Neurogenic and Neuroprotective Potential of Stem/Stromal Cells Derived from Adipose Tissue

Currently, the number of stem-cell based experimental therapies in neurological injuries and neurodegenerative disorders has been massively increasing. Despite the fact that we still have not obtained strong evidence of mesenchymal stem/stromal cells’ neurogenic effectiveness in vivo, research may need to focus on more appropriate sources that result in more therapeutically promising cell populations. In this study, we used dedifferentiated fat cells (DFAT) that are proven to demonstrate more pluripotent abilities in comparison with standard adipose stromal cells (ASCs). We used the ceiling culture method to establish DFAT cells and to optimize culture conditions with the use of a physioxic environment (5% O2). We also performed neural differentiation tests and assessed the neurogenic and neuroprotective capability of both DFAT cells and ASCs. Our results show that DFAT cells may have a better ability to differentiate into oligodendrocytes, astrocytes, and neuron-like cells, both in culture supplemented with N21 and in co-culture with oxygen–glucose-deprived (OGD) hippocampal organotypic slice culture (OHC) in comparison with ASCs. Results also show that DFAT cells have a different secretory profile than ASCs after contact with injured tissue. In conclusion, DFAT cells constitute a distinct subpopulation and may be an alternative source in cell therapy for the treatment of nervous system disorders.

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

Reproduction ◽

10.1530/rep-10-0483 ◽

2011 ◽

Vol 141 (6) ◽

pp. 809-820 ◽

Cited By ~ 50

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.

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Modulating the Substrate Stiffness to Manipulate Differentiation of Resident Liver Stem Cells and to Improve the Differentiation State of Hepatocytes

Stem Cells International ◽

10.1155/2016/5481493 ◽

2016 ◽

Vol 2016 ◽

pp. 1-12 ◽

Cited By ~ 38

Author(s):

Angela Maria Cozzolino ◽

Valeria Noce ◽

Cecilia Battistelli ◽

Alessandra Marchetti ◽

Germana Grassi ◽

...

Keyword(s):

Stem Cells ◽

Culture Method ◽

Cell Types ◽

Culture Conditions ◽

Precursor Cells ◽

Substrate Stiffness ◽

Growth And Survival ◽

Liver Stem Cells

In many cell types, several cellular processes, such as differentiation of stem/precursor cells, maintenance of differentiated phenotype, motility, adhesion, growth, and survival, strictly depend on the stiffness of extracellular matrix that,in vivo, characterizes their correspondent organ and tissue. In the liver, the stromal rigidity is essential to obtain the correct organ physiology whereas any alteration causes liver cell dysfunctions. The rigidity of the substrate is an element no longer negligible for the cultivation of several cell types, so that many data so far obtained, where cells have been cultured on plastic, could be revised. Regarding liver cells, standard culture conditions lead to the dedifferentiation of primary hepatocytes, transdifferentiation of stellate cells into myofibroblasts, and loss of fenestration of sinusoidal endothelium. Furthermore, standard cultivation of liver stem/precursor cells impedes an efficient execution of the epithelial/hepatocyte differentiation program, leading to the expansion of a cell population expressing only partially liver functions and products. Overcoming these limitations is mandatory for any approach of liver tissue engineering. Here we propose cell lines asin vitromodels of liver stem cells and hepatocytes and an innovative culture method that takes into account the substrate stiffness to obtain, respectively, a rapid and efficient differentiation process and the maintenance of the fully differentiated phenotype.

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Synaptic Organization of an organotypic slice culture of the mammalian retina

Visual Neuroscience ◽

10.1017/s0952523800008634 ◽

1996 ◽

Vol 13 (4) ◽

pp. 759-771 ◽

Cited By ~ 12

Author(s):

Marco Sassoè-Pognetto ◽

Andreas Feigenspan ◽

Joachim Bormann ◽

Heinz Wässle

Keyword(s):

Hot Spots ◽

Plexiform Layer ◽

Slice Culture ◽

Inner Plexiform Layer ◽

Synaptic Organization ◽

Glycine Receptors ◽

Ribbon Synapses ◽

Organotypic Slice Culture ◽

Transmitter Receptors

AbstractVertical Slices of postnatal day 6 (P6) rat retina were cut and cultured using the roller-tube technique. The organotypic differentiation during a culture period of up to 30 days has been described in a previous study (Feigenspan et al., 1993a). Here we concentrated on the synaptic organization in the retinal slice culture. Electron microscopy revealed the presence of ribbon synapses in the outer plexiform layer and conventional and ribbon syanpses in the inner plexiform layer. Immunofluroscence with antibodies that recognize specific subunits of GABAA or glycine receptors revealed a punctuate distribution of the receptors. They were aggregated in “hot spots” that correspond to a concentration of receptors at postsynaptic sites. Different isoforms of GABAA and glycine receptors occured in the slice cultures. The experiments show that there is a differentiation of synapses and a diversity of transmitter receptors in the slice cultures that is comparable to the in vivo retina.

