Christine Breynaert

Results of the 4th scientific workshop of the ECCO (I): Pathophysiology of intestinal fibrosis in IBD

The fourth scientific workshop of the European Crohns and Colitis Organization (ECCO) focused on the relevance of intestinal fibrosis in the disease course of inflammatory bowel disease (IBD). The objective was to better understand the pathophysiological mechanisms of intestinal fibrosis, to identify useful markers and imaging modalities of fibrosis in order to assess its presence and progression, and, finally, to point out possible approaches for the prevention and the treatment of fibrosis. The results of this workshop are presented in three separate manuscripts. This first section describes the most important mechanisms that contribute to the initiation and progression of intestinal fibrosis in IBD including the cellular and molecular mediators, the extracellular matrix molecules and matrix metalloproteinases/tissue inhibitors of metalloproteinases-system, the microbiota products, the role of fat, genetic and epigenetic factors, as well as the currently available experimental models. Furthermore, it identifies unanswered questions in the field of intestinal fibrosis and provides a framework for future research.

Characteristics of Skin Lesions Associated With Anti–Tumor Necrosis Factor Therapy in Patients With Inflammatory Bowel Disease: A Cohort Study

Context Tumor necrosis factor (TNF) antagonists are effective for the treatment of inflammatory bowel disease (IBD) that does not respond to conventional therapy. Some patients, however, have discontinued anti-TNF therapy because of the development of psoriasiform lesions. Contribution In this large study from a tertiary care center, nearly one third of patients with IBD who initiated anti-TNF therapy developed skin lesions. With dedicated dermatologic care, most were able to continue therapy without interruption. Caution Patients with IBD not receiving anti-TNF therapy were not studied. Implication Skin lesions seem to be commonly associated with anti-TNF therapy in patients with IBD but typically do not require treatment discontinuation for successful management. In the late 1990s, tumor necrosis factor (TNF) antagonists were introduced as treatment for several inflammatory disorders, including rheumatoid arthritis, ankylosing spondylitis, psoriasis, Crohn disease, and ulcerative colitis. Whereas conventional therapy primarily targets symptom control, anti-TNF therapy has been shown to reduce symptoms, heal mucosal ulcers, reduce hospitalizations and surgeries, and spare corticosteroids (110). Infliximab, the first biological agent targeted against TNF, was followed by other anti-TNF biologics, including adalimumab, golimumab, certolizumab pegol, and etanercept (which is not used for inflammatory bowel disease [IBD]). Anti-TNF therapy has also proved to have a favorable safety profile with respect to serious infections, cancer, and mortality as long as it is given for the right indication and comorbidities are taken into account (1120). However, psoriasiform skin manifestations have been reported (long after the completion of randomized, controlled trials) as adverse events associated with anti-TNF therapy (2130). The appearance of psoriasiform skin manifestations is intriguing and seems paradoxical because anti-TNF therapy is also successfully used in psoriasis treatment. Inflammatory bowel disease shares several pathophysiologic characteristics with psoriasis and atopic dermatitis. All of these conditions have a strong genetic component, with 163 loci identified for IBD, 36 identified for psoriasis, and 11 identified for atopic dermatitis thus far (3133). Of note, the reported regions of association overlap substantially (32, 34, 35). Because skin lesions may represent a significant burden to patients, it is important to advise patients on preventive measures or alternative therapeutic options, especially when they are at increased risk. This study aimed to accurately describe patients with IBD treated with anti-TNF therapy who did and did not develop skin lesions. We investigated the development of skin lesions in a retrospective cohort of such patients. We also looked at specific patient characteristics, autoimmune and genetic factors, and infliximab cumulative dose and trough levels (TLs) in patients with and without skin lesions. Methods Study Participants and Design We retrospectively analyzed a consecutive cohort of 917 patients with IBD for the occurrence of skin lesions during anti-TNF therapy. Diagnosis of IBD was well-established according to conventional endoscopic, radiologic, and histologic criteria (36, 37). A total of 955 patients initiated infliximab treatment at the University Hospitals Leuven between December 1994 and January 2009, of whom 917 agreed to participate in our research projects. At the time of initiation of infliximab treatment, they were naive to any other anti-TNF agent. A subset of patients were noted to have switched to another anti-TNF agent (adalimumab or certolizumab pegol) when they showed loss of response to infliximab. The Ethics Board of the University Hospitals Leuven approved the study before data collection (institutional review board approval number B322201213950/S53684). All participants provided written informed consent agreeing to take part in our current and future research projects when they were included in our ongoing patient registry. Diagnostic and Therapeutic Criteria for Patients With Skin Lesions At the Leuven IBD center, all patients were seen by the IBD gastroenterologists (S.V., G.V.A., P.R., and M.F.) every 8 weeks or more frequently if needed. Patients with