A novel method to generate and culture human mast cells: Peripheral CD34+ stem cell-derived mast cells (PSCMCs)

Schmetzer, Oliver; Valentin, Patricia; Smorodchenko, Anna; Domenis, Rossana; Gri, Giorgia; Siebenhaar, Frank; Metz, Martin; Maurer, Marcus

doi:10.1016/j.jim.2014.07.003

Cited by 35 publications

(38 citation statements)

References 17 publications

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“…35 In the current study, we used primary human MC derived from peripheral CD34 + cells, which may better represent qualities of CRC-infiltrating MC subtype (MC TC or submucosal MC) including the expression of FcɛRI, tryptase and chymase. 37 …”

Section: Discussionmentioning

confidence: 99%

“…37 Briefly, frozen stem cell concentrates were rapidly thawed at 37°C under sterile conditions, and poured in a large cell culture flask (Greiner, 661195). 20% human serum albumin clinical solution (HSA) (Sanquin), 6% hydroxyethyl starch clinical solution (Braun) and RMPI containing 10 U/ml Heparin (LEO pharma) were then added slowly and consecutively to the cell concentrate.…”

Section: Methodsmentioning

confidence: 99%

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Human mast cells promote colon cancer growth via bidirectional crosstalk: studies in 2D and 3D coculture models

Blokhuis

Derks

et al. 2018

OncoImmunology

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Chronic inflammation drives the development of colorectal cancer (CRC), where tumor-infiltrating immune cells interact with cancer cells in a dynamic crosstalk. Mast cells (MC), one of earliest recruited immune cells, accumulate in CRC tissues and their density is correlated with cancer progression. However, the exact contribution of MC in CRC and their interaction with colon cancer cells is poorly understood. Here, we investigated the impact of primary human MC and their mediators on colon cancer growth using 2D and 3D coculture models. Primary human MC were generated from peripheral CD34+ stem cells. Transwell chambers were used to analyze MC chemotaxis to colon cancer. Colon cancer cells HT29 and Caco2 differentially recruited MC by releasing CCL15 or SCF, respectively. Using BrdU proliferation assays, we demonstrated that MC can directly support colon cancer proliferation and this effect was mediated by their cellular crosstalk. 3D coculture models with cancer spheroids further confirmed the pro-tumor effect of MC on colon cancer growth, where direct cell-cell contact is dispensable and increased production of multiple soluble mediators was detected. Moreover, TLR2 stimulation of MC promoted stronger growth of colon cancer spheroids. By examining the transcriptome profile of colon cancer-cocultured MC versus control MC, we identified several MC marker genes, which were deregulated in expression. Our study provides an advanced in vitro model to investigate the role of human MC in cancer. Our data support the detrimental role of MC in CRC development and provide a molecular insight into the cellular crosstalk between MC and colon cancer cells.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Human mast cells promote colon cancer growth via bidirectional crosstalk: studies in 2D and 3D coculture models

Blokhuis

Derks

et al. 2018

OncoImmunology

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“…We also evaluated lymphocytapheresis as source of CD34 + cells. We thus describe a method for culturing huMC that uses total culture medium volumes of 20 to 30 ml for one culture to generate MCs by 7 weeks, against as much as 1L of culture medium for some other techniques [1, 4–6, 21]. This modified method is more cost effective (for cost comparison see Supplementary Table 1) and also less labor intensive since cultures only need to be attended to biweekly during the first 6 weeks of culture vs weekly in other methods.…”

Section: Discussionmentioning

confidence: 99%

An optimized protocol for the generation and functional analysis of human mast cells from CD34 + enriched cell populations

Yin

Bai

Olivera

et al. 2017

Journal of Immunological Methods

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The culture of mast cells from human tissues such a cord blood, peripheral blood or bone marrow aspirates has advanced our understanding of human mast cells (huMC) degranulation, mediator production and response to pharmacologic agents. However, existing methods for huMC culture tend to be laborious and expensive. Combining technical approaches from several of these protocols, we designed a simplified and more cost effective approach to the culture of mast cells from human cell populations including peripheral blood and cryopreserved cells from lymphocytapheresis. On average, we reduced by 30–50 fold the amount of culture media compared to our previously reported method, while the total MC number generated by this method (2.46 ± 0.63 × 106 vs. 2.4 ± 0.28 × 106, respectively, from 1.0 × 108 lymphocytapheresis or peripheral blood mononuclear blood cells [PBMCs]) was similar to previous method (2.36 ± 0.70 × 106), resulting in significant budgetary savings. In addition, we compared the yield of huMCs with or without IL-3 added to early cultures in the presence of stem cell factor (SCF) and interlukin-6 (IL-6) and found that the total MC number generated, while higher with IL-3 in the culture, did not reach statistical significance, suggesting that IL-3, often recommended in the culture of huMCs, is not absolutely required. We then performed a functional analysis by flow cytometry using standard methods and which maximized the data we could obtain from cultured cells. We believe these approaches will allow more laboratories to culture and examine huMC behavior going forward.

