Annual meeting—1980 Midwest society for pediatric research

ABSTRACTS

Annual meeting-1980 Midwest Society for Pediatric Research

Comparative bioavailability of intravenous chloramphenicol succinate and oral chloramphenicol palmitate R. E. Kauffman,* M. C. Thirnmoorthi,* J. Buckley,* and A. S. Dajani, Detroit, Mich. The relative bioavailability of intravenous (iv) chloramphenicol succinate (CAP-S) and oral chloramphenicol palmitate (CAPP) was compared in 8 children receiving chloramphenicol for known or suspected meningitis. The area under the serum concentration vs time curve (AUC) was estimated under steadystate conditions while receiving CAP-S and again while receiving CAP-P in each child. Chloramphenicol and CAP-S was determined by HPLC. In 7 of the 8 children, the A U C in patients while receiving oral CAP-P was significantly greater than in those while receiving iv CAP-S. The m e a n % increase in A U C with oral administration was 66% (_+ 21% SEM; P = 0.005) with a range of 27 to 184%. The dose of iv CAP-S did not correlate with the A U C (r = 0.06). However, the dose of oral CAP-P did correlate with A U C (r = 0.72). The bioavailability of oral CAP-P is superior to iv CAP-S. Furthermore, there is a greater correlation between dose and serum concentration when the oral preparation is used. Oral CAP-P appears to provide higher and more predictable serum concentrations than iv CAP-S and offers an attractive alternative in patients who require chloramphenicol and can tolerate oral medication. DISCUSSION Dr. Wilson, Shreveport: Can ~,ou assume that there is absolute bioavailability? Dr. Kauffman: It depends on the individual drug. When studying a drug, you are assuming that 100% of the drug measured is instantaneously entering the body. This is not true in the real situation. A prodrug is almost instantaneously changed to an active molecule. With this prodrug, the change does not occur completely and therefore it is impossible to predict, in a given patient, to what extent this is going to occur. Patients lose 10 to 70% of the dose in the urine. Dr. Hall, Kansas City: Was this a crossover study? Dr. Kauffman: No, each patient was first treated with iv chloramphenicol, and then given the oral form. Dr. Hall: How does that affect the pharmacokinetic data, since each patient received the iv drug first? Dr. Kauffman: We did not have the option to give the oral drug first. By comparing data from the same patient, you can eliminate patient variability. Dr. Keenan, St. Louis: Are there any new thoughts concerning *By invitation. Asterisk indicates the same throughout. 1014

TheJournalofPEDIATRICS VoL 98, No. 6, pp. 1014-1029

the relationship between oral chloramphenicol and the incidence of aplastic anemia. Is it related? Could it be that people taking the oral form received more drug than those who got it intravenous-

1: Dr. Kauffman: There are no data to prove one way or the other. W h e n the first cases of aplastic anemia were reported, larger doses of chloramphenicol were used compared to what is given now. These people probably developed a higher s e r u m concentration because of increased bioavailability with the oral drug. Such a toxic reaction would probably be with a higher dose o f drug.

Cerebral blood flow in neonatal puppies D. Camp,* U. Kotagal,* and L. Kleinman, Cincinnati, Ohio In an attempt to document autoregulation of cerebral blood flow in the newborn animal, cerebral blood flow was measured in 7 neonatal puppies, ranging in age from 2-13 days, by using the radioactive microsphere technique. Period I served as the control period. During Period It the puppies were m a d e hypotensive by withdrawing 15-25 m l / k g of blood. Period III consisted o f volume re-expansion with the previously removed blood. Arterial oxygen tension was maintained above 250 torr with 100% oxygen, and arterial carbon dioxide tension was maintained constant throughout aU periods by administering 5% CO_~ if necessary. During Period It mean blood pressure fell 21.9% _+ 2.7 SEM ( P < 0 . 0 1 ) and cardiac output decreased 25% _+ 6.3 SEM (P < 0.01). Cerebral blood flow during Periods I and II was 28.37 _+ 2.9 and 29.94 _+ 5.9 m l / m i n / 1 0 0 gm, respectively. The fraction o f cardiac output to the brain increased (P < 0.01) during hypovolemia. There were no significant differences in arterial pH, Pr or Po~ during hypovolemia, indicating that a higher fractional output to the brain was not caused by variations in arterial blood gases. During Period III cardiac output and m e a n blood pressure increased (P < 0.01) to previous control levels without changing cerebral blood flow. Thus, in the presence of decreased mean blood pressure and cardiac output, cerebral blood flow was maintained, demonstrating the presence of cerebral autoregulation in the newborn animal. DISCUSSION Dr. Hunt, Chicago: W h a t is the relationship between the maturity of the brain of the puppy and the h u m a n infant? Dr. Camp: I do not know. Dr. Hunt: W h a t were the extremes of Pa~,o2 in the various experimental groups? Dr. Camp: The range of Pac% was from 14 to the 40s. There was no correlation between blood flow and Pa~,o..,. Dr. Fleischmann, Detroit: Did you explore the limit o f autoregulation?

0022-3476/81/0601014+ 16501.60/0 9 198l The C. V. Mosby Co.

Volume 98 Number 6

Dr. Camp: I had planned to withdraw 20 to 25% of the blood volume to induce hypotension. 1 could not withdraw that volume without the development o f acidosis, hypotension, and tachycardia. Dr. Horowitz, Indianapolis: What anesthetic agent did you use? Dr. Camp: Local anesthesia was used. Dr. Horowitz: Does a neonatal puppy have a patent ductus arteriosus? Would that change your conclusion or hemodynamic evaluations? Dr. Camp: 1 cannot comment on that. Dr. Hoszman, Pittsburg: Others have suggested using two reference samples when using microspheres in the animal model. Did you investigate that? Dr. Camp: No, we are looking at that.

Factors affecting glycosylated hemoglobin (GHb) values in children with insulindependent diabetes (IDD) D. Daneman,* D. Wolfson,* D. Beeker, and A. Drash, Pittsburgh Pa. In 477 children with IDD treated by conventional methods, GHb (mierocolumn chromatography at 22 _+ 0.5 ~ C) and random blood glucose (BG) were measured over a 1-year period as indicators of metabolic control (once in 156, twice in 206, three times in 115). The data were analyzed to assess the effects o f patient's age, sex, disease duration, and, in a subgroup of 163, the number of daily insulin injections and insulin dose (U/kg). The mean • SEM %GHb over this period was 11.8 • 0.2%, and BG 237 ___ 9 mg/dl. Only 13 children (2.7%) had a normal %GHb (~< 7.5%). There was a highly significant correlation (P < 0.001) between %GHb and both BG and age, but not with sex or duration > 1 yr. BG did not correlate with any of these factors. In 206 children evaluated twice (mean interval 4.4 mo) there was a significant decrease in %GHb from evaluation 1 to 2 (12 --_ 0.2 to 11.4 ___0.2%; P < 0.001). In 115 with 3 evaluations, there was a similar fall from the first to second, but no further decrease to the third evaluation. Overall in 321 IDD children evaluated more than once, %GHb remained within +0.5% of the initial level in 17%, decreased in 53%, and increased in 30%. No relationship was found between %GHb and either the number o f daily insulin injections or insulin dose. There was no evidence of seasonal variation. These data indicate a closer relationship between metabolic control in children with IDD and age o f the child than with disease duration. This suggests that hormonaI changes and compliance problems during puberty may be very important factors in achieving good metabolic control. In our clinic using conventional methods, the ability to improve control over the short term as measured by changes in %GHb has been quite limited. This study helps to target those ID D children requiring a more aggressive therapeutic approach. DISCUSSION Dr. Pachman, Chicago: Do you think insulin-dependent diabetic children are a homogeneous group? Dr. Daneman: No. Dr. Tsang, Cincinnati: ls the assay itself affected by acute changes in blood sugar? Dr. Daneman: The major component o f the glycosylated

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hemglobin is stable, but there is a small reversible component. Dr. Tsang: With increasing age, children have higher glycosylated hemoglobins. Is this due to maturation or a sign o f gradual deterioration in metabolic control? Dr. Daneman: We feel that these changes are due to hormonal changes with increasing age. With adolescence, there may be decrease in compliance to therapy. Dr. Pachman: Does the diabetic mouse have the ability to glycosylate its hemoglobin? Dr. Daneman: All diabetic animals have this ability. Dr. Goldstein, Columbia: It has taken 3 or 4 years o f multiple measurements of hemoglobin AI~ to develop useful goals and use them as a feedback tool. We found a striking relationship with hemoglobin Ate levels and time, with low hemoglobin AI~ levels during the first several years and a plateau seen at 3 to 4 years. This relationship tends to correlate with loss of endogenous insulin. Do you have an explanation for our differing results? Dr. Daneman: We measured endogenous insulin secretion in patients with diabetes for more than 10 years and found no correlation with the hemoglobin Alo level. It may be related to the populations used in the two studies. Dr. Levitsky, Chicago: Since the technique for measurement o f glycosylated hemoglobin is not one which allows you to store samples and measure them all at one time, could there have been changes in your standards from assay to assay? Dr. Daneman: There is a small interassay variation. Using a water bath at a stable temperature enhances reproducibility. Dr. Levitsky: Would use of a chemical method to measure glycosylated hemoglobin, batch storing, and measurement of samples at one time change your values? Dr. Daneman: We are looking at that. Dr. Chesney, Madison: Does hemoglobin Ale increase with age in the control population? Dr. Daneman: No, not within the childhood population. It does increase in the adult population. Dr. Chesney: In your adolescent group, was there a difference in the correlation with glycosylated hemoglobin between the patient who was in good control versus the patient in poor control? Dr. Daneman: We have not separated this group. The study from Columbia suggests that the more insulin-dependent adolescent tends to have a better glycosylated hemoglobin than the insulin-independent adolescent because of differences in compliance.

