RESUMEN
The stem cells that maintain and repair the postnatal skeleton remain undefined. One model suggests that perisinusoidal mesenchymal stem cells (MSCs) give rise to osteoblasts, chondrocytes, marrow stromal cells, and adipocytes, although the existence of these cells has not been proven through fate-mapping experiments. We demonstrate here that expression of the bone morphogenetic protein (BMP) antagonist gremlin 1 defines a population of osteochondroreticular (OCR) stem cells in the bone marrow. OCR stem cells self-renew and generate osteoblasts, chondrocytes, and reticular marrow stromal cells, but not adipocytes. OCR stem cells are concentrated within the metaphysis of long bones not in the perisinusoidal space and are needed for bone development, bone remodeling, and fracture repair. Grem1 expression also identifies intestinal reticular stem cells (iRSCs) that are cells of origin for the periepithelial intestinal mesenchymal sheath. Grem1 expression identifies distinct connective tissue stem cells in both the bone (OCR stem cells) and the intestine (iRSCs).
Asunto(s)
Huesos/citología , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Intestino Delgado/citología , Células Madre Mesenquimatosas/citología , Animales , Cartílago/metabolismo , Intestino Delgado/metabolismo , Células Madre Mesenquimatosas/metabolismo , Ratones , Ratones Endogámicos C57BLRESUMEN
OBJECTIVE: Long-standing chronic pancreatitis is an established risk factor for pancreatic ductal adenocarcinoma (PDAC). Interleukin-1ß (IL-1ß) has been associated in PDAC with shorter survival. We employed murine models to investigate the mechanisms by which IL-1ß and chronic pancreatitis might contribute to PDAC progression. DESIGN: We crossed LSL-Kras+/G12D;Pdx1-Cre (KC) mice with transgenic mice overexpressing IL-1ß to generate KC-IL1ß mice, and followed them longitudinally. We used pancreatic 3D in vitro culture to assess acinar-to-ductal metaplasia formation. Immune cells were analysed by flow cytometry and immunohistochemical staining. B lymphocytes were adoptively transferred or depleted in Kras-mutant mice. B-cell infiltration was analysed in human PDAC samples. RESULTS: KC-IL1ß mice developed PDAC with liver metastases. IL-1ß treatment increased Kras+/G12D pancreatic spheroid formation. CXCL13 expression and B lymphocyte infiltration were increased in KC-IL1ß pancreata. Adoptive transfer of B lymphocytes from KC-IL1ß mice promoted tumour formation, while depletion of B cells prevented tumour progression in KC-IL1ß mice. B cells isolated from KC-IL1ß mice had much higher expression of PD-L1, more regulatory B cells, impaired CD8+ T cell activity and promoted tumorigenesis. IL-35 was increased in the KC-IL1ß pancreata, and depletion of IL-35 decreased the number of PD-L1+ B cells. Finally, in human PDAC samples, patients with PDAC with higher B-cell infiltration within tumours showed significantly shorter survival. CONCLUSION: We show here that IL-1ß promotes tumorigenesis in part by inducing an expansion of immune-suppressive B cells. These findings point to the growing significance of B suppressor cells in pancreatic tumorigenesis.
Asunto(s)
Linfocitos B/inmunología , Carcinoma Ductal Pancreático/etiología , Tolerancia Inmunológica/inmunología , Neoplasias Pancreáticas/etiología , Pancreatitis/complicaciones , Animales , Linfocitos T CD8-positivos/inmunología , Carcinoma Ductal Pancreático/inmunología , Citometría de Flujo , Interleucina-1beta/efectos adversos , Ratones , Ratones Transgénicos , Neoplasias Pancreáticas/inmunología , Pancreatitis/etiología , Pancreatitis/inmunologíaRESUMEN
BACKGROUND & AIMS: The intestinal epithelium is maintained by long-lived intestinal stem cells (ISCs) that reside near the crypt base. Above the ISC zone, there are short-lived progenitors that normally give rise to lineage-specific differentiated cell types but can dedifferentiate into ISCs in certain circumstances. However, the role of epithelial dedifferentiation in cancer development has not been fully elucidated. METHODS: We performed studies with Bhlha15-CreERT, Lgr5-DTR-GFP, Apcflox/flox, LSL-Notch (IC), and R26-reporter strains of mice. Some mice were given diphtheria toxin to ablate Lgr5-positive cells, were irradiated, or were given 5-fluorouracil, hydroxyurea, doxorubicin, or dextran sodium sulfate to induce intestinal or colonic tissue injury. In intestinal tissues, we analyzed the fate of progeny that expressed Bhlha15. We used microarrays and reverse-transcription PCR to analyze gene expression patterns in healthy and injured intestinal tissues and in tumors. We analyzed