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1.
Int J Mol Sci ; 24(23)2023 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-38069171

RESUMO

Micro-environmental factors, including stromal and immune cells, cytokines, and circulating hormones are well recognized to determine cancer progression. Melanoma cell growth was recently shown to be suppressed by cholecystokinin/gastrin (CCK) receptor antagonists, and our preliminary data suggested that melanoma patients with Helicobacter gastritis (which is associated with elevated serum gastrin) might have an increased risk of cancer progression. Therefore, in the present study, we examined how gastrin may act on melanoma cells. In 89 melanoma patients, we found a statistically significant association between circulating gastrin concentrations and melanoma thickness and metastasis, which are known risk factors of melanoma progression and prognosis. Immunocytochemistry using a validated antibody confirmed weak to moderate CCK2R expression in both primary malignant melanoma cells and the melanoma cell lines SK-MEL-2 and G361. Furthermore, among the 219 tumors in the Skin Cutaneous Melanoma TCGA Pan-Cancer dataset showing gastrin receptor (CCKBR) expression, significantly higher CCKBR mRNA levels were linked to stage III-IV than stage I-II melanomas. In both cell lines, gastrin increased intracellular calcium levels and stimulated cell migration and invasion through mechanisms inhibited by a CCK2 receptor antagonist. Proteomic studies identified increased MMP-2 and reduced TIMP-3 levels in response to gastrin that were likely to contribute to the increased migration of both cell lines. However, the effects of gastrin on tumor cell invasion were relatively weak in the presence of the extracellular matrix. Nevertheless, dermal fibroblasts/myofibroblasts, known also to express CCK2R, increased gastrin-induced cancer cell invasion. Our data suggest that in a subset of melanoma patients, an elevated serum gastrin concentration is a risk factor for melanoma tumor progression, and that gastrin may act on both melanoma and adjacent stromal cells through CCK2 receptors to promote mechanisms of tumor migration and invasion.


Assuntos
Melanoma , Neoplasias Cutâneas , Humanos , Melanoma/metabolismo , Gastrinas/farmacologia , Gastrinas/metabolismo , Proteômica , Receptores da Colecistocinina , Receptor de Colecistocinina B/genética , Receptor de Colecistocinina B/metabolismo
2.
Front Endocrinol (Lausanne) ; 12: 741887, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34867785

RESUMO

Background: Hypergastrinaemia occasionally indicates the presence of a gastrinoma. However it is much more commonly associated with various benign causes including proton pump inhibitor (PPI) use, Helicobacter pylori infection and/or atrophic gastritis. The extent to which these factors interact to influence fasting serum gastrin concentrations remains incompletely understood. Materials and Methods: Fasting serum gastrin concentrations were measured by radioimmunoassay in 1,400 patients attending for diagnostic oesophagogastro-duodenoscopy. After exclusions, 982 patients were divided into four groups and their results analysed. We compared gastrin concentrations in normal patients (no H. pylori infection, no PPI use and no histological evidence of gastric preneoplasia (n=233)), with those in patients who were taking regular PPIs (H. pylori negative with no gastric preneoplasia (n=301)), patients who had active H. pylori infection but no gastric preneoplasia (n=164) and patients with histologically confirmed gastric preneoplasia (n=284). Results: Median fasting gastrin concentration in the normal group was 20pM and was significantly increased in PPI users (46pM, p<0.0001), patients with active H. pylori infection (27pM, p<0.0001), and patients with antral (25pM, p<0.01) or corpus (48pM, p<0.0001) gastric preneoplasia. PPI use resulted in further significant increases in fasting serum gastrin concentrations in patients who were infected with H. pylori (50pM, n=56) or who had antral gastric preneoplasia (53pM, n=87), but did not significantly alter serum gastrin concentrations in patients with corpus preneoplasia (90pM, n=66). Conclusions: PPI use, H. pylori infection and atrophic gastritis all caused significant elevations of median fasting gastrin concentrations. However, several patients who had potential risk factors for hypergastrinaemia still demonstrated fasting serum gastrin concentrations within the normal range.


Assuntos
Gastrinas/sangue , Infecções por Helicobacter/tratamento farmacológico , Helicobacter pylori , Lesões Pré-Cancerosas/patologia , Inibidores da Bomba de Prótons/uso terapêutico , Neoplasias Gástricas/patologia , Idoso , Endoscopia do Sistema Digestório , Jejum , Feminino , Gastrite/complicações , Gastrite/tratamento farmacológico , Infecções por Helicobacter/microbiologia , Humanos , Masculino , Pessoa de Meia-Idade , Lesões Pré-Cancerosas/complicações , Neoplasias Gástricas/complicações
3.
Commun Biol ; 4(1): 1247, 2021 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-34728772

