RESUMO
According to conventional views, colon cancer originates from stem cells. However, inflammation, a key risk factor for colon cancer, has been shown to suppress intestinal stemness. Here, we used Paneth cells as a model to assess the capacity of differentiated lineages to trigger tumorigenesis in the context of inflammation in mice. Upon inflammation, Paneth cell-specific Apc mutations led to intestinal tumors reminiscent not only of those arising in patients with inflammatory bowel disease, but also of a larger fraction of human sporadic colon cancers. The latter is possibly because of the inflammatory consequences of western-style dietary habits, a major colon cancer risk factor. Machine learning methods designed to predict the cell-of-origin of cancer from patient-derived tumor samples confirmed that, in a substantial fraction of sporadic cases, the origins of colon cancer reside in secretory lineages and not in stem cells.
Assuntos
Carcinogênese , Linhagem da Célula , Neoplasias do Colo , Inflamação , Celulas de Paneth , Animais , Camundongos , Linhagem da Célula/genética , Celulas de Paneth/patologia , Humanos , Inflamação/genética , Inflamação/patologia , Carcinogênese/genética , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Mutação , Células-Tronco/patologia , Diferenciação Celular/genética , Transformação Celular Neoplásica/genética , Proteína da Polipose Adenomatosa do Colo/genética , Camundongos Endogâmicos C57BL , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/patologiaRESUMO
Paneth cells (PCs), a subset of intestinal epithelial cells (IECs) found at the base of small intestinal crypts, play an essential role in maintaining intestinal homeostasis. Altered PCs function is associated with diverse intestinal pathologies, including ileal Crohn's disease (CD). CD patients with ileal involvement have been previously demonstrated to display impairment in PCs and decreased levels of anti-microbial peptides. Although the immunosuppressive drug Azathioprine (AZA) is widely used in CD therapy, the impact of AZA on IEC differentiation remains largely elusive. In the present study, we hypothesized that the orally administered drug AZA also exerts its effect through modulation of the intestinal epithelium and specifically via modulation of PC function. AZA-treated CD patients exhibited an ileal upregulation of AMPs on both mRNA and protein levels compared to non-AZA treated patients. Upon in vitro AZA stimulation, intestinal epithelial cell line MODE-K exhibited heightened expression levels of PC marker in concert with diminished cell proliferation but boosted mitochondrial OXPHOS activity. Moreover, differentiation of IECs, including PCs differentiation, was boosted in AZA-treated murine small intestinal organoids and was associated with decreased D-glucose consumption and decreased growth rates. Of note, AZA treatment strongly decreased Lgr5 mRNA expression as well as Ki67 positive cells. Further, AZA restored dysregulated PCs associated with mitochondrial dysfunction. AZA-dependent inhibition of IEC proliferation is accompanied by boosted mitochondria function and IEC differentiation into PC.
Assuntos
Azatioprina , Diferenciação Celular , Doença de Crohn , Mucosa Intestinal , Celulas de Paneth , Doença de Crohn/tratamento farmacológico , Doença de Crohn/patologia , Doença de Crohn/metabolismo , Azatioprina/farmacologia , Celulas de Paneth/metabolismo , Celulas de Paneth/efeitos dos fármacos , Celulas de Paneth/patologia , Humanos , Diferenciação Celular/efeitos dos fármacos , Animais , Camundongos , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Feminino , Masculino , Íleo/efeitos dos fármacos , Íleo/metabolismo , Íleo/patologia , Adulto , Organoides/efeitos dos fármacos , Organoides/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Proliferação de Células/efeitos dos fármacos , Pessoa de Meia-Idade , Linhagem Celular , Índice de Gravidade de DoençaRESUMO
Epithelial barrier dysfunction and crypt destruction are hallmarks of inflammatory bowel disease (IBD). Intestinal stem cells (ISCs) residing in the crypts play a crucial role in the continuous self-renewal and rapid recovery of intestinal epithelial cells (IECs). However, how ISCs are dysregulated in IBD remains poorly understood. Here, we observe reduced DHX9 protein levels in IBD patients, and mice with conditional DHX9 depletion in the intestinal epithelium (Dhx9ΔIEC) exhibit an increased susceptibility to experimental colitis. Notably, Dhx9ΔIEC mice display a significant reduction in the numbers of ISCs and Paneth cells. Further investigation using ISC-specific or Paneth cell-specific Dhx9-deficient mice demonstrates the involvement of ISC-expressed DHX9 in maintaining epithelial homeostasis. Mechanistically, DHX9 deficiency leads to abnormal R-loop accumulation, resulting in genomic instability and the cGAS-STING-mediated inflammatory response, which together impair ISC function and contribute to the pathogenesis of IBD. Collectively, our findings highlight R-loop-mediated genomic instability in ISCs as a risk factor in IBD.
