Metabolic glycan labeling-assisted discovery of cell-surface markers for primary neural stem and progenitor cells

2018 ◽  
Vol 54 (43) ◽  
pp. 5486-5489
Author(s):  
Qing-Ran Bai ◽  
Lu Dong ◽  
Yi Hao ◽  
Xing Chen ◽  
Qin Shen
Keyword(s):  
Mass Spectrometry ◽  
Stem Cell ◽  
Cell Surface ◽  
Progenitor Cells ◽  
Neural Stem Cell ◽  
Surface Markers ◽  
Metabolic Labeling ◽  
Cell Surface Markers ◽  
Stem And Progenitor Cells

Metabolic labeling with azidosugars in a neural stem cell (NSC)-enriched endothelial coculture followed by mass-spectrometry profiling identifies sialoglycoproteins on NSCs.

Identification of Novel Surface Markers and Targets In Neoplastic Stem Cells In AML and CML: a Flow-Gene-Flow Screen Approach.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1575-1575 ◽  
Author(s):  
Harald Herrmann ◽  
Sabine Cerny-Reiterer ◽  
Irina Sadovnik ◽  
Viviane Winter ◽  
Katharina Blatt ◽  
...  
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Stem Cell ◽  
Cell Surface ◽  
Progenitor Cells ◽  
Gene Chip ◽  
Cytokine Receptors ◽  
Stem Cell Markers ◽  
Surface Markers ◽  
Leukemic Stem Cells

Abstract Abstract 1575 The concept of leukemic stem cells (LSC) is increasingly employed to explain the biology of various myeloid neoplasms and to screen for pivotal targets, with the hope to improve drug therapy through elimination of disease-initiating cells. Although the stem cell hypothesis may apply to all neoplasms, leukemia-initiating cells have so far only been characterized in some detail in myeloid leukemias. In an attempt to identify novel cell surface markers and targets on leukemic stem cells (LSC) in acute (AML) and chronic myeloid leukemia (CML), we examined CD34+/CD38- and CD34+/CD38+ populations of leukemic cells in a cohort of patients with AML (n=55) and CML (n=20). In a first step, cell surface antigen profiles were determined by multicolor flow cytometry. In this screen, we were able to show that CD34+/CD38- LSC in AML and CML consistently express certain cytokine receptors, including G-CSFR (CD114), SCFR/KIT (CD117), and IL-3RA (CD123). The low affinity IL-2R (CD25) was detectable on CD34+/CD38- stem cells in patients with CML, and in a subset of AML patients. Other cytokine receptors (R) such as FLT3, IGF-1R, endoglin (CD105), GM-CSFRA (CD116), and OSMR were expressed variably on CD34+/CD38- progenitor cells, whereas the EPOR was not detectable on LSC. We were also able to detect several established therapeutic targets on LSC, including CD33 and CD44. Whereas CD44 was consistently expressed on all LSC in all donors, CD33 was found to be expressed variably on subpopulations of LSC in AML and CML, depending on the phase and type of disease. By using cytokine ligands (G-CSF, IL-3, SCF, EPO) and targeted drugs, we were also able to confirm that identified cytokine receptors and targets were functionally active molecules and potentially relevant targets. In a next step, highly enriched (purity >98%) sorted CD34+/CD38- cells, CD34+/CD38+ cells, and CD34- cells were collected in patients with AML and CML, and in 3 cord blood samples as controls. Purified cells were subjected to gene chip analyses, qPCR, and functional analyses. The identity of leukemic progenitors was confirmed by FISH, and expression of markers and targets in CML stem cells and AML stem cells was confirmed by qPCR. In gene chip analyses, we screened for novel LSC markers and targets. Candidate genes were selected based and their specific expression in progenitor cell fractions and surface membrane location, which was confirmed by antibody staining experiments. Novel stem cell markers identified so far include ROBO4, NPDC-1, and CXCR7. The previously described surface markers MDR-1 and CLL-1 were also identified by flow cytometry, but were also found to be expressed on more mature hematopoietic cells. By contrast, ROBO4 was found to be expressed preferentially on CD34+/CD38- stem cells, but less abundantly on CD34+/CD38+ progenitor cells in CML. Interestingly, whereas ROBO4 was expressed on CD34+/CD38- stem cells in most patients with CML, ROBO4 expression on leukemic stem cells was only found in a subset of AML patients. By contrast, CD34+/CD38- stem cells in AML frequently expressed CLL-1 and NPDC-1 on their surface. In conclusion, we have identified novel markers and targets in CD34+/CD38- progenitor cells in AML and CML. These markers may be useful for the identification and isolation of leukemic stem cells in AML and CML, and for the validation of drug effects on these cells. Disclosures: De Angelis: Biopharm R&D, GSK: Employment. Holmes:Biopharm R&D, GSK: Employment. Valent:Domantis: Research Funding.