RESUMO

The gastric epithelium is often exposed to injurious elements and failure of appropriate healing predisposes to ulcers, hemorrhage, and ultimately cancer. We examined the gastric function of CD36, a protein linked to disease and homeostasis. We used the tamoxifen model of gastric injury in mice null for Cd36 (Cd36-/-), with Cd36 deletion in parietal cells (PC-Cd36-/-) or in endothelial cells (EC-Cd36-/-). CD36 expresses on corpus ECs, on PC basolateral membranes, and in gastrin and ghrelin cells. Stomachs of Cd36-/- mice have altered gland organization and secretion, more fibronectin, and inflammation. Tissue respiration and mitochondrial efficiency are reduced. Phospholipids increased and triglycerides decreased. Mucosal repair after injury is impaired in Cd36-/- and EC-Cd36-/-, not in PC-Cd36-/- mice, and is due to defect of progenitor differentiation to PCs, not of progenitor proliferation or mature PC dysfunction. Relevance to humans is explored in the Vanderbilt BioVu using PrediXcan that links genetically-determined gene expression to clinical phenotypes, which associates low CD36 mRNA with gastritis, gastric ulcer, and gastro-intestinal hemorrhage. A CD36 variant predicted to disrupt an enhancer site associates (p < 10-17) to death from gastro-intestinal hemorrhage in the UK Biobank. The findings support role of CD36 in gastric tissue repair, and its deletion associated with chronic diseases that can predispose to malignancy.


Assuntos
Antígenos CD36/genética , Mucosa Gástrica/metabolismo , Gastrite/genética , Hemorragia Gastrointestinal/genética , Úlcera Gástrica/genética , Animais , Antígenos CD36/metabolismo , Células Endoteliais/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL
4.
Cell Mol Gastroenterol Hepatol ; 10(1): 113-132, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32004755

RESUMO

BACKGROUND & AIMS: In patients with autoimmune atrophic gastritis and achlorhydria, hypergastrinemia is associated with the development of type 1 gastric neuroendocrine tumors (gNETs). Twelve months of treatment with netazepide (YF476), an antagonist of the cholecystokinin B receptor (CCKBR or CCK2R), eradicated some type 1 gNETs in patients. We investigated the mechanisms by which netazepide induced gNET regression using gene expression profiling. METHODS: We obtained serum samples and gastric corpus biopsy specimens from 8 patients with hypergastrinemia and type 1 gNETs enrolled in a phase 2 trial of netazepide. Control samples were obtained from 10 patients without gastric cancer. We used amplified and biotinylated sense-strand DNA targets from total RNA and Affymetrix (Thermofisher Scientific, UK) Human Gene 2.0 ST microarrays to identify differentially expressed genes in stomach tissues from patients with type 1 gNETs before, during, and after netazepide treatment. Findings were validated in a human AGSGR gastric adenocarcinoma cell line that stably expresses human CCK2R, primary mouse gastroids, transgenic hypergastrinemic INS-GAS mice, and patient samples. RESULTS: Levels of pappalysin 2 (PAPPA2) messenger RNA were reduced significantly in gNET tissues from patients receiving netazepide therapy compared with tissues collected before therapy. PAPPA2 is a metalloproteinase that increases the bioavailability of insulin-like growth factor (IGF) by cleaving IGF binding proteins (IGFBPs). PAPPA2 expression was increased in the gastric corpus of patients with type 1 gNETs, and immunohistochemistry showed localization in the same vicinity as CCK2R-expressing enterochromaffin-like cells. Up-regulation of PAPPA2 also was found in the stomachs of INS-GAS mice. Gastrin increased PAPPA2 expression with time and in a dose-dependent manner in gastric AGSGR cells and mouse gastroids by activating CCK2R. Knockdown of PAPPA2 in AGSGR cells with small interfering RNAs significantly decreased their migratory response and tissue remodeling in response to gastrin. Gastrin altered the expression and cleavage of IGFBP3 and IGFBP5. CONCLUSIONS: In an analysis of human gNETS and mice, we found that gastrin up-regulates the expression of gastric PAPPA2. Increased PAPPA2 alters IGF bioavailability, cell migration, and tissue remodeling, which are involved in type 1 gNET development. These effects are inhibited by netazepide.


Assuntos
Benzodiazepinonas/farmacologia , Tumores Neuroendócrinos/tratamento farmacológico , Compostos de Fenilureia/farmacologia , Proteína Plasmática A Associada à Gravidez/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Animais , Benzodiazepinas/farmacologia , Benzodiazepinonas/uso terapêutico , Linhagem Celular Tumoral , Modelos Animais de Doenças , Mucosa Gástrica/citologia , Mucosa Gástrica/patologia , Gastrinas/antagonistas & inibidores , Gastrinas/sangue , Gastrinas/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos Transgênicos , Tumores Neuroendócrinos/sangue , Tumores Neuroendócrinos/patologia , Organoides , Compostos de Fenilureia/uso terapêutico , Proteína Plasmática A Associada à Gravidez/análise , Proteína Plasmática A Associada à Gravidez/antagonistas & inibidores , Proteína Plasmática A Associada à Gravidez/genética , Cultura Primária de Células , Receptor de Colecistocinina B/antagonistas & inibidores , Receptor de Colecistocinina B/metabolismo , Neoplasias Gástricas/sangue , Neoplasias Gástricas/patologia , Resultado do Tratamento
5.
Data Brief ; 24: 103886, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31011600

