MesenCult™ Expansion Kit (Mouse)

For the culture of mouse MSCs and MEFs

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MesenCult™ Expansion Kit (Mouse)

For the culture of mouse MSCs and MEFs

1 Kit
Catalog #05513
234 USD

Required Products

Overview

MesenCult™ Expansion Kit (Mouse) is standardized for the culture of mouse mesenchymal stromal cells (MSCs; also known as mesenchymal stem cells) and mouse embryonic fibroblasts (MEFs). The kit includes MesenCult™ Basal Medium (Mouse), MesenCult™ 10X Supplement (Mouse), and MesenPure™. MesenCult™ Expansion Medium has been optimized for the derivation and expansion of mouse MSCs and MEFs in vitro as well as for the detection of colony-forming unit–fibroblasts (CFU­F). This kit was optimized using cells from the mouse strain C57BL/6.
To facilitate the enrichment of MSCs and MEFs during cell culture without serial passaging and frequent medium changes, simply add MesenPure™ to complete MesenCult™ Expansion Medium just prior to use. Although not required, the addition of MesenPure™ is strongly recommended, as the resulting MSC and MEF cultures are more homogeneous and exhibit more robust proliferation, differentiation, and colony formation when compared to complete MesenCult™ Expansion Medium alone.
NOTE: MesenCult™ Expansion Medium must be supplemented with L-Glutamine (Catalog #07100).
Advantages:
• Fast expansion of mouse MSCs with robust enrichment as early as passage 0.
• Optimized for use with mouse bone marrow-, compact bone- and adipose-derived MSCs and MEFs.
• Obtain homogeneous mouse MSC cultures while maintaining tri-lineage differentiation potential.
• Rigorous raw material screening and quality control minimize lot-to-lot variability and increase reproducibility between experiments.
Components:
  • MesenCult™ Basal Medium (Mouse), 450 mL
  • MesenCult™ 10X Supplement (Mouse), 50 mL
  • MesenPure™, 0.5 mL
Subtype:
Specialized Media
Cell Type:
Mesenchymal Stem and Progenitor Cells; Mouse Embryonic Fibroblasts
Species:
Mouse
Application:
Cell Culture; Colony Assay; Expansion
Brand:
MesenCult
Area of Interest:
Stem Cell Biology

Scientific Resources

Educational Materials

(5)

Product Applications

This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.

Data and Publications

Data

Procedure Summary for Hematopoietic CFU Assays

Figure 1. CFU-F Assay Comparing Mouse Bone Marrow (BM) MSCs Derived and Cultured in MesenCult™ Expansion Medium With and Without MesenPure™, and Other Commercially Available Media

Numerous CFU-F colonies were observed in cultures maintained in (A) MesenCult™ Expansion Medium (Control) and in (B) same medium containing MesenPure™. Few to no colony formation were observed when cultures were maintained in (C) Commercial Medium 1 or (D) Commercial Medium 2. Seeding density: 5x10^4 cells/cm^2.

Procedure Summary for Hematopoietic CFU Assays

Figure 2. Long-Term Expansion of Mouse BM-Derived MSCs is Observed When Cells are Cultured in MesenCult™ Expansion Medium

Mouse BM MSCs, derived and cultured in MesenCult™ Expansion Medium (Control), show superior long-term expansion rate compared to Commercial Medium 1 and 2. The addition of MesenPure™ enriches for MSCs as early as passage 0 and further improves the expansion rate beyond passage 8. The doubling time of mouse MSCs cultured with or without MesenPure™ are 2.29 and 3.01, respectively. BM MSCs culture-expanded using the MesenCult™ Expansion Kit, with or without MesenPure™, were done under hypoxic conditions. BM MSCs culture-expanded in Commercial Medium 1 and 2 were culture-expanded under normoxic conditions as recommended by their protocols. Data shown from one representative experiment (n=3).

