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Effects of chronic in utero hypoxemia on rat neonatal pulmonary arterial structure
Effects of chronic in utero hypoxemia on rat neonatal pulmonary arterial structure Idiopathic persistent pulmonary hypertension of the newborn infant (PPHN) is characterized by intrauterine structural remodeling of the pulmonary arterial bed, consisting of precocious d e v e l o p m e n t of muscle in intraacinar arte1"ies, proliferation of adventitial connective tissue, and sometimes medial hypertrophy of preacinar arteries. To evaluate whether gestational hypoxemia causes these changes, we studied p u l m o n a r y arterial Structure in two groups of newborn rats: one control, the other exposed to h y p o x e m i a produced by maternal hypoxia during the second half of gestation. Morphometric analysis of the p~Jlmonary arterial bed was performed after barium injection into the pulmonary arteries and formol saline expansion of the air spaces. Birth weight was similar in each group. Hematocrit was e l e v a t e d in the hypoxemia group (51% • 1.0% vs 46% • 0.8%, P <0.005): The structure of preacinar and intraacinar arteries was similar and normal in both groupS. Chronic fetal hypoxemia in the rat does not produce the pulmonary arterial structural changes identified in fatal cases of PPHN in human infants. (J PEDIATR1986;108(I):756-759) Robert L. G e g g e l , M.D., Mark J. Aronovitz, B.S., a n d Lynne M. Reid, M.D. From the Department Of Pathology, Children's Hospital Center and Harvard Medical School, Boston
Persistent pulmonary hypertension of the newborn infant is a syndrome characterized by central cyanosis produced by right-to-left shunting of blood across fetal channels (foramen ovale, ductus arteriosus) in the absence of alveolar or congenital heart disease. Although this syndrome can be associated with pulmonary hypoplasia (as in congenital diaphragmatic hernia) or perinatal failu1'e to increase comPliance of resistance arteries in a normally structured arterial bed, many fatal cases represent a third and unexplained variety? .3 In these cases elevated pulmonary vascular resistance is caused by precocious muscularization in utero of normally nonmuscular intraacinar arteries, sometimes with medial hypertrophy of preacinar arteries. In addition, there is usually a prominent adventitial sheath,
Supported bY Grant HL32363 for a Specialized Center of Research in Pulmonary Vascular Disease from the National Heart, Lung, and Blood Institute. Submitted for publication Sept. 16, 1985; accepted Nov. 26, 1985. Reprint requests: Robert L. Geggel, M.D., Children's Hospital Center, 300 Longwood Ave., Box 131, Boston, MA 02115.
756
the significance of which is unclear but which could restrict vessel dilation.2,3 The cause of this third variety of P P H N has not been determined. Because the postnatal pulmonary arterial bed constricts with alveolar hypoxia4 or pulmonary arterial hypoxemia 5 and undergoes structural remodeling with alveolar hypoxia6-8, attention has been focused on the role of fetal hypoxemia. ~ ~0 In the fetal lamb, the pulmonary Fio2 PPHN
Fi'action of inspired oxygen Persistent pulmonary hypertension of the newborn infant
arterial bed is reactive, dilating when mixed venoUs oxygen saturation i s increased ",~2 and constricting when it is decreased~0, ~3,14 TWO previous studies have evaluated the effect of intrauterine hypoxemia on the structure Of the pulmonary arterial bed. Goldberg et al. ~sreported an increased ratio of medial thickness to external diameter in muscular arteries (50 to 150 gm external diameter) from newborn rats that had been hypoxemic during gestation. Because they used
Volume 108 Number 5, Part 1
tissue in which the vessels had not been distended, they Could not distinguish between vasoconstriction and medial hypertrophy. 13Recently Murphy et al., 16applying morphometric techniques to the distended circulation, found that chronic fetal hypoxemia in the guinea pig did not cause neonatal pulmonary artery hypertension or increased muscularity of the pulmonary arterial bed, but did cause growth retardation. To evaluate whether species difference or the experimental technique accounts for the conflicting results, we applied morphometric methods to analyze the effect of gestational hypoxemia in the newborn rat.
