Search tips
Search criteria

Results 1-9 (9)

Clipboard (0)

Select a Filter Below

more »
Year of Publication
Document Types
1.  HMGA2 Moderately Increases Fetal Hemoglobin Expression in Human Adult Erythroblasts 
PLoS ONE  2016;11(11):e0166928.
Induction of fetal hemoglobin (HbF) has therapeutic importance for patients with beta-hemoglobin disorders. Previous studies showed that let-7 microRNAs (miRNAs) are highly regulated in erythroid cells during the fetal-to-adult developmental transition, and that targeting let-7 mediated the up-regulation of HbF to greater than 30% of the total globin levels in human adult cultured erythroblasts. HMGA2 is a member of the high-mobility group A family of proteins and a validated target of the let-7 family of miRNAs. Here we investigate whether expression of HMGA2 directly regulates fetal hemoglobin in adult erythroblasts. Let-7 resistant HMGA2 expression was studied after lentiviral transduction of CD34(+) cells. The transgene was regulated by the erythroid-specific gene promoter region of the human SPTA1 gene (HMGA2-OE). HMGA2-OE caused significant increases in gamma-globin mRNA expression and HbF to around 16% of the total hemoglobin levels compared to matched control transductions. Interestingly, no significant changes in KLF1, SOX6, GATA1, ZBTB7A and BCL11A mRNA levels were observed. Overall, our data suggest that expression of HMGA2, a downstream target of let-7 miRNAs, causes moderately increased gamma-globin gene and protein expression in adult human erythroblasts.
PMCID: PMC5115839  PMID: 27861570
2.  Erythroid-Specific Expression of LIN28A Is Sufficient for Robust Gamma-Globin Gene and Protein Expression in Adult Erythroblasts 
PLoS ONE  2015;10(12):e0144977.
Increasing fetal hemoglobin (HbF) levels in adult humans remains an active area in hematologic research. Here we explored erythroid-specific LIN28A expression for its effect in regulating gamma-globin gene expression and HbF levels in cultured adult erythroblasts. For this purpose, lentiviral transduction vectors were produced with LIN28A expression driven by erythroid-specific gene promoter regions of the human KLF1 or SPTA1 genes. Transgene expression of LIN28A with a linked puromycin resistance marker was restricted to the erythroid lineage as demonstrated by selective survival of erythroid colonies (greater than 95% of all colonies). Erythroblast LIN28A over-expression (LIN28A-OE) did not significantly affect proliferation or inhibit differentiation. Greater than 70% suppression of total let-7 microRNA levels was confirmed in LIN28A-OE cells. Increases in gamma-globin mRNA and protein expression with HbF levels reaching 30–40% were achieved. These data suggest that erythroblast targeting of LIN28A expression is sufficient for increasing fetal hemoglobin expression in adult human erythroblasts.
PMCID: PMC4684222  PMID: 26675483
3.  Iron dose-dependent differentiation and enucleation of human erythroblasts in serum-free medium 
Improvements in ex vivo generation of enucleated red blood cells are being sought for erythroid biology research, toward the ultimate goal of erythrocyte engineering for clinical use. Based upon the high levels of iron-saturated transferrin in plasma serum, it was hypothesized that terminal differentiation in serum-free media may be highly dependent on the concentration of iron. Here adult human CD34+ cells were cultured in a serum-free medium containing dosed levels of iron-saturated transferrin (holo-Tf, 0.1–1.0 mg/ml). Iron in the culture medium was reduced, but not depleted, with erythroblast differentiation into haemoglobinized cells. At the lowest holo-Tf dose (0.1 mg/ml), terminal differentiation was significantly reduced and the majority of the cells underwent apoptotic death. Cell survival, differentiation and enucleation were enhanced as the holo-Tf dose increased. These data suggest that adequate holo-Tf dosing is critical for terminal differentiation and enucleation of human erythroblasts generated ex vivo in serum-free culture conditions. Published 2013. This article is a US Government work and is in the public domain in the USA.
PMCID: PMC3883763  PMID: 23606586
erythropoiesis; serum-free media; holotransferrin; haemoglobin; enucleation; iron
4.  LIN28A Expression Reduces Sickling of Cultured Human Erythrocytes 
PLoS ONE  2014;9(9):e106924.
Induction of fetal hemoglobin (HbF) has therapeutic importance for patients with sickle cell disease (SCD) and the beta-thalassemias. It was recently reported that increased expression of LIN28 proteins or decreased expression of its target let-7 miRNAs enhances HbF levels in cultured primary human erythroblasts from adult healthy donors. Here LIN28A effects were studied further using erythrocytes cultured from peripheral blood progenitor cells of pediatric subjects with SCD. Transgenic expression of LIN28A was accomplished by lentiviral transduction in CD34(+) sickle cells cultivated ex vivo in serum-free medium. LIN28A over-expression (LIN28A-OE) increased HbF, reduced beta (sickle)-globin, and strongly suppressed all members of the let-7 family of miRNAs. LIN28A-OE did not affect erythroblast differentiation or prevent enucleation, but it significantly reduced or ameliorated the sickling morphologies of the enucleated erythrocytes.
PMCID: PMC4154803  PMID: 25188417
5.  A Synthetic Model of Human Beta-Thalassemia Erythropoiesis Using CD34+ Cells from Healthy Adult Donors 
PLoS ONE  2013;8(7):e68307.