gene expression patterns in human colorectal tumors using The Cancer Genome Atlas data set. RESULTS: Bhlha15 identified Paneth cells and short-lived secretory precursors (including pre-Paneth label-retaining cells) located just above the ISC zone in the intestinal epithelium. Bhlha15+ cells had no plasticity after loss of Lgr5-positive cells or irradiation. However, Bhlha15+ secretory precursors started to supply the enterocyte lineage after doxorubicin-induced epithelial injury in a Notch-dependent manner. Sustained activation of Notch converts Bhlha15+ secretory precursors to long-lived enterocyte progenitors. Administration of doxorubicin and expression of an activated form of Notch resulted in a gene expression pattern associated with enterocyte progenitors, whereas only sustained activation of Notch altered gene expression patterns in Bhlha15+ precursors toward those of ISCs. Bhlha15+ enterocyte progenitors with sustained activation of Notch formed intestinal tumors with serrated features in mice with disruption of Apc. In the colon, Bhlha15 marked secretory precursors that became stem-like, cancer-initiating cells after dextran sodium sulfate-induced injury, via activation of Src and YAP signaling. In analyses of human colorectal tumors, we associated activation of Notch with chromosome instability-type tumors with serrated features in the left colon. CONCLUSIONS: In mice, we found that short-lived precursors can undergo permanent reprogramming by activation of Notch and YAP signaling. These cells could mediate tumor formation in addition to traditional ISCs.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Neoplasias del Colon/genética , Enterocitos/patología , Mucosa Intestinal/metabolismo , Receptores Notch/metabolismo , Células Madre/metabolismo , Transcriptoma , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Antibióticos Antineoplásicos/farmacología , Antineoplásicos Hormonales/farmacología , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Antígeno CD24/metabolismo , Proteínas de Unión al Calcio , Proteínas de Ciclo Celular , Plasticidad de la Célula , Cromogranina A/genética , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Doxorrubicina/farmacología , Enterocitos/metabolismo , Expresión Génica , Perfilación de la Expresión Génica , Péptidos y Proteínas de Señalización Intercelular/genética , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/patología , Intestino Delgado/citología , Intestino Delgado/metabolismo , Ratones , Proteínas de Neoplasias/genética , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Proteínas Asociadas a Pancreatitis , Células de Paneth , Fosfoproteínas/metabolismo , Proteínas Proto-Oncogénicas c-kit/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Transducción de Señal , Células Madre/efectos de los fármacos , Células Madre/fisiología , Células Madre/efectos de la radiación , Tamoxifeno/farmacología , Proteínas Señalizadoras YAP , Familia-src Quinasas/metabolismoRESUMEN
OBJECTIVE: Progastrin is the incompletely cleaved precursor of gastrin that is secreted by G-cells in the gastric antrum. Both gastrin and progastrin bind to the CCK2 receptor (Cckbr or CCK2R) expressed on a subset of gastric epithelial cells. Little is known about how gastrin peptides and CCK2R regulate gastric stem cells and carcinogenesis. Interconversion among progenitors in the intestine is documented, but the mechanisms by which this occurs are poorly defined. DESIGN: We generated CCK2R-CreERT mice and performed inducible lineage tracing experiments. CCK2R+ antral cells and Lgr5+ antral stem cells were cultured in a three-dimensional in vitro system. We crossed progastrin-overexpressing mice with Lgr5-GFP-CreERT mice and examined the role of progastrin and CCK2R in Lgr5+ stem cells during MNU-induced carcinogenesis. RESULTS: Through lineage tracing experiments, we found that CCK2R defines antral stem cells at position +4, which overlapped with an Lgr5(neg or low) cell population but was distinct from typical antral Lgr5(high) stem cells. Treatment with progastrin interconverts Lgr5(neg or low) CCK2R+ cells into Lgr5(high) cells, increases CCK2R+ cell numbers and promotes gland fission and carcinogenesis in response to the chemical carcinogen MNU. Pharmacological inhibition or genetic ablation of CCK2R attenuated progastrin-dependent stem cell expansion and carcinogenesis. CONCLUSIONS: CCK2R labels +4 antral stem cells that can be activated and expanded by progastrin, thus identifying one hormonal trigger for gastric stem cell interconversion and a potential target for gastric cancer chemoprevention and therapy.
