B A Miller

Penn State Hershey Medical Center and Penn State College of Medicine, Hershey, Pennsylvania, United States

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Publications (54)636.54 Total impact

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    ABSTRACT: Erythropoietin (EPO) is a lineage-restricted growth factor that is required for erythroid proliferation and differentiation. EPO stimulates the phosphorylation and activation of p70 S6 kinase (p70 S6K), which is required for cell cycle progression. Here, the minimal cytoplasmic domains of the EPO receptor (EPO-R) required for p70 S6K activation were determined.Ba/F3 cells were stably transfected with wild-type (WT) EPO-R or EPO-R carboxyl-terminal deletion mutants, designated by the number of amino acids deleted from the cytoplasmic tail (-99, -131, -221). Transfected cells were growth factor deprived and then stimulated with EPO. p70 S6K, JAK2, IRS-2, and ERK1/2 phosphorylation/activation were examined. The ability of transfected 3-phosphoinositide-dependent protein kinase 1 (PDK1) to reconstitute p70 S6K phosphorylation in EPO-R mutants also was determined. Phosphorylation and activation of p70 S6K, JAK2, IRS-2, and ERK1/2 in Ba/F3 cells transfected with EPO-R-99 or EPO-R-99Y343F were similar to WT EPO-R. In contrast, EPO-dependent p70 S6K phosphorylation/activation, as well as IRS-2 and ERK1/2 phosphorylation, were minimal or absent in cells transfected with EPO-R-131 or EPO-R-221. JAK2 phosphorylation was reduced significantly in cells transfected with EPO-R-131 and abolished with EPO-R-221. To examine the role of PDK1, a kinase known to phosphorylate p70 S6K, Ba/F3 EPO-R-131 cells were transiently transfected with PDK1. WT constitutively active PDK1 restored p70 S6K phosphorylation in Ba/F3 EPO-R-131 cells but not in Ba/F3 EPO-R-221 cells. The results demonstrate that a minimal cytoplasmic subdomain of the EPO-R extending between -99 and -131 is required for p70 S6K phosphorylation and activation. The results also demonstrate that PDK1 is a critical component in this signaling pathway, which requires the presence of domains between -131 and -221 for its activation of p70 S6K.
    Experimental Hematology 05/2001; 29(4):432-40. · 2.91 Impact Factor
  • J Y Cheung, B A Miller
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    ABSTRACT: Erythropoietin is an obligatory growth factor for red blood cell production. The receptor for erythropoietin contains a single membrane-spanning domain with no intrinsic tyrosine kinase motifs. On binding to erythropoietin, the receptor dimerizes and activates multiple intracellular signaling molecules, including but not limited to JAK2, STAT5, PI 3-kinase, IRS-2, RAS, and Ca2+ channels. This review focuses on cytoplasmic signaling cascades involved in erythropoietin action.
    Nephron 04/2001; 87(3):215-22. · 13.26 Impact Factor
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    ABSTRACT: The filamentous fungus Aspergillus nidulans nudC (nuclear distribution C) gene is required for movement of nuclei following mitosis and for normal colony growth. It is highly conserved, structurally and functionally, throughout most of evolution. The human homolog, called HnudC, has been cloned and has an important role in cell proliferation. In hematopoiesis, HNUDC is highly expressed in early hematopoietic precursors and declines during normal differentiation. Stimulation of proliferation of the erythroleukemia cell line TF-1 with GM-CSF enhances HnudC protein and mRNA expression and treatment with antisense (but not sense) oligonucleotides to HnudC mRNA significantly reduces cell division. A significant increase in HNUDC is present in bone marrow aspirates from patients with acute lymphoblastic leukemia (ALL) and acute myelogenous leukemia (AML) compared to the level in normal cellular counterparts, demonstrating dysregulated expression in leukemia. These data support the conclusion that HnudC plays a functional role in promoting hematopoietic cell growth and that it is involved in leukemogenesis.
