RESUMO
Atlantic salmon exhibit seasonal reproduction. However, the mechanisms governing this are still unclear. Generally speaking, kisspeptin has been recognized as a regulator of reproduction. Here, we report a relationship between kisspeptin, GnRH and photoperiod in Atlantic salmon. The results demonstrated that the expression of the Atlantic salmon kisspeptin-receptor (skissr) was not always consistent with the expression pattern of Atlantic salmon GnRH3 (sGnRH3) during all developmental processes. Kisspeptin may exert its influence primarily in the early and later stages of gonad development by promoting the secretion of sGnRH3. Meanwhile, the expression levels of kissr were higher in fish with gonads at stage II and stage V under the long-day photoperiod regime than under the short-day regime. In addition, both skissr and sGnRH3 were also expressed in the saccus vasculosus (SV), an organ only found in fish. The SV might be a seasonal sensor regulating reproduction in addition to the hypothalamus (Hyp).
Assuntos
Proteínas de Peixes/biossíntese , Regulação da Expressão Gênica/fisiologia , Gônadas/crescimento & desenvolvimento , Hipotálamo/crescimento & desenvolvimento , Fotoperíodo , Receptores de Neuropeptídeos/biossíntese , Salmo salar/crescimento & desenvolvimento , Animais , Feminino , Hormônio Liberador de Gonadotropina/biossíntese , Masculino , Ácido Pirrolidonocarboxílico/análogos & derivadosRESUMO
Malignant mixed Müllerian tumors (MMMTs) are aggressive malignant neoplasms with a high recurrence rate and poor prognosis. Despite advances in adjuvant therapies in recent years, the prognosis of these tumors has not improved. Growth hormone-releasing hormone (GHRH) is produced by a variety of malignant tumors and acts as a growth factor in an autocrine/paracrine manner. Its function requires the presence of its receptors to exert its effects on neoplastic cells. In this study, we evaluated the expression of GHRH receptors (GHRH-R) in a group of MMMTs. Thirty-one examples of MMMTs from endometrium, ovary, uterine tube, and pelvic peritoneum were retrieved from the files of Department of Pathology at the University of Miami, Jackson Memorial Hospital. Immunohistochemistry for GHRH-R was performed on paraffin sections and the staining results were evaluated separately in both epithelial and mesenchymal components of each tumor. The presence of pituitary type growth hormone-releasing hormone receptor mRNA and that of its biologically active splice variant were also evaluated by RT-PCR in 6 of the tumors. Positive immunohistochemical reaction for GHRH-R was detected in 30 tumors (96%). The epithelial and sarcomatous components were positive in 30 (96%), whereas one endometrial tumor was negative in both components. The mRNA for GHRH-R and its splice variant was found in all 6 tested tumors. This study shows that GHRH-R is expressed by the majority of MMMTs in both epithelial and mesenchymal components. This finding could potentially serve as a basis for therapeutic approaches using synthetic peptide antagonists of GHRH-R that have shown significant efficacy with minimal side effects in experimental models.
Assuntos
Biomarcadores Tumorais/análise , Tumor Mulleriano Misto/metabolismo , Tumor Mulleriano Misto/patologia , Receptores de Neuropeptídeos/biossíntese , Receptores de Hormônios Reguladores de Hormônio Hipofisário/biossíntese , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Receptores de Neuropeptídeos/análise , Receptores de Hormônios Reguladores de Hormônio Hipofisário/análiseRESUMO
Prolonged stress and repeated activation of the hypothalamic-pituitary-adrenal axis can result in many sex-dependent behavioural and metabolic changes in rats, including alterations in feeding behaviour and reduced body weight. In adults, these effects of stress can be mimicked by corticosterone, a major output of the hypothalamic-pituitary-adrenal axis, and recapitulate the stress-induced sex difference, such that corticosterone-treated males show greater weight loss than females. Similar to adults, chronic stress during adolescence leads to reduced weight gain, particularly in males. However, it is currently unknown whether corticosterone mediates this somatic change and whether additional measures of neuroendocrine function are affected by chronic corticosterone exposure during adolescence in a sex-dependent manner. Therefore, we examined the effects of non-invasively administered corticosterone (150 or 300 µg/ml) in the drinking water of male and female rats throughout adolescent development (30-58 days of age). We found that adolescent animals exposed to chronic corticosterone gain significantly less weight than controls, which may be partly mediated by the effects of corticosterone on food consumption, fluid intake and gonadal hormone function. Our data further show that, despite similar circulating corticosterone levels, males demonstrate a greater sensitivity to these changes than females. We also found that Npy1 and Npy5 receptor mRNA expression, genes implicated in appetite regulation, was significantly reduced in the ventral medial hypothalamus of corticosterone-treated males and females compared to controls. Finally, parameters of gonadal function, such as plasma sex steroid concentrations and weight of reproductive tissues, were reduced by adolescent corticosterone treatment, although only in males. The data obtained in the present study indicate that chronic corticosterone exposure throughout adolescent development results in significant and sex-dependent somatic and neuroendocrine changes, and the results also provide an experimental framework for further investigating the impact of corticosterone on metabolic and neuroendocrine function during adolescence.
