Enrichment of Mesenchymal Progenitor Cells from Mouse Compact Bone.
Notice bibliographique
Résumé
Abstract Adult mammalian bone marrow (BM) contains at least two distinct stem cell populations; the stem cells of the hemopoietic lineage and a second population termed mesenchymal stem cells (MSC) whose function is to maintain the non-hemopoietic BM elements as well as skeletal homeostasis. MSC have been implicated as potential targets in a range of cellular therapies for treatment of defects of both the hemopoietic and skeletal systems, and as vehicles for gene therapy. In order to evaluate the potential of these cells in various therapies, a pre-clinical animal model in which both the biology and potential therapeutic applications of these cells can be assessed is of fundamental importance. The goal of the current study was to develop a robust and reproducible method for the isolation of MSC from murine hemopoietic tissues. Tibiae and femurs harvested from C57BL6/J mice were gently crushed with a pestle to release the marrow. The bone fragments were subsequently cut into small pieces with a scalpel and digested in a solution containing 3mg/ml Type I collagenase to yield a population of compact bone (CB) derived cells. Mesenchymal progenitor cells (MPC) were detected using an in vitro assay for fibroblast-colony forming cells (CFU-F). CB cells were plated at 1000 or 5000 cells per cm2 in complete MesenCult™ medium for 12 days, after which CFU-F-derived colonies were enumerated. We show that CFU-F are present at a significantly higher frequency in mouse CB than in the BM (433±225 vs 11.7±3.5 colonies/106 cells respectively, n=3). Based on these data we developed a simple and robust immunomagnetic selection method to highly enrich MPC from mouse CB by depleting essentially all nucleated hemopoietic cells (CD45+) and red blood cells (Ter119+) using magnetic particles and antibodies to CD45 and Ter119, respectively. Target CD45−Ter119− cells initially comprised 1.1±0.5% (n=9) of the total CB fraction as assayed by FACS. Following depletion, CD45−Ter119− cells comprised 74.5±16% (n=6) of the cells and were enriched 205 fold for CFU-F compared to the starting population, with a CFU-F frequency of 1 per 11 cells plated, and a total CFU-F recovery of 57.9 ± 18.5%. Analysis of CD45−Ter119−Sca-1+ cells, a phenotype previously shown to enrich for MPC, revealed that these cells were enriched 50 fold following depletion, from 0.53±0.5 to 26.5±8.23% (n=3). The enriched MPCs cultured at low O2 tension were devoid of hemopoietic contaminants at passage 1 and 2 as shown by lack of CD45, Ter119 and CD11b expression. The cultured CB-derived MPCs were capable of extensive in vitro proliferation and maintained the ability to differentiate into cells of the osteogenic, adipogenic and chondrogenic lineages. Furthermore, irradiated cultured mesenchymal cells supported long-term culture-initiating cells (LTC-IC) in 4-week cultures of Sca-1+ BM cells under limiting dilution conditions, at frequencies similar to those detected using irradiated primary BM feeders (i.e. 1 per 1600). These data provide a rapid, reproducible method by which multipotent mesenchymal cells devoid of contaminating hemopoietic cells can be readily obtained from limited numbers of mice to study the biology of MSC as well as the use of these cells as therapeutic agents in a preclinical animal model.
Récupéré en direct depuis OpenAlex et désinversé. Les résumés ne sont pas conservés dans cette base de données : les index inversés représentent 8,6 Go des 9,3 Go de texte de la base, et le serveur dispose de 13 Go libres.
Comment cette classification a été obtenuedéplier
Prédiction machine sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Le volet Gemma est une étiquette directe du modèle pour chaque travail de la base, lue sur la notice réduite au titre. Le volet Codex est un classifieur appris des 10 348 étiquettes directes de Codex et calibré sur les taux pondérés de l'échantillon; les champs sans appui suffisant ne portent aucun appel Codex. Le mode candidate est l'union des deux volets; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont pas des étiquettes humaines.
Scores du classifieur distillé par catégorie (deux têtes)
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,000 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,000 |
| Bibliométrie | 0,001 | 0,000 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,000 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,002 | 0,001 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule source (Gemma direct ou Codex distillé), pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».