METHODS Four pregnant Sprague-Dawley rats (Charles River Laboratories, Portage, Mich.) were obtained at an estimated 10 days of gestation. Two pregnant rats were maintained in room air, the other two from the twelfth day of gestation in a hypobaric oxygen chamber. The protocol was similar to that of Goldberg et al. ~5 except that hypobaric rather than normobaric hypoxia was used. On day 12 of gestation Fio2 was 0.19, and on each of the succeeding 4 days decreased an additional 0.01, so that on day 16 of gestation, Fio2 was 0.15. On day 17 of gestation, Fio2 was decreased to 0.13, and remained at this level until delivery. Each mother delivered her litter at 22 to 23 days estimated gestation. Eight newborn rats from each group were studied. Each group contained six males and two females, the sex determined by gross examination of gonadal structure. Newborn rats were weighed and killed within 4 hours of birth by intraperitoneal injection of sodium pentobarbital (30 mg/100 gm body weight). Within 1 minute of this injection, a deep incision was made in the left axilla, the area blotted dry, and freely flowing blood collected in capillary tubes for hematocrit determination. Through a midline sternotomy, the lungs, heart and trachea were identified. Silicone tubing (oD 0.0625 cm, ID 0.03 cm; Dow-Corning Corp, Midland, Mich.) was positioned by means of right ventriculotomy from the right ventricular outflow tract to the main pulmonary artery and secured with a suture. The pulmonary arterial bed was injected through the catheter with a barium-gelatin suspension, initially at 60 ~ C, by a hand-controlled pump that delivered increments of 0.02 ml. A total volume of 0.06 to 0.08 ml was infused into each lung. A separate suture had previously been tied around the descending thoracic aorta to prevent filling of this structure through the patent ductus arteriosus. The trachea was then cannulated in situ with PE-10 (oD 0.0580 cm, ID 0.0275 cm; Clay Adams, Parsippany, N.J.), and the lungs were expanded and fixed by instillation of formol saline solution at a pressure
Fetal hypoxemia re neonatal hypertension
757
of 25 cm H20. The heart and lungs were removed in their entirety, and fixation continued for 7 days at a pressure of 25 cm H20. A radiograph of the lungs was taken using 35 KeV for 24 seconds at a fixed tube distance of 60 cm. Because of the small lung size, one transverse block specimen was taken that included both lungs; three sections 4 #m thick and 1000/zm apart were cut from this block. Sections were stained for light microscopy by Miller elastin stain followed by van Gieson stain, to identify arterial smooth muscle and elastic laminae. Morphometric analysis of the pulmonary arterial bed was performed using previously described techniques. ~7 Sections were assigned a code number so that analysis was performed without knowledge of study group. For each animal, 75 intraacinar (the first 25 encountered on each of the three slides) and 30 preacinar arteries were examined. For each artery, external diameter, structure (nonmuscular, partially muscular, muscular), accompanying airway level (bronchiolus, terminal bronchiolus, respiratory bronchiole, alveolar duet, or alveolar wall), and medial wall thickness were assessed. Arteries smaller than 19/zm were not assessed. Statistical analysis. For statistical comparison of values for the two groups, the Student t test was used. Values are expressed as mean _+ SEM. RESULTS The newborn weights were similar: 6.19 + 0.17 gm for the control group and 6.20 ___ 0.15 gm for the hypoxemic group. Each mother had a similar weight gain during pregnancy. The hematocrit was higher in the hypoxemie newborn group than in the control newborn group (51% _+ 1.0% vs 46% + 0.8%, P <0.005). In both groups, postmortem arteriograms showed normal branching and similar lumen diameters of the major pulmonary arteries. Lung volumes also were similar.