Assuntos
Peso Corporal/efeitos dos fármacos , Corticosterona/administração & dosagem , Corticosterona/farmacologia , Tamanho do Órgão/efeitos dos fármacos , Receptores Acoplados a Proteínas G/biossíntese , Receptores de Neuropeptídeo Y/biossíntese , Receptores de Neuropeptídeos/biossíntese , Animais , Corticosterona/sangue , Relação Dose-Resposta a Droga , Estradiol/sangue , Feminino , Hormônios Esteroides Gonadais/sangue , Hipotálamo Médio/metabolismo , Masculino , Ratos , Glândulas Seminais/efeitos dos fármacos , Caracteres Sexuais , Testículo/efeitos dos fármacos , Testosterona/sangue , Útero/efeitos dos fármacosRESUMO
OBJECTIVE: to define the role of neurotransmitters and their receptors in the development of itch and in the maintenance of a skin inflammatory response in patients with psoriasis and atopic dermatitis. MATERIAL AND METHODS: Skin biopsy specimens from 30 patients with psoriasis and 30 patients with atopic dermatitis were investigated by histological, immunoperoxidase, and indirect immunofluorescence assays. The investigators determined the expression of protein gene product 9.5 (PGP9.5), amphiregulin, semaphorin 3A, calcitonin gene-related peptide (CGRP) and its receptor (CGRP-R), nerve growth factor (NGF) and its receptor TrkA, and substance P (SP) and its receptor SP-R. The indirect immunofluorescence assay was used for quantitative analysis. The findings were statistically analyzed using a Statistica 10 program. RESULTS: Immunoperoxidase examination of the skin biopsy specimens from patients with atopic dermatitis and psoriasis revealed enhanced expression of amphiregulin, NGF, and PGP9.5, appearance of positively stained epidermal nerve fibers, and decreased expression of the nerve reduction factor semaphorin 3A in all cases. Some patients with atopic dermatitis and psoriasis showed increased expression of CGRP and CGRP-R, SP, SP-R, and TrkA. A pronounced inflammatory response was generally observed in these cases. CONCLUSION: The investigation performed suggests that atopic dermatitis and psoriasis are characterized by a larger number of epidermal nerve fibers and by a direct correlation between this indicator, disease severity, and itch intensity. The production of neuropeptides and neurotrophins is closely related to the development of a skin inflammatory response irrespective of its cause and dysregulation of these processes is likely to favor the body's sensitization and the chronic pattern the course of diseases.
Assuntos
Dermatite Atópica/etiologia , Fatores de Crescimento Neural/biossíntese , Neuropeptídeos/biossíntese , Psoríase/etiologia , Receptores de Fator de Crescimento Neural/biossíntese , Receptores de Neuropeptídeos/biossíntese , Estudos de Casos e Controles , Dermatite Atópica/metabolismo , Dermatite Atópica/patologia , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Técnicas Imunoenzimáticas , Psoríase/metabolismo , Psoríase/patologia , Índice de Gravidade de DoençaRESUMO
Papillary thyroid cancer (PTC) is the most prevalent of all endocrine cancers. In recent studies, the presence of receptors for pituitary-type growth hormone-releasing hormone (pGHRH-R) has been demonstrated in various human cancers, including human prostate, brain, and other cancer lines. Thyroid malignancies, however, have not yet been investigated in this regard. In this study, we found that pGHRH-R and its functional splice variant, SV1, are present in normal thyroid and PTC cells. We also treated seven normal and PTC tumor thyroid cells in vitro with a GHRH antagonist, MIA-602, to compare its anti-proliferation and anti-invasion potential against vehicle-treated cells. We found that treatment with GHRH antagonist increases the expression of SV1 and pGHRH-R in tumor cells compared to tumor cells exposed to vehicle only, a response which may alter the sensitivity of signaling kinases within the cells. GHRH antagonist treatment of tumor cells also reduced activity of the tumor invasion marker, matrix metalloproteinase (MMP)-2, compared to tumor cells exposed to vehicle only. The expression of pGHRH-R and SV1, as well as MMP-2 activity, in normal thyroid cells remained unaffected by GHRH antagonist treatment. Similarly, cell proliferation rates for tumor or normal thyroid cells were not affected by GHRH antagonist treatment. Our findings have important implications for the therapeutic use of GHRH antagonist in cases of aggressive PTC refractory to conventional treatment modalities, and in which protein expression and MMP-2 activity in normal thyroid tissue is left unaltered.