    Leukemia and Lymphoma 12/2000; 39(5-6):447-54. · 2.61 Impact Factor
  • C D Gocke, S A Osmani, B A Miller
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    ABSTRACT: We recently identified a novel human gene, HnudC, homologous to an Aspergillus nidulans gene coding for a protein crucial to nuclear migration, cell wall morphogenesis, and cell growth. While mRNA for this gene is expressed in most tissues, HNUDC protein expression is highly regulated. To provide insight into the function of this protein, we performed immunohistochemical analysis of the distribution of HNUDC in 19 different human tissues. Intense immunolabeling was observed in proliferating cells, including spermatocytes at all stages, early hematopoietic cells, cortical thymocytes, immunoblasts, and basal colonic and esophageal mucosa. Within a given tissue, cells with different proliferative capacities demonstrated different levels of HNUDC expression. HNUDC was also highly expressed in ciliated epithelia including those found in ependyma, bronchial mucosa, and fallopian tubes. Immunolabeling was moderate in several non-proliferating tissues, but little or no labeling was observed in most other tissues examined. We also demonstrated by western blotting that most cell lines express extremely high levels of HNUDC compared to their normal counterparts. While this supports a role for HnudC in cell proliferation, these data indicate that cell lines are not a reliable measure of HNUDC protein expression in normal tissues. We conclude that HNUDC is highly expressed in cell lines and the proliferating cells of normal tissues, consistent with our hypothesis that HNUDC is conserved throughout evolution for a crucial function in cell division. In addition, the high level in ciliated cells suggests an important role in ciliary motility or assembly, analogous to its role in A. nidulans nuclear movement.
    Histochemie 11/2000; 114(4):293-301. · 2.61 Impact Factor
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    ABSTRACT: The pathogenesis of posttransplant erythrocytosis (PTE) has been elusive. Angiotensin converting enzyme inhibitors (ACEI) are efficacious in lowering the hematocrit of patients with PTE and angiotensin II (AII) type I receptors (AT1R) were recently detected on red blood cell precursors, burst-forming unit-erythroid- (BFU-E) derived cells. The purpose of this study was to determine whether there is increased expression of the AT1R on BFU-E-derived cells of patients with PTE, which might contribute to the pathogenesis of PTE. Twelve healthy volunteers and 25 transplant recipients (13 patients with and 12 without PTE) were studied. BFU-E from peripheral blood were cultured in methylcellulose and BFU-E-derived colonies were harvested on day 10. Western blotting was used to detect AT1R and erythropoietin receptor (EpoR) expression. Intracellular free calcium in response to AII and erythropoietin (Epo) was measured with digital video imaging. There were no differences between transplant patients, with and without PTE, with respect to weight, age, sex, blood pressure, serum creatinine, circulating renin, angiotensin II, and Epo levels. Hematocrit, red blood cell number, BFU-E-derived colony number,and size were significantly increased in PTE compared with other two groups. AT1R expression was increased by 44% on the erythroid progenitors of PTE versus non posttransplant erythrocytosis patients and by 32% in PTE patients versus normal volunteers. AT1R expression correlated significantly with the hematocrit in PTE (Spearman r=0.68, P=0.01). In contrast, EpoR expression was equivalent in all groups. The AT1R was functional since a significant increase in [Ca(i)] was observed in Fura-2 loaded day 10 cells when stimulated with AII (182%, P<0.0001). An increase in AT1R density was observed in erythroid precursors of transplant patients with PTE compared to those without PTE and normal volunteers, and the level of AT1R expression in PTE correlated significantly with the hematocrit. In contrast, EpoR expression was not different in PTE compared with non posttransplant erythrocytosis or normal controls. This study supports a role for the AT1 receptor signaling pathway in the pathogenesis of PTE.
