Andreas Stallmach

NOD2 (CARD15) mutations in Crohn's disease are associated with diminished mucosal alpha-defensin expression.

Background: Mutations in NOD2, a putative intracellular receptor for bacterial peptidoglycans, are associated with a subset of Crohn’s disease but the molecular mechanism linking this protein with the disease pathogenesis remains unclear. Human α defensins (HD-5 and HD-6) are antibiotic effector molecules predominantly expressed in Paneth cells of the ileum. Paneth cells also express NOD2. To address the hypothesis that the function of NOD2 may affect expression of Paneth cell defensins, we compared their expression levels with respect to NOD2 mutations in Crohn’s disease. Methods: Forty five Crohn’s disease patients (24 with NOD2 mutations, 21 with wild-type NOD2) and 12 controls were studied. Real time reverse transcription-polymerase chain reaction was performed with mucosal mRNA for HD-5, HD-6, lysozyme, secretory phospholipase A2 (sPLA2), tumour necrosis factor α, interleukin 8, and human hypoxanthine phosphoribosyltransferase (housekeeping gene). Immunohistochemistry with anti-HD-5 and histological Paneth cell staining were performed in 10 patients with NOD2 mutations or wild-type genotypes. Results: Ileal expression of HD-5 and HD-6, but not sPLA2 or lysozyme, were diminished in affected ileum, and the decrease was significantly more pronounced in patients with NOD2 mutations. In the colon, HD-5, HD-6, and sPLA2 were increased during inflammation in wild-type but not in NOD2 mutated patients. In both the colon and ileum, proinflammatory cytokines and lysozyme were unaffected by NOD2 status. Immunohistochemistry identified Paneth cells as the sole source of HD-5. Conclusion: As alpha defensins are important in the mucosal antibacterial barrier, their diminished expression may explain, in part, the bacterial induced mucosal inflammation and ileal involvement of Crohn’s disease, especially in the case of NOD2 mutations.

TGF-β and fibrosis in different organs — molecular pathway imprints

The action of transforming-growth-factor (TGF)-beta following inflammatory responses is characterized by increased production of extracellular matrix (ECM) components, as well as mesenchymal cell proliferation, migration, and accumulation. Thus, TGF-beta is important for the induction of fibrosis often associated with chronic phases of inflammatory diseases. This common feature of TGF-related pathologies is observed in many different organs. Therefore, in addition to the description of the common TGF-beta-pathway, this review focuses on TGF-beta-related pathogenetic effects in different pathologies/organs, i. e., arthritis, diabetic nephropathy, colitis/Crohns disease, radiation-induced fibrosis, and myocarditis (including their similarities and dissimilarities). However, TGF-beta exhibits both exacerbating and ameliorating features, depending on the phase of disease and the site of action. Due to its central role in severe fibrotic diseases, TGF-beta nevertheless remains an attractive therapeutic target, if targeted locally and during the fibrotic phase of disease.

Expression of interleukin‐12‐related cytokine transcripts in inflammatory bowel disease: Elevated interleukin‐23p19 and interleukin‐27p28 in Crohn's disease but not in ulcerative colitis

Background: It has been suggested that Crohns disease (CD) is associated with an exaggerated T‐helper 1 cytokine response manifested by increased production of interleukin (IL)‐12. IL‐12 is a heterodimeric protein comprising 2 disulfide‐linked subunits designated p35 and p40. Recently, IL‐12‐related cytokines, IL‐23 and IL‐27, were described. Biologically active IL‐23 is a heterodimer whose p40 subunit is identical to IL‐12p40 whereas its p19 subunit is distantly related to IL‐12p35. IL‐27 consists of EBI3, an IL‐12p40‐related protein, and p28, a newly discovered IL‐12p35‐related polypeptide. Aim: We sought to determine whether mucosal expression of IL‐23p19 and IL‐27p28 transcripts correlate with the inflammatory activity in inflammatory bowel disease (IBD). Patients/Methods: Messenger RNA expression in colonic mucosa from patients with Crohns disease (CD; n = 37) and ulcerative colitis (UC; n = 19), and in non‐IBD control subjects (specific colitis [SC]; n = 16) and normal, nondiseased control patients (n = 12) was measured by reverse‐transcribed real‐time polymerase chain reaction. Results: IL‐23p19 was significantly increased in inflamed mucosa in CD (P = 0.0377) and to a lesser extent also in UC patients but not in SC patients. Elevation of IL‐23p19 transcript levels in CD correlated with the severity of endoscopic lesions. IL‐27p28 transcripts and EBI3 transcripts were significantly elevated only in active CD. Discussion: IL‐23p19, IL‐27p28, and EBI3 transcripts are strongly up‐regulated in CD. The stimulatory effects of these cytokines on naive T cells in addition to a strongly synergistic action with IL‐12 to trigger interferon‐&ggr; production may contribute to the perpetuation of the inflammatory process in patients with CD. Notably, increased expression of IL‐23 and IL‐27 transcripts in CD suggests a T helper 1‐dominated immunologic function in this disease.