Microscopic analysis Preacinar arteries. At both the bronchiolar and terminal
bronehiolar level, arteries in the control and hypoxemic groups had similar and normal external diameter, percent medial thickness, and wall structure (Table). For a given external diameter, medial thickness was similar in each group. Intraaeinar arteries. At the alveolar wall, alveolar duct, and respiratory bronehiolar levels, arteries in the control and hypoxemic group had similar external diameter (Table). All intraaeinar arteries had external diameter of <50 ttm. For each group, wall structure was similar at the alveolar wall and alveolar duct levels. At the respiratory bronchiolar level, compared with the control group, the hypoxemic group contained more non-muscular, fewer
758
Geggel, Aronovitz, and Reid
The Journal of Pediatrics May 1986
Table. Structure of Preacinar and intra-acinar arteries Wall structure Accompanying airway landmark
No. of arteries examined per animal
Preaeinar arteries Bronchioli Control Hypoxemia Terminal bronchioli Control Hypoxemia Intra-acinar arteries Alveolar wall Control Hypoxemia Alveolar duct Control Hypoxemia Respiratory bronchiole Control Hypoxemia
External diameter (/~m)
Medial thickness (% Et))
Nonmuscular (mean %)
Partially muscular (mean %)
Muscular (mean %)
19 +_ 0.2 20 +__0.5
112.9 _+ 3.0 106.5 _+ 3.5
5.0 + 0.2 5.4 _+ 0.2
0 0
0 0
100 100
11 + 0.3 10 _+ 0.5
49.5 +_ 0.9 50.9 _+ 1.1
5.1 + 0.2 5.6 -+ 0.2
0 0
1 2
99 98
29 _+ 1 28 + 1
23.1 ___0.3 23.6 _+ 0.2
97.6 + 1.0 97.8 +- 1.7
2.0 + 0.8 2.2 + 1.7
0.4 +- 0.4 0
23 + 2 23 ___2
27.8 + 0.5 28.6 + 0.1
93.1 -+ 2.8 94.6 --- 3.2
6.9 -+ 2.8 5.4 - 3.2
23 + 1 24 +_ 2
35.1 + 0.7 35.8 +__0.3
63.3 +-- 4.3 78.6 + 4.9*
28.8 _+ 2.6 17.6 + 3.1]
0 0 7.9 + 2.8 3.8 - 2.0
*P <0.03. ~P <0.01.
partially muscular, but a similar small proportion of muscular arteries (Table). There was no abnormal precocious muscularization of arteries within the acinus; at each level more than 50% of arteries were nonmuscular. In both groups, the adventitial sheath was thin and normal. Miscellaneous findings. In both groups, pulmonary veins had normal size and structure. N o intravascular t h r o m b i were detected. Pulmonary vascular structure of male and female newborn rats was similar. DISCUSSION Our study demonstrates that prolonged gestational hypoxemia in the rat does not cause the pulmonary arterial structural changes characteristic of idiopathic P P H N in human neonates) ,3 There was no precociouS~extension of muscle within the acinus, medial hypertrophy of normally muscular preacinar arteries, or proliferation of adventitial connective tissue. That hypoxemia was present in the experimental group is indirectly confirmed by the elevation in hematocrit/s In the rat, gestational hypoxemia was not associated with growth retardation, as in the guinea pig.~6 Several previous studies in humans are consistent with our finding that gestational hypoxemia is not important in the pathogenesis of idiopathic P P H N . Polycythemia, produced by hypoxemia in the human fetus, 18 is usually not associated with idiopathic P P H N . 19,2~ The small pulmonary arteries of infants delivered to mothers residing at
high altitude (Leadville, Colorado, 10,150 feet, mean barometric pressure 525 m m Hg) have normal medial thickness at birth, although it is often increased by 3 weeks of age. 6 Infants delivered to mothers with hypoxemia associated with Eisenmenger syndrome can have polycythemia and right-to-left shunting through fetal channels that are corrected by partial exchange transfusion, 2t indicating that elevated pulmonary vascular resistance is caused by the increased hematocrit rather than by abnormal vessel structure. The findings of Goldberg et al. 15 probably represent vasoconstriction 13 rather than change in pulmonary arterial media. In that study of undistended vessels, only arteries larger than 50 /~m ~vere evaluated; these arteries would have been preacinar and thus normally muscular, because in our study all intraacinar arteries were less than 50 #m, although they had been distended before fixation. Vasoconstriction can contribute to idiopathic P P H N , 1,22 but does not represent the principal anatomic abnormality in fatal cases. 2,3 It has been suggested, but not proved, that P P H N is associated with fetal hypertension 23,24 or abnormalities in the prostaglandin 2528 or renin-angiotensin systems. 13 Inasmuch as fetal hypoxemia seems not to be the cause of the precocious muscularization associated with P P H N , further studies should concentrate on other mechanisms of pulmonary vascular control