Carcinogenic effect of herpes simplex viruses types 1 and 2 (HSV-1 and 2) in the mouse W. B. Wentz,* J. W. Reagan,* A. D. Heggie, Y. S. Fu,* and D. D. Anthony,* Cleveland, Ohio Epidemiologic studies of cervical carcinoma support the hypothesis that prior genital infection by HSV-2 is a cause of this important cancer in women. This hypothesis should interest pediatricians because of the high frequency of HSV infection in sexually active adolescents. The objective of this study was to test this hypothesis by determining if prolonged exposure of the mouse cervix to inactivated HSV-1 or HSV-2 would result in induction of cervical cancer. C~7 mice were exposed to formalininactivated HSV-1 or HSV-2, or to ultraviolet (UV)-inactivated HSV-2 by insertion of virus-saturated cotton pledgets into the

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Abstracts

vagina five times a week for 80 weeks. Control mice were exposed to control fluids prepared in the same way as virus stocks except for the absence of virus. Cervical cancer was detected in 24 to 60% and dysplasia in 18 to 66% of virus-exposed mice. All controls were normal. The frequency of cancer in the mice exposed to UV-inactivated HSV-2 was twice that associated with formalin-inactivated virus. In addition to cervical cancers, endometrial cancers were detected in 7 to 28% of virus-exposed mice, but not in controls. To determine if the oncogenic effect of HSV was specific for the genital tract in mice, formalininactivated HSV-1 or HSV-2 was instilled into the nasopharynx of mice for 12 to 32 weeks. No premalignant or malignant lesions of the naso- or oropharynx or lower respiratory tract were detected. This study shows that, in the mouse, (1) prolonged exposure of the female genital tract to inactivated HSV-1 or HSV-2 induces cervical dysplasia and carcinoma, (2) UVinactivated HSV-2 is more carcinogenic to the cervix than is formalin-inactivated virus, and (3) exposure to inactivated HSV1 or HSV-2 also induces endometrial carcinoma but with lower frequency than cervical carcinoma. DISCUSSION Dr. Dunkle, St. Louis: W h a t was the quantity of virus or titer of the culture material you had activated on the pledget? Does this bear any relationship to the quantity seen in natural infection? Dr. Wentz: The quantity on the pledget was 0.1 ml. The titer of the virus was 10~-10~ T I C D / m l , a m u c h greater exposure than in a natural infection. Dr. Paehman, Chicago: Are there some strains of mice that are more susceptible to infection than others? Dr. Wentz: We have used only two strains of mice, both of which were susceptible. Dr. Tsang, Cincinnati: Has there been any epidemiologic association in h u m a n s between herpes virus infection and carcin o m a of the genital tract? Dr. Wentz: There is an epidemiologic association between HSV-2 and cervical carcinoma. Dr. Green, Chicago: Are you implying that inactivated herpes virus is oncogenic? Is inactivated virus more oncogenic than live virus? Dr. Wentz: In in vitro cell cultures, inactive virus will cause oncogenic changes while the live virus will cause cell destruction. We used live virus in some experiments but 95% of the animals died. Attempts to immunize the animals with live virus against infection of the genital tract have not been successful. Dr. Green: Does the venereaUy transmitted infection seen in the h u m a n correspond to live virus or inactive virus? Dr. Wentz: The venereally transmitted disease is an active infection, but infections with herpes viruses are characterized by a latent infection followed by an active infection. This is the type of situation we are trying to simulate.

Initial l-thyroxine (T~) therapy for congenital hypothyroidism (CH) J. A. Germak,* T. P. Foley, Jr., D. C. Postellon,* and C. H. White,* Pittsburgh, Pa. In 13 infants with CH, serial thyroid function was assessed during initial T~ therapy. The group included 8 infants with

The Journal of Pediatrics June 1981

athyreosis, 3 with dyshormonogenesis (DH), and 1 each with ectopic and hypoplastic glands. Samples were collected at 0.5, 1, 2, 3, 5, 7, and 14 days, and monthly x 6 for total T, (TT,), free T~ (FT~), T3, and TSH. At diagnosis TSH was elevated in all; T~ was low in only 4 of 12, and TI'~ was low in 11 of 12. Eleven infants were treated with 50/~g/day (d) ofT4 (9-14/~g/kg/d in 10 and 18 /~g/kg/d in 1); 2 infants received 25 ~g/d o f T , (4-9/Lg/kg/d). In 3 infants with DH, the mean time in days of therapy to normalization of TT4 was 1.8 (range = 0.5-3) and T S H was 8 (range = 314); T3 was normal in all. In the remaining 10 infants the m e a n time in days of therapy to normalization of thyroid function for those tests that were abnormal at diagnosis was 5 days for TT~ (0.5-14), 11 days for FT4 (7-14), 43 days for TSH (14-150), and 3 days for T:~ (0.5-7). On T, therapy at 9-14 /~g/kg/d, "I'T~ was normal by a m e a n of 3 days. In 4 infants with athyreosis and 1 with hypoplasia on 9-14/~g/kg/d o f T,, the TT, and FT~ were transiently elevated at a m e a n of 40 days o f therapy, yet serum Ta was normal in all but the one infant with symptoms and signs o f hyperthyroidism. Conclusion. (1) Initial therapy for CH with 9-14/~g/kg/d o f T~ results in prompt normalization of TT4, FT4, and T3 with minimal risk for development of hyperthyroidism. (2) Infants with D H m a y have milder hypothyroidism that responds more rapidly to T,. (3) Serum T:~ was infrequently low at diagnosis, promptly normalized on T.,, and remained normal despite transient elevations in I7", and FT~ during therapy. DISCUSSION Dr. Green, Chicago: I disagree with your normal range o f serum T4 levels. In our laboratory, the normal range is up to 25 ~g/dl. Your cutoff is at 16/Lg/dl. Dr. Germak: The normals were for patients between l and 3 months of age. Dr. Green: We find serum T, levels, from 16 to 22 ~g/dl, in normal babies up to 6 months of age. With those levels it is not surprising to have only one infant that is thyrotoxic. Dr. Erenberg, Iowa City: W h y was the serum TSH level elevated for a longer period of time than the serum T, level? Dr. Germak: There m a y be a difference in the feedback mechanism. Dr. Green: ls the prolonged elevated serum TSH level an indication of inadequate therapy? Dr. Germak: No. If the other values are normal, we hesitate to increase the dose of T~. Dr. Green: To us, an excessively high serum TSH level is an indication to raise the T~ dose. Dr. Foley, Pittsburgh: In the rat and h u m a n there is abnormal feedback control for the regulation of TSH. It is important to maintain normal levels of total and free serum thyroxine and not to suppress serum TSH concentrations. Dr. Erenberg: W h y was the serum T~ level normal at birth? Dr. Germak: I feel there was a sufficient a m o u n t of T, to T3 conversion in the newborn. Dr. Winters, Chicago: Is the amplitude of the serum TSH elevation prior to diagnosis correlated to the length of time it takes for the serum TSH level to come back to normal? Dr. Germak: There does not appear to be any correlation. Dr. Rosenfield, Chicago: Previously, the recommendation was to start thyroxine replacement therapy at a low dose. Currently, the initial dose is equivalent to the replacement dose. Have you had any problems or complications with this method of treatment? Dr. Germak: Review of the fiterature reveals that the initial

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dose should be 10 /~g/kg/day. There have been no reports o f thyrotoxicosis in these patients. We had a problem with one hypothyroid patient who received the higher dose o f T,, which resolved following lowering o f the dose. Dr. Chesney, Madison: Was the TSH suppressed in this symptomatic patient? Dr. Germak: Yes. Dr. Dunkle, St. Louis: Were the hypothyroid infants symptomatic prior to treatment? Dr. Germak: Except for two infants, the patients were identified by the neonatal thyroid-screening program. They were started'on treatment within the first 2 months o f life. One infant, referred in by a physician, had mild symptoms of hypothyroidism at 2J& months o f age. The other infant was not identified until 7 months of age and was definitely symptomatic. Dr: Dunkle: How long do you follow the babies? Dr. Germak: We are testing them for development at l and 2 years of age.

"Physiologic hypercalcemia and hypermagnesemia'" associated with decreasing phosphorus intake in exclusively breast-fed infants F. R. Greer,* R. C. Tsang, and J. J. Steichen, Cincinnati, Ohio Simultaneous Ca, Mg, and P levels have not been reported prospectively in breast milk and in sera of nursing mothers and infants. Eighteen lactating mothers and their exclusively breastfed infants were followed for 6 months 0f life. Serum Ca, Mg, P, calcitonin (CT), and parathyroid hormone (PTH) were measured at 3, 6, 12, and 26 weeks. The average gestation was 40 weeks; birth weight, 3,494 gm. Over 6 months, breast milk Ca and Mg did not change, 26 _+ 1 SE vs 25 _+ I and 2.9 _++0.1 vs 3.3 + 0.2 mg/dl, respectively. However, breast milk P was significantly decreased by 6 weeks (t4.7 _+ 0.6 vs 12.7 +_ 0.4 mg/dI, P -< 0.02) falling to 10.7 _+ 0.4 mg/dl by 6 months (P < 0.001). Changes in breast milk minerals did not reflect changes in maternal serum. Maternal serum Ca and Mg increased from 8.6 _+ 0.2 to 9.6 + 0.3 mg/dl (P < 0.005) and 1.95 + 0.06 to 2.15 _+ 0.07 mg/dl (P < 0.05), respectively. Maternal serum P remained unchanged. However, as breast milk P decreased, infant serum P also decreased from 7.9 _+ 0.4 to 5.7 _4=_0.1 mg/dl (P < 0.001) by 6 months. By linear regression, infant P vs breast milk P, r = 0.388, P < 0.01. As infant serum P fell, Ca and Mg increased from 9_2 _+ 0.3 to 10.4 _+ 0.3 mg/dl (P < 0.005) and 1.92 _+ 0.07 to 2.47 _+ 0.08 mg/dl (P < 0.001), respectively. The values for Ca and M]g in infants at 6 months are relatively elevated compared with normal young adults (mean +_ SE Ca = 9.7 ___0.07 mg/dl and Mg = 2.19 _+ 0.02 mg/dl). Despite changes in minerals, infant serum PTH and CT remained unchanged. In conclusion, breast milk Ca, Mg, and P do not reflect changes in maternal cs~rum minerals; however, infant serum P decreases in association with decreases in'breast milk P. We speculate that relative hypercalcemia and hypermagnesemia in breast-fed infants at 6 months of age is related to decreasing P in breast milk. DISCUSSION Dr. Goldstein, Columbia: Have you looked at the same values prospectively in children who were receiving cow milk? Could

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these results be caused by a change in serum proteins? Dr. Green: We have not investigated infants fed cow milk. Ionized calcium in the mother did not change over the 26-week period. We did not measure maternal serum protein levels. Dr. Hall, Kansas City: In order to support Your hypothesis that the serum phosphorus level fell due to a decrease in phosphorus content of breast milk, you would have to know the infants' phosphorus intake. Do you know the phosphorus intake? Dr. Green: No, because we do not know the volume o f milk the infant was receiving. The volume of milk per kg decreases with time and we assume that the phosphorus intake was decreasing, because the phosphorus concentration o f breast milk decreases. Dr. Brown, Pittsburgh: Did you look at vitamin D content in the milk or in the infant's serum? Could your results be a sign of vitamin D deficiency that is resolving? Dr. Green: These babies were part of the study which compared babies on vitamin D supplement tO those not receiving vitamin D supplement. The babies who received vitamin D-had a higher serum vitamin D level. There is no correlation between the serum phosphorus and vitamin D levels. Dr. Lubin, Columbus: If you assume that the volume of milk ingested does not change and these results are not due to other trace metal interactions and represents n o r m a l levels for these children, would you postulate that these infants should be fed something other than breast milk between 4 and 6 months of age? Dr. Green: I do not know. We followed these infants for a year. All were started o n solid food at 6 months of age. The serum phosphorus level did not change from age 6 to 12 months.

Genetic association and linkage in congenital adrenal hyperplasia due to 21-hydroxylase deficiency D. O. Sobel,* J. P. Gutai, B. Smith,* and D. Wagener,* Pittsburgh, Pa. Twenty-nine families of patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency (21-OH def) were studied to evaluate specific HLA antigen association with the disease, and to further investigate the HLA linkage to 21-OH def gene. The association of 21-OH def and the HLA A~ antigen was established (P < 0.01) in our 29 affected families, and in combined reports of 54 families of North American-West European descent. In 23 unaffected siblings o f children with 21-OH def, there was a significant correlation coefficient (P < 0.01) between the biochemical and HLA determined genotype assignments, providing further evidence of 21-OH d e f gene linkage to the HLA complex_ However, in one family, we detected the first case of recombination between the 21-OH deficiency gene and the HLA-B locus. Studies in this family suggest that the 21-OH def gene is located between the HLA-B and glyoxlase loci, and that in the genotype assignment of high-risk families, both the hormonal and HLA methods of detection should be used. DISCUSSION Dr. Pachman, Chicago: If you do DR typing and associate it with your A and B typing, would you expect to further define your group? Dr. Sobel: In this family, we cannot tell the location o f the D locus on the gene. We do not have the data for the D typing.

