scholarly journals Anti-Cancer Activity Profiling of Chemotherapeutic Agents in 3D Co-Cultures of Pancreatic Tumor Spheroids with Cancer-Associated Fibroblasts and Macrophages

Cancers ◽  
2021 ◽  
Vol 13 (23) ◽  
pp. 5955
Author(s):  
So-Dam Jang ◽  
Jeeyeun Song ◽  
Hyun-Ah Kim ◽  
Chang-Nim Im ◽  
Iftikhar Ali Khawar ◽  
...  
Keyword(s):  
Stromal Cells ◽  
Stromal Cell ◽  
Pancreatic Tumor ◽  
Culture Conditions ◽  
Pancreatic Stellate Cells ◽  
Type I ◽  
M2 Macrophages ◽  
Tumor Spheroids ◽  
Activity Profiling ◽  
Anti Cancer

Activated pancreatic stellate cells (aPSCs) and M2 macrophages modulate tumor progression and therapeutic efficacy in pancreatic ductal adenocarcinoma (PDAC) via epithelial-mesenchymal transition (EMT). Here, our aim was to analyze the anti-invasion effects of anti-cancer agents where EMT-inducing cancer-stroma interaction occurs under three-dimensional (3D) culture conditions. We used microfluidic channel chips to co-culture pancreatic tumor spheroids (TSs) with aPSCs and THP-1-derived M2 macrophages (M2 THP-1 cells) embedded in type I collagen. Under stromal cell co-culture conditions, PANC-1 TSs displayed elevated expression of EMT-related proteins and increased invasion and migration. When PANC-1 TSs were exposed to gemcitabine, 5-fluorouracil, oxaliplatin, or paclitaxel, 30–50% cells were found unaffected, with no significant changes in the dose-response profiles under stromal cell co-culture conditions. This indicated intrinsic resistance to these drugs and no further induction of drug resistance by stromal cells. Paclitaxel had a significant anti-invasion effect; in contrast, oxaliplatin did not show such effect despite its specific cytotoxicity in M2 THP-1 cells. Overall, our findings demonstrate that the TS-stroma co-culture model of PDAC is useful for activity profiling of anti-cancer agents against cancer and stromal cells, and analyzing the relationship between anti-stromal activity and anti-invasion effects.

scholarly journals Dissecting the hematopoietic microenvironment. IX. Further characterization of murine bone marrow stromal cells

Blood ◽  
1993 ◽  
Vol 81 (5) ◽  
pp. 1205-1213 ◽  
Author(s):  
PE Penn ◽  
DZ Jiang ◽  
RG Fei ◽  
E Sitnicka ◽  
NS Wolf
Keyword(s):  
Bone Marrow ◽  
Smooth Muscle ◽  
Stromal Cell ◽  
Culture Conditions ◽  
Antigenic Determinants ◽  
Collagen Type Iv ◽  
Positive Staining ◽  
Type I ◽  
Stromal Fibroblasts ◽  
Murine Bone Marrow

Abstract We have previously shown the adherent nontransformed, nonimmortalized murine bone marrow stromal cell (BMSC) population to consist of phagocytic macrophage and endothelial-like cells and nonphagocytic fibroblasts. Both colonial and near confluent growth of each cell type was obtained following magnetic bead separation, subsequent passaging, and sustained culture with fetal bovine serum and cytokines. Monoclonal antibody staining of antigenic determinants was used to characterize the phenotype of the stromal cell population in primary platings of murine colony-forming unit fibroblast and long-term bone marrow cultures. The antibodies MECA-99, MECA-32, and MJ7–18, raised against murine vascular endothelial antigenic determinants, and von Willebrand's factor all stained selectively for the rounded endothelial- like cells. Endothelial-like cells as well as macrophages expressed the myeloid surface antigens F4/80, 7/4, and Mac-1 under our culture conditions. The cytoskeleton of the stromal fibroblasts in culture was shown to express smooth muscle-specific actin isoforms, as evidenced by positive staining of stress fibers for alpha smooth muscle-1, CGA-7 (alpha/gamma isoforms), and HHF-35 (recognizes all muscle-specific actins). Under culture conditions, stromal fibroblasts were also found to be positive for a polyclonal smooth muscle myosin. It was found that these fibroblasts stained for collagens type I, III, and IV in our cultures. Although collagen type IV is considered a by-product of endothelial cells, endothelial-like cells in our cultures did not stain for any of the collagen types. We propose a classification listing for murine BMSCs as macrophages, endotheliallike cells, and fibroblasts that display smooth muscle-like characteristics in culture.