Volume 108 N u m b e r 5, Part 1
We thank Karen Richter and Beth Little for technical assistance. REFERENCES 1. Geggel RL, Reid LM. The structural basis of PPHN. Clin Perinatol 1984;11:525-549. 2. Haworth SG, Reid L. Persistent fetal circulation: newly recognized structural features. J PEDIATR 1976;88:614-620. 3. Murphy JD, Rabinovitch M, Goldstein JD, Reid LM. The structural basis of persistent pulmonary hypertension of the newborn infant. J PEDIATR 1981;98:962-967. 4. James LS, Rowe RD. The pattern of response of pulmonary and systemic arterial pressures in newborn and older infants to short periods of hypoxia. J PEDJATR 1957;51:5-11. 5. Hughes JD, Rubin LJ. Relation between mixed venous oxygen tension and pulmonary vascular tone during normoxic, hyperoxic and hypoxic ventilation in dogs. Am J Cardiol 1984;54:1118-1123. 6. Naeye RL. Children at high altitude: pulmonary and renal abnormalities. Circ Res 1965;16:33-38. 7. Arias-Stella J, Saldana M. The terminal portion of the pulmonary arterial tree in people native to high altitudes. Circulation 1963;28:915-925. 8. Rabinovitch M, Gamble W, Nada AS, et al. Rat pulmonary circulation after chronic hypoxia; hemodynamic and structural features. Am J Physiol 1979;236:H818-27. 9. Siassi B, Goldberg S J, Emmanouilides GC, et al. Persistent pulmonary vascular obstruction in newborn infants. J PEDIATR 1971;78:610-615. 10. Soifer S J, Kaslow D, Heymann MA. Prolonged intrauterine hypoxia produces pulmonary hypertension in the newborn lamb [abst]. Pediatr Res 1983;17:336A. 11. Campbell AGM, Dawes GS, Fishman AP, Hyman A1. Pulmonary vasoconstriction and changes in heart rate during asphyxia in immature foetal lambs. J Physiol 1967;192:93110. 12. Campbell AGM, Cockburn F, Dawes GS, Milligan JE. Pulmonary vasoconstriction in asphyxia during cross-circulation between twin foetal lambs. J Physiol 1967;192:111121. 13. Drummond WH. Persistent pulmonary hypertension of the neonate (persistent fetal circulation syndrome). Adv Pediatr 1983;30:61-91. 14. Cohn HE, Sacks E, Heymann MA, Rudolph AM. Cardiovascular responses to hypoxemia and acidemia in fetal lambs. Am J Obstet Gynecol 1974;120:817-824.
759
F e t a l h y p o x e m i a re n e o n a t a l h y p e r t e n s i o n -.
15. Goldberg S J, Levy RA, Siassi B, Betten J. The effects o f maternal hypoxia and hyperoxia upon the neonatal pulmonary vasculature. Pediatrics 1971;48:528-533. 16. Murphy JD, Aronovitz M J, Reid LM. Effects of chronic in-utero hypoxia on the newborn guinea pig. Pediatr Res (in press). 17. Davies G, Reid L. Growth of the alveoli and pulmonary arteries in childhood. Thorax 1970;25:669-681. 18. Walker J, Turnbull EPN. Haemoglobin and red cells in the human foetus and their relation to the oxygen content of the blood in the vessels of the umbilical cord. Lancet 1953;2:312318. 19. Drummond WH, Peckham G J, Fox WW. The clinical profile of the newborn with persistent pulmonary hypertension. Clin Pediatr 1977;16:335-341. 20. Levin DL, Heymann MA, Kitterman JA, et al. Persistent pulmonary hypertension of the newborn infant. J PED1ATR 1976;89:626-630. 21. Mukhtar AI, Halliday HL. Eisenmenger syndrome in pregnancy: a possible cause of neonatal polycythemia and persistent fetal circulation. Obstet Gynecol 1982;60:651-652. 22. Gersony WM, Morishima HO, Daniel SS, et al. The hemodynamic effects of intrauterine hypoxia: an experimental model in newborn lambs. J PED~ATR 1976;89:631-635. 23. Levin DL, Hyman AI, Heyrnann MA, Rudolph AM. Fetal hypertension and the development of increased pulmonary vascular smooth muscle: a possible mechanism for persistent pulmonary hypertension of the newborn infant. J PEDIATR 1978;92:265-269. 24. Ruiz U, Piasecki G J, Balogh K, et al. An experimental model for fetal pulmonary hypertension. Am J Surg 1972;123:468471. 25. Levin DL, Mills L J, Weinberg AG. Hemodynamic, pulmonary vascular, and myocardial abnormalities secondary to pharmacologic constriction of the fetal dnctus arteriosus. Circulation 1979;60:360-364. 26. Harker LC, Kirkpatrick SE, Friedman WF, Bloor CM. Effects of indomethacin on fetal rat lungs: a possible cause of persistent fetal circulation (PFC). Pediatr Res 1981 ;15:147151. 27. Levin DL, Fixler DE, Morriss FC, Tyson J. Morphologic analysis of the pulmonary vascular bed in infants exposed in utero to prostaglandin inhibitors. J PEDIATR 1978;92:478483. 28. Perkin RM, Levin DL, Clark R. Serum salicylate levels and right-to-left ductus shunts in newborn infants with persistent pulmonary hypertension. J PEDIATR 1980;96:721-726.
