scholarly journals Role of microvascular endothelial cells on proliferation, migration and adhesion of hematopoietic stem cells

2020 ◽  
Vol 40 (3) ◽  
Author(s):  
Fanli Lin ◽  
Shuyue Wang ◽  
Hao Xiong ◽  
Yang Liu ◽  
Xiaoming Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Endothelial Cells ◽  
Hematopoietic Stem Cells ◽  
Ex Vivo ◽  
Microvascular Endothelial Cells ◽  
Cell Contact ◽  
Hematopoietic Stem ◽  
Direct Cell ◽  
Cell Cell

Abstract Background: The present study investigated the effects of microvascular endothelial cells (MECs) on the chemotaxis, adhesion and proliferation of bone marrow hematopoietic stem cells (HSCs) ex vivo. Methods and Results: MECs were collected from the lung tissue of C57BL/6 mice, and HSCs were isolated with immunomagnetic beads from bone marrow of GFP mice. MECs and HSCs were co-cultured with or without having direct cell–cell contact in Transwell device for the measurement of chemotaxis and adhesion of MECs to HSCs. Experimental results indicate that the penetration rate of HSCs from the Transwell upper chamber to lower chamber in ‘co-culture’ group was significantly higher than that of ‘HSC single culture’ group. Also, the HSCs in co-culture group were all adherent at 24 h, and the co-culture group with direct cell–cell contact had highest proliferation rate. The HSC number was positively correlated with vascular endothelial growth factor (VEGF) and stromal cell-derived factor-1 (SDF-1) levels in supernatants of the culture. Conclusions: Our study reports that MECs enhance the chemotaxis, adhesion and proliferation of HSCs, which might be related to cytokines SDF-1 and VEGF secreted by MECs, and thus MECs enhance the HSC proliferation through cell–cell contact. The present study revealed the effect of MECs on HSCs, and provided a basis and direction for effective expansion of HSCs ex vivo.

Mouse Jagged2 is differentially expressed in hematopoietic progenitors and endothelial cells and promotes the survival and proliferation of hematopoietic progenitors by direct cell-to-cell contact

Blood ◽  
2000 ◽  
Vol 96 (3) ◽  
pp. 950-957 ◽  
Author(s):  
Schickwann Tsai ◽  
Jutta Fero ◽  
Steve Bartelmez
Keyword(s):  
Stem Cells ◽  
Endothelial Cells ◽  
Hematopoietic Stem Cells ◽  
Full Length ◽  
Differentially Expressed ◽  
Cell Contact ◽  
Proliferative Potential ◽  
Hematopoietic Progenitors ◽  
Hematopoietic Stem ◽  
Direct Cell

To study the regulation of the early stages of hematopoiesis, cDNA representational difference analysis was used to isolate genes that were differentially expressed in primitive hematopoietic progenitors. The reasoning was that such genes were more likely to provide functions important to hematopoietic stem cells and progenitors. One of the genes identified through this approach encodes mouse Jagged2(mJagged2). Using quantitative reverse transcription–polymerase chain reaction, it was shown that mJagged2 was differentially expressed in c-kit+ hematopoietic progenitors, including those with the phenotypes of Lin− c-kit+Rhlo Holo and Lin−c-kit+ Rhhi Holo, and that they have been shown to be highly enriched for long-term and short-term repopulating hematopoietic stem cells, respectively. Western blot analyses showed that endothelial cells also expressed high levels of Jagged2, but stromal fibroblasts did not. Using a coculture system we found that exogenous, full-length mJagged2 promoted the survival and proliferation of hematopoietic progenitors, including the high-proliferative potential colony-forming cells. Direct cell-to-cell contact was required for this effect. Taken together, these findings indicate that both c-kit+ hematopoietic progenitors and endothelial cells express Jagged2 and that exogenous, full-length Jagged2 promotes the survival and proliferation of hematopoietic progenitors.