scholarly journals Stromal cells in myeloid and lymphoid long-term bone marrow cultures can support multiple hemopoietic lineages and modulate their production of hemopoietic growth factors

Blood ◽  
1986 ◽  
Vol 68 (6) ◽  
pp. 1348-1354 ◽  
Author(s):  
A Johnson ◽  
K Dorshkind
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Culture Conditions ◽  
B Lymphopoiesis ◽  
Bone Marrow Cultures ◽  
Factor Secretion ◽  
Marrow Cultures

Abstract Hemopoiesis in long-term bone marrow cultures (LTBMC) is dependent on adherent stromal cells that form an in vitro hemopoietic microenvironment. Myeloid bone marrow cultures (MBMC) are optimal for myelopoiesis, while lymphoid bone marrow cultures (LBMC) only support B lymphopoiesis. The experiments reported here have made a comparative analysis of the two cultures to determine whether the stromal cells that establish in vitro are restricted to the support of myelopoiesis or lymphopoiesis, respectively, and to examine how the different culture conditions affect stromal cell physiology. In order to facilitate this analysis, purified populations of MBMC and LBMC stroma were prepared by treating the LTBMC with the antibiotic mycophenolic acid; this results in the elimination of hemopoietic cells while retaining purified populations of functional stroma. Stromal cell cultures prepared and maintained under MBMC conditions secreted myeloid growth factors that stimulated the growth of granulocyte-macrophage colonies, while no such activity was detected from purified LBMC stromal cultures. However, this was not due to the inability of LBMC stroma to mediate this function. Transfer of LBMC stromal cultures to MBMC conditions resulted in an induction of myeloid growth factor secretion. When seeded under these conditions with stromal cell- depleted populations of hemopoietic cells, obtained by passing marrow through nylon wool columns, the LBMC stromal cells could support long- term myelopoiesis. Conversely, transfer of MBMC stroma to LBMC conditions resulted in a cessation of myeloid growth factor secretion; on seeding these cultures with nylon wool-passed marrow, B lymphopoiesis, but not myelopoiesis, initiated. These findings indicate that the stroma in the different LTBMC are not restricted in their hemopoietic support capacity but are sensitive to culture conditions in a manner that may affect the type of microenvironment formed.

scholarly journals A macrophage and theca cell-enriched stromal cell population influences growth and survival of immature murine follicles in vitro

Reproduction ◽  
2011 ◽  
Vol 141 (6) ◽  
pp. 809-820 ◽  
Author(s):  
Candace M Tingen ◽  
Sarah E Kiesewetter ◽  
Jennifer Jozefik ◽  
Cristina Thomas ◽  
David Tagler ◽  
...  
Keyword(s):  
Stromal Cells ◽  
Culture System ◽  
Stromal Cell ◽  
Ovarian Follicle ◽  
3D Culture ◽  
Culture Conditions ◽  
Growth And Survival ◽  
3D Culture System

