The HIF family member EPAS1/HIF-2α is required for normal hematopoiesis in mice

Blood ◽  
2003 ◽  
Vol 102 (5) ◽  
pp. 1634-1640 ◽  
Author(s):  
Marzia Scortegagna ◽  
Margaret A. Morris ◽  
Yavuz Oktay ◽  
Michael Bennett ◽  
Joseph A. Garcia
Keyword(s):  
Bone Marrow ◽  
Critical Role ◽  
Hypoxia Inducible Factor ◽  
Transcriptional Regulators ◽  
Hypoxic Stress ◽  
Pas Domain ◽  
Vascular Events ◽  
Family Function ◽  
Adult Mice

AbstractHypoxic stress plays a role in pathophysiologic states such as myocardial infarction and cerebral vascular events as well as in normal physiologic conditions including development and hematopoiesis. Members of the hypoxia inducible factor (HIF) family function as transcriptional regulators of genes involved in the hypoxic response. After generating adult mice that globally lack endothelial PAS domain protein 1 (EPAS1, also known as HIF-2α/HRF/HLF/MOP3), the second member of the HIF family, characterization of the hematopoietic cell population indicated that the loss of EPAS1/HIF-2α resulted in pancytopenia. Using bone marrow reconstitution experiments of lethally irradiated hosts, we have defined the extent and site of hematopoietic impairment in the EPAS1/HIF-2α null mice. These data suggest a critical role for EPAS1/HIF-2α in maintaining a functional microenvironment in the bone marrow for effective hematopoiesis.

Loss of Cxcl12/Sdf-1 in adult mice decreases the quiescent state of hematopoietic stem/progenitor cells and alters the pattern of hematopoietic regeneration after myelosuppression

Blood ◽  
2011 ◽  
Vol 117 (2) ◽  
pp. 429-439 ◽  
Author(s):  
Yi-Shiuan Tzeng ◽  
Hung Li ◽  
Yuan-Lin Kang ◽  
Wen-Cheng Chen ◽  
Wei-Cheng Cheng ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Adult Stage ◽  
Critical Role ◽  
Quiescent State ◽  
Hematopoietic Stem ◽  
Adult Mice ◽  
Conditional Deletion ◽  
Chemokine Cxcl12 ◽  
Stromal Cell Derived Factor ◽  
Quiescent Stem Cells

Abstract The C-X-C-type chemokine Cxcl12, also known as stromal cell–derived factor-1, plays a critical role in hematopoiesis during fetal development. However, the functional requirement of Cxcl12 in the adult hematopoietic stem/progenitor cell (HSPC) regulation was still unclear. In this report, we developed a murine Cxcl12 conditional deletion model in which the target gene can be deleted at the adult stage. We found that loss of stroma-secreted Cxcl12 in the adult led to expansion of the HSPC population as well as a reduction in long-term quiescent stem cells. In Cxcl12-deficient bone marrow, HSPCs were absent along the endosteal surface, and blood cell regeneration occurred predominantly in the perisinusoidal space after 5-fluorouracil myelosuppression challenge. Our results indicate that Cxcl12 is required for HSPC homeostasis regulation and is an important factor for osteoblastic niche organization in adult stage bone marrow.

scholarly journals The Coxiella burnetii QpH1 plasmid is a virulence factor for colonizing bone marrow-derived murine macrophages

2020 ◽  
Author(s):  
Shengdong Luo ◽  
Zhihui Sun ◽  
Huahao Fan ◽  
Shanshan Lu ◽  
Yan Hu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Coxiella Burnetii ◽  
Virulence Factor ◽  
Critical Role ◽  
Murine Macrophages ◽  
Infection Models

