- Email: [email protected]
Comparison of the different kinds of feeding on the level of fecal calprotectin
Early Human Development 90 (2014) 471–475
Contents lists available at ScienceDirect
Early Human Development journal homepage: www.elsevier.com/locate/earlhumdev
Comparison of the different kinds of feeding on the level of fecal calprotectin☆ Feng Li, Jingqiu Ma, Shanshan Geng, Junli Wang, Fang Ren, Xiaoyang Sheng ⁎ Department of Children and Adolescents Health Care, MOE-Shanghai Key Laboratory of Children's Environmental Health, Xin Hua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
a r t i c l e
i n f o
Article history: Received 7 May 2014 Accepted 14 June 2014 Available online xxxx Keywords: Fecal calprotectin Breast-feeding Formula-feeding Infants
a b s t r a c t Background: Controversial results have been reported on the effect of type of feeding on the level of fecal calprotectin in infants. Objective: To assess fecal calprotectin levels in breast fed or nonbreast fed healthy infants. Design: A study was conducted to compare fecal calprotectin in infants who were exclusively breastfed compared to those who were not breastfed in Shanghai, China. Stool samples were collected and analyzed, and the fecal calprotectin concentration was determined using a commercially available enzyme-linked immunosorbent assay. The infant's weight and length were measured. Parents were asked to fill in a brief questionnaire, with questions about several clinical and sociodemographic factors. Subjects: This study included 105 healthy infants aged 0–5 months. Results: Stool samples were obtained from 105 healthy infants (63 boys, 42 girls) with a median age of 2.86 months (range 1–5.88). The median fecal calprotectin concentration was significantly higher in breast fed infants (377 μg/g, range 35–937 μg/g) than that in nonbreast fed ones (233 μg/g, range 37–895 μg/g) (p = 0.001). A correlation was found that from 0 to 5 months, fecal calprotectin was negatively and significantly associated with age in both two kinds of feeding (breast fed: Spearman's rho − 0.346, p = 0.010; nonbreast fed: Spearman's rho −0.478, p b 0.001). Conclusions: Our findings show that the kind of feeding influences the fecal calprotectin concentration and breast fed infants have higher levels than nonbreast fed ones in the first months of life. This may represent that human milk influences the gut mucosa by immunomodulating factors. © 2014 Elsevier Ireland Ltd. All rights reserved.
1. Introduction Calprotectin, a 36-kDa calcium and zinc-binding protein, constitutes approximately 60% of the cytosolic protein of neutrophils, monocytes and macrophages [1], composed of 8 and 14 kDa subunits [2]. Therefore, the presence of calprotectin in feces can be seen as proportional to neutrophil migration to the gastrointestinal mucosa. Calprotectin has immunomodulating and antiproliferative effects as well as an important role in neutrophil defense against bacterial infections [3]. Neutrophil transepithelial migration and accumulation at mucosal surfaces and within the gut lumen are a hallmark of digestive inflammatory pathology [4]. The simplicity of the methods, as well as the sensitivity, propose Abbreviations: FC, fecal calprotectin. ☆ Financial disclosure: The authors have indicated that they have no financial relationships relevant to this article to disclose. ⁎ Corresponding author at: Department of Child and Adolescent Healthcare, Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine, Shanghai Institute for Pediatric Research, MOE-Shanghai Key Laboratory of Children's Environmental Health, Shanghai Key Laboratory of Pediatric Gastroenterology and Nutrition, 1665 Kongjiang Rd, Shanghai 200092, China. Tel.: +86 21 25077023; fax: +86 21 25078875. E-mail address: [email protected] (X. Sheng).
http://dx.doi.org/10.1016/j.earlhumdev.2014.06.005 0378-3782/© 2014 Elsevier Ireland Ltd. All rights reserved.
that fecal calprotectin (FC) may be an ideal routine screening tool for the diagnosis of organic intestinal disease [5]. Currently, the gold standard for assessing intestinal inflammation is endoscopy with biopsy sampling [6], but repeated endoscopic evaluations are not feasible, especially in children [7]. Fecal markers have been found to be an important clinic test in order to evaluate the flogistic state of the whole intestinal tract [8], more accurate than serum markers [9]. Measurement of fecal calprotectin is highly useful for the diagnosis of IBD and may serve as a surrogate marker of mucosal inflammation throughout the intestinal tract [10]. It is found a significant correlation between calprotectin concentration in gut lavage fluid and intestinal permeability, suggesting that increased intestinal permeability may be a consequence of increased transepithelial migration of neutrophils [11]. It is released from activated granulocytes and inflamed epithelia, as part of the initial innate immune response [10]. A high level of calprotectin in healthy neonates may be related to higher intestinal permeability, establishment of gut flora, and response to alimentary antigens [12]. Several factors, including postnatal age and feeding method, could play a role in the maturation process of the functional integrity of the small intestine mucosa resulting in a decrease in intestinal permeability [13]. The beginning of
472
F. Li et al. / Early Human Development 90 (2014) 471–475
enteral feeding and to the type of ingested milk (maternal or formula), as well as determining its levels in cases of altered gastrointestinal function, would allow confirmation of the usefulness of calprotectin as an easy and reproducible marker of intestinal involvement [14]. Until now, controversial results have been reported on the effect of type of feeding on the level of fecal calprotectin in infants. On the one hand, Savino et al. found higher levels of calprotectin in breastfed term infants compared with formula-fed term infants [15] and Dorosko et al. observed higher calprotectin levels in healthy, exclusively breast fed infants aged 0–6 months compared to mixed-fed infants [16]. On the other hand, other studies suggest that there is no significant difference in fecal calprotectin concentrations between breast fed and formula fed infants [17,18]. The aim of our study was to determine fecal calprotectin levels in healthy infants exclusively breast fed or nonbreast fed in Shanghai of China. 2. Materials and methods 2.1. Study population This study was performed at Department of Children and Adolescent's Health Care of Xinhua Hospital affiliated Shanghai Jiaotong University School of Medicine, where infants received routine physical examination. Parents of infants were consecutively recruited from April 2013 to November 2013, parents were asked to fill in a brief questionnaire, with questions about several clinical and sociodemographic factors. At enrolment, collected clinical parameters that included gestational age, birth weight, sex, Apgar score, mode of delivery (vaginal or cesarean section), postnatal age, type of enteral feeding (exclusively breast-feeding or formula-feeding), neonatal diseases, weaning foods, symptoms, and physical examination findings were recorded before each stool sample was collected. Inclusion criteria were as follows: infant age b6 months; gestational age N 37 completed weeks; 5 minute Apgar score N7; birth weight appropriate for gestational age (2500–4000 g), and exclusively breastfed or nonbreast fed; no illnesses in the last month. The exclusively breastfed infants had to have received only their mothers' milk from birth until the time when they were recruited. Nonbreastfed infants were chosen by mothers who decided not to breast-feed at birth or formula fed infants had received a standard formula at least one month until the time of recruitment. Exclusion criteria were: any intake of steroidal or non-steroidal anti-inflammatory drugs, antibiotics or any other drug during the 2 weeks before recruitment; infants with history of any sign or symptom of infection or gastrointestinal disease (diarrhea, vomiting, fever) were excluded. 2.2. Measurement of fecal calprotectin Informed consent was obtained from each mother to collect a spot sample of feces from their infant. Parents were instructed to collect a sample of stools into a plastic container, and store it in a refrigerator at 4 °C–8 °C; the sample had to be taken or sent to the hospital within 1 day. About 1 g feces was collected from the nappy and stored immediately at −80 °C until analysis. Before analysis, frozen stool samples were thawed at room temperature, and the calprotectin concentration was determined using a commercially available enzyme-linked immunosorbent assay (Bühlmann Laboratories AG, Schönenbuch, Switzerland) that measures quantitative calprotectin as previously described in our study [19] and another study [20]. In each sample run were included blanks, standards and controls. The calprotectin cut-off level representing a positive value was equal or greater than 50 μg/g as stated by the manufacturer. Results were expressed as μg/g stool. Informed consent was obtained by parents at enrolment. The study protocol was approved by the ethics committee of Xinhua Hospital affiliated to Shanghai Jiao Tong University School of Medicine.
