scholarly journals Increased IGF-IEc expression and mechano-growth factor production in intestinal muscle of fibrostenotic Crohn's disease and smooth muscle hypertrophy

2015 ◽  
Vol 309 (11) ◽  
pp. G888-G899 ◽  
Author(s):  
Chao Li ◽  
Kent Vu ◽  
Krystina Hazelgrove ◽  
John F. Kuemmerle
Keyword(s):  
Smooth Muscle ◽  
Crohn’S Disease ◽  
Growth Factor ◽  
Crohn's Disease ◽  
Muscle Hypertrophy ◽  
Smooth Muscle Actin ◽  
Specific Gene ◽  
Stricture Formation ◽  
Muscle Actin ◽  
Mechano Growth Factor

The igf1 gene is alternatively spliced as IGF-IEa and IGF-IEc variants in humans. In fibrostenotic Crohn's disease, the fibrogenic cytokine TGF-β1 induces IGF-IEa expression and IGF-I production in intestinal smooth muscle and results in muscle hyperplasia and collagen I production that contribute to stricture formation. Mechano-growth factor (MGF) derived from IGF-IEc induces skeletal and cardiac muscle hypertrophy following stress. We hypothesized that increased IGF-IEc expression and MGF production mediated smooth muscle hypertrophy also characteristic of fibrostenotic Crohn's disease. IGF-IEc transcripts and MGF protein were increased in muscle cells isolated from fibrostenotic intestine under regulation by endogenous TGF-β1. Erk5 and MEF2C were phosphorylated in vivo in fibrostenotic muscle; both were phosphorylated and colocalized to nucleus in response to synthetic MGF in vitro. Smooth muscle-specific protein expression of α-smooth muscle actin, γ-smooth muscle actin, and smoothelin was increased in affected intestine. Erk5 inhibition or MEF2C siRNA blocked smooth muscle-specific gene expression and hypertrophy induced by synthetic MGF. Conditioned media of cultured fibrostenotic muscle induced muscle hypertrophy that was inhibited by immunoneutralization of endogenous MGF or pro-IGF-IEc. The results indicate that TGF-β1-dependent IGF-IEc expression and MGF production in patients with fibrostenotic Crohn's disease regulates smooth muscle cell hypertrophy a critical factor that contributes to intestinal stricture formation.

scholarly journals P067 Proteins citrullination and Crohn’s disease: PAD4 but not PAD2 is a strong marker of ileal inflammation

2020 ◽  
Vol 14 (Supplement_1) ◽  
pp. S168-S169
Author(s):  
G Dragoni ◽  
B Creyns ◽  
G De Hertogh ◽  
B Verstockt ◽  
W J Wollants ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Crohn’S Disease ◽  
Crohn's Disease ◽  
Smooth Muscle Actin ◽  
Muscle Actin ◽  
Post Translational Modification ◽  
Right Colon Cancer ◽  
Expert Pathologist ◽  
Inflammatory Infiltrates ◽  
Activated Fibroblasts

Abstract Background Citrullination is a post-translational modification of proteins, mediated by enzymes called PAD (peptidylarginine deiminases). The immune system can attack citrullinated proteins, leading to autoimmune diseases such as rheumatoid arthritis, multiple sclerosis and ulcerative colitis, and the activity of PAD2 and PAD4 in innate immune cells has been demonstrated for these disorders. Recently, high levels of PAD2 have been described in activated fibroblasts in the context of liver fibrosis. We therefore investigated the role of PAD2 and PAD4, both in inflammatory and fibrotic contexts of ileal Crohn’s disease (CD). Methods We obtained ileal transmural samples from patients operated for stricturing ileal CD. Three different macroscopic areas within each resection specimen (i.e. proximal normal ileum, inflamed ileum and fibrotic ileum) were selected and histologically confirmed by an expert pathologist. Patients undergoing ileocolic resection for other conditions (e.g. right colon cancer) and with healthy terminal ileum were used as controls. For each region (normal CD, inflamed CD, fibrotic CD and control), immunohistochemistry (IHC), RNA and protein evaluations for PAD2 and PAD4 were performed. Multiplex immunofluorescence (IF) for PAD2, PAD4, myeloperoxidase, neutrophil elastase, CD68, vimentin and α-smooth muscle actin were carried out to investigate the enzymes-expressing cells. Additional IF was performed to study citrullinated histone 3 (H3cit) expression, the product of PAD4 activity in neutrophils and component of neutrophil extracellular traps (NETs). Statistical analysis was carried out with Kruskal–Wallis test and post hoc Mann–Whitney test. Results Resection specimens from 13 CD and 11 controls were included. IHC and IF showed an increased expression of both PAD2 and PAD4 in the neutrophils of inflamed areas, in cytoplasm and nucleus, respectively (Figure 1). Activated fibroblasts (vimentin+ and α-smooth muscle actin+) were negative for both enzymes. PAD4 mRNA expression was increased in inflamed tissue (p = 0.001, p = 0.008 and p = 0.028 vs. normal CD, fibrotic CD and controls, respectively), and confirmed using Western Blot (Figure 2). H3cit was increased in the ileal inflammatory infiltrates too (Figure 3), confirming high PAD4 expression. For PAD2, no significant changes were observed at RNA and protein level, mainly due to its reduced expression in epithelial cells from normal to diseased tissue (Figure 4). Conclusion Both PAD2 and PAD4 are strongly expressed in neutrophils of CD ileal resection specimens, but only PAD4 shows a significantly higher expression in the inflammatory context which translates in the formation of NETs. No direct correlation was observed between PAD enzymes and intestinal fibroblasts.