Innovations in in vitro ovarian follicle culture have revolutionized the field of fertility preservation, but the successful culturing of isolated primary and small secondary follicles remains difficult. Herein, we describe a revised 3D culture system that uses a feeder layer of ovarian stromal cells to support early follicle development. This culture system allows significantly improved primary and early secondary follicle growth and survival. The stromal cells, consisting mostly of thecal cells and ovarian macrophages, recapitulate the in vivo conditions of these small follicles and increase the production of androgens and cytokines missing from stromal cell-free culture conditions. These results demonstrate that small follicles have a stage-specific reliance on the ovarian environment, and that growth and survival can be improved in vitro through a milieu created by pre-pubertal ovarian stromal cell co-culture.

scholarly journals Stromal cells in myeloid and lymphoid long-term bone marrow cultures can support multiple hemopoietic lineages and modulate their production of hemopoietic growth factors

Blood ◽  
1986 ◽  
Vol 68 (6) ◽  
pp. 1348-1354 ◽  
Author(s):  
A Johnson ◽  
K Dorshkind
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Culture Conditions ◽  
B Lymphopoiesis ◽  
Bone Marrow Cultures ◽  
Factor Secretion ◽  
Marrow Cultures

Hemopoiesis in long-term bone marrow cultures (LTBMC) is dependent on adherent stromal cells that form an in vitro hemopoietic microenvironment. Myeloid bone marrow cultures (MBMC) are optimal for myelopoiesis, while lymphoid bone marrow cultures (LBMC) only support B lymphopoiesis. The experiments reported here have made a comparative analysis of the two cultures to determine whether the stromal cells that establish in vitro are restricted to the support of myelopoiesis or lymphopoiesis, respectively, and to examine how the different culture conditions affect stromal cell physiology. In order to facilitate this analysis, purified populations of MBMC and LBMC stroma were prepared by treating the LTBMC with the antibiotic mycophenolic acid; this results in the elimination of hemopoietic cells while retaining purified populations of functional stroma. Stromal cell cultures prepared and maintained under MBMC conditions secreted myeloid growth factors that stimulated the growth of granulocyte-macrophage colonies, while no such activity was detected from purified LBMC stromal cultures. However, this was not due to the inability of LBMC stroma to mediate this function. Transfer of LBMC stromal cultures to MBMC conditions resulted in an induction of myeloid growth factor secretion. When seeded under these conditions with stromal cell- depleted populations of hemopoietic cells, obtained by passing marrow through nylon wool columns, the LBMC stromal cells could support long- term myelopoiesis. Conversely, transfer of MBMC stroma to LBMC conditions resulted in a cessation of myeloid growth factor secretion; on seeding these cultures with nylon wool-passed marrow, B lymphopoiesis, but not myelopoiesis, initiated. These findings indicate that the stroma in the different LTBMC are not restricted in their hemopoietic support capacity but are sensitive to culture conditions in a manner that may affect the type of microenvironment formed.

scholarly journals Emerging data supporting stromal cell therapeutic potential in cancer: reprogramming stromal cells of the tumor microenvironment for anti-cancer effects

2020 ◽  
Vol 17 (4) ◽  
pp. 828-841
Author(s):  
Armel H. Nwabo Kamdje ◽  
Paul F. Seke Etet ◽  
Richard Tagne Simo ◽  
Lorella Vecchio ◽  
Kiven Erique Lukong ◽  
...  
Keyword(s):  
Tumor Microenvironment ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Therapeutic Potential ◽  
Anti Cancer

scholarly journals B lymphocyte precursors and myeloid progenitors survive in diffusion chamber cultures but B cell differentiation requires close association with stromal cells

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1418-1424 ◽  
Author(s):  
PC Kierney ◽  
K Dorshkind
Keyword(s):  
B Lymphocytes ◽  
Stromal Cells ◽  
Direct Contact ◽  
Stromal Cell ◽  
B Lymphocyte ◽  
Cell Layer ◽  
Close Association ◽  
Culture Conditions ◽  
B Lymphopoiesis ◽  
Hemopoietic Cells