Assuntos
Doenças Inflamatórias Intestinais , Estruturas R-Loop , Animais , Humanos , Camundongos , RNA Helicases DEAD-box/metabolismo , Células Epiteliais/metabolismo , Homeostase , Doenças Inflamatórias Intestinais/patologia , Mucosa Intestinal/metabolismo , Proteínas de Neoplasias/metabolismo , Celulas de Paneth/metabolismo , Células-Tronco/metabolismoRESUMO
Paneth cell metaplasia (PCM) typically arises in pre-existing gastrointestinal (GI) diseases; however, the mechanistic pathway that induces metaplasia and whether PCM is initiated exclusively by disorders intrinsic to the GI tract is not well known. Here, we describe the development of PCM in a murine model of chronic myelogenous leukemia (CML) that is driven by an inducible bcr-abl oncogene. Mechanistically, CML induces a proinflammatory state within the GI tract that results in the production of epithelial-derived IL-33. The binding of IL-33 to the decoy receptor ST2 leads to IL-9 production by type 2 innate lymphoid cells (ILC2) which is directly responsible for the induction of PCM in the colon and tissue remodeling in the small intestines, characterized by goblet and tuft cell hyperplasia along with expansion of mucosal mast cells. Thus, we demonstrate that an extra-intestinal disease can trigger an ILC2/IL-9 immune circuit, which induces PCM and regulates epithelial cell fate decisions in the GI tract.
Assuntos
Leucemia Mielogênica Crônica BCR-ABL Positiva , Celulas de Paneth , Animais , Camundongos , Interleucina-9/genética , Imunidade Inata , Interleucina-33/genética , Linfócitos , Intestino Delgado , MetaplasiaRESUMO
BACKGROUND: Intestinal epithelial cell (IEC) mitochondrial dysfunction involvement in inflammatory bowel diseases (IBD), including Crohn's disease affecting the small intestine, is emerging in recent studies. As the interface between the self and the gut microbiota, IECs serve as hubs of bidirectional cross-talk between host and luminal microbiota. However, the role of mitochondrial-microbiota interaction in the ileum is largely unexplored. Prohibitin 1 (PHB1), a chaperone protein of the inner mitochondrial membrane required for optimal electron transport chain function, is decreased during IBD. We previously demonstrated that mice deficient in PHB1 specifically in IECs (Phb1i∆IEC) exhibited mitochondrial impairment, Paneth cell defects, gut microbiota dysbiosis, and spontaneous inflammation in the ileum (ileitis). Mice deficient in PHB1 in Paneth cells (epithelial secretory cells of the small intestine; Phb1∆PC) also exhibited mitochondrial impairment, Paneth cell defects, and spontaneous ileitis. Here, we determined whether this phenotype is driven by Phb1 deficiency-associated ileal microbiota alterations or direct effects of loss of PHB1 in host IECs. RESULTS: Depletion of gut microbiota by broad-spectrum antibiotic treatment in Phb1∆PC or Phb1i∆IEC mice revealed a necessary role of microbiota to cause ileitis. Using germ-free mice colonized with ileal microbiota from Phb1-deficient mice, we show that this microbiota could not independently induce ileitis without host mitochondrial dysfunction. The luminal microbiota phenotype of Phb1i∆IEC mice included a loss of the short-chain fatty acid butyrate. Supplementation of butyrate in Phb1-deficient mice ameliorated Paneth cell abnormalities and ileitis. Phb1-deficient ileal enteroid models suggest deleterious epithelial-intrinsic responses to ileal microbiota that were protected by butyrate. CONCLUSIONS: These results suggest a mutual and essential reinforcing interplay of gut microbiota and host IEC, including Paneth cell, mitochondrial health in influencing ileitis. Restoration of butyrate is a potential therapeutic option in Crohn's disease patients harboring epithelial cell mitochondrial dysfunction. Video Abstract.