scholarly journals A novel human cell culture model to study visceral smooth muscle phenotypic modulation in health and disease

2018 ◽  
Vol 315 (4) ◽  
pp. C598-C607 ◽  
Author(s):  
Rianne D. W. Vaes ◽  
Linda van den Berk ◽  
Bas Boonen ◽  
David P. J. van Dijk ◽  
Steven W. M. Olde Damink ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Smooth Muscle ◽  
Growth Factor ◽  
Muscle Protein ◽  
Smooth Muscle Actin ◽  
Muscle Actin ◽  
Phenotypic Modulation ◽  
Culture Model ◽  
Health And Disease ◽  
Visceral Smooth Muscle

Adaptation of the smooth muscle cell (SMC) phenotype is essential for homeostasis and is often involved in pathologies of visceral organs (e.g., uterus, bladder, gastrointestinal tract). In vitro studies of the behavior of visceral SMCs under (patho)-physiological conditions are hampered by a spontaneous, uncontrolled phenotypic modulation of visceral SMCs under regular tissue culture conditions. We aimed to develop a new visceral SMC culture model that allows controlled phenotypic modulation. Human uterine SMCs [ULTR and telomerase-immortalized human myometrial cells (hTERT-HM)] were grown to confluency and kept for up to 6 days on regular tissue culture surfaces or basement membrane (BM) matrix-coated surfaces in the presence of 0–10% serum. mRNA and protein expression and localization of SMC-specific phenotype markers and their transcriptional regulators were investigated by quantitative PCR, Western blotting, and immunofluorescence. Maintaining visceral SMCs confluent for 6 days increased α-smooth muscle actin (1.9-fold) and smooth muscle protein 22-α (3.1-fold), whereas smooth muscle myosin heavy chain was only slightly upregulated (1.3-fold). Culturing on a BM matrix-coated surface further increased these proteins and also markedly promoted mRNA expression of γ-smooth muscle actin (15.0-fold), smoothelin (3.5-fold), h-caldesmon (5.2-fold), serum response factor (7.6-fold), and myocardin (8.1-fold). Whereas additional serum deprivation only minimally affected contractile markers, platelet-derived growth factor-BB and transforming growth factor β1 consistently reduced versus increased their expression. In conclusion, we present a simple and reproducible visceral SMC culture system that allows controlled phenotypic modulation toward both the synthetic and the contractile phenotype. This may greatly facilitate the identification of factors that drive visceral SMC phenotypic changes in health and disease.

scholarly journals Magnetic Resonance Imaging Predicts Histopathological Composition of Ileal Crohn’s Disease

2018 ◽  
Vol 12 (6) ◽  
pp. 718-729 ◽  
Author(s):  
Mathilde Wagner ◽  
Huaibin Mabel Ko ◽  
Manjil Chatterji ◽  
Cecilia Besa ◽  
Joana Torres ◽  
...  
Keyword(s):  
Magnetic Resonance Imaging ◽  
Smooth Muscle ◽  
Crohn’S Disease ◽  
Magnetic Resonance ◽  
Small Bowel ◽  
Crohn's Disease ◽  
Wall Thickness ◽  
Muscle Hypertrophy ◽  
Resonance Imaging ◽  
Post Contrast

Abstract Background and Aims Recently, smooth muscle hypertrophy has been suggested to be a contributor to small bowel lesions secondary to Crohn’s disease [CD], in addition to inflammation and fibrosis. Here, we assess the value of magnetic resonance imaging [MRI] for the characterisation of histopathological tissue composition of small bowel CD, including inflammation, fibrosis, and smooth muscle hypertrophy. Methods A total of 35 consecutive patients [male/female 17/18, mean age 33 years] with ileal CD, who underwent small bowel resection and a preoperative contrast-enhanced MRI examination within 1 month before surgery, were retrospectively included. Image assessment included qualitative [pattern/degree of enhancement, presence of ulcerations/fistulas/abscesses] and quantitative parameters [wall thickness on T2/T1-weighted images [WI], enhancement ratios, apparent diffusion coefficient [ADC], Clermont and Magnetic Resonance Index of Activity [MaRIA] scores). MRI parameters were compared with histopathological findings including active inflammation, collagen deposition, and muscle hypertrophy using chi square/Fisher or Mann-Whitney tests and univariate/multivariate logistic/linear regression analyses. Results Forty ileal segments were analysed in 35 patients. Layered pattern at early-post-contrast phase was more prevalent (odds ratio [OR] = 8; p = 0.008), ADC was significantly lower [OR = 0.005; p = 0.022], and MaRIA score was significantly higher [OR = 1.125; p = 0.022] in inflammation grades 2–3 compared with grade 1. Wall thickness on T2WI was significantly increased [OR = 1.688; p = 0.043], and fistulas [OR = 14.5; p = 0.017] were more prevalent in segments with disproportionately increased muscle hypertrophy versus those with disproportionately increased fibrosis. MaRIA/Clermont scores, wall thickness on T1WI and T2WI, and ADC were all significantly correlated with degree of muscular hypertrophy. Conclusions MRI predicts the degree of inflammation, and can distinguish prominent muscle hypertrophy from prominent fibrosis in ileal CD with reasonable accuracy (area under receiver operating characteristic curve [AUROC] > 0.7).

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