Updated German Guideline on Diagnosis and Treatment of Ulcerative Colitis, 2011

! Hintergrund Die Colitis ulcerosa (CU) ist neben dem Morbus Crohn die wichtigste chronisch-entzundliche Darmerkrankung (CED). Die Inzidenz fur die Colitis ulcerosa liegt in Deutschland bei bei 3,0–3,9 pro 100000 Einwohner [1, 2]. Die Pravalenz durfte in der westlichen Welt derzeit bei 160– 250 pro 100000 Einwohner liegen [3, 4]. Der hochste Gipfel der alterspezifischen Inzidenz liegt bei den 16bis 25-Jahrigen, wenn auch die Verteilung uber die Altersdekaden weit gleichmasiger ist als fruher beschrieben wurde [2]. Somit beginnt fur die meisten Patienten ihre Erkrankung wahrend der Schulzeit oder der Berufausbildung und dauert wahrend ihres gesamten beruflichen Lebens an. Daraus folgt, dass durch die Erkrankung nicht nur direkte Kosten (Medikamente, Arztbesuche, Operationen, Krankenhausaufenthalte etc.), sondern auch umfangreiche indirekte Kosten (Rente, Arbeitsausfalle etc.) entstehen. Es ist davon auszugehen, dass bei Patienten mit CU ca. die Halfte der Gesamtkosten den indirekten Kosten zuzuordnen sind [5]. An direkten medizinischen Kosten wurden dabei zuletzt zwischen 2500 und 5000€ pro Inhaltsverzeichnis! Hintergrund Die Colitis ulcerosa (CU) ist neben dem Morbus Crohn die wichtigste chronisch-entzündliche Darmerkrankung (CED). Die Inzidenz für die Colitis ulcerosa liegt in Deutschland bei bei 3,0–3,9 pro 100000 Einwohner [1, 2]. Die Prävalenz dürfte in der westlichen Welt derzeit bei 160– 250 pro 100000 Einwohner liegen [3, 4]. Der höchste Gipfel der alterspezifischen Inzidenz liegt bei den 16bis 25-Jährigen, wenn auch die Verteilung über die Altersdekaden weit gleichmäßiger ist als früher beschrieben wurde [2]. Somit beginnt für die meisten Patienten ihre Erkrankung während der Schulzeit oder der Berufausbildung und dauert während ihres gesamten beruflichen Lebens an. Daraus folgt, dass durch die Erkrankung nicht nur direkte Kosten (Medikamente, Arztbesuche, Operationen, Krankenhausaufenthalte etc.), sondern auch umfangreiche indirekte Kosten (Rente, Arbeitsausfälle etc.) entstehen. Es ist davon auszugehen, dass bei Patienten mit CU ca. die Hälfte der Gesamtkosten den indirekten Kosten zuzuordnen sind [5]. An direkten medizinischen Kosten wurden dabei zuletzt zwischen 2500 und 5000€ pro Inhaltsverzeichnis

Rapid mucosal CD4+ T-cell depletion and enteropathy in simian immunodeficiency virus-infected rhesus macaques