2.3. Statistical analysis The results are presented as median and range. All statistical analyses were performed with the Statistical Package for the Social Sciences for Windows software 17.0 (SPSS, Chicago, IL, USA). Statistical analyses between groups were carried out with Mann–Whitney U test and Kruskal–Wallis tests. Spearman's correlation test was used to evaluate the relationship between selected variables and fecal calprotectin values. Level of statistical significance was set at 0.05. 3. Result 3.1. Subjects characteristics The study population included 105 infants (63 boys, 42 girls) born at a median gestational age of 38.9 weeks (range 37–42 weeks) with a median birth weight of 3262 g (range 2500–4000 g). Forty were born by cesarean section and 65 by vaginal delivery. Fifty four newborns (32 boys, 22 girls) were breast fed, and Fifty one newborns (31 boys, 20 girls) were nonbreast fed. The age of the infants was 2.86 ± 1.31 months (range 1–5.88) (mean ± SD, breast-fed group 2.77 ± 1.29 months and nonbreast-fed group 2.96 ± 1.35 months). Gestational age, birth weight, gender of the two groups, age at the time of fecal sampling, and weight at sample collection were similar. The characteristics of the subjects are presented in Table 1. 3.2. Fecal calprotectin concentrations The median fecal calprotectin level calculated on all 105 infants was 288 μg/g of feces (inter quartile, 198-482 μg/g); high interindividual variations were observed, ranging between 35 and 937 μg/g. The fecal calprotectin concentrations (median: 288 μg/g) were mainly higher than the reference value. No significant differences were found between boys and girls (p = 0.592). Infants with breast fed (n = 54) had greatly increased levels of fecal calprotectin compared to infants with nonbreast fed (n = 51; p = 0.001). Breast fed infants had fecal calprotectin levels of 377 μg/g (range 35–937 μg/g) and nonbreast fed infants had 233 μg/g (range 37–895 μg/g). Fecal calprotectin levels for the two groups are displayed in Fig. 1. There was no significant correlation between calprotectin level and birth weight (r = 0.178, p = 0.059) or gestational age (r = 0.162, p = 0.084). A correlation was found between fecal calprotectin and age, from 0 to 5 months; fecal calprotectin was negatively and significantly associated with age in breast fed infants (Spearman's rho − 0.346, p = 0.010, seen in Fig. 2) and nonbreast fed infants (Spearman's rho − 0.478, p b 0.001, seen in Fig. 3). Figs. 2 and 3 depicts fecal calprotectin concentrations according to age in the two kinds of feeding. 4. Discussion In this study, we found higher fecal calprotectin levels in healthy, exclusively breast fed infants than in nonbreast fed ones in the first 5 months of life. This correlates similarly with previous research observing higher fecal calprotectin concentrations in breast fed infants compared to formula fed ones aged 0–3 months [15]. But in other study Rouge observed that fecal calprotectin levels were significantly higher in preterm infants who received formulas as their exclusive or predominant source of feeding (n = 21) than in those fed human milk (n = 104) [21]. It is well known that higher fecal calprotectin concentrations have been found in young infants compared with concentrations in adults and healthy children, showing wide interindividual and agedependent variation [17,22]. Olafsdottir et al. [23] found that the mean fecal calprotectin levels were significantly higher in healthy infants (2–10 weeks) than those in healthy children aged N 1 y (277
F. Li et al. / Early Human Development 90 (2014) 471–475
473
Table 1 Characteristics of the infants. Characteristic
Breast fed (n = 54)
Nonbreast fed (n = 51)
P value
F/χ2
Gestational age (weeks, mean ± SD) Birth weight (g) (mean ± SD) Vaginal delivery/cesarean section Boys/girls Age at the time of fecal sampling (months) Weight at sample collection (g)
39.1 ± 1.1 3232 ± 360 20/34 32/22 2.77 ± 1.29 6002 ± 1728
38.8 ± 1.1 3294 ± 392 20/31 31/20 2.96 ± 1.35 6115 ± 1510
0.534 0.400 0.818 0.873 0.294 0.368
0.389 0.528 0.053 0.025 1.114 0.546
μg/g vs. 40 μg/g). Hestvik et al. [22] analyzed fecal calprotectin levels in healthy children; the median fecal calprotectin concentrations were 345 μg/g in 0 b 3 months, 278 μg/g in 3 b 6 months, 183 μg/g in 6 b 12 months, 75 μg/g in 1 b 4 years and 28 μg/g in 4 b 12 years in that study. Our data confirm that infants in the first months of life have high calprotectin concentrations (288 μg/g) compared with healthy children. Dorosko et al. [16] reported that the median fecal calprotectin value was 207 μg/g in healthy infants (age range 3 days–6 months). Rugtveit et al. [24] found that the median fecal calprotectin concentrations were 269 μg/g in 6 weeks and 264 μg/g in 3 months. We found a little higher median concentration in our infants than previously published values in the normal infant population [2,16,17,24,25]. The factors influencing calprotectin values in stools are not yet fully understood [15]. Our study evaluated the utility of fecal calprotectin in children in Shanghai of China; the healthy subjects in our study had different variability of fecal calprotectin too. Samples exhibited high interindividual variation, which probably reflects varying individual fecal excretion. These interindividual variations may be attributed to the method of collection, in which the sample stayed in the baby's nappy. However, Olafsdottir et al. [23] showed that this sampling method, in which some water is absorbed by the nappy, increases the calprotectin concentration by no more than 30%; although this effect may also apply to this study, all fecal samples have been collected in the same way and this will not influence the results of our study. High calprotectin concentrations in stools reflect an increase in granulocytes in the gut lumen; this could be due to the greater intestinal permeability of newborns and young infants [15,26,27]. Taylor found that formula feeding and low human milk doses are associated with increased intestinal permeability which provides further insight into the manifestation of this disease and the protection provided by human milk feeding [13]. In that study, feeding type was associated with gut
Fig. 1. Fecal calprotectin levels in the two groups. Solid lines indicate median concentrations, and the dotted line indicates the upper reference limit of calprotectin concentration (50 μg/g).
