Embryonnaires humaines études de cellules souches dans le laboratoire Teitell sont soutenus par un California Institute for Regenerative Medicine (CIRM) Subvention de démarrage RS1-00313. Nous remercions les membres du grand centre de médecine régénérative et de recherche sur les cellules souches à l&#39;UCLA, en particulier le Dr Clark Amander, le Dr Jerome Zack, et des membres de l&#39;UCLA large souches Facilité Institut noyau cellulaire pour leur soutien de nos études.

l&#39;entretien de la culture quotidienne de CSEh <ol><li> milieux de culture des CSEh doivent être changés toutes les 24 heures. De la bouteille ES stock de médias stockés à 4 ° C, sortir la quantité de solution nécessaire (~ 20ml par 6-même plaque), placer dans un tube de 50ml Falcon, et chaud à 37 ° C dans un bain d&#39;eau. Une fois que le média est réchauffé, ajouter le bFGF conservés à 4 ° C à une concentration finale de 10ng/ml. Mettez le stock bFGF revenir à 4 ° C immédiatement après usage! </li><li> Retirer des plaques de 6 puits, mais tous les ~ 500μl de médias. Assurez-vous de tourbillonner la boîte de culture de suspendre pour l&#39;enlèvement des débris. Assurez-vous que le bFGF dans les milieux de culture des CSEh est bien mélangé et ajouter 3 ml de milieux frais de retour à chaque puits d&#39;une plaque à 6 puits. Remettez la plaque dans l&#39;incubateur. </li></ol> Fractionnement des CSEh MEFs sur Matrigel Habituellement, un confluentes 6-même assiette de CSEh le FAE peut être divisé 1:02-1:03 sur Matrigel plaques de 6 puits, avec les puits deviennent confluentes à nouveau 4-5 jours après fractionnement. <ol><li> Lorsque CSEh fractionnement sur Matrigel, MEF conditionné médias (CM) est utilisé pour maintenir la pluripotence. MEF-médias est conditionnée à l&#39;avance. Le premier jour, 1,5 × 10 7 γ-irradiés FAE sont ensemencées dans un flacon T75. Le deuxième jour, FAE sont lavés une fois avec une température ambiante x PBS, pH 7,4, puis 15ml de médias ES complétée par 5ng/ml bFGF est ajouté dans le ballon. (REMARQUE: FAE peuvent être étalées dans des flacons plus petits ou plus gros, mais la proportion entre la densité cellulaire et du volume des supports ES doit être constante.) Le flacon est incubé à 37 ° C pendant 24 heures. Le troisième jour, le CM est collectée et remplacé par 15 ml de médias ES frais complété avec 5ng/ml bFGF. Le CM collectées sont stockées à 4 ° C. MEF-conditionnée des médias d&#39;un placage de FAE sont récoltées pendant sept jours consécutifs de générer 15 × 7 = 105ml. Après sept jours, toutes les fractions sont combinées CM, filtration stérile, aliquotés et conservés à -20 ° C. CM des médias préparé comme décrit peut être stockée et utilisée pour 1 mois. </li><li> Un jour, avant de se séparer de CSEh, mis aliquotes dans des tubes eppendorf Matrigel à 4 ° C à dégeler la nuit (une aliquote contient 76.2mg de Matrigel, et ceci est la quantité nécessaire pour un 6-même plaque). Le jour de fractionnement, de prendre un flacon de Matrigel décongelées pour un 6-même assiette, mettre le flacon et 6ml de froid DMEM/F12 des médias sur la glace. Transférer le matrigel dans le DMEM/F12 médias et bien mélanger. Ajouter 1 ml de la solution à chaque puits d&#39;une plaque à 6 puits. Swirl la plaque de distribuer Matrigel en surface et incuber la plaque recouverte de Matrigel à température ambiante pendant au moins 1 heure avant de se séparer du CSEh. </li><li> CSEh le MEF sont lavées une fois avec 1 x PBS, pH 7,4. Par la suite 1ml d&#39;eau tiède collagénase IV (1mg/ml) est ajouté à chaque puits de la plaque de 6 puits, puis la plaque est incubée à 37 ° C pendant 5-10 min. </li><li> Ajouter 1ml de médias ES sans bFGF à chaque puits. Utiliser une pipette 1 ml de faire sauter les cellules souches de la plaque. Recueillir tous les médias contenant des touffes de CSEh et morts FAE dans un tube 50ml Falcon. Laissez gros amas de cellules se contenter de 5-10 min pour que les touffes de CSEh forment un culot au fond du tube tandis que la FAE restent en suspension dans le surnageant. Jeter le surnageant et laver à la remise en suspension le culot de CSEh ES en utilisant les médias manquent de bFGF, suivie par une centrifugation à température ambiante à 200g pendant 5 min. </li><li> Pour plaque de cellules souches sur des plaques de Matrigel, d&#39;abord laver les assiettes avec Matrigel DMEM/F12 température ambiante (1ml par puits) et en ajoutant 2,5 ml de CM complété avec 10ng/ml bFGF par puits. Reprendre le culot dans un volume approprié de CM complété avec 10ng/ml bFGF, une pipette la suspension cellulaire de haut en bas plusieurs fois jusqu&#39;à l&#39;amas devient uniforme et de petite taille. Aliquoter la suspension à 0,5 ml par puits sur la plaque de Matrigel. Notez que la densité des colonies sur Matrigel est plus élevée que la densité des colonies par fractionnement d&#39;habitude sur le FAE. Placer la plaque dans un 37 ° C de culture de tissu incubateur. </li><li> Pour maintenir la CSEh sur Matrigel, les médias CM complétée par 10ng/ml bFGF est changé toutes les 24 heures pour un maximum de 4-5 jours. </li></ol>

