scholarly journals 2057 L1 expression analysis in adipose-derived stem cells

2018 ◽  
Vol 2 (S1) ◽  
pp. 16-16
Author(s):  
Tiffany Kaul ◽  
Rachel Sabol ◽  
Maria E. Morales ◽  
Bruce Bunnell ◽  
Prescott Deininger
Keyword(s):  
Stem Cells ◽  
Cell Function ◽  
Cell Types ◽  
Full Length ◽  
Cell Populations ◽  
Potential Contribution ◽  
Adipose Derived Stem Cells ◽  
Bioinformatics Pipeline ◽  
Cytoplasmic Rna ◽  
L1 Element

OBJECTIVES/SPECIFIC AIMS: Long interspersed element-1s (L1s) are autonomous, mobile elements that are able to copy and insert themselves throughout the genome with their own reverse transcriptase and endonuclease. These elements make up 17% of the human genome with over 500,000 copies, though the vast majority of L1s are defective with only a few dozen potentially responsible for L1 activity. Full-length L1s have the potential to contribute to mutagenesis through random insertion and increased genetic instability. Here we set out to study L1 expression at the specific loci level in bone marrow-derived stem cells (bmSCs) and adipose-derived stem cells (ASCs) and compare the levels of expression from ASCs from donor patients who are young and lean, obese, and old. Our hypothesis is that L1-related damage may contribute to mutation and inflammation that alters the function of these stem cells throughout the life of an individual. METHODS/STUDY POPULATION: ASCs and bmSCs were isolated from patient donors. The following samples were collected: ASCs from 3 young (under the age of 59) and lean (BMI<30) patients, ASCs from 3 older patients (over the age of 59), ASCs from 3 patients with BMI>30, and bmSCs from 4 young and lean patients. Cytoplasmic RNA from the cell populations was isolated and sequenced by RNA-Seq from the cell populations. Using our recently developed bioinformatics pipeline, we set out to quantify L1 expression and identify the few culprit L1s at specific loci that are actively transcribing to RNA in the ASC and bmSC samples. RESULTS/ANTICIPATED RESULTS: Here we provide proof of concept with the application of this novel method in characterizing full-length expressed L1s at the specific loci level in ASCs and bmSCs. We identified L1 loci that are commonly expressed in these cell types and observed an increase in L1 expression in the obese and old ASC cells compared with the young, lean ASCs and bmSCs. DISCUSSION/SIGNIFICANCE OF IMPACT: ASCs hold the promise of broad application in the biomedical field including regenerative treatment. There are reports that ASCs cultivated from older and obese donors are less effective in regenerative treatments. By demonstrating an increased expression of the mutagenic L1 element in ASCs from obese and old donors, this study provides further evidence suggesting the preferable use of ASCs from young and lean donors for regenerative therapies. These studies will also help us to understand the potential contribution of L1 expression to loss of stem cell function during the aging process.

scholarly journals Stem cells: Aging and transcriptional fingerprints

2017 ◽  
Vol 217 (1) ◽  
pp. 79-92 ◽  
Author(s):  
Brice E. Keyes ◽  
Elaine Fuchs
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Cell Function ◽  
Cell Types ◽  
Cell Populations ◽  
Functional Abilities ◽  
Future Directions ◽  
Age Related ◽  
Transcriptional Changes ◽  
Stem Cell Populations

Stem cells are imbued with unique qualities. They have the capacity to propagate themselves through symmetric divisions and to divide asymmetrically to engender new cells that can progress to differentiate into tissue-specific, terminal cell types. Armed with these qualities, stem cells in adult tissues are tasked with replacing decaying cells and regenerating tissue after injury to maintain optimal tissue function. With increasing age, stem cell functional abilities decline, resulting in reduced organ function and delays in tissue repair. Here, we review the effect of aging in five well-studied adult murine stem cell populations and explore age-related declines in stem cell function and their consequences for stem cell self-renewal, tissue homeostasis, and regeneration. Finally, we examine transcriptional changes that have been documented in aged stem cell populations and discuss new questions and future directions that this collection of data has uncovered.