RESUMO

Radioimmunoassay data on the recharacterization of a gastrin polycolonal antibody first generated in 1973 is presented. The data include specificity, effect of matrix, and the establishment of a reference range for circulating fasting gastrin concentrations in normal subjects. For a discussion of the interpretation of the data, please see doi, 10.1016/j.peptides.2019.02.001 [1].

6.
Cancers (Basel) ; 11(2)2019 Feb 23.
Artigo em Inglês | MEDLINE | ID: mdl-30813438

RESUMO

Although hypoxia is known to contribute to several aspects of tumour progression, relatively little is known about the effects of hypoxia on cancer-associated myofibroblasts (CAMs), or the consequences that conditional changes in CAM function may have on tumour development and metastasis. To investigate this issue in the context of gastric cancer, a comparative multiomic analysis was performed on populations of patient-derived myofibroblasts, cultured under normoxic or hypoxic conditions. Data from this study reveal a novel set of CAM-specific hypoxia-induced changes in gene expression and secreted proteins. Significantly, these signatures are not observed in either patient matched adjacent tissue myofibroblasts (ATMs) or non-cancer associated normal tissue myofibroblasts (NTMs). Functional characterisation of different myofibroblast populations shows that hypoxia-induced changes in gene expression not only enhance the ability of CAMs to induce cancer cell migration, but also confer pro-tumorigenic (CAM-like) properties in NTMs. This study provides the first global mechanistic insight into the molecular changes that contribute to hypoxia-induced pro-tumorigenic changes in gastric stromal myofibroblasts.

7.
Clin Nutr ESPEN ; 30: 59-66, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30904230

RESUMO

BACKGROUND & AIMS: Bioelectrical impedance vector analysis (BIVA) is a non-invasive method of measuring human body composition. This offers the potential to evaluate nutritional and hydration states in cancer. Analysis of BIVA data using the z-score method (the number of standard deviations away from the mean value of the reference group) has the potential to facilitate comparisons between different cancer types. The aim of this study was to use the BIVA Reactance (R)/Reactance (Xc) z-score method to evaluate body composition differences in cancer, using data from previously published BIVA studies. METHODS: Previous studies using BIVA in cancer were identified from the literature. Bioimpedance measurements were analysed using the BIVA RXc z-score graph. The mean impedance vectors from the studied populations were transformed into standard deviates (with respect to the mean and standard deviation of the reference populations). Body composition was classified according to vector placement (i.e. normal, athletic, cachectic, oedematous and dehydrated). RESULTS: Seven male and three cancer female populations were evaluated. Body composition was classified as normal for the majority (n = 5), followed by cachexia (n = 4) and athletic (n = 1) respectively. Variation in body composition for the studied populations appeared to be related to gender, disease type and severity. CONCLUSIONS: The BIVA RXc z-score method has potential to evaluate body composition differences between cancer groups. This method can study body composition, according to cancer type, stage, gender and ethnicity. Limitations of the method relate to issues concerning the appropriate use of reference populations and variability between bioimpedance analysers. Better body composition assessment has the potential to personalise therapeutic, nutritional and hydration management. Further work is essential to facilitate in-depth evaluation in these areas, in order to achieve meaningful use of BIVA in clinical practice.


Assuntos
Composição Corporal , Caquexia/fisiopatologia , Impedância Elétrica , Neoplasias/fisiopatologia , Feminino , Humanos , Masculino , Estadiamento de Neoplasias , Neoplasias/patologia
8.
Oncotarget ; 10(2): 98-112, 2019 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-30719206

RESUMO

The chemokine-like peptide, chemerin, stimulates chemotaxis in several cell types. In this study we examined the expression of putative chemerin receptors in gastric cancer and the action of chemerin on cancer cell migration and invasion. Immunohistochemical studies of gastric tumors identified expression of two putative receptors, chemokine-like receptor-1 (CMKLR1) and G-protein coupled receptor 1(GPR1), in cancer cells; there was also some expression in stromal myofibroblasts although generally at a lower intensity. The expression of both receptors was detected in a gastric cancer cell line, AGS; chemerin itself was expressed in cultured gastric cancer myofibroblasts but not AGS cells. Chemerin stimulated (a) morphological transformation of AGS cells characterized by extension of processes and cell scattering, (b) migration in scratch wound assays and (c) both migration and invasion in Boyden chamber chemotaxis assays. These responses were inhibited by two putative receptor antagonists CCX832 and α-NETA. Inhibition of receptor expression by siRNA selectively reduced CMKLR1 or GPR1 and inhibited the action of chemerin indicating that both receptors contributed to the functional response. Using a proteomic approach employing stable isotope dynamic labeling of secretomes (SIDLS) to selectively label secreted proteins, we identified down regulation of tissue inhibitors of metalloproteinease (TIMP)1 and TIMP2 in media in response to chemerin. When cells were treated with chemerin and TIMP1 or TIMP2 the migration response to chemerin was reduced. The data suggest a role for chemerin in promoting the invasion of gastric cancer cells via CMKLR1 and GPR1at least partly by reducing TIMP1 and TIMP2 expression. Chemerin receptor antagonists have potential in inhibiting gastric cancer progression.

