A nomenclature for intestinal in vitro cultures

Many advances have been reported in the long-term culture of intestinal mucosal cells in recent years. A significant number of publications have described new culture media, cell formations, and growth patterns. Furthermore, it is now possible to study, e.g., the capabilities of isolated stem cells or the interactions between stem cells and mesenchyme. However, at the moment there is significant variation in the way these structures are described and named. A standardized nomenclature would benefit the ability to communicate and compare findings from different laboratories using the different culture systems. To address this issue, members of the NIH Intestinal Stem Cell Consortium herein propose a systematic nomenclature for in vitro cultures of the small and large intestine. We begin by describing the structures that are generated by preparative steps. We then define and describe structures produced in vitro, specifically: enterosphere, enteroid, reconstituted intestinal organoid, induced intestinal organoid, colonosphere, colonoid, and colonic organoid.

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Hypoxic and normoxic in vitro cultures maintain similar numbers of long-term reconstituting hematopoietic stem cells from mouse bone marrow

Experimental Hematology ◽

10.1016/j.exphem.2012.07.005 ◽

2012 ◽

Vol 40 (11) ◽

pp. 879-881 ◽

Cited By ~ 1

Author(s):

Yanjuan Tang ◽

Camilla Halvarsson ◽

Pernilla Eliasson ◽

Jan-Ingvar Jönsson

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

In Vitro Cultures ◽

Mouse Bone Marrow ◽

Hematopoietic Stem

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Strategies to Protect Hematopoietic Stem Cells from Culture-Induced Stress Conditions

Current Stem Cell Research & Therapy ◽

10.2174/1574888x15666200225091339 ◽

2020 ◽

Vol 15 ◽

Author(s):

Fatima Aerts-Kaya

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Ex Vivo ◽

Stress Conditions ◽

Stress Factors ◽

Hematopoietic Stem ◽

Induced Stress

: In contrast to their almost unlimited potential for expansion in vivo and despite years of dedicated research and optimization of expansion protocols, the expansion of Hematopoietic Stem Cells (HSCs) in vitro remains remarkably limited. Increased understanding of the mechanisms that are involved in maintenance, expansion and differentiation of HSCs will enable the development of better protocols for expansion of HSCs. This will allow procurement of HSCs with long-term engraftment potential and a better understanding of the effects of the external influences in and on the hematopoietic niche that may affect HSC function. During collection and culture of HSCs, the cells are exposed to suboptimal conditions that may induce different levels of stress and ultimately affect their self-renewal, differentiation and long-term engraftment potential. Some of these stress factors include normoxia, oxidative stress, extra-physiologic oxygen shock/stress (EPHOSS), endoplasmic reticulum (ER) stress, replicative stress, and stress related to DNA damage. Coping with these stress factors may help reduce the negative effects of cell culture on HSC potential, provide a better understanding of the true impact of certain treatments in the absence of confounding stress factors. This may facilitate the development of better ex vivo expansion protocols of HSCs with long-term engraftment potential without induction of stem cell exhaustion by cellular senescence or loss of cell viability. This review summarizes some of available strategies that may be used to protect HSCs from culture-induced stress conditions.

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Stability of Imprinting and Differentiation Capacity in Naïve Human Cells Induced by Chemical Inhibition of CDK8 and CDK19

Cells ◽

10.3390/cells10040876 ◽

2021 ◽

Vol 10 (4) ◽

pp. 876

Author(s):

Raquel Bernad ◽

Cian J. Lynch ◽

Rocio G. Urdinguio ◽

Camille Stephan-Otto Attolini ◽

Mario F. Fraga ◽

...

Keyword(s):

Stem Cells ◽

Pluripotent Stem Cells ◽

Primordial Germ Cell ◽

Normal Karyotype ◽

Cell Types ◽

Dna Demethylation ◽

Starting State ◽

Teratoma Formation

Pluripotent stem cells can be stabilized in vitro at different developmental states by the use of specific chemicals and soluble factors. The naïve and primed states are the best characterized pluripotency states. Naïve pluripotent stem cells (PSCs) correspond to the early pre-implantation blastocyst and, in mice, constitute the optimal starting state for subsequent developmental applications. However, the stabilization of human naïve PSCs remains challenging because, after short-term culture, most current methods result in karyotypic abnormalities, aberrant DNA methylation patterns, loss of imprinting and severely compromised developmental potency. We have recently developed a novel method to induce and stabilize naïve human PSCs that consists in the simple addition of a chemical inhibitor for the closely related CDK8 and CDK19 kinases (CDK8/19i). Long-term cultured CDK8/19i-naïve human PSCs preserve their normal karyotype and do not show widespread DNA demethylation. Here, we investigate the long-term stability of allele-specific methylation at imprinted loci and the differentiation potency of CDK8/19i-naïve human PSCs. We report that long-term cultured CDK8/19i-naïve human PSCs retain the imprinting profile of their parental primed cells, and imprints are further retained upon differentiation in the context of teratoma formation. We have also tested the capacity of long-term cultured CDK8/19i-naïve human PSCs to differentiate into primordial germ cell (PGC)-like cells (PGCLCs) and trophoblast stem cells (TSCs), two cell types that are accessible from the naïve state. Interestingly, long-term cultured CDK8/19i-naïve human PSCs differentiated into PGCLCs with a similar efficiency to their primed counterparts. Also, long-term cultured CDK8/19i-naïve human PSCs were able to differentiate into TSCs, a transition that was not possible for primed PSCs. We conclude that inhibition of CDK8/19 stabilizes human PSCs in a functional naïve state that preserves imprinting and potency over long-term culture.