313 UNSORTED, FRESHLY ISOLATED PORCINE ADIPOSE-DERIVED STEM CELLS ARE MORE EFFICACIOUS IN BONE HEALING COMPARED WITH PURIFIED CD34+ ADIPOSE-DERIVED STEM CELLS

2011 ◽  
Vol 23 (1) ◽  
pp. 253
Author(s):  
M. Bionaz ◽  
T. Jensen ◽  
E. Monaco ◽  
Z. Dymon ◽  
A. J. Maki ◽  
...  
Keyword(s):  
Stem Cells ◽  
Random Effect ◽  
Cell Types ◽  
Cell Populations ◽  
Adipose Derived Stem Cells ◽  
Cd34 Cells ◽  
Vitro Data ◽  
In Vitro Data

We have previously shown that heterologous transplantation of porcine adipose-derived stem cells (ADSC) enhances bone healing. Freshly harvested ADSC are a heterogeneous population that contains several types of cells other than stem cells. The isolation of highly purified ADSC could be of clinical importance. In this study, we compared the in vitro growth characteristics and in vivo healing potential of ADSC unsorted or separated using CD34 as a marker. The ADSC were extracted from the back fat of 4 male pigs at 6 months of age. For the in vitro experiment, aliquots of the ADSC were sorted by magnetic beads (Miltenyi Biotec, Gladbach, Germany) into CD34-positive (CD34+) and CD34-negative (CD34–) cell populations. The unsorted ADSC (uADSC), plus the CD34+, CD34–, and a 50:50 mixture of CD34+ and CD34– (MIX) were plated in 24-well plates and differentiated into osteocytes. A robotic stage inverted microscope was used to photograph the entire culture well, and then number, dimension, and density of bone nodules were assessed. Alizarin red (AR) staining was performed and quantified. Cells were harvested before cell plating and then on several time points during expansion, at confluence, and at 3, 6, or 18 days [d] of differentiation for cell counting and RNA extraction. Real-time RT-PCR was performed for CD34, COL1A1, and SPARC genes. For the in vivo experiment, freshly isolated ADSC were sorted by flow cytometry into CD34+ and CD34– cell populations. Unsorted and sorted cells were transplanted, in duplicate, into 10- or 25-mm mandible osteoctomies. Mandibles were harvested after 8 weeks for evaluation of healing by DEXA scanning. In vitro data were statistically analysed using a mixed model (SAS) with time and cell type as fixed effect and pig as the random effect. The in vivo data were assessed by ANOVA with cell types as the fixed effect and pig as the random effect. Freshly harvested ADSC contained 42.3 ± 11.0% CD34+ cells. The uADSC reached confluence at 6 days after plating, whereas other cell types reached confluence at 16 days. Expression of CD34 decreased after plating but was similar between cell types. Among osteogenic genes, only expression of SPARC increased during differentiation. The number of osteogenic nodules was higher (P < 0.05) in uADSC than the in other cell types, but the area and nodule density were similar to CD34– and greater (P < 0.05) than CD34+ and MIX. The amount of AR was higher (P < 0.05) in uADSC compared with CD34– and MIX but similar to CD34+. In the in vivo trial, uADSC had a greater (P < 0.05) healing compared with sorted cells. Among those, CD34– cells appeared to increase healing compared with CD34+ cells. Results indicate that CD34+ cells do not differ significantly from CD34– in the in vitro osteogenic differentiation but have lower in vivo healing capacity; however, in vitro data were confused by a lack of pure CD34– cells. The freshly isolated ADSC have a greater healing capacity than sorted cells, as indicated by in vitro and in vivo experiments. Overall our data indicate that the sorting of ADSC CD34+ cells is not of clinical relevance.

scholarly journals Multiple Roles for Cholinergic Signaling from the Perspective of Stem Cell Function

2021 ◽  
Vol 22 (2) ◽  
pp. 666
Author(s):  
Toshio Takahashi
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Cell Function ◽  
Cell Activity ◽  
Embryonic Stem ◽  
Cholinergic Neurons ◽  
Cell Types ◽  
Proliferative Potential ◽  
True Nature ◽  
Cholinergic Signaling