<ol>
	<li>Thomson, J. a. m. e. s. A., Itskovitz-Eldor, J. o. s. e. p. h., Shapiro, S. a. n. d. e. r. S., Waknitz, M. i. c. h. e. l. l. e. A., Swiergiel, J. e. n. n. i. f. e. r. J., Marshall, V. i. v. i. e. n. n. e. S., Jones, J. e. f. f. r. e. y. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Embryonic+Stem+Cell+Lines+Derived+from+Human+Blastocysts++.">Embryonic Stem Cell Lines Derived from Human Blastocysts .</a> Science. 282, 1145-1147 (1998).</li><li>Xu, C. h. u. n. h. u. i., Inokuma, M. a. r. g. a. r. e. t. S., Denham, J. e. r. r. o. d., Golds, K. a. t. h. a. l. e. e. n., Kundu, P. r. a. t. i. m. a., Gold, J. o. s. e. p. h. D., Carpenter, M. e. l. i. s. s. a. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Feeder-free+growth+of+undifferentiated+human+embryonic+stem+cells.">Feeder-free growth of undifferentiated human embryonic stem cells.</a> Nature Biotechnology. 19, 971-974 (2001).</li></ol>

Cette vidéo montre comment les CSEh passage de plaques MEF pour les plaques d&#39;alimentation Matrigel sans cellule. Notez que la densité des colonies sur Matrigel est plus élevée que la densité des colonies par fractionnement d&#39;habitude sur le FAE. Immunofluorescence et cytométrie de flux pour microscopie ou marqueurs de pluripotence des CSEh, tels que Oct-4, SSEA-4, sont nécessaires pour confirmer l&#39;entretien des CSEh dans un état indifférencié dans les conditions de culture nourricier libre.

