A Simplified 3D Culture System for Ovarian Follicles Utilizing a Solid Matrigel Drop to Create an In Vivo-Like Ovarian Microenvironment

2019 ◽  
Vol 9 (4) ◽  
pp. 558-561
Author(s):  
Jung Ah Yoon ◽  
Kyung-Ah Lee ◽  
Jung Kyu Choi
Keyword(s):  
Culture System ◽  
3D Culture ◽  
Ovarian Follicles ◽  
3D Culture System

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.

Implications of adipose-derived stromal cells in a 3D culture system for osteogenic differentiation: an in vitro and in vivo investigation

2013 ◽  
Vol 13 (1) ◽  
pp. 32-43 ◽  
Author(s):  
Francis H. Shen ◽  
Brian C. Werner ◽  
Haixiang Liang ◽  
Hulan Shang ◽  
Ning Yang ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Stromal Cells ◽  
Culture System ◽  
3D Culture ◽  
Adipose Derived Stromal Cells ◽  
3D Culture System

scholarly journals Lin28A Promotes the Amplification and Cancer Stemness of Lung Cancer Cells Via Activating MAPK Pathway Dependent on Let-7c Functions

2021 ◽  
Author(s):  
Rui Zhang ◽  
Pengpeng Liu ◽  
Xiao Zhang ◽  
Yingnan Ye ◽  
Jinpu Yu
Keyword(s):  
Lung Cancer ◽  
Stem Cells ◽  
Cancer Stem Cells ◽  
Culture System ◽  
3D Culture ◽  
Mapk Signaling ◽  
Cancer Stemness ◽  
Over Expression ◽  
3D Culture System

Abstract Background: Metastasis and relapse of lung cancer are the main cause of disease-related deaths. It’s reported that tumor metastasis and relapse originated from cancer stem cells (CSCs) which possess more potential in proliferation and invasion. In our previous studies, we established a conditional BME-based three-dimensional culture (3D culture) system to mimic the growth environment in vivo and further amplified lung cancer stem cells (LCSCs) in our system. However, the molecular mechanisms of the amplification and development of LCSCs in our 3D culture system are still not very clear. Methods: We tested the expression of Lin28 and let7 by western blot and qPCR, and constructed A549 cells either knockdown of Lin28 or overexpression of let7, followed by investigating the expression of stemness markers by flow cytometry and qPCR, and stem cell like phenotypes including cell proliferation, colony formation, mammosphere culture, cell apoptosis, migration, invasion and drug resistance in vitro, as well as tumorigenicity in vivo. Results: Here we observed Lin28A/let-7c was dysregulated in LCSCs both from the 3D culture system and from lung cancer tissues. Further, the abnormal expression of Lin28A/let-7c was correlated with poor survival outcomes. We found over-expression let-7c inhibited the maintenance of LCSC properties, while the results for knockdown of Lin28A showed Lin28A was critical for the enrichment and amplification of LCSCs via MAPK signaling pathway. Importantly, we found that either knockdown of Lin28A or over-expression of let-7c inhibited carcinogenesis and disrupted LCSC expansion in vivo. Conclusions: Our study uncovered the functions and mechanisms of the "Lin28A/let-7c/MAPK" signaling pathway in promoting the amplification and cancer stemness of LCSCs, which might be a potential therapeutic target for lung cancer therapy by reducing and even eliminating LCSCs in the future.

scholarly journals Dual-matrix 3D culture system as a biomimetic model of epithelial tissues

2019 ◽  
Author(s):  
Diana Bogorodskaya ◽  
Joshua S. McLane ◽  
Lee A. Ligon
Keyword(s):  
Cell Biology ◽  
Culture System ◽  
Drug Testing ◽  
Disease Modeling ◽  
Wide Spectrum ◽  
3D Culture ◽  
Cell Types ◽  
3D Cultures ◽  
3D Culture System

ABSTRACTRecent years have seen an unprecedented rise in the use of 3D culture systems, both in fundamental research and in more translational settings such as drug testing and disease modeling. However, 3D cultures often remain underused by cell biology labs, both due to technical difficulties in system setup and inherent drawbacks of many of the common systems. Here we describe an easy to use, inexpensive and rapidly assembled 3D culture system, suitable for generation of both normal polarized epithelial cysts and in-situ tumor spheroids. This system allows for exploration of many questions of normal and cancer cell biology, including morphogenesis, epithelial polarization, cell motility, intra- and intercellular communication, invasion, metastasis, and tumor-stoma interaction. The 3D cultures are made up of a stiffness tunable, dual-matrix model that can incorporate co-culture of multiple cell types. The model allows for increased physiological relevance by mimicking the organization, ligand composition and stiffness presentin-vivo. The setup allows for a wide spectrum of manipulation, including removing cells from the system for DNA/protein expression, transfection and high-resolution imaging of live or fixed cells.