skin symptoms (especially itching) or skin lesions that occurred during and were possibly linked to anti-TNF therapy were referred to a dermatologist (S.S.) experienced in anti-TNF therapies. Beginning in 2000, when the phenomenon of skin lesions associated with anti-TNF treatment became more clear, patients who came to consultation were systematically checked for lesions and referred to a dermatologist. We classified the patients in the following categories (listed in descending order of severity) according to the predominant or most distressing lesions (if 1 type of lesion was identified, the patient was placed in the most-severe category): palmoplantar pustulosis (pustules of the palms and soles with or without surrounding erythema or erythematosquamous lesions), psoriasis (sharply demarcated erythematosquamous lesion covered with silvery-white scales; histologic evaluation [when performed] showed psoriasis), psoriasiform eczema (dermatitis with some features of psoriasis, especially the orange-red color, but unsharply defined lesions, often superinfected; histologic evaluation [when performed] showed dermatitis), eczema (dermatitis without features of psoriasis), or xerosis (dryness of the skin without signs of inflammation). In cases of diagnostic uncertainty (especially psoriasis vs. psoriasiform eczema) (Appendix Figure 1), skin biopsy specimens were taken and histologic evaluation was done. Skin lesions that could not be classified into one of these categories, including infections, inflammatory skin disease, tumors, and alopecia areata, were classified as other. A full list of skin lesion types is provided in Appendix Table 1. Appendix Figure 1. Hematoxylin and eosin staining of psoriasis (top), psoriasiform eczema (middle), and eczema (bottom). All histologic pictures are original magnification50. Top. A skin lesion showing irregular acanthosis, focal parakeratosis, and crust/scale formation, resembling psoriasiform eczema. Middle. A skin lesion showing regular elongation and clubbing of rete ridges, thinning of suprapapillary epidermis, confluent parakeratosis, and superficial perivascular inflammation, resembling psoriasis. Bottom. A skin lesion showing irregular and plump acanthosis, multifocal parakeratosis and spongiosis, and superficial perivascular inflammation, resembling spongiotic eczema. Appendix Table 1. Types of Skin Lesions For dermatologic treatment of the most frequently occurring skin lesions, we used a standard protocol, which is provided in the Appendix. Patient Characteristics The medical records of the patients were systematically reviewed up to 1 July 2014. For each patient, sex, diagnosis, smoking (ever vs. never), age at diagnosis and at the start of infliximab treatment, start and stop dates of infliximab treatment, infliximab infusion dates, switch to other anti-TNF agent (adalimumab or certolizumab pegol) with respective start and stop dates, reason for discontinuing anti-TNF therapy, and cumulative infliximab dose (until development of skin lesion, discontinuation of therapy, or end of analysis) were retrieved. To correct for different durations of exposure to infliximab, the cumulative infliximab dose was divided by the duration of treatment. When the interval between infliximab infusions was more than 16 weeks (as happened in the first patients treated in the 1990s), the time between infusions was not included in the calculations of total duration of treatment. Loss of response to infliximab was assessed and was defined as the need to stop infliximab therapy or the need for surgery despite an attempt to optimize treatment. If a skin lesion was reported, date of onset, type, location, history of atopic dermatitis or psoriasis, and initiated treatment were obtained. Case patients who stopped anti-TNF treatment because of a lesion were also noted. For all patients with skin lesions, we noted at their last consultation whether they showed complete, partial, or no resolution of the lesions and whether they were still receiving treatment for them at that time. For patients in whom anti-TNF treatment was stopped because of lesions, we noted how the lesions subsequently evolved and, in cases of complete resolution, in what time frame. Measurement of Serum Trough Levels of Infliximab Infliximab TLs and anti-infliximab antibodies (ATIs) were measured at several time points during treatment by using enzyme-linked immunosorbent assays developed in-house (38, 39). We considered only samples from week 14 onward in patients receiving infliximab maintenance therapy (defined as receipt of 4 infliximab infusions and 6 months of therapy). A total of 8350 TL measurements in 433 patients were available (median, 18 [interquartile range {IQR}, 5 to 32] per patient). A patient was considered to have subtherapeutic TLs when at least 2 measurements were less than 3 g/mL and supratherapeutic TLs when at least 2 measurements were greater than 7 g/mL (40). A total of 2770 ATI measurements in 325 patients were available (median, 5 [IQR, 3 to 13] per patient). Anti-infliximab antibodies were evaluated only when TLs were not detectable. Patients were considered ATI-positive when they had at least 1 ATI concentration greater than 1 g/mL. Measurement of Antinuclear Antibodies and Double-Stranded DNA Antibodies Measurements at baseline (3 months before the first infliximab dose) and at several time points during follow-up were included. Antinuclear antibodies (ANAs) were determined on 1:40 diluted sera by an indirect immunofluorescent technique on Sjgren syndrome Atransfected HEp-2 cells. A total of 3820 ANA measurements in 847 patients were available (median, 3 [IQR, 1 to 5] per patien