permeability with a 2.8-fold higher composite median lactulose to mannitol ratio of formula-only fed infants compared with those who received human milk through the first postnatal month [13]. Catassi and Weaver have evaluated the relationship of feeding type and intestinal permeability in infants; they all demonstrated lower intestinal permeability in human milk-fed infants [27,28]. Taylor supported the theory that artificially fed infants lack human milk factors promoting intestinal tight junction closure and/or the artificial feedings cause mucosal damage leading to increased intestinal permeability [13]. Increased permeability of the mucosal barriers may be associated with increased migration of granulocytes into the gut lumen, a phenomenon reflected by high fecal concentrations of calprotectin [17]. The high calprotectin concentrations observed may reflect increased intestinal permeability related to transepithelial migration of neutrophils as observed in adults with inflammatory bowel disease [26]. But we demonstrate that fecal calprotectin concentrations from breast fed infants are higher than those from formula fed infants, consistent with other studies [15,16]. Dorosko did not consider these findings as increased intestinal inflammation in exclusively breast fed infants, but rather as a lack of inflammation in mixed-fed infants and perhaps a protective role for calprotectin in young exclusively breast fed infants [16]. Higher fecal calprotectin values in exclusively breast fed infants than in formula fed ones could be consistent with the promotion by human milk of the growth of the intestinal mucosa, even though breast milk has been shown to promote the reduction of intestinal permeability more than cow milk-based formulas [15,29]. We also speculated that calprotectin may serve a protective role for the immature infant intestine, until epithelial tight junction proteins and other factors of intestinal immunity up-regulate with age, consistent with the study of Dorosko [16]. During the first months of life, the composition of the intestinal microflora, which is involved in mucosal immunity maturation, depends on various factors, mainly the type of feeding [17,30]. Immaturity of the immune system and the gastrointestinal tract may play a pivotal
Fig. 2. Fecal calprotectin concentrations according to age in breast fed infants.
474
F. Li et al. / Early Human Development 90 (2014) 471–475
6. Conclusion This study suggests that healthy infants in the first months of life have high calprotectin concentrations compared with children and show wide interindividual variation. Breast fed infants have higher levels than nonbreast fed ones in the first months of life. Our study does not allow us to clearly identify the reason for our finding; this may represent that human milk influences the gut mucosa by immunomodulating factors. The mechanism underlying the relationship between feeding and fecal calprotectin concentrations in young infants is needed to be clarified in further studies. Conflict of interest The authors have no conflicts of interest. Acknowledgments Fig. 3. Fecal calprotectin concentrations according to age in nonbreast fed infants.
role in the susceptibility of infants [31]. The maturation of the infant's immune system is influenced by bacterial antigens from the microflora [32]. The intestinal epithelium, in addition to present a physical barrier to microbial penetration, plays a role by producing and secreting large quantities of antimicrobial peptides [2], keeping commensal bacteria from penetrating host tissues and causing serious problems as inflammation and sepsis. Feeding with breast milk affects the intestinal colonization, metabolic and immunologic reactions by the epithelial cells and the underlying lymphoid cells, and as a consequence, the intestinal inflammatory response, partially through the action of its oligosaccharides [31,33,34]. It can be ipotized that calprotectin contributes in healthy newborns to the host defense mechanism; increased levels of fecal calprotectin in the first period of life may be a reflection of a physiological maturation of the immune system [2,31]. Breast milk feeding may stimulate this maturation process of the immune system [31]. For fecal calprotectin, our results are in line with that study which found higher values of calprotectin in breast fed infants compared with formula infants [15]. The intestinal microbiota may play a role in this process [35]. Rouge provided evidence for an effect of intestinal bacterial colonization on fecal calprotectin excretion; fecal calprotectin correlated positively with intestinal colonization by Staphylococcus and Clostridium [21]. The intestinal microbiota may stimulate transepithelial migration of granulocytes through chemotactic agents [36]. In the immature gastrointestinal tract of infants, fecal calprotectin may provide a protective effect on the gut mucosa through its antimicrobial and antifungal effects [15,37]. Furthermore, fecal calprotectin may participate in the regulation of the intestinal bacterial colonization of the immature gut and may inhibit potential pathogenic bacterial growth [31].
5. Limitations Our study has several limitations. First, the sample was relatively small, and this did not allow reliable intra-group analysis, and calprotectin was the only marker studied. Therefore, we have no information about the permeability of the gut, but we aimed only to define calprotectin modifications according to the type of milk. Second, nonbreast fed infants may include some infants who had ever breast fed at birth or first days of life. But that did not influence our result. Third, we did not follow up the children with elevated fecal calprotectin concentrations to see if the concentrations normalized over time, so we do not have an explanation for fecal calprotectin higher than 600 μg/g. Also, we did not evaluate the effects on breastfed infants' calprotectin of a dairy product-free diet in their mothers.
