PubMed Health⌕ Search

Biomedical subjects

R D Christensen

Publications and source records attributed to R D Christensen.

At least 55 records · Page 3Linked to original sources
PubMed1999

A bone marrow biopsy technique suitable for use in neonates.

Thrombocytopenia and neutropenia are common among neonates in intensive care units. Bone marrow aspirations are sometimes performed as part of their evaluation. However, marrow biopsies have not been reported from living neonates. Since architecture and cellularity cannot generally be accurately assessed from marrow aspirates, we devised a biopsy technique which we successfully applied to five cytopenic neonates (three with severe persistent thrombocytopenia and two with idiopathic neutropenia). This technique used a 19 gauge, half-inch Osgood needle to obtain bone marrow clots from the tibias of small preterm neonates which enabled the assessment of marrow cellularity and architecture. On the basis of our initial experience we have ceased using the traditional bone marrow aspiration technique in neonates and now use this technique exclusively.

Biopsy, Needle↗
PubMed1999

Distribution of granulocyte colony-stimulating factor (G-CSF) and G-CSF-receptor mRNA and protein in the human fetus.

Granulocyte colony-stimulating factor (G-CSF) promotes neutrophil proliferation, differentiation, and survival. It acts by binding to specific cell-surface receptors (G-CSF-R), which are expressed on cells of granulocytic lineage, human endothelial cells, and placenta. It has been postulated that the administration of recombinant G-CSF (rG-CSF) to preterm neonates might be useful in treating infections or in reducing nosocomial infections. Whereas it is known that G-CSF and G-CSF-R are present in the developing fetal bone marrow and liver, no information is available as to the existence or distribution of nonhematopoietic G-CSF-R in other tissues of the developing human fetus. We hypothesized that G-CSF and its receptor might be expressed in various fetal tissues, as has been shown for other growth factors such as erythropoietin and fibroblast growth factor. Therefore, we studied the anatomical distribution of mRNA-encoding G-CSF and G-CSF-R, using RT-PCR and in situ RT-PCR in a variety of human fetal tissues ranging from 8 to 24 weeks postconception. The cellular distributions of the corresponding proteins were determined by immunohistochemistry. Both G-CSF and G-CSF-R were present in nearly every organ and tissue examined, but in discrete cellular localizations. G-CSF-R in kidney and intestine underwent changes in anatomical distribution with fetal development. These results indicate that G-CSF and G-CSF-R have wide anatomical expression in the developing human fetus.

Fetus↗
PubMed1999

The distribution of granulocyte-macrophage colony-stimulating factor and its receptor in the developing human fetus.

The purpose of the present study was to determine the distribution of granulocyte-macrophage colony-stimulating factor (GM-CSF) and its receptor (GM-CSF-R) in the human fetus. We used reverse transcription PCR to detect GM-CSF and GM-CSF-R mRNA in human fetal organs at 8 and 16 wk postconception, and cell-specific protein expression was localized in tissues by immunohistochemistry. GM-CSF was also measured by ELISA in paired samples of spinal fluid and plasma. GM-CSF mRNA and/or protein were detected in lung macrophages, spleen, adrenal cortex, placenta, and CNS including neurons and astrocytes. GM-CSF was detected by ELISA in 10 of the 39 cerebrospinal fluid samples tested. GM-CSF-R mRNA expression was present in all organs tested. Immunoreactivity for GM-CSF-R in most organs was limited to macrophages, but, brain, neurons and glial cells showed immunoreactivity. We conclude that GM-CSF is produced in lung, spleen, adrenal, placenta, and neural tissues during human fetal development and that GM-CSF-responsive cells include macrophages, neurons, and glial cells.

Adolescent↗
PubMed1999

Erythropoietin in the cerebrospinal fluid of neonates who sustained CNS injury.