Cellular and Molecular Mediators of Intestinal Fibrosis

Abstract Intestinal fibrosis is a major complication of the inflammatory bowel diseases (IBD) and although inflammation is necessary for its development, it would appear that it plays a minor role in its progression as anti-inflammatory treatments in IBD do not prevent fibrosis once it has started. The processes that regulate fibrosis would thus appear to be distinct from those regulating inflammation and, therefore, a detailed understanding of these pathways is vital to the development of anti-fibrogenic strategies. There have been several recent reviews exploring what is known, and what remains unknown, about the development of intestinal fibrosis. This review is designed to add to this literature but with a focus on the cellular components that are involved in the development of fibrogenesis and the major molecular mediators that impact on these cells. The aim is to heighten the understanding of the factors involved in intestinal fibrogenesis so that detailed research can be encouraged in order to advance the processes that could lead to effective treatments.

Tolerability of Shortened Infliximab Infusion Times in Patients With Inflammatory Bowel Diseases : A Single-Center Cohort Study

OBJECTIVES:Scheduled maintenance therapy with infliximab decreases the risk of infusion reactions. Many centers have accelerated infusion times to 1 h in selected patients who tolerate 5 mg/kg infliximab infusions. The aim of this study was to compare the tolerability of 1-h and 2-h infliximab infusions in patients with inflammatory bowel disease (IBD) in a large single-center cohort. The primary end point was the incidence of infusion reactions in both groups.METHODS:A retrospective chart analysis of all IBD patients treated with infliximab was performed. Infusions in scheduled maintenance for at least 6 months from December 1994 until March 2009 were included. All patients were treated at the infusion unit or during hospitalization under standard operating procedures. Infusion parameters were prospectively recorded. From 2004, in patients tolerating at least four 2-h infusions, infusions were given over 1 h.RESULTS:As of March 2009, 953 patients with IBD (77.6% Crohns disease, 22.4% ulcerative colitis) had been treated with infliximab. A total of 474 patients met the criteria of scheduled maintenance therapy. In total, 9,155 maintenance infusions were administered (4,307 over 1 h). No severe infusion reactions were documented. Mild acute reactions occurred in 0.6% (27/4,307) of the 1-h-infusion group and in 1.7% (80/4848) of the 2-h infusion group (P=0.0034). Delayed infusion reactions occurred in 0.2% of 1-h and 0.5% of 2-h infusion group patients (P=0.277). Loss of tolerability due to infusion reactions (1-h group 2.9% versus 2-h group 4.1%) was evenly distributed (P=0.34). None of the prespecified variables were predictive of infusion reactions in a multivariate analysis.CONCLUSIONS:In patients with IBD tolerating 2-h infusions of infliximab scheduled maintenance therapy, the infusion time can be shortened to 1 h with good tolerability. No severe reactions were observed and no predictors of infusion reactions were identified.

Infliximab restores the dysfunctional matrix remodeling protein and growth factor gene expression in patients with inflammatory bowel disease.