Persistent pulmonary hypertension of the newborn infant is a syndrome characterized by central cyanosis produced by right-to-left shunting of blood across fetal channels (foramen ovale, ductus arteriosus) in the absence of alveolar or congenital heart disease. Although this syndrome can be associated with pulmonary hypoplasia (as in congenital diaphragmatic hernia) or perinatal failu1'e to increase comPliance of resistance arteries in a normally structured arterial bed, many fatal cases represent a third and unexplained variety? .3 In these cases elevated pulmonary vascular resistance is caused by precocious muscularization in utero of normally nonmuscular intraacinar arteries, sometimes with medial hypertrophy of preacinar arteries. In addition, there is usually a prominent adventitial sheath,
Supported bY Grant HL32363 for a Specialized Center of Research in Pulmonary Vascular Disease from the National Heart, Lung, and Blood Institute. Submitted for publication Sept. 16, 1985; accepted Nov. 26, 1985. Reprint requests: Robert L. Geggel, M.D., Children's Hospital Center, 300 Longwood Ave., Box 131, Boston, MA 02115.
756
the significance of which is unclear but which could restrict vessel dilation.2,3 The cause of this third variety of P P H N has not been determined. Because the postnatal pulmonary arterial bed constricts with alveolar hypoxia4 or pulmonary arterial hypoxemia 5 and undergoes structural remodeling with alveolar hypoxia6-8, attention has been focused on the role of fetal hypoxemia. ~ ~0 In the fetal lamb, the pulmonary Fio2 PPHN
Fi'action of inspired oxygen Persistent pulmonary hypertension of the newborn infant
arterial bed is reactive, dilating when mixed venoUs oxygen saturation i s increased ",~2 and constricting when it is decreased~0, ~3,14 TWO previous studies have evaluated the effect of intrauterine hypoxemia on the structure Of the pulmonary arterial bed. Goldberg et al. ~sreported an increased ratio of medial thickness to external diameter in muscular arteries (50 to 150 gm external diameter) from newborn rats that had been hypoxemic during gestation. Because they used
Volume 108 Number 5, Part 1
tissue in which the vessels had not been distended, they Could not distinguish between vasoconstriction and medial hypertrophy. 13Recently Murphy et al., 16applying morphometric techniques to the distended circulation, found that chronic fetal hypoxemia in the guinea pig did not cause neonatal pulmonary artery hypertension or increased muscularity of the pulmonary arterial bed, but did cause growth retardation. To evaluate whether species difference or the experimental technique accounts for the conflicting results, we applied morphometric methods to analyze the effect of gestational hypoxemia in the newborn rat.
METHODS Four pregnant Sprague-Dawley rats (Charles River Laboratories, Portage, Mich.) were obtained at an estimated 10 days of gestation. Two pregnant rats were maintained in room air, the other two from the twelfth day of gestation in a hypobaric oxygen chamber. The protocol was similar to that of Goldberg et al. ~5 except that hypobaric rather than normobaric hypoxia was used. On day 12 of gestation Fio2 was 0.19, and on each of the succeeding 4 days decreased an additional 0.01, so that on day 16 of gestation, Fio2 was 0.15. On day 17 of gestation, Fio2 was decreased to 0.13, and remained at this level until delivery. Each mother delivered her litter at 22 to 23 days estimated gestation. Eight newborn rats from each group were studied. Each group contained six males and two females, the sex determined by gross examination of gonadal structure. Newborn rats were weighed and killed within 4 hours of birth by intraperitoneal injection of sodium pentobarbital (30 mg/100 gm body weight). Within 1 minute of this injection, a deep incision was made in the left axilla, the area blotted dry, and freely flowing blood collected in capillary tubes for hematocrit determination. Through a midline sternotomy, the lungs, heart and trachea were identified. Silicone tubing (oD 0.0625 cm, ID 0.03 cm; Dow-Corning Corp, Midland, Mich.) was positioned by means of right ventriculotomy from the right ventricular outflow tract to the main pulmonary artery and secured with a suture. The pulmonary arterial bed was injected through the catheter with a barium-gelatin suspension, initially at 60 ~ C, by a hand-controlled pump that delivered increments of 0.02 ml. A total volume of 0.06 to 0.08 ml was infused into each lung. A separate suture had previously been tied around the descending thoracic aorta to prevent filling of this structure through the patent ductus arteriosus. The trachea was then cannulated in situ with PE-10 (oD 0.0580 cm, ID 0.0275 cm; Clay Adams, Parsippany, N.J.), and the lungs were expanded and fixed by instillation of formol saline solution at a pressure
