GATA2 variants in patients with non-tuberculous mycobacterial or fungal infections without known immunodeficiencies

ABSTRACT Introduction Haploinsufficiency of the hematopoietic transcription factor GATA2 is associated with a broad spectrum of diseases, including infection susceptibility and neoplasms. We aimed to investigate GATA2 variants in patients with non-tuberculous mycobacterial (NTM) and/or fungal infections (FI) without known immunodeficiencies. Method We performed GATA2 genotyping in patients with NTM and/or FI. Results Twenty-two patients were enrolled (seventeen FI, four NTM and one with both infections). The pathogenic variant NG_029334.1:g.16287C>T was found in one patient (4.5%) and two asymptomatic offsprings. We also found the likely-benign variant NG_029334.1:g.12080G>A (rs2335052), the benign variant NG_029334.1:g.16225C>T (rs11708606) and the variant of uncertain significance NG_029334.1:g.16201G>A (rs369850507) in 18.2%, 27.3%, and 4.5% of the cases, respectively. Malignant diseases were additionally diagnosed in six patients. Conclusion Although detected in 45.4% of the patients, most GATA2 variants were benign or likely benign. Identifying a pathogenic variant was essential for driving both the patient's treatment and familial counseling. Pathogenic variants carriers should receive genetic counseling, subsequent infection prevention measures and malignancies surveillance. Additionally, case-control genotyping should be carried out in Brazil to investigate whether the observed variants may be associated with susceptibility to opportunistic infections and/or concurrent neoplasms.

Congenital deaf-mutism with pale complexion and anemia for 1 year in a school-aged girl / 中国当代儿科杂志

An 11-year-old girl was found to have pale complexion and anemia with gradual aggravation for one year. She was weak in the past and developed pneumonia in the right middle lung 3-5 times per year, which was improved after anti-infective therapy. She and her mother had congenital deaf-mutism. Physical examination showed the appearance of anemia, without bleeding, jaundice, hepatosplenomegaly, or lymph node enlargement. Routine blood test results showed reductions in all three blood cell lines, normocytic anemia, and megaloblastoid change in granulocytic and erythroid cell lines in bone marrow, with no obvious increase in primitive cells or metastatic tumor cells. Whole exome sequencing indicated the presence of a known pathogenic mutation for Emberger syndrome (ES), c.1084C>T (p.Arg362*) in the GATA2 gene. The girl was finally diagnosed with ES, and myelodysplastic syndrome (MDS) progressed to acute myeloid leukemia during follow-up. ES is a rare type of MDS with autosomal dominant inheritance in clinical practice, and it is difficult to make a confirmed diagnosis. ES should be considered for children with unexplained lymphedema and congenital deafness, and gene detection should be performed to make a confirmed diagnosis.

Clinical and molecular characteristics of GATA2 related pediatric primary myelodysplastic syndrome / 中华血液学杂志

Objective: To clarify the prevalence, clinical features and molecular characteristics of germline GATA2 mutations in pediatric primary myelodysplastic syndromes (MDS) . Methods: Next-generation sequencing technology was used to detect mutations in GATA2 and other myeloid malignancy genes in 129 children with primary MDS from Jan. 2007 to Jan. 2018. The relationship between genotypes and phenotypes was analyzed. Results: Germline GATA2 mutations accounted for 8.5% (11/129) of all primary MDS cases, and 14.0% (11/50) of MDS with excess blasts (MDS-EB) and acute myeloid leukaemia with myelodysplasia-related changes (AML-MRC) . Compared with GATA2 wild-type patients, GATA2 mutated patients were older at diagnosis[8 (1-16) years old vs 6 years old (range: 1 month old-18 years old) , P=0.035]and higher risk of monosomy 7 (72.7%vs 5.2%, P<0.001) and classified into MDS-EB and AML-MRC compared with refractory cytopenia of childhood (RCC) (63.6%vs 36.4%, P=0.111) . The multivariate analysis showed SETBP1 mutation (P=0.041, OR=9.003, 95%CI 1.098-73.787) and isolated monosomy 7 (P=0.002, OR=24.835, 95%CI 3.305-186.620) were significantly associated with germline mutated GATA2. Overall survival (OS) and outcomes of hematopoietic stem cell transplantation (HSCT) were not influenced by GATA2 mutational status. Conclusions: Our data identify germline GATA2 mutations have a high prevalence in older pediatric patients with monosomy 7, and high risk of progression into advanced MDS subtypes. GATA2 mutation status does not affect OS in pediatric primary MDS.

