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The erroneous adaptive immune response consequence of a highly specific interplay between selected gluten peptides and major histocompatibility complex class II HLA-DQ2/8-antigen restricted T cells plays a paramount role in CD pathogenesis . Dependent on the post-translational deamidation of gluten peptides by transglutaminase 2 (TG2), this interplay is influenced by the initial imprinting of the innate immune system through IL-15 upregulation that promotes the CD4+ T cell adaptive immune response [56, 57]. Presentation of gluten to CD4+ T cells carried out by dendritic cells as well as macrophages, B cells, and even enterocytes expressing HLA class II, can cause their recirculation in the lamina propria . The contact of CD4+ T cells in the lamina propria with gluten induces their activation and proliferation, with production of proinflammatory cytokines, metalloproteases, and keratinocyte growth factor by stromal cells, which induces cryptal hyperplasia and villous blunting secondary to intestinal epithelial cell death induced by intraepithelial lymphocytes (IELs) . Additionally, there is an overexpression of membrane-bound IL-15 on enterocytes in active CD causing over-expression of the natural killer (NK) receptors CD94 and NKG2D by CD3+ IELs . CD crypt hyperplasia has been hypothesized to be the consequence of an imbalance between continuous tissue damage due to the mucosal autoimmune insult described above and inability of the stem cells to compensate. We have recently provided a more mechanistic, evidence-based explanation for hyperplastic crypts in active CD by showing that the celiac hyperplastic crypt is characterized by an expansion of the immature progenitor cell compartment and downregulation of the Hedgehog signaling cascade . These data shed light on the molecular mechanisms underlying CD histopathology and illuminate the reason for the lack of enteropathy in the mouse models for CD. Indeed, lack of consistent CD-like enteropathy in humanized mice  supports the concept that the accelerated disruption of enterocytes secondary to the adaptive CD4+ T cell insult cannot fully explain CD pathogenesis, supporting the notion that an intrinsic defect of the stem cell compartment in subjects at risk of CD is a key element of CD enteropathy [60, 62].
crypts and things pdf 23
GvHD and allograft bowel rejection (AGBR) may be ruled out on clinical settings. GvHD may, however, come to the gastroenterologist or pathologist who are not provided with the history of bone marrow transplantation for instance. GvHD may have, although uncommonly, an increased IEL count in proximal small bowel biopsies. A decrescendo from base to apical villi and the finding of epithelial cell apoptosis in the deep crypts, with or without some degree of architectural disturbance, together with the clinical setting may help to address this diagnosis.
The study of somatic mutations with standard whole-genome sequencing requires isolating clonal groups of cells recently derived from a single cell8,13,14. To study somatic mutations across a diverse set of mammals, we isolated 208 individual intestinal crypts from 56 individuals across 16 species with a wide range of lifespans and body sizes: black-and-white colobus monkey, cat, cow, dog, ferret, giraffe, harbour porpoise, horse, human, lion, mouse, naked mole-rat, rabbit, rat, ring-tailed lemur and tiger (Supplementary Table 1). We chose intestinal crypts for several reasons. First, they are histologically identifiable units that line the epithelium of the colon and small intestine and are amenable to laser microdissection. Second, human studies have confirmed that individual crypts become clonally derived from a single stem cell and show a linear accumulation of mutations with age, which enables the estimation of somatic mutation rates through genome sequencing of single crypts8. Third, in most human crypts, most somatic mutations are caused by endogenous mutational processes common to other tissues, rather than by environmental mutagens8,18.
A colon sample was collected from each individual, with the exception of a ferret from which only a small intestine sample was available. This sample was included because results in humans have shown that the mutation rates of colorectal and small intestine epithelial stem cells are similar14,20 (Extended Data Fig. 1). We then used laser microdissection on histological sections to isolate individual crypts for whole-genome sequencing with a low-input library preparation method29 (Fig. 1a, Extended Data Fig. 2, Supplementary Table 2), with the exception of human crypts, for which sequencing data were obtained from a previous study8. A bioinformatic pipeline was developed to call somatic mutations robustly in all these species despite the variable quality of their genome assemblies (Methods). The distribution of variant allele fractions of the mutations detected in each crypt confirmed that crypts are clonal units in all species, enabling the study of somatic mutation rates and signatures (Extended Data Fig. 3).
