Antibody Array 3 Wheat/Gluten Proteome Reactivity & Autoimmunity

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1 2012 Cyrex Labratries, LLC. All Rights Reserved. 1

2 TABLE OF CONTENTS Overview Measuring Gluten-Reactivity Gluten Peptides Opiid Peptides Lectins Enzymes Influencing Factrs Genetic Envirnmental Histry Clinical Systemic Immune Effects Clinical Use f Antibdy Array 3 Clinical Interpretatin f Antibdy Array 3 Specimen Requirement Related Testing References 2012 Cyrex Labratries, LLC. All Rights Reserved. 2

3 WHEAT/GLUTEN PROTEOME REACTIVITY AND AUTOIMMUNITY OVERVIEW MEASURING GLUTEN-REACTIVITY Intrductin Current testing fr Gluten-Reactivity and Celiac disease (CD) includes serum IgG and IgA against gliadin and tissue transglutaminase-2 (ttg2). These antibdies are measured against minr cmpnents f a wheat prtein called alpha-gliadin. Hwever, wheat cnsists f multiple prteins and peptides including, alpha-gliadin, mega-gliadin, glutenin, glutemrphin, prdynrphin, and agglutinins. Any f these antigens has a capacity t challenge the immune system. Because f this hetergeneity f gluten prteins and peptides, multiple variatins in T- cell respnses may ccur against them. Recent medical research indicates that a large number f gluten epitpes, may be implicated in the develpment f Gluten-Reactivity, CD and ther assciated cnditins. The repertire and hierarchy f gluten peptides stimulate the intestinal T- cells and results in a significant elevatin f IgG and IgA prductin. The measurement f IgA and IgG against multiple gluten epitpes in bld can have imprtant implicatins in the accurate diagnsis and design f therapy fr Gluten-Reactive and CD patients. Fr a cmprehensive apprach t this prblem, pineering, patent-pending technlgies have been develped t measure IgA and IgG against varius wheat cmpnents including alphagliadins, -17-mer and native + deamidated -33-mer, gamma-gliadin-15-mer, mega-gliadin-17- mer, glutenin-21-mer, piid peptides (prdynrphin + glutemrphin), gliadin-transglutaminase cmplex, and tissue transglutaminases -2, -3 and -6. Research perfrmed in-huse 1 cnfirms that different Gluten-Reactive and CD patients recgnize an array f gluten antigens. Fr example, ne patient reacts t mega-gliadin, but nt t alphagliadin. The secnd patient reacts t all gliadin peptides, and the third patient reacts nly t the wheat germ agglutinin. Gluten-Reactivity is a systemic autimmune disease with diverse manifestatins. 2 CD r glutensensitive enterpathy, is nly ne aspect f a range f pssible manifestatins f reactivity t gluten. And yet, this enterpathy, ne f the mst cmmn lifelng disrders in bth the U.S. and Eurpe, 3 receives the lin's share f fcus t the pint f ignring ther manifestatins. Autimmune disease, the third leading cause f mrbidity and mrtality in the industrialized wrld, 4 is 10 times mre cmmn in a gluten-sensitive enterpathy than in the general ppulatin. 5 Thus, the burden n sciety frm Gluten-Reactivity cannt be verestimated. Earlier identificatin might result in earlier treatment, better quality f life, and an imprved prgnsis fr these patients. 6 The emphasis n Gluten-Sensitive Enterpathy (Celiac disease) as the main manifestatin f Gluten-Reactivity has been questined. It is nw accepted that Gluten-Reactivity is a systemic illness that can manifest in a range f rgan systems. Such manifestatins can ccur independently f the presence f the classic small-bwel lesin that defines CD. 7 That Gluten- Reactivity is regarded as principally a disease f the small bwel is a histrical miscnceptin Cyrex Labratries, LLC. All Rights Reserved. 3

