RU2736393C1 - Method of early preclinical diagnosis of allergic enteropathy in children - Google Patents

Abstract

FIELD: medicine.SUBSTANCE: invention refers to medicine, namely to allergology in pediatrics, pediatric gastroenterology, and can be used for early preclinical diagnosis of allergic enteropathy in children. Method involves urine concentration in the urine in children of the intestinal fraction of a fatty acid-binding protein 24 to 36 hours after introduction of the new product into the diet. If the level of the intestinal fraction of the protein which binds the fatty acids increases above 0.092 pg/ml, the developing aggravated allergic bowel disease is stated, damage to intestinal barrier and necessity to cancel further introduction of this product. If urine content of intestinal fraction of fatty acid binding protein is less than 0.092 pg/ml, there is no damage to intestinal barrier and possibility of further introduction of said product.EFFECT: use of the invention enables to obtain reliable information on the state of the intestinal barrier and development of its damage in a short period of time when introducing a new product to a child with food allergy during stages of ration expansion.1 cl, 1 dwg, 3 ex

Description

The invention relates to medicine, namely to allergology, pediatric gastroenterology and pediatrics, and can be used in the diagnosis of allergic enteropathy and staged diet therapy and dietary prophylaxis of food allergy in children.

The rapidly growing prevalence of food allergy (PA) is observed all over the world and is considered “the second wave of the epidemic of allergic diseases” [1, 3, 4]. Modern data indicate the involvement of various organs and systems in the formation of PA, a varied clinical picture with different severity and spectrum of manifestations [2; five; eight]. The diagnosis of PA is not difficult if the allergic reaction develops soon after taking the "causal" allergic product [1; 6]. However, diagnosis is difficult if this reaction occurs delayed, after a few hours or even days, or one has to suspect a frequently consumed food product that did not cause a reaction before [7; eight]. Clinical symptoms of allergic enteropathy affect mainly the gastrointestinal system, and are characterized by a delayed onset, from 2 to 5-7 days after ingestion of the allergen. Until now, there are significant difficulties in the early diagnosis of allergic enteropathy, there is no single diagnostic test, the diagnostic criteria are based on the analysis of anamnesis data, a complex clinical laboratory and instrumental research [3, 8]. Meanwhile, the severity of the course of the disease, the development of complications and the prognosis depend on the timing of the diagnosis and exclusion of the causally significant allergen from the diet. Establishing an accurate diagnosis is especially important in children in the first years of life, since an erroneous diagnosis can lead to the development of life-threatening reactions or the appointment of unreasonable restrictive diets [34; 54; 57; 71; 163].

Currently, according to numerous recommendatory documents, there is no generally accepted routine diagnostic test that can establish the diagnosis of PA without a doubt [39; 48; 64; 71; 103]. Published international clinical guidelines WAO and EAACI explain using algorithms the principles of diagnosis of PA [131; 285]. Diagnosis of food allergies should always begin with a detailed history, followed by diagnostic measures. The diagnosis can be made in the presence of a positive allergic anamnesis in combination with pathognomonic symptoms, positive results of allergy examination and the presence of clearly positive dynamics of the clinical picture after the elimination diet. In this case, the history and diagnostic diet play a major role [69; 102].

The generally accepted method for diagnosing allergic enteropathy is currently provocative dietary diagnostics [3]. This method is indirect, insufficiently informative and specific, the diagnostic process is extended in time, in addition, repeated provocation for diagnostic purposes is undesirable. The disadvantages of this method are the need to conduct a study during the period of remission of the disease, the need for long-term compliance with strict unreasonable elimination measures, the frequent development of deficiency states, especially in young children, an exacerbation of the process in case of positive results of this diagnostic method.

The known "Method for the diagnosis of allergy of the gastrointestinal tract in children" (RF Patent No. 21228836, authors Sazanova N.E., Varnacheva L.N., Novikova A.V., Khokhlova N.M.) by morphometric examination of the mucous membrane of the small intestine , at the same time, a combination of interepithelial eosinophils and tissue macrophages of villi is determined and with the first indicator from 0 to 1%, and the second from 3 to 10% diagnose delayed-type allergy, with the first indicator higher, and the second lower than the indicated values, an immediate-type allergy is diagnosed. However, this method refers to the topical diagnosis of diseases of the small intestine itself and does not allow one to judge about changes in other parts of the gastrointestinal tract caused by gastrointestinal allergy. In addition, the method is invasive and traumatic, therefore it has limitations on its use in children, and is also not indicated for mild cases of the disease. In addition, the morphometry method is rather laborious, has not received wide application in morphological laboratories and is usually used only for scientific research.