Background:Matrix metalloproteinases (MMPs), tissue inhibitors of metalloproteinases (TIMPs), a disintegrin and metalloprotease with thrombospondin motifs [ADAM(TS)s] and growth factors are involved in inflammation and tissue damage and repair, all occurring in inflammatory bowel disease (IBD). We studied the impact of anti-inflammatory therapy with infliximab on mucosal expression of these tissue remodeling genes in patients with IBD. Methods:Mucosal gene expression of 23 MMPs, 4 TIMPs, 50 ADAM(TS)s, and 158 growth factors was investigated in 61 patients with IBD before and after the first infliximab therapy and in 12 controls, with microarrays and quantitative RT-PCR. Protein localization, mucosal gelatinase levels, and net gelatinolytic activity were investigated by immunohistochemistry, zymography analysis, and gelatin degradation assay, respectively. Results:In patients with active IBD before infliximab versus controls, gene expression of many MMPs, TIMPs, ADAM(TS)s, and growth factors was upregulated, whereas colonic expression of MMP28 and TGFA and ileal expression of ADAMDEC1 and AGT were downregulated. After controlling inflammation with infliximab, most gene dysregulations observed at baseline were restored in responders. Increased ratio of MMP1/TIMP1 expression at baseline in active IBD was restored in responders with colonic mucosal healing. With immunohistochemistry, protein localization differences of MMP1, MMP3, REG1A, and TIMP1 were shown between active IBD and control mucosa. With zymography analysis and gelatin degradation assay, higher gelatinase levels and net gelatinolytic activity were measured before infliximab and levels normalized after infliximab. Conclusions:Our data suggest that suppression of inflammation results in the arrest of epithelial damage and subsequent mucosal healing. Therefore, the therapeutic potential of agents targeting MMPs or growth factors as primary therapy seems rather complex.

Unique gene expression and MR T2 relaxometry patterns define chronic murine dextran sodium sulphate colitis as a model for connective tissue changes in human Crohn's disease.

Introduction Chronically relapsing inflammation, tissue remodeling and fibrosis are hallmarks of inflammatory bowel diseases. The aim of this study was to investigate changes in connective tissue in a chronic murine model resulting from repeated cycles of dextran sodium sulphate (DSS) ingestion, to mimic the relapsing nature of the human disease. Materials and Methods C57BL/6 mice were exposed to DSS in drinking water for 1 week, followed by a recovery phase of 2 weeks. This cycle of exposure was repeated for up to 3 times (9 weeks in total). Colonic inflammation, fibrosis, extracellular matrix proteins and colonic gene expression were studied. In vivo MRI T 2 relaxometry was studied as a potential non-invasive imaging tool to evaluate bowel wall inflammation and fibrosis. Results Repeated cycles of DSS resulted in a relapsing and remitting disease course, which induced a chronic segmental, transmural colitis after 2 and 3 cycles of DSS with clear induction of fibrosis and remodeling of the muscular layer. Tenascin expression mirrored its expression in Crohn’s colitis. Microarray data identified a gene expression profile different in chronic colitis from that in acute colitis. Additional recovery was associated with upregulation of unique genes, in particular keratins, pointing to activation of molecular pathways for healing and repair. In vivo MRI T2 relaxometry of the colon showed a clear shift towards higher T2 values in the acute stage and a gradual regression of T2 values with increasing cycles of DSS. Conclusions Repeated cycles of DSS exposure induce fibrosis and connective tissue changes with typical features, as occurring in Crohn’s disease. Colonic gene expression analysis revealed unique expression profiles in chronic colitis compared to acute colitis and after additional recovery, pointing to potential new targets to intervene with the induction of fibrosis. In vivo T2 relaxometry is a promising non-invasive assessment of inflammation and fibrosis.

Inhibition of gelatinase B/MMP-9 does not attenuate colitis in murine models of inflammatory bowel disease

One third of patients with inflammatory bowel disease (IBD) inadequately respond to anti-TNF treatment and preclinical data suggest that matrix metalloproteinase-9 (MMP-9) is a novel therapeutic target. Here we show that IBD clinical and histopathological parameters found in wild type mice challenged with three different models of colitis, acute and chronic dextran sodium sulphate (DSS), and acute 2,4,6-trinitrobenzenesulfonic acid-induced colitis are not attenuated in MMP-9 knockout mice. We find similar colonic gene expression profiles in wild type and MMP-9 knockout mice in control and acute DSS conditions with the exception of eleven genes involved in antimicrobial response during colitis. Parameters of chronic colitis are similar in wild type and MMP-9 knockout mice. Pharmacological inhibition of MMP-9 with bio-active peptides does not improve DSS colitis. We suggest that MMP-9 upregulation is a consequence rather than a cause of intestinal inflammation and we question whether MMP-9 represents a disease target in IBD.

