Real-time quantitative RT-PCR analysis of human bone marrow stromal cells during osteogenic differentiation in vitro

2002 ◽  
Vol 85 (4) ◽  
pp. 737-746 ◽  
Author(s):  
Oliver Frank ◽  
Manuel Heim ◽  
Marcel Jakob ◽  
Andrea Barbero ◽  
Dirk Schäfer ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Osteogenic Differentiation ◽  
Real Time ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Marrow Stromal Cells ◽  
Human Bone ◽  
Pcr Analysis ◽  
Rt Pcr

Remodeling of Three-Dimensional Collagen I Matrices by Human Bone Marrow Stromal Cells during Osteogenic Differentiation In Vitro

2020 ◽  
Vol 3 (10) ◽  
pp. 6967-6978 ◽  
Author(s):  
Sarah Vogel ◽  
Franziska Ullm ◽  
Claudia Damaris Müller ◽  
Tilo Pompe ◽  
Ute Hempel
Keyword(s):  
Bone Marrow ◽  
Osteogenic Differentiation ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Three Dimensional ◽  
Marrow Stromal Cells ◽  
Human Bone ◽  
Collagen I ◽  
Human Bone Marrow

scholarly journals Human Bone Marrow Stromal Cells: A Reliable, Challenging Tool forIn VitroOsteogenesis and Bone Tissue Engineering Approaches

2016 ◽  
Vol 2016 ◽  
pp. 1-14 ◽  
Author(s):  
Ute Hempel ◽  
Katrin Müller ◽  
Carolin Preissler ◽  
Carolin Noack ◽  
Sabine Boxberger ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Osteogenic Differentiation ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Marrow Stromal Cells ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Peroxisome Proliferator ◽  
Differentiation Potential

Adult human bone marrow stromal cells (hBMSC) are important for many scientific purposes because of their multipotency, availability, and relatively easy handling. They are frequently used to study osteogenesisin vitro. Most commonly, hBMSC are isolated from bone marrow aspirates collected in clinical routine and cultured under the “aspect plastic adherence” without any further selection. Owing to the random donor population, they show a broad heterogeneity. Here, the osteogenic differentiation potential of 531 hBMSC was analyzed. The data were supplied to correlation analysis involving donor age, gender, and body mass index. hBMSC preparations were characterized as follows: (a) how many passages the osteogenic characteristics are stable in and (b) the influence of supplements and culture duration on osteogenic parameters (tissue nonspecific alkaline phosphatase (TNAP), octamer binding transcription factor 4, core-binding factor alpha-1, parathyroid hormone receptor, bone gla protein, and peroxisome proliferator-activated proteinγ). The results show that no strong prediction could be made from donor data to the osteogenic differentiation potential; only the ratio of induced TNAP to endogenous TNAP could be a reliable criterion. The results give evidence that hBMSC cultures are stable until passage 7 without substantial loss of differentiation potential and that established differentiation protocols lead to osteoblast-like cells but not to fully authentic osteoblasts.

Notch signaling enhances osteogenic differentiation while inhibiting adipogenesis in primary human bone marrow stromal cells

2009 ◽  
Vol 37 (7) ◽  
pp. 867-875.e1 ◽  
Author(s):  
Fernando Ugarte ◽  
Martin Ryser ◽  
Sebastian Thieme ◽  
Fernando A. Fierro ◽  
Katrin Navratiel ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Notch Signaling ◽  
Osteogenic Differentiation ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Marrow Stromal Cells ◽  
Human Bone ◽  
Human Bone Marrow

Isolation Yield and Osteogenic Potential of Human Bone Marrow Stromal Cells are Maintained Irrespective of Age or Gender

2007 ◽  
Vol 361-363 ◽  
pp. 1149-1152
Author(s):  
Jeong Joon Yoo ◽  
Jeon Hyun Bang ◽  
Kyung Hoi Koo ◽  
Kang Sup Yoon ◽  
Hee Joong Kim
Keyword(s):  
Bone Marrow ◽  
Osteogenic Differentiation ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Mononuclear Cells ◽  
Marrow Stromal Cells ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Osteogenic Potential ◽  
Donor Age

The relationships between donor age and gender and initial isolation yield and the osteogenic potentials of human bone marrow stromal cells (hBMSCs) have not been clearly elucidated. The authors investigated whether isolation yields and the osteogenic differentiation potentials of hBMSCs are indeed dependent on donor age or gender. Fresh bone marrow was aspirated from iliac crest of 72 donors (mean age 54.1 years; range, 23-84 years; 39 men and 33 women) undergoing total hip arthroplasty. Numbers of mononuclear cells, numbers of colony forming unit-fibroblasts (CFU-Fs) and alkaline phosphatase (ALP)-positive CFU-Fs, and numbers of BMSCs after isolation culture were not found to be significantly dependent on donor age or gender. Moreover, no significant age- or gender-related differences were observed in terms of the proliferation activities, ALP activities, and calcium contents of BMSCs during in vitro osteogenic differentiation. The data obtained from 72 human donors revealed no significant age- or genderrelated differences among hBMSCs in terms of isolation yields, proliferation activities, and osteogenic potentials.