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Isolation, Characterization, and Differentiation of Stem Cells From Various Dental Sources: An In Vitro Study

Journal of Advanced Oral Research ◽

10.1177/23202068211010768 ◽

2021 ◽

pp. 232020682110107

Author(s):

Sandeep S. Katti ◽

Kishore Bhat ◽

Chetana Bogar

Keyword(s):

Stem Cells ◽

Growth Factors ◽

Statistical Analysis ◽

Specific Growth ◽

Culture Media ◽

In Vitro Study ◽

Central Research ◽

Cell Lineages ◽

Apical Papilla

Aim: The aim of the current study was to isolate stem cells from various dental sources such as dental pulp, periodontal ligament (PDL), and apical papilla, and to characterize stem cells by staining for the presence/absence of specific surface markers and also to differentiate stem cells into osteogenic, chondrogenic, and adipogenic cell lineages by exposing them to specific growth factors under the ideal conditions. Materials and Methods: A total of 117 samples were included in the study, consisting of 30 pulp, 50 gingival, 35 PDL, and 2 apical papilla samples. The pulp was extirpated and transported to the Central Research Laboratory. Gingival connective tissue was collected from the participants undergoing any crown lengthening procedure or any gingivectomy procedure from the Department of Periodontology. A similar procedure was also followed for apical papilla and PDL. Isolation was done followed by the identification of the cells by immunocytochemistry using different markers. Once the identity of cells was confirmed, these cells were treated with different culture media to attain 70% to 100% confluency. Then the medium was replaced with a conditioning medium containing specific growth factors for differentiation into osteogenic, chondrogenic, and adipogenic cell lineages. Result: In our study, the number of samples collected and processed was 117. The isolation rate of stem cells from the above-collected samples was 70%. Statistical analysis—no statistical analysis was done as there was no variability expected. Conclusion: Our study showed that stem cells could be isolated, differentiated, and characterized from different dental sources.

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Expression Profile of New Marker Genes Involved in Differentiation of Canine Adipose-Derived Stem Cells into Osteoblasts

International Journal of Molecular Sciences ◽

10.3390/ijms22136663 ◽

2021 ◽

Vol 22 (13) ◽

pp. 6663

Author(s):

Maurycy Jankowski ◽

Mariusz Kaczmarek ◽

Grzegorz Wąsiatycz ◽

Claudia Dompe ◽

Paul Mozdziak ◽

...

Keyword(s):

Stem Cells ◽

In Vitro Culture ◽

Osteogenic Differentiation ◽

Molecular Mechanisms ◽

Marker Genes ◽

P Value ◽

Illumina Platform

Next-generation sequencing (RNAseq) analysis of gene expression changes during the long-term in vitro culture and osteogenic differentiation of ASCs remains to be important, as the analysis provides important clues toward employing stem cells as a therapeutic intervention. In this study, the cells were isolated from adipose tissue obtained during routine surgical procedures and subjected to 14-day in vitro culture and differentiation. The mRNA transcript levels were evaluated using the Illumina platform, resulting in the detection of 19,856 gene transcripts. The most differentially expressed genes (fold change >|2|, adjusted p value < 0.05), between day 1, day 14 and differentiated cell cultures were extracted and subjected to bioinformatical analysis based on the R programming language. The results of this study provide molecular insight into the processes that occur during long-term in vitro culture and osteogenic differentiation of ASCs, allowing the re-evaluation of the roles of some genes in MSC progression towards a range of lineages. The results improve the knowledge of the molecular mechanisms associated with long-term in vitro culture and differentiation of ASCs, as well as providing a point of reference for potential in vivo and clinical studies regarding these cells’ application in regenerative medicine.

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Chemically defined and xeno-free culture condition for human extended pluripotent stem cells

Nature Communications ◽

10.1038/s41467-021-23320-8 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Bei Liu ◽

Shi Chen ◽

Yaxing Xu ◽

Yulin Lyu ◽

Jinlin Wang ◽

...