Assuntos
Doença de Crohn , Microbioma Gastrointestinal , Ileíte , Doenças Inflamatórias Intestinais , Humanos , Animais , Camundongos , Ileíte/metabolismo , Inflamação/metabolismo , Doenças Inflamatórias Intestinais/metabolismo , Celulas de Paneth , Butiratos/metabolismo , Mitocôndrias/metabolismo , Mucosa Intestinal/metabolismoRESUMO
Rapid self-renewal of the intestinal epithelium requires the activity of intestinal stem cells (ISCs) that are intermingled with Paneth cells (PCs) at the crypt base. PCs provide multiple secreted and surface-bound niche signals and play an important role in the regulation of ISC proliferation. Here, we show that control of PC function by RNA-binding protein HuR via mitochondria affects intestinal mucosal growth by altering ISC activity. Targeted deletion of HuR in mice disrupted PC gene expression profiles, reduced PC-derived niche factors, and impaired ISC function, leading to inhibited renewal of the intestinal epithelium. Human intestinal mucosa from patients with critical surgical disorders exhibited decreased levels of tissue HuR and PC/ISC niche dysfunction, along with disrupted mucosal growth. HuR deletion led to mitochondrial impairment by decreasing the levels of several mitochondrial-associated proteins including prohibitin 1 (PHB1) in the intestinal epithelium, whereas HuR enhanced PHB1 expression by preventing microRNA-195 binding to the Phb1 mRNA. These results indicate that HuR is essential for maintaining the integrity of the PC/ISC niche and highlight a novel role for a defective PC/ISC niche in the pathogenesis of intestinal mucosa atrophy.
Assuntos
Proteína Semelhante a ELAV 1 , MicroRNAs , Mucosa , Celulas de Paneth , Animais , Humanos , Camundongos , Transporte Biológico , Fenômenos Fisiológicos Celulares , Mucosa Intestinal , MicroRNAs/genética , Proteínas Mitocondriais , Células-Tronco , Proteína Semelhante a ELAV 1/genéticaRESUMO
Introduction: The mechanism underlying radiation-induced gut microbiota dysbiosis is undefined. This study examined the effect of radiation on the intestinal Paneth cell α-defensin expression and its impact on microbiota composition and mucosal tissue injury and evaluated the radio-mitigative effect of human α-defensin 5 (HD5). Methods: Adult mice were subjected to total body irradiation, and Paneth cell α-defensin expression was evaluated by measuring α-defensin mRNA by RT-PCR and α-defensin peptide levels by mass spectrometry. Vascular-to-luminal flux of FITC-inulin was measured to evaluate intestinal mucosal permeability and endotoxemia by measuring plasma lipopolysaccharide. HD5 was administered in a liquid diet 24 hours before or after irradiation. Gut microbiota was analyzed by 16S rRNA sequencing. Intestinal epithelial junctions were analyzed by immunofluorescence confocal microscopy and mucosal inflammatory response by cytokine expression. Systemic inflammation was evaluated by measuring plasma cytokine levels. Results: Ionizing radiation reduced the Paneth cell α-defensin expression and depleted α-defensin peptides in the intestinal lumen. α-Defensin down-regulation was associated with the time-dependent alteration of gut microbiota composition, increased gut permeability, and endotoxemia. Administration of human α-defensin 5 (HD5) in the diet 24 hours before irradiation (prophylactic) significantly blocked radiation-induced gut microbiota dysbiosis, disruption of intestinal epithelial tight junction and adherens junction, mucosal barrier dysfunction, and mucosal inflammatory response. HD5, administered 24 hours after irradiation (treatment), reversed radiation-induced microbiota dysbiosis, tight junction and adherens junction disruption, and barrier dysfunction. Furthermore, HD5 treatment also prevents and reverses radiation-induced endotoxemia and systemic inflammation. Conclusion: These data demonstrate that radiation induces Paneth cell dysfunction in the intestine, and HD5 feeding prevents and mitigates radiation-induced intestinal mucosal injury, endotoxemia, and systemic inflammation.