Assuntos
Carcinoma/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Receptores de Neuropeptídeos/biossíntese , Receptores de Hormônios Reguladores de Hormônio Hipofisário/biossíntese , Neoplasias da Glândula Tireoide/metabolismo , Western Blotting , Carcinoma Papilar , Proliferação de Células , Feminino , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Humanos , Pessoa de Meia-Idade , Sermorelina/análogos & derivados , Sermorelina/farmacologia , Câncer Papilífero da TireoideRESUMO
Electroconvulsive therapy (ECT) remains one of the most effective treatments of major depression. It has been suggested that the mechanisms of action involve gene expression. In recent decades there have been several investigations of gene expression following both acute and chronic electroconvulsive stimulation (ECS). These studies have focused on several distinct gene targets but have generally included only few time points after ECS for measuring gene expression. Here we measured gene expression of three types of genes: Immediate early genes, synaptic proteins, and neuropeptides at six time points following an acute ECS. We find significant increases for c-Fos, Egr1, Neuritin 1 (Nrn 1), Bdnf, Snap29, Synaptotagmin III (Syt 3), Synapsin I (Syn 1), and Psd95 at differing time points after ECS. For some genes these changes are prolonged whereas for others they are transient. Npy expression significantly increases whereas the gene expression of its receptors Npy1r, Npy2r, and Npy5r initially decreases. These decreases are followed by a significant increase for Npy2r, suggesting anticonvulsive adaptations following seizures. In summary, we find distinct changes in mRNA quantities that are characteristic for each gene. Considering the observed transitory and inverse changes in expression patterns, these data underline the importance of conducting measurements at several time points post-ECS.
Assuntos
Eletroconvulsoterapia/efeitos adversos , Genes Precoces/genética , Hipocampo/metabolismo , Membranas Sinápticas/metabolismo , Animais , Perfilação da Expressão Gênica , Masculino , Modelos Animais , Proteínas Proto-Oncogênicas c-fos/biossíntese , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , RNA Mensageiro/biossíntese , Ratos , Ratos Sprague-Dawley , Receptores Acoplados a Proteínas G/biossíntese , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Neuropeptídeos/biossíntese , Receptores de Neuropeptídeos/genética , Receptores de Neuropeptídeos/metabolismo , Receptores de Neuropeptídeo Y/biossíntese , Receptores de Neuropeptídeo Y/genética , Receptores de Neuropeptídeo Y/metabolismo , Convulsões/genética , Membranas Sinápticas/genética , TranscriptomaRESUMO
Different classes of breast cancers were explored for their positivity for growth hormone-releasing hormone receptors (GHRH-R) in this pilot study, as no systematic evaluation of such tumors has been performed to date. Seventy-two small primary breast carcinomas were evaluated for GHRH-R expression by immunohistochemistry using a polyclonal antibody and a cutoff value of 10% staining. GHRH-R positivity was detected in 58% of all cases, 20/23 (87%) of invasive lobular carcinomas (ILC) and 22/46 (48%) of invasive ductal carcinomas (IDC). GHRH-R positivity was more frequent in grade 2 tumors (86%), as compared to grade 1 (18%) or grade 3 (47%) cancers. GHRH-R expression was not associated with mitotic scores, the Ki-67 labeling indices or nodal status. IDCs with casting-type calcifications on the mammogram showed positivity for GHRH-R in 9/12 (75%) cases. Most importantly, apocrine epithelium, and all 10 apocrine carcinomas added later to the study were GHRH-R-positive. These preliminary results suggest a greater than average GHRH-R expression in ILCs and IDCs associated with casting-type calcifications on the mammogram. Apocrine carcinomas seem uniformly positive for GHRH-R. Whether these findings could indicate a potential role of GHRH-antagonists in targeted treatment of these types of breast cancer requires further studies.
Assuntos
Neoplasias da Mama/patologia , Glândulas Mamárias Humanas/metabolismo , Receptores de Neuropeptídeos/metabolismo , Receptores de Hormônios Reguladores de Hormônio Hipofisário/metabolismo , Neoplasias da Mama/classificação , Neoplasias da Mama/diagnóstico , Carcinoma Ductal de Mama/patologia , Carcinoma Lobular/patologia , Feminino , Humanos , Imuno-Histoquímica , Antígeno Ki-67/análise , Mamografia , RNA Mensageiro/biossíntese , Receptor ErbB-2/metabolismo , Receptores de Neuropeptídeos/biossíntese , Receptores de Neuropeptídeos/genética , Receptores de Hormônios Reguladores de Hormônio Hipofisário/biossíntese , Receptores de Hormônios Reguladores de Hormônio Hipofisário/genética , Estudos RetrospectivosRESUMO
Human ß-defensins (hBDs) stimulate degranulation in rat peritoneal mast cells in vitro and cause increased vascular permeability in rats in vivo. In this study, we sought to determine whether hBDs activate murine and human mast cells and to delineate the mechanisms of their regulation. hBD2 and hBD3 did not induce degranulation in murine peritoneal or bone marrow-derived mast cells (BMMC) in vitro and had no effect on vascular permeability in vivo. By contrast, these peptides induced sustained Ca(2+) mobilization and substantial degranulation in human mast cells, with hBD3 being more potent. Pertussis toxin (PTx) had no effect on hBD-induced Ca(2+) mobilization, but La(3+) and 2-aminoethoxydiphenyl borate (a dual inhibitor of inositol 1,4,5-triphosphate receptor and transient receptor potential channels) caused substantial inhibition of this response. Interestingly, degranulation induced by hBDs was substantially inhibited by PTx, La(3+), or 2-aminoethoxydiphenyl borate. Whereas human mast cells endogenously express G protein-coupled receptor, Mas-related gene X2 (MrgX2), rat basophilic leukemia, RBL-2H3 cells, and murine BMMCs do not. Silencing the expression of MrgX2 in human mast cells inhibited hBD-induced degranulation, but had no effect on anaphylatoxin C3a-induced response. Furthermore, ectopic expression of MrgX2 in RBL-2H3 and murine BMMCs rendered these cells responsive to hBDs for degranulation. This study demonstrates that hBDs activate human mast cells via MrgX2, which couples to both PTx-sensitive and insensitive signaling pathways most likely involving Gαq and Gαi to induce degranulation. Furthermore, murine mast cells are resistant to hBDs for degranulation, and this reflects the absence of MrgX2 in these cells.
