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Delayed natural killer (NK) cell reconstitution after allogeneic stem cell transplantation (alloSCT) is associated with a higher risk of developing invasive aspergillosis. The interaction of NK cells with the human pathogen Aspergillus (A.) fumigatus is mediated by the fungal recognition receptor CD56, which is relocated to the fungal interface after contact. Blocking of CD56 signaling inhibits the fungal mediated chemokine secretion of MIP-1α, MIP-1β, and RANTES and reduces cell activation, indicating a functional role of CD56 in fungal recognition. We collected peripheral blood from recipients of an allograft at defined time points after alloSCT (day 60, 90, 120, 180). NK cells were isolated, directly challenged with live A. fumigatus germ tubes, and cell function was analyzed and compared to healthy age and gender-matched individuals. After alloSCT, NK cells displayed a higher percentage of CD56\(^{bright}\)CD16\(^{dim}\) cells throughout the time of blood collection. However, CD56 binding and relocalization to the fungal contact side were decreased. We were able to correlate this deficiency to the administration of corticosteroid therapy that further negatively influenced the secretion of MIP-1α, MIP-1β, and RANTES. As a consequence, the treatment of healthy NK cells ex vivo with corticosteroids abrogated chemokine secretion measured by multiplex immunoassay. Furthermore, we analyzed NK cells regarding their actin cytoskeleton by Structured Illumination Microscopy (SIM) and flow cytometry and demonstrate an actin dysfunction of NK cells shown by reduced F-actin content after fungal co-cultivation early after alloSCT. This dysfunction remains until 180 days post-alloSCT, concluding that further actin-dependent cellular processes may be negatively influenced after alloSCT. To investigate the molecular pathomechansism, we compared CD56 receptor mobility on the plasma membrane of healthy and alloSCT primary NK cells by single-molecule tracking. The results were very robust and reproducible between tested conditions which point to a different molecular mechanism and emphasize the importance of proper CD56 mobility.
The proteome profiles of the olfactory bulb of juvenile, adult and aged rats - an ontogenetic study
(2015)
Background:
In this study, we searched for proteins that change their expression in the olfactory bulb (oB) of rats during ontogenesis. Up to now, protein expression differences in the developing animal are not fully understood. Our investigation focused on the question whether specific proteins exist which are only expressed during different development stages. This might lead to a better characterization of the microenvironment and to a better determination of factors and candidates that influence the differentiation of neuronal progenitor cells.
Results:
After analyzing the samples by two-dimensional polyacrylamide gel electrophoresis (2DE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), it could be shown that the number of expressed proteins differs depending on the developmental stages. Especially members of the functional classes, like proteins of biosynthesis, regulatory proteins and structural proteins, show the highest differential expression in the stages of development analyzed.
Conclusion:
In this study, quantitative changes in the expression of proteins in the oB at different developmental stages (postnatal days (P) 7, 90 and 637) could be observed. Furthermore, the expression of many proteins was found at specific developmental stages. It was possible to identify these proteins which are involved in processes like support of cell migration and differentiation.
The Proteome Profiles of the Cerebellum of Juvenile, Adult and Aged Rats-An Ontogenetic Study
(2015)
In this study, we searched for proteins that change their expression in the cerebellum (Ce) of rats during ontogenesis. This study focuses on the question of whether specific proteins exist which are differentially expressed with regard to postnatal stages of development. A better characterization of the microenvironment and its development may result from these study findings. A differential two-dimensional polyacrylamide gel electrophoresis (2DE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis of the samples revealed that the number of proteins of the functional classes differed depending on the developmental stages. Especially members of the functional classes of biosynthesis, regulatory proteins, chaperones and structural proteins show the highest differential expression within the analyzed stages of development. Therefore, members of these functional protein groups seem to be involved in the development and differentiation of the Ce within the analyzed development stages. In this study, changes in the expression of proteins in the Ce at different postnatal developmental stages (postnatal days (P) 7, 90, and 637) could be observed. At the same time, an identification of proteins which are involved in cell migration and differentiation was possible. Especially proteins involved in processes of the biosynthesis and regulation, the dynamic organization of the cytoskeleton as well as chaperones showed a high amount of differentially expressed proteins between the analyzed dates.
Aspergillus (A.) fumigatus is an opportunistic fungal mold inducing invasive aspergillosis (IA) in immunocompromised patients. Although antifungal activity of human natural killer (NK) cells was shown in previous studies, the underlying cellular mechanisms and pathogen recognition receptors (PRRs) are still unknown. Using flow cytometry we were able to show that the fluorescence positivity of the surface receptor CD56 significantly decreased upon fungal contact. To visualize the interaction site of NK cells and A. fumigatus we used SEM, CLSM and dSTORM techniques, which clearly demonstrated that NK cells directly interact with A. fumigatus via CD56 and that CD56 is re-organized and accumulated at this interaction site time-dependently. The inhibition of the cytoskeleton showed that the receptor re-organization was an active process dependent on actin re-arrangements. Furthermore, we could show that CD56 plays a role in the fungus mediated NK cell activation, since blocking of CD56 surface receptor reduced fungal mediated NK cell activation and reduced cytokine secretion. These results confirmed the direct interaction of NK cells and A. fumigatus, leading to the conclusion that CD56 is a pathogen recognition receptor. These findings give new insights into the functional role of CD56 in the pathogen recognition during the innate immune response.