Highlights d Patients with severe COVID-19 accumulate HLA-DR Low monocytes and immature neutrophils in blood/lungs d Calprotectin level positively correlates with neutrophil count and disease severity d Loss of non-classical monocytes could identify high risk of severe COVID-19

Aldehyde dehydrogenase 1A1 (ALDH) activity is one hallmark of human bone marrow (BM), umbilical cord blood (UCB), and peripheral blood (PB) primitive progenitors presenting high reconstitution capacities in vivo. In this study, we have identified ALDH(+) cells within human skeletal muscles, and have analyzed their phenotypical and functional characteristics. Immunohistofluorescence analysis of human muscle tissue sections revealed rare endomysial cells. Flow cytometry analysis using the fluorescent substrate of ALDH, Aldefluor, identified brightly stained (ALDH(br)) cells with low side scatter (SSC(lo)), in enzymatically dissociated muscle biopsies, thereafter abbreviated as SMALD(+) (for skeletal muscle ALDH(+)) cells. Phenotypical analysis discriminated two sub-populations according to CD34 expression: SMALD(+)/CD34(-) and SMALD(+)/CD34(+) cells. These sub-populations did not initially express endothelial (CD31), hematopoietic (CD45), and myogenic (CD56) markers. Upon sorting, however, whereas SMALD(+)/CD34(+) cells developed in vitro as a heterogeneous population of CD56(-) cells able to differentiate in adipoblasts, the SMALD(+)/CD34(-) fraction developed in vitro as a highly enriched population of CD56(+) myoblasts able to form myotubes. Moreover, only the SMALD(+)/CD34(-) population maintained a strong myogenic potential in vivo upon intramuscular transplantation. Our results suggest that ALDH activity is a novel marker for a population of new human skeletal muscle progenitors presenting a potential for cell biology and cell therapy.

IntroductionThe Notch signaling pathway guides cell-fate decisions in multiple developmental processes. 1,2 Intercellular communications that control the developmental fate of multipotent cells are mediated by the Notch family of transmembrane receptors in several invertebrate and vertebrate developmental systems. The Notch proteins are single-pass receptors that are activated by the Delta (or Delta-like) and Jagged/Serrate families of membrane-bound ligands. 3 To date, 4 human Notch genes have been identified and all are expressed on hematopoietic cells. 2 Also, 5 human Notch ligands, Delta-like1/3/4, and Jagged1/2 were identified, and all were shown to bind to Notch-1, Notch-2, and Notch-4. 4,5 Notch receptors are matured in the secretory pathway and presented at the cell surface as heterodimeric molecules. Interaction with ligands leads to 2 additional proteolytic cleavages that liberate the Notch intracellular domain (NICD) from the plasma membrane. NICD enters the nucleus, where it interacts with the DNA-binding protein, CSL (C-promoter binding factor [CBF]-1, Suppressor of Hairless, LAG-1; also known as recombination signal-binding protein Jk [RBP-Jk]). 6 In the absence of NICD, CSL represses transcription through interactions with a corepressor complex, containing a histone deacetylase. Upon entering the nucleus, NICD displaces the corepressor complex from CSL and replaces it with a transcriptional activation complex that includes NICD, Mastermind (MAML-1), the histone acetyltransferase p300, and, possibly, PCAF p300/CBP (cyclic AMP response element-binding protein [CREB]-binding protein)-associated factor. Notch signaling, thus, converts CSL from a repressor to an activator, leading to the transcription of target genes. The target genes include members of the Hes and HRT/ HERP/Hey families of transcriptional repressors; therefore, Notch signaling is often viewed as a transcription cascade.The classical view holds that Notch signaling controls the balance between the progenitor pool and its differentiating progeny and thus is involved in the maintenance of stem cell fate. 1,2 In fact, a number of studies have provided evidence that ligand-induced Notch signaling favored hematopoietic stem cell (HSC) selfrenewal, increased the numbers of progenitors, and promoted HSC survival. Moreover, Notch signaling may be instructive for differentiation toward a particular fate. It plays a crucial role in the hematopoietic system, especially in the regulation of the T-cell lymphoid lineage commitment 7-9 and in late stages of B-cell development. 10 Because of the key role of Notch signaling in supporting early T-cell differentiation, it was generally established that Notch concomitantly negatively regulates myeloid lineage development. The megakaryocytic and erythroid lineages are extremely linked, because they share a common bipotent progenitor called the MEP (MK/erythroid progenitor). The role of Notch in megakaryocytic and erythroid development remains a matter of debate. While some data report that Notch signaling...