The known "Method of differential diagnosis of gastrointestinal food allergy" (Patent RU No. 2287166 (13), authors Varnacheva L.N., Sazanova N.E., Volkova E.A., Korkotashvili L.V., Mayanskaya I.V.), the essence of which is that the content of pre-β-lipoproteins and β-lipoproteins is determined in the blood serum. When the ratio of the levels of pre-β-lipoproteins to β-lipoproteins is higher than 0.31, the gastrointestinal form of food allergy is differentiated from another pathology of the gastrointestinal tract. However, this method does not provide early preclinical diagnosis of allergic enteropathy; it is recommended to carry out measures against the background of the formed clinical picture of the disease. In addition, blood serum is required for its implementation, which increases the invasiveness of the study, which is especially important for young children.

The closest analogue of the proposed method is a method of non-invasive diagnosis of enteropathy induced by cow's milk protein in infants (Patent RU No. 2613160, authors Zernova E.S., Shumatova T.A., Shishatskaya S.N., Grigoryan L.A. , Prikhodchenko N.G.), taken as a prototype, which consists in the fact that in children with suspected allergy to cow's milk proteins, before the start of the elimination diet against the background of clinical manifestations in coprofiltrates, the content of the following peptides is determined: β-defensin 2, zonulin, eosinophilic cationic protein. When β-defensin 2 values are more than 34.8 ng / ml, zonulin is higher than 1.59 ng / ml, and eosinophilic cationic protein is more than 455.57 ng / ml, the patient is diagnosed with allergic enteropathy induced by cow's milk protein. The disadvantage of this method is the need to study the combination of three fecal biomarkers, which increases the financial burden and reduces compliance. Also, the need to prepare coprofiltrate requires special equipment from laboratories and special skills from personnel, due to the high sensitivity of the method. Eosinophilic cationic protein can be negative in non-eosinophilic lesions of the gastrointestinal tract, zonulin is currently considered a proven marker of intestinal permeability, there is evidence that its increased numbers persist in patients in the stage of persistent remission, even with an elimination diet, which makes it difficult to use it as an informative marker of early preclinical diagnosis of the disease. In addition, these indicators are recommended for research at the stage of detailed clinical symptoms, the use of this method at the stage of prenosological diagnosis is impossible.

There are no analogues of the method for early preclinical diagnosis of allergic enteropathy in children.

The objective of the proposed method is to develop an effective, highly informative, non-invasive method for early preclinical diagnosis of allergic enteropathy in children, allowing to prevent the development of the disease at the stage of pre-illness or to prevent an exacerbation of the disease when expanding the diet in children with food allergies. The technical result from the implementation of the proposed method is that the claimed method allows in a short time to obtain reliable information about the state of the intestinal barrier and the development of its damage when a new product is introduced to a child with food allergies at the stages of diet expansion.

The essence of the method consists in determining the concentration of the intestinal fraction of the protein that binds fatty acids in the urine of children with food allergies 24-36 hours after the introduction of a new complementary food product into the diet. When determining the level of the intestinal fraction of the protein that binds fatty acids above 0.092 pg / ml, the development of acute damage to the intestinal barrier as a result of allergic inflammation and the need to cancel further administration of this product are judged. The content in the urine of the intestinal fraction of the protein that binds fatty acids below 0.092 pg / ml indicates the absence of damage to the intestinal barrier and the possibility of further administration of this product. EFFECT: method provides safe effective prevention of exacerbation of gastrointestinal forms of food allergy in children.

The technical result from the implementation of the proposed method:

- complete safety for the patient's health;

- the ability to conduct research at any age;

- the study requires a small amount of biological material (3 ml of urine);

- lack of reactogenicity on the part of the patient's body;

- the collection of material does not present any difficulties for parents, it can be done at home at any time, regardless of food intake, does not require special storage and transportation conditions.