We previously reported that erythropoietin (Epo) is present in human cerebrospinal fluid (CSF). It is not known whether CSF Epo concentrations change under conditions of CNS injury or, if so, whether this change reflects loss of blood-brain barrier integrity or increased CNS Epo synthesis. We hypothesized that CSF Epo increases in conditions of neural injury including hypoxia, meningitis, and intraventricular hemorrhage (IVH) and that CSF Epo concentrations are independent of plasma Epo concentrations. To test these hypotheses, Epo concentrations were measured in 122 paired CSF and blood samples obtained from neonates and children categorized as follows: 16, asphyxia; 31, meningitis; 11, IVH; 41, controls. Twelve infants treated with recombinant Epo (rEpo) and 11 additional samples from children with miscellaneous neurologic problems were also evaluated. CSF and plasma Epo concentrations were significantly higher in asphyxiated infants than in controls (225.0+/-155.0 versus 4.5+/-0.5 mU/mL; mean +/- SEM, p < 0.05, respectively, in CSF; 1806.7+/-1254 versus 5.2+/-0.5, p < 0.05 in plasma). Neonates with IVH had higher CSF Epo concentrations than controls (p < 0.01) but did not have higher plasma Epo concentrations than controls. Patients with meningitis did not have elevated CSF or plasma Epo concentrations. There was no correlation between CSF and plasma Epo concentrations in infants treated with rEpo. We conclude that Epo is selectively increased in the CSF by hypoxia, less so by IVH, and not at all by meningitis. rEpo treatment does not elevate CSF Epo. These findings suggest that rEpo does not cross the blood-brain barrier and that hypoxia induces increased CNS synthesis of Epo.

Asphyxia Neonatorum↗
PubMed1999

Concentrations of granulocyte colony-stimulating factor in human milk after in vitro simulations of digestion.

Human milk contains proteins that survive digestion in the neonatal gastrointestinal tract. Our group and others have reported that granulocyte colony-stimulating factor (G-CSF), a hematopoietic cytokine that influences neutrophil proliferation and differentiation, is present in human milk. We also reported that specific receptors for G-CSF are expressed on the villous enterocytes of neonates. However, the physiologic role of milk-borne G-CSF is not known. Thus, we sought to evaluate the capacity of human milk to protect G-CSF against proteolytic degradation after exposure to gastric secretions obtained from preterm (PT) and term (T) neonates at pH concentrations of 3.2, 5.8, and 7.4. Specifically, we examined degradation of 1) endogenous G-CSF in PT (n = 15) and T (n = 15) human milk; 2) recombinant human G-CSF (rhG-CSF) added to a protein-free buffer (n = 10, 5 PT and 5 T); and 3) rhG-CSF added to human milk (n = 12, 6 PT and 6 T), various commercially prepared infant formulas (n = 15), and cow's milk (n = 5). Endogenous G-CSF and rhG-CSF added to human milk resisted degradation at 1 and 2 h. However, when rhG-CSF was added to commercial formulas, >95% was degraded at 1 and 2 h at each pH level. Similarly, approximately 60% of rhG-CSF added to cow's milk was degraded at I and 2 h. We conclude that 1) endogenous G-CSF and rhG-CSF added to human milk are protected from degradation after exposure to gastric secretions at physiologic pH levels, 2) rhG-CSF added to infant formulas is not protected from degradation, and 3) it is likely that the G-CSF present in human milk is biologically available to the neonate.

Digestion↗
PubMed1998

A randomized pilot trial of administration of granulocyte colony-stimulating factor to women before preterm delivery.

OBJECTIVE: The object was to determine whether administering recombinant human granulocyte colony-stimulating factor to women before a preterm delivery (< or = 30 weeks' gestation) would improve fetal neutrophil production and neonatal outcome without adverse maternal effects. STUDY DESIGN: A single dose (25 microg/kg) of recombinant human granulocyte colony-stimulating factor or placebo was given to 26 women in preterm labor in a randomized, double-blind study. Neutrophil production was assessed by marrow aspirations of neonates 24 hours after delivery, blood neutrophil counts were measured in neonates for 1 week, and Scores for Neonatal Acute Physiologic State were calculated on days 1 and 7 of life. RESULTS: Of the 26 women enrolled, 16 were delivered of 20 infants within 3 weeks of receiving recombinant human granulocyte colony-stimulating factor. We found no apparent adverse effect on pregnancy duration or maternal comfort. The neonates born to mothers treated with recombinant human granulocyte colony-stimulating factor had a significantly greater marrow proliferative pool (P < .05 vs placebo) and a significant improvement in Scores for Neonatal Acute Physiologic State between days 1 and 7 (P < .02). CONCLUSIONS: Recombinant human granulocyte colony-stimulating factor administration to women before preterm delivery appears to have no significant immediate adverse effects on either the mother or the infant, and it could increase fetal neutrophil production and improve neonatal outcome.

