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  • br Conclusion In this work experiments were carried out

    2019-11-07


    Conclusion In this work, experiments were carried out to purify the protein with COX activity, using an anti-mouse COX-2 pAb and a COX activity assay as tools for monitoring the protein responsible for this activity in the parasite. Gp63 enrichment through the affinity column and identification by MS, together with immunoprecipitation with a commercial anti-gp63 mAb, and detection of COX activity with a recombinant gp63, undoubtedly showed that L. mexicana gp63 is the enzyme responsible for cyclooxygenase activity in this parasitic protozoon.
    Acknowledgments This work was supported by CONACYT grant No. 104108 to PTR. LAEF and JADG were recipients of a fellowship from CONACYT (204814 and 173691 respectively). We thank Dr. Sergio Encarnación-Guevara and Magdalena Hernández-Ortiz from the Centro de Ciencias Genómicas, UNAM, México for their help with the MS analysis of the immunoprecipitation sample. We also thank Mélida del Rosario Lizarazo-Taborda for her help with COX activity and immunoprecipitation experiments and Belém de Luna and Ma. Elena Cisneros-Albavera for their technical assistance.
    Introduction Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive and fibrosing interstitial lung disease [1], in which normal lung parenchyma is progressively replaced by altered extracellular matrix (ECM), and alveolar architecture is destroyed, leading to decreased lung compliance, disrupted gas exchange, and ultimately respiratory failure, and death [2,3]. IPF occurs worldwide and its prevalence appears to be increasing. The incidence of IPF has a range of 2.8–9.3 cases per 100, 000 person-years as estimated. Its prevalence is higher in North American and Europe (3–9 cases per 100, 000 person-years) than that in South America and East Asia (fewer than 4 cases per 100, 000 person-years) [1,4]. IPF is a quintessential aging-related disease because it occurs primarily in elderly with a median diagnosis age at 65 [5]. Aging-related mechanisms, such as genomic instability, telomere attrition, epigenetic alterations, mitochondrial dysfunction, and cellular senescence, have been proposed as a potential pathogenetic driver for the onset of IPF [6]. To date, the cause of idiopathic pulmonary fibrosis is unknown, and no pharmacologic therapies have been shown to improve survival [7]. Cellular senescence is a permanent Heparin and proliferative arrest, triggered by telomere attrition, DNA damage, and genomic and proteomic instability that plays a role in several complex biological processes, such as development, tissue repair, aging, and age-related disorders [8]. Senescent cells undergo growth arrest, and importantly, they retain viability and have metabolic activity, such as morphological transformation, activation of P16INK4a and P21, increase in senescence-associated β-galactosidase (SA-β-Gal) activity, and changes in dominant senescence-associated secretory phenotype (SASP). This event includes abundant cytokines and chemokines [9,10]. Mounting evidence has emerged that cellular senescence might be detrimental to IPF patients, and elimination of senescent cells might be beneficial for the treatment of IPF [11,12]. Though the mechanism underlying the cellular senescence has so far been mediated through interplay of multiple pathways, accumulating data has displayed that tumor suppressor protein 53 (P53) that modulates cell cycle, DNA damage response and apoptosis, could regulate cellular senescence [[13], [14], [15]]. However, whether the P53 could regulate cellular senescence in pulmonary fibrosis remains unexplored. Lung fibroblasts, as the predominant effector cells, play a vital role in the development of pulmonary fibrosis [16]. In response to various stimuli, quiescent lung fibroblasts are activated and trans-differentiate into myofibroblasts, producing large amounts of ECM, pro-inflammation cytokines, and profibrotic factors. Intriguingly, senescent fibroblasts overexpress α-SMA to exhibit a myofibroblast-like phenotype [17], and secrete lots of profibrotic factors to enhance the pulmonary fibrosis [18]. Thus, the development of new strategies to mediate senescent fibroblasts might be important for the treatment of pulmonary fibrosis. It has been reported that prostaglandin (PG) E2, as the major prostanoid in the lungs, is an important anti-proliferative and anti-fibrotic lipid mediator in pulmonary fibrosis [[19], [20], [21], [22], [23]]. In IPF patients, levels of PGE2 was reduced, due to the failure to up-regulate the expression of key inducible enzyme cyclooxygenase (COX)-2 [24,25]. Studies performed on the mechanism for the antifibrotic actions of the fibrinolytic pathway demonstrated that plasminogen activation exerted anti-fibrotic effects in pulmonary fibrosis via COX-2/PGE2 synthesis pathway [26]. Moreover, COX-2 was involved in the occurrence of cellular senescence [27,28]. While, there is still a lot of room to investigate the regulation of COX-2 on senescent fibroblasts in pulmonary fibrosis.