Stem cells have extensive proliferative potential and the ability to differentiate into one or more mature cell types. The mechanisms by which stem cells accomplish self-renewal provide fundamental insight into the origin and design of multicellular organisms. These pathways allow the repair of damage and extend organismal life beyond that of component cells, and they probably preceded the evolution of complex metazoans. Understanding the true nature of stem cells can only come from discovering how they are regulated. The concept that stem cells are controlled by particular microenvironments, also known as niches, has been widely accepted. Technical advances now allow characterization of the zones that maintain and control stem cell activity in several organs, including the brain, skin, and gut. Cholinergic neurons release acetylcholine (ACh) that mediates chemical transmission via ACh receptors such as nicotinic and muscarinic receptors. Although the cholinergic system is composed of organized nerve cells, the system is also involved in mammalian non-neuronal cells, including stem cells, embryonic stem cells, epithelial cells, and endothelial cells. Thus, cholinergic signaling plays a pivotal role in controlling their behaviors. Studies regarding this signal are beginning to unify our understanding of stem cell regulation at the cellular and molecular levels, and they are expected to advance efforts to control stem cells therapeutically. The present article reviews recent findings about cholinergic signaling that is essential to control stem cell function in a cholinergic niche.

scholarly journals Human Adipose-Derived Stem Cells Impair Natural Killer Cell Function and Exhibit Low Susceptibility to Natural Killer-Mediated Lysis

2012 ◽  
Vol 21 (8) ◽  
pp. 1333-1343 ◽  
Author(s):  
Olga DelaRosa ◽  
Beatriz Sánchez-Correa ◽  
Sara Morgado ◽  
Cristina Ramírez ◽  
Borja del Río ◽  
...  
Keyword(s):  
Stem Cells ◽  
Natural Killer Cell ◽  
Natural Killer ◽  
Cell Function ◽  
Killer Cell ◽  
Adipose Derived Stem Cells ◽  
Natural Killer Cell Function

scholarly journals Regional Anatomic and Age Effects on Cell Function of Human Adipose-Derived Stem Cells

2008 ◽  
Vol 60 (5) ◽  
pp. 538-544 ◽  
Author(s):  
Bret M. Schipper ◽  
Kacey G. Marra ◽  
Wei Zhang ◽  
Albert D. Donnenberg ◽  
J Peter Rubin
Keyword(s):  
Stem Cells ◽  
Cell Function ◽  
Age Effects ◽  
Adipose Derived Stem Cells

scholarly journals Aldehyde Dehydrogenases: Not Just Markers, but Functional Regulators of Stem Cells

2019 ◽  
Vol 2019 ◽  
pp. 1-15 ◽  
Author(s):  
Giuseppe Vassalli
Keyword(s):  
Stem Cells ◽  
Molecular Mechanisms ◽  
Cell Types ◽  
Cellular Function ◽  
Stem Cell Marker ◽  
Cell Populations ◽  
Aldh Activity ◽  
Cellular Functions ◽  
Stem And Progenitor Cells

Aldehyde dehydrogenase (ALDH) is a superfamily of enzymes that detoxify a variety of endogenous and exogenous aldehydes and are required for the biosynthesis of retinoic acid (RA) and other molecular regulators of cellular function. Over the past decade, high ALDH activity has been increasingly used as a selectable marker for normal cell populations enriched in stem and progenitor cells, as well as for cell populations from cancer tissues enriched in tumor-initiating stem-like cells. Mounting evidence suggests that ALDH not only may be used as a marker for stem cells but also may well regulate cellular functions related to self-renewal, expansion, differentiation, and resistance to drugs and radiation. ALDH exerts its functional actions partly through RA biosynthesis, as all-trans RA reverses the functional effects of pharmacological inhibition or genetic suppression of ALDH activity in many cell types in vitro. There is substantial evidence to suggest that the role of ALDH as a stem cell marker comes down to the specific isoform(s) expressed in a particular tissue. Much emphasis has been placed on the ALDH1A1 and ALDH1A3 members of the ALDH1 family of cytosolic enzymes required for RA biosynthesis. ALDH1A1 and ALDH1A3 regulate cellular function in both normal stem cells and tumor-initiating stem-like cells, promoting tumor growth and resistance to drugs and radiation. An improved understanding of the molecular mechanisms by which ALDH regulates cellular function will likely open new avenues in many fields, especially in tissue regeneration and oncology.