Incidence of acute severe infusion reactions to infliximab depends on definition used rather than assay

477 days, whereas 35% remained in sustained clinical remission. According to Hungarian regulations, biological therapy has to be discontinued after 1 year of treatment if the patient is in remission. Our aims were to assess the outcome of CD after the discontinuation of biologicals and to determine which factors might predict relapse. The data of 31 CD patients (17 women, 14 men; mean age at the diagnosis 31 years) receiving biological therapy (16 infliximab, 15 adalimumab) were assessed. The patients’ characteristics are in Table 1. Steroids were discontinued after an average of 13.7 weeks. Forty four per cent of the patients stopped immunosuppressants during biological therapy. Dose intensification was needed in four patients after an average of 7.5 months. The mean length of follow-up from the last infusion was 6 months. Biological therapy was restarted in eight patients (26%) within 4.4 months after the discontinuation and 63% of them achieved clinical remission again at the twelfth week. Two patients developed side effects during the second treatment period. No association was found between relapse and a range of variables: age at the diagnosis or the time of biological therapy, smoking, disease location, behaviour, perianal and extraintestinal manifestations, use of steroids or immunosuppressants, surgery, previous biological therapy, Best index or C-reactive protein concentration. Biological therapy had to be restarted within a short period in every fourth of our patients who were in remission after 1 year of anti-TNF-a treatment. Neither Waugh et al. nor our group could identify predictive factors for relapse and for the need for resumed treatment. Our national multicentre RASH (Relapse After Stopping biologicals in Hungary) study on the prognostic factors for the need to resume biological treatment is in progress.

Interleukin‐15 receptor α expression in inflammatory bowel disease patients before and after normalization of inflammation with infliximab

Interleukin‐15 (IL‐15) is a pro‐inflammatory cytokine thought to contribute to the inflammation in inflammatory bowel diseases (IBD). The specific receptor chain IL‐15Rα can be expressed as a transmembranous signalling receptor, or can be cleaved by a disintegrin and metalloprotease domain 17 (ADAM17) into a neutralizing, soluble receptor (sIL‐15Rα). The aim of this study is to evaluate the expression of IL‐15Rα in ulcerative colitis (UC) and Crohns disease (CD) patients before and after infliximab (IFX) therapy. Gene expression of IL‐15Rα, IL‐15 and ADAM17 was measured at the mRNA level by quantitative reverse transcription‐PCR in mucosal biopsies harvested before and after first IFX therapy. Concentrations of sIL‐15Rα were measured in sera of patients by ELISA and IL‐15Rα protein was localized in the gut by immunohistochemistry and immunofluorescence. Mucosal expression of IL‐15Rα is increased in UC and CD patients compared with controls and it remains elevated after IFX therapy in both responder and non‐responder patients. The concentration of sIL‐15Rα in serum is also increased in UC patients when compared with controls and does not differ between responders and non‐responders either before or after IFX. CD patients have levels of sIL‐15Rα comparable to healthy controls before and after therapy. In mucosal tissues, IL‐15Rα+ cells closely resemble activated memory B cells with a pre‐plasmablastic phenotype. To conclude, IBD patients have an increased expression of IL‐15Rα mRNA in the mucosa. Expression is localized in B cells, suggesting that IL‐15 regulates B‐cell functions during bowel inflammation. No change in release of sIL‐15Rα is observed in patients treated with IFX.

An abdominal pain syndrome in a lupus patient

Systemic lupus erythematosus can be complicated by the antiphospholipid syndrome (APS). The clinical manifestations of this syndrome most often documented thus far are recurrent deep venous thrombosis, recurrent spontaneous abortions, and cerebral vascular accidents. Abdominal ischemic events have received relatively little attention in prior reports. We report on a lupus patient with lupus anticoagulant positivity who presented with abdominal pain, anorexia, and weight loss who was subsequently diagnosed with gastric ulcers and pancreatitis. Computerized tomography of the abdomen in addition revealed splenic and kidney infarcts. We conclude that this patient had (ischemic) chronic pancreatitis with pseudocysts and splenic and renal infarcts probably due to secondary APS.