Keyword(s):

Stem Cells ◽

Pluripotent Stem Cells ◽

Culture Condition ◽

Human Fibroblast ◽

Culture System ◽

Disease Modeling ◽

Directed Differentiation

AbstractExtended pluripotent stem (EPS) cells have shown great applicative potentials in generating synthetic embryos, directed differentiation and disease modeling. However, the lack of a xeno-free culture condition has significantly limited their applications. Here, we report a chemically defined and xeno-free culture system for culturing and deriving human EPS cells in vitro. Xeno-free human EPS cells can be long-term and genetically stably maintained in vitro, as well as preserve their embryonic and extraembryonic developmental potentials. Furthermore, the xeno-free culturing system also permits efficient derivation of human EPS cells from human fibroblast through reprogramming. Our study could have broad utility in future applications of human EPS cells in biomedicine.

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MYOGENIC DIFFERENTIATION OF HUMAN MESENCHYMAL STEM CELLS IN VITRO AND LONG-TERM SURVIVAL IN A NUDE RAT MODEL FOR URINARY INCONTINENCE

The Journal of Urology ◽

10.1016/s0022-5347(09)61907-8 ◽

2009 ◽

Vol 181 (4) ◽

pp. 681

Author(s):

Gerhard Feil ◽

Adriana Drost ◽

Simon Baumann ◽

Jochen Schäfer ◽

Sibylle Weng ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Urinary Incontinence ◽

Rat Model ◽

Myogenic Differentiation ◽

Human Mesenchymal Stem Cells ◽

Term Survival ◽

Nude Rat

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71 TECHNIQUE FOR ESTABLISHMENT OF LONG TERM IN VITRO CULTURES OF PRIMARY MDS CELLS

Leukemia Research ◽

10.1016/s0145-2126(15)30072-2 ◽

2015 ◽

Vol 39 ◽

pp. S35

Author(s):

C. Ichim ◽

D. Koos ◽

T. Ichim ◽

R. Wells

Keyword(s):

In Vitro Cultures

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Effect of dexamethasone as osteogenic supplementation in in vitro osteogenic differentiation of stem cells from human exfoliated deciduous teeth

Journal of Materials Science Materials in Medicine ◽

10.1007/s10856-020-06475-6 ◽

2021 ◽

Vol 32 (1) ◽

Author(s):

Mariane Beatriz Sordi ◽

Raissa Borges Curtarelli ◽

Izabella Thaís da Silva ◽

Gislaine Fongaro ◽

Cesar Augusto Magalhães Benfatti ◽

...

Keyword(s):

Stem Cells ◽

Extracellular Matrix ◽

Osteogenic Differentiation ◽

Culture Media ◽

Deciduous Teeth ◽

Free Calcium ◽

Alizarin Red ◽

Matrix Mineralization ◽

Media Supplementation

AbstractIn in vitro culture systems, dexamethasone (DEX) has been applied with ascorbic acid (ASC) and β-glycerophosphate (βGLY) as culture media supplementation to induce osteogenic differentiation of mesenchymal stem cells. However, there are some inconsistencies regarding the role of DEX as osteogenic media supplementation. Therefore, this study verified the influence of DEX culture media supplementation on the osteogenic differentiation, especially the capacity to mineralize the extracellular matrix of stem cells from human exfoliated deciduous teeth (SHED). Five groups were established: G1—SHED + Dulbecco’s Modified Eagles’ Medium (DMEM) + fetal bovine serum (FBS); G2—SHED + DMEM + FBS + DEX; G3—SHED + DMEM + FBS + ASC + βGLY; G4—SHED + DMEM + FBS + ASC + βGLY + DEX; G5—MC3T3-E1 + α Minimal Essential Medium (MEM) + FBS + ASC + βGLY. DNA content, alkaline phosphatase (ALP) activity, free calcium quantification in the extracellular medium, and extracellular matrix mineralization quantification through staining with von Kossa, alizarin red, and tetracycline were performed on days 7 and 21. Osteogenic media supplemented with ASC and β-GLY demonstrated similar effects on SHED in the presence or absence of DEX for DNA content (day 21) and capacity to mineralize the extracellular matrix according to alizarin red and tetracycline quantifications (day 21). In addition, the presence of DEX in the osteogenic medium promoted less ALP activity (day 7) and extracellular matrix mineralization according to the von Kossa assay (day 21), and more free calcium quantification at extracellular medium (day 21). In summary, the presence of DEX in the osteogenic media supplementation did not interfere with SHED commitment into mineral matrix depositor cells. We suggest that DEX may be omitted from culture media supplementation for SHED osteogenic differentiation in vitro studies.

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