This study was funded by Shanghai Key Laboratory of Pediatric Gastroenterology and Nutrition (No. 11DZ2260500), National Natural Science Foundation of China (No. 81172686, No. 81302446) and Shanghai Science and Technology Commission Pujiang Talent Program of China (No. 08PJ14082). References [1] Dale I, Brandtzaeg P, Fagerhol MK, Scott H. Distribution of a new myelomonocytic antigen (L1) in human peripheral blood leukocytes. Immunofluorescence and immunoperoxidase staining features in comparison with lysozyme and lactoferrin. Am J Clin Pathol 1985;84:24–34. [2] Baldassarre ME, Altomare MA, Fanelli M, Carbone D, Di Bitonto G, Mautone A, et al. Does calprotectin represent a regulatory factor in host defense or a drug target in inflammatory disease? Endocr Metab Immune Disord Drug Targets 2007;7:1–5. [3] Chitkara DK, Talley NJ, Schleck C, Zinsmeister AR, Shah ND, Locke III GR. Recollection of childhood abdominal pain in adults with functional gastrointestinal disorders. Scand J Gastroenterol 2009;44:301–7. [4] Rasquin A, Di Lorenzo C, Forbes D, Guiraldes E, Hyams JS, Staiano A, et al. Childhood functional gastrointestinal disorders: child/adolescent. Gastroenterology 2006;130:1527–37. [5] Licata A, Randazzo C, Cappello M, Calvaruso V, Butera G, Florena AM, et al. Fecal calprotectin in clinical practice: a noninvasive screening tool for patients with chronic diarrhea. J Clin Gastroenterol 2012;46:504–8. [6] Diamanti A, Colistro F, Basso MS, Papadatou B, Francalanci P, Bracci F, et al. Clinical role of calprotectin assay in determining histological relapses in children affected by inflammatory bowel diseases. Inflamm Bowel Dis 2008;14:1229–35. [7] Turner D, Leach ST, Mack D, Uusoue K, McLernon R, Hyams J, et al. Faecal calprotectin, lactoferrin, M2-pyruvate kinase and S100A12 in severe ulcerative colitis: a prospective multicentre comparison of predicting outcomes and monitoring response. Gut 2010;59:1207–12. [8] Baldassarre ME, Fanelli M, Lasorella ML, Laneve A, Grosso R, Falcone MR, et al. Fecal calprotectin (FC) in newborns: is it a predictive marker of gastrointestinal and/or allergic disease? Immunopharmacol Immunotoxicol 2011;33:220–3. [9] Abraham BP, Kane S. Fecal markers: calprotectin and lactoferrin. Gastroenterol Clin North Am 2012;41:483–95. [10] Logan R. Faecal calprotectin for the diagnosis of inflammatory bowel disease. BMJ 2010;341:c3636. [11] Morisset M, Aubert-Jacquin C, Soulaines P, Moneret-Vautrin DA, Dupont C. A nonhydrolyzed, fermented milk formula reduces digestive and respiratory events in infants at high risk of allergy. Eur J Clin Nutr 2011;65:175–83. [12] Kapel N, Campeotto F, Kalach N, Baldassare M, Butel MJ, Dupont C. Faecal calprotectin in term and preterm neonates. J Pediatr Gastroenterol Nutr 2010;51:542–7. [13] Taylor SN, Basile LA, Ebeling M, Wagner CL. Intestinal permeability in preterm infants by feeding type: mother's milk versus formula. Breastfeed Med 2009;4:11–5. [14] Laforgia N, Baldassarre ME, Pontrelli G, Indrio F, Altomare MA, Di Bitonto G, et al. Calprotectin levels in meconium. Acta Paediatr 2003;92:463–6. [15] Savino F, Castagno E, Calabrese R, Viola S, Oggero R, Miniero R. High faecal calprotectin levels in healthy, exclusively breast-fed infants. Neonatology 2010;97:299–304. [16] Dorosko SM, Mackenzie T, Connor RI. Fecal calprotectin concentrations are higher in exclusively breastfed infants compared to those who are mixed-fed. Breastfeed Med 2008;3:117–9. [17] Campeotto F, Butel MJ, Kalach N, Derrieux S, Aubert-Jacquin C, Barbot L, et al. High faecal calprotectin concentrations in newborn infants. Arch Dis Child Fetal Neonatal Ed 2004;89:F353–5. [18] Rosti L, Braga M, Fulcieri C, Sammarco G, Manenti B, Costa E. Formula milk feeding does not increase the release of the inflammatory marker calprotectin, compared to human milk. Pediatr Med Chir 2011;33:178–81.
F. Li et al. / Early Human Development 90 (2014) 471–475 [19] Liu JR, Sheng XY, Hu YQ, Yu XY, Westcott J, Miller LV, et al. Fecal calprotectin levels are higher in rural than in urban Chinese infants and negatively associated with growth. BMC Pediatr 2012;12:129. [20] Manz M, Burri E, Rothen C, Tchanguizi N, Niederberger C, Rossi L, et al. Value of fecal calprotectin in the evaluation of patients with abdominal discomfort: an observational study. BMC Gastroenterol 2012;12:5. [21] Rouge C, Butel MJ, Piloquet H, Ferraris L, Legrand A, Vodovar M, et al. Fecal calprotectin excretion in preterm infants during the neonatal period. PLoS One 2010;5:e11083. [22] Hestvik E, Tumwine JK, Tylleskar T, Grahnquist L, Ndeezi G, Kaddu-Mulindwa DH, et al. Faecal calprotectin concentrations in apparently healthy children aged 0–12 years in urban Kampala, Uganda: a community-based survey. BMC Pediatr 2011;11:9. [23] Olafsdottir E, Aksnes L, Fluge G, Berstad A. Faecal calprotectin levels in infants with infantile colic, healthy infants, children with inflammatory bowel disease, children with recurrent abdominal pain and healthy children. Acta Paediatr 2002;91:45–50. [24] Rugtveit J, Fagerhol MK. Age-dependent variations in fecal calprotectin concentrations in children. J Pediatr Gastroenterol Nutr 2002;34:323–4 [author reply 4–5]. [25] Rhoads JM, Fatheree NY, Norori J, Liu Y, Lucke JF, Tyson JE, et al. Altered fecal microflora and increased fecal calprotectin in infants with colic. J Pediatr 2009;155:823–8 [e1]. [26] Berstad A, Arslan G, Folvik G. Relationship between intestinal permeability and calprotectin concentration in gut lavage fluid. Scand J Gastroenterol 2000;35:64–9. [27] Catassi C, Bonucci A, Coppa GV, Carlucci A, Giorgi PL. Intestinal permeability changes during the first month: effect of natural versus artificial feeding. J Pediatr Gastroenterol Nutr 1995;21:383–6.
475
[28] Weaver LT, Laker MF, Nelson R, Lucas A. Milk feeding and changes in intestinal permeability and morphology in the newborn. J Pediatr Gastroenterol Nutr 1987;6:351–8. [29] Wagner CL, Taylor SN, Johnson D. Host factors in amniotic fluid and breast milk that contribute to gut maturation. Clin Rev Allergy Immunol 2008;34:191–204. [30] Moreau MC, Gaboriau-Routhiau V. Influence of resident intestinal microflora on the development and functions of the gut-associated lymphoid tissue. Microb Ecol Health Dis 2001;13:65–86. [31] Westerbeek EA, Morch E, Lafeber HN, Fetter WP, Twisk JW, Van Elburg RM. Effect of neutral and acidic oligosaccharides on fecal IL-8 and fecal calprotectin in preterm infants. Pediatr Res 2011;69:255–8. [32] Field CJ. The immunological components of human milk and their effect on immune development in infants. J Nutr 2005;135:1–4. [33] Bode L. Recent advances on structure, metabolism, and function of human milk oligosaccharides. J Nutr 2006;136:2127–30. [34] Boehm G, Moro G. Structural and functional aspects of prebiotics used in infant nutrition. J Nutr 2008;138:1818S–28S. [35] Lu L, Walker WA. Pathologic and physiologic interactions of bacteria with the gastrointestinal epithelium. Am J Clin Nutr 2001;73:1124S–30S. [36] Nash S, Stafford J, Madara JL. Effects of polymorphonuclear leukocyte transmigration on the barrier function of cultured intestinal epithelial monolayers. J Clin Invest 1987;80:1104–13. [37] Steinbakk M, Naess-Andresen CF, Lingaas E, Dale I, Brandtzaeg P, Fagerhol MK. Antimicrobial actions of calcium binding leucocyte L1 protein, calprotectin. Lancet 1990;336:763–5.