    Transplantation 10/2000; 70(8):1188-94. · 3.78 Impact Factor
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    ABSTRACT: Erythropoietin (Epo) activates a voltage-independent Ca2+ channel that is dependent on tyrosine phosphorylation. To identify the domain(s) of the Epo receptor (Epo-R) required for Epo-induced Ca2+ influx, Chinese hamster ovary (CHO) cells were transfected with wild-type or mutant Epo receptors subcloned into pTracer-cytomegalovirus vector. This vector contains an SV40 early promoter, which drives expression of the green fluorescent protein (GFP) gene, and a cytomegalovirus immediate-early promoter driving expression of the Epo-R. Successful transfection was verified in single cells by detection of GFP, and intracellular Ca2+ ([Ca]i) changes were simultaneously monitored with rhod-2. Transfection of CHO cells with pTracer encoding wild-type Epo-R, but not pTracer alone, resulted in an Epo-induced [Ca]i increase that was abolished in cells transfected with Epo-R F8 (all eight cytoplasmic tyrosines substituted). Transfection with carboxyl-terminal deletion mutants indicated that removal of the terminal four tyrosine phosphorylation sites, but not the tyrosine at position 479, abolished Epo-induced [Ca]i increase, suggesting that tyrosines at positions 443, 460, and/or 464 are important. In CHO cells transfected with mutant Epo-R in which phenylalanine was substituted for individual tyrosines, a significant increase in [Ca]i was observed with mutants Epo-R Y443F and Epo-R Y464F. The rise in [Ca]i was abolished in cells transfected with Epo-R Y460F. Results were confirmed with CHO cells transfected with plasmids expressing Epo-R mutants in which individual tyrosines were added back to Epo-R F8 and in stably transfected Ba/F3 cells. These results demonstrate a critical role for the Epo-R cytoplasmic tyrosine 460 in Epo-stimulated Ca2+ influx.
    Journal of Biological Chemistry 08/1999; 274(29):20465-72. · 4.65 Impact Factor
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    ABSTRACT: The filamentous fungus Aspergillus nidulans nudC gene has an essential function in movement of nuclei following mitosis and is required for normal colony growth. Here, the molecular cloning and role in hematopoiesis of a human gene (designated HnudC) homologous to A. nidulans nudC is reported. The amino terminus of the larger human protein (HNUDC = 45 kDa) does not overlap with A. nidulans NUDC (22 kDa). However, NUDC and the C-terminal 94 amino acids of HNUDC are 67% identical. The C-terminal region of the HnudC gene fully complements the A. nidulans temperature-sensitive nudC3 mutation, suggesting that nudC has an essential function in cell growth that is conserved from filamentous fungi to humans. In initial studies, HNUDC levels were much higher in erythroid precursors compared to most other human tissues. Therefore, the potential role of HnudC in hematopoiesis was explored. In normal human bone marrow, HNUDC protein and mRNA are highly expressed in early myeloid and erythroid precursors and decline as these cells terminally differentiate. To determine whether hematopoietic growth factors induce HnudC expression, TF-1 cells were stimulated by granulocyte-macrophage colony-stimulating factor. This induced a significant increase in HNUDC protein and HnudC mRNA, suggesting that enhancement of HnudC expression in response to growth factor stimulation may be mediated at the transcription level. Furthermore, HNUDC was significantly enhanced in lysates of bone marrow aspirates from patients with acute myelogenous and acute lymphoblastic leukemia compared to aspirates from normal controls, suggesting that HnudC is involved in malignant hematopoietic cell growth as well. These data demonstrate that HNUDC is highly expressed in normal and malignant human hematopoietic precursors and suggest it is of functional importance in the proliferation of these cells.
    Experimental Hematology 05/1999; 27(4):742-50. · 2.91 Impact Factor
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    ABSTRACT: This article presents a space-time scan statistic, useful for evaluating space-time cluster alarms, and illustrates the method on a recent brain cancer cluster alarms in Los Alamos, NM. The space-time scan statistic accounts for the preselection bias and multiple testing inherent in a cluster alarm. Confounders and time trends can be adjusted for. The observed excess of brain cancer in Los Alamos was not statistically significant. The space-time scan statistic is useful as a screening tool for evaluating which cluster alarms merit further investigation and which clusters are probably chance occurrences.
