Secretome Analysis of Rabbit and Human Mesenchymal Stem and Endothelial Progenitor Cells: A Comparative Study

Human adipose tissue-derived mesenchymal stem cells (AT-MSCs) have been studied several years for their immunomodulatory effect through the paracrine mechanism and cytokine secretion. In combination with endothelial progenitor cells (EPCs), MSCs have great therapeutical potential for the repair of endothelium and wound healing. However, little is known about the cytokine profile of rabbit AT-MSCs or even EPCs. The aim of this study was to analyze the secretomes of these rabbit stem/progenitor cells. A large-scale human cytokine array (up to 80 cytokines) was used to identify and compare cytokines secreted into conditioned media of human and rabbit AT-MSCs as well as HUVECs and rabbit EPCs. Few cytokines were highly expressed by human AT-MSCs (TIMP-2, TIMP-1), HUVECs (MCP-1, TIMP-2, GRO, Angiogenin, IL-8, TIMP-1), or by rabbit EPCs (TIMP-2). Several cytokines have moderate expression by human (MCP-1, GRO, Angiogenin, TGF-β 2, IL-8, LIF, IL-6, Osteopontin, Osteoprotegerin) and rabbit AT-MSCs (TIMP-2, TGF-β 2, LIF, Osteopontin, IL-8, IL-5, IL-3) or by HUVECs (IL-6, MIF, TGF-β 2, GCP-2, IGFBP-2, Osteoprotegerin, EGF, LIF, PDGF-BB, MCP-3, Osteopontin, Leptin, IL-5, ENA-78, TNF- β) and rabbit EPCs (TGF-β 2, Osteopontin, GRO, LIF, IL-8, IL-5, IL-3). In conclusion, the proposed method seems to be useful for the secretome analysis of rabbit stem/progenitor cells.

In Vitro Secretome Analysis Suggests Differential Pathogenic Mechanisms between Fusarium oxysporum f. sp. cubense Race 1 and Race 4

Banana Fusarium wilt, caused by the fungus pathogen Fusarium oxysporum f. sp. cubense (Foc), is a devastating disease that causes tremendous reductions in banana yield worldwide. Secreted proteins can act as pathogenicity factors and play important roles in the Foc–banana interactions. In this study, a shotgun-based proteomic approach was employed to characterize and compare the secretomes of Foc1 and Foc4 upon banana extract treatment, which detected 1183 Foc1 and 2450 Foc4 proteins. Comprehensive in silico analyses further identified 447 Foc1 and 433 Foc4 proteins in the classical and non-classical secretion pathways, while the remaining proteins might be secreted through currently unknown mechanisms. Further analyses showed that the secretomes of Foc1 and Foc4 are similar in their overall functional characteristics and share largely conserved repertoires of CAZymes and effectors. However, we also identified a number of potentially important pathogenicity factors that are differentially present in Foc1 and Foc4, which may contribute to their different pathogenicity against banana hosts. Furthermore, our quantitative PCR analysis revealed that genes encoding secreted pathogenicity factors differ significantly between Foc1 and Foc4 in their expression regulation in response to banana extract treatment. To our knowledge, this is the first experimental secretome analysis that focused on the pathogenicity mechanism in different Foc races. The results of this study provide useful resources for further exploration of the complicated pathogenicity mechanisms in Foc.

Genome and Secretome Analysis of Staphylotrichum longicolleum DSM105789 Cultured on Agro-Residual and Chitinous Biomass

Staphylotrichum longicolleum FW57 (DSM105789) is a prolific chitinolytic fungus isolated from wood, with a chitinase activity of 0.11 ± 0.01 U/mg. We selected this strain for genome sequencing and annotation, and compiled its growth characteristics on four different chitinous substrates as well as two agro-industrial waste products. We found that the enzymatic mixture secreted by FW57 was not only able to digest pre-treated sugarcane bagasse, but also untreated sugarcane bagasse and maize leaves. The efficiency was comparable to a commercial enzymatic cocktail, highlighting the potential of the S. longicolleum enzyme mixture as an alternative pretreatment method. To further characterize the enzymes, which efficiently digested polymers such as cellulose, hemicellulose, pectin, starch, and lignin, we performed in-depth mass spectrometry-based secretome analysis using tryptic peptides from in-gel and in-solution digestions. Depending on the growth conditions, we were able to detect from 442 to 1092 proteins, which were annotated to identify from 134 to 224 putative carbohydrate-active enzymes (CAZymes) in five different families: glycoside hydrolases, auxiliary activities, carbohydrate esterases, polysaccharide lyases, glycosyl transferases, and proteins containing a carbohydrate-binding module, as well as combinations thereof. The FW57 enzyme mixture could be used to replace commercial enzyme cocktails for the digestion of agro-residual substrates.