9.
Carcinogenesis ; 40(4): 500-512, 2019 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-30624614

RESUMO

There is increasing evidence that stromal myofibroblasts play a key role in the tumour development however, the mechanisms by which they become reprogrammed to assist in cancer progression remain unclear. As cultured cancer-associated myofibroblasts (CAMs) retain an ability to enhance the proliferation and migration of cancer cells in vitro, it is possible that epigenetic reprogramming of CAMs within the tumour microenvironment may confer long-term pro-tumourigenic changes in gene expression. This study reports the first comparative multi-omics analysis of cancer-related changes in gene expression and DNA methylation in primary myofibroblasts derived from gastric and oesophageal tumours. In addition, we identify novel CAM-specific DNA methylation signatures, which are not observed in patient-matched adjacent tissue-derived myofibroblasts, or corresponding normal tissue-derived myofibroblasts. Analysis of correlated changes in DNA methylation and gene expression shows that different patterns of gene-specific DNA methylation have the potential to confer pro-tumourigenic changes in metabolism, cell signalling and differential responses to hypoxia. These molecular signatures provide new insights into potential mechanisms of stromal reprogramming in gastric and oesophageal cancer, while also providing a new resource to facilitate biomarker identification and future hypothesis-driven studies into mechanisms of stromal reprogramming and tumour progression in solid tumours.


Assuntos
Biomarcadores Tumorais/genética , Epigênese Genética , Neoplasias Esofágicas/patologia , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Miofibroblastos/patologia , Neoplasias Gástricas/patologia , Movimento Celular , Proliferação de Células , Metilação de DNA , Epigenômica , Neoplasias Esofágicas/genética , Humanos , Miofibroblastos/metabolismo , Neoplasias Gástricas/genética , Células Tumorais Cultivadas , Microambiente Tumoral
10.
Analyst ; 143(24): 5912-5917, 2018 Dec 03.
Artigo em Inglês | MEDLINE | ID: mdl-30191233

RESUMO

This work reports the first images obtained by combining an infrared aperture scanning near-field optical microscope (SNOM) with a quantum cascade laser (QCL). The future potential of this set-up is demonstrated by a preliminary study on an OE33 human oesophageal adenocarcinoma cell in which the cell is imaged at 1751 cm-1, 1651 cm-1, 1539 cm-1 and 1242 cm-1. In addition to the 1651 cm-1 image, three other images were acquired within the Amide I band (1689 cm-1, 1675 cm-1 and 1626 cm-1) chosen to correspond to secondary structures of proteins. The four images obtained within the Amide I band show distinct differences demonstrating the potential of this approach to reveal subtle changes in the chemical composition of a cell.


Assuntos
Adenocarcinoma/diagnóstico por imagem , Células Epiteliais/patologia , Lasers Semicondutores , Microscopia/instrumentação , Microscopia/métodos , Adenocarcinoma/patologia , Linhagem Celular Tumoral , Humanos
11.
Mol Cell Proteomics ; 17(9): 1837-1849, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29915148

RESUMO

Analysis of secretomes critically underpins the capacity to understand the mechanisms determining interactions between cells and between cells and their environment. In the context of cancer cell micro-environments, the relevant interactions are recognized to be an important determinant of tumor progression. Global proteomic analyses of secretomes are often performed at a single time point and frequently identify both classical secreted proteins (possessing an N-terminal signal sequence), as well as many intracellular proteins, the release of which is of uncertain biological significance. Here, we describe a mass spectrometry-based method for stable isotope dynamic labeling of secretomes (SIDLS) that, by dynamic SILAC, discriminates the secretion kinetics of classical secretory proteins and intracellular proteins released from cancer and stromal cells in culture. SIDLS is a robust classifier of the different cellular origins of proteins within the secretome and should be broadly applicable to nonproliferating cells and cells grown in short term culture.


