Search bioRxiv⌕ Search

bioRxiv · 10.1101/2020.11.06.348177

Pro chondrogenic effect of mesenchymal stromal cell-based treatment of chondral defects under physioxia in a novel ex vivo organ mode

Abstract

ObjectiveCartilage defect treatment strategies are dependent on the lesion size and severity. Osteochondral explants models are a platform to test cartilage repair strategies ex vivo. Current models lack in mimicking the variety of clinically relevant defect scenarios. In this controlled laboratory study, an automated device (artificial tissue cutter, ARTcut(R)) was implemented to reproducible create cartilage defects with controlled depth. In a pilot study, the effect of cartilage defect depth and oxygen tension on cartilage repair was investigated. DesignOsteochondral explants were isolated from porcine condyles. 4 mm chondral and full thickness defects were treated with either porcine chondrocytes (CHON) or co-culture of 20 % CHON and 80 % MSC (MIX) embedded in collagen hydrogel. Explants were cultured with tissue specific media (without TGF-{beta}) under normoxia (20 % O2) and physiological hypoxia (2 % O2). After 28 days, immune-histological stainings (Collagen II and X, Aggrecan) were scored (modified Bern-score, 3 independent scorer) to quantitatively compare treatments outcome. ResultsARTcut(R) represents a software-controlled device for creation of uniform cartilage defects. Comparing the scoring results of the MIX and the CHON treatment, a positive relation between oxygen tension and defect depth was observed. Low oxygen tension stimulated cartilaginous matrix deposition in MIX group in chondral defects and CHON treatment in full thickness defects. ConclusionARTcut(R) has proved a powerful tool to create cartilage defects and thus opens a wide range of novel applications of the osteochondral model, including the relation between oxygen tension and defect depth on cartilage repair.

Explore related subjects

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Schwab, A., Buss, A., Pullig, O., Ehlicke, F.. 2020-11-08. Pro chondrogenic effect of mesenchymal stromal cell-based treatment of chondral defects under physioxia in a novel ex vivo organ mode. https://doi.org/10.1101/2020.11.06.348177

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

Functional characterization of Rho GTPase activating proteins SYDE1 and SYDE2

The human genome encodes more than 60 proteins containing Rho GTPase activating protein (RhoGAP) domains, many of which remain understudied with respect to their target specificity and biological roles. SYDE1 and SYDE2 are two such orphan RhoGAPs, for which there are few studies characterizing their biochemical and cellular functions and conflicting reports identifying their cognate GTPases. We previously identified SYDE1 and SYDE2 in a screen for substrates of the c-Jun N-terminal kinases. Here, we show that SYDE1 and SYDE2 are preferentially phosphorylated by JNK1 relative to other mitogen-activated protein kinases (MAPKs) at sites proximal to a kinase docking region. Purified SYDE1 and SYDE2 are shown to have significant catalytic GAP activity toward RhoA, Rac1, and Cdc42. However, neither up- nor down-regulation of SYDE1/2 expression leads to detectable changes in bulk GTP loading of any of these GTPases. Nevertheless, we demonstrate that SYDE1 and SYDE2, in a partially GAP-dependent manner, increase cell spreading and number of focal adhesions, and promote more directionally persistent migration in HEK293 cells. Together, these findings establish SYDE1 and SYDE2 as robust JNK substrates with catalytic activity toward a set of Rho GTPases and reveal basic functions of SYDE1 and SYDE2 in regulating cell morphology, adhesion, and migration.

cell biology↗

The filopodial scaffold polyphosphate dictates cell adhesion-versus-invasion decisions

Inorganic polyphosphate (polyP) is an ancient polymer conserved across all life, serving cell type and location specific functions in every major compartment. Yet its role at the plasma membrane, where it accumulates to peak levels in many primary cells, is largely unknown. Here we identify polyP as a stabilizing component of filopodia, actin based membrane protrusions that govern cell adhesion, contact inhibition, and chemotaxis. Elevating cellular polyP increases filopodial stability and enhances cell adhesion, whereas reducing polyP accelerates filopodial disassembly and promotes cell migration. Mechanistically, we find that polyP acts as a structural filopodial scaffold, recruiting and organizing IRSp53, a membrane curvature inducing protein. We show that metastatic fibroblasts and breast cancer organoids carry markedly reduced and intracellularly reorganized polyP levels relative to their non transformed counterparts. Restoring endogenous polyP via lipid nanoparticle delivery suppresses their invasive phenotypes and reverses prometastatic gene expression signatures, implicating polyP as a primordial tumor suppressor.

cell biology↗

Mitochondrial transfer mediates metabolic communication between beta cells and islet macrophages

Pancreatic islet macrophages support islet homeostasis and adapt their metabolic program in response to environmental cues, including beta cell released factors. Intercellular mitochondrial transfer is a biological process that modulates cellular responses. To test whether beta cells, which are strongly secretory, transfer mitochondria to islet macrophages, we generated mice with beta cell-specific expression of mitochondrial GFP (PhAMfloxIns1Cre). We demonstrate that beta cells transfer mitochondria to islet macrophages in vivo and in vitro. Diabetogenic stressors did not alter the frequency of mitochondrial transfer and macrophages containing beta cell-derived GFP exhibit increased protein synthesis rates. RNA-seq identified upregulation of activity-regulated cytoskeleton associated protein (Arc) in macrophages receiving beta cell-derived mitochondria, while disruption of actin cytoskeleton dynamics prevented mitochondrial transfer. Together, these findings identify mitochondrial transfer as a previously unrecognized mechanism of beta cell-macrophage communication that may contribute to islet homeostasis and immune regulation.

cell biology↗