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3D Hepatic Organoid-Based Advancements in LIVER Tissue Engineering

Bioengineering ◽

10.3390/bioengineering8110185 ◽

2021 ◽

Vol 8 (11) ◽

pp. 185

Author(s):

Amit Panwar ◽

Prativa Das ◽

Lay Poh Tan

Keyword(s):

Tissue Engineering ◽

Self Assembly ◽

Liver Tissue ◽

3D Culture ◽

Culture Method ◽

Culture Conditions ◽

Phenotypic Properties ◽

Liver Tissue Engineering

Liver-associated diseases and tissue engineering approaches based on in vitro culture of functional Primary human hepatocytes (PHH) had been restricted by the rapid de-differentiation in 2D culture conditions which restricted their usability. It was proven that cells growing in 3D format can better mimic the in vivo microenvironment, and thus help in maintaining metabolic activity, phenotypic properties, and longevity of the in vitro cultures. Again, the culture method and type of cell population are also recognized as important parameters for functional maintenance of primary hepatocytes. Hepatic organoids formed by self-assembly of hepatic cells are microtissues, and were able to show long-term in vitro maintenance of hepato-specific characteristics. Thus, hepatic organoids were recognized as an effective tool for screening potential cures and modeling liver diseases effectively. The current review summarizes the importance of 3D hepatic organoid culture over other conventional 2D and 3D culture models and its applicability in Liver tissue engineering.

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Culture Conditions for Generating Human Bone Marrow Stromal Cells Influence Cell Immunophenotype and In Vivo Biodistribution in Immune Deficient Mice.

Blood ◽

10.1182/blood.v104.11.2334.2334 ◽

2004 ◽

Vol 104 (11) ◽

pp. 2334-2334

Author(s):

Joanna Vergidis ◽

Garnet Suck ◽

Xing-Hua Wang ◽

Peter W. Zandstra ◽

Armand Keating

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Stromal Cells ◽

Bone Marrow Stromal Cells ◽

Culture Conditions ◽

Femoral Bone ◽

Tibial Bone ◽

Low Levels ◽

Femoral Bone Marrow

Abstract Bone marrow stromal cells (MSCs) show promise for cell and gene therapies but their utility is currently limited by low levels of engraftment. A better understanding of factors underlying cell trafficking in vivo is likely to lead to enhanced MSC engraftment and tissue targeting. To address this issue, we studied the immunophenotype and transplantation potential of MSCs derived from two different culture systems, serially passaged adherent layers from standard long-term bone marrow cultures (LTBMC) and a novel stirred suspension bioreactor culture (SSBC) supplemented with IL-3 and SCF. MSCs were characterized as CD45 (−), VCAM-1 (+), CD44 (+), SH-3 (+), and CD49e (+) by flow cytometry. At earlier time points, the SSBC system generated 1.8x more MSCs compared with the LTBMC system when analyzing the CD45-negative fraction by flow cytometry. By the end of culture, however, the LTBMC system generated 7.3x more cells. Interestingly, 66% of the CD45 (−) cells from the bioreactor system were negative for both HLA Class I and II antigens. Furthermore, flow cytometry revealed that the bioreactor-derived cells expressed very low levels of the cell adhesion markers, VCAM-1 and CD44. These data suggest that the SSBC system generates cells that may have greater migratory freedom in vivo. Biodistribution patterns of the human MSCs derived from the two sources were examined in 11 completely unconditioned six-week old SCID mice. Six weeks after intravenous infusion of MSCs, femoral and tibial bone marrow, lung, liver, bone, spleen, brain, heart, and blood were analyzed for donor human cell engraftment by PCR and fluorescence in situ hybridization (FISH) against the murine background. Engraftment was determined by PCR with primers from the human alpha-satellite region (chromosome 17) that amplify a specific 850 bp fragment. One tibial bone marrow sample, two femoral bone marrow samples, one bone sample, and four lung samples from nine mice receiving LTBMC MSC were positive by PCR. In contrast, all of the same tissues from the mouse receiving SSBC MSC were negative for human donor cells, including the lung, with the intriguing exception of a positive PCR signal in heart tissue. The presence of donor MSC was confirmed in PCR+ specimens by FISH using a human Cy3- and a murine FITC-pan-centromeric probe. The frequencies of donor LTBMC MSC in the tibial bone marrow, femoral bone marrow, bone, and lung were 0.79%, 1.60%, 0.58%, and 1.99%, respectively. Donor bioreactor-derived MSCs accounted for 0.41% of the cardiomyocytes from one recipient, despite the absence of cardiac injury. Compared with LTBMC MSCs, SSBC-derived cells appear to display an unusual biodistribution pattern which may be attributable to the altered immunophenotype. The present study underscores the importance of culture conditions in influencing the immunophenotype of MSCs and holds promise for developing targeted cell therapy.

