Gisela K. Clemons

Circulating Erythroid Progenitors in the Anemia of Prematurity

We studied erythropoiesis in infants with the anemia of prematurity by counting the number of colonies derived from erythroid burst-forming units (BFU-E) in the blood of 11 premature infants before they received transfusions. Colony growth in blood from the infants was compared with growth in blood from adults and umbilical-cord blood from term infants, in the presence of erythropoietin, 0 to 2000 mU per milliliter. Addition of increasing concentrations of erythropoietin resulted in a stepwise increase in the number of colonies derived from BFU-E (P less than 0.0005) of all three groups of subjects. Cultures stimulated with 2000 mU of erythropoietin yielded 28.1 +/- 7.6, 88.0 +/- 19.4, and 121.0 +/- 22.5 bursts (mean +/- SE) per 10(5) cells plated in blood from adults, blood from premature infants, and cord blood, respectively. Although more BFU-E-derived colonies appeared when 200 or 2000 mU were present per milliliter in cultures of the infants blood and cord blood, the intrinsic responsiveness of BFU-E to erythropoietin was similar in all groups. Although the mean hematocrit was 26 percent, mean serum erythropoietin concentrations (+/- SD) in the infants (20.7 +/- 10.0 mU per milliliter) were not significantly different from those in the adult controls (24.0 +/- 6.5). We conclude that progenitor cells committed to erythroid differentiation are present during the anemia of prematurity, and that the intrinsic responsiveness of the circulating BFU-E pool to erythropoietin is normal. These results implicate inadequate production of erythropoietin as the cause of the anemia of prematurity and suggest that recombinant erythropoietin might provide a therapeutic alternative to transfusion for symptomatic babies with this condition.

Temporal Response of Immunoreactive Erythropoietin to Acute Hypoxemia in Fetal Sheep

ABSTRACT. Acute hypoxemia was produced in chronically catheterized sheep fetuses to determine the response time necessary to increase plasma immunoreactive erythropoietin (Ep) concentration. Sodium nitrite (0.2 mM) was infused via a fetal vein to induce fetal hypoxemia. The resultant fetal methemoglobinemia was associated with a predictable, incremental decrease in arterial oxygen content. Twelve nitrite infusions were performed in eight fetal sheep preparations (gestational ages 115-146 days). Mean methemoglobin level increased to 33% of total Hb after 1- 2 h of NaNO2 infusion. These results were compared to those obtained in nine control studies in eight fetuses in which no change was observed for plasma Ep, arterial oxygen content, Pao2, pHa, or whole blood lactate. In the nitrite infused group, however, a significant and progressive increase in mean plasma Ep level over baseline levels was observed during the 4th and 5th h of hypoxemia (p<0.01). This change in Ep was significantly greater compared to the control group. These results, however, were confounded by the concomitant development of a lactic acidemia secondary to the fetal hypoxemia. To examine the theoretic possibility that lactic acidemia may primarily affect fetal Ep levels, an additional group of five fetuses was infused with L-lactic acid for the same time period. Although the decrements in pHa and whole blood lactate levels achieved in these fetuses were in excess of those observed during the nitrite infusions, this possibility was ruled out since no change in fetal plasma Ep levels occurred. We conclude that during the 4th h of acute fetal hypoxemia a predictable, progressive increase in plasma Ep level is observed. This response of plasma Ep to hypoxemia in late gestation fetal sheep is qualitatively similar to that observed in adult animals, thus demonstrating developmental maturity of the fetus.

Direct relationship of antepartum glucose control and fetal erythropoietin in human Type 1 (insulin-dependent) diabetic pregnancy

SummaryIn the present study the antepartum relationship between maternal diabetic glucose control and fetal hypoxaemia was examined in 44 Type 1 (insulin-dependent) diabetic and 23 non-diabetic control pregnancies. Maternal HbA1C was used to assess maternal integrated blood glucose control while fetal metabolic control was evaluated by antepartum glucose, insulin, and C-peptide determinations in amniotic fluid at elective caesarean delivery. Fetal hypoxaemia was assessed indirectly by fetal umbilical vein plasma erythropoietin level at delivery. A prospectively developed statistical pathway model was used to examine the relationship of these variables. In applying forced stepwise multiple regression with this model, we observed in the diabetic subjects that mean maternal HbA1C during the last month of pregnancy correlated significantly with fetal umbilical venous erythropoietin at delivery (r=0.57, p<0.001). Additional significant contributions to umbilical venous erythropoietin were found for amniotic fluid glucose and amniotic fluid insulin when these two independent variables were added in stepwise fashion (p<0.01). We conclude that in diabetic pregnancy, antepartum control of maternal hyperglycaemia is a significant factor associated with fetal hypoxaemia. We speculate that this effect is mediated through perturbations which accelerate fetal metabolism and which is expressed by amniotic fluid levels of glucose and insulin.