Asunto(s)
Carcinogénesis , Antro Pilórico/citología , Receptor de Colecistoquinina B/fisiología , Células Madre/fisiología , Animales , Células Cultivadas , Gastrinas/fisiología , Ratones , Precursores de Proteínas/fisiologíaRESUMEN
OBJECTIVE: To investigate the role of obesity-associated inflammation and immune modulation in gastric carcinogenesis during Helicobacter-induced chronic gastric inflammation. DESIGN: C57BL/6 male mice were infected with H felis and placed on a high-fat diet (45% calories from fat). Study animals were analysed for gastric and adipose pathology, inflammatory markers in serum, stomach and adipose tissue, and immune responses in blood, spleen, stomach and adipose tissue. RESULTS: H felis-induced gastric carcinogenesis was accelerated in diet-induced obese mice compared with lean controls. Obesity increased bone marrow-derived immature myeloid cells in blood and gastric tissue of H felis-infected mice. Obesity also led to elevations in CD4 T cells, IL-17A, granulocyte macrophage colony-stimulating factor, phosphorylated STAT3 and prosurvival gene expression in gastric tissue of H felis-infected mice. Conversely, in adipose tissue of obese mice, H felis infection increased macrophage accumulation and expression of IL-6, C-C motif ligand 7 (CCL7) and leptin. Finally, the combination of obesity and gastric inflammation synergistically increased serum proinflammatory cytokines, including IL-6. CONCLUSIONS: Here, we have established a model to study the molecular mechanism by which obesity predisposes individuals to gastric cancer. In H felis-infected mice, obesity increased proinflammatory immune responses and accelerated gastric carcinogenesis. Interestingly, gastric inflammation augmented obesity-induced adipose inflammation and production of adipose-derived factors in obese, but not lean, mice. Our findings suggest that obesity accelerates Helicobacter-associated gastric cancer through cytokine-mediated cross-talk between inflamed gastric and adipose tissues, augmenting immune responses at both tissue sites, and thereby contributing to a protumorigenic gastric microenvironment.
Asunto(s)
Gastritis/complicaciones , Infecciones por Helicobacter/complicaciones , Helicobacter felis , Células Progenitoras Mieloides/fisiología , Obesidad/complicaciones , Neoplasias Gástricas/etiología , Células Th17/fisiología , Animales , Biomarcadores/metabolismo , Movimiento Celular , Citocinas/metabolismo , Dieta Alta en Grasa , Citometría de Flujo , Gastritis/diagnóstico , Gastritis/metabolismo , Gastritis/microbiología , Infecciones por Helicobacter/inmunología , Inflamación/complicaciones , Inflamación/metabolismo , Inflamación/microbiología , Masculino , Ratones , Ratones Endogámicos C57BL , Obesidad/inmunología , Factor de Transcripción STAT3/metabolismo , Neoplasias Gástricas/inmunología , Neoplasias Gástricas/microbiologíaRESUMEN
BACKGROUND & AIMS: Progastrin stimulates colonic mucosal proliferation and carcinogenesis through the cholecystokinin 2 receptor (CCK2R)-partly by increasing the number of colonic progenitor cells. However, little is known about the mechanisms by which progastrin stimulates colonic cell proliferation. We investigated the role of bone morphogenetic proteins (BMPs) in progastrin induction of colonic cell proliferation via CCK2R. METHODS: We performed microarray analysis to compare changes in gene expression in the colonic mucosa of mice that express a human progastrin transgene, gastrin knockout mice, and C57BL/6 mice (controls); the effects of progastrin were also determined on in vitro colonic crypt cultures from cholecystokinin 2 receptor knockout and wild-type mice. Human colorectal and gastric cancer cells that expressed CCK2R were incubated with progastrin or Bmp2; levels of ß-arrestin 1 and 2 were knocked down using small interfering RNAs. Cells were analyzed for progastrin binding, proliferation, changes in gene expression, and symmetric cell division. RESULTS: The BMP pathway was down-regulated in the colons of human progastrin mice compared with controls. Progastrin suppressed transcription of Bmp2 through a pathway that required CCK2R and was mediated by ß-arrestin 1 and 2. In mouse colonic epithelial cells, down-regulation of Bmp2 led to decreased phosphorylation of Smads1/5/8 and suppression of inhibitor of DNA binding 4. In human gastric and colorectal cancer cell lines, CCK2R was necessary and sufficient for progastrin binding and induction of proliferation; these effects were blocked when cells were incubated with recombinant Bmp2. Incubation with progastrin increased the number of CD44(+), bromodeoxyuridine+, and NUMB(+) cells, indicating an increase in symmetric divisions of putative cancer stem cells. CONCLUSIONS: Progastrin stimulates proliferation in colons of mice and cultured human cells via CCK2R- and ß-arrestin 1 and 2-dependent suppression of Bmp2 signaling. This process promotes symmetric cell division.