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		<th>Material Name</th>
		<th>Type</th>
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		<th>Catalogue Number</th>
		<th>Comment</th>
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<tr>
<td>Knockout Serum Replacer (KSR)</td>
<td>Reagent</td>
<td>Gibco</td>
<td>10828-028</td>
<td>&nbsp;</td>
<tr>
<td>DMEM/F12</td>
<td>Reagent</td>
<td>Gibco</td>
<td>11330-057</td>
<td>&nbsp;</td>
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<td> Non-essential Amino Acids</td>
<td>Reagent</td>
<td>Gibco</td>
<td>11140-050</td>
<td>&nbsp;</td>
<tr>
<td>GlutaMax </td>
<td>Reagent</td>
<td>Gibco</td>
<td>35050-061</td>
<td>&nbsp;</td>
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<td>DMEM</td>
<td>Reagent</td>
<td>Gibco</td>
<td>11995-065</td>
<td>&nbsp;</td>
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<td>FBS</td>
<td>Reagent</td>
<td>Clontech</td>
<td>631107</td>
<td>&nbsp;</td>
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<td>L-glutamine</td>
<td>Reagent</td>
<td>Gibco</td>
<td>25030-081</td>
<td>&nbsp;</td>
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<td>BME</td>
<td>Reagent</td>
<td>Fisher</td>
<td>BP176-100</td>
<td>&nbsp;</td>
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<td>bFGF</td>
<td>Reagent</td>
<td>R&D Systems</td>
<td>233-FB-025</td>
<td>&nbsp;</td>
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<td>Collagenase IV</td>
<td>Reagent</td>
<td>Gibco</td>
<td>17104-019</td>
<td>&nbsp;</td>
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<td>Dispase</td>
<td>Reagent</td>
<td>Stem Cell Technologies</td>
<td>17105-041</td>
<td>&nbsp;</td>
<tr>
<td>Penicillin / Streptomycin </td>
<td>Reagent</td>
<td>Gibco</td>
<td>15140-122</td>
<td>&nbsp;</td>
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<td>Gelatin</td>
<td>Reagent</td>
<td>Chemicon</td>
<td>ES-006-B</td>
<td>&nbsp;</td>
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<td>Matrigel</td>
<td>Reagent</td>
<td>BD Biosciences</td>
<td>354277</td>
<td>&nbsp;</td>
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<td>Oct-4 antibody</td>
<td>Reagent</td>
<td>Santa Cruz Biotechnology</td>
<td>SC-9081</td>
<td>&nbsp;</td>
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<td>anti-h/mSSEA-4 Phycoerythrin Conjugated Mouse IgG3</td>
<td>Reagent</td>
<td>R&D Systems</td>
<td>FAB1435P</td>
<td>&nbsp;</td>
<tr>
<td>FITCI-conjugated antirabbit IgG</td>
<td>Reagent</td>
<td>Jackson ImmunoResearch Laboratories. Inc.</td>
<td>715-095-152</td>
<td>&nbsp;</td>		</tbody>
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from mefs to matrigel 2: splitting hescs from mefs onto matrigel

Cette vidéo montre comment faire pousser des cellules souches embryonnaires humaines (CSEh) sur fibroblastes de souris embryonnaires (MEF) cellules nourricières, comment CSEh passage de plaques MEF pour chargeur de pile plaques sans Matrigel.

Watch this Scientific Journal Video about From MEFs to Matrigel 2: Splitting hESCs from MEFs onto Matrigel at JoVE.com

From MEFs to Matrigel 2: Splitting hESCs from MEFs onto Matrigel

Cette vidéo montre comment maintenir la croissance de cellules souches embryonnaires humaines (CSEh) dans le chargeur sans cellules conditions et la manière de CSEh passage en continu dans le chargeur sans cellules conditions. Confirmation de CSEh pluripotence grandi dans le chargeur sans cellules par microscopie d&#39;immunofluorescence conditions est également démontrée.

