Altered Functions of Neutrophils in Two Chinese Patients With Severe Congenital Neutropenia Type 4 Caused by G6PC3 Mutations

BackgroundSCN4 is an autosomal recessive disease caused by mutations in the G6PC3 gene. The clinical, molecular, and immunological features; function of neutrophils; and prognosis of patients with SCN4 have not been fully elucidated.MethodsTwo Chinese pediatric patients with G6PC3 mutations were enrolled in this study. Clinical data, genetic and immunologic characteristics, and neutrophil function were evaluated in patients and controls before and after granulocyte colony-stimulating factor (G-CSF) treatment.ResultsBoth patients had histories of pneumonia, inguinal hernia, cryptorchidism, and recurrent oral ulcers. Patient 1 also had asthma and otitis media, and patient 2 presented with prominent ectatic superficial veins and inflammatory bowel disease. DNA sequencing demonstrated that both patients harbored heterozygous G6PC3 gene mutations. Spontaneous and FAS-induced neutrophil apoptosis were significantly increased in patients, and improved only slightly after G-CSF treatment, while neutrophil respiratory burst and neutrophil extracellular traps production remained impaired in patients after G-CSF treatment.ConclusionG-CSF treatment is insufficient for patients with SCN4 patients, who remain at risk of infection. Where possible, regular G-CSF treatment, long-term prevention of infection, are the optimal methods for cure of SCN4 patients. It is important to monitor closely for signs of leukemia in SCN4 patients. Once leukemia occurs in SCN4 patients, hematopoietic stem cell transplantation is the most important choice of treatment.

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The chemokine GROβ mobilizes early hematopoietic stem cells characterized by enhanced homing and engraftment

Blood ◽

10.1182/blood-2006-06-031401 ◽

2007 ◽

Vol 110 (3) ◽

pp. 860-869 ◽

Cited By ~ 62

Author(s):

Seiji Fukuda ◽

Huimin Bian ◽

Andrew G. King ◽

Louis M. Pelus

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Hematopoietic Stem ◽

Platelet Recovery ◽

And Function ◽

Stimulating Factor ◽

Cxcr4 Axis

Abstract Mobilized peripheral blood hematopoietic stem cells (PBSCs) demonstrate accelerated engraftment compared with bone marrow; however, mechanisms responsible for enhanced engraftment remain unknown. PBSCs mobilized by GROβ (GROβΔ4/CXCL2Δ4) or the combination of GROβΔ4 plus granulocyte colony-stimulating factor (G-CSF) restore neutrophil and platelet recovery faster than G-CSF–mobilized PBSCs. To determine mechanisms responsible for faster hematopoietic recovery, we characterized immunophenotype and function of the GROβ-mobilized grafts. PBSCs mobilized by GROβΔ4 alone or with G-CSF contained significantly more Sca-1+-c-kit+-lineage− (SKL) cells and more primitive CD34−-SKL cells compared with cells mobilized by G-CSF and demonstrated superior competitive long-term repopulation activity, which continued to increase in secondary and tertiary recipients. GROβΔ4-mobilized SKL cells adhered better to VCAM-1+ endothelial cells compared with G-CSF–mobilized cells. GROβΔ4-mobilized PBSCs did not migrate well to the chemokine stromal derived factor (SDF)-1α in vitro that was associated with higher CD26 expression. However, GROβΔ4-mobilized SKL and c-Kit+ lineage− (KL) cells homed more efficiently to marrow in vivo, which was not affected by selective CXCR4 and CD26 antagonists. These data suggest that GROβΔ4-mobilized PBSCs are superior in reconstituting long-term hematopoiesis, which results from differential mobilization of early stem cells with enhanced homing and long-term repopulating capacity. In addition, homing and engraftment of GROβΔ4-mobilized cells is less dependent on the SDF-1α/CXCR4 axis.