Asunto(s)
Arrestinas/fisiología , Proteína Morfogenética Ósea 2/antagonistas & inhibidores , Proliferación Celular/efectos de los fármacos , Colon/efectos de los fármacos , Gastrinas/farmacología , Precursores de Proteínas/farmacología , Receptor de Colecistoquinina B/fisiología , Animales , Proteína Morfogenética Ósea 2/fisiología , Colon/citología , Ratones , Ratones Endogámicos C57BL , Transducción de Señal , Células Madre/efectos de los fármacos , beta-Arrestina 1 , beta-ArrestinasRESUMEN
While adult pancreatic stem cells are thought not to exist, it is now appreciated that the acinar compartment harbors progenitors, including tissue-repairing facultative progenitors (FPs). Here, we study a pancreatic acinar population marked by trefoil factor 2 (Tff2) expression. Long-term lineage tracing and single-cell RNA sequencing (scRNA-seq) analysis of Tff2-DTR-CreERT2-targeted cells defines a transit-amplifying progenitor (TAP) population that contributes to normal homeostasis. Following acute and chronic injury, Tff2+ cells, distinct from FPs, undergo depopulation but are eventually replenished. At baseline, oncogenic KrasG12D-targeted Tff2+ cells are resistant to PDAC initiation. However, KrasG12D activation in Tff2+ cells leads to survival and clonal expansion following pancreatitis and a cancer stem/progenitor cell-like state. Selective ablation of Tff2+ cells prior to KrasG12D activation in Mist1+ acinar or Dclk1+ FP cells results in enhanced tumorigenesis, which can be partially rescued by adenoviral Tff2 treatment. Together, Tff2 defines a pancreatic TAP population that protects against Kras-driven carcinogenesis.
Asunto(s)
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Neoplasias Pancreáticas/genética , Factor Trefoil-2/metabolismo , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Páncreas/metabolismo , Células Acinares/metabolismo , Carcinogénesis/genética , Carcinogénesis/metabolismoRESUMEN
The enteric neurotransmitter acetylcholine governs important intestinal epithelial secretory and immune functions through its actions on epithelial muscarinic Gq-coupled receptors such as M3R. Its role in the regulation of intestinal stem cell function and differentiation, however, has not been clarified. Here, we find that nonselective muscarinic receptor antagonism in mice as well as epithelial-specific ablation of M3R induces a selective expansion of DCLK1-positive tuft cells, suggesting a model of feedback inhibition. Cholinergic blockade reduces Lgr5-positive intestinal stem cell tracing and cell number. In contrast, Prox1-positive endocrine cells appear as primary sensors of cholinergic blockade inducing the expansion of tuft cells, which adopt an enteroendocrine phenotype and contribute to increased mucosal levels of acetylcholine. This compensatory mechanism is lost with acute irradiation injury, resulting in a paucity of tuft cells and acetylcholine production. Thus, enteroendocrine tuft cells appear essential to maintain epithelial homeostasis following modifications of the cholinergic intestinal niche.
Asunto(s)
Acetilcolina/metabolismo , Proteínas de Homeodominio/metabolismo , Mucosa Intestinal/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Animales , Quinasas Similares a Doblecortina , Células Enteroendocrinas/metabolismo , Femenino , Proteínas de Homeodominio/genética , Mucosa Intestinal/citología , Masculino , Ratones , Ratones Endogámicos C57BL , Neurotransmisores/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Proteínas Supresoras de Tumor/genéticaRESUMEN
TFF2 is a small, secreted protein with anti-inflammatory properties. We previously have shown that TFF2 gene delivery via adenovirus (Ad-Tff2) suppresses colon tumor growth in colitis associated cancer. Therefore, systemic administration of TFF2 peptide could potentially provide a similar benefit. Because TFF2 shows a poor pharmacokinetic, we sought to modify the TFF2 peptide in a manner that would lower its clearance rate but retain bioactivity. Given the absence of a sequence-based prediction of TFF2 functionality, we chose to genetically fuse the C-terminus of TFF2 with the carboxyl-terminal peptide of human chorionic gonadotropin ß subunit, and inserted into adenoviral vector that expresses Flag. The resulting Ad-Tff2-CTP-Flag construct translates into a TFF2 fused with two CTP and three Flag motifs. Administered Ad-Tff2-CTP-Flag decreased tumorigenesis and suppressed the expansion of myeloid cells in vivo. The fusion peptide TFF2-CTP-Flag delivered by adenovirus Ad-Tff2-CTP-Flag as well purified recombinant fusion TFF2-CTP-Flag was retained in the blood longer compared with wild-type TFF2 delivered by Ad-Tff2 or recombinant TFF2. Consistently, purified recombinant fusion TFF2-CTP-Flag suppressed expansion of myeloid cells by down-regulating cyclin D1 mRNA in vitro. Here, we demonstrate for the very first time the retained bioactivity and possible pharmacokinetic advantages of TFF2 with a modified C-terminus.