4 The Gluten-Reactivity has been prpsed t include nt nly CD, but als Gluten-Reactive patients withut mucsal lesins. Frm the skin (Dermatitis Herpetifrmis, 9 Psriatic arthritis, 10 Alpecia areata, Dermatmysitis, Cutaneus vasculitis 11 ), t the muscles (inflammatry mypathies 12 ), t the brain (Gluten Ataxia, 13 altered neurtransmitter prductin, 14 Schizphrenia, 15 peripheral neuralgias, 16 idipathic neurpathies, 17 ) and beynd, pathlgy t gluten expsure can ccur in multiple systems withut evidence f an enterpathy. 2 Negative serlgy shuld nt necessarily reassure the clinician 18 f neither negative immune activatin nr pathlgy. Several reprts shw that in the majrity f Celiac patients, antibdies t gliadin and transglutaminase may be negative In particular, sernegative CD seems t be quite frequent in patients with milder intestinal damage (Marsh I-IIIa lesins). 24 And these lesins ften present withut elevated Celiac markers. Sme reprts identify the sensitivity as lw as 27-31% 25 with lesser degrees f villus atrphy. Patients with nn-villus Atrphy Gluten-Reactivity (Marsh I, Marsh II) are mre likely than thers t test negative fr tissue transglutaminase and endmysial antibdies. 26 Despite the many published reprts n sernegative Celiacs, this subgrup understandably cntinues t be frgtten r nt included in diagnstic wrkup, unless presenting with Celiac crisis. Why wuld that be? This sernegativity is due t the measurement f antibdy IgG and/r IgA against nly ne antigen f wheat, alphagliadin. Numerus cmplicatins have arisen regarding an accurate identificatin and diagnsis f Gluten-Reactivity, with r withut the enterpathy CD. Clinicians have been frustrated with the high percentage f false negative serlgy. 27 Fr example, CD has been called the Unfrgiving Master f Nn-Specificity and Disguise. 28 Therefre, if Gluten-Reactivity and CD g undetected fr years, the results culd be devastating autimmune cnditins. Mechanisms f actin are shwn in the Figure 1. Immunlgical Mechanisms Underlying Gluten Reactivity and Intlerance Figure 1 The mechanisms behind Gluten-Reactivity can ignite either Th-1 r Th-2 r a cmbinatin f bth immune reactins. Therefre, shuld Healthcare Practitiners limit their diagnstic inquisitiveness slely t the wellreferenced indicatrs f a severe gluten enterpathy (anti-transglutaminase and endmysial antibdies)? Numerus researchers suggest nt Cyrex Labratries, LLC. All Rights Reserved. 4

5 Current serlgy testing althugh highly sensitive and specific fr severe gluten enterpathy des nt address the diversity f gluten peptides and the need fr mre sensitive markers f Gluten-Reactivity with r withut CD. GLUTEN PEPTIDES During digestin, gluten prteins are enzymatically brken dwn in the gastrintestinal tract. Hwever, because f the high prline cntent f gluten, the degradatin is nt efficient and relatively large gluten peptides can persist. 29 There are thusands f such gluten peptides prduced during the digestive prcess, and multiple peptides can stimulate an immune respnse in an individual. The prtease-resistant 33-amin acid peptide frm wheat -gliadin is the immundminant antigen in wheat, but little is knwn abut the hierarchy f immundminance and cnsistency f recgnitin f T-cell epitpes in viv. 30 Althugh the majrity f immune reactins t gluten peptides is due t binding t HLA-DQ2, sme gluten peptides have a different antigenic specificity frm that f CD and are independent f the actin f transglutaminase enzyme and HLA-DQ2/DQ8 31 and therefre are directly presented by antigen presenting cells t T cells. Further cperatin with B cells results in antibdy prductin t the antigens. Figure 2 The kernel f wheat is cmprised f hundreds f prteins. These mlecules can be classified as either water- r alchl-sluble. Prteins and peptides frm each categry can ptentially be pathgenic t the Gluten-Reactive patient. By assessing bth water- and alchl-sluble fractins f wheat, a clearer picture f Gluten-Reactivity can be btained Cyrex Labratries, LLC. All Rights Reserved. 5