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Enrichment of human hematopoietic stem cell activity in the CD34+Thy- 1+Lin- subpopulation from mobilized peripheral blood

Blood ◽

10.1182/blood.v85.2.368.368 ◽

1995 ◽

Vol 85 (2) ◽

pp. 368-378 ◽

Cited By ~ 136

Author(s):

L Murray ◽

B Chen ◽

A Galy ◽

S Chen ◽

R Tushinski ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Population ◽

Peripheral Blood ◽

Colony Stimulating Factor ◽

In Vivo Models ◽

Hematopoietic Stem ◽

Stimulating Factor ◽

Mobilized Peripheral Blood

Abstract The number of CD34+ cells in the peripheral blood of cancer patients is known to be increased following the administration of high dose chemotherapy and hematopoietic growth factors. These so-called peripheral blood stem cell grafts are now frequently used for autologous transplantation of patients with malignancies. In this report, we address the question of whether true long-term repopulating pluripotent hematopoietic stem cells (PHSC) are mobilized into peripheral blood following chemotherapy plus granulocyte/macrophage colony-stimulating factor (GM-CSF) or granulocyte colony-stimulating factor (G-CSF) mobilization. We have examined the presence of stem cells in mobilized peripheral blood (MPB) by using an antibody to the human Thy-1 molecule to stain the CD34+Lineage- (Lin-) population. The kinetics of mobilization of CD34+Thy-1+ Lin- cells into peripheral blood were studied, and the percentage of cells with this phenotype was found to vary widely depending on the day of leukapheresis. A CD34+Thy- 1+Lin- cell population, potentially containing PHSCs, was isolated by fluorescence activated cell sorting (FACS) and analyzed for activity. The multilineage differentiative capacity of this candidate stem cell- containing population in MPB was determined using an in vitro long-term culture system, in which cobblestone area formation was used as a means of detecting PHSCs. We also measured repopulating capacity by using two in vivo models in which severe combined immunodeficiency (SCID)-hu mice were implanted with human fetal bone or thymus grafts. Using these assays, we show that the highest frequency of cobblestone area-forming cells (CAFC) after 7 weeks of culture was observed in a subpopulation of CD34+Lin- cells, which expressed low levels of Thy-1. This cell population was capable of producing both B and myeloid cells, and maintaining CD34+Lin- cells in these long term cultures. Moreover, the CD34+Thy-1+Lin- cell subset possessed a higher ability to engraft and to demonstrate multilineage differentiative potential at 8 weeks in the SCID-hu bone assay. However, in the SCID-hu thymus model, both Thy-1+ and Thy-1- subpopulations were capable of donor T-cell engraftment at 6 weeks, suggesting the presence of cells capable of initiating T lymphopoiesis in both populations.(ABSTRACT TRUNCATED AT 400 WORDS)

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Alternative mechanisms with and without steel factor support primitive human hematopoiesis

Blood ◽

10.1182/blood.v81.6.1465.1465 ◽

1993 ◽

Vol 81 (6) ◽

pp. 1465-1470 ◽

Cited By ~ 56

Author(s):

HJ Sutherland ◽

DE Hogge ◽

D Cook ◽

CJ Eaves

Keyword(s):

Hematopoietic Stem ◽

Steel Factor ◽

Stem Cell Maintenance ◽

Murine Fibroblasts ◽

Normal Human ◽

Clonogenic Cell ◽

First Time ◽

Stimulating Factor

Abstract As a first approach to defining the molecular requirements for supporting human hematopoietic stem cell maintenance and differentiation in vitro, we have analyzed and compared the ability of various factors to support the maintenance and initial differentiation of human long-term culture-initiating cells (LTC-ICs), a distinct, rare primitive hematopoietic cell type whose progeny after 5 weeks include cells detectable as colony-forming cells. Normal human marrow cells highly enriched in LTC-ICs (approximately 1% pure) were placed in cultures containing either preestablished, irradiated human marrow adherent feeder layers, or feeders consisting of Steel factor-deficient SI/SI, or normal +/+ murine fibroblasts, or no feeders. In some groups, either Steel factor alone, granulocyte colony-stimulating factor (G- CSF) and interleukin-3 (IL-3), or all three factors combined were also added repeatedly. SI/SI murine fibroblasts were equivalent to +/+ controls and to normal human marrow feeders in supporting both LTC-IC maintenance and clonogenic cell output over a 5-week period. Soluble Steel factor alone could, however, effectively substitute for human marrow feeders to support LTC-IC maintenance, although clonogenic cell output was markedly reduced under these conditions. Conversely, soluble Steel factor with G-CSF and IL-3 or with feeders (or all together) did not further enhance (or depress) LTC-IC maintenance, although under these conditions clonogenic cell output was markedly increased. These findings confirm previous evidence that LTC-IC maintenance and clonogenic cell production are differentially regulated and show for the first time that LTC-IC maintenance can be supported by different nonsynergizing factors that may, but need not, include Steel factor.