Assuntos
Marcação por Isótopo/métodos , Neoplasias/metabolismo , Proteoma/metabolismo , Linhagem Celular Tumoral , Ontologia Genética , Humanos , Espaço Intracelular/metabolismo , Cinética , Reprodutibilidade dos Testes , Transdução de Sinais , Células Estromais/metabolismo , Fatores de Tempo
12.
Physiol Rep ; 6(10): e13683, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29845775

RESUMO

Matrix metalloproteinase (MMP)-7, unlike many MMPs, is typically expressed in epithelial cells. It has been linked to epithelial responses to infection, injury, and tissue remodeling including the progression of a number of cancers. We have now examined how MMP-7 expression changes in the progression to esophageal adenocarcinoma (EAC), and have studied mechanisms regulating its expression and its functional significance. Immunohistochemistry revealed that MMP-7 was weakly expressed in normal squamous epithelium adjacent to EAC but was abundant in epithelial cells in both preneoplastic lesions of Barrett's esophagus and EAC particularly at the invasive front. In the stroma, putative myofibroblasts expressing MMP-7 were abundant at the invasive front but were scarce or absent in adjacent tissue. Western blot and ELISA revealed high constitutive secretion of proMMP-7 in an EAC cell line (OE33) that was inhibited by the phosphatidylinositol (PI) 3-kinase inhibitor LY294002 but not by inhibitors of protein kinase C, or MAP kinase activation. There was detectable proMMP-7 in cultured esophageal myofibroblasts but it was undetectable in media. Possible metabolism of MMP-7 by myofibroblasts studied by proteomic analysis indicated degradation via extensive endopeptidase, followed by amino- and carboxpeptidase, cleavages. Myofibroblasts exhibited increased migration and invasion in response to conditioned media from OE33 cells that was reduced by MMP-7 knockdown and immunoneutralization. Thus, MMP-7 expression increases at the invasive front in EAC which may be partly attributable to activation of PI 3-kinase. Secreted MMP-7 may modify the tumor microenvironment by stimulating stromal cell migration and invasion.


Assuntos
Adenocarcinoma/metabolismo , Esôfago de Barrett/metabolismo , Neoplasias Esofágicas/metabolismo , Metaloproteinase 7 da Matriz/metabolismo , Adenocarcinoma/complicações , Idoso , Esôfago de Barrett/complicações , Linhagem Celular Tumoral , Progressão da Doença , Neoplasias Esofágicas/complicações , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Miofibroblastos/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo
13.
PLoS Pathog ; 13(11): e1006653, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29095917

RESUMO

Several conditions associated with reduced gastric acid secretion confer an altered risk of developing a gastric malignancy. Helicobacter pylori-induced atrophic gastritis predisposes to gastric adenocarcinoma, autoimmune atrophic gastritis is a precursor of type I gastric neuroendocrine tumours, whereas proton pump inhibitor (PPI) use does not affect stomach cancer risk. We hypothesised that each of these conditions was associated with specific alterations in the gastric microbiota and that this influenced subsequent tumour risk. 95 patients (in groups representing normal stomach, PPI treated, H. pylori gastritis, H. pylori-induced atrophic gastritis and autoimmune atrophic gastritis) were selected from a cohort of 1400. RNA extracted from gastric corpus biopsies was analysed using 16S rRNA sequencing (MiSeq). Samples from normal stomachs and patients treated with PPIs demonstrated similarly high microbial diversity. Patients with autoimmune atrophic gastritis also exhibited relatively high microbial diversity, but with samples dominated by Streptococcus. H. pylori colonisation was associated with decreased microbial diversity and reduced complexity of co-occurrence networks. H. pylori-induced atrophic gastritis resulted in lower bacterial abundances and diversity, whereas autoimmune atrophic gastritis resulted in greater bacterial abundance and equally high diversity compared to normal stomachs. Pathway analysis suggested that glucose-6-phospahte1-dehydrogenase and D-lactate dehydrogenase were over represented in H. pylori-induced atrophic gastritis versus autoimmune atrophic gastritis, and that both these groups showed increases in fumarate reductase. Autoimmune and H. pylori-induced atrophic gastritis were associated with different gastric microbial profiles. PPI treated patients showed relatively few alterations in the gastric microbiota compared to healthy subjects.


Assuntos
Acloridria/microbiologia , Mucosa Gástrica/microbiologia , Microbioma Gastrointestinal , Acloridria/induzido quimicamente , Acloridria/etiologia , Acloridria/imunologia , Adulto , Idoso , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/imunologia , Doenças Autoimunes/microbiologia , Análise por Conglomerados , Estudos de Coortes , Inglaterra/epidemiologia , Feminino , Mucosa Gástrica/efeitos dos fármacos , Mucosa Gástrica/imunologia , Gastrite Atrófica/tratamento farmacológico , Gastrite Atrófica/imunologia , Gastrite Atrófica/microbiologia , Microbioma Gastrointestinal/efeitos dos fármacos , Infecções por Helicobacter/tratamento farmacológico , Infecções por Helicobacter/imunologia , Infecções por Helicobacter/microbiologia , Helicobacter pylori/efeitos dos fármacos , Helicobacter pylori/crescimento & desenvolvimento , Helicobacter pylori/imunologia , Helicobacter pylori/isolamento & purificação , Hospitais Universitários , Humanos , Masculino , Pessoa de Meia-Idade , Inibidores da Bomba de Prótons/efeitos adversos , Inibidores da Bomba de Prótons/uso terapêutico , Risco , Neoplasias Gástricas/epidemiologia
14.
Physiol Rep ; 5(19)2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29038353

