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How to use faecal calprotectin in management of paediatric inflammatory bowel disease
  1. Amit Saha1,
  2. Mark P Tighe2,
  3. Akshay Batra1
  1. 1Paediatric Gastroenterology, Southampton University Hospital NHS Trust, Southampton, UK
  2. 2Department of Paediatrics, Poole Hospital NHS Trust, Dorset, UK
  1. Correspondence to Dr Akshay Batra, Paediatric Gastroenterology, Southampton University Hospital NHS Trust, MP 44, G level East Wing, Tremona Road, Southampton, SO16 6YD, UK; Akshay.Batra{at}uhs.nhs.uk

Abstract

Faecal calprotectin (FC) is a neutrophil-derived protein released in stool in response to mucosal inflammation. It is a simple, cheap and non-invasive test with high sensitivity and moderate specificity, which can be useful in the diagnosis and monitoring of inflammatory bowel disease (IBD). FC levels correlate well with bowel inflammation (both macroscopic and histological activity) and are not influenced by disease location or type of IBD. Despite the shortcoming with regards to specificity, it is the high sensitivity of FC that makes it a valuable screening tool in the diagnosis of IBD. It is especially effective in identifying children with low probability of IBD who would not benefit from further investigations. The cut-off value selected has a significant impact on the diagnostic accuracy of the test, influencing its sensitivity and specificity, and must be interpreted judiciously. Its role in disease monitoring is as an add-on test to Paediatric Ulcerative Colitis Activity Index and Paediatric Crohn's Disease Activity Index scores and can be used to differentiate disease relapse from functional symptoms. High levels of FC are also seen in a number of other conditions, such as gastrointestinal infections and coeliac disease. It is recommended that infective causes affecting the gut must be excluded first, before FC is measured.

  • Paediatric
  • IBD
  • Inflammatory Bowel disease
  • Calprotectin
  • diagnosis

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Introduction

The incidence of inflammatory bowel disease (IBD) in children less than 16 years of age in UK has been increasing and has gone up from 5.2/100 000/year in 2001 to 9.37/100 000/year over a period of 10 years.1 Crohn's disease can present with symptoms of abdominal pain (72%), weight loss (58%), anorexia (25%) and lethargy (27%) and children presenting with ulcerative colitis have a higher prevalence of diarrhoea (74%), bleeding (84%) and abdominal pain (62%),2 although there may be considerable clinical overlap. It is noteworthy that a significant proportion of children present with a range of non-specific symptoms such as lethargy and mild abdominal discomfort, with only about 25% presenting with the ‘classic triad’ of diarrhoea, pain, and weight loss.2 On the other hand, non-specific abdominal pain remains a common problem of childhood and represents a significant burden on primary and secondary care. It is therefore important to identify children with functional abdominal pain without using invasive investigations especially in the absence of ‘red flag’ symptoms; in whom investigations identify disease in <5% of children.3 In current clinical practice, a large number of endoscopies performed for suspected paediatric inflammatory bowel disease (PIBD) are negative, and there is an acute need to prioritise children who need it most.4

Early investigation in suspected cases of PIBD is imperative, as a delay in diagnosis can prolong the period of time patients suffer from symptoms. One study of 739 new paediatric IBD cases reported a median delay of 5 months (mean 11 months) from the onset of symptoms to the confirmation of diagnosis, with one-fifth of them having symptoms for more than a year.2 Sixty per cent of IBD cases were reported to have had a delay of less than 6 months, 19% a delay of 6–12 months, 14% a delay of 1–3 years, and 7% a delay of more than 3 years. Improved recognition of presenting features and judicious use of the screening, investigative and diagnostic tests may aid speedy diagnosis, prompt initiation of treatment and reduce the impact of disease.