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Microfluidics and multielectrode array-compatible organotypic slice culture method

Journal of Neuroscience Methods ◽

10.1016/j.jneumeth.2008.11.016 ◽

2009 ◽

Vol 178 (1) ◽

pp. 59-64 ◽

Cited By ~ 46

Author(s):

Yevgeny Berdichevsky ◽

Helen Sabolek ◽

John B. Levine ◽

Kevin J. Staley ◽

Martin L. Yarmush

Keyword(s):

Culture Method ◽

Multielectrode Array ◽

Slice Culture ◽

Organotypic Slice Culture

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MICROPROPOGATION AND ANALYSIS OF PHYTOCHEMICAL PROFILE OF TISSUE CULTURE GROWN PLANTAGO OVATA FORSK.

Asian Journal of Pharmaceutical and Clinical Research ◽

10.22159/ajpcr.2017.v10i4.16532 ◽

2017 ◽

Vol 10 (4) ◽

pp. 202 ◽

Cited By ~ 3

Author(s):

Mamta Sharma ◽

Amita Kumari ◽

Eshita Mahant

Keyword(s):

Tissue Culture ◽

Ecological Factors ◽

Culture Method ◽

Culture Conditions ◽

Culture Technique ◽

Plantago Ovata ◽

Tissue Culture Technique ◽

Grown Plant

Objectives : Plantago ovata is an important medicinal plant of Himalayan region greatly used in herbal dugs manufacturing. The plant is multipurpose and strictly present in the Himalaya. Plantago has many medicinal properties such as antioxidant, anti-inflammatory and hematopoiesis effects and protects the liver and is used for the treatment of cancer. The plant being medicinal possesses complex phytochemicals. The investigation of various Plantago organ (leaves, stem etc) revealed their high potential to produce a wide array of bioactive secondary metabolites. In present study the a new method of micropropagation through tissue culture was developed for Plantago so as to meet the future demand of plant. Futher a morphological and physiochemical comparison of tissue culture grown plant was done with in vivo grown plants.Methods: Plantago ovata was grown in -vitro through tissue culture technique using MS media and in-vivo in the nursery area of Shoolini University. In vitro culture of Plantago ovata forsk. were managed to restrict the ecological factors and to control the culture conditions. Experimental culture parameter including germination and phytochemical constituents of Plantago ovata in vivo and in vitro conditions were observed.Results: The result revealed changes in the concentration of phytochemical constituent’s in tissue culture grown Plantago. Phytochemicals constituents (carbohydrate, tannin, chlorophyll, saponin) was reduced in tissue culture grown plant where as some phytochemicals (phenol, alkaloid, flavanoid, protein, phytosterol) increased in tissue culture grown plant than in vivo plant. A reduction in morphological trait was found in tissue cultured plant.Conclusion: The developed tissue culture method for the micropropagation of Plantago ovata can be used as milestone to meet the industrial need in near future.Keywords: Plantago, Tissue Culture Technique, germination, phytochemicals.

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Exofucosylation of Adipose Mesenchymal Stromal Cells Alters Their Secretome Profile

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2020.584074 ◽

2020 ◽

Vol 8 ◽

Author(s):

David García-Bernal ◽

Mariano García-Arranz ◽

Ana I. García-Guillén ◽

Ana M. García-Hernández ◽

Miguel Blanquer ◽

...