RESUMO

The well-known action of the gastric hormone gastrin in stimulating gastric acid secretion is mediated by activation of cholecystokinin-2 receptors (CCK2R). The latter are expressed by a variety of cell types suggesting that gastrin is implicated in multiple functions. During wound healing in the stomach CCK2R may be expressed by myofibroblasts. We have now characterized CCK2R expression in cultured myofibroblasts. Immunocytochemistry showed that a relatively small proportion (1-6%) of myofibroblasts expressed the receptor regardless of the region of the gut from which they were derived, or whether from cancer or control tissue. Activation of CCK2R by human heptadecapeptide gastrin (hG17) increased intracellular calcium concentrations in a small subset of myofibroblasts indicating the presence of a functional receptor. Unexpectedly, we found over 80% of cells expressing CCK2R were also labeled with 5-ethynyl-2'-deoxyuridine (EdU) which is incorporated into DNA during S-phase of the cell cycle. hG17 did not stimulate EdU incorporation but increased migration of both EdU-labeled and unlabelled myofibroblasts; the migratory response was inhibited by a CCK2R antagonist and by an inhibitor of IGF receptor tyrosine kinase; hG17 also increased IGF-2 transcript abundance. The data suggest myofibroblasts express CCK2R in a restricted period of the cell cycle during S-phase, and that gastrin accelerates migration of these cells; it also stimulates migration of adjacent cells probably through paracrine release of IGF. Together with previous findings, the results raise the prospect that gastrin controls the position of dividing myofibroblasts which may be relevant in wound healing and cancer progression in the gastrointestinal tract.


Assuntos
Ciclo Celular , Movimento Celular , Miofibroblastos/metabolismo , Receptor de Colecistocinina B/metabolismo , Estômago/citologia , Cálcio/metabolismo , Células Cultivadas , Mucosa Gástrica/metabolismo , Gastrinas/farmacologia , Humanos , Fator de Crescimento Insulin-Like II/genética , Fator de Crescimento Insulin-Like II/metabolismo , Miofibroblastos/citologia , Miofibroblastos/fisiologia , Receptor de Colecistocinina B/agonistas , Receptor de Colecistocinina B/genética
15.
Peptides ; 91: 49-57, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28363795

RESUMO

Glucagon-like peptide (GLP)-2 stimulates intestinal epithelial proliferation by acting, in part, via IGF release from sub-epithelial myofibroblasts. The response of myofibroblasts to GLP-2 remains incompletely understood. We studied the action of GLP-2 on myofibroblasts from colon cancer and adjacent tissue, and the effects of conditioned medium from these cells on epithelial cell proliferation, migration and invasion. GLP-2 stimulated proliferation, migration and invasion of myofibroblasts and the proliferative and invasive responses of cancer-associated myofibroblasts were greater than those of myofibroblasts from adjacent tissue. The responses were inhibited by an IGF receptor inhibitor, AG1024. Conditioned medium from GLP-2 treated myofibroblasts increased proliferation, migration and invasion of SW480, HT29, LoVo epithelial cells and these responses were inhibited by AG1024; GLP-2 alone had no effect on these cells. In addition, when myofibroblasts and epithelial cells were co-cultured in Ibidi chambers there was mutual stimulation of migration in response to GLP-2. The latter increased both IGF-1 and IGF-2 transcript abundance in myofibroblasts. Moreover, a number of IGF binding proteins (IGFBP-4, -5, -7) were identified in myofibroblast medium; in the presence of GLP-2 there was increased abundance of the cleavage products of IGBBP-4 and IGFBP-5 suggesting activation of a degradation mechanism that might increase IGF bioavailability. The data suggest that GLP-2 stimulates cancer myofibroblast proliferation, migration and invasion; GLP-2 acts indirectly on epithelial cells partly via increased IGF expression in myofibroblasts and partly, perhaps, by increased bioavailability through degradation of IGFBPs.