Assuntos
Mastócitos/imunologia , Mastócitos/metabolismo , Proteínas do Tecido Nervoso/fisiologia , Receptores Acoplados a Proteínas G/fisiologia , Receptores de Neuropeptídeos/fisiologia , beta-Defensinas/fisiologia , Animais , Sinalização do Cálcio/genética , Sinalização do Cálcio/imunologia , Degranulação Celular/genética , Degranulação Celular/imunologia , Linhagem Celular , Células Cultivadas , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , Ratos , Receptores Acoplados a Proteínas G/biossíntese , Receptores Acoplados a Proteínas G/genética , Receptores de Neuropeptídeos/biossíntese , Receptores de Neuropeptídeos/genéticaRESUMO
Pheochromocytoma is a rare but potentially lethal chromaffin cell tumor with currently no effective treatment. Peptide hormone receptors are frequently overexpressed on endocrine tumor cells and can be specifically targeted by various anti-tumor peptide analogs. The present study carried out on mouse pheochromocytoma cells (MPCs) and a more aggressive mouse tumor tissue-derived (MTT) cell line revealed that these cells are characterized by pronounced expression of the somatostatin receptor 2 (sst2), growth hormone-releasing hormone (GHRH) receptor and the luteinizing hormone-releasing hormone (LHRH) receptor. We further demonstrated significant anti-tumor effects mediated by cytotoxic somatostatin analogs, AN-162 and AN-238, by LHRH antagonist, Cetrorelix, by the cytotoxic LHRH analog, AN-152, and by recently developed GHRH antagonist, MIA-602, on MPC and for AN-152 and MIA-602 on MTT cells. Studies of novel anti-tumor compounds on these mouse cell lines serve as an important basis for mouse models of metastatic pheochromocytoma, which we are currently establishing.
Assuntos
Neoplasias das Glândulas Suprarrenais/tratamento farmacológico , Feocromocitoma/tratamento farmacológico , Receptores de Neuropeptídeos/efeitos dos fármacos , 2-Hidroxifenetilamina/análogos & derivados , 2-Hidroxifenetilamina/farmacologia , Compostos de Anilina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/análogos & derivados , Doxorrubicina/farmacologia , Hormônio Liberador de Gonadotropina/análogos & derivados , Hormônio Liberador de Gonadotropina/antagonistas & inibidores , Hormônio Liberador de Gonadotropina/farmacologia , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Camundongos , Pirróis/farmacologia , Receptores LHRH/biossíntese , Receptores LHRH/efeitos dos fármacos , Receptores LHRH/metabolismo , Receptores de Neuropeptídeos/biossíntese , Receptores de Neuropeptídeos/metabolismo , Receptores de Hormônios Reguladores de Hormônio Hipofisário/biossíntese , Receptores de Hormônios Reguladores de Hormônio Hipofisário/efeitos dos fármacos , Receptores de Hormônios Reguladores de Hormônio Hipofisário/metabolismo , Receptores de Somatostatina/biossíntese , Receptores de Somatostatina/efeitos dos fármacos , Receptores de Somatostatina/metabolismo , Sermorelina/análogos & derivados , Sermorelina/farmacologia , Somatostatina/análogos & derivadosRESUMO
This study evaluated the effects of a modern antagonistic analog of GHRH on tumor growth and on expression of inflammatory cytokine genes in two models of human triple negative breast cancers (TNBC). The TNBC subtype is refractory to the treatment options available for other hormone-independent breast cancers. Inflammatory cytokines play a major role in the cellular signaling associated with breast cancer pathogenesis and enhance epithelial-mesenchymal transitions (EMT), drug resistance, and metastatic potential. Growth hormone-releasing hormone (GHRH) is a hypothalamic neuropeptide which regulates the synthesis and release of growth hormone by the pituitary and is an autocrine/paracrine growth factor for multiple human cancers. The effects of analogs of GHRH on tumoral cytokine expression have not been previously investigated. Animals bearing xenografts of the human TNBC cell lines, HCC1806 and MX-1, were treated with MIA-602, an antagonistic analog of GHRH. Treatment with MIA-602 significantly reduced tumor growth. We quantified transcript levels of the genes for several inflammatory cytokines. Expression of INFγ, IL-1α, IL-4, IL-6, IL-8, IL-10, and TNFα, was significantly reduced by treatment with MIA-602. We conclude that treatment of TNBC with GHRH antagonists reduces tumor growth through an action mediated by tumoral GHRH receptors and produces a suppression of inflammatory cytokine signaling. Silencing of GHRH receptors in vitro with siRNA inhibited the expression of GHRH-R genes and inflammatory cytokine genes in HCC1806 and MX-1 cells. Further studies on GHRH antagonists may facilitate the development of new strategies for the treatment of resistant cancers.