De FAE au Matrigel 2: Fractionnement des CSEh MEFs sur Matrigel

This video demonstrates how to grow human embryonic stem cells (hESCs) on mouse embryonic fibroblast (MEF) feeder cells, how to passage hESCs from MEF ...

from-mefs-to-matrigel-2-splitting-hescs-from-mefs-onto-matrigel

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Biology

11.4K vues.  University of California, Los Angeles. Cette vidéo montre comment maintenir la croissance de cellules souches embryonnaires humaines (CSEh) dans le chargeur sans cellules conditions et la manière de CSEh passage en continu dans le chargeur sans cellules conditions. Confirmation de CSEh pluripotence grandi dans le chargeur sans cellules par microscopie d'immunofluorescence conditions est également démontrée.

Vidéo: From MEFs to Matrigel 2: Splitting hESCs from MEFs onto Matrigel

From MEFs to Matrigel I: Passaging hESCs in the Presence of MEFs

This video demonstrates how to grow human embryonic stem cells (hESCs) on mouse embryonic fibroblast (MEF) feeder cells.


This video demonstrates how to grow human embryonic stem cells (hESCs) on mouse embryonic fibroblast (MEF) feeder cells. Part 1 of 3.

De MEFs à I Matrigel: CSEh repiquage en présence du MEF

This video demonstrates how to grow human embryonic stem cells (hESCs) on mouse embryonic fibroblast (MEF) feeder cells.
...

Recherche

Biologie

Generating iPS Cells from MEFS through Forced Expression of Sox-2, Oct-4, c-Myc, and Klf4

Pluripotency can be induced in differentiated murine by viral transduction of Oct4, Sox2, Klf4, and c-Myc (Takahashi and Yamanaka, 2006; Wernig, et al., 2007; Okita, et al., 2007; Maherali, et al., 2007). We have devised a reprogramming strategy in which these four transcription factors are expressed from doxycycline (dox)-inducible lentiviral vectors (Brambrink et al., 2008). Using these inducible constructs, we can derive induced pluripotent stem (iPS) cells from mouse embryonic fibroblasts (MEFs). In this video, we demonstrate the procedure for the generation of inducible lentiviruses that express the four transcription factors and show how to infect MEFs with these viruses in order to produce iPS cells. By using inducible lentiviruses, the expression of the four factors in controlled by the addition of doxycyline to the culture medium. The advantage of this system over the traditional retroviral infection is the ability to turn the genes on and off so that the kinetics of reprogramming and gene expression requirements can be analyzed in detail.

This video shows the procedure for generating induced pluripotent stem cells using inducible lentivirus that express Oct4, Sox2, c-Myc and Klf4.

Générer des cellules iPS de MEFS travers l&#39;expression forcée de Sox-2, Oct-4, c-Myc et Klf4

Pluripotency can be induced in differentiated murine by viral transduction of Oct4, Sox2, Klf4, and c-Myc (Takahashi and Yamanaka, 2006;  Wernig, et al., ...

From MEFs to Matrigel 3: Passaging hESCs from Matrigel onto Matrigel

This video demonstrates how to maintain the growth of human embryonic stem cells (hESCs) in feeder cell-free conditions and how to continuously passage hESCs in feeder cell-free conditions. Confirmation of hESC pluripotency grown in feeder cell-free conditions by immunofluorescence microscopy is also demonstrated.

This video demonstrates how to maintain the growth of human embryonic stem cells (hESCs) in feeder cell-free conditions and how to continuously passage hESCs in feeder cell-free conditions. Confirmation of hESC pluripotency grown in feeder cell-free conditions by immunofluorescence microscopy is also demonstrated. Part 3 of 3.

De FAE au Matrigel 3: Le repiquage du Matrigel CSEh sur Matrigel

This video demonstrates how to maintain the growth of human embryonic stem cells (hESCs) in feeder cell-free conditions and how to continuously passage ...