Assuntos
Movimento Celular , Neoplasias do Colo/patologia , Peptídeo 2 Semelhante ao Glucagon/fisiologia , Mucosa Intestinal/patologia , Miofibroblastos/patologia , Idoso de 80 Anos ou mais , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Técnicas de Cocultura , Neoplasias do Colo/metabolismo , Meios de Cultivo Condicionados/farmacologia , Células Epiteliais/efeitos dos fármacos , Feminino , Peptídeo 2 Semelhante ao Glucagon/farmacologia , Células HT29 , Humanos , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Redes e Vias Metabólicas/efeitos dos fármacos , Miofibroblastos/efeitos dos fármacos , Invasividade Neoplásica , Receptor IGF Tipo 1/antagonistas & inibidores , Receptor IGF Tipo 1/metabolismo , Receptor IGF Tipo 2/antagonistas & inibidores , Receptor IGF Tipo 2/metabolismo , Células Tumorais Cultivadas , Tirfostinas/farmacologia
16.
Br J Clin Pharmacol ; 83(3): 466-475, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-27704617

RESUMO

AIMS: Netazepide, a gastrin/cholecystokinin 2 receptor antagonist, once daily for 12 weeks reduced the number of tumours and size of the largest one in 16 patients with autoimmune chronic atrophic gastritis (CAG), achlorhydria, hypergastrinaemia and multiple gastric neuroendocrine tumours (type 1 gastric NETs), and normalized circulating chromogranin A (CgA) produced by enterochromaffin-like cells, the source of the tumours. The aim was to assess whether longer-term netazepide treatment can eradicate type 1 gastric NETs. METHODS: After a mean 14 months off netazepide, 13 of the 16 patients took it for another 52 weeks. Assessments were: gastroscopy; gene-transcript expression in corpus biopsies using quantitative polymerase chain reaction; blood CgA and gastrin concentrations; and safety assessments. RESULTS: While off-treatment, the number of tumours, the size of the largest one, and CgA all increased again. Netazepide for 52 weeks: cleared all tumours in 5 patients; cleared all but one tumour in one patient; reduced the number of tumours and size of the largest one in the other patients; normalized CgA in all patients; and reduced mRNA abundances of CgA and histidine decarboxylase in biopsies. Gastrin did not increase further, confirming that the patients had achlorhydria. Netazepide was safe and well tolerated. CONCLUSIONS: A gastrin/cholecystokinin 2 receptor antagonist is a potential medical and targeted treatment for type 1 gastric NETs, and an alternative to regular gastroscopy or surgery. Treatment should be continuous because the tumours will regrow if it is stopped. Progress can be monitored by CgA in blood or biomarkers in mucosal biopsies.


Assuntos
Doenças Autoimunes/tratamento farmacológico , Benzodiazepinonas/uso terapêutico , Gastrite Atrófica/tratamento farmacológico , Tumores Neuroendócrinos/tratamento farmacológico , Compostos de Fenilureia/uso terapêutico , Acloridria/complicações , Acloridria/tratamento farmacológico , Acloridria/metabolismo , Idoso , Doenças Autoimunes/sangue , Doenças Autoimunes/complicações , Benzodiazepinonas/efeitos adversos , Cromogranina A/biossíntese , Cromogranina A/sangue , Gastrinas/sangue , Gastrite Atrófica/sangue , Gastrite Atrófica/complicações , Histidina Descarboxilase/biossíntese , Humanos , Pessoa de Meia-Idade , Tumores Neuroendócrinos/sangue , Tumores Neuroendócrinos/complicações , Tumores Neuroendócrinos/metabolismo , Compostos de Fenilureia/efeitos adversos
17.
Cereb Cortex ; 27(3): 2318-2334, 2017 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-27073217

RESUMO

Cholecystokinin-expressing interneurons (CCK-INs) mediate behavior state-dependent inhibition in cortical circuits and themselves receive strong GABAergic input. However, it remains unclear to what extent GABAB receptors (GABABRs) contribute to their inhibitory control. Using immunoelectron microscopy, we found that CCK-INs in the rat hippocampus possessed high levels of dendritic GABABRs and KCTD12 auxiliary proteins, whereas postsynaptic effector Kir3 channels were present at lower levels. Consistently, whole-cell recordings revealed slow GABABR-mediated inhibitory postsynaptic currents (IPSCs) in most CCK-INs. In spite of the higher surface density of GABABRs in CCK-INs than in CA1 principal cells, the amplitudes of IPSCs were comparable, suggesting that the expression of Kir3 channels is the limiting factor for the GABABR currents in these INs. Morphological analysis showed that CCK-INs were diverse, comprising perisomatic-targeting basket cells (BCs), as well as dendrite-targeting (DT) interneurons, including a previously undescribed DT type. GABABR-mediated IPSCs in CCK-INs were large in BCs, but small in DT subtypes. In response to prolonged activation, GABABR-mediated currents displayed strong desensitization, which was absent in KCTD12-deficient mice. This study highlights that GABABRs differentially control CCK-IN subtypes, and the kinetics and desensitization of GABABR-mediated currents are modulated by KCTD12 proteins.