Quantitative Imaging of Femoral Bone Marrow Microenvironments Reveals a Heterogenous Distribution of Hematopoietic Stem and Progenitor Cells.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1455-1455
Author(s):  
Cesar Nombela-Arrieta ◽  
Brendan Harley ◽  
Gregory Pivarnik ◽  
John E Mahoney ◽  
Elena Levantini ◽  
...  
Keyword(s):  
Stem Cells ◽  
B Cells ◽  
Cell Biology ◽  
Conflicts Of Interest ◽  
Statistical Significance ◽  
Quantitative Imaging ◽  
Cell Types ◽  
Immunofluorescent Staining ◽  
Cell Populations ◽  
Hematopoietic Stem

Abstract Abstract 1455 Poster Board I-478 Sustained production of all mature blood cell types relies on the continuous proliferation and differentiation of a rare population of self-renewing, multipotent hematopoietic stem cells (HSCs). HSC maintenance and lineage differentiation are thought to be regulated by spatially confined niches, defined by cellular components, soluble regulators, and the extracellular matrix immediately surrounding stem cells. Identification of these microenvironments in which endogenous and transferred HSCs reside within the BM is a major challenge in stem cell biology with relevant clinical implications. Yet the extreme rarity of HSCs, their dynamic nature, and the lack of specific markers to identify them, have precluded an accurate definition of HSC niches to date. Quantitative imaging technologies such as Laser Scanning Cytometry (LSC) are designed for the automated analysis of large cell numbers at a single cell level with high resolution while preserving the morphological information lost in flow cytometry, therefore providing data of statistical significance even for rare cell populations such as HSCs. We have employed LSC to analyze the localization of both adoptively transferred and endogenous hematopoietic stem and progenitor cell (HSPC) populations inside whole longitudinal sections of murine femoral BM cavities. Our results indicate that, as previously suggested, purified HSPC (Lin−c-kit+Sca-1+) significantly accumulate in endosteal regions (ER) of BM cavities (within 100μm of inner bone surface) upon transplantation. Nevertheless, analysis of sufficient numbers of more differentiated cell subsets (Lin−c-kit+Sca-1− progenitors, pro B cells and mature B cells) indicated that these areas serve as homing sites for most hematopoietic cells, highlighting the limitations of any conclusions drawn on HSC niche identity from studies performed with transferred HSPC populations. Immunofluorescent staining of endogenous cell populations revealed a gradient in distribution of early hematopoietic progenitors (c-kit+), which accumulated in but were not restricted to ER regions. Of note, a vast majority (>80%) of HSPC (Bmi-GFPhic-kit+, or Lin−c-kit+Sca-1+),were found inside ER, although not directly adjacent to endosteal surfaces. Our studies define endosteal areas as tissue regions where HSPC reside in close proximity, but not necessarily in direct contact with a dense vascular network, osteoblastic cells and other potential niche cell types and growth factors currently under investigation. Disclosures No relevant conflicts of interest to declare.

scholarly journals Full-Length Transcriptome: A Reliable Alternative for Single-Cell RNA-Seq Analysis in the Spleen of Teleost Without Reference Genome

2021 ◽  
Vol 12 ◽  
Author(s):  
Lixing Huang ◽  
Ying Qiao ◽  
Wei Xu ◽  
Linfeng Gong ◽  
Rongchao He ◽  
...  
Keyword(s):  
Data Analysis ◽  
B Cells ◽  
Single Cell ◽  
Reference Genome ◽  
Immune Cell ◽  
Cell Types ◽  
Full Length ◽  
Cell Populations ◽  
Epinephelus Coioides ◽  
Rna Seq