Assuntos
Endotoxemia , Lesões por Radiação , alfa-Defensinas , Humanos , Adulto , Animais , Camundongos , Celulas de Paneth , Disbiose , Endotoxemia/etiologia , RNA Ribossômico 16S , Lesões por Radiação/etiologia , Citocinas , InflamaçãoRESUMO
As a family of cationic host defense peptides, defensins are mainly synthesized by Paneth cells, neutrophils, and epithelial cells, contributing to host defense. Their biological functions in innate immunity, as well as their structure and activity relationships, along with their mechanisms of action and therapeutic potential, have been of great interest in recent years. To highlight the key research into the role of defensins in human and animal health, we first describe their research history, structural features, evolution, and antimicrobial mechanisms. Next, we cover the role of defensins in immune homeostasis, chemotaxis, mucosal barrier function, gut microbiota regulation, intestinal development and regulation of cell death. Further, we discuss their clinical relevance and therapeutic potential in various diseases, including infectious disease, inflammatory bowel disease, diabetes and obesity, chronic inflammatory lung disease, periodontitis and cancer. Finally, we summarize the current knowledge regarding the nutrient-dependent regulation of defensins, including fatty acids, amino acids, microelements, plant extracts, and probiotics, while considering the clinical application of such regulation. Together, the review summarizes the various biological functions, mechanism of actions and potential clinical significance of defensins, along with the challenges in developing defensins-based therapy, thus providing crucial insights into their biology and potential clinical utility.
Assuntos
Doenças Inflamatórias Intestinais , Celulas de Paneth , Animais , Humanos , Celulas de Paneth/metabolismo , Doenças Inflamatórias Intestinais/metabolismo , Defensinas/genética , Defensinas/metabolismoRESUMO
Crohn's disease (CD) is an inflammatory bowel disease characterized by immune-mediated flares affecting any region of the intestine alternating with remission periods. In CD, the ileum is frequently affected and about one third of patients presents with a pure ileal type. Moreover, the ileal type of CD presents epidemiological specificities like a younger age at onset and often a strong link with smoking and genetic susceptibility genes. Most of these genes are associated with Paneth cell dysfunction, a cell type found in the intestinal crypts of the ileum. Besides, a Western-type diet is associated in epidemiological studies with CD onset and increasing evidence shows that diet can modulate the composition of bile acids and gut microbiota, which in turn modulates the susceptibility of the ileum to inflammation. Thus, the interplay between environmental factors and the histological and anatomical features of the ileum is thought to explain the specific transcriptome profile observed in CD ileitis. Indeed, both immune response and cellular healing processes harbour differences between ileal and non-ileal CD. Taken together, these findings advocate for a dedicated therapeutic approach to managing ileal CD. Currently, interventional pharmacological studies have failed to clearly demonstrate distinct response profiles according to disease site. However, the high rate of stricturing disease in ileal CD requires the identification of new therapeutic targets to significantly change the natural history of this debilitating disease.
