Novel approaches: Tissue engineering and stem cells--In vitro modelling of the gut

In many intestinal diseases, the function of the epithelial lining is impaired. In this review, we describe the recent developments of in vitro intestinal stem cell cultures. When these stem cells are grown in 3D structures (organoids), they provide a model of the intestinal epithelium, which is closely similar to the growth and development of the in vivo gut. This model provides a new tool to study various diseases of malabsorption in functional detail and therapeutic applications, which could not be achieved with traditional cell lines. First, we describe the organization and function of the healthy small intestinal epithelium. Then, we discuss the establishment of organoid cultures and how these structures represent the healthy epithelium. Finally, we discuss organoid cultures as a tool for studying intrinsic properties of the epithelium, as a model for intestinal disease, and as a possible source for stem cell transplantations

Novel approaches : Tissue engineering and stem cells--In vitro modelling of the gut

In many intestinal diseases, the function of the epithelial lining is impaired. In this review, we describe the recent developments of in vitro intestinal stem cell cultures. When these stem cells are grown in 3D structures (organoids), they provide a model of the intestinal epithelium, which is closely similar to the growth and development of the in vivo gut. This model provides a new tool to study various diseases of malabsorption in functional detail and therapeutic applications, which could not be achieved with traditional cell lines. First, we describe the organization and function of the healthy small intestinal epithelium. Then, we discuss the establishment of organoid cultures and how these structures represent the healthy epithelium. Finally, we discuss organoid cultures as a tool for studying intrinsic properties of the epithelium, as a model for intestinal disease, and as a possible source for stem cell transplantations

Adult stem cells in the small intestine are intrinsically programmed with their location-specific function

Differentiation and specialization of epithelial cells in the small intestine are regulated in two ways. First, there is differentiation along the crypt-villus axis of the intestinal stem cells into absorptive enterocytes, Paneth, goblet, tuft, enteroendocrine, or M cells, which is mainly regulated by WNT. Second, there is specialization along the cephalocaudal axis with different absorptive and digestive functions in duodenum, jejunum, and ileum that is controlled by several transcription factors such as GATA4. However, so far it is unknown whether location-specific functional properties are intrinsically programmed within stem cells or if continuous signaling from mesenchymal cells is necessary to maintain the location-specific identity of the small intestine. Using the pure epithelial organoid technique, we show that region-specific gene expression profiles are conserved throughout long-term cultures of both mouse and human intestinal stem cells and correlated with differential Gata4 expression. Furthermore, the human organoid culture system demonstrates that Gata4-regulated gene expression is only allowed in absence of WNT signaling. These data show that location-specific function is intrinsically programmed in the adult stem cells of the small intestine and that their differentiation fate is independent of location-specific extracellular signals. In light of the potential future clinical application of small intestine-derived organoids, our data imply that it is important to generate GATA4-positive and GATA4-negative cultures to regenerate all essential functions of the small intestine

Many inflammatory bowel disease risk loci include regions that regulate gene expression in immune cells and the intestinal epithelium

BACKGROUND & AIMS: The contribution of genetic factors to the pathogenesis of inflammatory bowel disease (IBD) has been established by twin, targeted-sequencing, and genome-wide association studies. These studies identified many risk loci, and research is underway to identify causal variants. These studies have focused mainly on protein-coding genes. We investigated other functional elements in the human genome, such as regulatory regions. METHODS: Using acetylated histone 3 lysine 27 chromatin immunoprecipitation and sequencing, we identified tens of thousands of potential regulatory regions that are active in intestinal epithelium (primary intestinal crypts and cultured organoids) isolated from resected material and from biopsies collected during ileo-colonoscopies and immune cells (monocytes, macrophages, CD34(+), CD4(+), and CD8(+)). We correlated these regions with susceptibility loci for IBD. RESULTS: We have generated acetylated histone 3 lysine 27 profiles from primary intestinal epithelium and cultured organoids, which we have made publically available. We found that 45 of 163 single nucleotide polymorphisms (SNPs) associated with IBD overlap specifically with active regulatory elements. In addition, by taking strong linkage disequilibrium into account, another 47 IBD-associated SNPs colocalized with active regulatory elements through other SNPs in their vicinity. Altogether, 92 of 163 IBD-associated SNPs correlated with distinct active regulatory elements-a frequency 2.5- to 3.5-fold greater than that expected from random sampling. The variations in these SNPs often create or disrupt known binding motifs; they might affect the binding of transcriptional regulators to alter expression of regulated genes. CONCLUSIONS: In addition to variants in protein coding genes, variants in noncoding DNA regulatory regions that are active in intestinal epithelium and immune cells are potentially involved in the pathogenesis of IBD