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Long-term preoperative glycemic control restored the perioperative neutrophilic phagocytosis activity in diabetic mice

BMC Endocrine Disorders ◽

10.1186/s12902-020-00629-x ◽

2020 ◽

Vol 20 (1) ◽

Author(s):

Daichi Fujimoto ◽

Yuki Nomura ◽

Moritoki Egi ◽

Norihiko Obata ◽

Satoshi Mizobuchi

Keyword(s):

Glycemic Control ◽

Insulin Therapy ◽

Neutrophil Function ◽

Ros Production ◽

Diabetic Mice ◽

Short Term ◽

Chronic Hyperglycemia ◽

Before And After ◽

Phagocytosis Activity

Abstract Background The risk of surgical site infection has been reported to be higher in patients with poorly controlled diabetes. Since chronic hyperglycemia impairs neutrophil functions, preoperative glycemic control may restore neutrophil function. However, long-term insulin therapy may lead to a delay in surgery, which may be a problem, especially in cancer surgery. It is therefore unfortunate that there have been few studies in which the optimal duration of perioperative glycemic control for diabetes with chronic hyperglycemia was investigated. Therefore, we investigated the effects of preoperative long-term insulin therapy and short-term insulin therapy on perioperative neutrophil functions in diabetic mice with chronic hyperglycemia. Methods Five-week-old male C57BL/6 J mice were divided into four groups (No insulin (Diabetes Mellitus: DM), Short-term insulin (DM), Long-term insulin (DM), and Non-diabetic groups). Diabetes was established by administrating repeated low-dose streptozotocin. The Short-term insulin (DM) group received insulin therapy for 6 h before the operation and the Long-term insulin (DM) group received insulin therapy for 5 days before the operation. The No insulin (DM) group and the Non-diabetic group did not receive insulin therapy. At 14 weeks of age, abdominal surgery with intestinal manipulation was performed in all four groups. We carried out a phagocytosis assay with fluorescent microspheres and a reactive oxygen species (ROS) production assay with DCFH-DA (2′,7′-dichlorodihydrofluorescein diacetate) before and 24 h after the operation using FACSVerse™ with BD FACSuite™ software. Results Blood glucose was lowered by insulin therapy in the Short-term insulin (DM) and Long-term insulin (DM) groups before the operation. Neutrophilic phagocytosis activities before and after the operation were significantly restored in the Long-term insulin (DM) group compared with those in the No insulin (DM) group (before: p = 0.0008, after: p = 0.0005). However, they were not significantly restored in the Short-term insulin (DM) group. Neutrophilic ROS production activities before and after the operation were not restored in either the Short-term insulin (DM) group or Long-term insulin (DM) group. Conclusions Preoperative and postoperative phagocytosis activities are restored by insulin therapy for 5 days before the operation but not by insulin therapy for 6 h before the operation.

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Partially differentiated ex vivo expanded cells accelerate hematologic recovery in myeloablated mice transplanted with highly enriched long- term repopulating stem cells

Blood ◽

10.1182/blood.v88.9.3642.bloodjournal8893642 ◽

1996 ◽

Vol 88 (9) ◽

pp. 3642-3653 ◽

Cited By ~ 41

Author(s):

SJ Szilvassy ◽

KP Weller ◽

B Chen ◽

CA Juttner ◽

A Tsukamoto ◽

...