Being a relapsing–remitting illness, patients with IBD are at risk of relapse throughout the course of their disease. This risk is ill-defined and ill-understood, due to the waxing and waning nature of the disease, with no treatment being 100% effective in maintaining remission. The Paediatric Ulcerative Colitis Activity Index (PUCAI) and Paediatric Crohn's Disease Activity Index (PCDAI) scores are useful at high levels in identifying overt relapse, but at lower levels, are unable to differentiate between functional symptoms and mild relapse.5 Endoscopy is the gold standard for assessing recurrence of inflammation and relapse but its’ use is limited because of the invasive nature of the test and the common requirement in children for a general anaesthetic.

Faecal calprotectin (FC) is a simple, cheap and non-invasive test with high sensitivity, which can be useful in identifying such cases. It has been shown to be diagnostically more accurate than commonly used blood parameters such as C-reactive protein, erythrocyte sedimentation rate, haemoglobin, platelet count, total white cell count and albumin.6 In its diagnostic guidance (DG11) published in 2013, the National Institute for Health and Care Excellence (NICE) recommends FC testing to support clinicians with the differential diagnosis of IBD or non-IBD in children. This review looks at the role of measurement of FC in diagnosing and monitoring of paediatric IBD.

Physiological background

Calprotectin, a neutrophil-derived protein present in stool, is a 36 kDa calcium- and zinc-binding protein found in a number of tissues and released during inflammation, both acute and chronic.7 Calprotectin is produced by intestinal neutrophils but is not specific to gastrointestinal mucosa; it possesses chemotactic and antimicrobial properties (anticandidal) mediated by zinc chelation via its histidine-rich regions8 and is excreted in excess into the intestinal lumen during an inflammatory process of the gut. It accounts for up to 60% of the total protein content of neutrophil cytosol, and therefore can be directly proportional to neutrophil influx into the gut mucosa.9

In the early phase of inflammation, unknown triggers on the gut epithelium lead to the activation of the intestinal immune system, resulting in the influx of monocytes, macrophages and granulocytes. These cells actively secrete inflammatory mediators or release granule proteins by cell degranulation. The neutrophil-derived S100 proteins (S100A8/A9 (calprotectin)) are released from the cytosol by activated or damaged cells under conditions of cell stress. Cytokines such as IL-1 or TNF-α (secreted by macrophages in response to inflammatory trigger) and/or bacterial products also cause monocytes and intestinal epithelial cells to synthesise and secrete proteins of the S100 family. Studies have even implicated high local concentrations of calprotectin as injurious to inflamed cells, inducing apoptosis, especially in zinc-deficient conditions.10 FC then easily passes into the intestinal lumen due to the inflamed and hyperpermeable gut mucosa and is deposited onto the faecal stream (figure 1).

Figure 1

Faecal calprotectin in mucosal inflammation of gastrointestinal tract. (1) Unknown trigger causing activation of gut immune system generating faecal calprotectin from (A) epithelial cells (B) monocytes and (C) neutrophils. The released faecal calprotectin passes into the gut lumen and is taken up by the faecal stream.

Technological background

FC can be measured in stool by ELISA methods and requires less than 5 g of stool for reliable assay. FC is resistant to enzymatic degradation in stool and is stable for up to 7 days at room temperature.11 This makes it possible for a sample to be collected and transported to the laboratory in a plain container.

There are several available technologies to measure the level of calprotectin in stool sample, including quantitative laboratory-based tests, fully quantitative rapid tests and semiquantitative point-of-care tests (POCTs).12 The fully quantitative laboratory-based technologies available to the National Health Service in England mainly use ELISA to detect and quantify calprotectin in stool. Several manufacturers such as Bühlmann, Calpro, Eurospital, Immunodiagnostik and Phadia AB provide an array of quantitative ELISA and rapid tests with different ranges. Preventis also manufactures immunochromatographic rapid tests used as a POCT, both a semiquantitative one using three set ranges and a qualitative test using a cut-off value of 50 µg/g of stool.12

Clinical indications in PIBD

In children with abdominal pain and diarrhoea, can a low FC accurately identify children who do not need further investigations for IBD?