Increased immunoreactive erythropoietin in cord serum after labor

Since several hours of hypoxemia in fetal animals is sufficient to cause an increase in the plasma erythropoietin level and since labor may be associated with fetal hypoxemia, this study was undertaken to determine if erythropoietin levels in cord blood were higher in fetuses subjected to labor. Two groups of term (37 to 41 weeks) singleton pregnancies were compared: (1) those delivered by elective repeat cesarean section without prior labor (n = 18) and (2) those delivered vaginally (n = 23). Erythropoietin was measured by a radioimmunoassay in which a highly purified human erythropoietin (70,000 U/mg of protein) was used and which has a sensitivity limit of 4 to 5 mU/ml. The mean cord serum erythropoietin level was higher in pregnancies with labor (46 +/- 34 mU/ml, mean +/- SD) compared to those without (26 +/- 10, p less than 0.02). There were no differences between the two groups for maternal age, gestational age, birth weight, infant sex, or Apgar scores. No association of erythropoietin with either gestational age or sex was found. In 11 pregnancies without labor, comparisons were made among simultaneously obtained samples of umbilical arterial plasma, umbilical venous plasma, and mixed cord serum. Although there were no differences between umbilical arterial and umbilical venous plasma erythropoietin levels (21.3 +/- 9.3 versus 19.0 +/- 7.8 mU/ml), mixed cord serum was inexplicably higher (24.4 +/- 9.5 mU/ml, p less than 0.01). We concluded that in uncomplicated pregnancies the duration and intensity of labor are sufficient to cause an increase in the fetal erythropoietin level at delivery.

Postnatal changes in serum immunoreactive erythropoietin in relation to hypoxia before and after birth

To assess the immediate postnatal changes of serum immunoreactive erythropoietin (EP) in infants born after acute or chronic fetal hypoxia, and to estimate the rate of EP disappearance, we studied EP concentration, measured by double-antibody radioimmunoassay, in cord venous plasma and in serum at a mean age of 8 hours in a control group (n = 9) and in three patient groups: (1) infants with polycythemia (n = 10), (2) infants born to mothers with preeclampsia of pregnancy, without (n = 22) or with (n = 11) acidosis at birth, and (3) infants with acute birth asphyxia (n = 19), seven of whom had postnatal hypoxia. In all patient groups, cord venous EP was elevated in comparison with values in control infants. No change was found in EP level between birth and 8 hours in control infants (geometric mean in cord and 8-hour sample: 20 and 16 mU/ml, not significant) or in acutely asphyxiated infants with postnatal hypoxia (122 and 72 mU/ml, not significant), whereas the EP level decreased in all other groups: infants with polycythemia (123 to 24 mU/ml, p less than 0.001), nonacidotic infants (78 to 26 mU/ml, p less than 0.001) and acidotic infants (176 to 38 mU/ml, p less than 0.001) of the preeclampsia group, and acutely asphyxiated infants without postnatal hypoxia (58 to 30 mU/ml, p less than 0.001). The mean (+/- SD) half-time of EP disappearance was 2.6 +/- 0.5 hours in infants with polycythemia and 3.7 +/- 0.9 hours in infants of the preeclampsia group.