Contents lists available at ScienceDirect
Early Human Development journal homepage: www.elsevier.com/locate/earlhumdev
Comparison of the different kinds of feeding on the level of fecal calprotectin☆ Feng Li, Jingqiu Ma, Shanshan Geng, Junli Wang, Fang Ren, Xiaoyang Sheng ⁎ Department of Children and Adolescents Health Care, MOE-Shanghai Key Laboratory of Children's Environmental Health, Xin Hua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
a r t i c l e
i n f o
Article history: Received 7 May 2014 Accepted 14 June 2014 Available online xxxx Keywords: Fecal calprotectin Breast-feeding Formula-feeding Infants
a b s t r a c t Background: Controversial results have been reported on the effect of type of feeding on the level of fecal calprotectin in infants. Objective: To assess fecal calprotectin levels in breast fed or nonbreast fed healthy infants. Design: A study was conducted to compare fecal calprotectin in infants who were exclusively breastfed compared to those who were not breastfed in Shanghai, China. Stool samples were collected and analyzed, and the fecal calprotectin concentration was determined using a commercially available enzyme-linked immunosorbent assay. The infant's weight and length were measured. Parents were asked to fill in a brief questionnaire, with questions about several clinical and sociodemographic factors. Subjects: This study included 105 healthy infants aged 0–5 months. Results: Stool samples were obtained from 105 healthy infants (63 boys, 42 girls) with a median age of 2.86 months (range 1–5.88). The median fecal calprotectin concentration was significantly higher in breast fed infants (377 μg/g, range 35–937 μg/g) than that in nonbreast fed ones (233 μg/g, range 37–895 μg/g) (p = 0.001). A correlation was found that from 0 to 5 months, fecal calprotectin was negatively and significantly associated with age in both two kinds of feeding (breast fed: Spearman's rho − 0.346, p = 0.010; nonbreast fed: Spearman's rho −0.478, p b 0.001). Conclusions: Our findings show that the kind of feeding influences the fecal calprotectin concentration and breast fed infants have higher levels than nonbreast fed ones in the first months of life. This may represent that human milk influences the gut mucosa by immunomodulating factors. © 2014 Elsevier Ireland Ltd. All rights reserved.
1. Introduction Calprotectin, a 36-kDa calcium and zinc-binding protein, constitutes approximately 60% of the cytosolic protein of neutrophils, monocytes and macrophages [1], composed of 8 and 14 kDa subunits [2]. Therefore, the presence of calprotectin in feces can be seen as proportional to neutrophil migration to the gastrointestinal mucosa. Calprotectin has immunomodulating and antiproliferative effects as well as an important role in neutrophil defense against bacterial infections [3]. Neutrophil transepithelial migration and accumulation at mucosal surfaces and within the gut lumen are a hallmark of digestive inflammatory pathology [4]. The simplicity of the methods, as well as the sensitivity, propose Abbreviations: FC, fecal calprotectin. ☆ Financial disclosure: The authors have indicated that they have no financial relationships relevant to this article to disclose. ⁎ Corresponding author at: Department of Child and Adolescent Healthcare, Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine, Shanghai Institute for Pediatric Research, MOE-Shanghai Key Laboratory of Children's Environmental Health, Shanghai Key Laboratory of Pediatric Gastroenterology and Nutrition, 1665 Kongjiang Rd, Shanghai 200092, China. Tel.: +86 21 25077023; fax: +86 21 25078875. E-mail address: [email protected] (X. Sheng).
http://dx.doi.org/10.1016/j.earlhumdev.2014.06.005 0378-3782/© 2014 Elsevier Ireland Ltd. All rights reserved.
that fecal calprotectin (FC) may be an ideal routine screening tool for the diagnosis of organic intestinal disease [5]. Currently, the gold standard for assessing intestinal inflammation is endoscopy with biopsy sampling [6], but repeated endoscopic evaluations are not feasible, especially in children [7]. Fecal markers have been found to be an important clinic test in order to evaluate the flogistic state of the whole intestinal tract [8], more accurate than serum markers [9]. Measurement of fecal calprotectin is highly useful for the diagnosis of IBD and may serve as a surrogate marker of mucosal inflammation throughout the intestinal tract [10]. It is found a significant correlation between calprotectin concentration in gut lavage fluid and intestinal permeability, suggesting that increased intestinal permeability may be a consequence of increased transepithelial migration of neutrophils [11]. It is released from activated granulocytes and inflamed epithelia, as part of the initial innate immune response [10]. A high level of calprotectin in healthy neonates may be related to higher intestinal permeability, establishment of gut flora, and response to alimentary antigens [12]. Several factors, including postnatal age and feeding method, could play a role in the maturation process of the functional integrity of the small intestine mucosa resulting in a decrease in intestinal permeability [13]. The beginning of
472
F. Li et al. / Early Human Development 90 (2014) 471–475
enteral feeding and to the type of ingested milk (maternal or formula), as well as determining its levels in cases of altered gastrointestinal function, would allow confirmation of the usefulness of calprotectin as an easy and reproducible marker of intestinal involvement [14]. Until now, controversial results have been reported on the effect of type of feeding on the level of fecal calprotectin in infants. On the one hand, Savino et al. found higher levels of calprotectin in breastfed term infants compared with formula-fed term infants [15] and Dorosko et al. observed higher calprotectin levels in healthy, exclusively breast fed infants aged 0–6 months compared to mixed-fed infants [16]. On the other hand, other studies suggest that there is no significant difference in fecal calprotectin concentrations between breast fed and formula fed infants [17,18]. The aim of our study was to determine fecal calprotectin levels in healthy infants exclusively breast fed or nonbreast fed in Shanghai of China. 2. Materials and methods 2.1. Study population This study was performed at Department of Children and Adolescent's Health Care of Xinhua Hospital affiliated Shanghai Jiaotong University School of Medicine, where infants received routine physical examination. Parents of infants were consecutively recruited from April 2013 to November 2013, parents were asked to fill in a brief questionnaire, with questions about several clinical and sociodemographic factors. At enrolment, collected clinical parameters that included gestational age, birth weight, sex, Apgar score, mode of delivery (vaginal or cesarean section), postnatal age, type of enteral feeding (exclusively breast-feeding or formula-feeding), neonatal diseases, weaning foods, symptoms, and physical examination findings were recorded before each stool sample was collected. Inclusion criteria were as follows: infant age b6 months; gestational age N 37 completed weeks; 5 minute Apgar score N7; birth weight appropriate for gestational age (2500–4000 g), and exclusively breastfed or nonbreast fed; no illnesses in the last month. The exclusively breastfed infants had to have received only their mothers' milk from birth until the time when they were recruited. Nonbreastfed infants were chosen by mothers who decided not to breast-feed at birth or formula fed infants had received a standard formula at least one month until the time of recruitment. Exclusion criteria were: any intake of steroidal or non-steroidal anti-inflammatory drugs, antibiotics or any other drug during the 2 weeks before recruitment; infants with history of any sign or symptom of infection or gastrointestinal disease (diarrhea, vomiting, fever) were excluded. 2.2. Measurement of fecal calprotectin Informed consent was obtained from each mother to collect a spot sample of feces from their infant. Parents were instructed to collect a sample of stools into a plastic container, and store it in a refrigerator at 4 °C–8 °C; the sample had to be taken or sent to the hospital within 1 day. About 1 g feces was collected from the nappy and stored immediately at −80 °C until analysis. Before analysis, frozen stool samples were thawed at room temperature, and the calprotectin concentration was determined using a commercially available enzyme-linked immunosorbent assay (Bühlmann Laboratories AG, Schönenbuch, Switzerland) that measures quantitative calprotectin as previously described in our study [19] and another study [20]. In each sample run were included blanks, standards and controls. The calprotectin cut-off level representing a positive value was equal or greater than 50 μg/g as stated by the manufacturer. Results were expressed as μg/g stool. Informed consent was obtained by parents at enrolment. The study protocol was approved by the ethics committee of Xinhua Hospital affiliated to Shanghai Jiao Tong University School of Medicine.
