Establishment and Culture of Human Intestinal Organoids Derived from Adult Stem Cells

Human intestinal organoids derived from adult stem cells are miniature ex vivo versions of the human intestinal epithelium. Intestinal organoids are useful tools for the study of intestinal physiology as well as many disease conditions. These organoids present numerous advantages compared to immortalized cell lines, but working with them requires dedicated techniques. The protocols described in this article provide a basic guide to establishment and maintenance of human intestinal organoids derived from small intestine and colon biopsies. Additionally, this article provides an overview of several downstream applications of human intestinal organoids

Enteroendocrine cells switch hormone expression along the crypt-to-villus BMP signalling gradient.

Enteroendocrine cells (EECs) control a wide range of physiological processes linked to metabolism1. We show that EEC hormones are differentially expressed between crypts (for example, Glp1) and villi (for example, secretin). As demonstrated by single-cell mRNA sequencing using murine Lgr5+ cell-derived organoids, BMP4 signals alter the hormone expression profiles of individual EECs to resemble those found in the villus. Accordingly, BMP4 induces hormone switching of EECs migrating up the crypt-villus axis in vivo. Our findings imply that EEC lineages in the small intestine exhibit a more flexible hormone repertoire than previously proposed. We also describe a protocol to generate human EECs in organoids and demonstrate a similar regulation of hormone expression by BMP signalling. These findings establish alternative strategies to target EECs with therapeutically relevant hormone production through BMP modulation

Copy number alterations and allelic ratio in relation to recurrence of rectal cancer

BACKGROUND: In rectal cancer, total mesorectal excision surgery combined with preoperative (chemo)radiotherapy reduces local recurrence rates but does not improve overall patient survival, a result that may be due to the harmful side effects and/or co-morbidity of preoperative treatment. New biomarkers are needed to facilitate identification of rectal cancer patients at high risk for local recurrent disease. This would allow for preoperative (chemo)radiotherapy to be restricted to high-risk patients, thereby reducing overtreatment and allowing personalized treatment protocols. We analyzed genome-wide DNA copy number (CN) and allelic alterations in 112 tumors from preoperatively untreated rectal cancer patients. Sixty-six patients with local and/or distant recurrent disease were compared to matched controls without recurrence. Results were validated in a second cohort of tumors from 95 matched rectal cancer patients. Additionally, we performed a meta-analysis that included 42 studies reporting on CN alterations in colorectal cancer and compared results to our own data. RESULTS: The genomic profiles in our study were comparable to other rectal cancer studies. Results of the meta-analysis supported the hypothesis that colon cancer and rectal cancer may be distinct disease entities. In our discovery patient study cohort, allelic retention of chromosome 7 was significantly associated with local recurrent disease. Data from the validation cohort were supportive, albeit not statistically significant, of this finding. CONCLUSIONS: We showed that retention of heterozygosity on chromosome 7 may be associated with local recurrence in rectal cancer. Further research is warranted to elucidate the mechanisms and effect of retention of chromosome 7 on the development of local recurrent disease in rectal cancer. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12864-015-1550-0) contains supplementary material, which is available to authorized users

An organoid-derived bronchioalveolar model for SARS-CoV-2 infection of human alveolar type II-like cells

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19), which may result in acute respiratory distress syndrome (ARDS), multiorg

Druggable growth dependencies and tumor evolution analysis in patient-derived organoids of neuroendocrine neoplasms from multiple body sites

Neuroendocrine neoplasms (NENs) comprise well-differentiated neuroendocrine tumors (NETs) and poorly differentiated neuroendocrine carcinomas (NECs). Treatment options for patients with NENs are limited, in part due to lack of accurate models. We establish patient-derived tumor organoids (PDTOs) from pulmonary NETs and derive PDTOs from an understudied subtype of NEC, large cell neuroendocrine carcinoma (LCNEC), arising from multiple body sites. PDTOs maintain the gene expression patterns, intra-tumoral heterogeneity, and evolutionary processes of parental tumors. Through hypothesis-driven drug sensitivity analyses, we identify ASCL1 as a potential biomarker for response of LCNEC to treatment with BCL-2 inhibitors. Additionally, we discover a dependency on EGF in pulmonary NET PDTOs. Consistent with these findings, we find that, in an independent cohort, approximately 50% of pulmonary NETs express EGFR. This study identifies an actionable vulnerability for a subset of pulmonary NETs, emphasizing the utility of these PDTO models

Regulation and plasticity of intestinal stem cells during homeostasis and regeneration

The intestinal epithelium is the fastest renewing tissue in mammals and has a large flexibility to adapt to different types of damage. Lgr5+ crypt base columnar (CBC) cells act as stem cells during homeostasis and are essential during regeneration. Upon perturbation, the activity of CBCs is dynamically regulated to maintain homeostasis and multiple dedicated progenitor cell populations can reverse to the stem cell state upon damage, adding another layer of compensatory mechanisms to facilitate regeneration. Here, we review our current understanding of how intestinal stem and progenitor cells contribute to homeostasis and regeneration, and the different signaling pathways that regulate their behavior. Nutritional state and inflammation have been recently identified as upstream regulators of stem cell activity in the mammalian intestine, and we explore how these systemic signals can influence homeostasis and regeneration

Regulation and plasticity of intestinal stem cells during homeostasis and regeneration

The intestinal epithelium is the fastest renewing tissue in mammals and has a large flexibility to adapt to different types of damage. Lgr5+ crypt base columnar (CBC) cells act as stem cells during homeostasis and are essential during regeneration. Upon perturbation, the activity of CBCs is dynamically regulated to maintain homeostasis and multiple dedicated progenitor cell populations can reverse to the stem cell state upon damage, adding another layer of compensatory mechanisms to facilitate regeneration. Here, we review our current understanding of how intestinal stem and progenitor cells contribute to homeostasis and regeneration, and the different signaling pathways that regulate their behavior. Nutritional state and inflammation have been recently identified as upstream regulators of stem cell activity in the mammalian intestine, and we explore how these systemic signals can influence homeostasis and regeneration