6 Up t 86 % f patients recgnize a different array f peptides. 32 And yet, cmmercially, the nly peptide that is tested is alpha-gliadin 33 MER. A panel f gluten peptides, which includes a number f the mre cmmn immundminant antigens, wuld prvide new pprtunities t screen, prevent disease develpment in individuals at risk, 33 and increase the sensitivity f the test t identify Gluten-Reactivity (with r withut the enterpathy CD). Antibdy Array 3 includes testing fr antibdies t the gluten peptides: native + deamidated -Gliadin-33-mer, -Gliadin-17-mer, -Gliadin-15-mer, -Gliadin-17-mer and Glutenin-21- mer. OPIOID PEPTIDES Exrphins are peptides which may have activity similar t that f mrphine and ther piids Five distinct exrphins have been identified in the pepsin-digest f gluten. The inhibitry actin f the exrphins in wheat has a specific piate effect. 37 This mrphine-like psychactive nature f the peptides results frm the incmplete digestin f these dietary prteins binding t the piate receptrs in the brain, and ffers a pssible explanatin fr sme f the reprted psychiatric reactins t these gluten prteins, including the sense f brain fg, behaviral prblems, r md swings that ften accmpany immune reactins t these fds and which may fllw with panic attacks, depressin, r ther neurlgical cmplaints. Antibdy Array 3 includes testing fr antibdies t the gluten piid peptides: Glutemrphin + Prdynrphin. LECTINS Wheat germ agglutinins (WGAs) are lectins r carbhydrate-binding prteins with a capacity t bind t many cells and tissue antigens. Lectins bind t cells invlved in the immune system and induce txic damage, inflammatin, and autimmunity. Mst lectins, including WGAs, are resistant t prtelysis, the degradatin f prteins by cellular enzymes. WGAs prfundly interfere with enzyme functin and inhibit their digestive functin By the binding f lectins t different tissue antigens, WGA can enhance antibdy prductin against itself as well as against the tissue and cells t which it binds. Fr example, in humans, WGA binds t the same surface receptrs t which islet autantibdies bind. Therefre, an islet cell with bund lectins wuld be a sitting duck fr autimmune diseases In humans, the evidence incriminating wheat lectin as a cause f IgA nephrpathy (IgAN) is nw impressive. 48 Children with IgAN have high bld levels f anti-gliadin and anti-mesangium antibdies, and their IgA is unusually lectin-sensitive. Als, there is circulating WGA in the bld f children with active IgAN, which may suggest that it is, in fact, wheat lectin frm the diet that may be an initiatr f the autimmune respnse. A gluten-free diet in IgAN patients was shwn t reduce prteinuria, IgAimmune cmplexes and IgA fd antibdies. 50 Antibdy Array 3 includes testing fr antibdies t the wheat lectin: Wheat Germ Agglutinin Cyrex Labratries, LLC. All Rights Reserved. 6

7 ENZYMES Enzymes cmplete this panel. Transglutaminases are a family f enzymes that frm prtein plymers, like scafflding, which are vital in the frmatin f barriers and stable structures. Since humans have transglutaminase (ttg) in many ther tissues, including bne, antibdies prduced against epithelial cell ttg2 can crss-react with ther ttgs such as bne, brain and skin. In such cases, this crss-reactin leads t autimmune respnses against ther tissues and thus develps int steprsis, neurautimmunity and skin disrders. Generally, patients with elevated antibdies t ttg are susceptible t autimmunity. ttg2 has been shwn t frm cmplexes with gliadin. 51 The incubatin f ttg2 with gliadin results in the frmatin f cvalent ttg-peptide cmplexes, which can adhere t intestinal walls. This psitining allws the gliadin-ttg cmplex t be recgnized by antigen-presenting cells, which prduces an immune respnse cascade that results in autantibdies. The prductin f these autantibdies may perpetuate a pr-inflammatry gastrintestinal destructive cycle. Tissue Transglutaminase-3 (ttg3) is expressed mainly in the epidermis, and t a lesser extent in the placenta and the brain. ttg3 has been shwn t be up-regulated in a variety f degenerative diseases. In certain patients, gluten-sensitive enterpathy manifests as a disrder f the skin called dermatitis herpetifrmis (DH). 54 DH is characterized by granular IgA depsits in the papillary dermis, which cntribute t plymrphic papules and blisters ften lcated ver extensr surfaces f the majr jints. 54 Patients with Huntingtn s disease have been shwn t make elevated antibdy levels t Transglutaminase-2 and Transglutaminase is activated by xidative stress, during which inflammatry cytkine prductin increases, specifically tumr necrsis factralpha and interfern-gamma. Huntingtn patients have been shwn t prduce mre interfern-gamma and interleukin-2 than healthy cntrls. 3 Elevated ttg3 expressin is in esphageal cancer. 55 Tissue Transglutaminase-6 (ttg6) is expressed in neural tissue. 7 The ttg6 enzyme is nt cmmnly expressed in the small intestine but can be fund in mucsal antigen-presenting cells. 56 Due t its clse hmlgy t ttg2 and ttg3, it prvides a clear pssibility that ttg6 culd be invlved in the pathgenesis f gluten reactivity related neurlgical dysfunctin. 7 Researchers speculate that autimmunity against ttg6 may result frm early brain damage and assciated inflammatin. 57 Patients with high levels f antibdies against ttg6 are suspected f having autimmunity against neurnal tissue. Neurnal clinical cnditins may manifest as Cerebral Palsy, 57 Gluten Ataxia 7 r Peripheral Neurpathy. 7 Antibdies may appear in serum befre the clinical nset f symptms. Antibdy Array 3 includes testing fr antibdies t enzymes: Tissue Transglutaminases -2, -3 and -6 and Gliadin-Transglutaminase Cmplex Cyrex Labratries, LLC. All Rights Reserved. 7