AbstractCoxiella burnetii carries a large conserved plasmid or plasmid-like chromosomally integrated sequence of unknown function. Here we report the curing of QpH1 plasmid from C. burnetii Nine Mile phase II, the characterization of QpH1-deficient C. burnetii in in vitro and in vivo infection models, and the characterization of plasmid biology. A shuttle vector pQGK, which is composed of the CBUA0036-0039a region (predicted for QpH1 maintenance), an E. coli plasmid ori, eGFP and kanamycin resistance genes was constructed. The pQGK vector can be stably transformed into Nine Mile II and maintained at a similar low copy like QpH1. Importantly, transformation with pQGK cured the endogenous QpH1 due to plasmid incompatibility. Compared to a Nine Mile II transformant of a RSF1010-based vector, the pQGK transformant shows an identical one-step growth curve in axenic media, a similar growth curve in Buffalo green monkey kidney cells, an evident growth defect in macrophage-like THP-1 cells, and dramatically reduced ability of colonizing bone marrow-derived murine macrophages. In the SCID mouse infection model, the pQGK transformants caused a lesser extent of splenomegaly. Moreover, the plasmid biology was investigated by mutagenesis. We found CBUA0037-0039 are essential for plasmid maintenance, and CBUA0037-0038 account for plasmid compatibility. Taken together, our data suggest that QpH1 encodes factor(s) essential for colonizing murine macrophages, and to a lesser extent for colonizing human macrophages. This study highlights a critical role of QpH1 for C. burnetii persistence in rodents, and expands the toolkit for genetic studies in C. burnetii.Author summaryIt is postulated that C. burnetii recently evolved from an inherited symbiont of ticks by the acquisition of novel virulence factors. All C. burnetii isolates carry a large plasmid or have a chromosomally integrated plasmid-like sequence. The plasmid is a candidate virulence factor that contributes to C. burnetii evolution. Here we describe the construction of novel shuttle vectors that allow to make plasmid-deficient C. burnetii mutants. With this plasmid-curing approach, we characterized the role of the QpH1 plasmid in in vitro and in vivo C. burnetii infection models. We found that the plasmid plays a critical role for C. burnetii growth in macrophages, especially in murine macrophages, but not in axenic media and BGMK cells. Our work highlights an essential role of the plasmid for the acquisition of colonizing capability in rodents by C. burnetii. This study represents a major step toward unravelling the mystery of the C. burnetii cryptic plasmids.

Hypoxia Induces Imatinib Resistance in CML Cell Lines.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4387-4387
Author(s):  
Daniela Buglio ◽  
Piera La Cava ◽  
Fabio Stagno ◽  
Annalisa Chiarenza ◽  
Salvatore Berretta ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Lines ◽  
Kinase Inhibitor ◽  
Critical Role ◽  
Hypoxia Inducible Factor ◽  
Leukemic Cell ◽  
Reversible Inhibitor ◽  
Imatinib Resistance ◽  
Downstream Target ◽  
Hypoxic Conditions

Abstract Background: Imatinib is a potent tyrosine kinase inhibitor that is highly effective in the treatment of chronic myeloid leukemia. However, some patients are resistant to imatinib. Some of the mechanisms leading to imatinib resistance include amplification of the BCR-ABL gene, determining overexpression of the protein, and mutations in the BCR-ABL protein with alteration of imatinib binding sites. Imatinib uptake is an active process mediated by a group of transporters that includes the organic cation transporters (hOCT) and it has been shown that different expression of OCT1 may play a critical role on intracellular drug levels and, hence, resistance to imatinib. Hypoxia is another important factor that may contribute to drug resistance. Hypoxia-Inducible Factor (HIF-1α) and its downstream target, Vascular endothelial Growth Factor (VEGF), have been shown to be overexpressed in leukemic bone marrow specimens compared to normal bone marrow. The purpose of this study is to determine if the hypoxic conditions and OCT1 inhibition affect imatinib sensitivity. Methods: Chronic myeloid leukemic cell line K562 and LAMA84 were cultured in 10% serum RPMI medium under hypoxic (3% O2) or normoxic (21% O2) conditions. All samples were treated with imatinib 1μM ± prazosin 13 μM (OCT1 reversible inhibitor) for 24 hs. Results: Cells treated with imatinib and cultured under hypoxic conditions demonstrated decreased apoptosis and increased cell viability compared to normoxic conditions (K562 Annexin + cells 62% vs 94%, p= 0.003, LAMA84 Annexin + cells 61% vs 92%, p=0.0028). The addition of prazosin almost abrogated imatinib efficacy in normoxic environment but did not modify the effect of imatinib under hypoxic conditions. Conclusions: Our data confirm that OCT1 is the most important imatinib carrier. Exposure of CML cell lines to an hypoxic environment results in reduced sensitivity to imatinib and this effect wasn’t affected by OCT1 inhibition. Search for underlying mechanisms of these findings are in progress.