We found substantial variation in the number of somatic single-base substitutions across species and across individuals within each species (Fig. 1b). For five species with samples from multiple individuals (dog, human, mouse, naked mole-rat and rat), linear regression confirmed a clear accumulation of somatic mutations with age (Fig. 1c, Extended Data Fig. 4, Supplementary Table 3). All linear regressions were also consistent with a non-significant intercept. This resembles observations in humans20 and suggests that the time required for a single stem cell to drift to fixation within a crypt is a small fraction of the lifespan of a species. This facilitates the estimation of somatic mutation rates across species by dividing the number of mutations in a crypt by the age of the individual (Supplementary Table 4). The number of somatic insertions and deletions (indels) was consistently lower than that of substitutions in all crypts (Fig. 1b), in agreement with previous findings in humans8.
Previous analyses in humans have shown that most somatic mutations in colorectal crypts accumulate neutrally, without clear evidence of negative selection against non-synonymous mutations and with a low frequency of positively selected cancer-driver mutations8. To study somatic selection in our data, we calculated the exome-wide ratio of non-synonymous to synonymous substitution rates (dN/dS) in each of the 12 species with available genome annotation. To do so and to detect genes under positive selection, while accounting for the effects of trinucleotide sequence context and mutation rate variation across genes, we used the dNdScv model38 (Methods). Although the limited number of coding somatic mutations observed in most species precluded an in-depth analysis of selection, exome-wide dN/dS ratios for somatic substitutions were not significantly different from unity in any species, in line with previous findings in humans8 (Extended Data Fig. 9). Gene-level analysis did not find genes under significant positive selection in any species, although larger studies are likely to identify rare cancer-driver mutations8.
Using whole-genome sequencing of 208 colorectal crypts from 56 individuals, we provide insights into the somatic mutational landscape of 16 mammalian species. Despite their different diets and life histories, we found considerable similarities in their mutational spectra. Three main mutational signatures explain the spectra across species, albeit with varying contributions and subtle variations in the profile of signature SBSB. These results suggest that, at least in the colorectal epithelium, a conserved set of mutational processes dominate somatic mutagenesis across mammals.
High-resolution scans were obtained from representative sections of each species. Example images are shown in Fig. 1a, Extended Data Fig. 2. Individual colorectal crypts were isolated from sections on polyethylene naphthalate (PEN) membrane slides by LCM with a Leica LMD7 microscope. Haematoxylin and eosin histology images were reviewed by a veterinary pathologist. For some samples we also cut a section of muscle tissue from below the colorectal epithelium of the section to use as a germline control for variant calling (Supplementary Table 2). Pre- and post-microdissection images of the tissue were recorded for each crypt and muscle sample taken. Each microdissection was collected in a separate well of a 96-well plate.
To validate our variant calling strategy, we used LCM to microdissect two sections from the same mouse colorectal crypt. We expected to detect a high fraction of shared somatic variants in these two sections, as their cells should be derived from the same ancestral epithelial stem cell. Both sections were submitted for independent library preparation, genome sequencing, variant calling and filtering using our pipeline. The majority of substitution variant calls (2,742 of 2,933, 93.5%) were shared between both sections (Supplementary Fig. 1c). By contrast, when comparing five separate crypts from a mouse, a maximum of two variants were shared between two crypts, and no variants were shared by three or more crypts (Supplementary Fig. 1d).
Trinucleotide-context mutational spectra of signature SBSB, as inferred independently from somatic mutations in crypts from four representative species (top to bottom): human, naked mole-rat, rat and rabbit (Methods). Signatures are shown in a human-genome-relative representation. Cosine similarities between each signature and the COSMIC human signatures SBS5 and SBS40 are provided.
Species information. For each of the (sub)species in the study, the provides: common name, scientific name, number of individuals in the study, number of colorectal crypts sequenced, range of individual ages, and source institution.
The gold standard for the diagnosis of Strongyloides infection is serial stool examination. However, traditional stool examinations are insensitive and require up to seven stool exams to reach a sensitivity of 100%. Specialized stool exams include Baermann concentration, Horadi-Mori filter paper culture, quantitative acetate concentration technique, and nutrient agar plate cultures. Duodenal aspirate is more sensitive than stool examination, and duodenal biopsy may reveal parasites in the gastric crypts, in the duodenal glands, or eosinophilic infiltration in the lamina propria. Frequently, larvae can be seen by a simple wet-mount in fluid from a bronchoalveolar lavage (BAL).