8 Measurements f antibdies against varius wheat prtein peptides and enzymes assciated with autimmunities are shwn in Figure 3. Figure 3 Antibdy Array 3 measures IgG and IgA antibdies against wheat prteins, peptides and assciated enzymes. IN FLUENCING FACTORS: GENETIC Of the general ppulatin, 40-50% are carriers f DQ2/DQ8 genes, hwever clse t 90% f Celiac disease patients carry the gene DQ2 (DQA1*05/DQB1*02), and a minrity (10%) f the Celiac disease patients carry DQ8 (DQA1*03/DQB1*0302). Typically, gluten peptides bind t the DQ2 and DQ8 mlecules. Recent research hwever, has identified at least eight new genmic regins with rbust levels f disease assciatin t Gluten-Reactivity. ENVIRONMENTAL (CHEMICALS, FOODS, BIOTOXINS, DRUGS )? Envirnmental factrs that have an imprtant rle in the develpment f CD have been suggested by epidemilgic studies. These include a prtective effect f breast-feeding and the intrductin f gluten in relatin t weaning. Numerus envirnmental factrs have been hypthesized as being catalysts fr the develpment f nt nly the gluten enterpathy CD, 62 but als systemic manifestatins f Gluten-Reactivity with r withut the enterpathy. Sme f these catalysts include bacteria, 63 viruses, 64 dysbisis, 65 and crss-reactive fds. 66 HISTORY (FAMILY, MEDICAL) CD and Gluten-Reactivity are characterized by a variety f clinical manifestatins. These include the typical malabsrptin syndrme (classic symptms) and a spectrum f symptms ptentially affecting any rgan r bdy system (nn-classic symptms). Clinical manifestatins f Gluten-Reactivity and CD can present at any age: 2012 Cyrex Labratries, LLC. All Rights Reserved. 8