Implications of Adipose-Derived Stromal Cells in a 3D Culture System for Osteogenic Differentiation: An in Vitro and in Vivo Investigation

2012 ◽  
Vol 12 (9) ◽  
pp. S26-S27
Author(s):  
Francis H. Shen ◽  
Brian C. Werner ◽  
Haixiang Liang ◽  
Hulan Shang ◽  
Ning Yang ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Stromal Cells ◽  
Culture System ◽  
3D Culture ◽  
Adipose Derived Stromal Cells ◽  
3D Culture System

O-101 Neospermatogenesis benefits from a three-dimensional culture system

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
S Lawrence ◽  
M Haddad ◽  
Z Rosenwaks ◽  
G D Palermo
Keyword(s):  
Conventional Method ◽  
Seminiferous Tubule ◽  
Culture System ◽  
Germ Line ◽  
Three Dimensional ◽  
3D Culture ◽  
Cell Stage ◽  
3D Culture System ◽  
Germline Differentiation

Abstract Study question Does a three-dimensional (3D) culture system increase the efficiency of male germline differentiation of mouse embryonic stem cells (mESC) over a bidimensional method? Summary answer Our 3D culture system based on direct spherification proves superior to the standard bidimensional plating in promoting neogametogenesis of mESC into post-meiotic male germ cells. What is known already Two-dimensional monolayer cell cultures are common in stem cell research. However, this method does not replicate a physiological 3D spatial relationship and may provide an inaccurate replication of in vivo environments. A 3D spherical structure allows us to mimic the seminiferous tubule, the site of in vivo spermatogenesis. By using spheroids as a scaffold to replicate cell culture systems, we can study spermatogenesis in a controlled setting. Direct spherification, a technique commonly used in molecular gastronomy, provides an opportunity to create spheroids that mimic in vivo events that materialize in the lab Study design, size, duration mESCs were initially cultured on a 6-well plate coated with fibroblasts and inserted into sodium alginate spheres. To coax differentiation, spheres (3 to 6 mm in diameter) were plunged directly into differentiation medium (DM) while the control mESC in 6-well dishes were layered with it. Cells obtained from both culture systems were tested by biomarkers for different germ cell stages Participants/materials, setting, methods Bidimensional mESC at 80% confluence were differentiated either on a plate or spherified for a 3D culture. Both systems underwent the same timeline of exposure to EpiLC medium with Activin A, bFGF and KSR for 3 days and PGCLC medium with BMP4, LIF, SCF and EGF for 7 days. Differentiated cells were retrieved from each method at day 3 and day 10 to assess for germ line differentiation markers, DAZL, VASA and BOULE Main results and the role of chance Under optic visualization through the sphere wall, cellular aggregation was seen on day 2 of culturing in EpiLC medium while this phenomenon was not observed on bidimensional plating. In the conventional method, cells expressed 7% DAZL (spermatogonium cell stage) and 1% VASA (pre-spermatid cell stage) whereas in direct spherification, cells expressed 20% DAZL (P < 0.001) and 15% VASA positivity (P < 0.0001). To further compare the different methods in later stages of germ-line differentiation, the remaining spheres were cultured in PGCLC medium for 7 days. At day 10, isolated cells were assessed for VASA and DAZL again. In the conventional method, 23% of cells expressed positivity for VASA and 29% DAZL whereas direct spherification achieved a positivity rate of 43% for VASA (P < 0.005) and 45% for DAZL (P < 0.005). This increased expression in both VASA and DAZL signify the increased number of cells undergoing germline differentiation. Additionally, BOULE was assessed for the presence of meiotic cells such as the spermatocyte. The conventional method yielded < 1% BOULE positivity whereas in direct spherification, there was 10% positivity (P < 0.005). Direct spherifcation result shows that differentiation almost doubled in comparison to the conventional method, yielding more post-meiotic cells in the same amount of time Limitations, reasons for caution Despite a higher differentiation rate in direct spherification, these cells would still need to be tested for their fertilization potential. The ability to achieve fertilization, blastocysts and live pups would provide final proof and reliability of this method of neogametogenesis Wider implications of the findings Differentiating ESCs through direct spherification provides an alternative to studying intercellular relationships. This provides an opportunity to study spermatogenesis in more detail by replicating the microenvironment of the seminiferous tubule. Once embryo developmental competence of the de novo gamete is confirmed, this may open a new chapter in human reproduction Trial registration number N/A