Assuntos
Neoplasias da Mama/tratamento farmacológico , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Citocinas/biossíntese , Citocinas/genética , Feminino , Expressão Gênica/efeitos dos fármacos , Inativação Gênica , Hormônio Liberador de Hormônio do Crescimento/genética , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Humanos , Camundongos , Camundongos Nus , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/genética , Receptor ErbB-2/metabolismo , Receptores de Estrogênio/metabolismo , Receptores de Neuropeptídeos/biossíntese , Receptores de Neuropeptídeos/genética , Receptores de Hormônios Reguladores de Hormônio Hipofisário/biossíntese , Receptores de Hormônios Reguladores de Hormônio Hipofisário/genética , Receptores de Progesterona/metabolismo , Sermorelina/análogos & derivados , Sermorelina/farmacologia , Transdução de Sinais , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Orexin A and orexin B (also known as hypocretins) are closely related peptides synthesized by hypothalamic neurons. They orchestrate diverse central and peripheral processes by stimulation of two G-protein coupled receptors, OX(1)R and OX(2)R. Recent studies have demonstrated the ability of orexins to promote a robust apoptosis in different cancer cells in culture and a potent growth reduction of human colon tumors in mice xenografts. Here we report effects of orexins on survival of rat C6 glioma cells, an experimental model for studies on glioblastoma multiforme (GBM). Quantitative real-time PCR demonstrated the expression of both types of orexin receptors in C6 cells. Orexin A and orexin B did not affect rat C6 glioma cell proliferation as assessed by [(3)H]thymidine incorporation assay. Incubation of the cells with orexin A (0.001-1 µM) resulted in a marked decrease of cell viability. The observed effect was caspase-dependent, as it was blocked by Z-VAD-fmk, a pan caspase inhibitor. In addition to that, a parallel increase in caspase-3 activity was observed. It is suggested that stimulation of orexin receptors induces death of rat C6 glioma cells through activation of caspase pathway.
Assuntos
Caspases/fisiologia , Glioma/patologia , Peptídeos e Proteínas de Sinalização Intracelular/farmacologia , Proteínas de Neoplasias/efeitos dos fármacos , Neuropeptídeos/farmacologia , Receptores Acoplados a Proteínas G/análise , Receptores Acoplados a Proteínas G/efeitos dos fármacos , Receptores de Neuropeptídeos/análise , Receptores de Neuropeptídeos/efeitos dos fármacos , Clorometilcetonas de Aminoácidos/farmacologia , Animais , Apoptose/efeitos dos fármacos , Inibidores de Caspase/farmacologia , Divisão Celular/efeitos dos fármacos , Linhagem Celular Tumoral/química , Linhagem Celular Tumoral/efeitos dos fármacos , Proteínas de Neoplasias/análise , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Receptores de Orexina , Orexinas , RNA Mensageiro/análise , RNA Neoplásico/análise , Ratos , Receptores Acoplados a Proteínas G/biossíntese , Receptores Acoplados a Proteínas G/genética , Receptores de Neuropeptídeos/biossíntese , Receptores de Neuropeptídeos/genéticaAssuntos
Adenocarcinoma/tratamento farmacológico , Apoptose/genética , Neoplasias Colorretais/tratamento farmacológico , Receptores Acoplados a Proteínas G/genética , Receptores de Neuropeptídeos/genética , Adenocarcinoma/genética , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral/efeitos dos fármacos , Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica , Genes Supressores de Tumor , Humanos , Injeções Intraperitoneais , Peptídeos e Proteínas de Sinalização Intracelular/administração & dosagem , Peptídeos e Proteínas de Sinalização Intracelular/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/uso terapêutico , Camundongos , Camundongos Nus , Neuropeptídeos/administração & dosagem , Neuropeptídeos/farmacologia , Neuropeptídeos/fisiologia , Neuropeptídeos/uso terapêutico , Receptores de Orexina , Orexinas , Receptores Acoplados a Proteínas G/biossíntese , Receptores de Neuropeptídeos/biossíntese , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
MICA is a ligand of the activating receptor NKG2D, expressed by NK and T cells. MICA expression is induced in cancer cells favoring their elimination by the immune system; however, many advanced tumors shed soluble MICA (sMICA), which impairs NKG2D-mediated cytotoxicity. ERp5 and GRP78 are endoplasmic reticulum-resident proteins that are translocated to the surface of epithelial tumor cells where they interact with MICA and are involved in sMICA shedding. In this study, we analyze the role of ERp5 and GRP78 in sMICA shedding in chronic lymphocytic leukemia (CLL). Immunofluorescence and flow cytometry analyses showed that ERp5 and GRP78 were significantly expressed on the surface of B cells and leukemia cells, but they were not expressed on T cells. The expression of ERp5 and GRP78 was significantly higher in leukemia cells than in B cells from controls. ERp5 and GRP78 co-localized with MICA on the surface of leukemia cells and the levels of expression of ERp5 and GRP78 correlated with the level of expression of membrane-bound MICA in CLL patients. Associated with higher expression of membrane-bound ERp5 and GRP78, serum sMICA levels were approximately threefold higher in patients than in controls. Elevated sMICA levels in CLL patients were associated with the down-modulation of NKG2D surface expression on CD8 T cells. Finally, pharmacological inhibition of B cell lines and stimulated leukemia cells showed that ERp5 activity is involved in sMICA shedding in CLL. In conclusion, these results uncover a molecular mechanism which regulates MICA protein shedding and immune evasion in CLL.