Assuntos
Doença de Crohn , Doenças do Íleo , Ileíte , Humanos , Doença de Crohn/epidemiologia , Doença de Crohn/genética , Doença de Crohn/terapia , Íleo/patologia , Ileíte/patologia , Inflamação/patologia , Celulas de Paneth/metabolismo , Celulas de Paneth/patologia , Doenças do Íleo/patologiaRESUMO
Primary cilia are sensory antennae located at the cell surface which mediate a variety of extracellular signals involved in development, tissue homeostasis, stem cells and cancer. Primary cilia are found in an extensive array of vertebrae cells but can only be generated when cells become quiescent. The small intestinal epithelium is a rapidly self-renewing tissue organized into a functional unit called the crypt-villus axis, containing progenitor and differentiated cells, respectively. Terminally differentiated villus cells are notoriously devoid of primary cilia. We sought to determine if intestinal crypts contain a quiescent cell population that could be identified by the presence of primary cilia. Here we show that primary cilia are detected in a subset of cells located deep in the crypts slightly above a Paneth cell population. Using a normal epithelial proliferative crypt cell model, we show that primary cilia assembly and activity correlate with a quiescent state. These results provide further evidence for the existence of a quiescent cell population in the human small intestine and suggest the potential for new modes of regulation in stem cell dynamics.
Assuntos
Cílios , Intestino Delgado , Humanos , Duodeno , Divisão Celular , Celulas de PanethRESUMO
BACKGROUND: Amino acid transporters play an important role in supplying nutrition to cells and are associated with cell proliferation. L-type amino acid transporter 1 (LAT1) is highly expressed in many types of cancers and promotes tumor growth; however, how LAT1 affects tumor development is not fully understood. METHODS: To investigate the role of LAT1 in intestinal tumorigenesis, mice carrying LAT1 floxed alleles that also expressed Cre recombinase from the promoter of gene encoding Villin were crossed to an ApcMin/+ background (LAT1fl/fl; vil-cre; ApcMin/+), which were subject to analysis; organoids derived from those mice were also analyzed. RESULTS: This study showed that LAT1 was constitutively expressed in normal crypt base cells, and its conditional deletion in the intestinal epithelium resulted in fewer Paneth cells. LAT1 deletion reduced tumor size and number in the small intestine of ApcMin/+ mice. Organoids derived from LAT1-deleted ApcMin/+ intestinal crypts displayed fewer spherical organoids with reduced Wnt/ß-catenin target gene expression, suggesting a low tumor-initiation capacity. Wnt3 expression was decreased in the absence of LAT1 in the intestinal epithelium, suggesting that loss of Paneth cells due to LAT1 deficiency reduced the risk of tumor initiation by decreasing Wnt3 production. CONCLUSIONS: LAT1 affects intestinal tumor development in a cell-extrinsic manner through reduced Wnt3 expression in Paneth cells. Our findings may partly explain how nutrient availability can affect the risk of tumor development in the intestines.