Fish is considered as a supreme model for clarifying the evolution and regulatory mechanism of vertebrate immunity. However, the knowledge of distinct immune cell populations in fish is still limited, and further development of techniques advancing the identification of fish immune cell populations and their functions are required. Single cell RNA-seq (scRNA-seq) has provided a new approach for effective in-depth identification and characterization of cell subpopulations. Current approaches for scRNA-seq data analysis usually rely on comparison with a reference genome and hence are not suited for samples without any reference genome, which is currently very common in fish research. Here, we present an alternative, i.e. scRNA-seq data analysis with a full-length transcriptome as a reference, and evaluate this approach on samples from Epinephelus coioides-a teleost without any published genome. We show that it reconstructs well most of the present transcripts in the scRNA-seq data achieving a sensitivity equivalent to approaches relying on genome alignments of related species. Based on cell heterogeneity and known markers, we characterized four cell types: T cells, B cells, monocytes/macrophages (Mo/MΦ) and NCC (non-specific cytotoxic cells). Further analysis indicated the presence of two subsets of Mo/MΦ including M1 and M2 type, as well as four subsets in B cells, i.e. mature B cells, immature B cells, pre B cells and early-pre B cells. Our research will provide new clues for understanding biological characteristics, development and function of immune cell populations of teleost. Furthermore, our approach provides a reliable alternative for scRNA-seq data analysis in teleost for which no reference genome is currently available.

scholarly journals Changes in ionizing radiation dose rate affect cell cycle progression in adipose derived stem cells

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0250160
Author(s):  
Matthew Rusin ◽  
Nardine Ghobrial ◽  
Endre Takacs ◽  
Jeffrey S. Willey ◽  
Delphine Dean
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Radiation Therapy ◽  
Stem Cell ◽  
Regenerative Medicine ◽  
Ionizing Radiation ◽  
Dose Rate ◽  
Cell Types ◽  
High Energy ◽  
Adipose Derived Stem Cells

Biomedical use of radiation is utilized in effective diagnostic and treatment tools, yet can introduce risks to healthy tissues. High energy photons used for diagnostic purposes have high penetration depth and can discriminate multiple tissues based on attenuation properties of different materials. Likewise, the ability to deposit energy at various targets within tumors make the use of photons effective treatment for cancer. Radiation focused on a tumor will deposit energy when it interacts with a biological structure (e.g. DNA), which will result in cell kill should repair capacity of the tissue be overwhelmed. Likewise, damage to normal, non-cancerous tissues is a consequence of radiation that can lead to acute or late, chronic toxicity profiles. Adipose derived stem cells (ADSCs) are mesenchymal stem cells that have been proven to have similar characteristics to bone marrow derived stem cells, except that they are much easier to obtain. Within the body, ADSCs act as immunomodulators and assist with the maintenance and repair of tissues. They have been shown to have excellent differentiation capability, making them an extremely viable option for stem cell therapies and regenerative medicine applications. Due to the tissue ADSCs are derived from, they are highly likely to be affected by radiation therapy, especially when treating tumors localized to structures with relatively high ADSC content (eg., breast cancer). For this reason, the purpose behind this research is to better understand how ADSCs are affected by doses of radiation comparable to a single fraction of radiation therapy. We also measured the response of ADSCs to exposure at different dose rates to determine if there is a significant difference in the response of ADSCs to radiation therapy relevant doses of ionizing radiation. Our findings indicate that ADSCs exposed to Cesium (Cs 137)-gamma rays at a moderate dose of 2Gy and either a low dose rate (1.40Gy/min) or a high dose rate (7.31Gy/min) slow proliferation rate, and with cell cycle arrest in some populations. These responses ADSCs were not as marked as previously measured in other stem cell types. In addition, our results indicate that differences in dose rate in the Gy/min range typically utilized in small animal or cell irradiation platforms have a minimal effect on the function of ADSCs. The potential ADSCs have in the space of regenerative medicine makes them an ideal candidate for study with ionizing radiation, as they are one of the main cell types to promote tissue healing.

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