9 Infancy (less than 2 years ld) diarrhea, abdminal distentin, failure t thrive (lw weight, lack f fat, hair thinning), anrexia, vmiting, psychmtr impairment (muscle wasting) Childhd diarrhea, cnstipatin, anemia, lss f appetite, shrt stature, steprsis Adulthd diarrhea, cnstipatin, anemia, aphthus ulcers, sre tngue & muth (muth ulcers, glssitis, stmatitis), dyspepsia, abdminal pain, blating (weight lss), fatigue, infertility, neurpsychiatric symptms (anxiety, depressin, etc.), bne pain (steprsis), weakness (mypathy, neurpathy). Reviewing current medicatins (antibitics, sterids, NSAID s, etc.), supplements, diets, and a detailed medical histry are critically imprtant in determining wh may have gluten sensitivity. The crrelatin between fd ingestin and symptm nset is f great clinical imprtance. CLINICAL SYTEMIC IMMUNE EFFECTS Inclusin f the specific antigens cmprising Antibdy Array 3, r Cmprehensive Gluten- Reactivity and Autimmunity is based n recent medical research studies. Cmprehensive quantitative mapping f T-cell epitpes was determined in CD. 32 Results demnstrated that patients respnd t a hetergeneus array f peptides; sme recgnized many peptides frm single r multiple gliadin families, while thers reacted t nly ne peptide. These results cnfirmed that a large number f gluten epitpes may be implicated in the develpment f CD and assciated diseases. Indeed, a T-cell line frm ne Celiac patient failed t recgnize any f the 21 tested peptides, which cnfirmed that a large number f gluten and ther wheat prtein epitpes are implicated in develpment f CD and assciated disrders. This suggests that ther gliadin peptides and prteins are invlved in the pathgenesis f Gluten-Reactivity and CD. We extended this hetergeneity in T-cell respnses t gluten and ther peptides riginated frm wheat t humral immune respnses by measuring IgG and IgA antibdies against nine different wheat antigens and peptides as well as enzymes assciated with autimmunities. Hetergeneity in IgG and IgA antibdies against these twelve antigens was cnfirmed by variatin in antibdy respnse against varius wheat assciated antigens n individual bases. Therefre, Antibdy Array-3, with its measurement f IgG and IgA antibdies against a repertire f prteins, enzymes and peptides, riginated nt nly frm -gliadin but als frm - and -gliadin, glutenins, agglutinins, piid peptides (Glutemrphin + Prdynrphin), and specific enzymes invlved in the pathgenesis f CD and Gluten-Reactivity, enhances the clinical sensitivity and specificity fr the detectin f CD and Gluten-Reactivity. CLINICAL USE OF ANTIBODY ARRAY 3 Measuring a patient s immune respnse t an array f wheat antigens increases the sensitivity and specificity, and will prvide greater cnfidence in frmulatin f a diagnsis that allws fr better patient cmpliance with a gluten-free diet. Assessing wheat/gluten reactivity and intestinal autimmunity is recmmended fr patients wh: 2012 Cyrex Labratries, LLC. All Rights Reserved. 9

10 Have gut dysbisis, which appears t be resistant t standard therapy Are suspected f having intestinal mucsal damage Cmplain f fd allergy and intlerance Cmplain f chemical hypersensitivity Present multiple-symptm cmplaints (including Chrnic Fatigue Syndrme and Fibrmyalgia) Suffer frm abnrmal immune cell cunt and functin May suffer frm bld-brain barrier permeability, depressin, r neurautimmunity Neurautimmune patients t cnsider: Thyriditis Arthritis Mycarditis Dermatitis Endcrinpathy Plyendcrinpathy Ostearthritis Pernicius Anemia Other CLINICAL INTERPRETATION OF ANTIBODY ARRAY 3 When IgA reactins are predminant, it is an indicatin f pssible Celiac disease and ther autimmunities. When IgG reactins are predminant, it is an indicatin f wheat/gluten immune respnse and pssible autimmunity due t lack f digestive enzymes and/r ther factrs. When bth IgA and IgG reactins ccur, it is an indicatin f wheat/gluten immune respnse and its prgressin t Celiac disease and/r ther autimmune disrders Cyrex Labratries, LLC. All Rights Reserved. 10