Based upon the lack of clinical samples available for research in many laboratories worldwide, a significant gap exists between basic and clinical studies of beta-thalassemia major. To bridge this gap, we developed an artificially engineered model for human beta thalassemia by knocking down beta-globin gene and protein expression in cultured CD34+ cells obtained from healthy adults. Lentiviral-mediated transduction of beta-globin shRNA (beta-KD) caused imbalanced globin chain production. Beta-globin mRNA was reduced by 90% compared to controls, while alpha-globin mRNA levels were maintained. HPLC analyses revealed a 96% reduction in HbA with only a minor increase in HbF. During the terminal phases of differentiation (culture days 14–21), beta-KD cells demonstrated increased levels of insoluble alpha-globin, as well as activated caspase-3. The majority of the beta-KD cells underwent apoptosis around the polychromatophilic stage of maturation. GDF15, a marker of ineffective erythropoiesis in humans with thalassemia, was significantly increased in the culture supernatants from the beta-KD cells. Knockdown of beta-globin expression in cultured primary human erythroblasts provides a robust ex vivo model for beta-thalassemia.
PMCID: PMC3704632  PMID: 23861885
6.  Prevalence of Uterine Leiomyomas in Lymphangioleiomyomatosis 
Fertility and sterility  2011;96(3):711-714.e1.
To determine the frequency of uterine leiomyomas and hysterectomy in patients with lymphangioleiomyomatosis (LAM), a disease characterized by proliferation of abnormal-appearing smooth muscle-like cells.
Retrospective study.
456 patients with sporadic LAM and LAM associated with tuberous sclerosis complex (LAM/TSC).
Natural history study at the National Institutes of Health.
Review of records and pelvic computed axial tomography scans.
Main Outcome Measure(s)
prevalence of uterine leiomyomas and hysterectomy.
A total of 174 women had uterine leiomyomas (38%). One hundred eighteen were diagnosed by CT scan and 56 were diagnosed by hysterectomy. Among 323 patients who did not have hysterectomy, 105 of 270 patients (39%) with sporadic LAM and 13 of 53 (25%) with LAM/TSC had uterine leiomyomas. Hysterectomy was performed in 108 of 378 subjects with sporadic LAM and 25 of 78 with LAM/TSC. Fifty six patients were found to have uterine fibroids at hysterectomy. The most common indications for hysterectomy were uterine leiomyoma, lymphangioleiomyomatosis and endometriosis.
Uterine leiomyomas are not more common in LAM than in the general population. However, in LAM, the frequency of hysterectomy is higher because of it having been recommended for treatment of LAM.
PMCID: PMC3165169  PMID: 21880281
Uterine leiomyomas; hysterectomy; lymphangioleiomyomatosis
7.  Expression of growth differentiation factor 15 is not elevated in individuals with iron deficiency secondary to volunteer blood donation 
Transfusion  2010;50(7):1532-1535.
Low serum hepcidin levels provide a physiologic response to iron demand in patients with iron deficiency (ID). Based on a discovery of suppressed hepcidin expression by a cytokine named growth differentiation factor 15 (GDF15), it was hypothesized that GDF15 may suppress hepcidin expression in humans with ID due to blood loss.
To test this hypothesis, GDF15 and hepcidin levels were measured in peripheral blood from subjects with iron-deficient erythropoiesis before and after iron supplementation.
Iron variables and hepcidin levels were significantly suppressed in iron-deficient blood donors compared to healthy volunteers. However, ID was not associated with elevated serum levels of GDF15. Instead, iron-deficient subjects’ GDF15 levels were slightly lower than those measured in the control group of subjects (307 ± 90 and 386 ± 104 pg/mL, respectively). Additionally, GDF15 levels were not significantly altered by iron repletion.
ID due to blood loss is not associated with a significant change in serum levels of GDF15.
PMCID: PMC3282986  PMID: 20210929
Bju International  2011;107(4):678-679.
PMCID: PMC3277454  PMID: 21276178
9.  Sporadic Lymphangioleiomyomatosis and Tuberous Sclerosis Complex with Lymphangioleiomyomatosis: Comparison of CT Features1 
Radiology  2006;242(1):277-285.
To retrospectively compare the frequencies of computed tomographic (CT) findings in patients with lymphangioleiomyomatosis (LAM) and patients with tuberous sclerosis complex (TSC) and LAM.
Materials and Methods
Institutional review board approval and informed consent were obtained for the HIPAA-compliant study. In 256 patients with LAM (mean age, 44 years) and 67 patients with TSC/LAM (mean age, 40 years), CT scans of the chest, abdomen, and pelvis were reviewed by a single radiologist. The fraction of lung involvement with cysts was estimated from high-spatial-resolution CT scans. Other findings assessed included noncalcified pulmonary nodules, pleural effusion, thoracic duct dilatation, hepatic and renal angiomyolipomas (AMLs), lymphangioleiomyoma (LALM), ascites, nephrectomy, and renal embolization. Confidence intervals and hypothesis tests of differences in frequencies, comparison of age quartiles, RIDIT analysis, analysis of variance, and correlation coefficients were used in the statistical analysis.
Patients with LAM had more extensive lung involvement (RIDIT score, 0.36) and higher frequency of LALM (29% vs 9%, P < .001), thoracic duct dilatation (4% vs 0, P = .3), pleural effusion (12% vs 6%, P = .2), or ascites (10% vs 6%, P = .3). Patients with TSC/LAM had higher frequency of noncalcified pulmonary nodules (12% vs 1%, P < .01), hepatic (33% vs 2%, P < .001) and renal (93% vs 32%, P < .001) AMLs, nephrectomy (25% vs 7%, P < .001), or renal artery embolization (9% vs 2%, P < .05).
The extent of lung disease is greater in LAM than TSC/LAM. Hepatic and renal AMLs and noncalcified lung nodules are more common in TSC/LAM, while lymphatic involvement—thoracic duct dilatation, chylous pleural effusion, ascites, and LALM—is more common in LAM.
PMCID: PMC2940246  PMID: 17105849

Results 1-9 (9)