Of the neonates with complicated MI, 4 of 6 had a volvolus and 4 of 6 had antenatal intestinal perforation. Five of six carried the ΔF508 mutation on 1 or more alleles (Table 4). A 7-year-old patient (patient 5) had undergone a further intestinal resection following an intestinal volvolus. In this individual, ileum was studied from the second resection and the initial (MI) resection. Only “healthy” tissues most distant from the perforation/volvolus were scored histologically.
Myofibroblastic transformation of fibroblasts (positivity for both smooth muscle actin and desmin and negativity for vimentin) was again prominent in the submucosa of CF tissues (5/6 patients) and uncommon in non-CF tissues (1/6) (Fig. 3C and D). CD45ro positivity (UCHL1 antibody) within the submucosa was a feature of CF tissues (6/6 CF tissues vs weakly in 1/6 non-CF atresias) (Table 6). The combined finding of submucosal myofibroblast proliferation and submucosal DC45ro positivity was highly predictive of CF. These findings are similar to those seen in DIOS tissues.
The features apparent in patient 5m at birth were still present at the age of 7 years when a second ileal resection was performed (Table 7). By this age, the transmural lymphocyte predominant imflammatory process apparent in individuals with DIOS and in 1 of 6 patients with MI at birth (patient 2m) was obviously apparent.
DIOS is probably a multifactorial syndrome, which is the end result of a number of processes to which children with CF are predisposed. Although there has been a reduction in incidence since the introduction of more efficacious microsphere enzyme replacement therapies (23), it seems unlikely that it is simply due to inspissation of incompletely digested and hydrated intestinal luminal contents. In keeping with this is the observation that it occurs in both pancreatic-sufficient and pancreatic-insufficient individuals (14,24). MI is positively associated with the ΔF508 genotype and negatively associated with G551D (25). Both ΔF508 and G551D have similar net deleterious effects on CFTR-mediated chloride currents and in this respect are severe phenotypes. This discordance for MI suggests that factors other than chloride currents are important in the genesis of MI. Individuals with MI are at increased risk of developing DIOS in later life. The ΔF508 mutation activates NFκB, a proinflammatory nuclear transcription factor, to a greater extent than G551D, raising the possibility that DIOS is an inflammatory dysmotility syndrome (8).
Prominent myofibroblastic transformation in the submucosa seen in individuals with both DIOS and MI supports an inflammatory origin of the fibrotic reaction that is seen more prominently in children with DIOS. Precise definition of an activated or fibrogenic phenotype of intestinal mesenchymal cell may be difficult. We have used dual positivity for desmin (D) and α-smooth muscle actin (SMA) together with negativity for vimentin (V) as a marker for myofibroblasts (MF) (30–32). In normal intestine MF and fibroblasts in submucosa, serosa, and intermuscular connective tissue are the primary sites of collagen synthesis (31,33). V+/SMA−/D+ fibroblasts and V−/SMA+/D+ myofibroblasts are the major sites of collagen type 1 synthesis in Crohn disease (31). The myofibroblastic transformation and proliferation of mesenchymal cells in the submucosa of patients with CF is in keeping with the contemporary view that some types of chronic intestinal inflammation are able to cause muscularis overgrowth and a change in enteric smooth muscle phenotype towards a collagen-producing myofibroblast phenotype (31,32). Procollagen genes can be induced by fibrogenic cytokines such as transforming growth factor-β (TGF-β) (32). Abnormalities in the innate immune response, which are a consistent characteristic of CF epithelia (34,35), may also provide an inherently profibrogenic influence (36,37). Interstitial cells of Cajal (ICC), a myofibroblast related intestinal cell subtype which is important in the regulation of gastrointestinal motor activity, are capable of dedifferentiation into a collagen-producing fibroblast or myofibroblast phenotype in chronic transmural intestinal inflammatory conditions (30,38). In addition to promoting fibrosis, the fibroblastic transformation of ICC would be expected to adversely affect gastrointestinal motility.
That fibrosis may be an intrinsic feature of CF rather than a secondary event is illustrated by the observation that ileal fibrosis can be present at birth in infants with severe CF genotypes (39). Colonic fibrosis is recognized in CF within the context of the so-called fibrosing colonopathy (40). Pathological descriptions of fibrosing colonopathy allude to the finding (in the colon) of severe fibrosis of the submucosa, muscularis propria, and lamina propria, which is out of proportion to the mild degree of mucosal inflammation (41). Ileal involvement was either inconspicuous or absent in earlier descriptions of fibrosing colonopathy (40,42–45), although a more contemporary publication alludes to ileal involvement with fibrosis of the mucosa, submucosa, and muscularis propria, and infiltration of the mucosa with eosinophils, although not with lymphocytes (41). Our ileal histopathological findings are different from those described in association with fibrosing colonopathy. Theories concerning the genesis of fibrosing colonopathy, in particular whether it is related to the formulation or dose of pancreatic exocrine supplement, have been the subject of substantial, sometimes acrimonious, debate (17,46–52). Although the numbers of case reports of fibrosing colonopathy (53) have fallen since the introduction of guidelines that total daily dose of lipase should equate to <10,000 IU lipase per kilogram of body weight per day (54), our data and that of others (55) support the notion that intestinal fibrosis in CF need not necessarily be iatrogenic. The thickened bowel visible on abdominal ultrasound in individuals with CF (56,57) is also independent of both dose and formulation of enzyme replacement administered (58) and seems likely to represent a further manifestation of a chronic transmural inflammatory process. The observation of echogenic bowel in infants with CF in utero (57,59) and our findings of myofibroblastic proliferation in infants with CF at birth support the notion that this is an inherent feature of CF.
These observations support the hypothesis that intestinal fibrosis in CF is unrelated to PERT. They also provide an as yet unproven rationale for anti-inflammatory treatment(s) in individuals with early-onset CF-associated motility disturbances and anti-fibrotic therapies to alter the natural history of DIOS. We believe that our observations make a strong case for routine and early ileal mucosal and laparoscopic seromuscular ileal biopsy (60) in individuals with severe and recurrent DIOS. Specific anti-inflammatory treatments and/or antifibrotic therapies may become justifiable when further data become available about both the prevalence of these changes and also the efficacy of early anti-inflammatory therapies. This approach seems more appropriate than what is essentially symptomatic treatment with bowel cleansing agents, mucolytics, and increased doses of pancreatic enzymes (21,24,61,62). It seems likely that the potential association of intestinal colonic fibrosis with dose, duration, and brand of pancreatic enzyme administered has detracted the physician's focus of attention from an underlying gastrointestinal inflammatory process that is inherent to CF (17).
Research at the Institute of Child Health and Great Ormond Street Hospital for Children NHS Trust benefits from R&D funding received from the NHS Executive.
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