2.3. Statistical analysis The results are presented as median and range. All statistical analyses were performed with the Statistical Package for the Social Sciences for Windows software 17.0 (SPSS, Chicago, IL, USA). Statistical analyses between groups were carried out with Mann–Whitney U test and Kruskal–Wallis tests. Spearman's correlation test was used to evaluate the relationship between selected variables and fecal calprotectin values. Level of statistical significance was set at 0.05. 3. Result 3.1. Subjects characteristics The study population included 105 infants (63 boys, 42 girls) born at a median gestational age of 38.9 weeks (range 37–42 weeks) with a median birth weight of 3262 g (range 2500–4000 g). Forty were born by cesarean section and 65 by vaginal delivery. Fifty four newborns (32 boys, 22 girls) were breast fed, and Fifty one newborns (31 boys, 20 girls) were nonbreast fed. The age of the infants was 2.86 ± 1.31 months (range 1–5.88) (mean ± SD, breast-fed group 2.77 ± 1.29 months and nonbreast-fed group 2.96 ± 1.35 months). Gestational age, birth weight, gender of the two groups, age at the time of fecal sampling, and weight at sample collection were similar. The characteristics of the subjects are presented in Table 1. 3.2. Fecal calprotectin concentrations The median fecal calprotectin level calculated on all 105 infants was 288 μg/g of feces (inter quartile, 198-482 μg/g); high interindividual variations were observed, ranging between 35 and 937 μg/g. The fecal calprotectin concentrations (median: 288 μg/g) were mainly higher than the reference value. No significant differences were found between boys and girls (p = 0.592). Infants with breast fed (n = 54) had greatly increased levels of fecal calprotectin compared to infants with nonbreast fed (n = 51; p = 0.001). Breast fed infants had fecal calprotectin levels of 377 μg/g (range 35–937 μg/g) and nonbreast fed infants had 233 μg/g (range 37–895 μg/g). Fecal calprotectin levels for the two groups are displayed in Fig. 1. There was no significant correlation between calprotectin level and birth weight (r = 0.178, p = 0.059) or gestational age (r = 0.162, p = 0.084). A correlation was found between fecal calprotectin and age, from 0 to 5 months; fecal calprotectin was negatively and significantly associated with age in breast fed infants (Spearman's rho − 0.346, p = 0.010, seen in Fig. 2) and nonbreast fed infants (Spearman's rho − 0.478, p b 0.001, seen in Fig. 3). Figs. 2 and 3 depicts fecal calprotectin concentrations according to age in the two kinds of feeding. 4. Discussion In this study, we found higher fecal calprotectin levels in healthy, exclusively breast fed infants than in nonbreast fed ones in the first 5 months of life. This correlates similarly with previous research observing higher fecal calprotectin concentrations in breast fed infants compared to formula fed ones aged 0–3 months [15]. But in other study Rouge observed that fecal calprotectin levels were significantly higher in preterm infants who received formulas as their exclusive or predominant source of feeding (n = 21) than in those fed human milk (n = 104) [21]. It is well known that higher fecal calprotectin concentrations have been found in young infants compared with concentrations in adults and healthy children, showing wide interindividual and agedependent variation [17,22]. Olafsdottir et al. [23] found that the mean fecal calprotectin levels were significantly higher in healthy infants (2–10 weeks) than those in healthy children aged N 1 y (277
F. Li et al. / Early Human Development 90 (2014) 471–475
473
Table 1 Characteristics of the infants. Characteristic
Breast fed (n = 54)
Nonbreast fed (n = 51)
P value
F/χ2
Gestational age (weeks, mean ± SD) Birth weight (g) (mean ± SD) Vaginal delivery/cesarean section Boys/girls Age at the time of fecal sampling (months) Weight at sample collection (g)
39.1 ± 1.1 3232 ± 360 20/34 32/22 2.77 ± 1.29 6002 ± 1728
38.8 ± 1.1 3294 ± 392 20/31 31/20 2.96 ± 1.35 6115 ± 1510
0.534 0.400 0.818 0.873 0.294 0.368
0.389 0.528 0.053 0.025 1.114 0.546
μg/g vs. 40 μg/g). Hestvik et al. [22] analyzed fecal calprotectin levels in healthy children; the median fecal calprotectin concentrations were 345 μg/g in 0 b 3 months, 278 μg/g in 3 b 6 months, 183 μg/g in 6 b 12 months, 75 μg/g in 1 b 4 years and 28 μg/g in 4 b 12 years in that study. Our data confirm that infants in the first months of life have high calprotectin concentrations (288 μg/g) compared with healthy children. Dorosko et al. [16] reported that the median fecal calprotectin value was 207 μg/g in healthy infants (age range 3 days–6 months). Rugtveit et al. [24] found that the median fecal calprotectin concentrations were 269 μg/g in 6 weeks and 264 μg/g in 3 months. We found a little higher median concentration in our infants than previously published values in the normal infant population [2,16,17,24,25]. The factors influencing calprotectin values in stools are not yet fully understood [15]. Our study evaluated the utility of fecal calprotectin in children in Shanghai of China; the healthy subjects in our study had different variability of fecal calprotectin too. Samples exhibited high interindividual variation, which probably reflects varying individual fecal excretion. These interindividual variations may be attributed to the method of collection, in which the sample stayed in the baby's nappy. However, Olafsdottir et al. [23] showed that this sampling method, in which some water is absorbed by the nappy, increases the calprotectin concentration by no more than 30%; although this effect may also apply to this study, all fecal samples have been collected in the same way and this will not influence the results of our study. High calprotectin concentrations in stools reflect an increase in granulocytes in the gut lumen; this could be due to the greater intestinal permeability of newborns and young infants [15,26,27]. Taylor found that formula feeding and low human milk doses are associated with increased intestinal permeability which provides further insight into the manifestation of this disease and the protection provided by human milk feeding [13]. In that study, feeding type was associated with gut
Fig. 1. Fecal calprotectin levels in the two groups. Solid lines indicate median concentrations, and the dotted line indicates the upper reference limit of calprotectin concentration (50 μg/g).