BACKGROUND & AIMS Human immunodeficiency virus (HIV) infection leads to severe immunologic and functional disturbances in the intestinal tract in late stages of the disease. Information on mucosal pathology directly after infection is limited. We characterized this early phase in rhesus macaques infected with simian immunodeficiency virus (SIV). METHODS Eight rhesus macaques were infected with SIV. Upper endoscopy was performed at defined times before and after infection. Viral load, percentage of CD4(+) T cells, villus height, crypt depth, and Ki-67-positive crypt cells were analyzed in duodenal biopsy specimens. Serum beta-carotene and vitamin D levels were assessed. RESULTS A rapid increase of duodenal SIV core protein (p27) concentration and an almost complete loss of intestinal CD4(+) T cells was found within 2 weeks after infection. A decrease of villus height was observed, and the percentage of Ki-67-positive (proliferating) crypt cells increased. Serum concentrations of vitamin D decreased in 6 of 8 animals, and beta-carotene concentrations decreased in 3 of 8 animals after infection. CONCLUSIONS Mucosal SIV replication and intestinal CD4(+) T cell depletion are early events in SIV-infected rhesus macaques. The structural changes of the mucosa strongly support the concept of HIV/SIV-induced enteropathy. In contrast to late-stage human HIV infection, early small intestinal villous atrophy in SIV infection is associated with crypt hyperplasia.

Immunosuppressive Therapy in Whipple's Disease Patients is Associated with the Appearance of Gastrointestinal Manifestations

OBJECTIVES:Whipples disease is a rare chronic disorder, which is caused by systemic infection with Tropheryma whipplei. The first symptom of Whipples disease usually is a nondestructive polyarthritis resembling in many aspects seronegative rheumatoid arthritis. This polyarticular inflammatory arthropathy preceding the diagnosis of Whipples disease for several years frequently is treated with nonsteroidal antiinflammatory drugs (NSAIDs) and with immunosuppressive therapy. There is evidence that altered immune functions play a role in the manifestation of the disease and that Whipples disease is associated with opportunistic infections. We therefore asked whether immunosuppressive treatment for arthropathy may alter the course of Whipples disease.PATIENTS AND METHODS:In a series of 27 patients with Whipples disease clinical data were documented and the patients were followed for 3–4 yr. The patients were classified into three groups according to their medication: (i) patients with immunosuppressive therapy preceding the diagnosis, (ii) patients with NSAIDs before diagnosis, and (iii) patients without such therapies.RESULTS:Arthropathies occurred in the mean 8 yr before diagnosis and were the first symptom in 63% of the patients. Gastrointestinal involvement usually became evident later on and frequently led to the diagnosis of Whipples disease. In patients with immunosuppressive treatment, diarrhea occurred in the median 4 months after the initiation of such therapy and diagnosis of Whipples disease was made after another 2 months. In contrast, other medical treatments were not closely followed by the onset of diarrhea.CONCLUSIONS:These results indicate an association between immunosuppressive therapy and the onset of diarrhea in Whipples disease and thus support the concept that immunologic factors play a role in disease pathogenesis. Further investigation on the interaction of the immune system and Tropheryma whipplei infection are required to understand the factors contributing to the clinical manifestation of this rare disorder and possibly to introduce preventive interventions.

Adverse effects of biologics used for treating IBD

In the last decade, biologic agents, in particular anti-TNF agents such as infliximab, adalimumab, and certolizumab have substantially extended the therapeutic armamentarium of inflammatory bowel disease (IBD). Additional approaches include biologicals, such as natalizumab, that block leucocyte adhesion; those that target cytokines, such as interleukin-12/23 antibodies; or those that inhibit T-cell signaling, such as interleukin-6 receptor antibodies. However, these drugs have a number of contraindications and side effects, especially when used in combination with classical immunosuppressive agents or corticosteroids. Areas of concern include opportunistic infections, malignancies, and miscellaneous complications such as injection/infusion reactions and autoimmunity and contraindications, such as heart failure and acute infectious diseases. In this review, the indications of biologicals in IBD treatment are briefly reported, and the potential disadvantages of a more active therapeutic approach in IBD are discussed. We have learned in the last decade that anti-TNF-alpha therapy is an effective and relatively safe treatment option for selected patients that changes the natural course of severe IBD. However, despite these changed therapeutic paradigms and goals in IBD, clinicians should be aware that the powerful immunosuppressive capacity of biologicals necessitates a rigorous long-term safety follow-up.