FC can be measured in children presenting with gastrointestinal symptoms suggestive of PIBD and is useful in distinguishing IBD from functional gastrointestinal disorders. It has been shown to correlate well with macroscopic and histological activity.13 In children 4 years and older, FC levels are not influenced by sex or age of the child and are equally reliable for Crohn's disease as well as ulcerative colitis as they are not affected by disease location.6

Children with new-onset PIBD have a significantly elevated FC at the time of diagnosis, compared with controls undergoing endoscopy, as shown by multiple studies.5 ,6 ,13 The cut-off values selected have a significant impact on the diagnostic accuracy of the test, influencing its sensitivity and specificity. A lower cut-off value increases sensitivity, and as higher cut-off values are selected sensitivity reduces and specificity increases (table 1). With a cut-off of 50 µg/g, the test has a sensitivity of 98%, with negative predictive value of 96%.

Table 1

Measures of diagnostic accuracy for increasing levels of faecal calprotectin in children with suspected inflammatory bowel disease

A recent systematic review by Henderson et al14 included eight studies in the paediatric population with a total of 715 patients, including 394 PIBD and 321 non-PIBD controls. This showed a pooled sensitivity of 0.978 (95% CI 0.947 to 0.996) and specificity of 0.682 (95% CI 0.502 to 0.863) for the diagnostic utility of FC in PIBD.14

In a diagnostic meta-analysis published in 2010, Rheenen et al showed that in children with suspected PIBD, an elevated FC level would increase the pre-test probability of having IBD from 61% to 86%; but more importantly, a normal FC level reduces it from 61% to 15%. The same study demonstrated that FC screening in children with suspected PIBD would reduce the number requiring endoscopy by 35%.13

In children with low clinical suspicion of IBD and normal serological markers, a FC <50 µg/g makes the diagnosis of IBD unlikely and would not warrant any further investigations. With judicious interpretation, there is growing evidence that FC is an asset in secondary care in determining which children do not need referral to tertiary care with suspected PIBD; and in tertiary care in identifying children who do not require endoscopy for suspected PIBD. The utility of testing FC must be balanced against patients with PIBD who have a false negative FC test and adding FC in the diagnostic pathway would result in up to 8% of children having a delayed diagnosis.13

In children presenting with signs and symptoms consistent with IBD, does an elevated FC confirm diagnosis?

Raised FC levels are associated with inflammation of the mucosal lining and are not very specific for IBD. The specificity increases with increasing value and levels >800 µg/g are 95% specific for IBD (table 1). Raised FC is unable to differentiate between various causes of gut inflammation and its use is limited to differentiating inflammatory and functional causes of symptoms. Hence other causes, such as infections and coeliac disease, must be ruled out before further investigations are performed. In children who present with classic symptoms of IBD with a high index of clinical suspicion, FC does not add more to the investigations as all these children need an endoscopy to confirm the diagnosis.15 It also has limited role in children presenting with rectal bleeding as most cases causing bleeding, such as infections, rectal polyps and solitary rectal ulcer syndrome, would lead to an elevated FC.

In children with IBD, is there a role of FC measurements in monitoring of disease activity?

Children with quiescent disease present with FC concentrations that are higher than the traditionally accepted normal reference range16 and therefore there is no defined role of measuring FC levels routinely. There is some evidence that FC has some utility as a surrogate marker for mucosal healing in IBD and it can be used in assessing the degree of inflammation and gives an indication of the efficacy of treatment.17 Studies have shown that measured FC levels correlate well with endoscopic severity at colonoscopy15 and are useful in identifying disease relapse.18 In one study published in 2008, patients with relapse showed significantly elevated FC levels compared with non-relapsed ones, with a FC value of 275 µg/g achieving a sensitivity and negative predictive value of 97%, and specificity and positive predictive value of 85% in predicting histological relapse.19 The specificity of FC in identifying relapse can be further increased by using it as an add-on test to PUCAI and PCDAI scores (with a cut-off at 500 µg/g).20

FC, when used in conjunction with clinical and serological markers,5 is useful in differentiating mild relapse from functional symptoms and correlates well with mucosal inflammation but there is currently no robust data to support regular measurement of FC as part of monitoring of PIBD.