11 INTERPRETATION OF ANTIBODIES AGAINST WHEAT, GLUTEN, ENZYME ANTIGENS POSITIVE REACTION TO: Wheat WGA γ Gliadin 15 mer α Gliadin 17 mer α Gliadin 33 mer ω Gliadin 17 mer Glutenin 21 mer Glutemrhpin + Prdynrphin GLUTEN SENSITIVITY WHEAT & GLUTEN SENSITIVITY WHEAT SENSITIVITY LECTIN SENSITIVITY AUTO IMMUNE REACTION INTERPRETATION Wheat sensitivity due t lack f digestive enzymes Sensitivity t wheat germ and spruted wheat One r any cmbinatin f psitives means sensitivity t specific gluten epitpes due t lack f enzymes, in particular DPPIV Immune reactin t piid peptides. Due t lack f digestive enzymes, in particular DPPIV CLINICAL APPROACH Wheat free diet Heal gut Check fr ther lectin sensitivity Gluten free diet Heal gut Check fr extra intestinal autimmunity Patient is addicted t wheat GliadintTG2 Cmplex ttg2 ttg3 ttg6 Autimmunity assciated with wheat sensitivity Pssible GI autimmunity assciated with wheat sensitivity, if wheat antigens are als psitive Pssible skin autimmunity assciated with wheat sensitivity, if wheat antigens are als psitive Pssible brain autimmunity assciated with wheat sensitivity, if wheat antigens are als psitive Remve trigger Use antiinflammatries 2012 Cyrex Labratries, LLC. All Rights Reserved. 11

12 SPECIMEN REQUIREMENT 2 ml Serum Ambient RELATED TESTING Antibdy Array 2 - Intestinal Antigenic Permeability Screen (Serum) Antibdy Array 4 - Gluten-Assciated Crss-Reactive Fds and Fds Sensitivity (Serum) Antibdy Array 5 Systemic Autimmune Reactivity Screen (Serum) 2012 Cyrex Labratries, LLC. All Rights Reserved. 12

13 REFERENCES 1 Vjdani, A. The characterizatin f the repertire f wheat antigens and peptides invlved in the humral immune respnses in patients with gluten sensitivity and Crhn s disease. ISRN Allergy, 2011; 2011: Hadjivassiliu, M., Sanders, D.S., Grünewald, R.A., et al. "Gluten sensitivity: frm gut t brain." Lancet Neurl, 2010;9(3): Fasan, A. "Celiac disease-hw t handle a clinical chamelen." NEnglJMed, 2003; 348: Arnsn, Y., Amital, H., Shenfeld, Y. "Vitamin D and autimmunity: new aetilgical and therapeutic cnsideratins." J Immunl, 2005; 175: Alaedini, A., Okamt H., Briani, C., Wllenberg, K., et al. "Immune crss-reactivity in Celiac disease: anti-gliadin antibdies bind t neurnal Synapsin I." JImmunl, 2007; 178: Green, P., Alaedini, A., Sander, H.W., et al. "Mechanisms underlying Celiac disease and its neurlgic manifestatins." Cell Ml Life Sci, 2005; 62: Hadjivassiliu, M., Aeschlimann, P., Strigun, A., et al. "Autantibdies in gluten ataxia recgnize a nvel neurnal transglutaminase." Ann Neurl, 2008;64(3): Hadjivassiliu, M., Grünewald, R.A., Davies-Jnes, G.A. "Gluten sensitivity as a neurlgical illness." J NeurlNeursurgPsychiatry, 2002;72(5): Marietta, E., Black, K., Camilleri, M., et al. "A new mdel fr dermatitis herpetifrmis that uses HLA-DQ8 transgenic NOD Mice." J Clin Invest, 2004;114(8): Lindqvist, U., Rudsander, A., Bström, A., et al. "IgA antibdies t gliadin and Celiac disease in psriatic arthritis." Rheumatlgy, 2002;41(1): Humbert, P., Pelletier, F., Dren, B., et al. "Gluten intlerance and skin diseases." Eur J Dermatl, 2006; 16(1): Selva-O'Callaghan, A., Casellas, F., De Trres, I., et al. "Celiac disease and antibdies assciated with Celiac disease in patients with inflammatry mypathy." Muscle Nerve, 2007;35(1): Hadjivassiliu, M., Grünewald, R., Sharrack, B., et al. "Gluten ataxia in perspective: epidemilgy, genetic susceptibility and clinical characteristics." Brain, 2003;126(Pt 3): Hadjivassiliu, M., Aeschlimann, D., Grünewald, R.A., et al. "GAD antibdy-assciated neurlgical illness and its relatinship t gluten sensitivity." ActaNeurlScand, 2010; 123(3): Eatn, W., Mrtensen, P.B., Agerb, E., et al. "Celiac disease and schizphrenia: ppulatin based case cntrl study with linkage f Danish natinal registers." Br Med J, 2004; 328(7437): Cyrex Labratries, LLC. All Rights Reserved. 13