Keyword(s):

Stem Cells ◽

Blood Cells ◽

Ex Vivo ◽

Granulocyte Colony Stimulating Factor ◽

Short Term ◽

Hematopoietic Stem ◽

Colony Forming Units ◽

Sustained Recovery ◽

Stimulating Factor

The ability of an infusion of ex vivo expanded hematopoietic cells to ameliorate cytopenia following transplantation of hematopoietic stem cells (HSCs) is controversial. To address this issue, we measured the recovery of circulating leukocytes, erythrocytes, and platelets in lethally irradiated mice transplanted with 10(3) enriched HSCs, with or without their expanded equivalent (EE) generated after 7 days of culture in interleukin-3 (IL-3), IL-6, granulocyte colony-stimulating factor and Steel Factor. Two HSC populations differing in their content of short-term repopulating progenitors were evaluated. Thy-1loLIN-Sca- 1+ (TLS) bone marrow (BM) is enriched in colony-forming cells (CFCs), day 8 and day 12 spleen colony-forming units (CFU-S) (435 +/- 19, 170 +/- 30, and 740 +/- 70 per 10(3) cells, respectively), and stem cells with competitive long-term repopulating potential (> or = 1 per 43 cells). Thy-1loSca-1+H-2Khl cells (TSHFU) isolated from BM 1 day after treatment of donor mice with 5-fluorouracil (5-FU) are also highly enriched in competitive repopulating units (CRU, > or = 1 per 55 cells), but are depleted of CFCs, day 8 and day 12 CFU-S (171 +/- 8, 0 and 15 +/- 4 per 10(3) cells, respectively). Recipients of 10(3) TLS cells transiently recovered leukocytes to > or = 2,000/microL in 12 days, but sustained engraftment required 25 days. Platelets recovered to > or = 200,000/microL in 15 days, and erythrocytes never decreased below 50% of normal. Mice transplanted with 10(3) TSHFU cells recovered leukocytes in 15 days, and platelets and erythrocytes in 18 days. Recipients of unseparated normal or 5-FU-treated BM cells (containing 10(3) TLS or TSHFU cells) recovered safe levels of blood cells in 9 to 12 days, suggesting that unseparated marrow contains early engrafting cells that were depleted by sorting. Upon ex vivo expansion, total cells, CFCs and day 12 CFU-S were amplified 2,062-,83- and 13-fold, respectively, from TLS cells; and 1,279-, 259- and 708-fold, respectively, from TSHFU cells. Expanded cells could regenerate the majority of lymphocytes and granulocytes in primary (17 weeks) and secondary (26 weeks) hosts and were only moderately impaired compared to fresh HSCs. The EE of TSHFU cells was more potent than that of TLS cells, suggesting that more highly enriched HSCs are more desirable starting populations for this application. When mice were transplanted with 10(3) TSHFU cells and their EE, the duration of thrombocytopenia was shortened from 18 to 12 days, and anemia was abolished. Leukocytes were also elevated on days 9 to 12, although sustained recovery was not accelerated. Anemia was also abrogated in recipients of 10(3) TLS cells and their EE. Early platelet counts were slightly higher than with TLS cells alone, but leukocyte recovery was not improved. These data confirm that TLS cells contribute to early and sustained hematopoiesis, and demonstrate a benefit of ex vivo expanded cells in accelerating engraftment of more primitive TSHFU stem cells depleted of progenitors.

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Clinical phenotypes and genotypic characteristics of infantile-onset inflammatory bowel disease with interleukin-10 receptor A mutations in 22 Chinese children

10.21203/rs.3.rs-34174/v2 ◽

2020 ◽

Author(s):

Dexiu Guan ◽

Jing Zhang ◽

Shu Guo ◽

Feihong Yu ◽

Jin Zhou ◽

...