Mouse Jagged2 is differentially expressed in hematopoietic progenitors and endothelial cells and promotes the survival and proliferation of hematopoietic progenitors by direct cell-to-cell contact

Blood ◽  
2000 ◽  
Vol 96 (3) ◽  
pp. 950-957 ◽  
Author(s):  
Schickwann Tsai ◽  
Jutta Fero ◽  
Steve Bartelmez
Keyword(s):  
Stem Cells ◽  
Endothelial Cells ◽  
Hematopoietic Stem Cells ◽  
Full Length ◽  
Differentially Expressed ◽  
Cell Contact ◽  
Proliferative Potential ◽  
Hematopoietic Progenitors ◽  
Hematopoietic Stem ◽  
Direct Cell

Abstract To study the regulation of the early stages of hematopoiesis, cDNA representational difference analysis was used to isolate genes that were differentially expressed in primitive hematopoietic progenitors. The reasoning was that such genes were more likely to provide functions important to hematopoietic stem cells and progenitors. One of the genes identified through this approach encodes mouse Jagged2(mJagged2). Using quantitative reverse transcription–polymerase chain reaction, it was shown that mJagged2 was differentially expressed in c-kit+ hematopoietic progenitors, including those with the phenotypes of Lin− c-kit+Rhlo Holo and Lin−c-kit+ Rhhi Holo, and that they have been shown to be highly enriched for long-term and short-term repopulating hematopoietic stem cells, respectively. Western blot analyses showed that endothelial cells also expressed high levels of Jagged2, but stromal fibroblasts did not. Using a coculture system we found that exogenous, full-length mJagged2 promoted the survival and proliferation of hematopoietic progenitors, including the high-proliferative potential colony-forming cells. Direct cell-to-cell contact was required for this effect. Taken together, these findings indicate that both c-kit+ hematopoietic progenitors and endothelial cells express Jagged2 and that exogenous, full-length Jagged2 promotes the survival and proliferation of hematopoietic progenitors.

The role of children's bone marrow mesenchymal stromal cells in the ex vivo expansion of autologous and allogeneic hematopoietic stem cells

2015 ◽  
Vol 39 (10) ◽  
pp. 1099-1110 ◽  
Author(s):  
Iordanis Pelagiadis ◽  
Eftichia Stiakaki ◽  
Christianna Choulaki ◽  
Maria Kalmanti ◽  
Helen Dimitriou
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Hematopoietic Stem

Hemangiogenic Role of ELF4 in Bone Marrow Post Myeloablation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1783-1783
Author(s):  
Mariela Sivina ◽  
Takeshi Yamada ◽  
Natalie Dang ◽  
H. Daniel Lacorazza
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Endothelial Cells ◽  
Hematopoietic Stem Cells ◽  
Blood Vessels ◽  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Bone Marrow Cells ◽  
Hematopoietic Stem ◽  
Huvec Cells

Abstract Bone marrow suppression is an important cause of death in patients exposed to radiation or in cancer patients treated with conventional chemotherapeutic agents. Myeloablative treatments (i.e. 5-fluorouracil administration) lead to apoptosis of blood forming cells and to regression of blood vessels in bone marrow. It is well known that hematological recovery post-bone marrow insult depends on the capacity of hematopoietic stem cells to regenerate the entire hematopoietic system, however, the transcriptional machinery involved in the regeneration of sinusoidal blood vessels in bone marrow from endothelial progenitor cells is largely unknown. Endothelial cells express the Tie2 receptor tyrosine kinase (a.k.a. Tek), which is involved in the angiogenic remodeling and vessel stabilization. Gene targeting of Tie2 showed that it is not required for differentiation and proliferation of definitive hematopoietic lineages in the embryo although Tie2 is needed during postnatal bone marrow hematopoiesis. ELF is a subgroup of the ETS family of transcription factors composed by ELF1, ELF2 (a.k.a. NERF), ELF3, ELF4 (a.k.a. MEF) and ELF5. ELF1 and ELF2 have been shown to regulate Tie2 expression in vitro. Recently we showed that ELF4 modulates the exit of hematopoietic stem cells (HSC) from quiescence (Lacorazza et al., Cancer Cell2006, 9:175–187). Given the high homology between ELF1 and ELF4 and the same origin of HSC and endothelial progenitor cells, we hypothesize that ELF4 regulates proliferation and Tie2 expression of endothelial cells. We used a luciferase gene reporter system in COS-7 and HEK cells to examine the capacity of ELF proteins to activate Tie2. ELF4 is the strongest activator of Tie2 expression following the hierarchy ELF4>ELF1>ELF2 variant 1>ELF2 variant 2. Site directed mutagenesis of each of the five ETS-binding sites (EBS) present in the Tie2 promoter shows that ELF4 binds preferentially to EBS 1, 3 and 5. Binding of ELF4 to the Tie2 promoter was confirmed by chromatin immunoprecipitation and EMSA. Although Elf1 gene expression is essentially normal in Elf4−/− bone marrow cells collected after 5-FU treatment, we detected diminished Tie2 expression compared to Elf4+/+ bone marrow cells. The association of this effect to human endothelial cells derived from umbilical cord (HUVEC cells) was investigated. All-trans retinoic acid (ATRA) and vascular-endothelial growth factor (VEGF) induced ELF4 expression in HUVEC cells in a dose and time dependent manner which was followed by increased Tie2 expression, suggesting that expression of ELF4 is modulated by angiogenic signals. Moreover, endothelial cells treated with ATRA showed rapid wound colonization in a wound assay. Expression of the pan-endothelial marker MECA-32 was determined by immunohistochemistry to correlate Tie2 with the regeneration of blood vessels: myeloablated Elf4−/− femurs exhibited a reduction of MECA-32 positive arterioles. Finally, temporal and spatial expression of Tie2 during hematological recovery post ablation was measured in bone marrow using transgenic Tie2-LacZ mice crossed to Elf4−/− mice. Collectively, our data suggests that ELF4 regulates Tie2 expression in endothelial cells but most importantly their proliferative capacity in response to angiogenic signals.