Developing Cell-Specific Antibodies to Endothelial Progenitor Cells Using Avian Immune Phage Display Technology

2011 ◽  
Vol 16 (7) ◽  
pp. 744-754 ◽  
Author(s):  
Tyrone Bowes ◽  
Shirley A. Hanley ◽  
Aaron Liew ◽  
Marc Eglon ◽  
Kaveh Mashayekhi ◽  
...  
Keyword(s):  
Cell Surface ◽  
Phage Display ◽  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Peripheral Blood ◽  
Surface Markers ◽  
Cell Surface Markers ◽  
Endothelial Progenitor ◽  
Phage Display Technology ◽  
Display Technology

This study aims at generating immune chicken phage display libraries and single-chain antibodies (scFvs) specifically directed against cell surface markers of cultured peripheral blood mononuclear cells (PBMCs) that contain endothelial progenitor cells (EPCs). In contrast to previous approaches that use well-defined recombinant antigens attached to plastic surfaces that may alter the structure of the proteins, the authors describe a method that maintains the cell surface markers on live cells while providing the opportunity to rapidly screen entire libraries for antibodies that bind to unknown cell surface markers of progenitor/stem cells. Chickens immunized with live EPCs, consisting of a heterogeneous population of lymphocytes and monocytes, demonstrated a robust immune response. After three rounds of biopanning, the authors purified and characterized three unique scFvs called UG1-3. Codon-optimized recombinant UG1 (gUG-1) shows binding by flow cytometry to circulating CD14-positive cells in peripheral blood consistent with predominant expression of a target protein on monocyte subsets. The authors describe the successful use of immunization of chickens for the generation of scFvs against a heterogenous population of EPCs displaying unknown cell surface markers and demonstrate the strong potential of phage display technology in the development of reagents for the isolation and characterization of stem/progenitor cells.

scholarly journals Gingival-Derived Mesenchymal Stem Cell from Rabbit (Oryctolagus cuniculus): Isolation, Culture, and Characterization

2020 ◽  
Author(s):  
Alexander Patera Nugraha ◽  
Fedik Abdul Rantam ◽  
Ida Bagus Narmada ◽  
Diah Savitri Ernawati ◽  
Igo Syaiful Ihsan
Keyword(s):  
Flow Cytometry ◽  
Stem Cell ◽  
Cell Surface ◽  
Osteogenic Differentiation ◽  
Oryctolagus Cuniculus ◽  
Surface Markers ◽  
Cell Surface Markers ◽  
Alizarin Red ◽  
Regenerative Dentistry

Abstract Objective This study aims to confirm whether the GDMSCs isolated from rabbit’s (Oryctolagus cuniculus) gingiva are mesenchymal stem cells (MSCs). Materials and Methods This study design was partly quasi-experimental with an observational design. GDMSCs were isolated from the gingiva of healthy male rabbits (O. cuniculus) (n = 2), 6 months old, and 3 to 5 kg of body weight. The specific cell surface markers of MSCs; clusters of differentiation (CD), namely, CD44, CD73, CD90, CD105, and CD200 expressions; and hematopoietic stem cell surface markers CD34 and CD45 were examined using flow cytometry and immunohistochemistry with immunofluorescence. The osteogenic differentiation of isolated GDMSCs was examined using alizarin red staining. Results GDMSCs in the fourth passage showed a spindle-like formation and fibroblast-like cells that attached to the base of the culture plate. GDMSCs were MSCs that positively expressed CD44, CD73, CD90, CD105, and CD200 but did not express CD34 and CD45 when examined using flow cytometry and immunohistochemical analysis. GDMSCs had osteogenic differentiation confirmed by calcified deposits in vitro with a red–violet and brownish color after alizarin red staining. Conclusion GDMSCs isolated from the rabbits (O. cuniculus) were confirmed as MSCs in vitro documented using immunohistochemistry and flow cytometry. GDMSCs can differentiate into osteogenic lineage in vitro that may be suitable for regenerative dentistry.