permeability with a 2.8-fold higher composite median lactulose to mannitol ratio of formula-only fed infants compared with those who received human milk through the first postnatal month [13]. Catassi and Weaver have evaluated the relationship of feeding type and intestinal permeability in infants; they all demonstrated lower intestinal permeability in human milk-fed infants [27,28]. Taylor supported the theory that artificially fed infants lack human milk factors promoting intestinal tight junction closure and/or the artificial feedings cause mucosal damage leading to increased intestinal permeability [13]. Increased permeability of the mucosal barriers may be associated with increased migration of granulocytes into the gut lumen, a phenomenon reflected by high fecal concentrations of calprotectin [17]. The high calprotectin concentrations observed may reflect increased intestinal permeability related to transepithelial migration of neutrophils as observed in adults with inflammatory bowel disease [26]. But we demonstrate that fecal calprotectin concentrations from breast fed infants are higher than those from formula fed infants, consistent with other studies [15,16]. Dorosko did not consider these findings as increased intestinal inflammation in exclusively breast fed infants, but rather as a lack of inflammation in mixed-fed infants and perhaps a protective role for calprotectin in young exclusively breast fed infants [16]. Higher fecal calprotectin values in exclusively breast fed infants than in formula fed ones could be consistent with the promotion by human milk of the growth of the intestinal mucosa, even though breast milk has been shown to promote the reduction of intestinal permeability more than cow milk-based formulas [15,29]. We also speculated that calprotectin may serve a protective role for the immature infant intestine, until epithelial tight junction proteins and other factors of intestinal immunity up-regulate with age, consistent with the study of Dorosko [16]. During the first months of life, the composition of the intestinal microflora, which is involved in mucosal immunity maturation, depends on various factors, mainly the type of feeding [17,30]. Immaturity of the immune system and the gastrointestinal tract may play a pivotal
Fig. 2. Fecal calprotectin concentrations according to age in breast fed infants.
474
F. Li et al. / Early Human Development 90 (2014) 471–475
6. Conclusion This study suggests that healthy infants in the first months of life have high calprotectin concentrations compared with children and show wide interindividual variation. Breast fed infants have higher levels than nonbreast fed ones in the first months of life. Our study does not allow us to clearly identify the reason for our finding; this may represent that human milk influences the gut mucosa by immunomodulating factors. The mechanism underlying the relationship between feeding and fecal calprotectin concentrations in young infants is needed to be clarified in further studies. Conflict of interest The authors have no conflicts of interest. Acknowledgments Fig. 3. Fecal calprotectin concentrations according to age in nonbreast fed infants.
role in the susceptibility of infants [31]. The maturation of the infant's immune system is influenced by bacterial antigens from the microflora [32]. The intestinal epithelium, in addition to present a physical barrier to microbial penetration, plays a role by producing and secreting large quantities of antimicrobial peptides [2], keeping commensal bacteria from penetrating host tissues and causing serious problems as inflammation and sepsis. Feeding with breast milk affects the intestinal colonization, metabolic and immunologic reactions by the epithelial cells and the underlying lymphoid cells, and as a consequence, the intestinal inflammatory response, partially through the action of its oligosaccharides [31,33,34]. It can be ipotized that calprotectin contributes in healthy newborns to the host defense mechanism; increased levels of fecal calprotectin in the first period of life may be a reflection of a physiological maturation of the immune system [2,31]. Breast milk feeding may stimulate this maturation process of the immune system [31]. For fecal calprotectin, our results are in line with that study which found higher values of calprotectin in breast fed infants compared with formula infants [15]. The intestinal microbiota may play a role in this process [35]. Rouge provided evidence for an effect of intestinal bacterial colonization on fecal calprotectin excretion; fecal calprotectin correlated positively with intestinal colonization by Staphylococcus and Clostridium [21]. The intestinal microbiota may stimulate transepithelial migration of granulocytes through chemotactic agents [36]. In the immature gastrointestinal tract of infants, fecal calprotectin may provide a protective effect on the gut mucosa through its antimicrobial and antifungal effects [15,37]. Furthermore, fecal calprotectin may participate in the regulation of the intestinal bacterial colonization of the immature gut and may inhibit potential pathogenic bacterial growth [31].
5. Limitations Our study has several limitations. First, the sample was relatively small, and this did not allow reliable intra-group analysis, and calprotectin was the only marker studied. Therefore, we have no information about the permeability of the gut, but we aimed only to define calprotectin modifications according to the type of milk. Second, nonbreast fed infants may include some infants who had ever breast fed at birth or first days of life. But that did not influence our result. Third, we did not follow up the children with elevated fecal calprotectin concentrations to see if the concentrations normalized over time, so we do not have an explanation for fecal calprotectin higher than 600 μg/g. Also, we did not evaluate the effects on breastfed infants' calprotectin of a dairy product-free diet in their mothers.