Non-Random Lentiviral Vector Insertions in Bone Marrow Progenitors from Fanconi Anemia Patients

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2358-2358
Author(s):  
Ali Nowrouzi ◽  
Africa Gonzales-Murillo ◽  
Anna Paruzynski ◽  
Ariana Jacome ◽  
Paula Rio ◽  
...  
Keyword(s):  
Stem Cells ◽  
Gene Therapy ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Fanconi Anemia ◽  
Large Scale ◽  
Random Distribution ◽  
Ex Vivo ◽  
Hematopoietic Stem ◽  
In Cis

Abstract Improved protocols using lentiviral vectors have been established with minimal cytokine exposure and short transduction times proving more suitable for overcoming the disease-specific challenge in correcting functionally defective hematopoietic stem cells (HSCs) of Fanconi Anemia (FA) patients. Bone marrow (BM) cells from FA patients were transduced ex vivo with lentiviral vectors (LVs) expressing FANCA and/or EGFP using optimized conditions to preserve the repopulating properties of the primitive hematopoietic stem cells (manuscript submitted). In a forward preclinical screening of possible LV-induced side effects we analyzed the insertional inventory in colonies generated by FA BM cells previously transduced with the LVs. We have established and optimized DNA and RNA isolation procedures for minimal cell numbers, suitable for large scale screening of colony forming cell (CFC) derived colonies by linear amplification-mediated PCR (LAM-PCR) and massive parallel pyrosequencing (454 GS Flx system; Roche). This approach is applicable for detecting early indicators of clonal selection, and is based on the analysis of common integration sites (CIS) and non-random distribution of vector insertions in particular genomic loci. From a total of 180 CFC-derived colonies expressing the EGFP LV marker gene, 298 vector insertions could be sequenced and mapped to the human genome. The analysis of vector targeted gene coding regions showed a non-random genomic distribution of LV insertions, with a significant overrepresentation of RefSeq genes that are part of distinct functional categories. Accordingly vector associated genes are predominantly involved in cellular signal cascades regulated by the MAP Kinase family known to be involved in a wide variety of cellular processes such as proliferation, differentiation, transcription regulation and development. Apart from the observed high integration frequency in genes (>80%), partial loss of vector LTR nucleotides was detected in >10% of the integrants (3–25bp). Notably, >20% of the lentiviral insertions were found to be located in CIS of predominantly 2nd order. Further screening assays of LV transduced CFC-derived colonies will allow a deeper investigation in the functional consequences of such CIS targeting in gene therapy protocols of FA. However our results suggest that the LV transduction of FA BM progenitors leads to a relatively high frequency of insertions in CIS which may be indicative of an insertion based (specific) selection mechanism. We herby show that the ex vivo large scale integration site analyses of CFC-derived colonies from patients considered to undergo gene therapeutic treatments constitutes a robust approach, which combined with mouse preclinical biosafety studies will help to improve the safety of clinical gene therapy protocols. The non-random distribution of LV integrations in CIS associated genes and in genes involved in particular cellular pathways may be indicative for the altered biochemical pathways characteristic of FA stem cells, with reported defects in DNA repair and self-renewal.

