Project description:Echinoderms, possessing outstanding regenerative capability, provide unique model system for the study of the response to injury. However, there is little known about the proteomic composition of coelomic fluid, an important biofluid circulating through the whole body and reflecting an overall biological status of the organism. In this study, we used LC-MALDI tandem mass spectrometry to characterize proteome of cell-free coelomic fluid of starfish Asterias rubens and follow the changes occurring in response to puncture wound and blood loss. Our study demonstrated significant changes of CF proteome during the first hours after injury and presented a series of candidate proteins involved in early response to injury, providing interesting targets for future functional studies.
Project description:The origin of cells involved in regeneration in Echinoderms is an open question till now. Most experimental data support the process of transdifferentiation as the main mechanism, although the activity of progenitor/stem cells is also considered. The renewal of coelomocytes, the cells which occupy the coelomic cavity in Asterias rubens is an example of physiological regeneration. Although it is considered that coelomic epithelium is the main source of coelomocytes, many details of this process remain unclear. In this study, we provide evidence of the origin of several types of coelomocytes in A. rubens from the coelomic epithelium, with particular emphasis on small undifferentiated cells. The concept of a pool of marginal coelomocytes is introduced as cells sequestered on the surface of the coelomic epithelium and characterized by a special cellular composition. Proteomic analysis by LC-MALDI TOF/TOF MS identified in total 403 proteins in coelomocytes (COE), coelomic epithelium (CE) and coelomic epithelium subpopulation enriched with undifferentiated cells (CEW). Proteins common and associated with each cell pool were revealed and the intermediate status of cells on the border of coelomic epithelium and coelom was confirmed. The potential players involved in regenerative processes were identified.
Project description:Coelomic epithelium (CE), or coelothelium, is a mesodermal-derived tissue lining coelomic cavities and enveloping inner organs in all coelomates, including vertebrates, where it forms both peritoneum and pericardium, as well as pleura, in mammals. Due to its histogenetic potential, CE is considered a key-tissue characterized by unique features. At present, in adult echinoderms, CE is supposed to play fundamental key roles acting as: 1) source of stem/multipotent cells (undifferentiated coelomocytes), particularly during regenerative phenomena (Candia Carnevali et al., 2009; Candia Carnevali and Burighel, 2010; Hernroth et al., 2010; García-Arrarás et al., 2011). 2) haematopoietic tissue, being the direct source of circulating elements (differentiated coelomocytes) including immune cells, indispensable elements during the first repair events of clotting and defence against pathogens (Boassche and Jangoux, 1976; Munoz-Chapuli et al., 2005; Pinsino et al., 2007). Despite these indications, the histological approach alone, microscopic or ultrastructural, is not sufficient to fully attribute a haematopoietic role to the CE and even the presumptive production/release process of stem/pluripotent cells from coelothelium during regeneration need to be confirmed as well. As a matter of fact, it is still unclear if the intense proliferation detectable in regenerating CE is functional to the production of wandering coelomocytes necessary to restore the post-traumatic fluid loss or rather to produce truly regeneration-competent cells used as building blocks of the regenerating tissues. In order to overcome some of these issues and deeply understand the complex physiological role of this epithelial tissue in echinoderm biology, it is important to integrate morphological evidences with detailed molecular data. In particular, proteomics is becoming a powerful tool to deepen the problem of cell and tissue functions during both tissue homeostasis and regeneration by depicting the existing protein activities involved in the regrowth processes. However, proteomics investigations focussed on echinoderm CE are quite limited. Specific molecular information are due to the works by Gabre and co-workers (2015) and by Kim and co-workers (2018), which are addressed to the characterization of CE transcriptome from the clonal starfish Coscinasterias muricata and the blue bat starfish Patiria pectinifera, respectively. The analysis of the biological processes associated to the transcripts identified in these studies suggests the involvement of CE in the development of anatomical units. More recently the CE proteome of the common starfish Asterias rubens was also published (Sharlaimova et al., 2021). Despite these evidences, there is a great lack of integrated multidisciplinary studies that can contribute to unravel the complex roles exerted by this tissue both in regeneration and other physiological processes. In order to partially unravel this gap, the present work is addressed to analyse the CE of the starfish Marthasterias glacialis in both standard physiological (homeostatic) and regenerating conditions, by combining histological/ultrastructural analyses with appropriate proteomics approaches, the former to provide a detailed cell and tissue perspective, the latter to contribute a base-line molecular view of CE multifunctional implications. This integrated approach can significantly help to shed new light into the intriguing role of this apparently simple but fundamental tissue.
Project description:Background: Atopic dermatitis (AD) is the most common chronic inflammatory skin disease, but its complex pathogenesis is only insufficiently understood, resulting in still limited treatment options. Objective: We sought to characterize AD on both transcriptomic and proteomic levels in humans. Methods: We used skin suction blistering, a painless and nonscarring procedure that can simultaneously sample skin cells and interstitial fluid. We then compared results with conventional biopsies. Results: Suction blistering captured epidermal and most immune cells equally well as biopsies, except for mast cells and nonmigratory CD163+ macrophages that were only present in biopsy isolates. Using single-cell RNA sequencing, we found comparable transcriptional profiles of key inflammatory pathways between blister and biopsy AD, but suction blistering was superior in cell-specific resolution for high-abundance transcripts (KRT1/KRT10, KRT16/KRT6A, S100A8/S100A9), which showed some background signals in biopsy isolates. Compared with healthy controls, we found characteristic upregulation of AD-typical cytokines such as IL13 and IL22 in Th2 and Th22 cells, respectively, but we also discovered these mediators in proliferating T cells and natural killer T cells, that also expressed the antimicrobial cytokine IL26. Overall, not T cells, but myeloid cells were most strongly enriched in AD, and we found dendritic cell (CLEC7A, amphiregulin/AREG, EREG) and macrophage products (CCL13) among the top upregulated proteins in AD blister fluid proteomic analyses. Conclusion: These data show that by using cutting-edge technology, suction blistering offers several advantages over conventional biopsies, including better transcriptomic resolution of skin cells, combined with proteomic information from interstitial fluid, unraveling novel inflammatory players that shape the cellular and proteomic microenvironment of AD.
Project description:It was found that Vibrio splendidus could survive under high concentration of tetracycline, and the coelomic fluid of sea cucumber increased the tolerance of Vibrio splendidus to tetracycline. Therefore, the transcriptome was determined to find the cause of drug resistance in Vibrio splendidus.