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Dr. Chesney, Madison: The Eskimos have a high incidence of this deficiency. Have you done any HLA typing in them? Have you looked at other factors that are known to be linked with this deficiency, such as complement components? Dr~ Sobel: In the Eskimo the incidence of that gene frequency is very high (45 per 1,000), but I do not know of any HLA data in that group. We did not look at complement components. Dr. Keenan, St. Louis: What is the biochemical definition tbr defining this group? Dr. Sobel: We are trying to determine ~l" they carry one gene for 21-hydroxylase deficiency.

Growth hormone (GH) stimulates adrenal steroidogenesis in vivo and in vitro in fetal (F) sheep (S) U. P. Devaskar,* S. U. Devaskar,* N. Velayo,* S. Voina,* and M. A. Sperling, Cincinnati, Ohio We have previously demonstrated that hGH directly stimulates F but not maternal adrenal steroidogenesis in rabbit (Pediatr Res 14:310, 1980). We extended this study in S which permits in vivo as well as in vitro assessment. Infusion of 20 /zg of hGH into chronically catheterized F (n = 4) produced a 60-350% increase above basal plasma corticosteroid (CS) concentrations P < 0.01). No such increase above basal was observed in adult S after hGH infusion (n = 3) or in saline-infused F (n = 2). Similarly, hGHstimulated CS production only in F and not maternal isolated adrenocrotical ceils in a dose-response fashion. Maternal and F cells responded to ACTH1 ~, (see Table I). Contamination of hGH by a melanocyte-stimulating hormone (aMSH) cannot account for these findings since we could not stimulate steroidogenesis from c~MSH stimulation in F in vivo or in vitro. We conclude that hGH directly stimulates adrenal steroidogenesis in the F and not in adult sheep. DISCUSSION Dr. Thompson, Iowa City: Was the dose of growth hormone given the same in the fetus as in the adult? Dr. Devaskar: Yes, both in vitro and in vivo. Dr. Thompson: Did you obtain your growth hormone concentrations at 60 or 90 minutes? Dr. Devaskar: Sixty minutes. Dr. Erenberg, Iowa City: W h a t was the cross reaction with cortisone in your radioimmunioassay? Dr. Devasker: There was no cross reaction with cortisone. Dr. Erenberg: Did you measure ovine growth hormone levels? ff you tied off"one of the carotid arteries, you may have affected the fetal pituitary gland and may have decreased endogenous fetal ovine growth hormone production or secretion. Dr. Devaskar: We did not measure ovine growth hormone levels. Dr. Green, Chicago: What you have demonstrated is that the adrenal receptor in the fetal gland is not discriminatory. In human fetal adrenal gland, there is response to hCG and c~MSH. We have early evidence that it may als0 respond to LH. This may indicate that the receptor is nondiscriminatory. Did you activate an adenyl cyclase? Dr. Devaskar: We measured in vitro adenyl cyclase and did not see any change.

The Journal of Pediatrics June 1981

T a b l e I. C S p r o d u c t i o n (X +_ SE) ( n g / 1 0 ~ c e l l s / 2

I Basal Increment above basal ACTH, ~ (25 pg/ml) hGH (1.25 gg/ml) hGH (2.5/~g/ml)

Fetal (n = 5) 1.79 _+ 0.42

hr)

Maternal (n = 3)

1.92 P < 0.02 2,87 +_ 0.63 5.04 3.08 _+ 0.86 0.15 P < 0.02 5.33 _+ 1.1 0.13

_+ 0.59 _+ 1.54 + 0.17 • 0.19

Dr. Wilson, Shreveport: The use of controls in this type of experience is very important. Inactivated growth hormone should have been used for the in vivo and in vitro systems. Dr. Devaskar: We did not use it. Dr. Wilson: Did you look at maternal corticosteroids? Did you have any information on the turnover of these steroids? Dr. Devaskar: We did not do metabolic clearance rates.

Prediction of apnea in siblings of sudden infant death syndrome victims using brainstem auditory evoked potentials J. P. Orlowski,* D. Lonsdale,* and R. H. Nodar, Cleveland, Ohio Epidemiologic studies have suggested that siblings of previous sudden infant death syndrome (SIDS) victims are at a 4 to 7 times increased risk of SIDS. We evaluated 10 neonates who were siblings of autopsy-confirmed SIDS victims with brainstem auditory evoked potential (BAEP) testing. The 10 neonates were all less than 4 weeks of age when evaluated and had not had any clinically detected apnea. Five of the infants had abnormal BAEP responses and each of these infants also had abnormal flexor and extensor neck tone. The remaining 5 neonates had normal BAEP test results and only one o f these infants had abnormal neck tone. After instruction of the parents in infant cardiopulmonary resuscitation and the use of a home apnea monitor, the patients were monitored at home for a period of 6 months. During the home apnea monitoring, the 5 infants with abnormal BAEP responses each developed clinically important apneas of greater than 20 seconds at 6 to 16 weeks of age. The remaining 5 infants with normal BAEP test responses never developed clinically important aPneas during 6 months of home apnea monitoring. Improvement in BAEP test results performed at 1-month intervals in the 5 infants with abnormal initial results, correlated with the subsequent disappearance o f clinically important apneas. These results suggest that BAEP testing may be able to predict the development o f clinically ~important apneas in neonates at increased risk f o r SIDS before the apneas occur, and that improvement in BAEP resuits to normal heralds the resolution o f apnea. DISCUSSION Dr. Cohen, Detroit: Would you define apnea? How do you define the beginning of the apneic spell? Dr. Orlowski: Apnea was defined, by the monitoring done both in and outside of the hospital, as cessation of breathing for

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more than 20 seconds. We do sleep studies in our hospital to define central versus obstructive or combinations o f types of apnea. The beginning of the 20-second time period is defined as the end of any respiratory motion followed by 20 seconds without spontaneous breathing. S o m e people feel 10 seconds of apnea may be abnormal in infants. Twenty seconds may be conservative. Dr. Hunt, Chicago: Other investigators have had difficuity in detecting the same differences as you have in term of controls versus near-misses of S1DS and now siblings of S1DS victims. Are you doing something differently than other investigators? Dr. Orlowski: Auditory evoked potential physiology is a relatively new field. There are differences among interpreters as to what is abnormal. Some say the presence of any peaks at all in infants is normal. We have defined specific criteria which appear to be statistically significant, such as amplitude, wave shape, presence of peaks, inter- and intra-ear peak latencies, and response stability. Dr. Strauss, Iowa City: Is it Correct to assume that these have become normal in time in all the infants that you have studied? Do you feel that this is a developmental delay problem? Dr. Orlowski: That is our hypothesis at this time. We cannot say that they have all become normal. Some of the children are over 1 year of age and continue to have apnea and have abnormal brainstem potentials. The general trend is that the brainstem potentials tend to become normal 1 to 3 months prior to complete cessation Of apnea. We believe this is a maturational phenomenon. The brainstem matures electrophysiologically and then we see the disappearance of apnea. Dr. Strauss: Have these immature patterns been reproduced in premature infants? Dr. Orlowski: We have not looked at this. Dr. Keenan, St: Louis: With any test which has the potential for screening applications, you must be careful as far as the definition of the population that you are testing. I am assuming the incidence of SIDS in 2 per 1,000 in the northern Ohio area and you are studying children who have a 50% abnormality, which is 500 per 1,000. What are your ideas and concerns about preselection bias in this population, since they really were not normal and were selected ~ study for other reasons besides sibship, and about postselection bias, in terms of the definition o f apnea versus controls versus children with the defined immaturity by gestation? Dr. Orlowski: This study is too premature to be used as a screening tool. We are in the process o f trying to organize a statewide study now to get a large number of normal patients since our normal population is relatively small. Preselection is a problem in any nonrandomized, nOncontrolled study. Our study is not randomized or controlled at this stage. There is no statistical significance to any of our findings at this stage. Dr. Rich, Chicago: Are the abnormalities in the brainstem evoked potentials corresponding to a defined lesion in the brainstem? Dr. Orlowski: We are looking at a specific function of the brainstem which includes the auditory pathway. The examina' tion will be abnormal if a child does not have normal hearing. All of Our children are screened by an audiologist for normal hearing. Many premature infants are exposed to ototoxic drugs which may affect this test, since they may not have normal auditory pathways. We have no idea as to how the abnormalities found correlate with lesions in the proposed respiratory centers or other specific areas in the brainstem.

Abstracts

10 19

Iron status of breast-fed and formula-fed infants at age 6 months A. H. Lubin,* R. O. Shrock,* and J. L. Bonnet,* Columbus, Ohio (Introduced by S. B. Kontras, Columbus, Ohio) We compared iron status o f 23 breast-fed and 23 formula-fed infants, whose mothers were enrolled in the second trimester of pregnancy in a study determining effects of maternal nutritional and environmental factors on the infant's subsequent growth and development. Term infants, receiving no supplementary iron, fed at least 95% of caloric requirements for 6 months from either breast milk (Group I) or commercial iron-fortified formula (Group II) were matched for race, sex, and socioeconomic status. Results indicate significantly higher hematocrit levels for Group I, a trend toward higher hemoglobin levels for Group I, and no significant differences in percent saturation between the two groups (Table II). Percent saturation levels in both groups approach minimum acceptable standards at age 6 months. Our data suggest that iron from human milk is absorbed as well as iron from fortified formula a n d ' t h a t breast-fed infants are not generally iron deficient bY 4 months. The controversy regarding the need for prophylactic administration of iron to all breast-fed infants by age 4 months and possibly the need for supplementary iron (iron-fortified cereals or iron-containing drops) for infants on iron-fortified formula, alone, may be resolved by the continued dietary and biochemical observation of iron status in these same infants through their first year of life, now i n progress, DISCUSSION Dr. Tsang, Cincinnati: It is not easy to dissect out all of the variables. Both groups are different by the measures you have described, all of which presumably make some difference. The conclusions to be drawn are diffuse. Why couldn't you design a study that would look at each factor individually? Dr. Lubin: There have been studies which have been done in that way. We did focus on whether the women were breast or bottle feeding. All the children were then subsequently matched for sex, age, race, and, as best as possible, for socioeconomic status. All of the women in this study received medical attention throughout their pregnancy from the same clinic or a group o f private physicians. That bias appeared to be controlled. Dr. Tsang: In multiple regression analysis, it is not possible to sort out all the various factors. When were the solids introduced? Dr. Lubin: Most of the women from both groups were providing 95% of the calories by milkonly for 90% of the children at both 1 and 2 months of age. By 4 months o f a g e , 80% of breast-feeding mothers were providing calories from the breast only. At 4 months of age, breast milk-fed babies received a greater percentage of their calories from breast milk only.The addition of solids accounted for only 15 or 20% of the iron intake for both groups of infants. Dr. Owen, Ann Arbor: How do you know that 17% of the calories were coming from food, other formula, or breast milk? Dr. Lubin: This is calculated by determining the number o f calories from solid foods and then extrapolating back and determining the number of calories needed for the weight gain that they demonstrated. Dr. Keenan, St. Lou{s: This is an implied balance study. You

1020

Abstracts

The Journal of Pediatrics June 1981

Table II

[No. ] Hgb Group I Group II

23 23

12.6 + 12.0 _+

X

Hct

0.96 0.73

37.9 + 35.6 +

X

%Sat ] X

1 . 9 5 16.5 • 1 . 6 3 17.3 •

9.1 6.3

did not show us data that may cause an increased loss of iron, such as an intercurrent infection, especially gastroenteritis. Dr. Lubin: We had a very low incidence of that. In the 46 infants, there were no significant episodes of gastroenteritis. We did not measure iron lost in the stool. That is another variable if this were to be a true balance study. Dr. Bhatia, Iowa City: How did you record the volume and caloric intake of the formula-fed group? Dr. Lubin: Dietary records were taken by 24-hour recall and in-home assessments monthly until 6 months of age and at the health care visits. Dr. Bhatia: Did you assume that the iron content is that stated by the manufacturer? Dr. Lubin: Yes.

