Optimization of<i> Ex Vivo</i> Murine Bone Marrow Derived Immature Dendritic Cells: A Comparative Analysis of Flask Culture Method and Mouse CD11c Positive Selection Kit Method

Gosavi, Rahul Ashok; Salwe, Sukeshani; Mukherjee, Sandeepan; Dahake, Ritwik; Kothari, Sweta; Patel, Vainav; Chowdhary, Abhay; Deshmukh, Ranjana

doi:10.1155/2018/3495086

Cited by 8 publications

(4 citation statements)

References 30 publications

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“…For differentiation into bone-marrow derived macrophages (BMDM), non-adherent cells were moved to bacteriological Petri dishes the next day and differentiated over 1 week in cDMEM containing 20 ng/mL recombinant murine M-CSF (Biolegend,576404). Generation of murine bone-marrow derived dendritic cells (BMDC) has been previously described (Gosavi et al, 2018). In brief, bone marrow progenitor cells were collected in cRPMI and replated in cRPMI containing 20 ng/mL recombinant murine GM-CSF (Peprotech, 315-03) (DC media).…”

Section: Cell Linesmentioning

confidence: 99%

Phagocytic ‘teeth’ and myosin-II ‘jaw’ power target constriction during phagocytosis

Vorselen

Barger

Wang

et al. 2021

eLife

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Phagocytosis requires rapid actin reorganization and spatially controlled force generation to ingest targets ranging from pathogens to apoptotic cells. How actomyosin activity directs membrane extensions to engulf such diverse targets remains unclear. Here, we combine lattice light-sheet microscopy (LLSM) with microparticle traction force microscopy (MP-TFM) to quantify actin dynamics and subcellular forces during macrophage phagocytosis. We show that spatially localized forces leading to target constriction are prominent during phagocytosis of antibody-opsonized targets. This constriction is largely driven by Arp2/3-mediated assembly of discrete actin protrusions containing myosin 1e and 1f (‘teeth’) that appear to be interconnected in a ring-like organization. Contractile myosin-II activity contributes to late-stage phagocytic force generation and progression, supporting a specific role in phagocytic cup closure. Observations of partial target eating attempts and sudden target release via a popping mechanism suggest that constriction may be critical for resolving complex in vivo target encounters. Overall, our findings present a phagocytic cup shaping mechanism that is distinct from cytoskeletal remodeling in 2D cell motility and may contribute to mechanosensing and phagocytic plasticity.

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Section: Cell Linesmentioning

confidence: 99%

Phagocytic ‘teeth’ and myosin-II ‘jaw’ power target constriction during phagocytosis

Vorselen

Barger

Wang

et al. 2021

eLife

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“…In addition, CD11c is a type I transmembrane protein that is expressed on monocytes, granulocytes, a subset of B cells, DCs and macrophages. Highly enriched CD11c+ DCs can be differentiated from mouse bone marrow [ 27 ] to assess their activation or maturation status by various agents ex vivo.…”

Section: Introductionmentioning

confidence: 99%

A Novel Thienopyrimidine Analog, TPH104, Mediates Immunogenic Cell Death in Triple-Negative Breast Cancer Cells

Tukaramrao

Malla

Saraiya

et al. 2021

Cancers

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Enhancing the tumor immunogenic microenvironment has been suggested to circumvent triple-negative breast cancer (TNBC) resistance and increase the efficacy of conventional chemotherapy. Here, we report a novel chemotherapeutic compound, TPH104, which induces immunogenic cell death in the TNBC cell line MDA-MB-231, by increasing the stimulatory capacity of dendritic cells (DCs), with an IC50 value of 140 nM. TPH104 (5 µM) significantly increased ATP levels in the supernatant and mobilized intracellular calreticulin to the plasma membrane in MDA-MB-231 cells, compared to cells incubated with the vehicle. Incubating MDA-MB-231 cells for 12 h with TPH104 (1–5 µM) significantly increased TNFα mRNA levels. The supernatants of dying MDAMB-231 cells incubated with TPH104 increased mouse bone marrow-derived DC maturation, the expression of MHCII and CD86 and the mRNA expression of TNFα, IL6 and IL12. Overall, these results indicate that TPH104 induces immunogenic cell death in TNBC cells, in part, by activating DCs.

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“…In fact, when culturing cells, it is necessary to adjust the external environmental conditions, such as pH, CO 2 concentration, and temperature of the culture medium, for cells to grow in an environment most suitable for them, and these conditions will vary depending on cell type and experimental system. It is necessary to select the cell culture vessel, such as culture dishes or flasks, according to the desired purpose [7,8]. However, what is widely used in laboratories are the commercially available culture dishes sold by manufacturers [9].…”

Section: Introductionmentioning

confidence: 99%

Fabrication of a Novel Culture Dish Adapter with a Small Recess Structure for Flow Control in a Closed Environment

Yasuda

Adachi

Okonogi³

et al. 2019

Applied Sciences

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Cell culture medium replacement is necessary to replenish nutrients and remove waste products, and perfusion and batch media exchange methods are available. The former can establish an environment similar to that in vivo, and microfluidic devices are frequently used. However, these methods are hampered by incompatibility with commercially available circular culture dishes and the difficulty in controlling liquid flow. Here, we fabricated a culture dish adapter using polydimethylsiloxane that has a small recess structure for flow control compatible with commercially available culture dishes. We designed U-shaped and I-shaped recess structure adapters and we examined the effects of groove structure on medium flow using simulation. We found that the U-shaped and I-shaped structures allowed a uniform and uneven flow of medium, respectively. We then applied these adaptors to 293T cell culture and examined the effects of recess structures on cell proliferation. As expected, cell proliferation was similar in each area of a dish in the U-shaped structure adapter, whereas in the early flow area in the I-shaped structure adapter, it was significantly higher. In summary, we succeeded in controlling liquid flow in culture dishes with the fabricated adapter, as well as in applying the modulation of culture medium flow to control cell culture.

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Optimization of Ex Vivo Murine Bone Marrow Derived Immature Dendritic Cells: A Comparative Analysis of Flask Culture Method and Mouse CD11c Positive Selection Kit Method

Cited by 8 publications

References 30 publications

Phagocytic ‘teeth’ and myosin-II ‘jaw’ power target constriction during phagocytosis

Phagocytic ‘teeth’ and myosin-II ‘jaw’ power target constriction during phagocytosis

A Novel Thienopyrimidine Analog, TPH104, Mediates Immunogenic Cell Death in Triple-Negative Breast Cancer Cells

Fabrication of a Novel Culture Dish Adapter with a Small Recess Structure for Flow Control in a Closed Environment

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