Adult↗
PubMed1998

Tissue distribution of erythropoietin and erythropoietin receptor in the developing human fetus.

OBJECTIVE: Erythropoietin receptors (Epo-R) have been demonstrated on several nonhematopoietic cell types in animal models and in cell culture. Our objective was to determine the tissue distribution and cellular specificity of erythropoietin (Epo) and its receptor in the developing human fetus. STUDY DESIGN: The expression of Epo and Epo-R mRNA was ascertained by RT-PCR for organs ranging in maturity from 5 to 24 weeks postconception. The cellular location of protein immunoreactivity was then determined using specific antiEpo and antiEpo-R antibodies. Antibody specificity was established by Western analysis. RESULTS: mRNA for Epo and Epo-R was found in all organs in the first two trimesters. Immunolocalization of Epo was limited to the liver parenchymal cells, kidney interstitial cells and proximal tubules, neural retina of the eye, and adrenal cortex. As development progressed, immunoreactivity in the kidney became more prominent. In contrast, immunoreactivity for Epo-R was widespread throughout the body, in cell types including endothelial cells, myocardiocytes, macrophages, retinal cells, cells of the adrenal cortex and medulla, as well as in small bowel, spleen, liver, kidney, and lung. CONCLUSIONS: The distribution of Epo and its receptor is more widespread in the developing human than was initially postulated. Epo-R is expressed on many cell types during early fetal development, leading us to speculate that Epo acts in concert with somatic growth and development factors during this period. Further investigation is required to understand the nonhematopoietic role of Epo during human development.

Adrenal Cortex↗
PubMed1998

The first-appearance of neutrophils in the human fetal bone marrow cavity.

We sought to define the time of first-appearance of neutrophils within the developing human bone marrow cavity, and to compare this with the time of appearance of G-CSF and its receptor (G-CSF-R) at that site. We hypothesized that the onset of G-CSF production is an initiation signal for neutrophil production within the marrow cavity, and that therefore G-CSF mRNA and G-CSF protein in the marrow cavity would immediately precede the first-appearance of neutrophils. To test this, we determined the time of first-appearance of neutrophils in the clavicular marrow space using a monoclonal antibody against myeloperoxidase (MPOAb), and then validated these findings by flow cytometric analyses, for neutrophil cell-surface markers, of cells flushed from the marrow cavity. After thus defining the time of first-appearance of neutrophils, specific mRNA transcripts for G-CSF and G-CSF-R were sought from clavicles of varying gestational ages, using RT-PCR, and the presence of these proteins in the clavicles were sought using immunohistochemistry. We observed that; (1) MPO+ cells first appeared in the clavicular marrow cavity between the 10 to 11th weeks post-conception, (2) Flow cytometric analyses confirmed that these MPO+ marrow cells included CD11b+, CD15+ neutrophils, (3) Transcripts for G-CSF and G-CSF-R, and the specific G-CSF and G-CSF-R proteins, were present in the clavicles by 6 weeks post-conception, 4 to 5 weeks before the first-appearance of neutrophils. Thus, neutrophils first appear in the human clavicular marrow at 10-11 weeks post-conception, and G-CSF and G-CSF-R are present in the developing bone rudiment preceding the appearance of neutrophils. It is unclear whether neutrophils arise in the marrow cavity in response to the onset of production of G-CSF or to other initiation signals.

Bone Marrow↗
PubMed1998

"Idiopathic neutropenia" in very low birthweight infants.