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Adenoviridae/genética , Neoplasias Colorrectales/terapia , Terapia Genética , Vectores Genéticos , Factor Trefoil-2/genética , Animales , Gonadotropina Coriónica/genética , Colitis/inducido químicamente , Colitis/complicaciones , Neoplasias Colorrectales/etiología , Ratones , Oligopéptidos/genética , Fragmentos de Péptidos/genética , TransgenesRESUMEN
Cancers of the gastrointestinal (GI) tract are among the most frequent and most lethal cancers worldwide. An important reason for this high mortality is that early disease is typically asymptomatic, and patients often present with advanced, incurable disease. Even in high-risk patients who routinely undergo endoscopic screening, lesions can be missed due to their small size or subtle appearance. Thus, current imaging approaches lack the sensitivity and specificity to accurately detect incipient GI tract cancers. Here we report our finding that a single dose of a high-sensitivity surface-enhanced resonance Raman scattering nanoparticle (SERRS-NP) enables reliable detection of precancerous GI lesions in animal models that closely mimic disease development in humans. Some of these animal models have not been used previously to evaluate imaging probes for early cancer detection. The studies were performed using a commercial Raman imaging system, a newly developed mouse Raman endoscope, and finally a clinically applicable Raman endoscope for larger animal studies. We show that this SERRS-NP-based approach enables robust detection of small, premalignant lesions in animal models that faithfully recapitulate human esophageal, gastric, and colorectal tumorigenesis. This method holds promise for much earlier detection of GI cancers than currently possible and could lead therefore to marked reduction of morbidity and mortality of these tumor types.
Asunto(s)
Endoscopía/métodos , Neoplasias Gastrointestinales/diagnóstico , Tracto Gastrointestinal/metabolismo , Nanopartículas/química , Espectrometría Raman/métodos , Animales , Modelos Animales de Enfermedad , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
Catecholamines stimulate epithelial proliferation, but the role of sympathetic nerve signaling in pancreatic ductal adenocarcinoma (PDAC) is poorly understood. Catecholamines promoted ADRB2-dependent PDAC development, nerve growth factor (NGF) secretion, and pancreatic nerve density. Pancreatic Ngf overexpression accelerated tumor development in LSL-Kras+/G12D;Pdx1-Cre (KC) mice. ADRB2 blockade together with gemcitabine reduced NGF expression and nerve density, and increased survival of LSL-Kras+/G12D;LSL-Trp53+/R172H;Pdx1-Cre (KPC) mice. Therapy with a Trk inhibitor together with gemcitabine also increased survival of KPC mice. Analysis of PDAC patient cohorts revealed a correlation between brain-derived neurotrophic factor (BDNF) expression, nerve density, and increased survival of patients on nonselective ß-blockers. These findings suggest that catecholamines drive a feedforward loop, whereby upregulation of neurotrophins increases sympathetic innervation and local norepinephrine accumulation.
Asunto(s)
Catecolaminas/farmacología , Factores de Crecimiento Nervioso/metabolismo , Neoplasias Pancreáticas/tratamiento farmacológico , Adrenérgicos/farmacología , Animales , Carcinoma in Situ/metabolismo , Línea Celular Tumoral , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacología , Ratones Transgénicos , Neoplasias Pancreáticas/patología , Gemcitabina , Neoplasias PancreáticasRESUMEN
In many solid tumors, parasympathetic input is provided by the vagus nerve, which has been shown to modulate tumor growth. However, whether cholinergic signaling directly regulates progression of pancreatic ductal adenocarcinoma (PDAC) has not been defined. Here, we found that subdiaphragmatic vagotomy in LSL-Kras +/G12D;Pdx1-Cre (KC) mice accelerated PDAC development, whereas treatment with the systemic muscarinic agonist bethanechol restored the normal KC phenotype, thereby suppressing the accelerated tumorigenesis caused by vagotomy. In LSL-Kras +/G12D;LSL-Trp53 +/R172H;Pdx1-Cre mice with established PDAC, bethanechol significantly extended survival. These effects were mediated in part through CHRM1, which inhibited downstream MAPK/EGFR and PI3K/AKT pathways in PDAC cells. Enhanced cholinergic signaling led to a suppression of the cancer stem cell (CSC) compartment, CD11b+ myeloid cells, TNFα levels, and metastatic growth in the liver. Therefore, these data suggest that cholinergic signaling directly and indirectly suppresses growth of PDAC cells, and therapies that stimulate muscarinic receptors may be useful in the treatment of PDAC.Significance: Subdiaphragmatic vagotomy or Chrm1 knockout accelerates pancreatic tumorigenesis, in part via expansion of the CSC compartment. Systemic administration of a muscarinic agonist suppresses tumorigenesis through MAPK and PI3K/AKT signaling, in early stages of tumor growth and in more advanced, metastatic disease. Therefore, CHRM1 may represent a potentially attractive therapeutic target. Cancer Discov; 8(11); 1458-73. ©2018 AACR. This article is highlighted in the In This Issue feature, p. 1333.