    American Journal of Public Health 10/1998; 88(9):1377-80. · 3.93 Impact Factor
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    ABSTRACT: Bcl-3 is a proto-oncogene involved in the chromosomal translocation t(14;19) found in some patients with chronic lymphocytic leukemia. It shares structural similarities with and is a member of the IkappaB family of proteins. In this report, involvement of Bcl-3 in hematopoietic growth factor-stimulated erythroid proliferation and differentiation was examined. In TF-1 cells, an erythroleukemia cell line, granulocyte-macrophage colony-stimulating factor (GM-CSF) and erythropoietin (Epo) greatly enhanced Bcl-3 expression at both the protein and mRNA levels in association with stimulation of proliferation. Bcl-3 protein was also highly expressed in early burst-forming unit-erythroid (BFU-E)-derived erythroid precursors (day 7) and decreased during maturation (days 10 and 14), suggesting that Bcl-3 is involved in normal erythroid proliferation. In these hematopoietic cells, Bcl-3 was hyperphosphorylated. GM-CSF and Epo modulated the subcellular localization of Bcl-3. Upon stimulation of TF-1 cells with GM-CSF or Epo, the nuclear translocation of Bcl-3 was dramatically enhanced. Overexpression of Bcl-3 in TF-1 cells by transient transfection along with the NF-kappaB factors p50 or p52 resulted in significant induction of an human immunodeficiency virus-type 1 (HIV-1) kappaB-TATA-luceriferase reporter plasmid, demonstrating that Bcl-3 has a positive role in transactivation of kappaB-containing genes in erythroid cells. Stimulation with GM-CSF enhanced c-myb mRNA expression in these cells. Bcl-3 in nuclear extracts of TF-1 cells bound to a kappaB enhancer in the c-myb promoter together with NF-kappaB2/p52 and this binding activity was enhanced by GM-CSF stimulation. Furthermore, cotransfection of Bcl-3 with p52 or p50 in TF-1 cells resulted in significant activation of a c-myb kappaB-TATA-luceriferase reporter plasmid. These findings suggest that Bcl-3 may participate in the transcriptional regulation of certain kappaB-containing genes involved in hematopoiesis, including c-myb.
    Blood 09/1998; 92(4):1225-34. · 9.78 Impact Factor
  • M Y Zhang, S C Sun, L Bell, B A Miller
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    ABSTRACT: NF-kappaB/Rel designates a widely distributed family of transcription factors involved in immune and acute phase responses. Here, the expression and function of NF-kappaB factors in erythroid proliferation and differentiation were explored. In an erythroleukemia cell line, TF-1, high levels of p105/p50, p100/p52, p65, and IkappaBalpha were detected 24 hours after growth factor deprivation. In response to granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulation, significant induction of p52 expression was observed. GM-CSF also induced nuclear translocation of both p52 and p65. No induction of NF-kappaB factors was observed with erythropoietin stimulation of TF-1 cells. Overexpression of p52 and p65 in TF-1 cells by transient transfection resulted in significant induction of a kappaB-TATA-luciferase reporter plasmid, showing that these factors are functional in vivo in erythroid cells. To determine whether NF-kappaB factors may play a role in normal erythropoiesis, levels of these factors were determined in burst-forming unit-erythroid (BFU-E)-derived cells at different stages of differentiation. The NF-kappaB factors p105/p50, p100/p52, and p65 were highly expressed in early BFU-E-derived precursors, which are rapidly proliferating, and declined during maturation. Furthermore, nuclear levels of NF-kappaB factors p50, p52, and p65 were higher in less mature precursors (day 10 BFU-E-derived cells) compared with more differentiated (day 14) erythroblasts. In nuclear extracts from day 10 BFU-E-derived cells, p50, p52, and p65 were able to form complexes, which bound to kappaB sites in the promoters of both the c-myb and c-myc genes, suggesting that c-myb and c-myc may be among the kappaB-containing genes regulated by NF-kappaB factors in normal erythroid cells. Taken together, these data show that NF-kappaB factors are modulated by GM-CSF and suggest they function to regulate specific kappaB containing genes involved in erythropoiesis.
    Blood 07/1998; 91(11):4136-44. · 9.78 Impact Factor
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    ABSTRACT: During the decade between 1980-1990, the rate of cancer in children in the U.S. increased. It is unknown whether cancer in infancy, which is biologically and clinically different from cancer in older children, also increased. To evaluate changes in cancer incidence in infants in the U.S. age < 1 year, data from the Surveillance, Epidemiology, and End Results (SEER) program and the U.S. Bureau of the Census were used to construct age specific, population-based cancer incidence rates. Overall, the annual cancer rate in infants increased from 189 cases per million infants between 1979-1981 to 220 between 1989-1991. At both timepoints, female infants had higher cancer rates than male infants. Although the rates for female infants remained stable at 223 between 1979-1981 versus 236 between 1989-1991, rates for male infants increased from 158 to 205 during the same timepoints. Male infants had increased rates of central nervous system (CNS) tumors (P < 0.05), neuroblastoma, and retinoblastoma; female infants had increased rates of teratomas (P < 0.01) and hepatoblastomas. Between 1979-1981, the three most common types of cancer in infants were neuroblastoma, leukemia, and renal tumors (27%, 15%, and 14%, respectively), and were neuroblastoma, CNS tumors, and leukemia between 1989-1991 (27%, 15%, and 13%, respectively). This study shows that the rate of certain types of cancer in infants in the U.S. is increasing. Studies of both genetic and environmental factors are needed to explain these increased rates and the changing distribution of cancer in the first year of life.