Keyword(s):

Inflammatory Bowel Disease ◽

Mortality Rate ◽

Bowel Disease ◽

Early Onset ◽

Gene Mutations ◽

Interleukin 10 ◽

Clinical Phenotypes ◽

Oral Ulcers ◽

Inflammatory Bowel ◽

Genotypic Characteristics

Abstract Background : Infantile-onset inflammatory bowel disease (IO-IBD) patients with interleukin-10/interleukin-10 receptor (IL-10/IL-10R) mutations suffer from more severe and intractable disease. We aimed to investigate the clinical phenotypes and genotypic characteristics of IO-IBD patients with IL-10RA gene mutations. Methods : Data on 22 patients with IO-IBD with IL-10RA gene mutations were retrospectively analyzed, and high-throughput sequencing was used to identify the IL-10RA gene mutations. Results : In 22 patients with IO-IBD with IL-10RA mutations, c.C301T (p.R101W) (86.4%, 19/22) and c.G537A (p.T179T) (36.4%, 8/22) were the most common ones, and one novel mutation was identified (c.635G>C (p.R212P)). Patients had extremely early onset of symptoms, with 81.8% (18/22) having disease onset within 1 month after birth, and median onset time was 8.5 (interquartile range: 3.0–24.0) days. In addition, 77.3% (17/22) of patients had recurrent perianal lesions. Oral ulcers and skin rashes were common extra-intestinal manifestations, accounting for 72.7% (16/22) and 63.6% (14/22), respectively. In this study, three patients underwent enterostomy and one experienced intestinal perforation repair. Thalidomide was used in five patients in this study: one achieved clinical remission, three were clinical improvement and one still had disease activity. Two patients underwent umbilical cord blood transplantation (UCBT) and remained stable. Follow-up showed that the mortality rate was as high as 45% (9/20). Conclusions : In 22 IO-IBD patients with IL-10RA mutations, the most common mutations were c.C301T (p.R101W) and c.G537A (p.T179T). Patient characteristics included extremely early onset of symptoms and extra-intestinal manifestations such as recurrent perianal lesions, oral ulcers, and skin rashes, which were common. UCBT and thalidomide might be effective treatments, although the mortality rate was high in this study.

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Rapid and Efficient Mobilization of Murine Hematopoietic Stem and Progenitor Cells with Nox-A12, a New Spiegelmers®-Based CXCR4/SDF-1(CXCL12) Antagonist

Blood ◽

10.1182/blood.v118.21.2995.2995 ◽

2011 ◽

Vol 118 (21) ◽

pp. 2995-2995 ◽

Cited By ~ 1

Author(s):

Marina Scheller ◽

Frank Schwoebel ◽

Doerte Vossmeyer ◽

Achim Leutz

Keyword(s):

Progenitor Cells ◽

Conflicts Of Interest ◽

Mirror Image ◽

Hematopoietic Stem ◽

Combined Administration ◽

Stem And Progenitor Cells ◽

Clinical Potential ◽

Progenitor Compartment ◽

Stimulating Factor

Abstract Abstract 2995 Mobilization of hematopoietic stem cells (HSCs) and progenitor cells (HPCs) is important in many hematological therapies. However, up to 30% of the patients respond poorly to standard granulocyte colony-stimulating factor (G-CSF) treatment, highlighting the need for more effective mobilizing strategies. The CXCR4/stromalcell-derived factor 1 (SDF-1) axis plays a crucial role in the interaction between HSCs and the marrow niche and is involved in HSC mobilization. NOX-A12 is a structured mirror-image RNA oligonucleotide, a so-called Spiegelmer®, that was identified to bind SDF-1 thereby inhibiting its activity with subnanomolar IC50. HSC/HPC mobilization by NOX-A12 was examined in the mouse. Single NOX-A12 administration induced reversible mobilization of HSC/HPC populations within a few hours. NOX-A12 synergized with G-CSF to strongly enhance HSC/HPC mobilization. In particular, the progenitor compartment mobilized by single NOX-A12 administration contained more differentiated short-term HSCs (ST-HSCs), and combined administration of NOX-A12 and G-CSF mobilized a significantly higher proportion of primitive and more potent murine long-term repopulating cells that successfully engrafted primary and secondary lethally-irradiated recipients. These results characterize NOX-A12 as a potent HSCs/HPCs mobilizing therapeutic in mammals and suggest its clinical potential. Disclosures: No relevant conflicts of interest to declare.

