RESUMO
Eosinophilic esophagitis (EoE) is a chronic inflammatory condition of the esophagus characterized by increased number of eosinophils. Currently, EoE diagnosis is based on endoscopic procedures for histopathological examination, eosinophils' counting and, often, in clinical practice, the challenge is the differentiation between EoE and gastroesophageal reflux disease (GERD). Our aim was to develop novel peptide ligand to Eosinophil cationic protein (ECP) present in EoE biopsies of patients with potential to be used for detection. We performed a comparative proteomic analysis using liquid chromatography-tandem mass spectrometry (LC-MS/MS) of esophageal biopsies from pediatric patients with eosinophilic esophagitis, gastroesophageal reflux disease and control individuals. Then, phage display technology was used to select peptides against specific up-regulated protein from EoE patients. Twelve phage clones were selected after three biopanning rounds, and the best phage clone reactivity was evaluated by phage-ELISA assay using esophageal mucus samples from 94 pediatric patients. Mass spectrometry showed that eosinophil cationic protein (ECP) was one of the most up-regulated proteins in EoE patients, which is an eosinophil granule protein usually deposited on tissues to mediate remodeling, but in excess may cause fibrosis and hypertrophy, especially in allergic responses. A highly reactive ECP-ligand peptide (E5) was able to distinguish reactive mucus of EoE patients from GERD and the control individuals by Phage-ELISA, achieving a sensitivity of 84.62%, and a specificity of 82.72%. This is the first study that successfully demonstrated an antibody-like peptide targeting ECP at the esophagus mucus as a useful auxilliary tool for EoE diagnosis with a significant association with atopic disorders and dysphagia.ClinicalTrials.gov no.: NCT03069573.
Assuntos
Esofagite Eosinofílica , Refluxo Gastroesofágico , Criança , Cromatografia Líquida , Enterite , Proteína Catiônica de Eosinófilo , Eosinofilia , Esofagite Eosinofílica/diagnóstico , Esofagite Eosinofílica/patologia , Eosinófilos/metabolismo , Gastrite , Refluxo Gastroesofágico/complicações , Humanos , Ligantes , Muco/metabolismo , Peptídeos , Proteômica , Espectrometria de Massas em TandemRESUMO
Epidermal growth factor receptor is a cancer driver whose nuclear localization has been associated with the progression of prostate cancer to the castration-resistant phenotype. Previous reports indicated a functional interaction between this receptor and the protein Annexin A1, which has also been associated with aggressive tumors. The molecular pathogenesis of castration-resistant prostate cancer remains largely unresolved, and herein we have demonstrated the correlation between the expression levels and localization of the epidermal growth factor receptor and Annexin A1 in prostate cancer samples and cell lines. Interestingly, a higher expression of both proteins was detected in castration-resistant prostate cancer cell lines and the strongest correlation was seen at the nuclear level. We verified that Annexin A1 interacts with the epidermal growth factor receptor, and by using prostate cancer cell lines knocked down for Annexin A1, we succeeded in demonstrating that Annexin A1 promotes the nuclear localization of epidermal growth factor receptor. Finally, we showed that Annexin A1 activates an autocrine signaling in castration-resistant prostate cells through the formyl peptide receptor 1. The inhibition of such signaling by Cyclosporin H inhibits the nuclear localization of epidermal growth factor receptor and its downstream signaling. The present work sheds light on the functional interaction between nuclear epidermal growth factor receptor and nuclear Annexin A1 in castration-resistant prostate cancer. Therefore, strategies to inhibit the nuclear localization of epidermal growth factor receptor through the suppression of the Annexin A1 autocrine loop could represent an important intervention strategy for castration-resistant prostate cancer.
Assuntos
Anexina A1/metabolismo , Núcleo Celular/metabolismo , Neoplasias de Próstata Resistentes à Castração/metabolismo , Receptores Androgênicos/metabolismo , Idoso , Anexina A1/genética , Comunicação Autócrina/fisiologia , Linhagem Celular Tumoral , Receptores ErbB/genética , Receptores ErbB/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/genética , Transdução de SinaisRESUMO
Cyclo-Gly-Pro (CGP) attenuates nociception, however its effects on salivary glands remain unclear. In this study, we investigated the acute effects of CGP on salivary flow and composition, and on the submandibular gland composition, compared with morphine. Besides, we characterized the effects of naloxone (a non-selective opioid receptor antagonist) on CGP- and morphine-induced salivary and glandular alterations in mice. After that, in silico analyses were performed to predict the interaction between CGP and opioid receptors. Morphine and CGP significantly reduced salivary flow and total protein concentration of saliva and naloxone restored them to the physiological levels. Morphine and CGP also reduced several infrared vibrational modes (Amide I, 1687-1594cm-1; Amide II, 1594-1494cm-1; CH2/CH3, 1488-1433cm-1; C = O, 1432-1365cm-1; PO2 asymmetric, 1290-1185cm-1; PO2 symmetric, 1135-999cm-1) and naloxone reverted these alterations. The in silico docking analysis demonstrated the interaction of polar contacts between the CGP and opioid receptor Cys219 residue. Altogether, we showed that salivary hypofunction and glandular changes elicited by CGP may occur through opioid receptor suggesting that the blockage of opioid receptors in superior cervical and submandibular ganglions may be a possible strategy to restore salivary secretion while maintaining antinociceptive action due its effects on the central nervous system.