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Classical BCR-ABL-negative myeloproliferative neoplasms (MPN) are clonal disorders of hematopoietic stem cells (HSC) caused mainly by recurrent mutations in genes encoding JAK2 (JAK2), calreticulin (CALR), or the thrombopoietin receptor (MPL). Interferon alpha (IFNα) has demonstrated some efficacy in inducing molecular remission in MPN. In order to determine factors that influence molecular response rate, we evaluated the long-term molecular efficacy of IFNα in MPN patients by monitoring the fate of cells carrying driver mutations in a prospective observational and longitudinal study of 48 patients over more than 5 years. We measured several times per year the clonal architecture of early and late hematopoietic progenitors (84,845 measurements) and the global variant allele frequency in mature cells (409 measurements). Using mathematical modeling and hierarchical Bayesian inference, we further inferred the dynamics of IFNα-targeted mutated HSC. Our data support the hypothesis that IFNα targets JAK2V617F HSC by inducing their exit from quiescence and differentiation into progenitors. Our observations indicate that treatment efficacy is higher in homozygous than heterozygous JAK2V617F HSC and increases with high IFNα dosage in heterozygous JAK2V617F HSC. Besides, we found that the molecular responses of CALRm HSC to IFNα were heterogeneous, varying between type 1 and type 2 CALRm, and high dosage of IFNα correlates with worse outcomes. Together, our work indicates that the long-term molecular efficacy of IFNα implies an HSC exhaustion mechanism and depends on both the driver mutation type and IFNα dosage.

Initial cell transplantation trials dedicated to the repair of striated muscles in muscular dystrophies produced mitigated results and underlined some limitations of cellular candidates under study. The research and identification of new stem cell candidates, the invention of new molecular strategies for correction of gene expression, the development of complementary approaches to improve transplantation success, have been justified by the unmet medical needs. These efforts led to new preclinical and clinical trials based on these concepts.

We investigated whether Notch signaling pathways have a role in human developmental hematopoiesis. In situ histochemistry analysis revealed that Notch1, 2, and 4 and Notch ligand (Delta1-4, and Jagged1) proteins were not expressed in the yolk sac blood islands, the para-aortic splanchnopleure, the hematopoietic aortic clusters, and at the early stages of embryonic liver hematopoiesis. Notch1-2, and Delta4 were eventually detected in the embryonic liver, from 34 until 38 days postconception. Fluorescenceactivated cell sorter analysis showed that first-trimester embryonic liver CD34 + CD38 low cells expressed both Notch1 and Notch2. When these cells were cultured on S17 stroma stably expressing Delta4, a 2.6-fold increase in BFU-E number was observed at day 7, as compared with cultures with control stroma, and this effect was maintained for 2 weeks. Importantly, exposure of these cells to Delta4 under these conditions maintained the original frequency and quality of long-term culture-initiating cells (LTC-ICs), while control cultures quickly resulted in the extinction of this LTC-IC potential. Furthermore, short-term exposure of embryonic liver adherent cells to erythropoietin resulted in a dose-dependent increase in Delta4 expression, almost doubling the expression observed with untreated stroma. This suggests that Delta4 has a role in the regulation of hematopoiesis after a hypoxic stress in the fetus. Stem Cells 2005;23:550-560

Background Aldehyde dehydrogenases (ALDHs) are key players in cell survival, protection, and differentiation via the metabolism and detoxification of aldehydes. ALDH activity is also a marker of stem cells. The skeletal muscle contains populations of ALDH‐positive cells amenable to use in cell therapy, whose distribution, persistence in aging, and modifications in myopathic context have not been investigated yet. Methods The Aldefluor® (ALDEF) reagent was used to assess the ALDH activity of muscle cell populations, whose phenotypic characterizations were deepened by flow cytometry. The nature of ALDH isoenzymes expressed by the muscle cell populations was identified in complementary ways by flow cytometry, immunohistology, and real‐time PCR ex vivo and in vitro . These populations were compared in healthy, aging, or Duchenne muscular dystrophy (DMD) patients, healthy non‐human primates, and Golden Retriever dogs (healthy vs. muscular dystrophic model, Golden retriever muscular dystrophy [GRMD]). Results ALDEF + cells persisted through muscle aging in humans and were equally represented in several anatomical localizations in healthy non‐human primates. ALDEF + cells were increased in dystrophic individuals in humans (nine patients with DMD vs. five controls: 14.9 ± 1.63% vs. 3.6 ± 0.39%, P = 0.0002) and dogs (three GRMD dogs vs. three controls: 10.9 ± 2.54% vs. 3.7 ± 0.45%, P = 0.049). In DMD patients, such increase was due to the adipogenic ALDEF + /CD34 + populations (11.74 ± 1.5 vs. 2.8 ± 0.4, P = 0.0003), while in GRMD dogs, it was due to the myogenic ALDEF + /CD34 − cells (3.6 ± 0.6% vs. 1.03 ± 0.23%, P = 0.0165). Phenotypic characterization associated the ALDEF + /CD34 − cells with CD9, CD36, CD49a, CD49c, CD49f, CD106, CD146, and CD184, some being associated with myogenic capacities. Cytological and histological analyses distinguished several ALDH isoenzymes (ALDH1A1, 1A2, 1A3, 1B1, 1L1, 2, 3A1, 3A2, 3B1, 3B2, 4A1, 7A1, 8A1, and 9A1) expressed by different cell populations in the skeletal muscle tissue belonging to multinucleated fibres, or myogenic, endothelial, interstitial, and neural lineages, designing them as potential new markers of cell type or of metabolic activity. Important modifications were noted in isoenzyme expression between healthy and DMD muscle tissues. The level of gene expression of some isoenzymes (ALDH1A1, 1A3, 1B1, 2, 3A2, 7A1, 8A1, and 9A1) suggested their specific involvement in muscle stability or regeneration in situ or in vitro . Conclusions ...