Iron absorption from human milk and formula P. J. Garry,* E. M. Hooper,* B. A. Gilbert,* Albuquerque, N. M., and G. M. Owen, Ann Arbor, Mich. An estimation of iron absorption during the first 6 months of life was based on changes in body weight and total body iron (TBI) calculated as the sum of hemoglobin iron (Hgb) and body storage iron (BSI) from plasma ferritin (Pediatr Res 13:143, 1979). In another study, 232 healthy newborn infants (2,750 gm at birth) were enrolled in 4 feeding groups; exclusively breast-fed (B), breast-fed plus supplemental iron (B+), formula (F), or iron-fortified formula ( F + ) . Between birth and age 3 months, increments of TBI were: 49 mg (B), 59 mg (B+), 24 mg (F), and 28 mg (F + ). Between 3 and 6 months, TBI increments were: - 3 mg (B), 26 mg (B +), 7 mg (F), and 23 mg (F +). Percentage of iron intake absorbed was 81% (B), 10% (B+), 97%, (F), and 6% (F + ) during the first 3 months and 10% (B), 40% (B + ), 22% (F), and 3% ( F + ) between 3 and 6 months. Iron absorption is a function of weight gain: hence, greater absorption from birth to 3 months than from 3 to 6 months, irrespective of feeding. Consumption of foods other than human milk was limited and therefore not a factor influencing iron absorption. At ages 6 and 9 months, B + infants had significantly higher mean hemoglobin and ferritin levels than did B infants. Our data confirm that iron in human milk is of high bioavailability, yet exclusive breastfeeding without iron supplementation before age 6 months is not a certain means of preventing iron deficiency in midinfancy. DISCUSSION Dr. Strauss, Iowa City: The University of Iowa pediatric nutrition group advocates supplementation of breast-fed infants from birth. Hemoglobin, iron, and iron saturation levels, relatively insensitive ways of looking at total body iron, will be normal ' for the first 6 months of life regardless of feeding as long as infants are getting ordinary supplementation. Even term healthy babies run out of storage iron at 4 to 6 months of age. If you are interested in preventing the depletion of storage iron, supplemen-

tal iron must be given even to exclusively breast-fed infants. The exclusively breast-fed infant will not receive sufficient amounts of iron from breast milk. Dr. Tsang, Cincinnati: The retention in breast-fed babies at 3 months of age was 81% but at 6 months was only 10%. Retention in infants fed formula were 97% at 3 months of age and dropped to 22% at 6 months. Why was there such a tremendous drop in the percent absorption? Dr. Garry: Part of the explanation is that the rate o f absorption is in part a function o f growth. This decrease in rate of absorption and apparent retention reflects the fact that babies between 3 and 6 months of age are growing a} a relatively slow rate compared to the first 3 months of life.

Excretion of apparently normal acidic c~-mannosidase by mannosidosis fibroblasts Y. Ben-Yoseph, C. L. DeFraneo,* E. E. Pahl, ~' J. Charrow,* and H. L. Nadler, Chicago, Ill.

Cultured skin fibroblasts from a number of patients with mannosidosis have documented deficiency o f intracellular acidic a-mannosidase activity and normal extracellular activity, in contrast to most other lysosomal disorders in which a single enzyme deficiency is evident both intracellularly and extracellularly. In order to rule out the possibility that the normal extracellular activity is due to mannosidase derived from fetal calf serum (FCS), the cells were cultivated in medium containing FCS from which acidic a-mannosidase was adsorbed out by concanavalin A, and then grown in FCS-free medium for 24 hours prior to harvesting. Normal amounts of excreted acidic a-mannosidase were determined in the media whereas the activity in the cell lysates was diminished. In contrast to the heat lability and low affinity of the intracellular enzyme for mannoside substrates, the extracellular enzyme exhibited both normal thermostability and normal kinetics. The possible presence o f an inhibitor or absence of an activator in mannosidosis cell lysates was ruled out by mixing experiments with normal cell lysates. Incubation of the mannosidosis extracellular enzyme with either normal or patient cell lysate resulted in a partial loss of activity, whereas an additive value was observed when normal extracellular enzyme was incubated with either mannosidosis or normal cell lysates. These findings suggest that the mutation in this disease results in altered catalytic properties and thermostability only after complete intracellular "processing." This processing presumably takes place within the lysosomes, and the extracellular enzyme which has escaped this step does not express the defect. DISCUSSION Dr. Cohen, Detroit: How pure is the enzyme that you are working with? Dr. Ben-Yoseph: The system is a crude one, and we are only measuring the acidic a-mannosidase activity. Dr. Chesney, Madison: Why are the serum levels normal if they are excreting normal enzymes? Dr. Ben-Yoseph: This may be explained if we assume that the enzyme in the serum is coming from the liver. Dr. Chesney: Do you think this type of scheme can help explain some of the discrepancies in the enzymatic properties in other disease processes such as cystinosis? Dr. Ben-Yoseph: Yes.

VohLme 98 Number 6

~-A drenergic receptors and adenylate cyclase in developing rat myocardium J. A. Whitsett, J. Hall, and C. Darovee-Beckerman,

Cincinnati, Ohio /3-Adrenergic receptors (BAR) were identified in rat myocardium during development by the binding of [:3H] dihydroalprenolol ([~H]DHA) and [~I]iodohydroxybenzylpindolol([~2"~]HYP). Heart weight, protein, and the number ot~BAR per cell increased with age; however, binding capacity (Bmax) per protein was lower in the adult rat (220 gm body weight), 36.8 +_ 4.1 as compared to the fetal or neonatal rat, 58.3 • 7.2 fmoles/mg protein, m _+ SE, P < 0.01. While protein content, Na +, K § ATPase, 5'-nucleotidase, and adenylate cyclase (AC) increased with ventricular and body weight, the developmental increase in [:~H]DHA binding sites per heart did not keep up with increases in ventricular mass. BAR subtype was/32 in both fetal and adult membranes; the ratio was 75% fll and 25%/32 in fetal samples. Guanine nucleotides (GN) decreased agonist affinity for the inhibition of [I~]HYP binding equally in adult and fetal samples, and enhanced the activation of AC by catecholamines. The lower specific activity (per mg protein) of BAR in adult myocardium may be related to the decreasing proportion of sarcolemmal protein as compared to total cell protein which occurs during hypertrophic cardiac growth, supporting the hypothesis that sarcolemmal area is a major determinant of BAR number. The function of BAR is mediated by GNs early in development, and observed age-dependent differences in the activation of catecholamine-sensitive AC by GTP and Gpp(NH)p may relate to developmental differences in the properties of GN-dependent factors which mediate receptor occupancy and c-AMP formation or "coupling." DISCUSSION Dr. Chesney, Madison: Have you looked at cholera toxin? Dr. Whitsett: Yes. We will report it later. Dr. Erenberg, Iowa City: What effect does the concentration of the hormone coming to the receptor have on this system? Is there a difference between the fetal level and the maternal level with your schema? Dr. Whitsett: The overall receptor affinity is modulated equally in the fetus and adult by GTP. The same dose response curve would be expected in both mother and fetus. Dr. Chesney: Is there is a discrepancy between the fetal response to exogenous GTP and to the inhibitor? Dr. Whitsett: Yes.

In vitro effect of prednisolone on antibodydependent celLmediated cytotoxicity (ADCC) in pediatric autoimmune disease M. L. Miller* and L. M. Paehman, Chicago, Ill. Steroids, accepted therapy for several pediatric autoimmune diseases, inhibit a variety of immune functions. One function, ADCC, mediated by Fc receptor bearing cells, may play a role in autoimmune diseases by eliminating autoantibody-coated cells. ADCC and its alteration by steroids have not been studied in children with lupus (SLE) or dermatomyositis (DMS). The

Abstracts

10 2 1

purpose of this study is to determine the effect of prednisolone on ADCC in normal adults and children with SLE and DMS, and to describe a child in whom ADCC correlated with disease activity. Chang cells labeled with chromium 51 were used as targets. Prednisolone added in vitro to leukocytes from 10 healthy adults reduced ADCC from 67.3 • 13.3 by 17.8% (100/~g/ml added) and 16.6% (50/~g/ml added) at 64:1 effector:target ratio (E:T) (P < 0.05). The decrease at 32:1 E:T and effect of 10 /Lg/ml prednisolone were not significant. Neither 100 nor 50 /~g/ml prednisolone significantly decreased ADCC in 3 children with SLE (76.4 • 23.1 and 73.3 • 23.6 at 64:1 and 32:1 E:T) or 4 children with DMS (70.3 _+ 14.3 and 61.5 _+ 16.1 at 64:1 and 32:1 E:T). A 1089 black girl with active SLE had ADCC values of 33.0 and 27.5 at 64:1 and 32:1 E:T. One week after intravenous administration of prednisolone, clinical and serologic findings improved; ADCC increased to 83.6 and 50.7, respectively. It is concluded that in vitro prednisolone inhibits ADCC from normal adults but not 3 children with SLE or 4 with DMS. ADCC correlated in one SLE patient w~th disease activity. It is speculated that, because of therapy or underlying disease in children with SLE and DMS, lack of ADCC response to in vitro prednisolone may reflect altered cell subpopulations. DISCUSSION Dr. Strauss, Iowa City: Were all the patients receiving steroids? Was there some similarity in their therapy? Dr. Miller: All the patients were receiving steroids for a long period of time. Dr. Strauss: In the mononuclear fraction that you used, I assume these were an admixture of monocytes. Were the number of null cells equal in each plate? Dr. Miller: The effector to target ratio was kept constant in the assay at 64:1. I could not tell if there was a change in the null cell population. Dr. Chesney, Madison: Do you know the percentage of null cells in your patients? Dr. Miller: No. Dr. Chesney: Do you know the suppressor cell status of your patients with systemic lupus? Dr. Miller: No. Dr. Chesney: What is the equivalent in vivo daily dosage of prednisolone which would correspond to the amount used in vitro? Dr. Miller: All except one patient had inactive disease. They were on less than 0.5 mg/kg/day. The dosage use in the in vitro system would be equivalent to the peak concentration of 1 mg/kg. Dr. Whitsett, Cincinnati: Most steroid-dependent functions take 24 hours. Why is your system so rapid? Dr. Miller: That may be true in general but steroid effect on cell lymphocyte populations is more rapid than 24 hours. In vivo, you can see an effect in 4 to 6 hours. Dr. Whitsett: How do you know that your receptor is already saturated with steroids? Dr. Miller: We cannot say for sure. It would be difficult to explain some of the changes if all of the receptor sites were saturated. Dr. Kallen, Cleveland: How do you explain the clinical improvement? Dr. Miller: I am not sure why killer cell activity improved. Perhaps a subcell population that had been suppressed responded to therapy. Alternately, null cells coated with antibody and immune complexes were no longer coated and could resume

1 022

Abstracts

function. What happens in vitro may not always be what happens in vivo.