scholarly journals Genome-scale analyses of transcriptional start sites in Mycobacterium marinum under normoxic and hypoxic conditions

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Shaojia Huang ◽  
Wei Zhou ◽  
Wei Tang ◽  
Yong Zhang ◽  
Yangbo Hu ◽  
...  
Keyword(s):  
Adaptive Response ◽  
Consensus Sequence ◽  
Critical Role ◽  
Transcriptional Level ◽  
Hypoxic Stress ◽  
Rna Seq ◽  
Hypoxic Conditions ◽  
Transcriptional Start Sites ◽  
Genome Scale

Abstract Background Hypoxic stress plays a critical role in the persistence of Mycobacterium tuberculosis (Mtb) infection, but the mechanisms underlying this adaptive response remain ill defined. Material and methods In this study, using M. marinum as a surrogate, we analyzed hypoxic responses at the transcriptional level by Cappable-seq and regular RNA-seq analyses. Results A total of 6808 transcriptional start sites (TSSs) were identified under normoxic and hypoxic conditions. Among these TSSs, 1112 were upregulated and 1265 were downregulated in response to hypoxic stress. Using SigE-recognized consensus sequence, we identified 59 SigE-dependent promoters and all were upregulated under hypoxic stress, suggesting an important role for SigE in this process. We also compared the performance of Cappable-seq and regular RNA-seq using the same RNA samples collected from normoxic and hypoxic conditions, and confirmed that Cappable-seq is a valuable approach for global transcriptional regulation analyses. Conclusions Our results provide insights and information for further characterization of responses to hypoxia in mycobacteria, and prove that Cappable-seq is a valuable approach for global transcriptional studies in mycobacteria.

scholarly journals Isolation and characterization of bone marrow-derived mesenchymal progenitor cells with myogenic and neuronal properties

2007 ◽  
Vol 313 (5) ◽  
pp. 1008-1023 ◽  
Author(s):  
Mitsutaka Shiota ◽  
Toshio Heike ◽  
Munetada Haruyama ◽  
Shiro Baba ◽  
Atsunori Tsuchiya ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Mesenchymal Progenitor Cells ◽  
Isolation And Characterization

scholarly journals Ultrafiltration Method for Plasma Protein Binding Studies and Its Limitations

Processes ◽  
2021 ◽  
Vol 9 (2) ◽  
pp. 382
Author(s):  
Camelia-Maria Toma ◽  
Silvia Imre ◽  
Camil-Eugen Vari ◽  
Daniela-Lucia Muntean ◽  
Amelia Tero-Vescan
Keyword(s):  
Protein Binding ◽  
Plasma Protein Binding ◽  
Plasma Protein ◽  
Specific Binding ◽  
Critical Role ◽  
Volume Ratio ◽  
Binding Studies ◽  
Experimental Conditions ◽  
Key Aspects

Plasma protein binding plays a critical role in drug therapy, being a key part in the characterization of any compound. Among other methods, this process is largely studied by ultrafiltration based on its advantages. However, the method also has some limitations that could negatively influence the experimental results. The aim of this study was to underline key aspects regarding the limitations of the ultrafiltration method, and the potential ways to overcome them. The main limitations are given by the non-specific binding of the substances, the effect of the volume ratio obtained, and the need of a rigorous control of the experimental conditions, especially pH and temperature. This review presents a variety of methods that can hypothetically reduce the limitations, and concludes that ultrafiltration remains a reliable method for the study of protein binding. However, the methodology of the study should be carefully chosen.

scholarly journals The Role of [18F]Fluciclovine PET/CT in the Characterization of High-Risk Primary Prostate Cancer: Comparison with [11C]Choline PET/CT and Histopathological Analysis

