bromochloroacetic-acid and Inflammatory-Bowel-Diseases

The colon mucosa is lined with crypts of circa 300 cells, forming a continuous barrier whose roles include absorption of water, recovery of metabolic energy sources (notably short chain fatty acids), secretion of a protective mucus barrier, and physiological signalling. There is high turnover and replenishment of cells in the mucosa, disruption of this may lead to bowel pathologies including cancer and inflammatory bowel disease. Keratins have been implicated in the processes of cell death, epithelial integrity, response to inflammation and as a result are often described as guardians of the colonic epithelium. Keratin proteins carry extensive post-translational modifications, the cofactors for kinases, acetyl transferases and other modification-regulating enzymes are themselves products of metabolism. A cluster of studies has begun to reveal a bidirectional relationship between keratin form and function and metabolism. In this paper we hypothesise a mechanistic interaction between keratins and metabolism is governed through regulation of post-translational modifications and may contribute significantly to the normal functioning of the colon, placing keratins at the centre of a nutrition-metabolism-health triangle.

Topics: Colon; Colorectal Neoplasms; Humans; Inflammatory Bowel Diseases; Intestinal Mucosa; Keratins; Rectum

Keratins are cytoskeletal proteins that provide structural support to epithelial cells and tissues. Perturbation causes cell and tissue fragility and accounts for a large number of genetic disorders in humans. In humans, 54 functional keratin genes exist and 21 different keratin genes including hair keratins and hair follicle-specific epithelial keratins have been associated with hereditary disorders. Moreover, keratins have been implicated in more complex traits such as liver disease and inflammatory bowel disease. Understanding the molecular basis of keratin disorders has been the basis for improved diagnosis with prognostic implications, genetic counseling and prenatal testing for severe disorders. Besides their mechanical role, keratins have newly identified functions in apoptosis, cell growth, tissue polarity, wound healing and tissue remodeling. Improved understanding of the regulatory functions of keratins may offer novel approaches to overcome current treatment limitations.

Topics: Corneal Dystrophies, Hereditary; Humans; Inflammatory Bowel Diseases; Keratins; Liver Diseases; Mutation; Skin Diseases, Genetic

Topics: Animals; Chromosome Mapping; Genetic Predisposition to Disease; Humans; Inflammatory Bowel Diseases; Keratin-8; Keratins; Phenotype

Whilst the importance of mutations in a wide range of keratins in skin fragility disorders is now well established, there is much less evidence for simple epithelial keratin involvement in disease. Some simple epithelial keratin mutations have been reported in liver cirrhosis and pancreatitis patients, and recently mutations in the simple epithelial keratin K8 were identified in a group of patients with inflammatory bowel disease (Crohn disease or ulcerative colitis). In comparison with the mutations seen in epidermal keratins, these simple epithelial mutations would be predicted to have mild consequences, although analysis shows that they do have a distinct effect. This review article discusses the evidence that these mutations are a predisposing factor for inflammatory bowel disease.

Topics: Colon; Cytoskeleton; Epithelial Cells; Genetic Predisposition to Disease; Humans; Inflammatory Bowel Diseases; Keratin-18; Keratin-8; Keratins; Mutation; Stress, Mechanical

We evaluated the usefulness of cytokeratin 20 (CK20) mRNA expression in the quantitative detection of circulating tumor cells in the blood of patients with colorectal cancer (CRC). Blood samples from healthy volunteers (HVs; n = 37), patients with localized (n = 42) and metastatic colorectal cancer (n = 40), and patients with chronic inflammatory bowel disease (CID; n = 15) were examined. After immunomagnetic enrichment using microbeads against human epithelial antigen, total RNA was extracted, reverse transcribed, and analyzed by real-time reverse transcriptase-polymerase chain reaction using the LightCycler instrument. CK20 expression in peripheral blood was found in 46 of 82 (56%) patients with CRC, 8 of 37 (22%) HVs, and 9 of 15 (60%) patients with CID. Levels of CK20 mRNA were significantly higher in blood samples from CRC patients (median 681) than in blood samples from HVs (median 0) (P = 0.001), whereas no difference could be detected between patients with CRC and CID. Although the present technique could not distinguish CRC from CID, the method warrants further efforts to improve sample preparation and tumor cell enrichment, which may render real-time CK20 reverse transcriptase-polymerase chain reaction a feasible technique in identifying circulating tumor cells in peripheral blood of cancer patients.