Assuntos
Proteína da Polipose Adenomatosa do Colo , Sistema y+L de Transporte de Aminoácidos , Neoplasias Intestinais , Celulas de Paneth , Animais , Camundongos , Transformação Celular Neoplásica/genética , Mucosa Intestinal/patologia , Neoplasias Intestinais/metabolismo , Intestino Delgado/patologia , Intestinos , Celulas de Paneth/metabolismo , Celulas de Paneth/patologia , Proteína da Polipose Adenomatosa do Colo/metabolismo , Sistema y+L de Transporte de Aminoácidos/metabolismoAssuntos
Doença de Crohn , Humanos , Doença de Crohn/diagnóstico , Doença de Crohn/tratamento farmacológico , Doença de Crohn/metabolismo , Celulas de Paneth/metabolismo , Fatores de Proteção , Autofagia , Transporte Biológico , Proteínas Relacionadas à Autofagia/genética , Proteínas Relacionadas à Autofagia/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Reguladoras de Apoptose/metabolismoRESUMO
Intestinal homeostasis depends on interactions between the intestinal epithelium, the immune system and the microbiota. Because of these complicated connections, there are many problems that need to be solved. Current research has indicated that genes targeted by Wnt signaling are responsible for controlling intestinal stem cell fate and for modulating intestinal homeostasis. Our data show that loss of frizzled 7 (Fzd7), an important element in Wnt signaling, interrupts the differentiation of mouse intestinal stem cells into absorptive progenitors instead of secretory progenitors (precursors of goblet and Paneth cells). The alteration in canonical Wnt and Notch signaling pathways interrupts epithelial homeostasis, resulting in a decrease in physical protection in the intestine. Several phenotypes in our Fzd7-deleted model were similar to the features of enterocolitis, such as shortened intestines, decreased numbers of goblet cells and Paneth cells, and severe inflammation. Additionally, loss of Fzd7 exacerbated the defects in a chemical-induced colitis model and could initiate tumorigenesis. These findings may provide important information for the discovery of efficient therapeutic methods to treat enterocolitis and related cancers in the intestines.
Assuntos
Enterocolite , Celulas de Paneth , Animais , Camundongos , Diferenciação Celular , Enterocolite/metabolismo , Células Caliciformes/metabolismo , Homeostase , Mucosa Intestinal/metabolismo , Intestinos , Via de Sinalização WntRESUMO
Adult stem cells are responsible for homoeostasis and regeneration of epithelial tissues. Stem cell function is regulated by both cell autonomous mechanisms as well as the niche. Deregulated stem cell function contributes to diseases such as cancer. Epithelial organoid cultures generated from tissue-resident adult stem cells have allowed unprecedented insights into the biology of epithelial tissues. The subsequent adaptation of organoid technology enabled the modelling of the communication of stem cells with their cellular and non-cellular niche as well as diseases. Starting from its first model described in 2009, the murine small intestinal organoid, we discuss here how epithelial organoid cultures have been become a prime in vitro research tool for cell and developmental biology, bioengineering, and biomedicine in the last decade.
Assuntos
Células-Tronco Adultas , Organoides , Animais , Camundongos , Células-Tronco , Epitélio , Celulas de Paneth , Células EpiteliaisRESUMO
Menopausal women often face long-term estrogen treatment. G protein-coupled estrogen receptor (GPER) expressed in intestinal crypt was activated by estrogen therapy, but it was unclear whether chronic GPER activation during menopause had an effect on intestinal stem cells (ISCs). We tested the effect of chronic GPER activation on ISCs of ovariectomized (OVX) mice by injection of the selective GPER agonist G-1 for 28 days, or G-1 stimulation of organoids derived from crypts of OVX mice. G-1 up-regulated crypt depth, the number of Ki67+, bromodeoxyuridine+ cells and Olfm4+ ISCs, and the expression of ISCs marker genes (Lgr5, Olfm4 and Axin2). G-1 administration promoted organoid growth, increased the number of EdU+ cells per organoid and protein expression of Cyclin D1 and cyclin B1 in organoids. After G-1 treatment in vivo or in vitro, Paneth cell-derived Wnt3, Wnt3 effector ß-catenin and Wnt target genes c-Myc and Cyclin D1 increased in ileum or organoids. Once blocking the secretion of Wnt3 from Paneth cells, the effects of G-1 on organoids growth, ISCs marker genes and Wnt/ß-catenin signaling were abolished. G-1 did not affect the number of Paneth cells in ex vivo organoids, while activated Mmp7/cryptdin program in Paneth cells, promoted their maturation, and increased the expression of lysozyme protein. G-1 pretreatment in OVX mice inhibited radiation-induced ISCs proliferation injury and enhanced the resistance of mice to intestinal injury. In conclusion, chronic GPER activation prompted the Wnt3 synthesis in Paneth cells, thus increased the proliferation of ISCs via activation of Wnt3/ß-catenin signaling in OVX mice.
