

AGA

Next is an enzyme-linked immunosorbent AGA assay. This is known to be the least valid of diagnosing methods. It is much less accurate than the anti-tTG and EMA assays (Brusca et al). The AGA test may provide false positive results for other gastrointestinal diseases like lactose intolerance (Fasano, Catassi). Because of afore mentioned disadvantages and the AGAs low positive predictive value (PPV), it is no longer recommended for use in diagnosing CD in adults. However, it is useful for testing children under the age of 18 months (Provider points:Testing for Celiac Disease) .

DGP DGP (deamidated gliadin peptides) tests have greatly improved in recent history. They produce higher specificity levels than AGA tests (Brusca et al) and may be the best method for diagnosing CD in patients that are IgA deficient (Provider points:Testing for Celiac Disease) . When combined with the anti-tTG test, accuracy is greatly improved in young children, DGP plus tTG could be the most effective test combination. IgA deficiency occurs in 3-5% of CD patients, this deficiency can cause TTG and EMA results to be falsely negative (Blood Testing for Celiac Disease). DGP should be used if a patient is IgA deficient.

tTGA The tTGA is an enzyme located in and outside of cells and has many functions. tTG is the target of an autoimmune humoral response that results in both secreted and circulating antibodies predominantly of the immunoglobulin (Ig) A isotype (Rostom et al). The tTGA (anti-tissue transglutaminase) test is also less accurate in very young patients, but when celiac autoantibodies IgA-TTG and EMA are found in high amounts within an

individual experiencing typical symptoms, the chance of diagnosing celiac from a small intestine biopsy is nearly 100% (Catassi, Fasano). tTG is measured by an enzyme-linked immunosorbent assay that utilizes guinea pig liver, human recombinant or red-cell derived tTG as the substrate (Rostom et al).

AAA The anti-actin IgA (AAA) assay is useful because this specific antibody concentration is directly related to the degree of intestinal damage (Brusca et al). Therefore, it provides information about the mucosal status without a biopsy (Brusca et al). Figure 2 Serological Result Values (Brusca et al)

tTGIgA Sensitivity 77.9 Specificity 100 PPV 100

Intestinal Biopsy

tTG IgG 54.7 100 100

EMA IgA 77.9 100 100

DGP IgA 45.3 100 100

DGP IgG 60.0 100 100

AGA IgA 45.3 96.4 95.6

AGA IgG 50.5 74.6 77.4

AAA IgA 77.9 83.6 89.2

A histological analysis of small bowel biopsy specimens, initially taken by capsule and then by standard upper endoscopy, has become the gold standard for celiac disease diagnosis (Catassi, Fasano). Changes in the small intestine mucosa can vary from total to partial villous atrophy and are judged based on the Marsh scale (Rostom et al). A description of each level in the Marsh scale is provided below in figure 3. Damage in the intestine can be irregular so its important to take multiple endoscopic biopsy specimens from the small intestine (Rostom et al). Results can be misleading if the disease is erratic,

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not enough specimens are taken or if the biopsy specimen was oriented poorly and tissue sections were cut tangentially (Rostom et al). Figure 3- Marsh Scale (Rostom et al) Marsh 0 Marsh I Normal mucosal and villous architecture Infiltrative Normal mucosal and villous architecture Increased numbers of intraepithelial lymphocytes Hyperplastic Similar to above, but with enlarged crypts and with increased crypt cell division a. Partial villous atrophy Shortened blunt villi Mild lymphocyte infiltration Enlarged hyperplastic crypts b. Subtotal villous atrophy Clearly atrophic villi, but still recognizable Enlarged crypts whose immature epithelial cells are generated at an increased rate Influx of inflammatory cells c. Total villous atrophy Complete loss of villi Severe crypt hyperplastic, and infiltrative inflammatory lesion Hypoplastic Total villous atrophy Normal crypt depth, but hypoplasia Normal intraepithelial lymphocyte count Many feel this does not exist and represents severe malnutrition

Marsh II Marsh III

Marsh IV

Discussion: According to the results presented in figure 2, the tTG-igA and EMA tests are the most accurate for diagnosing CD because of their high sensitivity and specificity values. These are followed by the DGP-IgG and AAA test. AGA has the worst sensitivity and specificity values, the AGA test falsely diagnoses CD 45.3% of the time.

