Digoxin, a steroidal glycoside that inhibits Na⁺/K⁺-ATPase, is the most commonly prescribed cardiac medication in North America. Blood levels of this drug are routinely measured to reduce the risks of toxicity. Reports questioning the specificity of antisera used in radioimmunoassays for serum digoxin measurements began to appear after 1975¹ when plasma from patients with renal failure, not on glycoside therapy, showed false-positive digoxin levels. Since then, digoxin-like immunoreactive substances (DLIS) have been found in sera from patients with hepatic failure, hypertension, pre-eclampsia, in amniotic fluid and cord blood. Some of the highest values for DLIS have been detected in premature infants, where levels have often exceeded the therapeutic range (0.2-2.0 µg/L) for digoxin. Cord blood has been identified as a rich source of DLIS. Dahl et al² were the first to suggest that a circulating saluretic substance "endoxin", may cause hypertension in salt sensitive rats. Gruber et al³ reported on the existence of digoxin-like factor(s) in the plasma of volume-expanded dogs. Plasma from these dogs inhibited Na⁺/K⁺ATPase activity. A number of other studies have supported the concept that such digoxin-like factors may be of etiological significance in hypertension⁴. In view of these observations, a study was undertaken to isolate and fractionate DLIS from mixed cord blood and determine whether or not any of this digoxin-like material possessed Na⁺/K⁺-ATPase inhibitory properties. Cord blood collected in the Grace Hospital Maternity Unit (Vancouver, BC), was pooled and DLIS extracted using C₁₈,R-Sep Paks. Extracts were resolved by high performance liquid chromatography (HPLC) into several fractions containing digoxin equivalent immunoactivity as measured by radioimmunoassay (RIA). A number of steroids and bile acids (dehydroepi-androsterone-sulfate, cortisone, Cortisol, deoxycortisone, ∆⁴androstene-dione, progesterone and glycochenodeoxycholic acid) cross-reacted with digoxin antisera and had HPLC retention times similar to DLIS-containing fractions. The ability of HPLC generated DLIS positive cord blood fractions to inhibit Na⁺/K⁺-ATPase activity was determined in three different assay systems; red cell ⁸⁶Rb uptake canine kidney-Na⁺/K⁺-ATPase and red cell membrane-Na⁺/K⁺-ATPase. At least six fractions contained DLIS and inhibited Na⁺/K⁺-ATPase activity. Inhibition varied with the assay system used but none of the fractions inhibited ⁸⁶Rb uptake by erythocytes. One fraction (which eluted at 29 minutes) contained progesterone; 72% of the inhibitory activity present in this fraction was attributable to this steroid. Another inhibitory fraction co-eluted with dehydroepiandrosterone-sulfate (DHEAS-S). The only fractions found to inhibit both the red cell membrane and canine kidney Na⁺ /K⁺-ATPase enzymes eluted at 7 and 29 minutes. In summary, a number of digoxin-like immunoreactive substances were isolated from cord blood by HPLC fractionation and found to inhibit Na⁺/K⁺-ATPase activity. Inhibition varied with the assay system used. There was no apparent correlation between inhibition and digoxin immunoreactivity. Very large quantities (500 mL) of cord blood were extracted to demonstrate these properties. It remains to be determined whether or not DLIS isolated during the perinatal period is of physiological significance. / Medicine, Faculty of / Pathology and Laboratory Medicine, Department of / Graduate

This study explored infants' immediate and residual responses to holding and rocking, and how these responses relate to previously proposed mechanisms to explain long term benefits found when infants are repeatedly exposed to tactile and vestibular stimulation. This form of stimulation has been proposed to increase infants' ability to control and organize l) their behavioral state, 2) their arousal and autonomic functioning, or 3) that there is no clear relationship between immediate responses and long term benefits. Behavioral state and heart rate were collected on 40 infants who were randomly assigned to either a control group where infants were briefly repositioned twice but otherwise lay undisturbed for 90 minutes or an experimental group where infants were held and rocked for 30 minutes during the middle of a 90-minute observation. Results of analyses showed that, when infants were held and rocked they 1) displayed a lower Heart Rate Mean and Standard Deviation, 2) displayed a lower Mean Heart Rate During Active Sleep, 3) spent less time in a FussCry State, 3) were less likely to cry continuously, and 4) displayed nominally Smoother State Transitions and greater Stability Within States. Following the cessation of the rocking stimulus infants in the Experimental Group l) displayed a lower Mean Heart Rate, 2) displayed a lower Mean Heart Rate while in a Quiet Alert State, 3) were more likely to spend some time in a Quiet Sleep State, and 4) were less likely to cry continuously. In addition, all infants displayed Smoother State Transitions and greater Stability Within States during the first 30 minutes than during the final 30 minutes of the observation. Finally, across the 90-minute observation, the infants who were not rocked spent progressively more time in a Quiet Alert State, whereas infants who were rocked spent less time in a Quiet Alert State. The results were the most consistent with the hypothesis that the mechanism leading to both the immediate and residual effects of the stimulation was an increase in control and organization of infants’ arousal and autonomic functioning. / Ph. D.

Indiana University-Purdue University Indianapolis (IUPUI) / Objectives: To determine (1) whether prescribed and delivered energy and protein intakes during the first two weeks of life met Ziegler’s estimated requirements for Very Low Birth Weight (VLBW) infants, (2) if actual energy during the first week of life correlated with time to regain birth weight and reach full enteral nutrition (EN) defined as 100 kcal/kg/day, (3) if growth velocity from time to reach full EN to 36 weeks’ postmenstrual age (PMA) met Ziegler’s estimated fetal growth velocity (16 g/kg/day), and (4) growth outcomes at 36 weeks’ PMA. Study design: Observational study of feeding, early nutrition and early growth of 40 VLBW infants ≤ 30 weeks GA at birth in three newborn intensive care units NICUs. Results: During the first week of life, the percentages of prescribed and delivered energy (69% [65 kcal/kg/day]) and protein (89% [3.1 g/kg/day]) were significantly less than theoretical estimated requirements. Delivered intakes were 15% less than prescribed because of numerous interruptions in delivery and medical complications. During the second week, the delivered intakes of energy (90% [86 kcal/kg/day]) and protein (102% [3.5 g/kg/day]) improved although the differences between prescribed and delivered were consistently 15%. Energy but not protein intake during the first week was significantly related to time to reach full EN. Neither energy nor protein intake significantly correlated with days to return to birth weight. The average growth velocity from the age that full EN was attained to 36 weeks’ PMA (15 g/kg/day) was significantly less than the theoretical estimated fetal growth velocity (16 g/kg/day) (p<0.03). A difference of 1 g/kg/day represents a total deficit of 42 - 54 grams over the course of a month. At 36 weeks’ PMA, 53% of the VLBW infants had extrauterine growth restriction, or EUGR (<10th percentile) on the Fenton growth grid and 34% had EUGR on the Lubchenco growth grid. Conclusions: The delivered nutrient intakes were consistently less than 15% of the prescribed intakes. Growth velocity between the age when full EN was achieved and 36 weeks’ PMA was 6.7% lower than Ziegler’s estimate. One-third to one-half of the infants have EUGR at 36 weeks’ PMA.