Renin-aldosterone responses in a boy with defective renal K § excretion S. Sauder, R. P. Kelch,* R. J. Grekin, and R. C. Kelsch, Ann Arbor, Mich. Hyperkalemia and acidosis were found in a 131~ boy during evaluation of delayed growth and development. Only two similar cases have been reported. We have added information about this disorder through studies o1" anterior pituitary function, urinary K § excretion, and renin-aldosterone responses to postural changes and Na + restriction. Other than short stature and delayed development, the patient was asymptomatic (height age 10 years, weight age 9%2 years, skeletal age 10 years, Tanner stage I). Hyperkalemic hyperchloremic metabolic acidosis with partial respiratory compensation was documented (Na 139, K 7.1, CI 116, CO~ 19 mEq/L; pH 7.31, Pr 32 mm Hg). Gtucocorticoid and thyroxine secretion were normal. During sleep, he had an early-pubertal pattern o f pulsatile LH secretion and a normal increase in serum growth hormone (max hGH 12.1 ng/ml). Distal tubular H + excretion and inulin and PAH clearance were normal. Urinary K + excretion was low for the degree of hyperkalemia. Infusion of Na,,SO~, a nonabsorbable anion, increased urinary K 9 excretion and decreased serum K § to 5.5 mEq/L. Plasma renin activity (PRA) was below the normal range for each posture and diet (Na + 120 m E q / d a y - s u p i n e 0, 4-hr-upright 0.06; N a ' 10 m E q / d a y - s u p i n e 0.6, 2-hr-upright 2.27 ng/ml/hr). Although serum aldosterone was within the range found in normokalemic individuals, it was low for the degree of hyperkalemia in this patient (Na + 120 mEq/day-supine 18, 4-hr-upright 39; Na + 10 m E q / d a y - s u p i n e 30, 2hr-upright 56 ng/dl). During Na + restriction, serum K + rose to 8 m E q / L and the patient became tachycardic and hypotensive. Our studies support the previous hypothesis that renal K 9 excretion is defective. The alterations in renin-aldo responses may be attributed to known effects of hyperkalemia. Although it suppresses PRA, hyperkalemia directly increases aldo and thus partially compensates for decreased renin stimulation. We speculate that the patient's symptoms during Na + restriction were due to tow PRA and hence reduced vasoconstriction and Na + retention from angiotensin II. DISCUSSION Dr. Chesney, Madison: This is the Spitzer/Weinstein syndrome. How does this differ from type IV RTA? Dr. Sauder: The major difference was that this child has a better aldosterone response. Dr. Chesney: Have you seen any change in the growth pattern after putting the patient on sodium supplementation and chlorothiazides? Dr. Sauder: Since correcting his hyperkalemia 2 months ago, he has grown a centimeter. We will need more time to see the change in growth pattern. Dr. Fleischmann, Detroit: What happened to the urinary sodium when you have restricted sodium? Dr. Sauder: The urinary sodium fell to about 30 mEq/24 hours on a 10 mEq diet on the third day. He had not really completely shut down his urinary excretion of sodium.

The Journal of Pediatrics June 1981

Dr. Kallen, Cleveland: Do you interpret lack of improvement after Florinef administration to target organ unresponsiveness? Dr. Sauder: I think he has an abnormal system excreting potassium that can be stimulated if he had more aldosterone but that cannot be totally corrected. If he had resisted aldosterone, his serum aldosterone level would be higher.

Absence of isolated testicular relapse (ITR) in childhood acute lymphoblastic leukemia (ALL) with a repetitive reinduction protocol Y. Ravindranath and Raj Warrier, Detroit, Mich. ITR has emerged as a major new problem in children with ALL both on and off"therapy. After treatment, overt ITR as high as 40% has been reported. Testicular biopsy has become a routine at the time of cessation of therapy, and occult ITR o f up to 10% has been noted. Our data are in sharp contrast to the published reports of ITR. We have not encountered ITR as the sole initial relapse in any of 101 boys with ALL treated at our institution over an 1l-year period. Ten patients had mediastival mass at presentation and 41 had WBC > 20,000/mm ~ at diagnosis. All were treated (for 5 years) with identical chemotherapy and the last 62 patients in addition received intermittent intrathecal methotrexate (MTX) and fractional irradiatio n (IMFRA) central nervous system (CNS) prophylaxis. Remission was induced with vincristine (VCR) 2 mg/mL plus 6-mercaptopurine (6-MP) 2.5 mg/kg, and prednisone (Pred) 60 mg/m '~ and maintained with alternating cycles of biweekly oral MTX 30 m g / m ~ for 6 weeks, and VCR + 6-MP + Pred for4 weeks. I M F R A CNS prophylaxis is given every 10 weeks. The median duration of complete remission was 18 months (range 1-20 months) for patients prior to the introduction of CNS prophylaxis, and it has not yet been reached in those receiving CNS prophylaxis (range 1-95 months). Five of 26 patients off therapy had relapse (3 bone marrow, 1 CNS, and 1 simultaneous BM and CNS). Thus in a group of boys with ALL treated with a repetitive reinduction protocol, as described above, ITR as the sole initial recurrence was not seen on or off therapy. This suggests that an effective maintenance program prevents ITR and that the testis may not be a sanctuary site. DISCUSSION Dr. Gilchrist, Rochester: The outlook for boys with ALL is worse than that for girls, most likely not related to testicular involvement but to some other biologic difference in the male population leading to higher incidence of relapse. Age and white count are independent variables that are mentioned as prognostic factors. What was the age o f diagnosis in your patients? Dr. Ravindranath: Our distribution is similar to what you have reported. Dr. Strauss, Iowa City: How did you determine testicular relapse in a late sense? How long were the cases followed after cessation of therapy? Did they all have a testicular biopsy or was it by clinical criteria? Dr. Ravindranath: Patients were followed for 2 to 6 years after cessation of therapy. In this group o f patients, the median was 24 months. We have not done testicular biopsies because we have not found isolated testicular relapse to be a problem. I cannot say if we missed a few microscopic foci.

Volume 98 Number 6

Prognostic factors and management of histiocytosis X P. S. Simmons,* L. E. Wold,* L. R. Elveback,* D. C. Dahlin,* and G. S. Gilchrist, Rochester, Minn. We studied 68 patients with biopsy-proven histiocytosis X diagnosed at the Mayo Clinic between 1962 and 1979. At diagnosis 46 (68%) had disease confined to bone (39 solitary and 7 multiple); 12 had osseous involvement as part of multisystem disease, and 10 had only extraosseous disease. Four had pulmonary involvement, 4 had diabetes insipidus (D1) at diagnosis, and 6 developed D1 during the course of their disease. Of the 51 patients with single organ disease, 12 had surgical treatment alone, 33 radiotherapy with or without surgery, 5 chemotherapy (2 skin, 3 lung), and 1 had radiation plus chemotherapy. Of 17 patients with multisystem disease, 4 received radiation, 5 chemotherapy, and 8 both. Of the 18 patients treated with vinblastine as a single agent, 14 achieved partial or complete remission, 2 had stable disease, and only 2 progressed. At median follow-up of 106 months 55 are alive and disease-free, 4 are alive with DI only, and 4 have stable and 2 active disease. Two patients died of their disease and 1 died of unrelated causes. Forty-three (63%) had no disease exacerbations, 13 (19%) one, and 12 (18%) multiple relapses. Histologic review in 53 cases evaluated mitotic activity, eosinophilic infiltrate, multinucleated giant cells, necrosis, and "syncytial" quality. Age (under 2 years), presence of DI, and multiple organ system involvement correlated positively (P < 0.05) with unfavorable course or outcome but histologic pattern did not correlate with prognosis. We conclude that vinblastine used as a single agent is effective therapy in bistiocytosis X, that certain clinical features are of prognostic significance, hut we failed to confirm the observations of others that course or outcome can be predicted by histologic pattern. DISCUSSION Dr. Sutton, Rochester: How did you define young age? Dr. Simmons: Children less than 2 years o f age. Dr. Cohen, Detroit: Was diabetes insipidus found at the time of diagnosis or anytime during the clinical course? Dr. Simmons: At any time during the clinical course. Dr. Strauss, Iowa City: Was the histology of those with multisystemic disease looked at separately? Dr. Simmons: The pathology was looked at without any clinical history knowledge. The pathologist could not tell the difference between an isolated bony lesion versus the biopsy from a patient with multisystemic disease.

Serum alkaline phosphatase-a sensitive index of zinc undernutrition R. J. Rothbaum* and P. R. Maur* (introduced by W. Balistreri), Cincinnati, Ohio Zinc is an essential component of total parenteral nutrition solutions; however, exact requirements for infants and children are not known. Assessment of total body zinc status is difficult since measurement of serum, red cell, hair, and urinary zinc levels are cumbersome and are not consistent or reliable indices. Zinc losses in diarrheal fluid have been found to be high in adults. We studied 7 infants (age 3 weeks to 3 years) with profuse secretory diarrhea--in 6 cases enteropathogenic E. coli (adhesive

Abstracts

1023

strain 0119) was isolated; the oldest child had protracted diarrhea of unknown etiology. Elemental zinc (100 iLg/kg/day) was provided in central venous alimentation solutions along with 110-120 cal/kg/day, 3-4 gm protein/kg/day, and more than 5% of total calories as linoleic acid (as Intralipid). In spite of this, all infants developed an "acrodermatitis enteropathica-like" rash. Isolated serum zinc values were often n o r m a l - o r were low only in conjunction with hypoalbuminemia. Urinary zinc concentrations were high. An increase in the rate of zinc supplementation (200 ~g/kg/day) was associated with disappearance o f the rash. Serum alkaline phosphatase concentration was inversely related to the severity of the rash, being low during progression of the rash and increasing with healing of the skin lesion. Conelusion. (1) Serum alkaline phosphatase, a zinc metalloenzyme, may serve as a bioassay of zinc status in the ill infant. Low enzyme values indicate that higher doses o f zinc are needed for repletion of zinc losses. (2) The commonly recommended dose (t00/~g/kg/day) o f elemental zinc may not be adequate replacement for zinc losses in infants with severe watery diarrhea and mucosal atrophy. DISCUSSION Dr. Becker, Pittsburgh: These events you showed were very similar to kwashiorkor. In kwashiorkor the administration of zinc alone did not cure the skin lesions. Are you sure that it was the zinc that cured the skin lesions or was it related to other factors such as time? In kwashiorkor you have a low serum alkaline phosphatase level. As they clinically improve, their serum alkaline phosphatase levels improve without the addition of zinc. Dr. Rothbaum: It is very difficult to determine who is and is not zinc deficient. The clinical picture, serum zinc values, and possibly serum alkaline phosphatase levels are fairly dependable factors to help determine the need for zinc. We did skin biopsies on several infants and could not confirm the diagnosis. Our patients received adequate calories, protein intake, and essential fatty acid intake during their hospitalization; therefore, we presumed that the adequate replacement o f zinc was responsible for the improvement. Dr. Gilchrist, Rochester: Was the alkaline phosphatase of bone or hepatic origin? Did you do any isoenzyme studies? Dr. Rothbaum: No. Dr. Lonsdale, Cleveland: It appears that the problem in these babies was loss and nonutilization of zinc. Would you discuss the roles o f the ligand, picolinic acid, pyridoxine, and copper in relationship to the utilization o f zinc? Dr. Rothbaum: Our patients received routine supplemental copper. According to their serum copper levels, it appeared that they had adequate replacement. Picolinic acid is important in the intestinal absorption of zinc. In these infants, it would be important in whatever enterohepatic circulation there is. We have no idea as to their level of picolinic acid. Pyridoxine is part of our routine supplementation. Dr. Hunt, Chicago: I think that you are correct in that the doses of zinc that are being recommended for oral or parenteral supplementation are probably too low. We found that 80 to 100 mg/kg/day is not enough for preterm infants on parenteral nutrition or oral feeding. Zinc deficiency may accompany essential fatty acid deficiency unless you supplement both. Did you give Intralipid? Did you document essential fatty acid deficiency? Dr. Rothbaum: Most of these infants received 40-50% of their calories per day as fat as they received a sufficient amount of linoleic acid. We did not do any serum analyses of fatty acids.

