scholarly journals Notch Ligand Jagged1 Is a Fetal Liver Niche Factor for the Function of Embryonic Hematopoietic Stem Cells

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 203-203
Author(s):  
Kostandin V. Pajcini ◽  
Lijian Shao ◽  
Na Yoon Paik ◽  
Kilian Sottoriva
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Endothelial Cells ◽  
Hematopoietic Stem Cells ◽  
Notch Signaling ◽  
Fetal Liver ◽  
Sequencing Analysis ◽  
Hematopoietic Stem ◽  
Cell Fate Decisions ◽  
Niche Factor

Abstract Embryonic hematopoietic stem cells (HSC) expand rapidly during development in the fetal liver. Notch1 is required for emergence of the definitive hematopoietic stem cells (HSCs) from the hemogenic endothelium, and is essential for survival and function of HSCs in the fetal liver. The identity of the ligand and the ligand-presenting cell during hematopoietic development would provide valuable information of the Notch signaling mechanism in HSCs as well as the identity of key niche cells that drive the expansion and cell fate decisions of embryonic HSCs. In the present study, we have taken a comprehensive approach to determine the ligands and cells that initiate Notch signaling in the mouse fetal liver. To this end, we have performed single-cell analysis for all Notch signaling proteins and many known targets in E14.5 fetal HSCs and adult bone marrow HSCs as well as fetal liver endothelial cells. We determined that Jagged1 (Jag1) is highly expressed in both endothelial cells as well as in fetal HSCs but not in adult HSCs. We have performed conditional loss-of-function analysis of Jag1 in fetal endothelial cells as well as in fetal hematopoietic lineages, where both myeloid and megakaryocytic progenitors are shown to express high levels of Jag1. Our results indicate that while loss of endothelial Jag1 has severe effects in embryonic vascular development, loss of hematopoietic Jag1 allows for normal fetal morphology, yet severely impedes the functional ability of fetal liver HSCs to expand and differentiate. RNA-Sequencing analysis of long-term fetal HSCs in Jag1-mutant embryos (VavCreJag f/f) revealed reduced expression of Gata2, Mllt3, Hoxa7, Angpt1 and IL-12a genes in fetal HSCs, which are well-known regulators of self-renewal and expansion. Our findings indicate that Jag1 is an essential niche factor for development of HSCs in the fetal liver and for functional potential of fetal HSCs once in the bone marrow microenvironment. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

scholarly journals Hematopoietic Jagged1 Is Required for the Transition of Hematopoietic Stem Cells from the Fetal Liver to the Adult Bone Marrow Niche

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 10-11
Author(s):  
Lijian Shao ◽  
Na Yoon Paik ◽  
Kostandin V. Pajcini
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Endothelial Cells ◽  
Hematopoietic Stem Cells ◽  
Notch Signaling ◽  
Fetal Liver ◽  
Hematopoietic Stem ◽  
Hematopoietic Development ◽  
Adult Bone Marrow ◽  
Adult Bone

Notch signaling is known to play important roles in hematopoietic development and differentiation. Notch1 is required for emergence of the definitive hematopoietic stem cells (HSCs) from the hemogenic endothelium, and we have previously shown that Notch signaling is essential for survival and function of HSCs in the fetal liver. Activation of canonical Notch signaling requires direct cellular contact; thus, the identity of the ligand and the ligand-presenting cell during hematopoietic development would provide valuable information of the Notch signaling mechanism in HSCs as well as the identity of key niche cells that drive the expansion and cell fate decisions of embryonic HSCs. In the present study, we have taken a comprehensive approach to determine the ligands and cells that initiate Notch signaling in the mouse fetal liver. To this end, we have performed single-cell PCR analysis for all Notch signaling proteins in E14.5 fetal HSCs and compared the findings to the adult bone marrow HSCs. We also have analyzed fetal liver endothelial cells for surface expression of all Notch ligands. We determined that Jagged1 (Jag1) is highly expressed in both endothelial cells as well as in fetal HSCs but not adult HSCs. We have performed conditional loss-of-function analysis of Jag1 in fetal endothelial cells using inducible Ve-cadherinCreERT2 as well as in fetal hematopoietic lineages using constitutive VavCre. Our results indicate that while loss of endothelial Jag1 has severe effects in embryonic vascular development, loss of hematopoietic Jag1 allows for normal fetal morphology, yet severely impedes the functional ability of fetal liver HSCs to expand and differentiate both in vitro and in vivo. Fetal to adult transplantation of VavCre+Jag1f/f HSCs indicated a defect in reconstitution potential of fetal HSCs that lack Jag1 expression. Our findings indicate that hematopoietic Jag1 is essential for maturation of HSCs in the fetal liver and for homing and reconstitution potential of HSCs into the post-natal bone marrow microenvironment. Disclosures No relevant conflicts of interest to declare.