Lovastatin reduces stemness via epigenetic reprograming of BMP2 and GATA2 in human endometrium and endometriosis

Objective: The stem cell theory in the endometriosis provides an advanced avenue of targeting these cells as a novel therapy to eliminate endometriosis. In this regard, studies showed that lovastatin alters the cells from a stem-like state to more differentiated condition and reduces stemness. The aim of this study was to investigate whether lovastatin treatment could influence expression and methylation patterns of genes regulating differentiation of endometrial mesenchymal stem cells [eMSCs] such as BMP2, GATA2 and RUNX2 as well as eMSCs markers

Materials and Methods: In this experimental investigation, MSCs were isolated from endometrial and endometriotic tissues and treated with lovastatin and decitabin. To investigate the osteogenic and adipogenic differentiation of eMSCs treated with the different concentration of lovastatin and decitabin, BMP2, RUNX2 and GATA2 expressions were measured by real-time polymerase chain reaction [PCR]. To determine involvement of DNA methylation in BMP2 and GATA2 gene regulations of eMSCs, we used quantitative Methylation Specific PCR [qMSP] for evaluation of the BMP2 promoter status and differentially methylated region of GATA2 exon 4

Results: In the present study, treatment with lovastatin increased expression of BMP2 and RUNX2 and induced BMP2 promoter demethylation. We also demonstrated that lovastatin treatment down-regulated GATA2 expression via inducing methylation. In addition, the results indicated that CD146 cell marker was decreased to 53% in response to lovastatin treatment compared to untreated group

Conclusion: These findings indicated that lovastatin treatment could increase the differentiation of eMSCs toward osteogenic and adiogenic lineages, while it decreased expression of eMSCs markers and subsequently reduced the stemness

Expression of NF-E1b in colorectal cancer tissues and its clinical significance / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To explore the expression of NF-E1b in colorectal cancer tissues and its association with various clinicopathological parameters and prognosis of the patients.</p><p><b>METHODS</b>Clinicopathological and follow-up data of 168 colorectal cancer patients undergoing radical operation at Department of Gastrointestinal Surgery, Peking University Cancer Hospital and Institute from 2005 to 2012 were retrospectively analyzed, including 96 males and 72 females, with mean age of (57.8±11.2) years. The expression of NF-E1b protein was detected in samples of 168 resected colorectal cancer tissues and 45 adjacent non-cancerous tissues by immunohistochemistry. The expression rates of NF-E1b were compared among different clinicopathological features. Moreover, the association between NF-E1b expression and prognosis was analyzed.</p><p><b>RESULTS</b>The expression of NF-E1b protein located mainly in cytoplasm. Positive rate of NF-E1b expression in adjacent non-cancerous tissues was 17.8% (8/45), which was obviously lower than 67.9%(114/168) of cancer tissues with significant difference (χ(2)=36.376, P=0.000). Clinicopathological parameters analysis suggested that the expression level of NF-E1b in cancer tissues was associated with age (χ(2)=4.862, P=0.030), TNM staging (χ(2)=10.969, P=0.002), lymph node metastasis (χ(2)=7.390, P=0.008) and distal metastasis (χ(2)=17.887, P=0.000). The median follow-up time was 23(1-77) months. The overall 5-year survival of this cohort was 33.3%. Colorectal cancer patients with high levels of NF-E1b expression showed a worse overall survival compared with those with low levels of NF-E1b expression (18.4% vs. 56.6%, P=0.000). Univariate Cox regression analysis showed that tumor location (P=0.034), tumor size (P=0.003), TNM staging (P=0.000), depth of tumor invasion (P=0.009), lymph node metastasis (P=0.000), distant metastasis (P=0.000) and NF-E1b expression level (P=0.001) were associated with the prognosis of colorectal cancer patients. Multivariate Cox regression analysis revealed that tumor diameter >4 cm (HR=2.193,95% CI:1.334 to 3.603, P=0.002), distant metastasis (HR=2.064, 95% CI:1.160 to 3.672, P=0.014) and high NF-E1b expression (HR=1.994,95% CI:1.068 to 3.724, P=0.030) were independent risk factors of predicting poor prognosis of colorectal cancer patients.</p><p><b>CONCLUSIONS</b>NF-E1b expression up-regulates in colorectal cancer tissues. High expression of NF-E1b is associated with poor prognosis of colorectal cancer patients. NF-E1b may serve as a potential target of the treatment for colorectal cancer.</p>