Assuntos
Antígenos de Histocompatibilidade Classe I/metabolismo , Leucemia Linfocítica Crônica de Células B/metabolismo , Isomerases de Dissulfetos de Proteínas/biossíntese , Receptores de Neuropeptídeos/biossíntese , Evasão Tumoral/fisiologia , Idoso , Chaperona BiP do Retículo Endoplasmático , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Imunofluorescência , Humanos , Leucemia Linfocítica Crônica de Células B/imunologia , Masculino , Microscopia ConfocalRESUMO
BACKGROUND: Recurrent abdominal pain is reported by a third of patients with diverticulosis, particularly those with previous episodes of acute diverticulitis. The current understanding of the etiology of this pain is poor. Our aim was to assess visceral sensitivity in patients with diverticular disease and its association with markers of previous inflammation and neuropeptides. METHODS: Patients with asymptomatic and symptomatic diverticular disease underwent a flexible sigmoidoscopy and biopsy followed 5-10 days later by visceral sensitivity testing with barostat-mediated rectal distension. Inflammation was assessed by staining of serotonin (5HT) and CD3 positive cells. mRNA levels of tumor necrosis factor alpha (TNF α) and interleukin-6 (IL-6) were quantitated using RT-PCR. Neuropeptide expression was assessed from percentage area staining with substance P (SP) and mRNA levels of the neurokinin 1 & 2 receptors (NK1 & NK2), and galanin 1 receptor (GALR1). KEY RESULTS: Thirteen asymptomatic and 12 symptomatic patients were recruited. The symptomatic patients had a lower first reported threshold to pain (28.4 mmHg i.q.r 25.0-36.0) than the asymptomatic patients (47 mmHg i.q.r 36.0-52.5, P < 0.001). Symptomatic patients had a higher median overall pain rating for the stimuli than the asymptomatic patients (P < 0.02). Symptomatic patients had greater median relative expression of NK1 and TNF alpha mRNA compared with asymptomatic patients. There was a significant correlation between barostat VAS pain scores and NK 1 expression (Figure 4, r(2) 0.54, P < 0.02). CONCLUSIONS & INFERENCES: Patients with symptomatic diverticular disease exhibit visceral hypersensitivity, and this may be mediated by ongoing low grade inflammation and upregulation of tachykinins.
Assuntos
Dor Abdominal/etiologia , Doença Diverticular do Colo/complicações , Diverticulose Cólica/complicações , Neuropeptídeos/biossíntese , Dor Abdominal/metabolismo , Dor Abdominal/patologia , Idoso , Doença Diverticular do Colo/metabolismo , Doença Diverticular do Colo/patologia , Diverticulose Cólica/metabolismo , Diverticulose Cólica/patologia , Feminino , Humanos , Hiperestesia/etiologia , Hiperestesia/metabolismo , Hiperestesia/patologia , Inflamação/complicações , Inflamação/metabolismo , Inflamação/patologia , Masculino , Pessoa de Meia-Idade , Limiar da Dor/fisiologia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase em Tempo Real , Receptores de Neuropeptídeos/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Vísceras/metabolismo , Vísceras/patologiaRESUMO
Growth hormone secretion from the anterior pituitary gland is controlled by interactions between three hormone receptors, between GHRH and GHRH receptor (GHRH-R), between ghrelin and growth hormone secretagogue receptor (GHS-R1a), and between somatostatin and somatostatin receptors in the hypothalamus and anterior pituitary gland. Ghrelin-GHS-R1a is involved in many important functions, including GH secretion and appetite. We investigated age-related changes in the expressions of GHS-R1a, GHS-R1b (the truncated-type receptor), and GHRH-R mRNAs by real-time reverse transcription-PCR using 16 tissues, leukocytes, oocytes, and cumulus cells in Holstein-Friesian cattle. The tissue samples were divided into three age classes: 1) 19 to 26 d of age (preweaning calves), 2) 2 mo to 6.5 mo of age (postweaning calves), and 3) 3.2 to 8.1 yr of age (cows). The GHS-R1a mRNA was highly (P < 0.05) expressed in the arcuate nucleus, pituitary gland, and liver compared with that of the other tissues in all age classes. Expression of GHS-R 1a mRNA in the arcuate nucleus of postweaning calves was > 10-fold greater (P < 0.01) than those of preweaning calves and cows, and its expression level was the greatest (P < 0.01) in all tissues examined in age group 2. GHS-R1a and GHRH-R mRNA expressions in the pituitary gland of preweaning calves tended to be greater (P < 0.20 and P < 0.17, respectively) than those of postweaning calves and cows. GHS-R1b mRNA expression was detected in all tissues examined, and abundance was greater (P < 0.05) in the pancreas, pituitary gland, spleen, arcuate nucleus, adipose tissue, and leukocyte compared with that of the other tissues examined in age group 3. Interestingly, a relatively large animal-to-animal variation was observed in pancreas GHS-R 1b mRNA expression. The GHRH-R mRNA was markedly increased (P < 0.01) in the pituitary gland in all age groups compared with that of the other tissues. GHRH-R mRNA abundance in the arcuate nucleus, pituitary gland, liver, spleen, adipose tissue, and heart of preweaning calves tended to be greater than those of postweaning calves and cows. The GHRH-R mRNA was not detected in the mammary gland and adipose tissue of nonlactating cows.