Cargo Loading onto Kinesin Powered Molecular Shuttles

Cells have evolved sophisticated molecular machinery, such as kinesin motor proteins and microtubule filaments, to support active intracellular transport of cargo. While kinesins tail domain binds to a variety of cargoes, kinesins head domains utilize the chemical energy stored in ATP molecules to step along the microtubule lattice. The long, stiff microtubules serve as tracks for long-distance intracellular transport.
These motors and filaments can also be employed in microfabricated synthetic environments as components of molecular shuttles 1. In a frequently used design, kinesin motors are anchored to the track surface through their tails, and functionalized microtubules serve as cargo carrying elements, which are propelled by these motors. These shuttles can be loaded with cargo by utilizing the strong and selective binding between biotin and streptavidin. The key components (biotinylated tubulin, streptavidin, and biotinylated cargo) are commercially available.
Building on the classic inverted motility assay 2, the construction of molecular shuttles is detailed here. Kinesin motor proteins are adsorbed to a surface precoated with casein; microtubules are polymerized from biotinylated tubulin, adhered to the kinesin and subsequently coated with rhodamine-labeled streptavidin. The ATP concentration is maintained at subsaturating concentration to achieve a microtubule gliding velocity optimal for loading cargo 3. Finally, biotinylated fluorescein-labeled nanospheres are added as cargo. Nanospheres attach to microtubules as a result of collisions between gliding microtubules and nanospheres adhering to the surface.
The protocol can be readily modified to load a variety of cargoes such as biotinylated DNA4, quantum dots 5 or a wide variety of antigens via biotinylated antibodies 4-6.

Molecular shuttles consisting of functionalized microtubules gliding on surface-adhered kinesin motor proteins can serve as a nanoscale transport system. Here, the assembly of a typical shuttle system is described.

Chargement du fret sur la kinésine Powered navettes moléculaires

Cells have evolved sophisticated molecular machinery, such as kinesin motor proteins and microtubule filaments, to support active intracellular transport ...

An Introduction to Worm Lab: from Culturing Worms to Mutagenesis

This protocol describes procedures to maintain nematodes in the laboratory and how to mutagenize them using two alternative methods: ethyl methane sulfonate (EMS) and 4, 5', 8-trimethylpsoralen combined with ultraviolet light (TMP/UV). Nematodes are powerful biological systems for genetics studies because of their simple body plan and mating system, which is composed of self-fertilizing hermaphrodites and males that can generate hundreds of progeny per animal. Nematodes are maintained in agar plates containing a lawn of bacteria and can be easily transferred from one plate to another using a pick. EMS is an alkylating agent commonly used to induce point mutations and small deletions, while TMP/UV mainly induces deletions. Depending on the species of nematode being used, concentrations of EMS and TMP will have to be optimized. To isolate recessive mutations of the nematode Pristionchus pacificus, animals of the F2 generation were visually screened for phenotypes. To illustrate these methods, we mutagenized worms and looked for Uncoordinated (Unc), Dumpy (Dpy) and Transformer (Tra) mutants.

Screening for mutants with phenotypic defects is a straightforward method for identifying genes that function in a given biological process. In this article we describe how to culture free living worms (e.g., Pristionchus pacificus) in the laboratory and show two different mutagenesis methods, EMS and TMP/UV.

Une introduction au laboratoire Worm: La culture de Worms à la mutagenèse

This protocol describes procedures to maintain nematodes in the laboratory and how to mutagenize them using two alternative methods: ethyl methane ...

A Matrigel-Based Tube Formation Assay to Assess the Vasculogenic Activity of Tumor Cells