BLOS2 maintains hematopoietic stem cells in the fetal liver via repressing Notch signaling

2017 ◽  
Vol 51 ◽  
pp. 1-6.e2 ◽  
Author(s):  
Qiuping He ◽  
Suwei Gao ◽  
Junhua Lv ◽  
Wei Li ◽  
Feng Liu
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Notch Signaling ◽  
Fetal Liver ◽  
Hematopoietic Stem

scholarly journals Molecular Modulation of Fetal Liver Hematopoietic Stem Cell Mobilization into Fetal Bone Marrow in Mice

2020 ◽  
Vol 2020 ◽  
pp. 1-12
Author(s):  
Huihong Zeng ◽  
Jiaoqi Cheng ◽  
Ying Fan ◽  
Yingying Luan ◽  
Juan Yang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Vascular Endothelial Cells ◽  
Fetal Liver ◽  
Bone Marrow Niche ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Fetal Bone

Development of hematopoietic stem cells is a complex process, which has been extensively investigated. Hematopoietic stem cells (HSCs) in mouse fetal liver are highly expanded to prepare for mobilization of HSCs into the fetal bone marrow. It is not completely known how the fetal liver niche regulates HSC expansion without loss of self-renewal ability. We reviewed current progress about the effects of fetal liver niche, chemokine, cytokine, and signaling pathways on HSC self-renewal, proliferation, and expansion. We discussed the molecular regulations of fetal HSC expansion in mouse and zebrafish. It is also unknown how HSCs from the fetal liver mobilize, circulate, and reside into the fetal bone marrow niche. We reviewed how extrinsic and intrinsic factors regulate mobilization of fetal liver HSCs into the fetal bone marrow, which provides tools to improve HSC engraftment efficiency during HSC transplantation. Understanding the regulation of fetal liver HSC mobilization into the fetal bone marrow will help us to design proper clinical therapeutic protocol for disease treatment like leukemia during pregnancy. We prospect that fetal cells, including hepatocytes and endothelial and hematopoietic cells, might regulate fetal liver HSC expansion. Components from vascular endothelial cells and bones might also modulate the lodging of fetal liver HSCs into the bone marrow. The current review holds great potential to deeply understand the molecular regulations of HSCs in the fetal liver and bone marrow in mammals, which will be helpful to efficiently expand HSCs in vitro.

Induction of Adipocyte Differentiation and Reduction of Hematopoesis-Supporting Activity of Mouse Stromal Cells by Suppression of Transcription Factor GATA-2.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1392-1392
Author(s):  
Yoko Okitsu ◽  
Hideo Harigae ◽  
Masanori Seki ◽  
Toru Fujiwara ◽  
Shinichiro Takahashi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Transcription Factor ◽  
Hematopoietic Stem Cells ◽  
Stromal Cells ◽  
Colony Formation ◽  
Adipocyte Differentiation ◽  
Fetal Liver ◽  
Expression Level ◽  
Hematopoietic Stem

Abstract (Introduction) Aplastic anemia (AA) is characterized by peripheral pancytopenia and fatty bone marrow. An immunological attack to hematopoietic stem cells has been thought to be responsible for the development of the disease. Previously, we reported the expression of transcription factor GATA-2 is significantly decreased in CD34 positive cells in AA. Together with the phenotypes of hematopoietic stem cells in GATA-2 hetero-knockout mice, GATA-2 down-regulation may play a role in the reduction of a stem cell pool observed in AA. On the other hand, GATA-2 has been shown to be essential for the maintenance of immaturity of preadipocytes. If a pathological immune response in AA decreases the level of GATA-2 expression in not only hematopoietic stem cells but also stromal preadipocytes, it may accelerate the maturation of preadipocytes, leading to the formation of fatty bone marrow. To explore this possibility, the phenotypic change of stromal preadipocytes by suppression of GATA-2 was examined in this study. (Method) The GATA-2 expression level was suppressed by using siRNA for GATA-2 in mouse stromal preadipocyte cell lines, TBR9 and TBR343. After the treatment with siRNA, the adipocyte differentiation was induced by the incubation with insulin and dexamethasone for 7days. Then, the maturation level was examined by oil drops formation judged by oil red staining, and by the expression level of adipcin and PPAR-γ mRNA. Supporting activity of hematopoietic colony formation was also evaluated by using mouse fetal liver cells after siRNA treatment. (Results) By using designed siRNA, the GATA-2 expression was suppressed to 30% of control, whereas the expression level of GATA-3, which is co-expressed in preadipocytes, was unchanged. When GATA-2 was suppressed by siRNA, the oil drop formation and adipocyte-specific gene expression was significantly accelerated in both of stromal cells. Furthermore, the number of fetal liver hematopoetic colonies was significantly decreased by suppression of GATA-2, suggesting that GATA-2 down-regulation in stromal preadipocytes results in not only the acceleration of the maturation but also the reduced supporting activity of hematopoietic colony formation (Conclusion) These results suggest that suppression of GATA-2 in hematopoietic tissues induces the characteristic features of AA, i.e., decreased the number of hematopoietic stem cells and increased number of mature adipocytes.

