RESUMO
Extracellular nucleotides can regulate cell proliferation in both normal and tumorigenic tissues. Here, we studied how extracellular nucleotides regulate the proliferation of ZL55 cells, a mesothelioma-derived cell line obtained from bioptic samples of asbestos-exposed patients. ADP and 2-MeS-ADP inhibited ZL55 cell proliferation, whereas ATP, UTP, and UDP were inactive. The nucleotide potency profile and the blockade of the ADP-mediated inhibitory effect by the phospholipase C inhibitor U-73122 suggest that P2Y1 receptor controls ZL55 cell proliferation. The activation of P2Y1 receptor by ADP leads to activation of intracellular transduction pathways involving [Ca2+ ]i , PKC-δ/PKC-α, and MAPKs, ERK1/2 and JNK1/2. Cell treatment with ADP or 2-MeS-ADP also provokes the activation of p53, causing an accumulation of the G1 cyclin-dependent kinase inhibitors p21WAF1 and p27Kip . Inhibition of ZL55 cell proliferation by ADP was completely reversed by inhibiting MEK1/2, or JNK1/2, or PKC-δ, and PKC-α. Through the inhibition of ADP-activated transductional kinases it was found that PKC-δ was responsible for JNK1/2 activation. JNK1/2 has a role in transcriptional up-regulation of p53, p21WAF1/CIP1 , and p27kip1 . Conversely, the ADP-activated PKC-α provoked ERK1/2 phosphorylation. ERK1/2 increased p53 stabilization, required to G1 arrest of ZL55 cells. Concluding, the importance of the study is twofold: first, results shed light on the mechanism of cell cycle inhibition by ADP; second, results suggest that extracellular ADP may inhibit mesothelioma progression.
Assuntos
Difosfato de Adenosina/farmacologia , Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Mesotelioma/tratamento farmacológico , Proteína Quinase C-alfa/metabolismo , Proteína Quinase C-delta/metabolismo , Agonistas do Receptor Purinérgico P2Y/farmacologia , Receptores Purinérgicos P2Y1/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Difosfato de Adenosina/análogos & derivados , Amianto/efeitos adversos , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Ativação Enzimática , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Mesotelioma/enzimologia , Mesotelioma/genética , Mesotelioma/patologia , Fosforilação , Proteína Quinase C-alfa/genética , Proteína Quinase C-delta/genética , Estabilidade Proteica , Interferência de RNA , Receptores Purinérgicos P2Y1/metabolismo , Tionucleotídeos/farmacologia , Fatores de Tempo , Transfecção , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
Ulcerative colitis (UC) and Crohn's disease (CD) represent the two main forms of chronic inflammatory bowel diseases (IBD). The exact IBD etiology is not yet revealed but CD and UC are likely induced by an excessive immune response against normal constituents of the intestinal microbial flora. IBD diagnosis is based on clinical symptoms often combined with invasive and costly procedures. Thus, the need for more non-invasive markers is urgent. Several routine laboratory investigations have been explored as indicators of intestinal inflammation in IBD, including blood testing for C-reactive protein, erythrocyte sedimentation rate, and specific antibodies, in addition to stool testing for calprotectin and lactoferrin. However, none has been universally adopted, some have been well-characterized, and others hold great promise. In recent years, the technological developments within the field of mass spectrometry (MS) and bioinformatics have greatly enhanced the ability to retrieve, characterize, and analyze large amounts of data. High-throughput research allowed enhancing the understanding of the biology of IBD permitting a more accurate biomarker discovery than ever before. In this review, we summarize currently used IBD serological and stool biomarkers and how proteomics and lipidomics are contributing to the identification of IBD biomarkers.