Over the past several decades, a tube formation assay using growth factor-reduced Matrigel has been typically employed to demonstrate the angiogenic activity of vascular endothelial cells in vitro1-5. However, recently growing evidence has shown that this assay is not limited to test vascular behavior for endothelial cells. Instead, it also has been used to test the ability of a number of tumor cells to develop a vascular phenotype6-8. This capability was consistent with their vasculogenic behavior identified in xenotransplanted animals, a process known as vasculogenic mimicry (VM)9. There is a multitude of evidence demonstrating that tumor cell-mediated VM plays a vital role in the tumor development, independent of endothelial cell angiogenesis6, 10-13. For example, tumor cells were found to participate in the blood perfused, vascular channel formation in tissue samples from melanoma and glioblastoma patients8, 10, 11. Here, we described this tubular network assay as a useful tool in evaluation of vasculogenic activity of tumor cells. We found that some tumor cell lines such as melanoma B16F1 cells, glioblastoma U87 cells, and breast cancer MDA-MB-435 cells are able to form vascular tubules; but some do not such as colon cancer HCT116 cells. Furthermore, this vascular phenotype is dependent on cell numbers plated on the Matrigel. Therefore, this assay may serve as powerful utility to screen the vascular potential of a variety of cell types including vascular cells, tumor cells as well as other cells.

A tube formation assay is used to evaluate vascular activity of tumor cells.

Un test basé sur Matrigel Tube Formation d&#39;évaluer l&#39;activité Vasculogenic de cellules tumorales

Over the past several decades, a tube formation assay using growth factor-reduced Matrigel has been typically employed to demonstrate the angiogenic ...

Generation of Myospheres From hESCs by Epigenetic Reprogramming

Generation of a homogeneous and abundant population of skeletal muscle cells from human embryonic stem cells (hESCs) is a requirement for cell-based therapies and for a "disease in a dish" model of human neuromuscular diseases. Major hurdles, such as low abundance and heterogeneity of the population of interest, as well as a lack of protocols for the formation of three-dimensional contractile structures, have limited the applications of stem cells for neuromuscular disorders. We have designed a protocol that overcomes these limits by ectopic introduction of defined factors in hESCs - the muscle determination factor MyoD and SWI/SNF chromatin remodeling complex component BAF60C - that are able to reprogram hESCs into skeletal muscle cells. Here we describe the protocol established to generate hESC-derived myoblasts and promote their clustering into tridimensional miniaturized structures (myospheres) that functionally mimic miniaturized skeletal muscles7.

Here, we describe a protocol based on epigenetic reprogramming of human embryonic stem cells (hESCs) toward generating a homogeneous population of skeletal muscle progenitors that under permissive culture conditions form three-dimensional clusters of contractile myofibers (myospheres), which recapitulate biological features of human skeletal muscles.

Génération de Myospheres De CSEh par Reprogrammation épigénétique

Generation of a homogeneous and abundant population of skeletal muscle cells from human embryonic stem cells (hESCs) is a requirement for cell-based ...

Xenopus laevis as a Model to Identify Translation Impairment

Protein synthesis is a fundamental process to gene expression impacting diverse biological processes notably adaptation to environmental conditions. The initiation step, which involves the assembly of the ribosomal subunits at the mRNA initiation codon, involved initiation factor including eIF4G1. Defects in this rate limiting step of translation are linked to diverse disorders. To study the potential consequences of such deregulations, Xenopus laevis oocytes constitute an attractive model with high degrees of conservation of essential cellular and molecular mechanisms with human. In addition, during meiotic maturation, oocytes are transcriptionally repressed and all necessary proteins are translated from preexisting, maternally derived mRNAs. This inexpensive model enables exogenous mRNA to become perfectly integrated with an effective translation. Here is described a protocol for assessing translation with a factor of interest (here eIF4G1) using stored maternal mRNA that are the first to be polyadenylated and translated during oocyte maturation as a physiological readout. At first, mRNA synthetized by in vitro transcription of plasmids of interest (here eIF4G1) are injected in oocytes and kinetics of oocyte maturation by Germinal Vesicle Breakdown detection is determined. The studied maternal mRNA target is the serine/threonine-protein-kinase mos. Its polyadenylation and its subsequent translation are investigated together with the expression and phosphorylation of proteins of the mos signaling cascade involved in oocyte maturation. Variations of the current protocol to put forward translational defects are also proposed to emphasize its general applicability. In light of emerging evidence that aberrant protein synthesis may be involved in the pathogenesis of neurological disorders, such a model provides the opportunity to easily assess this impairment and identify new targets.