Novel three-dimensional long-term bone marrow culture system using polymer particles with grafted epoxy-polymer-chains supports the proliferation and differentiation of hematopoietic stem cells

2011 ◽  
Vol 236 (11) ◽  
pp. 1342-1350 ◽  
Author(s):  
Yukio Hirabayashi ◽  
Yoshihiro Hatta ◽  
Jin Takeuchi ◽  
Isao Tsuboi ◽  
Tomonori Harada ◽  
...  
Keyword(s):  
Stromal Cells ◽  
Culture System ◽  
3D Structure ◽  
Three Dimensional ◽  
Hematopoietic Cells ◽  
3D Culture ◽  
Polymer Chains ◽  
Polymer Particles ◽  
Hematopoietic Stem ◽  
3D Culture System

Hematopoiesis occurs in the bone marrow, where primitive hematopoietic cells proliferate and differentiate in close association with a three-dimensional (3D) hematopoietic microenvironment composed of stromal cells. We examined the hematopoietic supportive ability of stromal cells in a 3D culture system using polymer particles with grafted epoxy polymer chains. Umbilical cord blood-derived CD34+ cells were co-cultivated with MS-5 stromal cells. They formed a 3D structure in the culture dish in the presence of particles, and the total numbers of cells and the numbers of hematopoietic progenitor cells, including colony-forming unit (CFU)-Mix, CFU-granulocyte-macrophage, CFU-megakaryocyte and burst-forming unit-erythroid, were measured every seven days. The hematopoietic supportive activity of the 3D culture containing polymer particles and stromal cells was superior to that of 2D culture, and allowed the expansion and maintenance of hematopoietic progenitor cells for more than 12 weeks. Various types of hematopoietic cells, including granulocytes, macrophages and megakaryocytes at different maturation stages, appeared in the 3D culture, suggesting that the CD34+ cells were able to differentiate into a range of blood cell types. Morphological examination showed that MS-5 stromal cells grew on the surface of the particles and bridged the gaps between them to form a 3D structure. Hematopoietic cells slipped into the 3D layer and proliferated within it, relying on the presence of the MS-5 cells. These results suggest that this 3D culture system using polymer particles reproduced the hematopoietic phenomenon in vitro, and might thus provide a new tool for investigating hematopoietic stem cell–stromal cell interactions.

scholarly journals Mixed enzymatic-explant protocol for isolation of mesenchymal stem cells from Wharton’s jelly and encapsulation in 3D culture system

2012 ◽  
Vol 05 (10) ◽  
pp. 580-586 ◽  
Author(s):  
Saeed Azandeh ◽  
Mahmoud Orazizadeh ◽  
Mahmoud Hashemitabar ◽  
Ali Khodadadi ◽  
Ali Akbar Shayesteh ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Culture System ◽  
3D Culture ◽  
Wharton’S Jelly ◽  
Wharton's Jelly ◽  
3D Culture System

Regeneration of insulin-producing islets from dental pulp stem cells using a 3D culture system

2018 ◽  
Vol 13 (6) ◽  
pp. 673-687 ◽  
Author(s):  
Hiromi Yagi Mendoza ◽  
Tomomi Yokoyama ◽  
Tomoko Tanaka ◽  
Hisataka Ii ◽  
Ken Yaegaki
Keyword(s):  
Stem Cells ◽  
Dental Pulp ◽  
Culture System ◽  
3D Culture ◽  
Dental Pulp Stem Cells ◽  
3D Culture System
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