Assuntos
Ciclina D1 , Celulas de Paneth , Camundongos , Feminino , Animais , Celulas de Paneth/metabolismo , Ciclina D1/metabolismo , beta Catenina/metabolismo , Íleo/metabolismo , Células-Tronco , Via de Sinalização Wnt , Proliferação de Células , Estrogênios/farmacologia , Estrogênios/metabolismo , Mucosa Intestinal/metabolismo , Proteína Wnt3/metabolismo , Proteína Wnt3/farmacologiaRESUMO
Intestinal stem cell maturation and development coincide with gut microbiota exposure after birth. Here, we investigated how early life microbial exposure, and disruption of this process, impacts the intestinal stem cell niche and development. Single-cell transcriptional analysis revealed impaired stem cell differentiation into Paneth cells and macrophage specification upon antibiotic treatment in early life. Mouse genetic and organoid co-culture experiments demonstrated that a CD206+ subset of intestinal macrophages secreted Wnt ligands, which maintained the mesenchymal niche cells important for Paneth cell differentiation. Antibiotics and reduced numbers of Paneth cells are associated with the deadly infant disease, necrotizing enterocolitis (NEC). We showed that colonization with Lactobacillus or transfer of CD206+ macrophages promoted Paneth cell differentiation and reduced NEC severity. Together, our work defines the gut microbiota-mediated regulation of stem cell niches during early postnatal development.
Assuntos
Enterocolite Necrosante , Microbioma Gastrointestinal , Camundongos , Animais , Celulas de Paneth/fisiologia , Diferenciação Celular/fisiologia , MacrófagosRESUMO
Calorie restriction has been recently shown to increase intestinal stem cell competition and to reduce mutation fixation in young mice. However, the impact of aging on this process is unknown. By employing Confetti reporter mice, here we show that, unexpectedly, old mice have more intestinal stem cell (ISC) competition than young mice. Moreover, differently from what observed in young mice, calorie restriction, when applied at late-life, decreases this process. Importantly, we also observed a strong correlation between the ISC competition and Paneth cell number. In vivo analysis and in vitro organoid experiments indicated that Paneth cells play a major role in driving intestinal stem cell competition and crypt clonality. Taken together, our results provide evidence that increasing the number of Paneth cells can increase the number of competitive ISCs, representing a valuable therapeutic target to delay fixation of mutated intestinal stem cells.
Assuntos
Restrição Calórica , Celulas de Paneth , Camundongos , Animais , Competição entre as Células , Intestinos , Células-Tronco , Mucosa IntestinalRESUMO
Superoxide dismutase 1 (SOD1) modulates intestinal barrier integrity and intestinal homeostasis as an antioxidant enzyme. Intestinal homeostasis is maintained by the intestinal stem cells (ISCs). However, whether and how SOD1 regulates ISCs is unknown. In this study, we established intestinal organoids from tamoxifen-inducible intestinal epithelial cell-specific Sod1 knockout (Sod1f/f; Vil-creERT2) mice. We found that loss of Sod1 in organoids suppressed the proliferation and survival of cells and Lgr5 gene expression. SOD1 is known for nearly half a century for its canonical role as an antioxidant enzyme. We identified its enzyme-independent function in ISC: inhibition of SOD1 enzymatic activity had no impact on organoid growth, and enzymatically inactive Sod1 mutants could completely rescue the growth defects of Sod1 deficient organoids, suggesting that SOD1-mediated ISC growth is independent of its enzymatic activity. Moreover, Sod1 deficiency did not affect the ROS levels of the organoid, but induced the elevated WNT signaling and excessive Paneth cell differentiation, which mediates the occurrence of growth defects in Sod1 deficient organoids. In vivo, epithelial Sod1 loss induced a higher incidence of apoptosis in the stem cell regions and increased Paneth cell numbers, accompanied by enhanced expression of EGFR ligand Epiregulin (EREG) in the stromal tissue, which may compensate for Sod1 loss and maintain intestinal structure in vivo. Totally, our results show a novel enzyme-independent function of SOD1 in ISC growth under homeostasis.