    Cancer 04/1998; 82(7):1396-400. · 5.20 Impact Factor
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    ABSTRACT: Basic helix-loop-helix proteins, which are tissue specific (SCL) or broadly expressed (E proteins), interact positively to regulate erythroid specific genes. Here, expression of SCL and two broadly expressed E proteins, E47 and HEB, was high early in erythroid differentiation and declined during maturation. Stimulation of erythroid progenitors/precursors with stem cell factor (SCF) enhanced SCL and E protein levels, one mechanism by which SCF may increase erythroid proliferation. Interactions between SCL and E proteins are competed by Id2, which binds and sequesters E proteins. Upregulation of Id2, demonstrated here late in erythroid differentiation, may downregulate genes involved in erythroid proliferation/differentiation. We examined expression of bHLH proteins in transfusion-dependent patients with Diamond-Blackfan anemia (DBA) to determine if these interactions are disrupted. In erythroblasts from patients, expression of SCL protein and mRNA was normal and SCL increased in response to SCF. However, E47 and HEB protein levels were significantly decreased. Id2 was strongly expressed in patients. Through reduction of SCL/E protein heterodimer formation, abnormal levels of bHLH transcription factors may affect expression of erythroid specific genes, such as beta globin. Stimulation of Diamond-Blackfan cells with SCF partially compensated for this defect, enhancing expression of E47, HEB, and SCL. SCF may function to increase SCL/E protein heterodimer formation, which may be one of the mechanisms through which SCF stimulates erythroid proliferation/differentiation in DBA.
    Blood 10/1997; 90(5):2068-74. · 9.78 Impact Factor
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    ABSTRACT: High breast cancer mortality rates have been reported in the northeastern part of the United States, with recent attention focused on Long Island, New York. In this study, the authors investigate whether the high breast cancer mortality is evenly spread over the Northeast, in the sense that any observed clusters of deaths can be explained by chance alone, or whether there are clusters of statistical significance. Demographic data and age-specific breast cancer mortality rates for women were obtained for all 244 counties in 11 northeastern states and for the District of Columbia for 1988-1992. A recently developed spatial scan statistic is used, which searches for clusters of cases without specifying their size or location ahead of time, and which tests for their statistical significance while adjusting for the multiple testing inherent in such a procedure. The basic analysis is adjusted for age, with further analyses examining how the results are affected by incorporating race, urbanicity, and parity as confounding variables. There is a statistically significant and geographically broad cluster of breast cancer deaths in the New York City-Philadelphia, Pennsylvania, metropolitan area (p = 0.0001), which has a 7.4% higher mortality rate than the rest of the Northeast. The cluster remains significant when race, urbanicity, and/or parity are included as confounding variables. Four smaller subclusters within this area are also significant on their own strength: Philadelphia with suburbs (p = 0.0001), Long Island (p = 0.0001), central New Jersey (p = 0.0001), and northeastern New Jersey (p = 0.0001). The elevated breast cancer mortality on Long Island might be viewed less as a unique local phenomenon and more as part of a more general situation involving large parts of the New York City-Philadelphia metropolitan area. The several known and hypothesized risk factors for which we could not adjust and that may explain the detected cluster are most notably age at first birth, age at menarche, age at menopause, breastfeeding, genetic mutations, and environmental factors.