Tissue-specific mutation accumulation in human adult stem cells during life

The gradual accumulation of genetic mutations in human adult stem cells (ASCs) during life is associated with various age-related diseases, including cancer. Extreme variation in cancer risk across tissues was recently proposed to depend on the lifetime number of ASC divisions, owing to unavoidable random mutations that arise during DNA replication. However, the rates and patterns of mutations in normal ASCs remain unknown. Here we determine genome-wide mutation patterns in ASCs of the small intestine, colon and liver of human donors with ages ranging from 3 to 87 years by sequencing clonal organoid cultures derived from primary multipotent cells. Our results show that mutations accumulate steadily over time in all of the assessed tissue types, at a rate of approximately 40 novel mutations per year, despite the large variation in cancer incidence among these tissues. Liver ASCs, however, have different mutation spectra compared to those of the colon and small intestine. Mutational signature analysis reveals that this difference can be attributed to spontaneous deamination of methylated cytosine residues in the colon and small intestine, probably reflecting their high ASC division rate. In liver, a signature with an as-yet-unknown underlying mechanism is predominant. Mutation spectra of driver genes in cancer show high similarity to the tissue-specific ASC mutation spectra, suggesting that intrinsic mutational processes in ASCs can initiate tumorigenesis. Notably, the inter-individual variation in mutation rate and spectra are low, suggesting tissue-specific activity of common mutational processes throughout life

Loss of syntaxin 3 causes variant microvillus inclusion disease

Microvillus inclusion disease (MVID) is a disorder of intestinal epithelial differentiation characterized by life-threatening intractable diarrhea. MVID can be diagnosed based on loss of microvilli, microvillus inclusions, and accumulation of subapical vesicles. Most patients with MVID have mutations in myosin Vb that cause defects in recycling of apical vesicles. Whole-exome sequencing of DNA from patients with variant MVID showed homozygous truncating mutations in syntaxin 3 (STX3). STX3 is an apical receptor involved in membrane fusion of apical vesicles in enterocytes. Patient-derived organoid cultures and overexpression of truncated STX3 in Caco-2 cells recapitulated most characteristics of variant MVID. We conclude that loss of STX3 function causes variant MVID

Maternal and perinatal outcomes by planned place of birth among women with low-risk pregnancies in high-income countries: A systematic review and meta-analysis

Background: The comparative safety of different birth settings is widely debated. Comparing research across high-income countries is complex, given differences in maternity service provision, data discrepancies, and varying research techniques and quality. Studies of births planned at home or in birth centres have reported both better and poorer outcomes than planned hospital births. Previous systematic reviews have focused on outcomes from either birth centres or home births, with inconsistent attention to quality appraisal. Few have attempted to synthesise findings. Objective: To compare maternal and perinatal outcomes from different places of birth via a systematic review of high-quality research, and meta-analysis of appropriate data (Prospero registration CRD42016042291). Design: Reviewers searched CINAHL, Embase, Maternity and Infant Care, Medline and PsycINFO databases to identify studies comparing selected outcomes by place of birth among women with low-risk pregnancies in high-income countries. They critically appraised identified studies using an instrument specific to birth place research and then combined outcome data via meta-analysis, using RevMan software. Findings: Twenty-eight articles met inclusion criteria, yielding comparative data on perinatal mortality, mode of birth, maternal morbidity and/or NICU admissions. Meta-analysis indicated that women planning hospital births had statistically significantly lower odds of normal vaginal birth than in other planned settings. Women experienced severe perineal trauma or haemorrhage at a lower rate in planned home births than in obstetric units. There were no statistically significant differences in infant mortality by planned place of birth, although most studies had limited statistical power to detect differences for rare outcomes. Differences in location, context, quality and design of identified studies render results subject to variation. Conclusions and implications for practice: High-quality evidence about low-risk pregnancies indicates that place of birth had no statistically significant impact on infant mortality. The lower odds of maternal morbidity and obstetric intervention support the expansion of birth centre and home birth options for women with low-risk pregnancies.Full Tex