1 024

Abstracts

The Journal of Pediatrics June 1981

Table HI. Results o f 2 daily I N S injections ( m e a n _+ S E M with p e r c e n t o f p a t i e n t s in p a r e n t h e s e s

No reactions HbAlo (%)* Insulin (U/kg)

9.6 _+ 0.55 (85) 0.60 -+ 0.05

[

Mild to occasional

l

8.5 + 0.70 (10) 0.71 • 0.03

Frequent 7.7 • 0.17 (4) 0.83 • 0.04

]

Severe~ 7.2 _+ 0.28 (1) 0.96 _+ 0.14

*Normal HbA,,. = 5.42 • 0.34. tSeizures or altered consciousness.

Radionuclide angiography in evaluation of left-to-right shunt in atrial septal defects R. A. Hurwitz, R. L. Caldwell,* D. A. Girod, and A. Siddiqui,* Indianapolis, Ind. The value of radionuclide angiocardiography (RAC) in estimating pulmonary flow, systemic flow (Qp:Qs) in children with secundum atrial septal defects (ASD) was investigated by comparing results of RAC to (1) data obtained during cardiac catheterization (CC) and (2) estimate of Qp:Qs from measurements of right ventricle by M mode echocardiography. CC was performed on 17 patients to quantitate shunt and evaluate the possibility of associated pulmonary stenosis and/or anomalous pulmonary veins. Estimate of Qp:Qs by first pass RAC had good correlation (r = 0.88) with data obtained at CC. All 12 patients requiring surgery had a Qp:Qs > 1.8 when estimated by RAC; no child had significant underestimation of shunt by RAC. M mode echocardiograms demonstrated only mild enlargement of the fight ventricle in 2/12 children undergoing surgical correction on the basis of CC results. Echo measurement of right ventricular dimension overestimated Qp:Qs in 2 children. This study shows: (I) good correlation between RAC estimate and CC calculation of shunt in children with ASD, and (2) better correlation between RAC and CC than between M mode echocardiography and CC, suggesting that RAC be used for noninvasive assessment of Qp:Qs in children with ASD. DISCUSSION Dr. Feldt, Rochester: In any other indicator system, we have a decreased confidence level when the shunts are at 20 to 30%. What is your confidence level for shunts at about 20 to 30%? How many of your patients had two-dimensional echocardiography? Dr. Hurwitz: Any shunt 3:1 or greater is reported as 3:1. Any shunt considered 1.2:1 or less is reported as insignificant. We did croSs sectional echocardiography on a number of patients trying to measure the size of the defect. There seemed to be a fairly good correlation. We did not do measurements of the right ventricle. Dr. Ostrea, Detroit: Have you tried this technique in infants? How reliable is this technique when there is significant pulmonary disease? Dr. Hurwitz: This technique will only determine a left-to-right shunt. If there is significant pulmonary disease to cause an elevated pulmonary resistance, the shunt will be different when compared to the situation when there is no significant pulmonary disease. If the patient is critically ill and has an elevated pulmonary resistance, the time o f the bolus injection may be lengthened, which will significantly deter the evaluation. Those with significant pulmonary disease can have a problem with this type of determination. We have not looked at very many infants.

The relationship between hypoglycemia and glucose control in children and adolescents with diabetes mellitus S. B. Peth,* D. E. Goldstein, B. Walker,* S. Rawlings,* J. DaCosta,* R. Hess,* and J. England,* Columbia, Mo. Hypoglycemia is considered a serious complication of insulin treatment (INS) o f children and adolescents with diabetes mellitus (DM), limiting the degree to which blood glucose can be normalized. We have determined prospectively over 18 months the incidence of reported hypoglycemic reactions (HR) in 171 patients with DM evaluated at 540 clinic visits. All patients were treated with 2 daily INS injections. Result s are summarized in Table III. Conclusion. Very few patients treated with twice daily INS reported HR. Only 5% of patients reported frequent or severe HR during the 18-month period. The incidence of HR correlated significantly with glycemic control estimated by hemoglobin A~ (HbA~,) (P < 0.0001). Patients with HbAI,. < 8.5% and INS dose > 0.7 U/kg showed increased incidence of HR as compared to patients with either higher HbA~,, level or lower Ins dosages. The level of HbAI~ and insulin dosage predict risk for serious HR in patients with DM. DISCUSSION Dr. Simmons, Rochester: How is the insulin dose chosen for a patient? Were any of the patients on home glucose monitoring? Dr. Peth: Patients' families are taught to regulate insulin dosage at home based on urine tests and hypoglycemic symptoms. At clinic visits, we reassess insulin dosage using hemoglobin AL~. Home glucose monitoring is used by a few patients but we have no data to report. Dr. Winters, Chicago: On one hand, you suggest 75% of patients in the severe group had reactions because they were not adhering to their dietary schedule. You are suggesting that we may use these data to reduce the insulin dose even in the patient who is not having a reaction. I find this contradictory. Dr. Peth: We would like to prevent reactions and still have the patient maintain a realistic goal for blood glucose regulation. These patients do not have normal blood glucose regu'lation. We did not find symptomatic hypoglycemia in the patient whose control is poor. These patients may be different than patients from other centers. Patients with the best compliance and the lowest AI~ levels are at the greatest risk of having hypoglycemic reactions. Dr. Becket, Pittsburgh: Many feel that if you want to get your patient under good control, you should have some degree of symptomatic hypoglycemia. Without it, your patient is not in good control. How many of your patients with your low hemo-

Volume 98 Number 6

globin A,c levels did not have any degree of symptomatic hypoglycemia? Dr. Peth: There are m a n y patients with low A,~ levels that do not have symptomatic hypoglycemia. The A~c level is a good guide to the overall blood glucose regulation, in our experience.

Granulocyte adherence in newborn infants S. Rao,* R. L. Olesinski,* U. Doshi,* and D. Vidyasagar, Chicago, Ilk Adherence to the vascular endothelium is an important cell property of the granulocyte which determines the degree of inflammation and host defense against bacterial invasion. In order to determine if granulocyte adherence (GA) is altered during the newborn period, we studied 52 healthy neonates during their first week of life. Mothers of the infants chosen were not on any antenatal medications. GA was asssayed by modified nylon-fiber columns. The m e a n _+ SEM for 80 healthy adult volunteers was 69.6 _+ 1.2% while the GA for the 52 newborns was 92.0 _+ 0.9%. There was a significant difference between the two (P < 0.01 by Welch t test). There was no significant influence of either race or sex in both adult and neonatal populations studied. A small but significant negative correlation between GA and hematocrit existed for the adults (r = 0.242, P < 0.05), but no such correlation was demonstrable for the neonatal group. There was no significant correlation for the adults between GA and total granulocyte counts, and a small but significant negative correlation was noted for the neonates (r = -0.285, P < 0.05). Conclusion. (1) Granulocyte adherence is not defective in newborns. This cell property is greatly enhanced in this population and thus unlikely to contribute to defective chemotaxis. (2) Increased adherence m a y explain the more frequently noted granulocytopenia seen in neonatal sepsis, in which it contributes to increased granulocyte margination in the presence of endotoxin. DISCUSSION Dr. Ostrea, Detroit: Was the total granulocyte count for the normal newborn low? Dr. Rao: No, the m e a n value was 6,800 cells/ram :~. Dr. Strauss, Iowa City: Would you please define granulocyte? You should be able to determine in the pre- and postfilter samples the percentage of bands and other granulocyte forms. Did you compare the adherence of mature versus immature neutrophils? ls it an intrinsic or acquired neutrophil defect? You may be able to define that by determining the adherence of maternal cells. Dr. Rao: We have included bands with our mature neutrophils in all our counts. We did not study them separately. We did not study the adherence of bands versus mature cells. We studied cells from the newborn, since cord blood studies would be altered significantly because of epinephrine released during the stress of delivery. We do not know how pregnancy affects granulocyte adherence in the mother. In order to establish if it is due to a humoral a n d / o r intrinsic factor, we would have to do other studies. Some of these studies require a large a m o u n t of blood which we cannot get from our neonates. Dr. Hall, Kansas City: Did you obtain the granulocyte counts

Abstracts

10 2 5

on the same samples that you did these studies or were they from separate samples? I a m still impressed with the m e a n granulocyte count being only 6,800 cells/mm ~. Dr. Rao: They were all done on venipuncture samples.

Antigen specific antibody production by human mononuclear leukocyte cultures K. C. Rich, J. Verrier-Jones,* and J. Friedman,* Chicago, Ill., and Dallas, Texas Bacteriophage ~X174 is a well-standardized i m m u n o g e n for quantitating h u m a n antibody production in vivo. W e report on preliminary studies of in vitro ~X174 antibody production with the eventual goal of assessing cell cooperation in response to a specific antigen. Antibody is detected by a bacterial plaque assay which quantitates the neutralization of a standard n u m b e r of phage particles. Phage particles are either added directly to mononuclear leukocyte cultures and neutralizing activity compared to phage incubated in media without cells, or the neutralizing activity of cell-free culture supernatant is determined. Substantial neutralization is observed by cultures from i m m u n e donors within 48 hours, while little or none is observed from n o n i m m u n e donors. Neutralization in culture supernatants is due to antibody since it is abolished by incubation with solid phase anti-immunoglobulin antibody. It is specific for the i m m u n o g e n since culture supernatants from animals immunized with 0XI74 do not neutralize phage T2. Phage neutralization is observed in cultures drawn 1 week after iv immunization while supernatant antibody is detectable at 5 weeks. These studies suggest that in vitro production of ~X174 antibody can be assayed. This m a y prove useful in examining the role of cell cooperation in specific antibody formation in humans. DISCUSSION Dr. Pachman, Chicago: Are there any adverse effects from immunizing with bacteriophage? Dr. Rich: Several hundred patients have been immunized and there have been no adverse side effects.