First Korean case of Emberger syndrome (primary lymphedema with myelodysplasia) with a novel GATA2 gene mutation

No abstract available.

Effect of simulated microgravity on erythroid differentiation of K562 cells and the mechanism / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the effect of simulated microgravity on erythroid differentiation of K562 cells and explore the possible mechanism.</p><p><b>METHODS</b>The fourth generation rotating cell culture system was used to generate the simulated microgravity environment. Benzidine staining was used to evaluate the cell inhibition rate, and real-time quantitative PCR (qRT-PCR) was used to detect GATA-1, GATA-2, Ets-1, F-actin, β-Tubulin and vimentin mRNA expressions. The changes of cytoskeleton were observed by fluorescence microscopy, and Western blotting was employed to assay F-actin, β-tubulin and vimentin protein expression levels.</p><p><b>RESULTS</b>Benzidine staining showed that simulated microgravity inhibited erythroid differentiation of K562 cells. K562 cells treated with Hemin presented with increased mRNA expression of GATA-1 and reduced GATA-2 and Ets-1 mRNA expressions. Simulated microgravity treatment of the cells resulted in down-regulated GATA-1, F-actin, β-tubulin and vimentin mRNA expressions and up-regulated mRNA expressions of GATA-2 and Ets-1, and reduced F-actin, β-tubulin and vimentin protein expressions. Exposure to simulated microgravity caused decreased fluorescence intensities of cytoskeletal filament F-actin, β-tubulin and vimentin in the cells.</p><p><b>CONCLUSION</b>Simulated microgravity inhibits erythroid differentiation of K562 cells possibly by causing cytoskeleton damages to result in down-regulation of GATA-1 and up-regulation of GATA-2 and Ets-1 expressions.</p>

MonoMAC syndrome / 中国当代儿科杂志

MonoMAC syndrome is a newly discovered immune deficiency syndrome caused by GATA-2 mutation, which is an autosomal dominant genetic disease. MonoMAC syndrome has typical immune cell abnormalities, with severe infection and is prone to develop into a hematological disease. Therapeutics for this disease mainly relies on symptomatic treatment and hematopoietic stem cell transplantation. In this paper, the research advances in clinical manifestations, laboratory tests, pathogenesis, diagnosis and treatment of MonoMAC syndrome are reviewed.

Effect of GATA-2 overexpression on function of mouse fetal liver hematopoietic stem cells / 中国实验血液学杂志

This study was aimed to investigate the effect of GATA-2 over-expression on function of mouse fetal liver hematopoietic stem cells. GATA-2 was introduced into mouse fetal liver cells via retrovirus mediated transduction with GFP as a detecting marker. Flow cytometry, colony-forming assay and cell cycle assay were used to detect the biologic changes of these retrovirus infected mouse fetal liver hematopoietic stem cells. The results showed that GATA-2 over-expression increased the Lin(-)Sca1(+)C-Kit(+) (LSK) population dramatically. Cell cycle of LSK cells didn't show abnormal, while colony forming ability decreased significantly. These data indicated that GATA-2 over-expression inhibited definitive differentiation of mouse fetal liver hematopoietic stem cells. It is concluded that over-expression of GATA-2 can significantly raise the LSK cell proportion in mouse fetal liver and inhibit the differentiation capability, the underlying mechanisms may be related to up-regulation of Hes-1, which may lead to the blocking of cell differentiation at the stem/progenitor cell stage.