Procedure Summary for Hematopoietic CFU Assays

Figure 3. Mouse BM- and Compact Bone (CB)-Derived MSCs Culture-expanded in MesenCult™ Expansion Medium With or Without MesenPure™ Maintain Multi-Lineage Differentiation Potential

Enriched populations of MSCs were observed at earlier passages upon addition of MesenPure™, which showed increased and more dense differentiation than control cultures. (A) Mouse BM MSCs culture-expanded in MesenCult™ Expansion Medium (Control) differentiated into (B) adipocytes; and (C) osteoblasts. (D) Mouse BM-derived MSCs culture-expanded with MesenPure™ differentiated into (E) adipocytes; and (F) osteoblasts . Differentiation of mouse BM MSCs into chondrocytes is in progress. (G) Mouse CB MSCs culture-expanded in MesenCult™ Expansion Medium (Control) differentiated into (H) adipocytes, (I) osteoblasts and (J) chondrocytes. Adipose-derived mesenchymal stem and progenitor cells, and mouse embryonic fibroblasts (MEFs) were derived and culture-expanded using the MesenCult™ Expansion Kit. These cells were also differentiated towards the adipogenic and osteogenic lineages (data not shown). Adipocytes were stained with Oil Red O staining. Osteoblasts were stained with Alkaline phosphatase and silver nitrate (von Kossa). Chondrocytes were stained with Alcian Blue and Nuclear Fast Red. Images were taken at passage 2.

Procedure Summary for Hematopoietic CFU Assays

Figure 4. Flow Cytometric Analysis of Culture-Expanded Mouse BM-Derived MSCs Using the MesenCult™ Expansion Kit

Mouse BM MSCs were culture-expanded in MesenCult™ Expansion Medium (Control) or with MesenPure™. MSCs from passage 2 were stained for the mesenchymal surface markers, CD106 and Sca1, and the hematopoietic marker, CD45. Stained cells were then analyzed by flow cytometry. MSCs culture-expanded in Control medium show distinct populations of CD45+ hematopoietic cells and CD45- (CD106+ and Sca1+) MSCs. Upon addition of MesenPure™ to the Control Medium, an enriched and homogenous population of CD45- (CD106+ and Sca1+) MSCs are obtained.

Publications

(11)
Nature communications 2020 jan

Gastric squamous-columnar junction contains a large pool of cancer-prone immature osteopontin responsive Lgr5-CD44+ cells.

D.-J. Fu et al.

Abstract

Areas of a junction between two types of epithelia are known to be cancer-prone in many organ systems. However, mechanisms for preferential malignant transformation at the junction areas remain insufficiently elucidated. Here we report that inactivation of tumor suppressor genes Trp53 and Rb1 in the gastric squamous-columnar junction (SCJ) epithelium results in preferential formation of metastatic poorly differentiated neoplasms, which are similar to human gastroesophageal carcinoma. Unlike transformation-resistant antral cells, SCJ cells contain a highly proliferative pool of immature Lgr5-CD44+ cells, which are prone to transformation in organoid assays, comprise early dysplastic lesions, and constitute up to 30{\%} of all neoplastic cells. CD44 ligand osteopontin (OPN) is preferentially expressed in and promotes organoid formation ability and transformation of the SCJ glandular epithelium. OPN and CD44 overexpression correlate with the worst prognosis of human gastroesophageal carcinoma. Thus, detection and selective targeting of the active OPN-CD44 pathway may have direct clinical relevance.
Cell reports 2019 sep

Natural Killer Cells Suppress T Cell-Associated Tumor Immune Evasion.

A. J. Freeman et al.

Abstract

Despite the clinical success of cancer immunotherapies, the majority of patients fail to respond or develop resistance through disruption of pathways that promote neo-antigen presentation on MHC I molecules. Here, we conducted a series of unbiased, genome-wide CRISPR/Cas9 screens to identify genes that limit natural killer (NK) cell anti-tumor activity. We identified that genes associated with antigen presentation and/or interferon-$\gamma$ (IFN-$\gamma$) signaling protect tumor cells from NK cell killing. Indeed, Jak1-deficient melanoma cells were sensitized to NK cell killing through attenuated NK cell-derived IFN-$\gamma$-driven transcriptional events that regulate MHC I expression. Importantly, tumor cells that became resistant to T cell killing through enrichment of MHC I-deficient clones were highly sensitive to NK cell killing. Taken together, we reveal the genes targeted by tumor cells to drive checkpoint blockade resistance but simultaneously increase their vulnerability to NK cells, unveiling NK cell-based immunotherapies as a strategy to antagonize tumor immune escape.
JCI insight 2019 sep

Comparative pathogenesis of Ebola virus and Reston virus infection in humanized mice.