Partial Characterization and In Vitro Expansion of Putative CLL Precursor/Stem Cells Which Are Dependent on Bone Marrow Microenvironment for Survival

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2433-2433
Author(s):  
Medhat Shehata ◽  
Rainer Hubmann ◽  
Martin Hilgarth ◽  
Susanne Schnabl ◽  
Dita Demirtas ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Marrow Stromal Cells ◽  
Rt Pcr ◽  
Hematopoietic Stem ◽  
Healthy Persons

Abstract Abstract 2433 Chronic lymphocytic leukemia (CLL) is characterized by the clonal expansion of B lymphocytes which typically express CD19 and CD5. The disease remains incurable and recurrence often occurs after current standard therapies due to residual disease or probably due to the presence of therapy-resistant CLL precursors. Based on the growing evidence for the existence of leukemia stem cells, this study was designed to search for putative CLL precursors/stem cells based on the co-expression of CLL cell markers (CD19/CD5) with the hematopoietic stem cell marker (CD34). Forty seven CLL patients and 17 healthy persons were enrolled in the study. Twenty four patients had no previous treatment and 23 had pre-therapy. Twenty two patients were in Binet stage C and 25 patients in B. Twenty two patients had unmutated and 18 mutated IgVH gene (7: ND). Cytogenetic analysis by FISH showed that 14 patients had del 13q, 8 had del 11q, 4 had del 17p and 9 had trisomy 12. Peripheral blood and bone marrow mononuclear cells were subjected to multi-colour FACS analysis using anti-human antibodies against CD34, CD19 and CD5 surface antigens. The results revealed the presence of triple positive CD34+/CD19+/CD5+ cells in CLL samples (mean 0.13%; range 0.01–0.41) and in healthy donors (0.31%; range 0.02–0.6) within the CD19+ B cells. However, due to the high leukocyte count in CLL patients, the absolute number of these cells was significantly higher in CLL samples (mean: 78.7; range 2.5–295 cells /μL blood) compared to healthy persons (mean: 0.45: range 0.04–2.5 cells/μl)(p<0,001). These triple positive “putative CLL stem cells” (PCLLSC) co-express CD133 (67%), CD38 (87%), CD127 (52%), CD10 (49%), CD20 (61%), CD23 (96%), CD44 (98%) and CD49d (74%). FISH analysis on 4 patients with documented chromosomal abnormalities detected the corresponding chromosomal aberrations of the mature clone in the sorted CD34+/CD5+/CD19+ and/or CD34+/CD19-/CD5- cells but not in the CD3+ T cells. Multiplex RT-PCR analysis using IgVH family specific primer sets confirmed the clonality of these cells. Morphologically, PCLLSC appeared larger than lymphocytes with narrow cytoplasm and showed polarity and motility in co-culture with human bone marrow stromal cells. Using our co-culture microenvironment model (Shehata et al, Blood 2010), sorted cell fractions (A: CD34+/19+/5+, B: CD34+/19-/5- or C: CD34-/CD19+/5+) from 4 patients were co-cultured with primary autologous human stromal cells. PCLLSC could be expanded in the co-culture to more than 90% purity from fraction A and B but not from fraction C. These cells remained in close contact or migrated through the stromal cells. PCLLSC required the contact with stromal cells for survival and died within 1–3 days in suspension culture suggesting their dependence on bone marrow microenvironment or stem cell niches. RT-PCR demonstrated that these cells belong to the established CLL clone. They also eexpress Pax5, IL-7R, Notch1, Notch2 and PTEN mRNA which are known to play a key role in the early stages of B cells development and might be relevant to the early development of the malignant clone in CLL. Using NOD/SCID/IL2R-gamma-null (NOG) xenogeneic mouse system we co-transplanted CLL cells from 3 patients (5 million PBMC/mouse) together with autologous bone marrow stromal cells (Ratio: 10:1). The percentage of PCLLSC in the transplanted PBMC was 0.18% (range 0.06–0.34%). Using human-specific antibodies, human CD45+ cells were detected in peripharal blood of the mice (mean 0.9 % range 0.47–1.63%) after 2 months of transplantation. More than 90% of the human cells were positive for CD45 and CD5. Among this population, 26% (range 15–35%) of the cells co-expressed CD45, CD19, CD5 and CD34 and thus correspond to the PCLLSC. In conclusion, our data suggest the existence of putative CLL precursors/stem cells which reside within the CD34+ hematopoietic stem cell compartment and carry the chromosomal aberrations of the established CLL clone. These cells could be expanded in vitro in a bone marrow stroma-dependent manner and could be engrafted and significantly enriched in vivo in NOG xenotransplant system. Further characterization and selective targeting and eradication of these cells may pave the way for designing curative therapeutic strategies for CLL. Disclosures: No relevant conflicts of interest to declare.

Selection of Common Markers for Bone Marrow Stromal Cells from Various Bones Using Real-Time RT-PCR: Effects of Passage Number and Donor Age

2007 ◽  
Vol 13 (10) ◽  
pp. 2405-2417 ◽  
Author(s):  
Akira Igarashi ◽  
Kazumi Segoshi ◽  
Yuhiro Sakai ◽  
Haiou Pan ◽  
Masami Kanawa ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Real Time ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Marrow Stromal Cells ◽  
Donor Age ◽  
Rt Pcr ◽  
Passage Number ◽  
Selection Of
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