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.

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.

In Utero Depletion Of Fetal Host Hematopoietic Stem Cells Improves Engraftment Following Neonatal Transplantation In Mice

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3240-3240
Author(s):  
Chris Derderian ◽  
Charmin King ◽  
Priya Togarrati ◽  
Agnieszka Czechowicz ◽  
Ninnia Lescano ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Peripheral Blood ◽  
Hematopoietic Cell Transplantation ◽  
Mononuclear Cells ◽  
Fetal Liver ◽  
In Utero ◽  
Hematopoietic Stem ◽  
In Utero Transplantation

Abstract Introduction In utero hematopoietic cell transplantation (IUHCTx) is a promising strategy to treat congenital disorders as the fetal host can potentially be tolerized to transplanted cells early in gestation. However, levels of engraftment have been low and fetal host conditioning strategies to increase space in hematopoietic niches have not been widely explored. We hypothesized that depletion of fetal host hematopoietic stem cells (HSC) using an antibody against the c-kit receptor (ACK2), a strategy which selectively depletes HSC by disrupting stem cell factor (SCF) signaling, would improve engraftment after HSC transplantation. Methods Fetal C57B6.CD45.2 (B6) mice were injected with increasing doses of ACK2 (2.5-50 µg/fetus) or isotype control antibody on E14.5 and surviving pups were transplanted with congenic B6.CD45.1 fetal liver mononuclear cells (2.5×106 cells/pup) on day of life 1 (P1, 7 days after in utero injection), allowing post-transplantation host monitoring. Host HSC depletion and residual serum ACK2 concentration were examined on P1. Peripheral blood chimerism, defined as donor/(donor+host) CD45 cells, as well as the lineage distribution of chimeric cells, were determined beginning 4 weeks after transplantation. Results Survival to birth among fetuses injected with 2.5, 5, or 10 µg of ACK2 was similar to controls (control: 74%; 2.5 µg: 80%; 5 µg: 71%; 10 µg: 60%, p=0.2 by chi-square test, n≥45/group) but was significantly lower at higher concentrations (20 µg: 37%; 50 µg: 31%, p<0.001 vs. control, n≥70/group). Transient anemia and leukopenia were observed on P1 with doses ≥ 5 µg which resolved by P7 (n=17). Four of 19 pups previously treated with ACK2 (2.5-10 µg) and observed long-term had patchy coat discoloration, possibly a manifestation of disruption of C-kit+ melanocyte migration. In utero ACK2 treatment resulted in significant and dose-dependent depletion of host HSCs (defined as Lin-Sca-1+C-kit+, KLS) in the bone marrow of treated animals by P1 (Figure 1A). There was no depletion of KLS cells in the liver. Residual ACK2 antibody was undetectable in the serum by P1, validating our strategy of in utero depletion and neonatal transplantation. In animals receiving neonatal transplantation, ACK2 depletion resulted in a significant increase in levels of engraftment 4 weeks after transplantation compared to controls (control: 3.3±0.3%; 2.5 µg: 13±1.4%; 5 µg: 10±2.4%; 10 µg: 11±2.0%, p<0.05 for each dose vs control by ANOVA). Accordingly, we detected an increased number total bone marrow KLS cells 7 days after transplantation in ACK2 treated animals compared to controls (412±45.9 vs. 933±112 cells, p=0.01, n≥3/group). Moreover, levels of chimerism increased over time in treated animals (Figure 1B; 12 weeks: 2.5 µg: 190%; 5 µg: 170%; 10 µg: 160%) while they remained unchanged in controls. Overall, levels of chimerism achieved with ACK2 treatment were significantly higher than that observed in animals that received in utero transplantation without ACK2 depletion. Lineage analysis of peripheral blood for granulocytes, B cells, and T cells indicated an equal increase in all lineages, suggesting ACK2 depletes true HSCs and not committed progenitors. Interestingly, ACK2 depletion at doses 2.5-10 µg did not result in engraftment of allogeneic BALB/c cells (n=11), indicating that allogeneic neonatal transplantation, unlike in utero transplantation, is limited by a host immune response which is unaffected by ACK2. Conclusion We have demonstrated that fetal HSC depletion using ACK2 can lead to clinically relevant levels of donor cell engraftment with minimal toxicity. In previous studies with this antibody, host HSC depletion required either immunodeficient animals or concurrent irradiation, whereas we achieved depletion in wild-type fetal hosts, suggesting differences in fetal vs. adult HSC sensitivity to SCF signaling. Future studies should explore this strategy to improve engraftment in large animals models of IUHCTx. Disclosures: Weissman: Amgen, Systemix, Stem cells Inc, Cellerant: Consultancy, Employment, Equity Ownership, Membership on an entity’s Board of Directors or advisory committees.

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