B. Escudero-P\'erez et al.

Abstract

Filoviruses of the genus Ebolavirus include five species with marked differences in their ability to cause disease in humans. From the highly virulent Ebola virus to the seemingly nonpathogenic Reston virus, case-fatality rates can range between 0-90{\%}. In order to understand the molecular basis of these differences it is imperative to establish disease models that recapitulate human disease as faithfully as possible. Non-human primates are the gold-standard models for filovirus pathogenesis, but comparative studies are skewed by the fact that Reston virus infection can be lethal for NHP. Here we have used HLA-A2 transgenic, NOD-scid-interleukin 2$\gamma$ receptor knockout (NSG-A2) mice reconstituted with human hematopoiesis to compare Ebola virus and Reston virus pathogenesis in a human-like environment. While significantly less pathogenic than Ebola virus, Reston virus killed 20{\%} of infected mice, a finding that was linked to exacerbated inflammation and viral replication in the liver. In addition, 'humanized' mice recapitulated the case-fatality ratios of different Ebolavirus species in humans. Our findings point out at humanized mice as a putative model to test the pathogenicity of newly discovered filoviruses, and warrants further investigations on Reston virus pathogenesis in humans.
The Journal of cell biology 2019 nov

Loss of myosin Vb promotes apical bulk endocytosis in neonatal enterocytes.

A. C. Engevik et al.

Abstract

In patients with inactivating mutations in myosin Vb (Myo5B), enterocytes show large inclusions lined by microvilli. The origin of inclusions in small-intestinal enterocytes in microvillus inclusion disease is currently unclear. We postulated that inclusions in Myo5b KO mouse enterocytes form through invagination of the apical brush border membrane. 70-kD FITC-dextran added apically to Myo5b KO intestinal explants accumulated in intracellular inclusions. Live imaging of Myo5b KO-derived enteroids confirmed the formation of inclusions from the apical membrane. Treatment of intestinal explants and enteroids with Dyngo resulted in accumulation of inclusions at the apical membrane. Inclusions in Myo5b KO enterocytes contained VAMP4 and Pacsin 2 (Syndapin 2). Myo5b;Pacsin 2 double-KO mice showed a significant decrease in inclusion formation. Our results suggest that apical bulk endocytosis in Myo5b KO enterocytes resembles activity-dependent bulk endocytosis, the primary mechanism for synaptic vesicle uptake during intense neuronal stimulation. Thus, apical bulk endocytosis mediates the formation of inclusions in neonatal Myo5b KO enterocytes.
Stem cells translational medicine 2019 jun

Mesenchymal Stromal Cells Modulate Corneal Alloimmunity via Secretion of Hepatocyte Growth Factor.

S. K. Mittal et al.

Abstract

Mesenchymal stromal cells (MSCs) are multipotent stem cells that participate in tissue repair and posses considerable immunomodulatory potential. MSCs have been shown to promote allograft survival, yet the mechanisms behind this phenomenon have not been fully defined. Here, we investigate the capacity of MSCs to suppress the allogeneic immune response by secreting the pleiotropic molecule hepatocyte growth factor (HGF). Using an in vivo mouse model of corneal transplantation, we report that MSCs promote graft survival in an HGF-dependent manner. Moreover, our data indicate that topically administered recombinant HGF (1) suppresses antigen-presenting cell maturation in draining lymphoid tissue, (2) limits T-helper type-1 cell generation, (3) decreases inflammatory cell infiltration into grafted tissue, and (4) is itself sufficient to promote transplant survival. These findings have potential translational implications for the development of HGF-based therapeutics. Stem Cells Translational Medicine 2019.
PRODUCTS ARE FOR RESEARCH USE ONLY AND NOT INTENDED FOR HUMAN OR ANIMAL DIAGNOSTIC OR THERAPEUTIC USES UNLESS OTHERWISE STATED. FOR ADDITIONAL INFORMATION ON QUALITY AT STEMCELL, REFER TO WWW.STEMCELL.COM/COMPLIANCE.