Abstract The aim of this study was to investigate the relative contribution of direct contact with stromal cells v stromal cell-derived soluble mediators to the differentiation of B lymphocytes and cells from other hematopoietic lineages. This was investigated by making a comparison between hemopoietic cells grown in direct contact with stroma to those in diffusion chambers (DCs) placed over purified populations of stroma. The source of stromal cells was adherent layers from myeloid or lymphoid long-term bone marrow cultures that had been treated with mycophenolic acid, an antibiotic that depletes hemopoietic cells from the cultures but retains a functional stroma. The cells seeded into the chambers were fresh marrow cells that had been passed through two consecutive nylon wool columns to deplete cell populations capable of forming an adherent cell layer in vitro. DCs were placed in wells in which the adherent stroma, growing under myeloid or lymphoid conditions, was present. The results indicate that progenitors of granulocytes and macrophages survived and differentiated in DCs under myeloid culture conditions, as the number of cells and absolute number of CFU-GM increased over that present in the reseed population. These levels, however, were markedly less than in parallel cultures in which the cells were seeded directly onto stroma. Hematopoiesis in DCs placed over hemopoietically active stroma was not optimal, suggesting that factors were used by those hemopoietic cells closest to the stroma. A B lymphocyte precursor survived in DCs under myeloid but not lymphoid conditions, and its differentiation into B lymphocytes was dependent on close association with stromal cells; B lymphopoiesis initiated when cells from DCs grown under myeloid conditions were harvested from the chambers and seeded directly onto stroma initiated and maintained under lymphoid bone marrow culture conditions. B lymphopoiesis did not initiate if the DC from the myeloid conditions was left intact and placed directly over a lymphoid stromal cell layer in lymphoid conditions.

scholarly journals Dissecting the hematopoietic microenvironment. IX. Further characterization of murine bone marrow stromal cells

Blood ◽  
1993 ◽  
Vol 81 (5) ◽  
pp. 1205-1213
Author(s):  
PE Penn ◽  
DZ Jiang ◽  
RG Fei ◽  
E Sitnicka ◽  
NS Wolf
Keyword(s):  
Bone Marrow ◽  
Smooth Muscle ◽  
Stromal Cell ◽  
Culture Conditions ◽  
Antigenic Determinants ◽  
Collagen Type Iv ◽  
Positive Staining ◽  
Type I ◽  
Stromal Fibroblasts ◽  
Murine Bone Marrow

We have previously shown the adherent nontransformed, nonimmortalized murine bone marrow stromal cell (BMSC) population to consist of phagocytic macrophage and endothelial-like cells and nonphagocytic fibroblasts. Both colonial and near confluent growth of each cell type was obtained following magnetic bead separation, subsequent passaging, and sustained culture with fetal bovine serum and cytokines. Monoclonal antibody staining of antigenic determinants was used to characterize the phenotype of the stromal cell population in primary platings of murine colony-forming unit fibroblast and long-term bone marrow cultures. The antibodies MECA-99, MECA-32, and MJ7–18, raised against murine vascular endothelial antigenic determinants, and von Willebrand's factor all stained selectively for the rounded endothelial- like cells. Endothelial-like cells as well as macrophages expressed the myeloid surface antigens F4/80, 7/4, and Mac-1 under our culture conditions. The cytoskeleton of the stromal fibroblasts in culture was shown to express smooth muscle-specific actin isoforms, as evidenced by positive staining of stress fibers for alpha smooth muscle-1, CGA-7 (alpha/gamma isoforms), and HHF-35 (recognizes all muscle-specific actins). Under culture conditions, stromal fibroblasts were also found to be positive for a polyclonal smooth muscle myosin. It was found that these fibroblasts stained for collagens type I, III, and IV in our cultures. Although collagen type IV is considered a by-product of endothelial cells, endothelial-like cells in our cultures did not stain for any of the collagen types. We propose a classification listing for murine BMSCs as macrophages, endotheliallike cells, and fibroblasts that display smooth muscle-like characteristics in culture.