The proposed assessment method is technically simple, non-invasive, informative, its implementation does not require large economic costs.

The claimed method for early preclinical diagnosis of allergic enteropathy in children is new and has not been described in the public literature. The technical result is achieved as a result of the implementation of the proposed method, the essence of which lies in the fact that the child's urine is examined with the determination of the content of the intestinal fraction of the protein that binds fatty acids in it to assess the consistency of the intestinal barrier and the possibility of introducing new products and further expanding the diet in children. The intestinal fraction of fatty acid binding protein (I-FABP) is determined in urine by enzyme linked immunosorbent assay (ELISA) on an automatic enzyme immunoassay analyzer using reagents from Cloud-Clone Corp (CCC, USA). When determining the level of the intestinal fraction of the protein that binds fatty acids above 0.092 pg / ml, the development of acute damage to the intestinal barrier as a result of allergic inflammation and the need to cancel further administration of this product are judged. The content in the urine of the intestinal fraction of the protein that binds fatty acids below 0.092 pg / ml indicates the absence of damage to the intestinal barrier and the possibility of further administration of this product.

The claimed distinctive features are new, since the determination of I-FABP in urine has not previously been used to diagnose allergic enteropathy in children. For the purpose of early diagnosis of gastrointestinal forms of food allergy and prevention of exacerbations of allergic diseases in children, methods for diagnosing damage to the intestinal barrier using various techniques are being intensively developed [1, 8, 9, 10, 11]. However, the high invasiveness of the main diagnostic methods limits their routine use in clinical practice and delays the diagnosis, contributes to the formation of chronic diseases, polydeficiency states. In this regard, the development and introduction into clinical practice of non-invasive diagnostic methods, early diagnosis of an exacerbation of the disease at the prenosological stage and predictive therapy are especially relevant in pediatric practice. Allergic enteropathy is associated with acute damage to the mucous membrane of the small intestine and enterocytes. Intestinal fatty acid binding protein (I-FABP) is an intracellular protein that is specifically and abundantly expressed in the epithelial cells of the mucous membrane of the small and large intestine [5]. It has been shown that I-FABP is released into the bloodstream after damage to the mucous membrane of the small intestine, and then it is eliminated by the kidneys [7]. Therefore, I-FABP can be measured in both serum (or plasma) and urine. The concentration of I-FABP is very low in the urine of healthy subjects, but its value significantly increases in the blood 60 minutes after damage to intestinal enterocytes [5-7].

The method is carried out as follows. A child with a gastrointestinal form of food allergy at the 3rd stage of diet therapy (the stage of expanding the diet), 24-36 hours after the diagnostic introduction of a new product, urine is taken in an amount of 3 ml into a sterile bottle. The sampling of material can be carried out at any time of the day, regardless of the meal. The biological material is centrifuged for 10 minutes, after which the material is poured into epindorfs. The intestinal fraction of proteins that bind fatty acids in urine is examined by enzyme linked immunosorbent assay (ELISA) on an automatic enzyme immunoassay analyzer using reagents from Cloud-Clone Corp (CCC, USA). When determining the level of the intestinal fraction of the protein that binds fatty acids above 0.092 pg / ml, the development of acute damage to the intestinal barrier as a result of allergic inflammation and the need to cancel further administration of this product are judged. The content in the urine of the intestinal fraction of the protein that binds fatty acids below 0.092 pg / ml indicates the absence of damage to the intestinal barrier and the possibility of further administration of this product.