We employed a standardized investigative approach to evaluate four cases of "idiopathic neutropenia" in very low birthweight infants. The evaluation included maternal anti-neutrophil antibodies, a marrow aspiration, and a three-day trial of recombinant granulocyte colony-stimulating factor (rG-CSF). All patients had neutropenia at or shortly following delivery, and remained neutropenic (generally <500/microl) for 1-9 weeks until rG-CSF was administered. Blood and bone marrow studies in all 4 indicated that the neutropenia was: (i) the kinetic result of diminished neutrophil production; (ii) not alloimmune; (iii) not cyclic; and (iv) not associated with recognized inborn errors, bacterial or viral infections, or medications. All responded to rG-CSF by transiently increasing their blood neutrophils to normal, and the neutropenia resolved in all cases with time. It is likely that "idiopathic neutropenia" in this population represents a common phenotype of several different causes, rather than a single entity. Some cases, perhaps the majority, will respond to a short course of rG-CSF administration.

Diseases in Twins↗
PubMed1998

Recent advances in the pathogenesis and treatment of nonimmune neutropenias in the neonate.

Neutropenia in the neonate is defined as a significantly lower than normal concentration of neutrophils in the peripheral blood and can be broadly classified as immune or nonimmune. Nonimmune neutropenias are those that are not the direct or indirect result of antibodies directed against neutrophils. Recent advances have been made in the pathogenesis and treatment of several types of neonatal neutropenia, including reticular dysgenesis, Schwachman-Diamond syndrome, Kostmann syndrome, neutropenia in infants of hypertensive women, and "idiopathic neutropenia." This review of nonimmune neutropenias highlights these developments.

Female↗
PubMed1998

In vitro effect of dexamethasone phosphate on hematopoietic progenitor cells in preterm infants.

The in vitro effect of dexamethasone on the clonal growth of haematopoietic progenitors in preterm infants was investigated. Concentrations of 10(6)M to 10(9)M were associated with a dose dependent inhibition of colony formation, with the most clinically important effects seen on the earliest erythroid and granulocyte-macrophage colonies. Because of the potential clinical implications of these observations, studies are needed to determine the effects of dexamethasone on haematopoiesis in preterm infants.

Adult↗
PubMed1998

Erythropoietin and erythropoietin receptor in the developing human central nervous system.

We have previously shown the presence of erythropoietin (Epo) within the spinal fluid of normal preterm and term infants, and the presence of Epo receptor (Epo-R) in the spinal cords of human fetuses. It is not known, however: 1) whether cells within the fetal central nervous system (CNS) express Epo; 2) if so, whether this expression changes with development; 3) which cells within the CNS express Epo-R; 4) whether Epo-R expression within the CNS changes with development; and 5) whether Epo-R within the fetal CNS are functional. Expression of mRNA for Epo and Epo-R was sought by reverse transcription-PCR in mixed primary cultures of fetal spinal cords as well as NT2 and hNT cells, human cell lines of neuronal precursors and mature neurons, respectively. Epo was measured by ELISA in spent media from primary cell culture, and immunohistochemistry was used to identify Epo-R on neurons and glia in cell culture, and in brain sections. Developmental changes in Epo and Epo-R expression were sought in spinal cords and brains from fetuses of 7-24 wk postconception by semiquantitative PCR. To assess Epo-R function, NT2 cells were exposed to conditions which stimulate programmed cell death, and rescue from apoptosis by the addition of recombinant Epo was evaluated by nuclear matrix protein ELISA, cell counts, and by Klenow labeling of DNA fragments. Epo and Epo-R mRNA were expressed in mixed primary cultures of neural tissues and NT2 and hNT cells. Epo was detected by ELISA in media removed from mixed cell cultures, and immunohistochemical staining confirmed the presence of Epo-R on neurons and their supporting cells. Semiquantitative PCR revealed no significant change in expression of either Epo or Epo-R in spinal cords between 7 and 16 wk of gestation, with increased expression of Epo and Epo-R in brains from 8 to 24 wk of gestation. Epo mRNA expression from neurons doubled under conditions of hypoxia. Recombinant Epo decreased apoptotic cell death of neurons under conditions of hypoxia. Protein and mRNA for Epo and its receptor are expressed by human neurons and glial cells in spinal cord and brain during fetal development. These receptors appear to have a neuroprotective effect in conditions of hypoxia.

Central Nervous System↗
PubMed1998

Hematopoiesis in the liver and marrow of human fetuses at 5 to 16 weeks postconception: quantitative assessment of macrophage and neutrophil populations.