scholarly journals B lymphocyte precursors and myeloid progenitors survive in diffusion chamber cultures but B cell differentiation requires close association with stromal cells

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1418-1424
Author(s):  
PC Kierney ◽  
K Dorshkind
Keyword(s):  
B Lymphocytes ◽  
Stromal Cells ◽  
Direct Contact ◽  
Stromal Cell ◽  
B Lymphocyte ◽  
Cell Layer ◽  
Close Association ◽  
Culture Conditions ◽  
B Lymphopoiesis ◽  
Hemopoietic Cells

The aim of this study was to investigate the relative contribution of direct contact with stromal cells v stromal cell-derived soluble mediators to the differentiation of B lymphocytes and cells from other hematopoietic lineages. This was investigated by making a comparison between hemopoietic cells grown in direct contact with stroma to those in diffusion chambers (DCs) placed over purified populations of stroma. The source of stromal cells was adherent layers from myeloid or lymphoid long-term bone marrow cultures that had been treated with mycophenolic acid, an antibiotic that depletes hemopoietic cells from the cultures but retains a functional stroma. The cells seeded into the chambers were fresh marrow cells that had been passed through two consecutive nylon wool columns to deplete cell populations capable of forming an adherent cell layer in vitro. DCs were placed in wells in which the adherent stroma, growing under myeloid or lymphoid conditions, was present. The results indicate that progenitors of granulocytes and macrophages survived and differentiated in DCs under myeloid culture conditions, as the number of cells and absolute number of CFU-GM increased over that present in the reseed population. These levels, however, were markedly less than in parallel cultures in which the cells were seeded directly onto stroma. Hematopoiesis in DCs placed over hemopoietically active stroma was not optimal, suggesting that factors were used by those hemopoietic cells closest to the stroma. A B lymphocyte precursor survived in DCs under myeloid but not lymphoid conditions, and its differentiation into B lymphocytes was dependent on close association with stromal cells; B lymphopoiesis initiated when cells from DCs grown under myeloid conditions were harvested from the chambers and seeded directly onto stroma initiated and maintained under lymphoid bone marrow culture conditions. B lymphopoiesis did not initiate if the DC from the myeloid conditions was left intact and placed directly over a lymphoid stromal cell layer in lymphoid conditions.

scholarly journals Contact- and growth factor-dependent survival in a canine marrow- derived stromal cell line

Blood ◽  
1995 ◽  
Vol 85 (9) ◽  
pp. 2414-2421 ◽  
Author(s):  
R Huss ◽  
CA Hoy ◽  
HJ Deeg
Keyword(s):  
Growth Factor ◽  
Cell Line ◽  
Cell Density ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Culture Conditions ◽  
Autocrine Factors ◽  
Stromal Cell Line ◽  
Lower Cell Density ◽  
Local Cell

Cell-cell interactions and the presence of growth factors such as stem cell factor (SCF; or c-kit ligand) or interleukin-6 (IL-6) are involved in the proliferation and differentiation of the canine marrow-derived stromal cell line DO64. In the presence of SCF, stromal cells are induced to differentiate, but not to proliferate. In contrast, in the presence of IL-6, stromal cells are induced to proliferate rather than to differentiate in culture. Both SCF and IL-6 are produced by the stromal cells themselves and, thus, act as autocrine factors. In addition, DO64 cells also interact physically with each other in culture when grown under optimal culture conditions (70% to 90% cell confluence and in the presence of serum), thereby supporting proliferation and maintaining viability. Under conditions of lower cell density or low serum or growth factor concentrations in culture, DO64 cells tend to aggregate and form clusters. This increase in local cell concentration is associated with preservation of viability, presumably because of the accumulation of autocrine factors. If no signal, neither intercellular nor soluble, is provided, and DO64 cells are not able to reach a critical cell density or to produce sufficient factors in an autocrine fashion, the cells cease to proliferate and eventually die.

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