Limitations

Although the utility of FC has now been well established in the diagnosis of PIBD, it is, however, not a specific test for IBD; especially in children who present with a broad spectrum of symptoms. It also has no role in identifying the site or type if disease. High levels of FC in children are also seen in a number of other conditions, notably gastrointestinal infections, coeliac disease, immunodeficiency and non-steroidal anti-inflammatory drug induced enteropathy.9 Indeed it is recommended that infective causes affecting the gut must be excluded first, before FC is considered, in order to avoid false positive results that might prompt unnecessary endoscopies.

Future research

Although FC testing has shown promising results for children with suspected PIBD in secondary care, further research is needed to establish the place of FC testing within robust, evidence-based clinical management pathways. This includes further research into the optimal cut-off values, interpretation and guidance on ‘intermediate range’ results and its role in monitoring disease activity and predicting relapse. Finally, further prospective studies are needed to fully determine the effect of FC measurement and its interpretation in the reduction of endoscopy rates.

Test your knowledge

  1. Faecal calprotectin (FC) level is useful in differentiating inflammatory bowel disease (IBD) from functional gastrointestinal disorders if the value is

    1. <50 µg/g

    2. 50–100 µg/g

    3. 100–200 µg/g

    4. >200 µg/g

  2. Faecal calprotectin levels are influenced by

    1. Age

    2. Gender

    3. Disease location

    4. All of the above

    5. None of the above

  3. FC levels should be checked routinely for children being followed up for IBD

    1. True

    2. False

  4. FC levels are likely to be normal in which of the following condition

    1. Coeliac disease

    2. Clostridium difficile colitis

    3. Small bowel Crohn's disease

    4. Ulcerative colitis

    5. None of the above

  5. The main role of FC measurement is

    1. To rule out IBD in children with low clinical suspicion

    2. To confirm IBD in children with high clinical suspicion

    Answers to the quiz are at the end of the references.

Answers to the multiple choice questions

  • (1) A; (2) E; (3) B; (4) E; (5) A.

Clinical bottom line

  • High levels of faecal calprotectin (FC) are also seen in a number of other conditions associated with mucosal inflammation, such as gastrointestinal infections and coeliac disease. Indeed, it is recommended that infective causes affecting the gut must be excluded first, before FC is measured.

  • FC correlates well with bowel inflammation (both macroscopic and histological activity) and is not influenced by sex, age, irritable bowel disease (IBD) type or disease location.

  • The cut-off values selected has a significant impact on the diagnostic accuracy of the test, influencing its sensitivity and specificity, and must be interpreted judiciously by an experienced clinician.

  • The high negative predictive value of a normal FC can be utilized in risk stratification and screening of children with suspected IBD, that is, children with low clinical suspicion and levels <50 µg/g do not need further investigations.

  • FC screening is particularly cost-effective when baseline clinical suspicion for IBD is moderate-to-low. Conversely, direct endoscopic evaluation is more cost-effective when the pre-test probability of true IBD in children is high, and there is little value in measuring FC in these patients.

  • FC is most effective when used as an add-on test to Paediatric Ulcerative Colitis Activity Index and Paediatric Crohn's Disease Activity Index scores in differentiating disease relapse from functional symptoms.

References

Footnotes

  • Contributors The literature search was done by AS and AB. The layout of the article was done by AB and was written by AS, AB and MPT.

  • Competing interests None declared.

  • Provenance and peer review Commissioned; externally peer reviewed.

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