Topics: Adult; Aged; Aged, 80 and over; Case-Control Studies; Colorectal Neoplasms; Female; Health; HT29 Cells; Humans; Hydroxymethylbilane Synthase; Inflammatory Bowel Diseases; Keratin-20; Keratins; Male; Middle Aged; Reference Standards; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; ROC Curve; Sensitivity and Specificity

We have identified miss-sense mutations in keratin 8 in a subset of patients with inflammatory bowel disease (Crohn disease and ulcerative colitis). Inflammatory bowel diseases are a group of disorders that are polygenic in origin and involve intestinal epithelial breakdown. We investigated the possibility that these keratin mutations might contribute to the course of the disease by adversely affecting the keratin filament network that provides mechanical support to cells in epithelia. The mutations (Gly62 to Cys, Ile63 to Val and Lys464 to Asn) all lie outside the major mutation hotspots associated with severe disease in epidermal keratins, but using a combination of in vitro and cell culture assays we show that they all have detrimental effects on K8/K18 filament assembly in vitro and in cultured cells. The G62C mutation also gives rise to homodimer formation on oxidative stress to cultured intestinal epithelial cells, and homodimers are known to be polymerization incompetent. Impaired keratin assembly resulting from the K8 mutations found in some inflammatory bowel disease patients would be predicted to affect the maintenance and re-establishment of mechanical resilience in vivo, as required during keratin cytoskeleton remodeling in cell division and differentiation, which may lead to epithelial fragility in the gut. Simple epithelial keratins may thus be considered as candidates for genes contributing to a risk of inflammatory bowel disease.

Topics: Actin Cytoskeleton; Animals; Antibodies, Monoclonal; Base Sequence; Cell Differentiation; Chromosomes, Human, Pair 12; Colitis, Ulcerative; Crohn Disease; Dimerization; Electrophoresis, Polyacrylamide Gel; Humans; Inflammation; Inflammatory Bowel Diseases; Keratin-8; Keratins; Mice; Models, Genetic; Molecular Sequence Data; Mutation; Oxidative Stress; Polymers; Protein Binding; Protein Conformation; Sequence Analysis, DNA; Time Factors; Transfection; Xenopus

Topics: Cell Cycle Proteins; GTP-Binding Proteins; Humans; Inflammatory Bowel Diseases; Keratin-8; Keratins; Mutation; Pyoderma Gangrenosum

Keratin 8 is a major component of intermediate filaments in single-layered epithelia of the gastrointestinal tract. Keratin 8 deficient mice display signs of colitis and diarrhoea characteristic for inflammatory bowel disease. Very recently, two keratin 8 mutations, Y54H and G62C, were identified.. We investigated if these keratin 8 missense mutations were associated with inflammatory bowel disease.. In total, 217 German patients with Crohn' s disease, 131 German patients with ulcerative colitis, and 560 German control subjects were enrolled in this study.. Samples were analysed by PCR amplification and subsequent melting curve analysis using fluorescence resonance energy transfer probes.. The G62C mutation was detected in five (2.3%) patients presenting with Crohn's disease and in three (2.3%) with ulcerative colitis. In comparison, 9 (1.6%) out of 560 controls were heterozygous for this mutation. No patient or control was homozygous for this mutation. Patients carrying one mutant allele did not show any noticeable characteristics in their corresponding phenotype. In contrast, the Y54H mutation was observed in neither any of the 348 patients with inflammatory bowel disease nor in any control subject.. Our data indicate that both keratin 8 mutations, G62C and Y54H, do not play a relevant pathogenic role in inflammatory bowel disease.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Case-Control Studies; DNA Mutational Analysis; Female; Heterozygote; Humans; Inflammatory Bowel Diseases; Keratin-8; Keratins; Male; Middle Aged; Mutation, Missense

Fractalkine (CX3CL1) is synthesized as a type I transmembrane protein. Its unique CX(3)C chemokine domain is attached to a 241-amino acid mucin stalk, a 19-amino acid transmembrane domain, and a 37-amino acid intracellular domain of unknown function. A soluble form of fractalkine can be generated by proteolytic cleavage at the base of the mucin stalk. Novel monoclonal and polyclonal antibodies that specifically recognize only the amino- or carboxyl-terminal ends of the human fractalkine molecule have revealed that epithelial cells are the predominant cell type expressing transmembrane forms of fractalkine in human skin, the tonsil, and the large intestine. Using these specific anti-fractalkine reagents we do not detect high-level expression of fractalkine on endothelial cells in normal or inflamed colon samples obtained from patients with Crohn's disease or ulcerative colitis. In contrast to previous reports we do not detect fractalkine expression by Langerhans cells or immature dendritic cells in mucosal-associated lymphoid tissues in vivo. We show that the reagent used in previous studies, an anti-fractalkine N-terminal peptide antisera, cross-reacts with human CD84. Finally we discuss potential roles for fractalkine in constitutive leukocyte trafficking based on its observed pattern of expression in epithelia.

Topics: Adenocarcinoma; Animals; Antibodies; Antibody Specificity; Antigens, CD; Cell Line; Chemokine CX3CL1; Chemokines, CX3C; Chemokines, CXC; CHO Cells; Colon; Colorectal Neoplasms; Cricetinae; Cross Reactions; Epidermis; Epithelial Cells; Humans; Inflammatory Bowel Diseases; Keratins; Membrane Glycoproteins; Membrane Proteins; Palatine Tonsil; Peptides; Protein Isoforms; Signaling Lymphocytic Activation Molecule Family; Tumor Cells, Cultured