During human development, the liver and marrow both function as hematopoietic organs, but little is known about differences in the production of macrophages and neutrophils by these two organs. We used immunohistochemical stains to quantify the ratio of neutrophils to macrophages within the liver and the marrow of 16 fetuses from 5 to 16 wk postconception. At 5 wk the liver had a ratio of one granulocyte [myeloperoxidase (MPO)-positive cell] to every 9 +/- 5 (X +/- SD) macrophages (KP-1-positive cells). Between 5 and 16 wk, the granulocyte to macrophage ratio in the liver was constant, whereas it changed markedly in the marrow. Before 8 wk no MPO-positive or KP-1-positive cells were observed in bones. At 10 wk, bones still had no MPO-positive cells, but KP-1-positive cells were abundant. At 11-12 wk the granulocyte to macrophage ratio was 1 to 1 +/- 1, but by 13-16 wk it had increased to 8 +/- 3 MPO-positive cells to one KP-1-positive cell. We hypothesized that at 13-16 wk the abundance of MPO-positive cells in the marrow and their scarcity in the liver was the result of production of granulocyte colony-stimulating factor (G-CSF) and its receptor (G-CSF-R) in the marrow and their absence in the liver. However, by reverse transcriptase-PCR mRNAs for G-CSF and G-CSF-R were positive in both organs at all gestations, and G-CSF and G-CSF-R proteins (by immunohistochemistry) were also abundant in all liver and marrow specimens. We then hypothesized that progenitors in the fetal liver were intrinsically different from those in the marrow, and were unable to generate clones of neutrophils. However, progenitors from the liver produced neutrophils abundantly in culture. Thus, the explanation is likely related to as yet undescribed environmental differences between the fetal liver and marrow.

Bone Marrow↗
PubMed1997

The effect of erythropoietin on the transfusion requirements of preterm infants weighing 750 grams or less: a randomized, double-blind, placebo-controlled study.

BACKGROUND: Clinical trials of erythropoietin (EPO) administration to preterm infants have not focused on infants weighing 750 gm or less, the population most likely to receive multiple transfusions because of large phlebotomy losses. It is unknown whether preterm infants weighing 750 gm or less will respond to EPO by accelerating erythropoiesis, or whether EPO administered to this population will decrease blood transfusions. METHODS: We randomly assigned 28 extremely low birth weight preterm infants (mean +/- SEM: 24.7 +/- 0.3 weeks' gestation, 662 +/- 14 gm birth weight), in the first 72 hours of life, to receive either EPO (200 U/kg/day) or placebo for 14 days and administered transfusions only according to protocol over a 21-day study period. All infants received 1 mg/kg/day iron dextran in their total parenteral nutrition solution during the 14-day treatment period. RESULTS: During the 21-day study period, a lower number and volume of transfusions were received by the EPO recipients (4.7 +/- 0.7 transfusions per patient and 70 +/- 11 ml/kg per patient) than by the placebo recipients (7.5 +/- 1.1 transfusions per patient and 112 +/- 17 ml/kg per patient; p < 0.05, EPO vs placebo), whereas hematocrits remained similar in the two groups. Reticulocyte counts were similar in both groups on day 1 but were greater in the EPO recipients on day 14 (EPO day 1, 351 +/- 53; EPO day 14, 359 +/- 40 x 10(3)/microl; placebo day 1, 334 +/- 64; placebo day 14, 120 +/- 10 x 10(3)/microl; p < 0.01, EPO vs placebo). Serum ferritin concentrations were similar in both groups at the beginning of the study but were greater in the placebo recipients by day 14 (EPO, 262 +/- 44 microg/L; placebo, 593 +/- 92 microg/L; p < 0.01). No adverse effects of EPO or iron were noted. CONCLUSION: The combination of EPO and parenteral iron stimulates erythropoiesis in preterm infants weighing 750 gm or less and results in fewer transfusions during their first 3 weeks of life.

Anemia, Hemolytic↗
PubMed1997

Erythropoietin is present in the cerebrospinal fluid of neonates.