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15 30 Tye-Din, J.A, Stewart, J.A., Drmey, J.A., et al. "Cmprehensive, quantitative mapping f T- cell epitpes in gluten in Celiac disease." SciTransl Med, 2010; 2(41):41ra Samar, D., Dickersn, F., Kasarda, D.D., et al. "Nvel immune respnse t gluten in individuals with schizphrenia." Schizphr Res, 2010;118(1-3): Camarca, A., Andersn, R.P., Mamne, G., et al. "Intestinal T-cell respnses t gluten peptides are largely hetergeneus: implicatins fr a peptide-based therapy in Celiac disease." J Immunl, 2009;182(7): Vader, W., Ky, Y., Van Veelen, P., et al. "The gluten respnse in children with Celiac disease is directed tward multiple gliadin and glutenin peptides." Gastrenterlgy, 2002;122(7): Fukudme, S. and Yshikawa, M. "Opiid derived frm wheat gluten: their islatin and characterizatin." FEBS Let,1992; 296: Fukudme, S., Shimatsu, A., Suganuma, H., Yshikawa, M. "Effect f gluten exrphins A5 and B5 n the pstprandial plasma insulin level in cnscius rats." Life Sci, 1995;57(7): Mycrft, F.J., Wei, E.T., Bernardin, J.E., Kasarda, D.D. "MIF-like sequences in milk and wheat prteins." N Engl J Med, 1982;307(14): Ziudru, C., Streaty, R.A., Klee, W.A. "Opiid peptides derived frm fd prteins: The exrphins. J BilChem, 1979;254(7): Dhan, F.C. "Genetic hypthesis f idipathic schizphrenia: its exrphin cnnectin." Schizphr Bull, 1988;14(4): Saelid, G., Haug, J.O., Heiberg, T., Reichelt, K.L. "Peptide-cntaining fractins in depressin." BilPsychiatry, 1985;20(3): Hggan, R. "Abslutism's hidden message fr medical scientism." Interchange, 1997; 28(2-3): Bardcz S., Grant G., Brwn D.S., et al. "Plyamine metablism and uptake during Phaselus vulgaris lectin induced grwth in rat small intestine." Digestin, 1990; 46(suppl 2): Wright, H.T., Sandrasageram, G., Wright, C.S. "Evlutin f a family f Gnac-binding prteins cntaining the disulphide-rich dmain f wheat germ agglutinin. J MlEvl, 1991; 33: Pusztai, A., Ewen, S.W., Grant, G., et al. "Relatinship between survival and binding f plant lectins during small intestinal passage and their effectiveness as grwth factrs." Digestin, 1990; 46(suppl 2): Ericksn, R.H., Kim, J., Sleisinger, M.H., Him, Y.S. "Effect f lectins n the activity f brush brder membrane-bund enzymes f rat small intestine." J PediatrGastrenterlNutr, 1985; 4: Cyrex Labratries, LLC. All Rights Reserved. 15