Assuntos
Núcleo Arqueado do Hipotálamo/fisiologia , Bovinos/genética , Regulação da Expressão Gênica , Hormônio do Crescimento/biossíntese , Adeno-Hipófise/fisiologia , Receptores de Grelina/biossíntese , Receptores de Neuropeptídeos/biossíntese , Receptores de Hormônios Reguladores de Hormônio Hipofisário/biossíntese , Fatores Etários , Animais , Bovinos/metabolismo , Feminino , Hormônio do Crescimento/genética , Hormônio do Crescimento/metabolismo , Análise dos Mínimos Quadrados , Masculino , Adeno-Hipófise/metabolismo , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Receptores de Grelina/genética , Receptores de Neuropeptídeos/genética , Receptores de Hormônios Reguladores de Hormônio Hipofisário/genéticaRESUMO
BACKGROUND: Delayed gastric emptying (DGE) is still a common postoperative complication after pancreaticoduodenectomy (PD). Because different reconstruction techniques after PD and the influence of motilin receptor expression are controversially discussed, the present study analyzed the influence of a total orthotopic reconstruction technique on DGE after PD. METHODS: Data from patients undergoing PD and reconstruction using a total orthotopic technique were reviewed, and correlations between DGE and clinico-pathological variables were analyzed. Motilin receptor expression was measured within the duodenum, jejunum, and terminal ileum. RESULTS: Three hundred seven patients received orthotopic reconstruction using a single jejunal loop. DGE grade B or C could be observed in 16.6% of the patients. DGE was significantly associated with the severity of a postoperative pancreatic fistula, the need for a reoperation, wound infections, and vascular complications. Furthermore, these parameters correlated significantly with the grade of DGE. The density of motilin receptor expression decreased significantly behind the duodenum in aboral direction. CONCLUSIONS: The orthotopic reconstruction after PD is the shortest distance without resection of a jejunal segment, preserves the greatest length of jejunum and thus the highest density of motilin receptors, and should therefore be recommended to reduce the incidence of DGE after PD.
Assuntos
Esvaziamento Gástrico/fisiologia , Gastroparesia/etiologia , Jejuno/metabolismo , Pancreaticoduodenectomia/efeitos adversos , Procedimentos de Cirurgia Plástica/efeitos adversos , Receptores dos Hormônios Gastrointestinais/biossíntese , Receptores de Neuropeptídeos/biossíntese , Biomarcadores/metabolismo , Feminino , Seguimentos , Gastroparesia/metabolismo , Gastroparesia/fisiopatologia , Humanos , Imuno-Histoquímica , Jejuno/cirurgia , Masculino , Pessoa de Meia-Idade , Pancreaticoduodenectomia/métodos , Estudos Prospectivos , Procedimentos de Cirurgia Plástica/métodosRESUMO
Resistance to apoptosis is a recurrent theme in colon cancer. We have shown previously that the 7-transmembrane spanning receptor OX1R for orexins promotes robust apoptosis in the human colon cancer cell line HT29 through an entirely novel mechanism involving phosphorylation of tyrosine-based motifs in OX1R. Here, we investigated the status of OX1R in a large series of human colorectal tumors and hepatic metastases. All primary colorectal tumors regardless of their localization and Duke's stages and all hepatic metastases tested expressed OX1R mRNA and/or protein. In sharp contrast, adjacent normal colonocytes or hepatocytes as well as control normal tissues were negative. Next, we showed that nine human colon cancer cell lines established from primary tumors or metastases expressed OX1R mRNA and underwent important apoptosis on orexin-A challenge. Most interestingly, orexin-A also promoted robust apoptosis in cells that are resistant to the most commonly used drug in colon cancer chemotherapy, 5-fluorouracil. When human colon cancer cells were xenografted in nude mice, orexin-A administered at day 0 strongly slowed the tumor growth and even reversed the development of established tumors when administered 7 days after cell inoculation. Orexin-A also acts by promoting tumor apoptosis in vivo because caspase-3 is activated in tumors on orexin treatment of nude mice. These findings support that OX1R is an Achilles heel of colon cancers, even after metastasis or chemoresistance. They suggest that OX1R agonists might be novel candidates for colon cancer therapy.