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Long-term support of hematopoiesis by a single stem cell clone in patients with paroxysmal nocturnal hemoglobinuria

Blood ◽

10.1182/blood.v99.8.2748 ◽

2002 ◽

Vol 99 (8) ◽

pp. 2748-2751 ◽

Cited By ~ 27

Author(s):

Jun-ichi Nishimura ◽

Toshiyuki Hirota ◽

Yuzuru Kanakura ◽

Takashi Machii ◽

Takashi Kageyama ◽

...

Keyword(s):

Stem Cell ◽

Paroxysmal Nocturnal Hemoglobinuria ◽

Somatic Mutations ◽

Cell Clone ◽

Gene Mutations ◽

Polymorphonuclear Cells ◽

Hematopoietic Stem ◽

Cell Disorder ◽

Pnh Clone

Abstract Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired hematopoietic stem cell disorder characterized by clonal blood cells that are deficient in glycosylphosphatidylinositol-anchored proteins because of somatic mutations of the PIG-A gene. Many patients with PNH have more than one PNH clone, but it is unclear whether a single PNH clone remains dominant or minor clones eventually become dominant. Furthermore, it is unknown how many hematopoietic stem cells (HSCs) sustain hematopoiesis and how long a single HSC can support hematopoiesis in humans. To understand dynamics of HSCs, we reanalyzed the PIG-A gene mutations in 9 patients 6 to 10 years after the previous analyses. The proportion of affected peripheral blood polymorphonuclear cells (PMNs) in each patient was highly variable; it increased in 2 (from 50% and 65% to 98% and 97%, respectively), was stable in 4 (changed less than 20%), and diminished in 3 (94%, 99%, and 98% to 33%, 57%, and 43%, respectively) patients. The complexity of these results reflects the high variability of the clinical course of PNH. In all patients, the previously predominant clone was still present and dominant. Therefore, one stem cell clone can sustain hematopoiesis for 6 to 10 years in patients with PNH. Two patients whose affected PMNs decreased because of a decline of the predominant PNH clone and who have been followed up for 24 and 31 years now have an aplastic condition, suggesting that aplasia is a terminal feature of PNH.

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Characterization of primitive hematopoietic cells from patients with dyskeratosis congenita

Blood ◽

10.1182/blood-2007-10-120204 ◽

2008 ◽

Vol 111 (9) ◽

pp. 4523-4531 ◽

Cited By ~ 34

Author(s):

Frederick D. Goldman ◽

Geraldine Aubert ◽

Al J. Klingelhutz ◽

Mark Hills ◽

Sarah R. Cooper ◽

...

Keyword(s):

Dyskeratosis Congenita ◽

Normal Numbers ◽

Hematopoietic Stem ◽

Cd34 Cells ◽

Normal Differentiation ◽

Stimulating Factor ◽

Mobilized Peripheral Blood

Abstract Dyskeratosis congenita (DC) is an inherited bone marrow (BM) failure syndrome associated with mutations in telomerase genes and the acquisition of shortened telomeres in blood cells. To investigate the basis of the compromised hematopoiesis seen in DC, we analyzed cells from granulocyte colony-stimulating factor mobilized peripheral blood (mPB) collections from 5 members of a family with autosomal dominant DC with a hTERC mutation. Premobilization BM samples were hypocellular, and percentages of CD34+ cells in marrow and mPB collections were significantly below values for age-matched controls in 4 DC subjects. Directly clonogenic cells, although present at normal frequencies within the CD34+ subset, were therefore absolutely decreased. In contrast, even the frequency of long-term culture-initiating cells within the CD34+ DC mPB cells was decreased, and the telomere lengths of these cells were also markedly reduced. Nevertheless, the different lineages of mature cells were produced in normal numbers in vitro. These results suggest that marrow failure in DC is caused by a reduction in the ability of hematopoietic stem cells to sustain their numbers due to telomere impairment rather than a qualitative defect in their commitment to specific lineages or in the ability of their lineage-restricted progeny to execute normal differentiation programs.

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