Effects of transcription factor GATA-2 on transcriptive regulation of iASPP gene / 中国实验血液学杂志

iASPP can prompt the cell proliferation and inhibit the apoptosis of many cells. There are putative binding sites of transcription factor GATA-2 upstream of iASPP transcription start site. GATA-2 plays an important role in the proliferation and differentiation of hematopoietic stem cells (HSC) and progenitors. This study was aimed to explore the role of GATA-2 protein in iASPP gene transcription. Firstly, the expression of iASPP and GATA-2 protein in some leukemia cell lines was detected by Western blot. Second, The expressive vector of pCMV5-GATA2 and the luciferase reporter vectors containing possible binding sites of GATA-2 were constructed and co-transfected into HEK293 and CV-1 cells. Then the luciferase activity was assayed by luminometer. Also, ChIP assays were performed to further confirm the specific binding of GATA-2 to iASPP promoter. The results showed that GATA-2 was overexpressed in most cell lines with high level of iASPP. GATA-2 exhibited a significant effect on luciferase activity of reporter gene iASPP and in a dose-dependant manner. The relative luciferase activity was up-regulated to about two-fold of the empty vector control when the transfection dose of pCMV5-GATA2 plasmid was increased to 100 ng. While the effect was more significant in CV-1 cells and showed a 6.7-fold increase. The ChIP assay demonstrated the in vivo specific binding of GATA-2 to iASPP. The binding sites of GATA2 were located between nt -361 ∼ -334 in upstream of iASPP gene transcription start site. It is concluded that transcription factor GATA-2 can bind with the cis-regulatory region of the iASPP promoter and up-regulate iASPP expression.

Spontaneous differentiation of human embryonic stem cells into hematopoietic cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To characterize the time course of spontaneous differentiation of in vitro cultured human embryonic stem cells (hESCs) into hematopoietic cells to provide experimental evidence for induction of hematopoietic commitment of hESCs.</p><p><b>METHODS</b>In human embryoid bodies (hEBs) derived from spontaneous differentiation of chESC3, a hESC cell line we established previously, the expressions of such genes as KDR, Bmi1, Scl and gata2 were detected by RT-PCR every other day during the 12-day differentiation to monitor the process of the hematopoiesis. The hematopoietic stem cell marker CD34 was examined using flow cytometry to evaluate the efficiency of hematopoietic differentiation of the cells on days 6, 8, 10 and 12. The spontaneously differentiated hESCs were seeded in the hematopoietic colony culture system to study the hematopoietic colony forming ability. Immunocytochemical staining for CD45 was performed on the hEBs to examine the emergence of mature hematopoietic cells.</p><p><b>RESULTS</b>The expressions of the hematopoietic stem cell-related genes KDR and Bmi-1 were detected in the hESCs, and on days 4 to 6, the two genes were upregulated with prolonged cuture of the hEBs. Scl and gata2 gene expressions were detected since 6-8 days of culture and maintained high expressions till day 12. Flow cytometry revealed a gradual increase in CD34-positive cells in the culture, with positivity rates on days 6, 8, 10, and 12 of (1.4-/+0.4)%, (3.4-/+1.3)%, (5.5-/+2.2)%, and (5.1-/+1.7)%, respectively. The numbers of CD43-positive cell colonies on days 6, 8, 10, and 12 were 0, 7-/+2, 37-/+11, and 89-/+29 in each 10(5) cells, respectively. Immunocytochemical staining identified CD45-positive cells on days 10, 12, 15, and 18 in the cell colonies, with the positive cell numbers of 0, 40.5-/+15.09, 178.6-/+55.89, and 253.0-/+52.04, respectively.</p><p><b>CONCLUSION</b>The hESCs undergo spontaneous hematopoietic differentiation in 3 stages, including the differentiation into germ layer-specific cells (days 6-8), expansion period of the hematopoitic progenitors (days 8-12), and maturation of the hematopoietic cells (after day 15).</p>