Asunto(s)
Carcinoma Ductal Pancreático/prevención & control , Transformación Celular Neoplásica/efectos de los fármacos , Colinérgicos/farmacología , Células Madre Neoplásicas/efectos de los fármacos , Neoplasias Pancreáticas/prevención & control , Receptor Muscarínico M1/fisiología , Animales , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patología , Transformación Celular Neoplásica/metabolismo , Transformación Celular Neoplásica/patología , Genes ras , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones Noqueados , Ratones SCID , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Transducción de Señal , Neoplasias PancreáticasRESUMEN
Overexpression of human progastrin increases colonic mucosal proliferation and colorectal cancer progression in mice. The G-protein coupled receptor 56 (GPR56) is known to regulate cell adhesion, migration, proliferation and stem cell biology, but its expression in the gut has not been studied. We hypothesized that the promotion of colorectal cancer by progastrin may be mediated in part through GPR56. Here, we found that GPR56 expresses in rare colonic crypt cells that lineage trace colonic glands consistent with GPR56 marking long-lived colonic stem-progenitor cells. GPR56 was upregulated in transgenic mice overexpressing human progastrin. While recombinant human progastrin promoted the growth and survival of wild-type colonic organoids in vitro, colonic organoids cultured from GPR56-/- mice were resistant to progastrin. We found that progastrin directly bound to, and increased the proliferation of, GPR56-expressing colon cancer cells in vitro, and proliferation was increased in cells that expressed both GPR56 and the cholecystokinin-2 receptor (CCK2R). In vivo, deletion of GPR56 in the mouse germline abrogated progastrin-dependent colonic mucosal proliferation and increased apoptosis. Loss of GPR56 also inhibited progastrin-dependent colonic crypt fission and colorectal carcinogenesis in the azoxymethane (AOM) mouse model of colorectal cancer. Overall, we found that progastrin binds to GPR56 expressing colonic stem cells, which in turn promotes their expansion, and that this GPR56-dependent pathway is an important driver and potential new target in colorectal carcinogenesis.
Asunto(s)
Neoplasias del Colon/metabolismo , Gastrinas/metabolismo , Precursores de Proteínas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Células Madre/metabolismo , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Carcinogénesis/efectos de los fármacos , Carcinogénesis/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Colon/metabolismo , Neoplasias del Colon/genética , Neoplasias del Colon/patología , Gastrinas/genética , Gastrinas/farmacología , Humanos , Mucosa Intestinal/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Unión Proteica , Precursores de Proteínas/genética , Precursores de Proteínas/farmacología , Receptor de Colecistoquinina B/genética , Receptor de Colecistoquinina B/metabolismo , Receptores Acoplados a Proteínas G/genéticaRESUMEN
The colorectal tumor microenvironment contains a diverse population of myeloid cells that are recruited and converted to immunosuppressive cells, thus facilitating tumor escape from immunoediting. We have identified a genetically and functionally distinct subset of dynamic bone marrow myeloid cells that are characterized by histidine decarboxylase (HDC) expression. Lineage tracing in Hdc-CreERT2;R26-LSL-tdTomato mice revealed that in homeostasis, there is a strong bias by HDC+ myeloid cells toward the CD11b+Ly6Ghi granulocytic lineage, which was accelerated during azoxymethane/dextran sodium sulfate (AOM/DSS)-induced colonic carcinogenesis. More importantly, HDC+ myeloid cells strongly promoted colonic tumorigenesis, and colon tumor progression was profoundly suppressed by diphtheria toxin A (DTA)-mediated depletion of HDC+ granulocytic myeloid cells. In addition, tumor infiltration by Foxp3+ regulatory T cells (Tregs) was markedly impaired following HDC+ myeloid cell depletion. We identified an HDC+ myeloid-derived Cxcl13/Cxcr5 axis that mediated Foxp3 expression and Treg proliferation. Ablation of HDC+ myeloid cells or disruption of the Cxcl13/Cxcr5 axis by gene knockdown impaired the production and recruitment of Tregs. Cxcl13 induction of Foxp3 expression in Tregs during tumorigenesis was associated with Stat3 phosphorylation. Overall, HDC+ granulocytic myeloid cells affect CD8+ T cells directly and indirectly through the modulation of Tregs and thus appear to play key roles in suppressing tumoricidal immunity.