This study was funded by Shanghai Key Laboratory of Pediatric Gastroenterology and Nutrition (No. 11DZ2260500), National Natural Science Foundation of China (No. 81172686, No. 81302446) and Shanghai Science and Technology Commission Pujiang Talent Program of China (No. 08PJ14082). References [1] Dale I, Brandtzaeg P, Fagerhol MK, Scott H. Distribution of a new myelomonocytic antigen (L1) in human peripheral blood leukocytes. Immunofluorescence and immunoperoxidase staining features in comparison with lysozyme and lactoferrin. Am J Clin Pathol 1985;84:24–34. [2] Baldassarre ME, Altomare MA, Fanelli M, Carbone D, Di Bitonto G, Mautone A, et al. Does calprotectin represent a regulatory factor in host defense or a drug target in inflammatory disease? Endocr Metab Immune Disord Drug Targets 2007;7:1–5. [3] Chitkara DK, Talley NJ, Schleck C, Zinsmeister AR, Shah ND, Locke III GR. Recollection of childhood abdominal pain in adults with functional gastrointestinal disorders. Scand J Gastroenterol 2009;44:301–7. [4] Rasquin A, Di Lorenzo C, Forbes D, Guiraldes E, Hyams JS, Staiano A, et al. Childhood functional gastrointestinal disorders: child/adolescent. Gastroenterology 2006;130:1527–37. [5] Licata A, Randazzo C, Cappello M, Calvaruso V, Butera G, Florena AM, et al. Fecal calprotectin in clinical practice: a noninvasive screening tool for patients with chronic diarrhea. J Clin Gastroenterol 2012;46:504–8. [6] Diamanti A, Colistro F, Basso MS, Papadatou B, Francalanci P, Bracci F, et al. Clinical role of calprotectin assay in determining histological relapses in children affected by inflammatory bowel diseases. Inflamm Bowel Dis 2008;14:1229–35. [7] Turner D, Leach ST, Mack D, Uusoue K, McLernon R, Hyams J, et al. Faecal calprotectin, lactoferrin, M2-pyruvate kinase and S100A12 in severe ulcerative colitis: a prospective multicentre comparison of predicting outcomes and monitoring response. Gut 2010;59:1207–12. [8] Baldassarre ME, Fanelli M, Lasorella ML, Laneve A, Grosso R, Falcone MR, et al. Fecal calprotectin (FC) in newborns: is it a predictive marker of gastrointestinal and/or allergic disease? Immunopharmacol Immunotoxicol 2011;33:220–3. [9] Abraham BP, Kane S. Fecal markers: calprotectin and lactoferrin. Gastroenterol Clin North Am 2012;41:483–95. [10] Logan R. Faecal calprotectin for the diagnosis of inflammatory bowel disease. BMJ 2010;341:c3636. [11] Morisset M, Aubert-Jacquin C, Soulaines P, Moneret-Vautrin DA, Dupont C. A nonhydrolyzed, fermented milk formula reduces digestive and respiratory events in infants at high risk of allergy. Eur J Clin Nutr 2011;65:175–83. [12] Kapel N, Campeotto F, Kalach N, Baldassare M, Butel MJ, Dupont C. Faecal calprotectin in term and preterm neonates. J Pediatr Gastroenterol Nutr 2010;51:542–7. [13] Taylor SN, Basile LA, Ebeling M, Wagner CL. Intestinal permeability in preterm infants by feeding type: mother's milk versus formula. Breastfeed Med 2009;4:11–5. [14] Laforgia N, Baldassarre ME, Pontrelli G, Indrio F, Altomare MA, Di Bitonto G, et al. Calprotectin levels in meconium. Acta Paediatr 2003;92:463–6. [15] Savino F, Castagno E, Calabrese R, Viola S, Oggero R, Miniero R. High faecal calprotectin levels in healthy, exclusively breast-fed infants. Neonatology 2010;97:299–304. [16] Dorosko SM, Mackenzie T, Connor RI. Fecal calprotectin concentrations are higher in exclusively breastfed infants compared to those who are mixed-fed. Breastfeed Med 2008;3:117–9. [17] Campeotto F, Butel MJ, Kalach N, Derrieux S, Aubert-Jacquin C, Barbot L, et al. High faecal calprotectin concentrations in newborn infants. Arch Dis Child Fetal Neonatal Ed 2004;89:F353–5. [18] Rosti L, Braga M, Fulcieri C, Sammarco G, Manenti B, Costa E. Formula milk feeding does not increase the release of the inflammatory marker calprotectin, compared to human milk. Pediatr Med Chir 2011;33:178–81.
F. Li et al. / Early Human Development 90 (2014) 471–475 [19] Liu JR, Sheng XY, Hu YQ, Yu XY, Westcott J, Miller LV, et al. Fecal calprotectin levels are higher in rural than in urban Chinese infants and negatively associated with growth. BMC Pediatr 2012;12:129. [20] Manz M, Burri E, Rothen C, Tchanguizi N, Niederberger C, Rossi L, et al. Value of fecal calprotectin in the evaluation of patients with abdominal discomfort: an observational study. BMC Gastroenterol 2012;12:5. [21] Rouge C, Butel MJ, Piloquet H, Ferraris L, Legrand A, Vodovar M, et al. Fecal calprotectin excretion in preterm infants during the neonatal period. PLoS One 2010;5:e11083. [22] Hestvik E, Tumwine JK, Tylleskar T, Grahnquist L, Ndeezi G, Kaddu-Mulindwa DH, et al. Faecal calprotectin concentrations in apparently healthy children aged 0–12 years in urban Kampala, Uganda: a community-based survey. BMC Pediatr 2011;11:9. [23] Olafsdottir E, Aksnes L, Fluge G, Berstad A. Faecal calprotectin levels in infants with infantile colic, healthy infants, children with inflammatory bowel disease, children with recurrent abdominal pain and healthy children. Acta Paediatr 2002;91:45–50. [24] Rugtveit J, Fagerhol MK. Age-dependent variations in fecal calprotectin concentrations in children. J Pediatr Gastroenterol Nutr 2002;34:323–4 [author reply 4–5]. [25] Rhoads JM, Fatheree NY, Norori J, Liu Y, Lucke JF, Tyson JE, et al. Altered fecal microflora and increased fecal calprotectin in infants with colic. J Pediatr 2009;155:823–8 [e1]. [26] Berstad A, Arslan G, Folvik G. Relationship between intestinal permeability and calprotectin concentration in gut lavage fluid. Scand J Gastroenterol 2000;35:64–9. [27] Catassi C, Bonucci A, Coppa GV, Carlucci A, Giorgi PL. Intestinal permeability changes during the first month: effect of natural versus artificial feeding. J Pediatr Gastroenterol Nutr 1995;21:383–6.
475
[28] Weaver LT, Laker MF, Nelson R, Lucas A. Milk feeding and changes in intestinal permeability and morphology in the newborn. J Pediatr Gastroenterol Nutr 1987;6:351–8. [29] Wagner CL, Taylor SN, Johnson D. Host factors in amniotic fluid and breast milk that contribute to gut maturation. Clin Rev Allergy Immunol 2008;34:191–204. [30] Moreau MC, Gaboriau-Routhiau V. Influence of resident intestinal microflora on the development and functions of the gut-associated lymphoid tissue. Microb Ecol Health Dis 2001;13:65–86. [31] Westerbeek EA, Morch E, Lafeber HN, Fetter WP, Twisk JW, Van Elburg RM. Effect of neutral and acidic oligosaccharides on fecal IL-8 and fecal calprotectin in preterm infants. Pediatr Res 2011;69:255–8. [32] Field CJ. The immunological components of human milk and their effect on immune development in infants. J Nutr 2005;135:1–4. [33] Bode L. Recent advances on structure, metabolism, and function of human milk oligosaccharides. J Nutr 2006;136:2127–30. [34] Boehm G, Moro G. Structural and functional aspects of prebiotics used in infant nutrition. J Nutr 2008;138:1818S–28S. [35] Lu L, Walker WA. Pathologic and physiologic interactions of bacteria with the gastrointestinal epithelium. Am J Clin Nutr 2001;73:1124S–30S. [36] Nash S, Stafford J, Madara JL. Effects of polymorphonuclear leukocyte transmigration on the barrier function of cultured intestinal epithelial monolayers. J Clin Invest 1987;80:1104–13. [37] Steinbakk M, Naess-Andresen CF, Lingaas E, Dale I, Brandtzaeg P, Fagerhol MK. Antimicrobial actions of calcium binding leucocyte L1 protein, calprotectin. Lancet 1990;336:763–5.