16 45 Brady, P.G., Vannier, A.M., Banwell, J.G. "Indentificatin f the dietary lectin wheat germ agglutinin in human intestinal cntents." Gastrenterlgy, 1978; 75: Kitan, N., Taminat, T., Ida, T., et al. "Detectin f antibdies against WGA bund glycprteins n the islet-cell membrane." Diabet Med, 1988; 5: Uchigata, Y., Spitalnik, S.L., Tachiwaki, O., et al. "Pancreatic islet cell surface glycprteins cntaining Gal 1-4 GNAc-R identified by a cytxic mnclnal autantibdy." J Exp Med, 1987; 165: Cpp, R., Amre, A., Rccatell, D. "Dietary antigens and primary IgA nephrpathy." J Am ScNephl, 1992; 2(suppl 10):s173-s Cpp, R., Amre, A., Giangli, B., et al. "Macrmlecular IgA and abnrmal IgA reactivity in sera frm children with IgA nephrpathy." ClinNephrl, 1995; 43: Amre, A., Cavall, F., Bcchiett, E., et al. "Cytkine mrna expressin by cultured rat mesangial cells after cntact with envirnmental lectins." Kidney Internat, 1993; 39(suppl):S41- S Matthias, T., Pfeiffer, S., Selmi, C., Gershwin, M.E. Diagnstic challenges in Celiac disease and the rle f the tissue transglutaminase-new-epitpe. Clinic Rev AllergImmunl, 2010; 38: Ahvaz,i B., Beshans, K.M.,Rastinejad, F. The emerging structural understanding f transglutaminase 3. J Structural Bil, 2004; 147: Kim, S-Y., Jeitner, T.M., Steinert, P.M. Transglutaminases in Disease. NeurchemIntnl, 2002; 40: Sárdy, M., Kárpárti, S., Merkl, B.,et al. Epidermal Transglutaminase (TGase 3) Is the autantigen f dermatitis herpetifrmis. J Exp Med, 2002; 196(6): Uemura, N., Nakanishi, Y., Kat, H.,et al. Transglutaminase 3 as a prgnstic bimarker in esphageal cancer revealed by prtemics. Int J Cancer, 2009; 124: Dubis, P. C. and van Heel, D. A. "Translatinal mini-review series n the immungenetics f gut disease: immungenetics fceliac disease." ClinExpImmunl, 2008; 153: Stenberg, R., Kaukinen, K., Bengtssn, M., et al. Early develping celiac disease in children with cerebral palsy. J PediatrGastrenterlNutr, 2011; 53(6): Plenge, R. "Unlcking the pathgenesis f Celiac disease." Nat Genet, 2010; 42(4): Ivarssn, A., Hernell, O.,Stenlund, H.,ÅkePerssn, L. "Breast-feeding prtects against Celiac disease." Am J ClinNutr, 2002;75: Cyrex Labratries, LLC. All Rights Reserved. 16

17 60 Nrris, J.M,Barriga, K., Hffenberg, E.J.,et al. "Risk f Celiac disease autimmunity and timing f gluten intrductin in the diet f infants at increased risk f the disease." JAMA, 2005; 19: Naiyer, A.J. and Green, P.H.R. "Hw imprtant is the timing f gluten intrductin fr children with Celiacdisease?" Nat ClinPractGastrenterlHepatl, 2005; 2(10): Betterle, C., and Zanchetta, R. "Update n autimmune plyendcrine syndrmes (APS)." ACTA BIO MEDICA, 2003; 74: Verdu, E.F., Maur, M., Burgeis, J., Armstrng, D., "Clinical nset f Celiac disease after an episde f Campylbacter jejuni enteritis." Can J Gastrenterl,2007; 21(7): Zanni, G., Navne, R., Lunardi, C., et al. "In Celiac disease, a subset f autantibdies against transglutaminase binds tll-like receptr 4 and induces activatin f mncytes." PLS Med, 3(9):e358.DOI: /jurnal.pmed Tursi, A., Brandiman, G., Girgelli, GM. "High prevalence f small intestinal bacterial vergrwth in Celiac patients with persistence f gastrintestinal symptms after gluten withdrawal." Am J Gastr, 2003; 98(4): Bnds, R., Midr-Hriuti, T., Gldblum R. "A structural basis fr fd allergy: the rle f crss-reactivity." Curr Opinin AllerImmunl, 2008;8: Jnes, R. and Sleet S. "Easily missed?:celiac disease. Br Med J, 2009;338:a Jnes, S., D'Suza, C., Habubi, N. "Patterns f clinical presentatin f adult Celiac disease in a rural setting." Nutri J, 2006; 5: Feighery, C. "Clinical review: frtnightly review Celiac disease." Br Med J, 1999; 319: Fasan, A. "Clinical presentatin f Celiac disease in the pediatric ppulatin." Gastrenterlgy, 2005;128(suppl 1):S68 S Hill, I.D., Direks, M.H, Liptak, G.S., et al. "Guideline fr the diagnsis and treatment f Celiac disease in children: recmmendatins f the Nrth American sciety fr pediatric gastrenterlgy, hepatlgy and nutritin." J PediatrGastrenterlNutr, 2005; 40(1): Cyrex Labratries, LLC. All Rights Reserved. 17

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