Assuntos
Apoptose/fisiologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/secundário , Neuropeptídeos/metabolismo , Receptores Acoplados a Proteínas G/biossíntese , Receptores de Neuropeptídeos/biossíntese , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/secundário , Animais , Células CACO-2 , Linhagem Celular Tumoral , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/genética , Resistencia a Medicamentos Antineoplásicos , Fluoruracila/farmacologia , Células HCT116 , Células HT29 , Humanos , Neoplasias Hepáticas/genética , Receptores de Orexina , Orexinas , Ratos , Receptores Acoplados a Proteínas G/genética , Receptores de Neuropeptídeos/genéticaRESUMO
In search for the new polypeptides responsible for energy homeostasis which are also involved in regulating the growth and function of the human prostate, we assessed the expression of orexins (OXs) and of orexin receptors (OXRs) in human normal prostate and in benign prostatic hyperplasia (BPH). Conventional RT-PCR revealed the expression of OXR2 in all studied samples obtained either from normal prostates or BPH ones while neither preproorexin (ppOX)nor OXR1 mRNA were detected. In adenomatous prostates, expression levels of OXR2 were 30- to 40-fold higher compared to controls. Western blot analysis demonstrated the presence of OXR2 protein in the studied samples and its expression levels were 4-fold higher in tissue samples from BPH. In normal glands, presence of OXR2-like immunoreactivity was found in the apical parts of epithelial cells as well as in smooth muscle cells of the stroma. Immunostaining for OXR2 was more intense in sections obtained from BPH. Immunohistochemistry did not detect the expression of OXR1-like protein. OXA serum concentrations were lowered in BPH patients (mean ± SE 56±4 ng/ml, n=12; P<0.01) and unaltered in prostate cancer (79±7 ng/ml, n=18) compared to the controls (69±2 ng/ml, n=16). On the contrary, serum OXB levels were similar in all studied groups of patients. We thus have demonstrated the mRNA and protein expression of OXR2, but not of ppOX and OXR1 in both normal and BPH human prostate glands. We also demonstrated notable up-regulation of OXR2 in benign prostatic hyperplasia, an alteration accompanied by lowered serum OXA concentrations. These findings suggest that both OXA and OXR2 may be involved in the pathogenesis and/or maintenance of BPH.
Assuntos
Células Epiteliais/metabolismo , Regulação da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intracelular/sangue , Miócitos de Músculo Liso/metabolismo , Neuropeptídeos/sangue , Hiperplasia Prostática/sangue , Receptores Acoplados a Proteínas G/biossíntese , Receptores de Neuropeptídeos/biossíntese , Idoso , Células Epiteliais/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Miócitos de Músculo Liso/patologia , Receptores de Orexina , Orexinas , Hiperplasia Prostática/patologiaRESUMO
Hypothalamic growth hormone (GH)-releasing hormone (GHRH) regulates the release of GH from the pituitary gland. The receptors for GHRH (GHRH-R) are expressed predominantly in the pituitary. Recent evidence demonstrates that splice variants of the GHRH receptor are also expressed in several nonpituitary tissues, both normal and tumoral, as well as in cancer cell lines. The aim of this study was to investigate the expression of the splice variant 1 (SV-1) of GHRH-R in colorectal cancer (CRC). Seventy patients who underwent partial colectomy for CRC were enrolled in the study. Immunohistochemical expression of SV-1 was studied in paraffin-embedded sections of patient tumor tissue. A cytoplasmic supranuclear expression of SV-1 was observed in CRC as well as in the normal colon mucosa. Tumor grade and pathological stage were negatively correlated with expression of SV-1 (P = 0.012 and P = 0.013, respectively). CRCs metastatic to the liver showed a lower expression of SV-1 than did primary tumors, but this difference was not statistically significant. Kaplan-Meier and Cox univariate survival analyses indicated an improved survival time in patients with high SV-1 compared with those with low GHRH-R expression, but this difference was not statistically significant. The immunohistochemical expression of SV-1 seems to be a favorable prognostic factor in CRC.
Assuntos
Biomarcadores Tumorais/biossíntese , Neoplasias Colorretais/metabolismo , Receptores de Neuropeptídeos/biossíntese , Receptores de Hormônios Reguladores de Hormônio Hipofisário/biossíntese , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteína Axina , Caderinas/metabolismo , Estudos de Coortes , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/patologia , Proteínas do Citoesqueleto/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Metástase Neoplásica , Estadiamento de Neoplasias , Prognóstico , Modelos de Riscos Proporcionais , Isoformas de Proteínas , Estatísticas não ParamétricasRESUMO
Triple-negative breast cancers do not express receptors for estrogen or progesterone and do not overexpress HER2. These tumors have an unfavorable prognosis and at present chemotherapy is the only treatment option. Because the antagonists of growth hormone-releasing hormone (GHRH) have been shown to inhibit growth of a variety of cancers by endocrine and paracrine/autocrine mechanisms, we evaluated the expression of GHRH receptors in human specimens of triple-negative breast cancers and the response to GHRH by in vitro models. In samples of triple-negative breast cancers we found mRNA expression for the GHRH receptor and its functional splice variant SV1 in 25 and 70% of the cases, respectively and for GHRH in 80% of the samples. Immunoreaction of SV1 was detected in the human triple-negative breast cancer cell line HCC1806 while HCC1937 was negative. The growth of HCC1806 was stimulated by GHRH(1-44)NH(2) and inhibited by GHRH antagonist MZ-J-7-118. In addition, in HCC1806 MAP-kinases ERK-1/2 were activated by GHRH. Our findings suggest the existence of an autocrine loop consisting of GHRH and GHRH receptors in triple-negative breast cancers. Our in vitro studies demonstrate that targeting the GHRH receptor may be a therapeutic option which should be evaluated in studies in vivo.