Cancers ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1575
Author(s):  
Lucia Zanoni ◽  
Riccardo Mei ◽  
Lorenzo Bianchi ◽  
Francesca Giunchi ◽  
Lorenzo Maltoni ◽  
...  
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Malignant Lesion ◽  
Lesion Detection ◽  
Histopathological Analysis ◽  
Choline Pet ◽  
Primary Prostate Cancer ◽  
Pet Ct

The primary aim of the study was to evaluate the role of [18F]Fluciclovine PET/CT in the characterization of intra-prostatic lesions in high-risk primary PCa patients eligible for radical prostatectomy, in comparison with conventional [11C]Choline PET/CT and validated by prostatectomy pathologic examination. Secondary aims were to determine the performance of PET semi-quantitative parameters (SUVmax; target-to-background ratios [TBRs], using abdominal aorta, bone marrow and liver as backgrounds) for malignant lesion detection (and best cut-off values) and to search predictive factors of malignancy. A six sextants prostate template was created and used by PET readers and pathologists for data comparison and validation. PET visual and semi-quantitative analyses were performed: for instance, patient-based, blinded to histopathology; subsequently lesion-based, un-blinded, according to the pathology reference template. Among 19 patients included (mean age 63 years, 89% high and 11% very-high-risk, mean PSA 9.15 ng/mL), 45 malignant and 31 benign lesions were found and 19 healthy areas were selected (n = 95). For both tracers, the location of the “blinded” prostate SUVmax matched with the lobe of the lesion with the highest pGS in 17/19 cases (89%). There was direct correlation between [18F]Fluciclovine uptake values and pISUP. Overall, lesion-based (n = 95), the performance of PET semiquantitative parameters, with either [18F]Fluciclovine or [11C]Choline, in detecting either malignant/ISUP2-5/ISUP4-5 PCa lesions, was moderate and similar (AUCs ≥ 0.70) but still inadequate (AUCs ≤ 0.81) as a standalone staging procedure. A [18F]Fluciclovine TBR-L3 ≥ 1.5 would depict a clinical significant lesion with a sensitivity and specificity of 85% and 68% respectively; whereas a SUVmax cut-off value of 4 would be able to identify a ISUP 4-5 lesion in all cases (sensitivity 100%), although with low specificity (52%). TBRs (especially with threshold significantly higher than aorta and slightly higher than bone marrow), may be complementary to implement malignancy targeting.

scholarly journals Self-Assembling Polypeptide Hydrogels as a Platform to Recapitulate the Tumor Microenvironment

Cancers ◽  
2021 ◽  
Vol 13 (13) ◽  
pp. 3286
Author(s):  
Dariusz Lachowski ◽  
Carlos Matellan ◽  
Ernesto Cortes ◽  
Alberto Saiani ◽  
Aline F. Miller ◽  
...  
Keyword(s):  
Cell Culture ◽  
Tumor Microenvironment ◽  
Cancer Cell ◽  
Critical Role ◽  
Hypoxia Inducible Factor ◽  
Pancreatic Cancer Cell Line ◽  
Cancer Cell Line ◽  
Culture Systems ◽  
A Cell

The tumor microenvironment plays a critical role in modulating cancer cell migration, metabolism, and malignancy, thus, highlighting the need to develop in vitro culture systems that can recapitulate its abnormal properties. While a variety of stiffness-tunable biomaterials, reviewed here, have been developed to mimic the rigidity of the tumor extracellular matrix, culture systems that can recapitulate the broader extracellular context of the tumor microenvironment (including pH and temperature) remain comparably unexplored, partially due to the difficulty in independently tuning these parameters. Here, we investigate a self-assembled polypeptide network hydrogel as a cell culture platform and demonstrate that the culture parameters, including the substrate stiffness, extracellular pH and temperature, can be independently controlled. We then use this biomaterial as a cell culture substrate to assess the effect of stiffness, pH and temperature on Suit2 cells, a pancreatic cancer cell line, and demonstrate that these microenvironmental factors can regulate two critical transcription factors in cancer: yes-associated protein 1 (YAP) and hypoxia inducible factor (HIF-1A).

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