RESUMEN
Within the gastrointestinal stem cell niche, nerves help to regulate both normal and neoplastic stem cell dynamics. Here, we reveal the mechanisms underlying the cancer-nerve partnership. We find that Dclk1+ tuft cells and nerves are the main sources of acetylcholine (ACh) within the gastric mucosa. Cholinergic stimulation of the gastric epithelium induced nerve growth factor (NGF) expression, and in turn NGF overexpression within gastric epithelium expanded enteric nerves and promoted carcinogenesis. Ablation of Dclk1+ cells or blockade of NGF/Trk signaling inhibited epithelial proliferation and tumorigenesis in an ACh muscarinic receptor-3 (M3R)-dependent manner, in part through suppression of yes-associated protein (YAP) function. This feedforward ACh-NGF axis activates the gastric cancer niche and offers a compelling target for tumor treatment and prevention.
Asunto(s)
Acetilcolina/fisiología , Factor de Crecimiento Nervioso/fisiología , Transducción de Señal/fisiología , Neoplasias Gástricas/etiología , Proteínas Adaptadoras Transductoras de Señales/fisiología , Animales , Proteínas de Ciclo Celular , Quinasas Similares a Doblecortina , Mucosa Gástrica/inervación , Ratones , Ratones Endogámicos C57BL , Fosfoproteínas/fisiología , Proteínas Serina-Treonina Quinasas/análisis , Receptor Muscarínico M3/fisiología , Proteínas Señalizadoras YAPRESUMEN
Mist1 was recently shown to identify a discrete population of stem cells within the isthmus of the oxyntic gland within the gastric corpus. Chief cells at the base of the gastric corpus also express Mist1. The relevance of Mist1 expression as a marker of specific cell populations within the antral glands of the distal stomach, however, is unknown. Using Mist1-CreERT mice, we revealed that Mist1+ antral cells, distinct from the Mist1+ population in the corpus, comprise long-lived progenitors that reside within the antral isthmus above Lgr5+ or CCK2R+ cells. Mist1+ antral progenitors can serve as an origin of antral tumors induced by loss of Apc or MNU treatment. Mist1+ antral progenitors, as well as other antral stem/progenitor population, express Cxcr4, and are located in close proximity to Cxcl12 (the Cxcr4 ligand)-expressing endothelium. During antral carcinogenesis, there is an expansion of Cxcr4+ epithelial cells as well as the Cxcl12+ perivascular niche. Deletion of Cxcl12 in endothelial cells or pharmacological blockade of Cxcr4 inhibits antral tumor growth. Cxcl12/Cxcr4 signaling may be a potential therapeutic target.
RESUMEN
Myeloid-biased hematopoietic stem cells (MB-HSCs) play critical roles in recovery from injury, but little is known about how they are regulated within the bone marrow niche. Here we describe an auto-/paracrine physiologic circuit that controls quiescence of MB-HSCs and hematopoietic progenitors marked by histidine decarboxylase (Hdc). Committed Hdc+ myeloid cells lie in close anatomical proximity to MB-HSCs and produce histamine, which activates the H2 receptor on MB-HSCs to promote their quiescence and self-renewal. Depleting histamine-producing cells enforces cell cycle entry, induces loss of serial transplant capacity, and sensitizes animals to chemotherapeutic injury. Increasing demand for myeloid cells via lipopolysaccharide (LPS) treatment specifically recruits MB-HSCs and progenitors into the cell cycle; cycling MB-HSCs fail to revert into quiescence in the absence of histamine feedback, leading to their depletion, while an H2 agonist protects MB-HSCs from depletion after sepsis. Thus, histamine couples lineage-specific physiological demands to intrinsically primed MB-HSCs to enforce homeostasis.