Expression of GATA-1 and GATA-2 in the bone marrow of patients with Monge's disease / 中华血液学杂志

<p><b>OBJECTIVE</b>To find out how GATA-1 and GATA-2 behave in the bone marrow of patients with Monge's disease.</p><p><b>METHODS</b>The levels of mRNA in mononuclear cells (MNC) and proteins of GATA-1 and GATA-2 in the bone marrow of patients with Monge's disease and controls were determined by RT-PCR and immune cytolysis chemical method.</p><p><b>RESULTS</b>(1) All patients and controls expressed GATA-1 mRNA (Monge's disease 1.033 +/- 0.146, Control 0.458 +/- 0.076) and GATA-2 mRNA (Monge's disease 0.451 +/- 0.073, Control 0.185 +/- 0.074). All patients expressed both GATA-1 (positive cell counts 77.3 +/- 33.3, positive score 135.4 +/- 75.4) and GATA-2 ( positive cell counts 29.4 +/- 11.4, positive score 48.4 +/- 19.7). All the controls expressed GATA-1 (positive cell counts 18.1 +/- 11.3, positive score 24.2 +/- 13.4) while 12 of 20 controls expressed GATA-2 ( positive cell counts 5.4 +/- 3.0, positive score 7.3 +/- 4.2). The expression of mRNA and proteins of GATA-1 and GATA-2 in Monge's disease were higher than in controls (P < 0.01). (2) There was a positive correlation between GATA-1 and Hb (P < 0.01), as did between mRNA and proteins of GATA-1 and GATA-2. (3) Both the proteins of GATA-1 and GATA-2 located only in the cytoplasm but not the nucleus.</p><p><b>CONCLUSIONS</b>Two of inherent genes, GATA-1 and GATA-2 which were expressed at higher levels in patients with Monge's disease than in controls might play significant roles in the pathogenesis of Monge's disease.</p>

Expressions of transcription factor GATA-1 and GATA-2 genes in bone marrow stromal cells from patients with leukemia / 中国实验血液学杂志

In order to investigate expressions of transcription factor GATA-1 and GATA-2 genes in the bone marrow stromal cells (BMSCs) from patients with leukemia or normal controls, bone marrow stromal cells from 34 normal cases and 42 cases with leukemia were cultured long-term in vitro. Nonadherent cells (bone marrow hematopoietic cells) and amplified adherent cells (BMSC) were collected separately. Expressions of GATA-1 and GATA-2 genes were analyzed by using RT-PCR-ELISA; the semi-quantitative expression levels of GATA genes in the BMSCs from patients with leukemia were compared with normal controls. The results showed that expressions of GATA-1 and GATA-2 genes could be detected in the BMSCs and the bone marrow hematopoietic cells from both normal controls and the cases of leukemia. The expression ratio of GATA-1 in the BMSCs from acute lymphocytic leukemia (ALL) (85.7%) was similar to the normal controls (88.2%), whereas the expression ratios in BMSCs from acute myelocytic leukemia (AML) (55.6%) and chronic myelocytic leukemia (CML) (41.2%) were significant lower than the normal controls (P < 0.05). The rank of expression level of GATA-1 gene in the BMSCs was "ALL>AML>normal>CML". There was no difference in the expression level of GATA-2 gene within the BMSCs from normal controls and patients with leukemia. The ranks of expression levels of GATA-1 and GATA-2 genes in bone marrow hematopoietic cells were "AML>normal>ALL>CML" and "AML>CML>ALL>normal". The dominant expression of GATA-2 gene was found in the BMSCs from AML, CML or normal controls. It is inferred that the expressions of GATA-1 and GATA-2 genes in the BMSCs of normal controls and patients with leukemia may influence the regulation of hematopoiesis in the bone marrow stroma and it is worthy of further study to explore their roles in pathogenesis and development of leukemia.