Creatine phosphokinase and its isoenzymes in normal and stressed neonates T. M. Sutton,* J. F. O'Brien,* F. Kleinberg,* R. F. House,* and R. H. Feldt, Rochester, Minn. Twenty-six newborn infants (13 male, 13 female) with normal prenatal histories, no perinatal stress, and normal vaginal deliveries had determination of creatine phosphokinase (CPK) activity and isoenzyme activity in cord blood and in serum at 24 hours of age. Total CPK activity was high in cord blood (mean = 156.8 U / L ) and Was significantly higher at 24 hours of age (mean = 438.6 U / L , P = 0.0001). There was a significant increase in the skeletal muscle isoenzyme ( C P K - M M ) from a m e a n of 125.6 U / L in cord blood to a m e a n of 405.8 U / L at 24 hours of age (P = 0.0001). There was a significant increase in the cardiac muscle isoenzyme from a mean o f 3.1 U / L at birth to 19.2 U / L at 24 hours although this was not significant (P = 0.0004). There was no significant change in the brain isoenzyme (CPK-

10 26

Abstracts

BB) from a mean of 28.9 U / L in cord .blood to 13.6 U / L at 24 hours o f age (P = 0.095). These results were compared to similar values for a group of 10 neonates with severe cardiac problems. Three o f the ill neonates had significant elevation of total serum CPK and the skeletal muscle isoenzyme (CPK-MM) When compared to the normal newborn infants. There were no significant differences between the normal infants and the ill neonates for t h e cardiac isoenzyme (CPK-MB) and the brain isoenzyme (CPK-BB). These data suggest that caution should be used in the diagnosis of certain neonatal syndromes based on serum creatine phosphokinase levels and isoenzymes alone. More data ~tre needed before definite conclusions regarding the specific importance of these enzyme studies on certain neonatal syndromes can be reached. DISCUSSION Dr_ Raju, Chicago: Were there any differences in the type of delivery in the normal group? in children with meningitis and asphyxia, the CSF portion of CPK is elevated. Do you have any data on these types o f infants? Dr. Sutton: All of the infants were delivered by normal spontaneous vaginal delivery. Five were delivered by elective outlet forceps and had values similar to the other infants. There are no controlled studies measuring CPK-BB after a CNS insult. Dr. Hunt, Chicago: How transient are these rises? Is it related to the hemodynamie situations in your patients? Dr. Sutton: We are not sure how transient the rise in the MB fraction is. As for the cardiac isoenzymes, we do not have long-term follow-up.

Variable effect of increased Ca, P, and vitamin D (D) intake on bone mineralization in preterm infants fed formula with polycose and medium-chain triglyceride (MCT) F. R. Greer,* J. J. Steichen, and R. C. Tsang, Cincinnati, Ohio The ideal postnatal rate of bone mineralization in preterm infants is unknown. The "intrauterine" bone mineral content (IUBMC) was determined by us previously using direct photon absorptiometry. Previously, high Ca, P, and vitamin D supplementation of a standard formula had resulted in a bone mineralization rate comparable to IUBMC. In this study, we studied high Ca (140 mg/dl), P (75 mg/dl), and D (120 [U/dl) in a 24 cal/ounce formula with 50% carbohydrate as polycose and 50% fat as MCT. After establishing full feeds with Similac 20 cai/ounce, the modified formula plus an additional supplement of 400 IU D/day was begun at 2 weeks o f life for an average of 4.75 weeks. BMC of the radius, serum Ca, P, Mg, 250H:vit D, parathyroid hormone (PTH), and calcitonin (CT) were measured at the onset and again after 3-4 weeks. Intake of Ca, P, and D averaged 204 mg/kg/day, 116 mg/kg/day, and 453 IU/kg/day, respectively. Mean caloric intake was 118 kcal/kg/day with a mean weight increase _+ SE of 18.5 _+ 0.7 gmCkg/day. There were no significant changes in serum Ca (8.9 vs 9.2 mg/dl), P (7.1 vs 6.8 mg/d!), Mg (2.17 vs 2.17 mg/dl), 250H-D (36.4 vs 39.9

The Journal of Pediatrics June 1981

ng/ml), PTH (58 vs 69/*l-Eq/ml), and CT (78 vs 45 pg/mi). Postnatal BMC increased and equaled the rate of increase of IUBMC in 8 infants, 48.2 _+ 3.9 mg/cm to 63.5 _+ 5.1 mg/cm, P < 0.01. In 4 infants, however, BMC decreased from 56.3 + 7.0 m g / c m to 41.6 _+ 2.7 mg/cm, P < 0.05, with BMC only approximating the IUBMC for infants of 29 weeks' gestation (40 mg/cm). Serum Ca, P, Mg, 250H-D, PTH, and CT were not different between these two groups of infants nor were intakes of Ca, P, D, and calories. We conclude that this high calcium and phosphorus, 24-calorie formula with polycose and MCT has variable effects on BMC in preterm infants. We speculate that polycose may adversely affect Ca absorption in some preterm infants. DISCUSSION Dr. Erenberg, Iowa City: Did you compare the serum calcitrophic hormone levels in the responders and nonresponders? Dr. Green: There were no differences. Dr. Raju, Chicago: You measured the bone mineral content after 3 to 4 weeks when the infants Were gaining 18.5 gin/day in weight. How much did they increase their length in that time? Since the bone is growing, you may not be doing your repeat bone densitometry at the same site. Dr. Green: We did try to measure the same point on the forearm with each reading, but the baby was growing. We do not look at the rate of increase in length. Dr. Callenbaugh, Kansas City: The data are similar to our data from a 6-year retrospecti,)e review of all low-birth-weight infants less than 1,500 gm. As far as calcium, phosphorus, vitamin D, and caloric intake, there were no differences between the groups that did and did not develop radiographic evidence of rickets. The serum phosphorus level began to drop at 10 to 14 days of age, while feedings were only begun at 10 days o f age. Sixty percent o f the infants who developed rickets were black, while only 30% of the infants who did not develop rickets were black. Do you have information on the maternal nutritional status, maternal and cord blood 25 hydroxy vitamin D levels, or on bone mineral content of the mothers at the time of delivery? Dr. Green: No. Dr. Huntl Chicago: How reproducible are your bone mineral content measurements? Dr. Green: Each bone is scanned four times. There is about a 5% variability. Dr. Whitsett , Cincinnati: Does it pay to have heavy bones in a baby? Dr. Green: I don't know.

Changes in tissue p H (tpH), cardiac output (CO), and blood lactate (BL) in puppies R. Bhat,* B. Braverman,* P. Justice,* D. Vidyasagar, and A. Ivankovich,* Chicago, Ill. We have previously reported the clinical usefulness of tpH in sick neonates: In the present study the effect of sudden hypovolemia on tpH was studied in 5 puppies under anesthesia. Mean blood pressure (MBP), heart rate (HR), right atrial pressure, tpH, and body ternp were monitored continuously. Hypovolemia was induced by bleeding 20 ml/kg o f blood. CO, BL, and blood gases were analyzed every 15 minutes during study period. After 30 minutes of hypovolemia blood was reinfused and all the above

Volume 98 Number 6

Abstracts

1 027

Table IV

Mse Steady state . 15 min of hypovolemia 30 rain of hypovolemia Recovery 15 rain 30 rain

co)

BL

tpH

ApH

(ram Hg)

HR/ml

(1/mt)

(m tool~L)

7.37 _+ 0.03 7.27 _+ 0.05 7.22 _+ 0.06

7.40 • 0.10 7.31 • 0.05 7.27 • 0.05

82 ___ 6.0 68 _+ 4.0 71 • 7.0

206 • 8.0 171 • 8.0 173 _+ 15

0.98 • 0.1 0.60 • 0.04 0.57 _+ 0.06

0.98 • 0.14 3.5 • 1.1 4.52 • 1.2

7.26 • 0.05 7.28 • 0.04

7.31 • 0.03 7.34 + 0.04

79 • 6.0 80 +_ 6.0

192 • 5 196 • 6

0.93 • 0.17 0.84 • 0.10

3.35 • 1.33 2.19 +_ 0.79

values were monitored for the next 30 minutes. Data are given in Table IV. The tpH correlated with the ApH (n = 25, r = 0.88, P = 0.001) Following hypovolemia HR, MBP, HR, CO, ApH, and tpH decreased significantly until reinfusion. MBP, HR, CO, ApH returned to normal within 30 minutes of reinfusion but tpH remained lower than ApH. Correlation o f t p H with BL was better (r = 0.66, P = 0.007) than ApH and BL. tpH changes did not correlate with CO (r = -0.12). Conclusions. (1) tpH changes following bypovolemia appeared earlier and were significantly lower than ApH. (2) In spite of rapid recovery of MBP, CO, and HR, the tpH was slow to recover. (3) tpH changes correlated well with BL. Speculation. Slow recovery of tpH following hypovolemia indicates slow clearance of lactate from tissues. Thus, tpH along with ApH will be a better guide to clinical management. DISCUSSION Dr. Ostrea, Detroit: Your data show that there was a decreased stroke volume. Why was there no increase in the heart rate? Dr. Bhat: The lack of increase in heart rate is probably due to an initiat high heart rate in the puppies. Dr. Fleischmann, Detroit: Was this a muscle surface glass electrc;de? Dr. Bhat: No, this is a subcutaneous tissue electrode with a 1 mm depth under the Skin. Dr. Fleischmann: Do you have any difficulty with the stability of the glass electrode? Dr. Bhat: So far we have h a d n o problems. We have kept them in infants for up to 5 days. Dr. Donn, Ann Arbor: Several investigators utilizing transcutaneous oxygen analyzers have suggested that the relative heating power mode is a good indicator of both local perfusion and central arterial pressure. Have you had the opportunity to do simultaneous studies? Dr. Bhat: No, we have not measured simultaneous Pao~ and pH. Dr. Seeger, Madison: Did you. have any blood gases to evaluate the respiratory compensation of the metabolic acidosis? Dr. Bhat: The paco~ remains steady but the range of the Paco.~ from puppy to puppy was 27 to 47 mm Hg. It remained steady in each puppy. The Pao., demonstrated a slight drop at 30 minutes following the infusion. Dr. Seeger: You would expect some drop in Paco~ to indicate some respiratory compensation for the metabolic acidosis. That finding combined with the paradoxical response in cardiac output suggests that perhaps there was something that happened

during the anesthesia that prevented the animals from normally responding to the stress to which they were subjected. Dr. Bhat: The animals recovered for at least an hour after the placement of the catheters. Dr. Erenberg, Iowa City: Even though the Pao~ remained relatively stable, you removed approximately 30% o f the blood volume and therefore decreased the oxygen-carrying capacity by 30%. Perhaps the changes you are seeing in pH were related to hypoxia at the tissue level. If you express your data as volumes percent instead of Pao._,, you may see a better correlation with pH. Dr. Bhat: We have not looked at those data yet.

Ontogeny and thyroid dependent maturation of fl-adrenergic receptors in rat lung C. Darovec-Beckerman,* J. Pollinger,* K. Adams,* H. Needelman,* and J. A. Whitsett Cincinnati, Ohio Catecholamines mediate surfactant release and smooth muscle tone in the lung, by interaction with fl-adrenergic receptors (BAR) and adenylat e cye!ase (AC). The present study describes the normal ontogeny of BAR in rat lung membrane and the role Of thyroid hormone in the postnatal increase in BAR. BAR were identified with [~H] dihydroalprenolol (['~H] DHA). Bmax increased progressively from 46 • 4.9 fmole/mg protein to 491 _+ 39 m • SE from day 18 of gestation to postnatal day 28, at which time adult Bmax was attained. A great increase in lung BAR occurred between postnatal days 14 and 28, coincident with the known increase in T~ and T~ in the rat; we therefore studied tung BAR in hypothyroidism (Hypo) induced with propyhhiouracil from birth, in normal age-maiched control pups, and in euthyroid pups (Euth) on PTU but treated with thyroxine (T~). Normal and Euth pups grew normally and by day 28, lung weight, DNA, protein, and BAR were identical. Hypo pups grew normally to day 14 but grew poorly thereafter; by day 28 somatic and lung weight, lung DNA, and protein were markedly decreased in Hypo pups. Lung BARs were similar to controls up to day 14, but were decreased in Hypo pups at day 28,290 • 15 n = 13 vs 489 • 31 fmoles/mg protein n = 8 in Euth~ m • SE, P < 0.001.1 n addition, treatment of hypothyroid pups with T~ on days 25-27 doubled lung fl-adrenergic receptors on day 28. The fl._,lfi, adrenergic subtype ratio and protein/DNA ratios were not altered in Hypo at 28 days. We conclude that thyroid hormone or thyroid dependent factors are required for the normal postnatal maturation of the fl-adrenergic system in the rat lung.