The efficiency of the method has been proved by clinical and laboratory examination of 28 children aged 1 month to 3 years, who, as a result of a comprehensive examination, established allergic enteropathy induced by cow's milk proteins. The comparison group consisted of 20 healthy children, matched by sex and age, with an unburdened allergic history. In all children, the intestinal fraction of fatty acid binding proteins (I-FABP) was determined in urine by enzyme-linked immunosorbent assay (ELISA) on an automatic enzyme-linked immunosorbent analyzer using Cloud-Clone Corp reagents (CCC, USA). Average concentrations of I-FABP in urine in children with food allergies and in children from the comparison group (control group) were 0.164 ± 0.031 pg / ml and 0.039 ± 0.009 pg / ml, respectively (p <0.05). Strong direct correlations were revealed (Spearman's correlation coefficient S = 0.634, p <0.05) between the level of I-FABP in urine and the severity of gastrointestinal manifestations of food allergy. All children strictly adhered to the prescribed elimination diet with the exclusion of cow's milk protein during the first and second stages of elimination diet therapy. When studying I-FABP in urine in the dynamics of the disease, it was found to be significantly reduced to 0.066 ± 0.004 pg / ml, respectively (p <0.05). During the 3 stage of elimination diet therapy (diet expansion stage) with dynamic observation, all children with CMPA were divided into 2 subgroups: 1a subgroup consisted of 10 children who, 24-72 hours after the introduction of a new complementary food product, developed symptoms of an exacerbation of the disease in the form of frequent stools watery character with a sour odor, flatulence and flatulence, in this subgroup the level of I-FABP in 24-36 hours after the introduction of the new product was 0.149 ± 0.026 pg / ml. In 16 children, with dynamic observation for 24-72 hours, there were no symptoms from the gastrointestinal tract and other body systems, the introduction of the product was continued, they made up subgroup 1b. The I-FABP level 24-36 hours after the introduction of the new product in this subgroup was 0.052 ± 0.013 pg / ml (statistical significance p = 0.041 compared with subgroup 1b, p = 0.086 compared with the control group of healthy children).

Thus, the determination of I-FABP in urine is a non-invasive criterion for assessing the structural and functional state of the intestine in children and can be used as an objective diagnostic marker indicating damage to enterocytes and the development of allergic enteropathy at the preclinical and pre-nosological stage. These observations were confirmed using ROC analysis when comparing the Area Under Curve (AUC) (Fig. 1).

The AUC indicator in the study of I-FABP in urine samples is 0.78, the optimal threshold (cut-off point) is 0.092 pg / ml, the sensitivity is 91%, and the specificity is 76%. Thus, the determination of the content of I-FABP in urine is an objective criterion for damage to the intestinal barrier in children with gastrointestinal food allergy and can be used as an effective method for early preclinical diagnosis of allergic enteropathy in children.

Clinical example 1.

Boy A., 5 months old, since 2 months has been observed with a diagnosis of Allergic dermatitis, allergic enteropathy. From the moment of diagnosis, she has been on an elimination diet with the exception of BCM-based products by a nursing mother. The condition is stable, there are no complaints, physical and neuropsychic development corresponds to age. Given the age of the child and the need to introduce complementary foods, it was decided to introduce vegetable puree into the diet. After 26 hours from the moment of introducing vegetable puree from zucchini into the diet (complementary foods were introduced according to the National Program for Optimizing Feeding of Children of the First Year of Life in the Russian Federation, 2019), urine was collected and the content of I-FABP in urine was determined, the result was 0.067 pkg / ml. According to the proposed method for early preclinical diagnosis of allergic enteropathy in children, it was concluded that there was no damage to the intestinal barrier, remission of the allergic disease, and the possibility of further administration of this product. The subsequent increase in the volume of the administered product did not exacerbate the disease.

Clinical example 2.

Boy I., 7 months old, from 1 month old is observed with a diagnosis of allergic enteropathy. From the moment of diagnosis, he is on an elimination diet with the exclusion of BCM-based products, the child is bottle-fed, receives a mixture based on high hydrolysis of cow's milk protein. The condition is stable, there are no complaints, physical and neuropsychic development corresponds to age. Taking into account the child's age and the duration of the elimination diet (6 months), according to the principles of staged, elimination diet therapy [4], it was decided to transfer the child to a formula based on partial hydrolysis of cow's milk protein. After 24 hours from the moment the mixture was introduced into the diet based on partial hydrolysis of cow's milk protein, urine was collected and I-FABP was determined in urine, the result was 0.303 pkg / ml. According to the proposed method for early preclinical diagnosis of allergic enteropathy in children, it was concluded that acute damage to the intestinal barrier as a result of allergic inflammation, the risk of exacerbation of allergic enteropathy and the need to cancel further administration of this product. The child was returned to the previous stage of diet therapy (a mixture based on high hydrolysis of whey protein); further observation of the child for 2 weeks did not show an aggravation of food allergy in the child.