Hematopoietic Stem Cells Mediate the Repair of Damaged Endothelium

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 5458-5458
Author(s):  
Dana L. Pfaffle ◽  
Shuguang Jiang ◽  
Devorah C. Goldman ◽  
William H. Fleming
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Endothelial Cells ◽  
Portal Vein ◽  
Hematopoietic Stem Cells ◽  
Retention Capacity ◽  
Hematopoietic Stem ◽  
Chase Period ◽  
Portal Veins ◽  
Short Term Experiment

Abstract Recent studies indicate that vascular endothelium is an important component of the hematopoietic niche. As endothelial cells (ECs) are sensitive to radiation-induced damage, we evaluated the potential role of hematopoietic stem cells to enhance EC proliferation and repair. To test this hypothesis, lethally irradiated mice were transplanted with either 200–500 c-kit+, Sca+, lineage- (KSL) cells or an equivalent dose of unfractionated bone marrow (BM) cells (1×106 cells). Control groups included irradiated, non-transplanted, and non-irradiated, non-transplanted mice. Immediately after irradiation, all recipients were maintained on 0.8mg/ml Bromodeoxyuridine (BrdU) -containing water. Eleven days following irradiation, liver tissue was harvested and the fraction of proliferating BrdU+ ECs in the portal vein was assessed by immunostaining using both light and fluorescence microscopy. In irradiated, non-transplanted mice, 0.95% ± 0.17 SEM of portal vein ECs demonstrated the incorporation of BrdU. Transplantation of KSL cells increased the frequency of proliferating endothelial cells 2.5-fold to 2.5% ± 0.20 (p<0.0006). The transplantation of an equivalent number of unfractionated BM cells further increased the frequency of proliferating ECs by more than 3.5-fold (3.75% ± 0.21; p<0.0005). In non-transplanted, non-irradiated mice, BrdU+ ECs were detected at an intermediate level (2.30% ± 0.24) that is significantly higher than irradiated nontransplant recipients (p<0.006). To gain a better understanding of how hematopoietic stem cells (HSCs) influence the label retention capacity of ECs, we performed a BrdU pulse-chase experiment. Lethally irradiated mice were transplanted with 200 KSL cells, allowed 4 weeks for recovery, and then maintained on BrdU drinking water for 4 weeks. Consistent with our findings from the short term experiment described above, significantly more BrdU+ ECs were detected in the portal veins of KSL transplanted mice (15.36% ± 2.07) compared to those in non-transplanted, non-irradiated mice (8.68% ± 0.54; p<0.04) at the start of the chase period. During the first 24 weeks of the washout phase, BrdU+ ECs declined at a greater rate in the KSL recipients than in controls, indicating increased EC turnover. Interestingly, however, in both experimental groups, BrdU retention plateaued at 24 weeks and remained constant through 36 weeks. Taken together, our results indicate that radiation damage suppresses the incorporation of BrdU into ECs compared to steady state conditions and that this suppression can be reversed by the transplantation of either hematopoietic stem cells or unfractionated bone marrow. The extent to which BM derived ECs contribute to the proliferating EC pool will be addressed in future studies.

scholarly journals Dynamic roles of angiopoietin-like proteins 1, 2, 3, 4, 6 and 7 in the survival and enhancement of ex vivo expansion of bone-marrow hematopoietic stem cells

2013 ◽  
Vol 4 (3) ◽  
pp. 220-230 ◽  
Author(s):  
Shahina Akhter ◽  
Md. Mashiar Rahman ◽  
Hyun Seo Lee ◽  
Hyeon-Jin Kim ◽  
Seong-Tshool Hong
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Hematopoietic Stem
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