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“…Intestinal mast cells were evaluated by toluidine blue staining as previously described (28). Briefly, cells were fixed with 4% paraformaldehyde (PFA) (Sigma-Aldrich) for 10 min at room temperature (RT) and then stained with 1% toluidine blue (Sigma-Aldrich) for 1 h at RT, and subsequently, cells were washed in distilled water 3 times and covered by use of a coverslip and mounting medium.…”

Section: Methodsmentioning

confidence: 99%

Human Mucosal Mast Cells Capture HIV-1 and Mediate Viral trans -Infection of CD4 ⁺ T Cells

Jiang

Wei

et al. 2016

J Virol

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Despite great advances in antiretroviral therapies, human immunodeficiency virus type 1 (HIV-1) infection still remains a major global epidemic. Sexual transmission is the principal route of HIV-1 acquisition, making the genital and rectal mucosae the major sites of viral transmission. The intestinal mucosa is also the primary site where HIV-1 amplifies to disseminate virus throughout the host and is critical in the early events in the establishment of infection and evasion of immune defenses. However, the mechanisms contributing to the establishment of HIV-1 primary infection remain largely unexplored. Cell-associated viral dissemination has been proposed to play pivotal roles in HIV-1 primary infection, and multiple cell types, such as dendritic cells (DCs) and macrophages, have been reported to be hijacked by HIV-1 for local and systemic viral spread (1-4). DCs provide one of the best-described cell models for understanding cell-mediated HIV-1 capture and dissemination (1,(5)(6)(7)(8).Mast cells are derived from hematopoietic progenitor cells and undergo final maturation in vascularized tissues. Mast cells are strategically in close contact with the host-environment interface, such as the skin, airway, gastrointestinal tract, and urinary tract. They express numerous pathogen-associated molecular patterns and play an important role in the early immunosurveillance for many pathogens (9, 10). Mast cells can interact with various immune cells in complex ways, including release of soluble factors and direct contact (11), and are important immune effector and modulatory cells that help to link innate and adaptive immunity in the fight against pathogens (9,(12)(13)(14). They have been shown to be important for host defense against various viruses, such as vesicular stomatitis virus, Sendai virus, hantavirus, reovirus, dengue virus, influenza virus, herpes simplex virus, and murine cytomegalovirus (15-23). Additionally, mast cells can serve as antigenpresenting cells and participate in traditional immunologic synapse formation with T cells to mediate antigen-specific T cell activation (24).

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A novel method to generate and culture human mast cells: Peripheral CD34+ stem cell-derived mast cells (PSCMCs)

Cited by 35 publications

References 17 publications

Human mast cells promote colon cancer growth via bidirectional crosstalk: studies in 2D and 3D coculture models

Human mast cells promote colon cancer growth via bidirectional crosstalk: studies in 2D and 3D coculture models

An optimized protocol for the generation and functional analysis of human mast cells from CD34 + enriched cell populations

Human Mucosal Mast Cells Capture HIV-1 and Mediate Viral trans -Infection of CD4 ⁺ T Cells

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A novel method to generate and culture human mast cells: Peripheral CD34+ stem cell-derived mast cells (PSCMCs)

Cited by 35 publications

References 17 publications

Human mast cells promote colon cancer growth via bidirectional crosstalk: studies in 2D and 3D coculture models

Human mast cells promote colon cancer growth via bidirectional crosstalk: studies in 2D and 3D coculture models

An optimized protocol for the generation and functional analysis of human mast cells from CD34 + enriched cell populations

Human Mucosal Mast Cells Capture HIV-1 and Mediate Viral trans -Infection of CD4 + T Cells

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Product

Resources

About

Human Mucosal Mast Cells Capture HIV-1 and Mediate Viral trans -Infection of CD4 ⁺ T Cells