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1.
Nat Cell Biol ; 22(12): 1399-1410, 2020 12.
Article in English | MEDLINE | ID: mdl-33230302

ABSTRACT

Severe infections are a major stress on haematopoiesis, where the consequences for haematopoietic stem cells (HSCs) have only recently started to emerge. HSC function critically depends on the integrity of complex bone marrow (BM) niches; however, what role the BM microenvironment plays in mediating the effects of infection on HSCs remains an open question. Here, using a murine model of malaria and combining single-cell RNA sequencing, mathematical modelling, transplantation assays and intravital microscopy, we show that haematopoiesis is reprogrammed upon infection, whereby the HSC compartment turns over substantially faster than at steady-state and HSC function is drastically affected. Interferon is found to affect both haematopoietic and mesenchymal BM cells and we specifically identify a dramatic loss of osteoblasts and alterations in endothelial cell function. Osteo-active parathyroid hormone treatment abolishes infection-triggered HSC proliferation and-coupled with reactive oxygen species quenching-enables partial rescuing of HSC function.


Subject(s)
Hematopoiesis/physiology , Hematopoietic Stem Cells/physiology , Malaria/physiopathology , Stem Cell Niche/physiology , Animals , Bone Marrow Cells/cytology , Bone Marrow Cells/metabolism , Bone Marrow Cells/physiology , Endothelial Cells/cytology , Endothelial Cells/metabolism , Endothelial Cells/physiology , Gene Expression Profiling/methods , Hematopoiesis/drug effects , Hematopoiesis/genetics , Hematopoietic Stem Cells/cytology , Hematopoietic Stem Cells/metabolism , Humans , Malaria/parasitology , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Mesenchymal Stem Cells/physiology , Mice, Inbred C57BL , Mice, Knockout , Osteoblasts/cytology , Osteoblasts/metabolism , Osteoblasts/physiology , Parathyroid Hormone/pharmacology , Plasmodium berghei/physiology , Reactive Oxygen Species/metabolism , Stem Cell Niche/genetics
2.
Polymers (Basel) ; 12(8)2020 Aug 06.
Article in English | MEDLINE | ID: mdl-32781503

ABSTRACT

Cell migration has a central role in osteochondral defect repair initiation and biomaterial-mediated regeneration. New advancements to reestablish tissue function include biomaterials and factors promoting cell recruitment, differentiation and tissue integration, but little is known about responses to mechanical stimuli. In the present pilot study, we tested the influence of extrinsic forces in combination with biomaterials releasing chemoattractant signals on cell migration. We used an ex vivo mechanically stimulated osteochondral defect explant filled with fibrin/hyaluronan hydrogel, in presence or absence of platelet-derived growth factor-BB or stromal cell-derived factor 1, to assess endogenous cell recruitment into the wound site. Periodic mechanical stress at early time point negatively influenced cell infiltration compared to unloaded samples, and the implementation of chemokines to increase cell migration was not efficient to overcome this negative effect. The gene expression at 15 days of culture indicated a marked downregulation of matrix metalloproteinase (MMP)13 and MMP3, a decrease of ß1 integrin and increased mRNA levels of actin in osteochondral samples exposed to complex load. This work using an ex vivo osteochondral mechanically stimulated advanced platform demonstrated that recurrent mechanical stress at early time points impeded cell migration into the hydrogel, providing a unique opportunity to improve our understanding on management of joint injury.

3.
J Control Release ; 309: 220-230, 2019 09 10.
Article in English | MEDLINE | ID: mdl-31369767

ABSTRACT

Articular cartilage is frequently injured by trauma or osteoarthritis, with limited and inadequate treatment options. We investigated a new strategy based on hydrogel-mediated delivery of a locked nucleic acid microRNA inhibitor targeting miR-221 (antimiR-221) to guide in situ cartilage repair by endogenous cells. First, we showed that transfection of antimiR-221 into human bone marrow-derived mesenchymal stromal cells (hMSCs) blocked miR-221 expression and enhanced chondrogenesis in vitro. Next, we loaded a fibrin/hyaluronan (FB/HA) hydrogel with antimiR-221 in combination or not with lipofectamine carrier. FB/HA strongly retained functional antimiR-221 over 14 days of in vitro culture, and provided a supportive environment for cell transfection, as validated by flow cytometry and qRT-PCR analysis. Seeding of hMSCs on the surface of antimiR-221 loaded FB/HA led to invasion of the hydrogel and miR-221 knockdown in situ within 7 days. Overall, the use of lipofectamine enhanced the potency of the system, with increased antimiR-221 retention and miR-221 silencing in infiltrating cells. Finally, FB/HA hydrogels were used to fill defects in osteochondral biopsies that were implanted subcutaneously in mice. FB/HA loaded with antimiR-221/lipofectamine significantly enhanced cartilage repair by endogenous cells, demonstrating the feasibility of our approach and the need to achieve highly effective in situ transfection. Our study provides new evidence on the treatment of focal cartilage injuries using controlled biomaterial-mediated delivery of antimicroRNA for in situ guided regeneration.


Subject(s)
Chondrogenesis , Drug Delivery Systems/methods , Hydrogels/chemistry , MicroRNAs/administration & dosage , Aged , Animals , Cartilage, Articular/injuries , Cartilage, Articular/physiology , Cells, Cultured , Female , Fibrin/chemistry , Humans , Hyaluronic Acid/chemistry , Mesenchymal Stem Cells/metabolism , Mice , MicroRNAs/genetics , MicroRNAs/therapeutic use , Middle Aged , Regeneration
4.
Open Biol ; 6(6)2016 06.
Article in English | MEDLINE | ID: mdl-27335321

ABSTRACT

Haematopoiesis is the complex developmental process that maintains the turnover of all blood cell lineages. It critically depends on the correct functioning of rare, quiescent haematopoietic stem cells (HSCs) and more numerous, HSC-derived, highly proliferative and differentiating haematopoietic progenitor cells (HPCs). Infection is known to affect HSCs, with severe and chronic inflammatory stimuli leading to stem cell pool depletion, while acute, non-lethal infections exert transient and even potentiating effects. Both whether this paradigm applies to all infections and whether the HSC response is the dominant driver of the changes observed during stressed haematopoiesis remain open questions. We use a mouse model of malaria, based on natural, sporozoite-driven Plasmodium berghei infection, as an experimental platform to gain a global view of haematopoietic perturbations during infection progression. We observe coordinated responses by the most primitive HSCs and multiple HPCs, some starting before blood parasitaemia is detected. We show that, despite highly variable inter-host responses, primitive HSCs become highly proliferative, but mathematical modelling suggests that this alone is not sufficient to significantly impact the whole haematopoietic cascade. We observe that the dramatic expansion of Sca-1(+) progenitors results from combined proliferation of direct HSC progeny and phenotypic changes in downstream populations. We observe that the simultaneous perturbation of HSC/HPC population dynamics is coupled with early signs of anaemia onset. Our data uncover a complex relationship between Plasmodium and its host's haematopoiesis and raise the question whether the variable responses observed may affect the outcome of the infection itself and its long-term consequences on the host.


Subject(s)
Hematopoiesis , Malaria/parasitology , Sporozoites/pathogenicity , Animals , Cell Proliferation , Disease Models, Animal , Hematopoietic Stem Cells/cytology , Hematopoietic Stem Cells/parasitology , Malaria/blood , Mice
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