Cord Plasma Vasopressin, Erythropoietin, and Hypoxanthine as Indices of Asphyxia at Birth

ABSTRACT: To assess the value of cord plasma arginine vasopressin (AVP), erythropoietin (EP), and hypoxanthine (HX) as indices of asphyxia, we studied 62 infants of mothers with preeclampsia, 34 acutely asphyxiated infants, with 5-min Apgar score ≤6 and/or umbilical arterial pH ≤7.05, and 38 control infants. Umbilical arterial AVP in the asphyxia group (geometric mean; 95% confidence interval: 180; 92–350 pg/ml) was higher than in the control group (23; 8–466, p = 0.002) and correlated with umbilical arterial pH (r = −0.447, p = 0.028). AVP levels in the preeclampsia group did not differ from controls. Cord venous EP was higher in infants delivered by elective cesarean section from women with severe preeclampsia (115; 75–177 mU/ml, p < 0.001) than in control infants (23; 18–27); in the whole group EP correlated with pH (r = −0.493, p < 0.001). EP in the asphyxia group was similar (46; 35–65) to controls (40; 33–47) and did not correlate with pH. Cord arterial HX in the preeclampsia group was similar to controls (12.3; 9.5–16.0 μmol/liter), but elevated in the asphyxia group (23.7; 17.6–31.8, p = 0.001), in which HX correlated with pH (r = 0.558, p = 0.008) and AVP (r = 0.588, p = 0.005). EP did not correlate with AVP or HX in any group, nor did any of the variables correlate with the Apgar score. We conclude that cord plasma AVP and HX reflect acute asphyxia, whereas EP is elevated after more prolonged hypoxia.

Development and preliminary application of a homologous radioimmunoassay for bullfrog prolactin

Abstract A homologous radioimmunoassay (RIA) with high sensitivity was developed for bullfrog prolactin (fPRL). The fPRL used for immunization, iodination, and reference preparation was obtained by disc gel electrophoresis of frog anterior pituitaries. Bullfrog growth hormone (fGH) showed minimal cross-reactivity, and plasma from hypophysectomized frogs had no detectable immunoreactive PRL. The antiserum to fPRL inactivated biologically active PRL in tadpoles and caused accelerated metamorphosis [Clemons, G. K., and Nicoll, C. S. (1977). Gen. Comp. Endocrinol. 31, 495–497.]. The levels of immunoreactive fPRL were determined in individual whole pituitaries and in pooled plasma samples from Rana catesbeiana tadpoles through metamorphic climax (Taylor-Kollros stages XXI–XXV). Pituitary PRL rose slowly from the emergence of the forelegs through stage XXIII and then increased markedly in stage XXIV. Circulating levels paralleled pituitary content, except that the rise during stage XXIV was less pronounced.

Amniotic fluid erythropoietin predicts fetal distress in Rh-immunized pregnancies

Repeated amniotic fluid erythropoietin measurements in 23 Rh-immunized pregnancies were done to evaluate erythropoietin levels of amniotic fluid as an indicator of fetal distress (umbilical artery, pH 7.14 or less, or 1-minute Apgar score of 4 or less). Amniotic fluid erythropoietin levels did not vary significantly between 168 and 273 gestational days in the pregnancies without fetal distress. Increasing levels of amniotic fluid erythropoietin predicted highly reliably severe fetal distress at birth. Whether erythropoietin levels of amniotic fluid can also predict fetal distress in other pathologic pregnancies needs further study.

Effect of mammalian thyrotropin releasing hormone on prolactin secretion by bullfrog adenohypophyses in vitro

The effects of the mammalian thyrotropin-releasing hormone (TRH) on the secretion of prolactin by bullfrog adenohypophysis in vitro were investigated in short-term incubation and 24-hr organ culture experiments. Prolactin in the medium or in the incubated tissue was measured by either polyacrylamide disc gel electrophoresis and densitometry or by a homologous radioimmunoassay. The TRH was consistently effective in promoting prolactin release in vitro in concentrations of 10 ng/ml to 10 μg/ml. The tripeptide also caused an increase in the tissue prolactin content over a wide range of concentrations. These results indicate that TRH may function as a prolactin-releasing factor in the bullfrog.

Effects of antisera to bullfrog prolactin and growth hormone on metamorphosis of Rana catesbeiana tadpoles

Abstract Rabbit antisera to bullfrog prolactin (fPRL) or growth hormone (fGH), or normal rabbit serum (NRS) were injected intraperitoneally into late prometamorphic tadpoles of Rana catesbeiana . The injections were given once a week for 5 weeks at a dose of 50 μl per animal. The fPRL antiserum accelerated metamorphic climax and the GH antiserum did not. The results indicate that endogenous PRL has antimetamorphic effects in this species.