Clinical example 3.

Boy P., 1 year old. The family history of allergies is aggravated by the mother's cousin - eczema, the older sister of the boy - neurodermatitis. Manifestations of food intolerance in the form of unstable stool, persistent diarrhea were observed from 2 months of age when the child was transferred to artificial feeding based on CMP, from 4 months it was observed for allergic enteropathy, from 6 months he was transferred to artificial feeding (a mixture based on highly hydrolyzed whey protein). Within 2 weeks, the condition stabilized, there were no complaints, physical and neuropsychic development was age-appropriate. Considering the child's age and the need to introduce complementary foods, it was decided to introduce cereal complementary foods (buckwheat porridge) into the diet. 30 hours after the introduction of buckwheat porridge into the diet (complementary foods were introduced according to the National Program for the Optimization of Feeding of Children of the First Year of Life in the Russian Federation, 2019), urine was collected and the content of I-FABP in urine was determined, the result was 0.155 pkg / ml. According to the proposed method for early preclinical diagnosis of allergic enteropathy in children, it was concluded that acute damage to the intestinal barrier as a result of allergic inflammation, the risk of exacerbation of allergic enteropathy and the need to cancel further administration of this product. Despite the recommendations received, the mother continued the introduction of complementary foods, given the absence of clinical symptoms. On the 5th day after the introduction of complementary foods, the child was admitted to the hospital with complaints of constant anxiety during and after feeding, unmotivated crying, regurgitation after eating, bloating, diarrheal syndrome.

Sources of information:

1. Pampura A.N. Food allergy in young children / A.N. Pampura // Pediatrics. Journal them. G.N. Speransky. - 2016. - T. 95, No. 3. -FROM. 152-157.

2. RU 2621312 (FGAU "SCCH" of the Ministry of Health of Russia), 01.06.2017 "A method for predicting the formation of tolerance in case of allergy to cow's milk protein in young children."

3. RU 2613160 (GBOU VO "TSMU" of the Ministry of Health of Russia), 03/15/2017 "Method for non-invasive diagnosis of enteropathy induced by cow's milk protein in infants."

4. Baranov A.A., Namazova-Baranova L.S. Federal clinical guidelines for the provision of medical care to children with allergies to cow's milk proteins M., 2015. [electronic resource http://www.pediatr-mssia.ru/sites/default/files/file/kr_abkm.pdf]

5. Yavuz S. T., Buyuktiryaki B. et al. Factors that predict the clinical reactivity and tolerance in children with cow's milk allergy // Ann Allergy Asthma Immunol. - 2013 Apr. - V. 110 (4). - P. 284-289.

6. Fasano A. Zonulin and ist regulation of intestinal barrier function: the biological door to inflammation, autoimmunity, and cancer // Physiol Rev. - 2011. -V. 91, no. 1. - P. 151-175.

7. Vassilopoulou E., Konstantinou G., Kassimos D. et al. Reintroduction of cow's milk in milk-allergic children: safety and risk factors / // Int Arch Allergy Immunol. - 2008: 146: 156-61.

8. Steele L. et al. Mucosal immunology of tolerance and allergy in the gastrointestinal tract // Immunol Res. - 2012. - V. 1, No. 54. - P. 75-82.

9. Mishra A., Prakash S., Sreenivas V. et al. Structural and Functional Changes in the Tight Junctions of Asymptomatic and Serology-negative First-degree Relatives of Patients With Celiac Disease / // J Clin Gastroenterol. - 2016. - V. 50, No. 7. - P. 551-560.

Claims (1)

A method for early preclinical diagnosis of allergic enteropathy in children, including the determination in the urine of children of the concentration of the intestinal fraction of the protein that binds fatty acids, 24-36 hours after the introduction of a new product into the diet, characterized in that with an increase in the level of the intestinal fraction of the protein that binds fatty acids , above 0.092 pg / ml, it is judged about the development of an exacerbation of an allergic bowel disease, damage to the intestinal barrier and the need to cancel further administration of this product, if the content of the intestinal fraction of the protein that binds fatty acids in the urine is below 0.092 pg / ml, it is judged that there is no damage to the intestinal barrier and the possibility further introduction of this product.

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