Erythropoietin (Epo) was measured by enzyme-linked immunosorbent assay in 80 cerebrospinal fluid (CSF) samples to determine whether Epo is present in the CSF of infants, CSF Epo concentrations correlate with age, and CSF Epo concentrations correlate with Epo therapy. Epo was present in the CSF of normal neonates. CSF Epo concentrations correlated negatively with increasing age. Recombinant Epo therapy did not affect CSF Epo concentrations, although values ranged somewhat higher in this group.

Adolescent↗
PubMed1996

Assessment of the contribution of the spleen to granulocytopoiesis and erythropoiesis of the mid-gestation human fetus.

Several current textbooks of hematology describe the spleen of the mid-gestation human fetus as a granulocytopoietic and an erythropoietic organ. Although studies in fetal rodents support this view, a convincing demonstration of such in normal human fetuses is lacking. We tested the hypothesis that the human mid-gestation fetal spleen is normally (1) a site of active granulocytopoiesis and erythropoiesis, or (2) a site of production of specific granulocytopoietic or erythropoietic growth factors. This was accomplished using the spleens, livers, and long-bones of 18 human fetuses, 13-22 weeks gestation, immediately following elective, induced, pregnancy terminations. Organs of some of the abortuses were placed directly into formalin for histologic evaluation. Organs from others were subjected to RNA extraction for subsequent probing for specific hematopoietic growth factor mRNA. Cell suspensions were created from the organs of other abortuses for quantification of the absolute number of neutrophils and erythrocytes and their precursors and progenitors. Evidence of active hematopoiesis was present in marrow and liver but not spleen. Transcripts for granulocyte colony-stimulating factor (G-CSF) were detected in the marrow but not the spleen or liver, and transcripts for erythropoietin (Epo) were detected in the liver but not the spleen or marrow. The populations of hematopoietic progenitor cells and normoblasts in the fetal spleen cell were similar to those in fetal blood. Thus, it is likely that the hematopoietic progenitors recovered from fetal spleen cell suspensions are the result of blood within the spleen, rather than from hematopoiesis within the organ. The spleen of mid-gestation human fetuses, unlike the spleen of fetal rats, does not normally function as an active site of granulocytopoiesis or erythropoiesis, nor is it an active site for production of G-CSF or Epo.

Abortion, Induced↗
PubMed1996

Transplacental passage of recombinant human granulocyte colony-stimulating factor in women with an imminent preterm delivery.

OBJECTIVE: We attempted to determine, in a pilot study, whether recombinant human granulocyte colony-stimulating factor, administered to women with an imminent delivery at < or = 30 weeks' estimated gestational age, crosses to the fetal circulation and stimulates fetal neutrophil production. STUDY DESIGN: We measured granulocyte colony-stimulating factor and neutrophil concentrations in the blood of the mothers and in the umbilical venous cord blood after a single intravenous dose (25 micrograms/kg) of recombinant granulocyte colony-stimulating factor given to 11 women in whom preterm delivery was imminent and compared these with values from 34 control women and 35 cord sera. RESULTS: Ten infants were delivered within 30 hours (10.8 +/- 8.9, mean +/- SD) of the granulocyte colony-stimulating factor administration ("early delivery"), and two were delivered after 54 and 108 hours, respectively. In the early delivery group maternal granulocyte colony-stimulating factor concentrations and blood neutrophil levels were higher than in controls. However, no difference was seen in the cord blood neutrophil concentrations. In the late delivery group, although maternal serum and cord blood granulocyte colony-stimulating factor concentrations did not differ from controls, cord blood neutrophil levels were higher (25,900 and 17,700 cells/microliters) than controls (3500 +/- 2000 cells/microliters, p < 0.05) and remained elevated for 1 week. Specifically, the blood neutrophil levels on days 5 and 8 were 12,000 and 17,000, respectively, for these patients, whereas the reference range upper limit for controls was 6000 cells/microliters. CONCLUSION: Administration of recombinant granulocyte colony-stimulating factor to pregnant women with an imminent preterm delivery may result in the transplacental passage of a measurable quantity of granulocyte colony-stimulating factor, an amount that can have a biologic effect on the fetus. These events were most noticeable in those patients who received granulocyte colony-stimulating factor at least 30 hours before being delivered of their infants.

Bacterial Infections↗