Asunto(s)
Médula Ósea/metabolismo , Células Madre Hematopoyéticas/metabolismo , Histamina/metabolismo , Células Mieloides/metabolismo , Animales , Médula Ósea/efectos de los fármacos , Trasplante de Médula Ósea , Citometría de Flujo , Células Madre Hematopoyéticas/efectos de los fármacos , Lipopolisacáridos/farmacología , Ratones , Células Mieloides/efectos de los fármacosRESUMEN
CD11b(+)Gr-1(+) myeloid-derived suppressor cells (MDSCs) expand in the spleen during cancer and promote progression through suppression of cytotoxic T cells. An anti-inflammatory reflex arc involving the vagus nerve and memory T cells is necessary for resolution of acute inflammation. Failure of this neural circuit could promote procarcinogenic inflammation and altered tumour immunity. Here we show that splenic TFF2, a secreted anti-inflammatory peptide, is released by vagally modulated memory T cells to suppress the expansion of MDSCs through CXCR4. Splenic denervation interrupts the anti-inflammatory neural arc, resulting in the expansion of MDSCs and colorectal cancer. Deletion of Tff2 recapitulates splenic denervation to promote carcinogenesis. Colorectal carcinogenesis could be suppressed through transgenic overexpression of TFF2, adenoviral transfer of TFF2 or transplantation of TFF2-expressing bone marrow. TFF2 is important to the anti-inflammatory reflex arc and plays an essential role in arresting MDSC proliferation. TFF2 offers a potential approach to prevent and to treat cancer.
Asunto(s)
Proliferación Celular/genética , Colitis/genética , Neoplasias Colorrectales/genética , Mucinas/genética , Proteínas Musculares/genética , Células Mieloides/inmunología , Péptidos/genética , Receptores CXCR4/inmunología , Bazo/inmunología , Linfocitos T Citotóxicos/inmunología , Nervio Vago , Traslado Adoptivo , Animales , Western Blotting , Trasplante de Médula Ósea , Colitis/inmunología , Colon/metabolismo , Colon/patología , Neoplasias Colorrectales/inmunología , Citocinas/inmunología , Desnervación , Progresión de la Enfermedad , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Inmunohistoquímica , Inflamación , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mucinas/inmunología , Mucinas/metabolismo , Proteínas Musculares/inmunología , Proteínas Musculares/metabolismo , Neoplasias/genética , Neoplasias/inmunología , Péptidos/inmunología , Péptidos/metabolismo , Permeabilidad , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Bazo/inervación , Subgrupos de Linfocitos T/inmunología , Factor Trefoil-2 , Vagotomía Troncal , Estimulación del Nervio VagoRESUMEN
WNT/ß-catenin signalling is crucial for intestinal homoeostasis. The intestinal epithelium and stroma are the major source of WNT ligands but their origin and role in intestinal stem cell (ISC) and epithelial repair remains unknown. Macrophages are a major constituent of the intestinal stroma. Here, we analyse the role of macrophage-derived WNT in intestinal repair in mice by inhibiting their release using a macrophage-restricted ablation of Porcupine, a gene essential for WNT synthesis. Such Porcn-depleted mice have normal intestinal morphology but are hypersensitive to radiation injury in the intestine compared with wild-type (WT) littermates. Porcn-null mice are rescued from radiation lethality by treatment with WT but not Porcn-null bone marrow macrophage-conditioned medium (CM). Depletion of extracellular vesicles (EV) from the macrophage CM removes WNT function and its ability to rescue ISCs from radiation lethality. Therefore macrophage-derived EV-packaged WNTs are essential for regenerative response of intestine against radiation.
Asunto(s)
Vesículas Extracelulares/metabolismo , Macrófagos/metabolismo , Traumatismos por Radiación/metabolismo , Células Madre/metabolismo , Proteínas Wnt/metabolismo , Vía de Señalización Wnt , Aciltransferasas/genética , Aciltransferasas/metabolismo , Animales , Células Cultivadas , Femenino , Mucosa Intestinal/citología , Mucosa Intestinal/metabolismo , Estimación de Kaplan-Meier , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones Endogámicos ICR , Ratones Noqueados , Ratones Transgénicos , Traumatismos por Radiación/genéticaRESUMEN
The existence of adult pancreatic progenitor cells has been debated. While some favor the concept of facultative progenitors involved in homeostasis and repair, neither a location nor markers for such cells have been defined. Using genetic lineage tracing, we show that Doublecortin-like kinase-1 (Dclk1) labels a rare population of long-lived, quiescent pancreatic cells. In vitro, Dclk1+ cells proliferate readily and sustain pancreatic organoid growth. In vivo, Dclk1+ cells are necessary for pancreatic regeneration following injury and chronic inflammation. Accordingly, their loss has detrimental effects after cerulein-induced pancreatitis. Expression of mutant Kras in Dclk1+ cells does not affect their quiescence or longevity. However, experimental pancreatitis converts Kras mutant Dclk1+ cells into potent cancer-initiating cells. As a potential effector of Kras, Dclk1 contributes functionally to the pathogenesis of pancreatic cancer. Taken together, these observations indicate that Dclk1 marks quiescent pancreatic progenitors that are candidates for the origin of pancreatic cancer.