    American Journal of Epidemiology 08/1997; 146(2):161-70. · 4.78 Impact Factor
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    ABSTRACT: Erythropoietin (Epo) induces a dose-dependent increase in intracellular free Ca2+ ([Ca2+]i) in human erythroblasts, which is dependent on extracellular Ca2+ and blocked by high doses of nifedipine or Ni2+. In addition, pretreatment of human erythroblasts with mouse antihuman erythropoietin receptor antibody but not mouse immunopure IgG blocked the Epo-induced [Ca2+]i increase, indicating the specificity of the Ca2+ response to Epo stimulation. In this study, the erythropoietin-regulated calcium channel was identified by single channel recordings. Use of conventional whole cell patch-clamp failed to detect Epo-induced whole cell Ca2+ current. To minimize washout of cytosolic constituents, we next used nystatin perforated patch, but did not find any Epo-induced whole cell Ca2+ current. Using Ba2+ (30 mmol/L) as charge carrier in cell-attached patches, we detected single channels with unitary conductance of 3.2 pS, reversal potential of +72 mV, and whose unitary current (at +10 mV) increased monotonically with increasing Ba2+ concentrations. Channel open probability did not appreciably change over the voltage range (-50 to +30 mV) tested. Epo (2 U/mL) increased both mean open time (from 4.27 +/- 0.75 to 11.15 +/- 1.80 ms) and open probability (from 0.26 +/- 0.06 to 2.56 +/- 0.59%) of this Ba(2+)-permeable channel. Our data strongly support the conclusion that the Epo-induced [Ca2+]i increase in human erythroblasts is mediated via Ca2+ entry through a voltage-independent Ca2+ channel.
    Blood 02/1997; 89(1):92-100. · 9.78 Impact Factor
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    ABSTRACT: Clinical trials have demonstrated that use of mammographic screening and advances in therapy can improve prognosis for women with breast cancer. We determined the trends in breast cancer mortality rates, as well as incidence and survival rates by extent of disease at diagnosis, for white women in the United States and considered whether these trends are consistent with widespread use of such beneficial medical interventions. We examined mortality data from the National Center for Health Statistics and incidence and survival data by extent of disease from the Surveillance, Epidemiology, and End Results Program of the National Cancer Institute, all stratified by patient age, using statistical-regression techniques to determine changes in the slope of trends over time. The age-adjusted breast cancer mortality rate for U.S. white females dropped 6.8% from 1989 through 1993. A significant decrease in the slope of the mortality trend of approximately 2% per year was observed in every decade of age from 40 to 79 years of age. Trends in incidence rates were also similar among these age groups: localized disease rates increased rapidly from 1982 through 1987 and stabilized or increased more slowly thereafter; regional disease rates decreased after 1987; and distant disease rates have remained level over the past 20 years. Three-year relative survival rates increased steadily and significantly for both localized and regional disease from 1980 through 1989 in all ages, with no evidence of an increase in slope in the late 1980s. The decrease in the diagnosis of regional disease in the late 1980s in women over the age of 40 years likely reflects the increased use of mammography earlier in the 1980s. The increase in survival rates, particularly for regional disease, likely reflects improvements in systemic adjuvant therapy. Statistical modeling indicates that the recent drop in breast cancer mortality is too rapid to be explained only by the increased use of mammography; likewise, there has been no equivalent dramatic increase in survival rates that would implicate therapy alone. Thus, indications are that both are involved in the recent rapid decline in breast cancer mortality rates in the United States.
    JNCI Journal of the National Cancer Institute 12/1996; 88(21):1571-9. · 14.34 Impact Factor
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    ABSTRACT: Erythropoietin induces a dose-dependent increase in cytosolic calcium in human erythroblasts that is mediated by a voltage-independent Ca2+ channel. Inhibition of this response to erythropoietin by pertussis toxin suggests involvement of guanine nucleotide-binding regulatory proteins (G-proteins). The role of G-proteins in regulation of the erythropoietin-modulated Ca2+ channel was delineated here by microinjection of G-protein modulators or subunits into human erythroid precursors. This is the first report on the use of microinjection to study erythropoietin signal transduction in normal precursor cells. Fura-2 loaded day-10 burst-forming units-erythroid-derived erythroblasts were used for microinjection and free intracellular calcium concentration ([Ca(i)]) was measured with digital video imaging. BCECF (1,2',7'-bis(2-carboxyethyl)-5-(and -6-)-carboxyfluorescein) was included in microinjectate, and an increase in BCECF fluorescence was evidence of successful microinjection. Cells were microinjected with nonhydrolyzable analogues of GTP, GTPgammaS or GDPbetaS, which maintain the alpha subunit in an activated or inactivated state, respectively. [Ca(i)] increased significantly in a dose-dependent manner after microinjection of GTPgammaS. However, injection of GDPbetaS blocked the erythropoietin-induced calcium increase, providing direct evidence that activation of a G-protein is required. To delineate which G-protein subunits are involved, alpha or betagamma transducin subunits were purified and microinjected as a sink for betagamma or alpha subunits in the erythroblast, respectively. Transducin betagamma, but not alpha, subunits eliminated the calcium response to erythropoietin, demonstrating the primary role of the alpha subunit. Microinjected antibodies to Gi(alpha)2, but not Gi(alpha)1 or Gi(alpha)3, blocked the erythropoietin-stimulated [Ca(i)] rise, identifying Gi(alpha)2 as the subunit involved. This was confirmed by the ability of microinjected recombinant myristoylated Gi(alpha)2, but not Gi(alpha)1 or Gi(alpha)3 subunits, to reconstitute the response of pertussis toxin-treated erythroblasts to erythropoietin. These data directly demonstrate a physiologic function of G-proteins in hematopoietic cells and show that Gi(alpha)2 is required in erythropoietin modulation of [Ca(i)] via influx through calcium channels.