Up-regulation of transcription factors GATA-1 and GATA-2 induced by Panax notoginosides in hematopoietic cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To observe the role of Panax notoginosides (PNS) in up-regulation of GATA family transcription factors, and explore intracellular signal pathway of PNS in the proliferation of hematopoietic cells.</p><p><b>METHODS</b>Human bone marrow cells were incubated with different concentrations of PNS for colony-forming assay. Human cell lines HL-60, K562, CHRF-288 and Meg-01 were incubated with PNS (10 mg/L) for 14 days. The cell nuclear proteins were extracted and analyzed by Western blot with antibodies against GATA-1, GATA-2. Electrophoretic mobility shift assay (EMSA) and antibody gel supershift assay was performed using (32)P labeled GATA consensus oligonucleotide which contains binding site for GATA transcription factors.</p><p><b>RESULTS</b>PNS could promote the proliferation of CFU-GM and CFU-E and induce the expression of GATA-1, GATA-2. The nuclear proteins of both GATA-1 and GATA-2 in K562, CHRF-288 and Meg-01 cells treated by PNS were increased by (1.5 - 2.8) and (2.0 - 3.1)-fold over untreated cells respectively. GATA binding activity initiated by PNS was apparently elevated to form higher density band of GATA-DNA complex. While there was no detectable change in HL-60 cells before and after PNS treatment. The predominant GATA binding complex was mainly attributable to both GATA-1 and GATA-2 proteins being in phosphorylated status.</p><p><b>CONCLUSION</b>PNS can induce the synthesis of transcription factors GATA-1 and GATA-2 and enhance their DNA binding activity, which could play a role in the up-regulation of the expression genes related to proliferation and differentiation in hematopoietic cells.</p>

The investigation of hematopoietic capacity of HPP-CFC derived from murine embryonic stem cells in vitro and in vivo / 生物工程学报

The hematopoietic system of the mouse arises from extraembryonic mesoderm that migrate through primitive streak to the presumptive yolk sac at day 7.0 of gestation. However, the mechanisms regulating mesoderm commitment to hematopoietic lineages remain poorly understood. Previous studies demonstrated that the development kinetics and growth factor responsiveness of hematopoietic precursors derived from embryonic stem cells (ES cells) is similar to that found in the yolk sac, indicating that the onset of hematopoiesis within the embryoid bodies (EBs) parallels that found in the embryo. Furthermore, in vitro differentiation of ES cells to hematopoietic cells is valuable for establishment of therapeutic clone against a variety of hematological disorders. Despite the identification of multipotential hematopoietic progenitors in EBs, a subset of more primitive progenitors, identical to the high proliferative potential colony-forming cells (HPP-CFC) derived from human and murine hematopoietic tissues, have not been clearly identified regarding particular their replating potential in vitro. HPP-CFC is among the most primitive hematopoietic multipotent precursors cultured in vitro. In this study, our aim was to investigate the in vitro and in vivo hematopoietic capacity of HPP-CFC within the day 12 EBs, rather than the expansion of more committed progenitors. In this study the HPP-CFC could be detected within EBs differentiated for 5 to 14 days of murine ES cells, but the development dynamics of the HPP-CFC differed greatly among distinct serum lots. Qualitatively HPP-CFC is capable of forming secondary colonies. As to our expectation the ES cells-derived HPP-CFC demonstrated similar regeneration capacity to those from yolk sac, giving rise to secondary granulocyte, erythrocyte, macrophage and mast cells, however largely differed from the counterparts of adult bone marrow. In addition, by RT-PCR ES cells-derived HPP-CFC were found to express transcription factors associated closely with stem cell proliferation including SCL, GATA-2 and AML1 as well as various receptors of hematopoietic growth factors such as c-kit, GM-CSF receptor and interleukin 3 receptor et al. Finally, in order to understand the in vivo hematopoietic capacity of the ES cells-derived HPP-CFC, spleen colony-forming unit (CFU-S) assay was performed. Nevertheless, typical CFU-S was not observed after transplantation of the day 12 EB cells or HPP-CFC colonies into lethally irradiated adult murine. In conclusion the HPP-CFC differentiated from murine ES cells displayed robust hematopoietic activity in vitro, however their in vivo reconstitution ability was not detected. The difference between in vitro and in vivo hematopoietic activities of ES cells-derived primitive hematopoietic precursors deserves further investigation.