Assuntos
Neoplasias Intestinais , Superóxido Dismutase , Camundongos , Animais , Superóxido Dismutase-1/genética , Superóxido Dismutase-1/metabolismo , Epirregulina/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Ligantes , Antioxidantes/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Células-Tronco/metabolismo , Celulas de Paneth/metabolismo , Organoides/metabolismo , Neoplasias Intestinais/metabolismo , Receptores ErbB/metabolismo , Tamoxifeno/farmacologia , Mucosa Intestinal/metabolismo , Proliferação de CélulasRESUMO
Loss of Paneth cells and their antimicrobial granules compromises the intestinal epithelial barrier and is associated with Crohn's disease, a major type of inflammatory bowel disease1-7. Non-classical lymphoid cells, broadly referred to as intraepithelial lymphocytes (IELs), intercalate the intestinal epithelium8,9. This anatomical position has implicated them as first-line defenders in resistance to infections, but their role in inflammatory disease pathogenesis requires clarification. The identification of mediators that coordinate crosstalk between specific IEL and epithelial subsets could provide insight into intestinal barrier mechanisms in health and disease. Here we show that the subset of IELs that express γ and δ T cell receptor subunits (γδ IELs) promotes the viability of Paneth cells deficient in the Crohn's disease susceptibility gene ATG16L1. Using an ex vivo lymphocyte-epithelium co-culture system, we identified apoptosis inhibitor 5 (API5) as a Paneth cell-protective factor secreted by γδ IELs. In the Atg16l1-mutant mouse model, viral infection induced a loss of Paneth cells and enhanced susceptibility to intestinal injury by inhibiting the secretion of API5 from γδ IELs. Therapeutic administration of recombinant API5 protected Paneth cells in vivo in mice and ex vivo in human organoids with the ATG16L1 risk allele. Thus, we identify API5 as a protective γδ IEL effector that masks genetic susceptibility to Paneth cell death.
Assuntos
Proteínas Reguladoras de Apoptose , Doença de Crohn , Predisposição Genética para Doença , Linfócitos Intraepiteliais , Proteínas Nucleares , Celulas de Paneth , Animais , Humanos , Camundongos , Proteínas Reguladoras de Apoptose/metabolismo , Morte Celular , Doença de Crohn/genética , Doença de Crohn/metabolismo , Doença de Crohn/patologia , Predisposição Genética para Doença/genética , Mucosa Intestinal/patologia , Proteínas Nucleares/metabolismo , Celulas de Paneth/patologia , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos Intraepiteliais/imunologia , Linfócitos Intraepiteliais/metabolismo , Sobrevivência Celular , Organoides , AlelosRESUMO
PURPOSE OF REVIEW: Paneth cells are specialized, secretory epithelial cells located in the small intestine. Although their existence was first described in 1872, their precise role in the gut remained unclear for over a century. Over the past few decades, elegant studies have shown Paneth cells play a key role enhancing gut barrier function, as niche cells for the intestinal stem cell compartment and via secreting antimicrobial peptides to establish an antimicrobial barrier at the epithelial surface. This review describes what is known about Paneth cell biology from human and animal studies with a focus on their putative role in clinical gastrointestinal disease. RECENT FINDINGS: Recent work has demonstrated important associations of dysfunctional Paneth cells with several gastrointestinal disorders. These include Crohn's disease, enteric infections, graft-versus-host disease, necrotizing enterocolitis, and environmental enteric dysfunction. Ongoing studies are examining precisely how Paneth cell biology is altered in these various disease states. SUMMARY: By understanding the mechanisms of Paneth cell regulation - and how these processes go awry in specific gastrointestinal diseases - we set the stage for using Paneth cells as biomarkers for disease progression and developing novel therapeutics that augment Paneth cell function to treat a spectrum of gastrointestinal disorders.