10 28

Abstracts

DISCUSSION Dr. Erenberg, Iowa City: Did you measure serum T~ levels in the mother and her pups? Dr. Darovec-Beckerman: In the pups, it was below the detectable level of the assay for the hypothyroid pups. Dr. Erenberg: Do you know how T, works in your system? Dr. Darovec-Beckerman: 1 do not know. T, definitely affects receptor number. Dr. Pachman, Chicago: Is the emergence of these receptors a permanent factor? If you withdraw therapy and make the pup somewhat less than euthyroid, will the receptor status change? Dr. Darovec-Beckerman: W h e n pups that were hypothryroid up to day 25 gestation are given high doses of T~ from day 25 to day 27 gestation, we did bring the receptor levels up to normal. We do not know if withdrawal of therapy will change the n u m b e r of receptors. Dr. Erenberg: Have you looked at the lung histologically to see if there was a difference in the n u m b e r of type 1 or type 11 cells? Dr. Darovec-Beckerman: No.

The influence of glucose infusion on serum bilirubin and bilirubin binding to albumin E. M. Ostrea, Jr., A. Bartos,* C. A. Jesurun,* and C. Fleury,* Detroit, Mich. The jaundiced neonate is at risk to bilirubin (bil) toxicity if the serum albumin (alb) has a diminished capacity to further bind bil as indicated by a high saturation index (SI). A m o n g competitors of bil for alb binding sites, nonesterified fatty acid (NEFA) has been shown to play a prominent role. This study initially reports 12 jaundiced neonates (bil = 13.9 • 0.5 mg/dl) with a high SI (7.8 _+ 3.7%) and no hemolytic disease or drug intake. Glucose infusion was given at a rate of 1 g m / k g / h r for 1-2 hours. After infusion, there was a significant (P < 0.01) fall in S1 (A = --4.1%), serum bil (A = - - 0 . 9 mg/dl) and N E F A (A = - 0 . 4 8 mEq/L). These changes were not due to simple dilution since there was no significant (P > 0.10) fall in serum total proteins (A = - 0 . 2 g/dl). In 6 infants, serum insulin was measured by RIA which showed a rise in insulin level after glucose infusion (A = + 17.8 u U / m l , P < 0.005). Analysis of serum N E F A by GLC showed that the fall in N E F A and SI was associated with a significant (P < 0.005) fall in the % composition of oleic (A = --6.5%) and linoleic (A = --3.2%) acids. Subsequently, SI was routinely measured in infants with serum bil _> 10 mg/dl. Out of 2,392 SI tests, 8% had a high SI ( > 7%). Glucose infusion was given to 81 of these infants ( m e a n bil = 14.5 mg/dl; m e a n SI = 8%). Postinfusion, there was a significant (P < 0.005) drop in serum bil (A = --1.86 mg/dl) and SI (A = -3.8%) in 79/81 of the infants treated. Conclusion. Our study indicates that a high percentage (15%) of infants with serum bil _> 10 m g / d l m a y be at or near risk to bil encephalopathy as indicated by an SI _> 7%. These infants should be identified since treatment with glucose is simple and highly effective in lowering both serum bil and S1. The mechanisms of action of glucose m a y be: (1) stimulates the release of insulin which is a lipogenic agent, a n d / o r (2) possible enhancement of bil clearance by the liver, Such as through increased U D P G A formation, to explain the fall in serum bil despite the fall in SI. The favorable response in most cases to glucose

The Journal of Pediatrics June 1981

treatment indicates that serum NEFA, particularly the unsaturated FA (oleic and finoleic), play a major role clinically in bil binding to alb. DISCUSSION Dr. Dunkel, St. Louis: If you follow the serum bilirubin levels, do they decrease after the glucose infusions? Dr. Ostrea: They continue to remain the same. The saturation index does not result in high levels. Dr. Hoszman, Pittsburgh: Have you infused saline rather than glucose? Dr. Ostrea: No. Dr. Pildes, Chicago: W h a t serum glucose levels did you reach during the 2 hours? Dr. Ostrea: At 30 minutes into the infusion, they ranged from 135 to 175 mg/dl, as measured by Dextrostix. Dr. Pitdes: You are giving twice the a m o u n t of glucose that we normally give. Are you creating hyperglycemia which m a y be more dangerous than reducing your level of serum bilirubin from 13.9 to 13.0 mg/dl? Dr. Ostrea: We are more concerned about the saturation. T h e urine glucose range from + 1 to + 3. Dr: Hunt, Chicago: It is clear that your saturation index is changing but I a m not convinced that the patient is changing. How confident are you that a saturation index above a certain point indicates a child with a greater risk for encephalopathy? Dr. Ostrea: This is based on the clinical study of Odell et al, which demonstrated that a saturation index greater than 8% was associated with a higher risk of encephalopathy.

Increases in blood pressure (BP) and heart rate (HR) in response to parenteral calcium (Ca) therapy (Rx) in the hypocalcemic neonate Donna J. Salsburey* and David R. Brown, Pittsburgh, Pa. Twenty-four hypocalcemic neonates (serum Ca < 7.0 mg/dl) with a m e a n gestational age of 31,3 weeks and a m e a n postnatal age of 26 hours had BP and HR measured serially from 20 minutes before to 300 minutes after receiving their first dose of Ca. Each patient received 18 m g / k g of elemental Ca as a 9% Ca gluconate solution given at a rate of I m l / k g / m i n . Before Ca Rx, systolic BP was 56.0 _+ 1.6 torr (mean _+ SE), m e a n BP was 40.9 • 1.3 torr and diastolic BP was 28.7_+_ 1.4 torr. The maximal increases were 7.5 • 1.0 torr for systolic BP, 3.7 + 0.6 torr for mean BP, and 2.3 _+ 0.6 torr lbr diastolic BP (P < 0.001 for all 3, paired t test). The maximal increase occurred in all 3 values at 20 minutes after Ca Rx and all 3 returned to baseline values by 75 minutes. H R was 143.2 + 2.8 beats/minute before Ca Rx, increased 8.8 • 2.6 beats/minute (P < 0.005, paired t test) at 35 minutes ( m a x i m u m ) and remained elevated through 90 minutes posttreatment. No H R or BP increase occurred in 7 of these patients when they received 2 m l / k g of 0.9% saline at a rate of 1 m l / k g / m i n u t e , suggesting that the increase in H R and BP in response to Ca infusion were not due to volume expansion. Local effects at the site of Ca infusion also did not appear to be responsible for the BP and HR changes, since 9 patients received Ca intravenously and 15 received Ca intra-arterially with no differences between these two groups in the magnitude of the BP

Volume 98 Number 6

or H R responses. The etiology of this response to Ca Rx may be due to a direct effect of Ca on cardiac output or peripheral vascular resistance, or to an indirect effect o f Ca on catecholamine release. Regardless of the cause, these data suggest that Ca Rx in the hypocalcemic neonate may have a salutary effect on cardiovascular performance. DISCUSSION Dr. Erenberg, Iowa City: Did you measure calcium levels preand postinfusion and correlate them with the changes in blood pressure and heart rate? Dr. Salsburey: We measured both total and ionized calcium before and 5, 20, 90, and 300 minutes postinfusion. Compared to the pretreatment level, there was a significant increase in the serum total calcium level at 5 minutes, with a slow decline to the pretreatment level at 300 minutes po~tinfusion. Dr. Erenberg: How did you take your blood pressure reading, since the blood pressure values can vary with sleep state and activity? Dr. Salsburey: We waited until the blood pressure was stable for 5 to 10 seconds before recording the value. Dr. Erenberg: Was there a change in spontaneous activity of the infants following the infusion of calcium? Dr. Salsburey: No. All the hypocalcemic infants were asymptomatic. Dr. Raju, Chicago: The heart rate variability that you showed is in the physiologic range of beat-to-beat variability that is expected in the normal newborn. Dr. Sa~sburey: You are correct. This may explain why the heart rate responses were not consistent with what you would expect with calcium infusions.

Single dose treatment (RX) of urinary tract infections (UTI) in children E. D. Shapiro* and E. R. Wald, Pittsburgh, Pa. Afebrile children with dysuria, frequency, or abdominal pain, no clinical evidence of pyelonephritis, and two clean-catch urine cultures with > 100,000 bacterial colonies/ml were randomly assigned to a Rx group. Rx was with oral amoxicillin (Amx) in either a single dose (SD) of 50 m g / k g or a conventional dose (CD) of 40 m g / k g / d a y at 8-hour intervals for 10 days. Follow-up urine cultures were obtained 2 days, 12 days, t month, and 3 months after initiation of Rx. Cure was defined as relief from symptoms and a negative urine culture 48 hours after the last dose of Amx. Reinfection (Rl) was defined as two positive urine cultures after documentation of cure (Table V). Of 13 children in the SD group (median age 5.3 years), II were cured. One SD failure was not cured by C D treatment either. The other SD failu{e had a UT1 with Amx-resistant bacteria. Two children in the SD group had RI. O f 14 children in the CD group (median

Abstracts

1 029

cu,'ed I

No

Table V. R e s u l t s

Theroy

I No 're te

SD CD

13 14

I

11 13

2 1

RI = Reinfection. age 6.8 years), 13 were cured. There was one CD failure and one RI. Treatment of lower UT1 with a SD of A m x is effective ha selected children. DISCUSSION Dr. Lubin, Columbus: Do you find that there arc other individuals who responded who did have Proteus mirabilis infections? Dr. Shapiro: Only one child had a Proteus mirabilis infection. Dr. Lubin: Do you think other drugs besides amoxicillin might be useful in this single dose therapy proves to be successful? Dr. Shapiro: Yes. Dr. Fleischmann, Detroit: Were all o f these first infections? Dr. Shapiro: No. The majority were first infections, but one-third of the patients had repeat infections. None of the patients with repeat infections had been infected in the previous several weeks or month. Dr. Fleischmann: Did you routinely do radiologic investigation of the girls? Dr. Shapiro: No, we generally do it following the first infection in children less than 3 years of age. Dr. Lubin: There is a recent suggestion that the cutoff of a hundred thousand colonies is m u c h too rigid. Should this same kind o f approach be applied to those infants who have greater than 10,000 colonies/ml? Dr. Shapiro: That study was done primarily in sexually active women. There are no data to answer that question in children.

0 ~ Consumption (V02), COs production (VC02), and insensible water loss (IWL) in premature infants in single (S IV) versus double-wall (D IV) incubators T. F. Yell, S. Voora,* and R. S. Pildes, Chicago, Ill. The complete study has been published under the title of "Oxygen consumption and insensible water loss in premature infants in single- versus double-walled incubators," by T. F. Yeh, S. Voora, L. D. Lilien, J. Matwynshyn, G. Srinivasan, and R. S. Pildes, in the December, 1980, issue of THE JOURNAL OF PEDIATRICS (J PEDIATR 97:967, 1980).