scholarly journals Single-cell RNA-Sequencing uncovers transcriptional states and fate decisions in haematopoiesis

2017 ◽  
Author(s):  
Emmanouil I. Athanasiadis ◽  
Jan G. Botthof ◽  
Helena Andres ◽  
Lauren Ferreira ◽  
Pietro Lio ◽  
...  
Keyword(s):  
Cell Surface ◽  
Specific Cell ◽  
Surface Markers ◽  
Evolutionary Analysis ◽  
Cell Surface Markers ◽  
Lineage Differentiation ◽  
Lineage Tree ◽  
Stem And Progenitor Cells ◽  
A Cell ◽  
Fate Decision

ABSTRACTThe success of marker-based approaches for dissecting haematopoiesis in mouse and human is reliant on the presence of well-defined cell-surface markers specific for diverse progenitor populations. An inherent problem with this approach is that the presence of specific cell surface markers does not directly reflect the transcriptional state of a cell. Here we used a marker-free approach to computationally reconstruct the blood lineage tree in zebrafish and order cells along their differentiation trajectory, based on their global transcriptional differences. Within the population of transcriptionally similar stem and progenitor cells our analysis revealed considerable cell-to-cell differences in their probability to transition to another, committed state. Once fate decision was executed, the suppression of transcription of ribosomal genes and up-regulation of lineage specific factors coordinately controlled lineage differentiation. Evolutionary analysis further demonstrated that this haematopoietic program was highly conserved between zebrafish and higher vertebrates.

Breast cancer stem cells: A novel therapy for breast cancer.

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. e13030-e13030
Author(s):  
Soma Mukhopadhyay ◽  
Sudeshna Gangopadhyay ◽  
Swati Dasgupta ◽  
Saubhik Sengupta ◽  
Ujjal Kanti Ray ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Stem Cells ◽  
Stem Cell ◽  
Cancer Stem Cells ◽  
Cell Surface ◽  
Surface Markers ◽  
Breast Cancer Stem Cells ◽  
Cell Surface Markers ◽  
Primary Tumors ◽  
Free Survival

e13030 Background: Simple BRCA screening is insufficient for ‘event-free survival’ as breast cancer is clinically and pathologically an extremely heterogeneous disease. Targeting Breast Cancer Stem Cells (BCSCs) present in bone marrow and breast tissues is a lucrative alternative. Identification of BCSCs is salient aspect of our research. Invasive and mesenchymal property of BCSCs with CD44+/CD24low/ALDH1+ phenotype has made them a promising target for eliminating metastatic capacity of primary tumors. We hypothesize that ability to therapeutically attack stem cell hinges upon identifying unique targets like cell surface markers and this will decide development of specific target therapies. Methods: A total of 10 early chemo-naive patients with biopsy proven triple-negative metastatic breast cancer in the age group of 18-36 yr.s (mean age 28 yr.s) were selected randomly and tested for CD44/CD24 cell surface markers following immunosorting using magnetic cell sorter and immunophenotyping by flowcytometric analysis. Isolated BCSCs were cultured for in vitro drug sensitivity towards platinum, anthracycline and docetaxel. Correlation was drawn between cell differentiation, % of stem cells and drug response. Accordingly chemotherapy was designed for a particular patient. % of BCSCs in pre- and post-chemotherapeutic condition was further compared. Results: We have detected BCSCs in 90% of cases. Among positive samples, 89% patients showed platinum sensitivity and rest were found to be anthracycline sensitive. No sensitivity to docetaxel was observed. In lieu of this, cisplatin was applied in vivo and % of BCSCs came down to 6.58% from initial 11.16% (for a representative case). Conclusions: Thus primary aim to target BCSCs at the onset of tumors in breast cancer patients to control metastasis and relapse of disease was somewhat obtained. We further plan to correlate ratio of selected markers present in patients in pre- and post-chemotherapeutic condition with time to recurrence, mortality, morbidity and progression-free survival. Finally, if no BCSCs prevail after chemotherapy, then patients would be kept under observation and if traces are found, we would proceed to targeted therapy trial like PARP inhibitor or autologous stem cell replacement.

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