Assuntos
Colecistocinina/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/metabolismo , Potenciais Pós-Sinápticos Inibidores/fisiologia , Interneurônios/metabolismo , Canais de Potássio/metabolismo , Receptores de GABA-A/metabolismo , Animais , Região CA1 Hipocampal/metabolismo , Região CA1 Hipocampal/ultraestrutura , Dendritos/metabolismo , Dendritos/ultraestrutura , Imuno-Histoquímica , Interneurônios/ultraestrutura , Masculino , Microscopia Imunoeletrônica , Técnicas de Patch-Clamp , Ratos Wistar , Técnicas de Cultura de Tecidos
18.
Cereb Cortex ; 27(4): 2453-2468, 2017 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-27102657

RESUMO

Although extensively studied postnatally, the functional differentiation of cholecystokinin (CCK)-containing interneurons en route towards the cerebral cortex during fetal development is incompletely understood. Here, we used CCKBAC/DsRed mice encoding a CCK promoter-driven red fluorescent protein to analyze the temporal dynamics of DsRed expression, neuronal identity, and positioning through high-resolution developmental neuroanatomy. Additionally, we developed a dual reporter mouse line (CCKBAC/DsRed::GAD67gfp/+) to differentiate CCK-containing interneurons from DsRed+ principal cells during prenatal development. We show that DsRed is upregulated in interneurons once they exit their proliferative niche in the ganglionic eminence and remains stably expressed throughout their long-distance migration towards the cerebrum, particularly in the hippocampus. DsRed+ interneurons, including a cohort coexpressing calretinin, accumulated at the palliosubpallial boundary by embryonic day 12.5. Pioneer DsRed+ interneurons already reached deep hippocampal layers by embryonic day 14.5 and were morphologically differentiated by birth. Furthermore, we probed migrating interneurons entering and traversing the cortical plate, as well as stationary cells in the hippocampus by patch-clamp electrophysiology to show the first signs of Na+ and K+ channel activity by embryonic day 12.5 and reliable adult-like excitability by embryonic day 18.5. Cumulatively, this study defines key positional, molecular, and biophysical properties of CCK+ interneurons in the prenatal brain.


Assuntos
Diferenciação Celular/fisiologia , Córtex Cerebral/citologia , Colecistocinina/metabolismo , Interneurônios/citologia , Neurogênese/fisiologia , Animais , Movimento Celular , Córtex Cerebral/embriologia , Córtex Cerebral/metabolismo , Imuno-Histoquímica , Hibridização In Situ , Interneurônios/metabolismo , Camundongos , Camundongos Transgênicos , Microscopia Confocal , Técnicas de Patch-Clamp
19.
Oncotarget ; 8(1): 203-214, 2017 Jan 03.
Artigo em Inglês | MEDLINE | ID: mdl-27448962

RESUMO

OBJECTIVE: The incidence of esophageal adenocarcinoma (EAC) is increasing, but factors contributing to malignant progression of its precursor lesion, Barrett's esophagus (BE), have not been defined. Hypergastrinemia caused by long-term use of proton pump inhibitors (PPIs), has been suggested as one possible risk factor. The gastrin receptor, CCK2R, is expressed in the cardia and upregulated in BE, suggesting the involvement of the gastrin-CCK2R pathway in progression. In the L2-IL-1ß mouse model, Barrett's-like esophagus arises from the gastric cardia. Therefore, we aimed to analyze the effect of hypergastrinemia on CCK2R+ progenitor cells in L2-IL-1ß mice. DESIGN: L2-IL-1ß mice were mated with hypergastrinemic (INS-GAS) mice or treated with PPIs to examine the effect of hypergastrinemia in BE progression. CCK2R-CreERT crossed with L2-IL-1ß mice were used to analyze the lineage progenitor potential of CCK2R+ cells. Cardia glands were cultured in vitro, and the effect of gastrin treatment analyzed. L2-IL-1ß mice were treated with a CCK2R antagonist YF476 as a potential chemopreventive drug. RESULTS: Hypergastrinemia resulted in increased proliferation and expansion of Barrett's-like esophagus. Lineage tracing experiments revealed that CCK2R+ cells are long-lived progenitors that can give rise to such lesions under chronic inflammation. Gastrin stimulated organoid growth in cardia culture, while CCK2R inhibition prevented Barrett's-like esophagus and dysplasia. CONCLUSIONS: Our data suggest a progression model for BE to EAC in which CCK2R+ progenitor cells, stimulated by hypergastrinemia, proliferate to give rise to metaplasia and dysplasia. Hypergastrinemia can result from PPI use, and the effects of hypergastrinemia in human BE should be studied further.


Assuntos
Esôfago de Barrett/etiologia , Esôfago de Barrett/metabolismo , Gastrinas/metabolismo , Regulação da Expressão Gênica , Mioblastos Cardíacos/metabolismo , Receptor de Colecistocinina B/genética , Animais , Esôfago de Barrett/patologia , Biomarcadores , Modelos Animais de Doenças , Progressão da Doença , Gastrinas/farmacologia , Hiperglicemia , Imuno-Histoquímica , Metaplasia , Camundongos , Camundongos Transgênicos , Mioblastos Cardíacos/efeitos dos fármacos , Fenótipo , Receptor de Colecistocinina B/metabolismo
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