Keyword(s):

Mesenchymal Stromal Cells ◽

Stromal Cells ◽

Culture Conditions ◽

Peripheral Tissues ◽

Standard Culture ◽

Anti Inflammatory ◽

Selectin Binding ◽

The Impact

Mesenchymal stromal cells (MSCs) constitute the cell type more frequently used in many regenerative medicine approaches due to their exclusive immunomodulatory properties, and they have been reported to mediate profound immunomodulatory effects in vivo. Nevertheless, MSCs do not express essential adhesion molecules actively involved in cell migration, a phenotypic feature that hampers their ability to home inflamed tissues following intravenous administration. In this study, we investigated whether modification by fucosylation of murine AdMSCs (mAdMSCs) creates Hematopoietic Cell E-/L-selectin Ligand, the E-selectin-binding CD44 glycoform. This cell surface glycan modification of CD44 has previously shown in preclinical studies to favor trafficking of mAdMSCs to inflamed or injured peripheral tissues. We analyzed the impact that exofucosylation could have in other innate phenotypic and functional properties of MSCs. Compared to unmodified counterparts, fucosylated mAdMSCs demonstrated higher in vitro migration, an altered secretome pattern, including increased expression and secretion of anti-inflammatory molecules, and a higher capacity to inhibit mitogen-stimulated splenocyte proliferation under standard culture conditions. Together, these findings indicate that exofucosylation could represent a suitable cell engineering strategy, not only to facilitate the in vivo MSC colonization of damaged tissues after systemic administration, but also to convert MSCs in a more potent immunomodulatory/anti-inflammatory cell therapy-based product for the treatment of a variety of autoimmune, inflammatory, and degenerative diseases.

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POSCAbilities: The Application of the Prion Organotypic Slice Culture Assay to Neurodegenerative Disease Research

Biomolecules ◽

10.3390/biom10071079 ◽

2020 ◽

Vol 10 (7) ◽

pp. 1079

Author(s):

Hailey Pineau ◽

Valerie Sim

Keyword(s):

Neurodegenerative Diseases ◽

Prion Diseases ◽

In Vitro Models ◽

Slice Culture ◽

Advantages And Disadvantages ◽

Organotypic Slice Culture ◽

Drug Treatments

Prion diseases are fatal, transmissible neurodegenerative disorders whose pathogenesis is driven by the misfolding, self-templating and cell-to-cell spread of the prion protein. Other neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis and Huntington’s disease, share some of these prion-like features, with different aggregation-prone proteins. Consequently, researchers have begun to apply prion-specific techniques, like the prion organotypic slice culture assay (POSCA), to these disorders. In this review we explore the ways in which the prion phenomenon has been used in organotypic cultures to study neurodegenerative diseases from the perspective of protein aggregation and spreading, strain propagation, the role of glia in pathogenesis, and efficacy of drug treatments. We also present an overview of the advantages and disadvantages of this culture system compared to in vivo and in vitro models and provide suggestions for new directions.

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Clinical-grade production of human mesenchymal stromal cells: occurrence of aneuploidy without transformation

Blood ◽

10.1182/blood-2009-05-219907 ◽

2010 ◽

Vol 115 (8) ◽

pp. 1549-1553 ◽

Cited By ~ 317

Author(s):

Karin Tarte ◽

Julien Gaillard ◽

Jean-Jacques Lataillade ◽

Loic Fouillard ◽

Martine Becker ◽

...

Keyword(s):

Mesenchymal Stromal Cells ◽

Stromal Cells ◽

Calf Serum ◽

Human Leukocyte ◽

Culture Conditions ◽

Human Mscs ◽

Clinical Grade ◽

Human Mesenchymal Stromal Cells

Abstract Clinical-grade human mesenchymal stromal cells (MSCs) have been expanded in vitro for tissue engineering or immunoregulatory purposes without standardized culture conditions or release criteria. Although human MSCs show poor susceptibility for oncogenic transformation, 2 recent studies described their capacity to accumulate chromosomal instability and to give rise to carcinoma in immunocompromised mice after long-term culture. We thus investigated the immunologic and genetic features of MSCs expanded with fetal calf serum and fibroblast growth factor or with platelet lysate in 4 cell-therapy facilities during 2 multicenter clinical trials. Cultured MSCs showed a moderate expression of human leukocyte antigen-DR without alteration of their low immunogenicity or their immunomodulatory capacity. Moreover, some transient and donor-dependent recurring aneuploidy was detected in vitro, independently of the culture process. However, MSCs with or without chromosomal alterations showed progressive growth arrest and entered senescence without evidence of transformation either in vitro or in vivo.

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