Xenopus laevis as a Model to Identify Translation Impairment

Protein synthesis control occurs mainly at the translation initiation step, deficiencies in which are linked to diverse disorders. To better understand their etiology, we described here a protocol using Xenopus laevis oocytes assessing the translation of mos transcript in the presence of a mutant of translation initiation factor eIF4G1.

Xenopus laevis comme un modèle pour identifier traduction Dépréciation

Protein synthesis is a fundamental process to gene expression impacting diverse biological processes notably adaptation to environmental conditions. The ...

Using Caco-2 Cells to Study Lipid Transport by the Intestine

Studies of dietary fat absorption are generally conducted by using an animal model equipped with a lymph cannula. Although this animal model is widely accepted as the in vivo model of dietary fat absorption, the surgical techniques involved are challenging and expensive. Genetic manipulation of the animal model is also costly and time consuming. The alternative in vitro model is arguably more affordable, timesaving, and less challenging. Importantly, the in vitro model allows investigators to examine the enterocytes as an isolated system, reducing the complexity inherent in the whole organism model. This paper describes how human colon carcinoma cells (Caco-2) can serve as an in vitro model to study the enterocyte transport of lipids, and lipid-soluble drugs and vitamins. It explains the proper maintenance of Caco-2 cells and the preparation of their lipid mixture; and it further discusses the valuable option of using the permeable membrane system. Since differentiated Caco-2 cells are polarized, the main advantage of using the permeable membrane system is that it separates the apical from the basolateral compartment. Consequently, the lipid mixture can be added to the apical compartment while the lipoproteins can be collected from the basolateral compartment. In addition, the effectiveness of the lentivirus expression system in upregulating gene expression in Caco-2 cells is discussed. Lastly, this paper describes how to confirm the successful isolation of intestinal lipoproteins by transmission electron microscopy (TEM).

Caco-2 cells can serve as an in vitro model to study the enterocyte transport of lipids, and lipid-soluble drugs/vitamins. The permeable membrane system separates the apical from the basolateral compartment, while the lentivirus expression system offers an effective gene overexpression method. The isolation of lipoproteins is confirmed by TEM.

Utilisation de cellules Caco-2 à étudier Lipid Transport par l&#39;intestin

Studies of dietary fat absorption are generally conducted by using an animal model equipped with a lymph cannula. Although this animal model is widely ...

Magnetic-Activated Cell Sorting Strategies to Isolate and Purify Synovial Fluid-Derived Mesenchymal Stem Cells from a Rabbit Model

Mesenchymal stem cells (MSCs) are the main cell source for cell-based therapy. MSCs from articular cavity synovial fluid could potentially be used for cartilage tissue engineering. MSCs from synovial fluid (SF-MSCs) have been considered promising candidates for articular regeneration, and their potential therapeutic benefit has made them an important research topic of late. SF-MSCs from the knee cavity of the New Zealand white rabbit can be employed as an optimized translational model to assess human regenerative medicine. By means of CD90-based magnetic activated cell sorting (MACS) technologies, this protocol successfully obtains rabbit SF-MSCs (rbSF-MSCs) from this rabbit model and further fully demonstrates the MSC phenotype of these cells by inducing them to differentiate to osteoblasts, adipocytes, and chondrocytes. Therefore, this approach can be applied in cell biology research and tissue engineering using simple equipment and procedures.

This article presents a simple and economic protocol for the straightforward isolation and purification of mesenchymal stem cells from New Zealand white rabbit synovial fluid.

Cellule magnétique activé tri des stratégies visant à isoler et purifier Synovial fluide mésenchymateuses cellules souches dérivées d’un modèle de lapin

Mesenchymal stem cells (MSCs) are the main cell source for cell-based therapy. MSCs from articular cavity synovial fluid could potentially be used for ...