Long-Term Effectiveness of Azathioprine in IBD Beyond 4 Years: A European Multicenter Study in 1176 Patients

In Crohn’s disease the optimal duration of azathioprine treatment is still controversial and for ulcerative colitis only limited data are available to support its efficacy. Charts of 1176 patients with IBD from 16 European centers were analyzed. Flare incidences and steroid dosages were assessed for the time before and during treatment and after discontinuation. Within the first 4 years, azathioprine suppressed flare incidence and steroid consumption in both diseases (P < 0.001). While in CD discontinuation after 3–4 years did not lead to reactivation, this was the case in UC. However, continuation beyond 4 years further improved clinical activity in CD and steroid requirement in both diseases (P < 0.001). Discontinuation of azathioprine may thus be considered after 3–4 years in CD patients in complete remission without steroid requirement. In all other CD patients and for UC patients in general, continuation seems beneficial. These results support a novel differential algorithm for long-term azathioprine therapy in IBD.

Nano- and microscaled particles for drug targeting to inflamed intestinal mucosa—A first in vivo study in human patients

Most of the drugs used in the treatment of inflammatory bowel disease (IBD) become systemically bioavailable and potentially bear strong adverse effects. Targeting the inflamed areas of the intestine and keeping the drug localised at its site of action can reduce adverse effects. In animal studies, luminal uptake into inflamed mucosal areas has been shown to be size dependent. We investigated the potential of nano- and microparticle uptake into the rectal mucosa of human IBD patients. Fluorescently labelled placebo nanoparticles (NP) 250nm in size and microparticles (MP) 3.0μm in size were prepared. 2h after rectal application to patients with Crohns disease (CD) or ulcerative colitis (UC), confocal laser endomicroscopy was performed to visualise the particles in inflamed mucosal areas. In biopsies, ex vivo mucosal transport processes were investigated in miniaturised Ussing chambers. We examined 33 patients with IBD (19 patients with CD, 14 patients with UC) and 6 healthy controls. A significantly enhanced accumulation of MP in ulcerous lesions was observed (covered area=1.28% (range 0.83%-3.45%) vs. 0% in controls; p=0.011), while NP were visible only in traces on mucosal surfaces of all patients. The Ussing chamber experiments suggest persorption of particles through cellular voids; statistical significance was only reached for NP. Drug-containing particles may have great potential to more specifically target intestinal lesions to maximise therapeutic efficacy and minimise potential side effects. Nanoparticles may not be required for local drug delivery to intestinal lesions in humans, thereby minimising the risk of unintended translocation into the blood system.

Fibrogenesis and Fibrolysis in Collagenous Colitis Patterns of Procollagen Types I and IV, Matrix- Metalloproteinase-1 and -13, and TIMP-1 Gene Expression

Collagenous colitis is characterized by the deposition of a superficial subepithelial collagenous layer, the pathogenesis of which is unknown. Because the excess matrix deposition is potentially reversible, a labile imbalance between fibrogenesis and fibrolysis may be suspected. Expression of procollagen alpha1(I) and alpha1(IV), matrix-metalloproteinase (MMP)-1 and -13, and tissue inhibitor of metalloproteinase (TIMP)-1 genes was semiquantitated by in situ hybridization on serial biopsies of 12 patients with collagenous colitis and compared to controls. Collagen types I, III, IV, and VI, tenascin, undulin/collagen XIV, and alpha-actin were localized by immunohistology. The superficial collagen layer stained strongly for collagen types I, III, and VI, and particularly for tenascin, but not for undulin. Elevated procollagen alpha1(I), procollagen alpha1(IV), and TIMP-1 transcript levels were found in alpha-actin-positive cells with linear distribution underneath the superficial collagenous layer, whereas MMP-1 RNA expression was variable and restricted to cell clusters. MMP-13 expression was undetectable. The patterns of procollagen alpha1(I)- and alpha1(IV)-specific labeling, combined with an intense tenascin- but absent undulin-specific staining, indicate deposition of an immature interstitial matrix that may be susceptible to degradation. The restricted MMP-1 RNA expression, counteracted by increased TIMP-1 expression, suggests locally impaired fibrolysis as a relevant factor in the pathogenesis of collagenous colitis.