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In general, I can conclude that an intestinal biopsy is the best way to diagnose celiac disease. It is consistently accurate regardless of age, symptoms and other factors. A biopsy will always positively diagnose someone with celiac and will always negatively diagnose someone that does not have celiac. However, since a biopsy is an expensive surgical procedure I think that under specific conditions, the EMA and tTG- IgA tests could be used for a definite diagnosis. On the other hand, there are several factors that need to be taken into consideration. For example, if the patient is under 2 years of age, the DGP, tTG and AGA tests are useful. Also, if the patient is IgA deficient, the DGP and tTG-IgG tests should be utilized. Signs and symptoms that the patient is exhibiting should also be taken into consideration for diagnosing. For the most accurate results, patients must be on a gluten containing diet leading up to the test. Although celiac diagnosing methods have advanced significantly over the past decade, an intestinal biopsy remains the gold standard. I believe with more research we can develop an inclusive, efficient, accurate method for diagnosing CD without a surgical procedure.

Conclusion: During this study I aimed to look at a variety of diagnosing methods for celiac disease. Celiac disease has been a trending epidemic over the last ten years. Because some people may be asymptomatic, screening and effective diagnosing methods are imperative. I concluded that the pre-existing gold standard of an intestinal biopsy is the best way to diagnose CD because it is precise regardless of age and other factors. However, since surgery is expensive and inconvenient a lot of research has been devoted to determining the specificity and sensitivity of blood and serum tests. From analyzing an array of

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research, I determined that the EMA and tTG tests were the best from a specificity and sensitivity point of view. Given the permanent nature of the celiac condition, diagnostic accuracy remains a must (Catassi, Fasano).

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References Ada Aita, Elisa Rossi, Daniela Basso, Graziella Guariso, Dania Bozzato, Michela Pelloso, Matilde Pescarin, Carlo-Federico Zambon, Filippo Navaglia, Eliana Greco, Marco Gasparetto, Paola Fogar, Andrea Padoan, Stefania Moz, Mario Plebani, Chemiluminescence and ELISA-based serum assays for diagnosing and monitoring celiac disease in children: A comparative study, Clinica Chimica Acta, Volume 421, 5 June 2013, Pages 202-207, ISSN 0009-8981, 10.1016/j.cca.2013.03.024. (http://www.sciencedirect.com/science/article/pii/S0009898113001241) Alaa Rostom, Joseph A. Murray, Martin F. Kagnoff, American Gastroenterological Association (AGA) Institute Technical Review on the Diagnosis and Management of Celiac Disease, Gastroenterology, Volume 131, Issue 6, December 2006, Pages 19812002, ISSN 0016-5085, 10.1053/j.gastro.2006.10.004. (http://www.sciencedirect.com/science/article/pii/S001650850602227X) Alessio Fasano, Carlo Catassi, Current Approaches to Diagnosis and Treatment of Celiac Disease: An Evolving Spectrum, Gastroenterology, Volume 120, Issue 3, February 2001, Pages 636-651, ISSN 0016-5085, 10.1053/gast.2001.22123. (http://www.sciencedirect.com/science/article/pii/S0016508501251877) Brusca I, Carroccio A, Bizzaro N, et al. The old and new tests for celiac disease: which is the best test combination to diagnose celiac disease in pediatric patients?. Clinical Chemistry & Laboratory Medicine [serial online]. January 2012;50(Brusca et al):111117. Available from: Academic Search Premier, Ipswich, MA. Accessed April 29, 2013. Canadian Celiac Association. Blood Testing for Celiac Disease. N.p.: Canadian Celiac Association, n.d. Celiac.ca. 2008. Print. 2013. Carlo Catassi, Alessio Fasano, Celiac Disease Diagnosis: Simple Rules Are Better Than Complicated Algorithms, The American Journal of Medicine, Volume 123, Issue 8, August 2010, Pages 691-693, ISSN 0002-9343, 10.1016/j.amjmed.2010.02.019. (http://www.sciencedirect.com/science/article/pii/S0002934310003591) National Institute of Health. Provider Points:Testing for Celiac Disease. N.p.: National Institute of Health, 2008. Celiac.nih.gov. Celiac Disease Awareness Campaign. Web. 2013.

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