    Journal of Clinical Investigation 11/1996; 98(8):1728-36. · 12.81 Impact Factor
  • L M Wun, E J Feuer, B A Miller
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    ABSTRACT: A number of studies have attributed much of the sharp increase in breast cancer incidence in the United States during the 1980s to the increased detection through mammography. The most recent breast cancer data from the US National Cancer Institute's Surveillance, Epidemiology, and End Results (SEER) program show that the incidence trend has slowed, while results from the National Health Interview Survey (NHIS) of 1987 and 1990 indicate that the percentage of women receiving mammograms continues to increase. This phenomenon suggested the need to reassess the relationship between increasingly early detection of breast cancer and overall incidence trends. A polynomial age-cohort model was used to establish the secular trend in incidence rates excluding the impact of recent increases in detection due to the rising use of mammography. Based on the model, the incidence trend in the youngest age group (40 to 49 years) would peak and then begin to decline in the early 1980s. This pattern would manifest itself later in successively older age groups as these younger cohorts age. Breast cancer trends are seen to be generally consistent with the impact of the increased use of mammography when its effect is superimposed upon the background of declining or slowing secular trends. These results support previous reports linking incidence rates with the increase in screening-mammography.
    Cancer Causes and Control 04/1995; 6(2):135-44. · 3.20 Impact Factor
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    ABSTRACT: Cancer incidence rates have been reported to be increasing in the United States, although trends vary according to form of cancer. We identify the cancers accounting for the rising incidence, quantify the changes that have occurred from the mid-1970s to the early 1990s, and contrast incidence and mortality trends to provide clues to the determinants of the temporal patterns. Sex-, race-, and age-specific and age-adjusted incidence rates for the 5-year periods 1987-1991 versus 1975-1979 were calculated for 28 cancers among men and 30 cancers among women using data from the Surveillance, Epidemiology, and End Results (SEER) Program of cancer registration covering about 10% of the U.S. population. Similar rates were computed using national mortality data. Cancers were ranked according to the change in incidence rates over the two periods. Age-adjusted incidence rates for all cancers combined increased by 18.6% among males and 12.4% among females from 1975-1979 to 1987-1991, due largely to rising rates for prostate cancer among men and for breast and lung cancers among women. National mortality rates for all cancers combined rose less steeply, 3% and 6% among men and women, respectively, driven mostly by continuing increases in lung cancer mortality, while death rates for the majority of the cancers were steady or declining. Total cancer incidence rose at all ages, but with different tumors responsible for the increases at different ages: leukemia and brain/nervous system cancer among children; testicular cancer, nonmelanoma skin cancer (largely Kaposi's sarcoma), non-Hodgkin's lymphoma, and melanoma among young and middle-aged adults; and prostate, breast, and lung cancers among older individuals. In contrast, mortality rates for all cancers combined declined among both males and females under age 55 years, increasing only among older persons. Trends in cancer incidence and mortality differ. For most cancers, incidence rates are rising, while mortality rates are generally stable or declining. Much of the recent increase in cancer incidence can be explained by known factors. Improved detection appears to account for most of the increases in breast cancer among women and prostate cancer among men. On the other hand, cigarette smoking is the major determinant of the rise in lung cancer among women, acquired immunodeficiency syndrome has led to increases in non-Hodgkin's lymphoma and Kaposi's sarcoma among young and middle-aged men, and sunlight exposure patterns have affected the trends in melanoma. Some trends remain unexplained, however, and may reflect changing exposures to carcinogens yet to be identified and clarified.
