Original ArticlePlasma citrulline as a quantitative biomarker of HIV-associated villous atrophy in a tropical enteropathy population
Introduction
Tropical enteropathy (TE) is a population shift in intestinal mucosal morphology and function which resembles an asymptomatic villous atrophy of the small bowel. It is characterised by subclinical malabsorption and increased permeability, but its clinical significance is unknown. It is not even clear whether it is truly an abnormality or an adaptive response to environmental factors. It is most prevalent in the developing world within 30° of either side of the equator, but it is not limited to indigenous populations as it also affects European and American expatriates living in tropical regions. Mild reduction in villous height, broadening of the villi and increased crypt depth are the most common histological features described in TE.1 An increase in inflammatory cell infiltrate may occur, as well as mucosal T cell activation and crypt hyperplasia.2 Functional changes have been described, including subclinical malabsorption of carbohydrates,3 fat and vitamin B12,4 and increased mucosal permeability.5, 6
TE is observed in asymptomatic individuals, which distinguishes it from tropical sprue, a severe but now rare clinical illness. The histopathological changes of TE resemble those occurring in the early stages of coeliac disease and in other immune mediated enteropathies. It is likely that mucosal immune activation, perhaps in response to repeated intestinal infections, results in the observed microscopic changes described in TE. We have previously observed that Citrobacter rodentium may play such a role.7 In many tropical countries, the high prevalence of HIV leads to HIV enteropathy being superimposed on TE. On the whole the morphological and functional changes become significant in advanced AIDS.7 There are surprisingly few data on the responsiveness of the enteropathy to treatment of intestinal infection and to treatment with anti-retroviral drugs, and a non-invasive marker of intestinal mucosal health would be very useful in analysing the response of the mucosa to treatment.
A relationship between plasma citrulline and intestinal function has been proposed from studies examining rejection of small bowel allografts, in which plasma citrulline concentrations were found to be reduced in cases of mucosal damage, and with an inverse correlation to severity on biopsy.8, 9 Lower plasma citrulline concentrations have also been recorded in patients with villous atrophy than in healthy subjects or patients with anorexia nervosa. A range of thresholds was suggested: <10 μmol/L representing diffuse total villous atrophy; through 10–30 μmol/L for non-extensive or partial villous atrophy; to normal subjects who have levels >30 μmol/L10
One of the most exciting potential applications is the use of the circulating citrulline concentration as an objective tool for determining the need for and route of nutritional support.10, 11, 12 Evidence from human and animal studies suggests that citrulline may prove useful as a more general marker of small bowel epithelial damage regardless of cause.13, 14
There are few data on citrulline levels in humans. In our previous work we reported a quadratic (non-linear) correlation between fasting citrulline concentrations and remnant small bowel length, indicating that decreased citrulline concentrations predominantly occur when intestinal disease (or loss) is severe.15 This observation supported the earlier work from Paris and elsewhere.10, 13, 14. However, Luo et al.16 reported no relation between fasting plasma citrulline concentrations and intestinal absorptive capacity in short bowel syndrome, Peters et al.17 described poor diagnostic value of citrulline concentrations in intestinal energy absorption in enterocyte damage, and Miceli et al.18 were unable to confirm the value of plasma citrulline as a sensitive marker of intestinal atrophy. The position of citrulline as a marker of intestinal function cannot therefore be considered fully established.
In the current study we hypothesize that serum citrulline concentration might be a reliable marker of early morphological and functional changes in HIV enteropathy.
Section snippets
Patients
Endoscopic jejunal biopsy specimens and peripheral blood samples were collected from 150 adult patients with TE to investigate whether serum citrulline concentrations mirrored the degree of mucosal injury. TE was defined histologically from the presence of reduction in villous height, broadening of the villi and increased crypt depth in the absence of serological evidence for coeliac disease or documentable gastrointestinal infection TE.1 We included both HIV seropositive and HIV seronegative
Results
Our data set included 150 subjects (64 men and 86 women) affected by TE. HIV status was available for 145 subjects and only these individuals are considered further. Some 30.3% of the total sample were HIV seropositive (44 of 145; 21% in men:13 of 62; 37.3% in women: 31 of 83). One quarter (n = 11) of the HIV seropositive patients had a CD4 count below 200. Citrulline concentrations were very similar in all HIV seronegative subjects (Table 1). The median villus-height-to-crypt depth ratio was
Discussion
Citrulline contained in food is absorbed by the intestine. Its presence in the lumen, leads to citrulline accumulation in enterocytes, especially in the middle and lower ileum.21 A broad set of transporters seems to be able to transport citrulline from the lumen to the cell.22 Most of the circulating citrulline however comes from glutamine conversion in enterocytes.23 Citrulline is excreted by the kidneys after conversion to arginine.
In severe villous atrophy or in massive intestinal resection
What is current knowledge
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Circulating citrulline concentration correlates with reduced enterocyte mass independently of nutritional and inflammatory status.
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Citrulline concentration does not seem to be a sufficiently sensitive marker for rejection or viral enteritis as its values decline only in more severe intestinal mucosal damage.
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The time course of blood citrulline following radiation is in agreement with well known radiation effects on intestinal mucosa and clinical observations of acute intestinal injury.
What is new here
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The citrulline concentration appears to be a reliable marker of small bowel structural and functional integrity. It warrants attention as a possible surrogate for evaluation of HIV related enteropathy.
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Citrulline concentrations show a significant inverse correlation with the Schenk and Klipstein score in HIV positive patients
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Serum citrulline measurement is feasible and repeatable. It has the potential to be used as a marker of response to therapy in patients with HIV infection.
Statement of authorship
All authors made a substantial contribution to the conception and design of the study, and to the interpretation of the results. All were involved in the drafting and/or revision of the manuscript, and all gave their approval for the submitted manuscript.
Conflict of interest
None of the authors has any conflict of interest to declare.
Acknowledgements
We gratefully acknowledge Roy Sherwood from King’s College Hospital who devised the citrulline assay method and carried out all the measurements. Many thanks to Simone Bartolini and Winnie Dhaliwal for their kind help. This study was supported in part by the Italian Government Research Fund. The support of the UK NIHR via the Comprehensive Biomedical Research Centre at UCL/UCLH is also gratefully acknowledged.
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