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Moura, P. L.

Publications and source records attributed to Moura, P. L..

2 recordsLinked to original sources

Erythroid differentiation intensifies RNA mis-splicing in SF3B1-mutant myelodysplastic syndromes with ring sideroblasts

Myelodysplastic syndromes with ring sideroblasts (MDS-RS) commonly originate from mutations in the splicing factor SF3B1 (SF3B1mt). SF3B1mt cause RNA mis-splicing, mechanistically established as the major driver of RS development. However, little is known about RS fate and biology after their initial formation in the human bone marrow. We here achieve isolation of viable RS from patient samples, enabling the first complete investigation of SF3B1mt development from stem cell to RS. We show that RS skew MACS-isolated CD34+ data towards erythroid features not recapitulated in single-cell RNAseq. We demonstrate that RS divide, differentiate, enucleate and actively respond to mis-splicing/oxidative stress, decreasing wildtype stem cell fitness via GDF15 overproduction. We identify circulating RS as a uniform clinical feature associated with disease burden. Finally, we establish that SF3B1mt mis-splicing intensifies during erythroid differentiation and demonstrate through combined transcriptomics/proteomics an uncoupling of RNA/protein biology in RS encompassing severe and dysfunctional mis-splicing of proapoptotic genes. Statement of significanceWe here combine a novel method for RS isolation with state-of-the-art multiomics to perform the first complete investigation of SF3B1mt MDS-RS hematopoiesis from stem cell to RS. We identify the survival mechanisms underlying SF3B1mt erythropoiesis and establish an active role for erythroid differentiation and RS themselves in SF3B1mt MDS-RS pathogenesis.

cancer biology↗

Characterizing the polarization continuum of macrophage subtypes M1, M2a and M2c

Macrophages are vital components of the inflammatory response and exhibit phenotypical plasticity through active conversion between pro- and anti-inflammatory cell subtypes, a feature which can be reproduced in ex vivo culture. We employed a multifaceted approach utilizing proteomics, flow cytometry, activity assays and livecell microscopy imaging to characterize four cultured macrophage subtypes: unstimulated MO, classically activated M1, alternatively activated M2a, and deactivated M2c macrophages. Whole cell proteomics identified a total of 5435 proteins, with >50% of these proteins exhibiting significant alterations in abundance between the different subtypes. This confirms that four distinct macrophage subtypes are induced from the same originating donor material through stimulation with specific cytokines. Additional surfaceome analysis revealed that M2c macrophages significantly upregulate pro-inflammatory markers compared to the MO baseline and thus appear to be activated or primed to activate, similar to M1. Surface protein expression provided further subtype characterization, in particular distinguishing between the M2a and M2c macrophages. We next explored the re-polarization capabilities of macrophages using dexamethasone, an anti-inflammatory glucocorticoid known to induce macrophage polarization towards the M2c de-activated phenotype. We show that activated M1 macrophages treated with dexamethasone for 48-hours upregulate the levels of CD163 and CD206, markers synonymous with a phenotypical shift from M1 to M2c yet retain key surface markers and display the functional phenotype of M1 macrophages. The observed repolarization of M1 pro-inflammatory macrophages provides a potential mechanism through which dexamethasone treatment improves COVID-19 prognosis and constitutes evidence of partial repolarization along the macrophage continuum. These proteomic and functional ex vivo macrophage datasets provide a valuable resource for studying macrophage polarity and the impact of dexamethasone on macrophage phenotype and function.

immunology↗