    JNCI Journal of the National Cancer Institute 03/1995; 87(3):175-82. · 14.34 Impact Factor
  • B A Miller, L L Bell, C J Lynch, J Y Cheung
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    ABSTRACT: We have reported that erythropoietin induces a dose-dependent increase in cytosolic calcium ([Cai]) in single human peripheral blood BFU-E derived erythroblasts which is specific for stage of differentiation and that this increase is modulated by erythropoietin through an ion channel permeable to Ca2+. Here, the role of protein phosphorylation in the increase in intracellular free calcium [Cai] stimulated by erythropoietin was studied with digital video imaging. Preincubation of day 10 erythroblasts with a broad inhibitor of serine/threonine and tyrosine kinases, staurosporine (100 nM), blocked the increase in [Cai] over 20 min following erythropoietin stimulation. However, erythropoietin-induced calcium influx was unaffected by preincubation of cells with specific inhibitions of protein kinase C (calphostin C) or the cAMP- or cGMP-dependent kinases (KT 5720, HA 1004), and [Cai] did not increase following stimulation with phorbol 12-myristate 13-acetate (PMA) or dibutyryl cAMP. These results suggest that neither protein kinase C nor protein kinase A mediate the erythropoietin-induced [Cai] increase. In contrast, preincubation with genistein, a tyrosine kinase inhibitor, blocked the erythropoietin induced increase in [Cai]. To further study calcium entry in erythroblasts, we determined mastoparan, a peptide from wasp venom, induced a dose-dependent rise in [Cai] in erythroblasts which required external calcium. Stimulation of erythroid precursors with 10 microM mastoparan resulted in an increase in [Cai] from 52 +/- 3 nM to 214 +/- 36 nM which peaked at 20 min. The mastoparan-induced [Cai] increase was also dependent on tyrosine phosphorylation since it was blocked by preincubation with genistein. These results demonstrate that both erythropoietin and mastoparan stimulate calcium entry by a mechanism which has a genistein sensitive step and suggest that tyrosine kinase activation is required for the rise in [Cai] to occur.
    Cell Calcium 01/1995; 16(6):481-90. · 4.33 Impact Factor
  • K C Chu, B A Miller, E J Feuer, B F Hankey
    [Show abstract] [Hide abstract]
    ABSTRACT: U.S. cancer mortality data derived from information recorded on death certificates are frequently relied upon as an indicator of progress against cancer. A limitation of this measure is the lack of information pertaining to the onset of disease, such as year-of-diagnosis, age-at-diagnosis, stage of disease at diagnosis and histology of lesions. However, population-based cancer registries collect these types of data and allow the calculation of an incidence-file based mortality rate. This incidence-based mortality rate allows a partitioning of mortality by variables associated with the cancer onset. Breast cancer incidence-based mortality measures are created and compared to mortality rates based on death certificates over a comparable time period. Novel mortality measures, such as mortality rates by stage-at-diagnosis, age-at-diagnosis and year-of-diagnosis, are used to illustrate the value of this approach.
    Journal of Clinical Epidemiology 01/1995; 47(12):1451-61. · 5.33 Impact Factor

Publication Stats

2k Citations
636.54 Total Impact Points

Institutions

  • 2000–2001
    • Penn State Hershey Medical Center and Penn State College of Medicine
      • • Pediatrics
      • • Department of Pathology
      Hershey, Pennsylvania, United States
    • University of Maryland, Baltimore
      • Department of Pathology
      Baltimore, MD, United States
  • 1988–2001
    • Pennsylvania State University
      • • Department of Pediatrics
      • • Department of Cellular and Molecular Physiology
      • • Department of Medicine
      University Park, MD, United States
  • 1998
    • National Institutes of Health
      • Division of Cancer Prevention
      Maryland, United States
  • 1991–1995
    • National Cancer Institute (USA)
      • Division of Cancer Control and Population Sciences
      Maryland, United States
  • 1984–1988
    • Dana-Farber Cancer Institute
      • Department of Pediatric Oncology
      Dana, NC, United States
  • 1987
    • Boston Children's Hospital
      Boston, Massachusetts, United States