Fetal hypoxemia re neonatal hypertension
757
of 25 cm H20. The heart and lungs were removed in their entirety, and fixation continued for 7 days at a pressure of 25 cm H20. A radiograph of the lungs was taken using 35 KeV for 24 seconds at a fixed tube distance of 60 cm. Because of the small lung size, one transverse block specimen was taken that included both lungs; three sections 4 #m thick and 1000/zm apart were cut from this block. Sections were stained for light microscopy by Miller elastin stain followed by van Gieson stain, to identify arterial smooth muscle and elastic laminae. Morphometric analysis of the pulmonary arterial bed was performed using previously described techniques. ~7 Sections were assigned a code number so that analysis was performed without knowledge of study group. For each animal, 75 intraacinar (the first 25 encountered on each of the three slides) and 30 preacinar arteries were examined. For each artery, external diameter, structure (nonmuscular, partially muscular, muscular), accompanying airway level (bronchiolus, terminal bronchiolus, respiratory bronchiole, alveolar duet, or alveolar wall), and medial wall thickness were assessed. Arteries smaller than 19/zm were not assessed. Statistical analysis. For statistical comparison of values for the two groups, the Student t test was used. Values are expressed as mean _+ SEM. RESULTS The newborn weights were similar: 6.19 + 0.17 gm for the control group and 6.20 ___ 0.15 gm for the hypoxemic group. Each mother had a similar weight gain during pregnancy. The hematocrit was higher in the hypoxemie newborn group than in the control newborn group (51% _+ 1.0% vs 46% + 0.8%, P <0.005). In both groups, postmortem arteriograms showed normal branching and similar lumen diameters of the major pulmonary arteries. Lung volumes also were similar.
Microscopic analysis Preacinar arteries. At both the bronchiolar and terminal
bronehiolar level, arteries in the control and hypoxemic groups had similar and normal external diameter, percent medial thickness, and wall structure (Table). For a given external diameter, medial thickness was similar in each group. Intraaeinar arteries. At the alveolar wall, alveolar duct, and respiratory bronehiolar levels, arteries in the control and hypoxemic group had similar external diameter (Table). All intraaeinar arteries had external diameter of <50 ttm. For each group, wall structure was similar at the alveolar wall and alveolar duct levels. At the respiratory bronchiolar level, compared with the control group, the hypoxemic group contained more non-muscular, fewer
758
Geggel, Aronovitz, and Reid
The Journal of Pediatrics May 1986
Table. Structure of Preacinar and intra-acinar arteries Wall structure Accompanying airway landmark
No. of arteries examined per animal
Preaeinar arteries Bronchioli Control Hypoxemia Terminal bronchioli Control Hypoxemia Intra-acinar arteries Alveolar wall Control Hypoxemia Alveolar duct Control Hypoxemia Respiratory bronchiole Control Hypoxemia
External diameter (/~m)
Medial thickness (% Et))
Nonmuscular (mean %)
Partially muscular (mean %)
Muscular (mean %)
19 +_ 0.2 20 +__0.5
112.9 _+ 3.0 106.5 _+ 3.5
5.0 + 0.2 5.4 _+ 0.2
0 0
0 0
100 100
11 + 0.3 10 _+ 0.5
49.5 +_ 0.9 50.9 _+ 1.1
5.1 + 0.2 5.6 -+ 0.2
0 0
1 2
99 98
29 _+ 1 28 + 1
23.1 ___0.3 23.6 _+ 0.2
97.6 + 1.0 97.8 +- 1.7
2.0 + 0.8 2.2 + 1.7
0.4 +- 0.4 0
23 + 2 23 ___2
27.8 + 0.5 28.6 + 0.1
93.1 -+ 2.8 94.6 --- 3.2
6.9 -+ 2.8 5.4 - 3.2
23 + 1 24 +_ 2
35.1 + 0.7 35.8 +__0.3
63.3 +-- 4.3 78.6 + 4.9*
28.8 _+ 2.6 17.6 + 3.1]
0 0 7.9 + 2.8 3.8 - 2.0
*P <0.03. ~P <0.01.
partially muscular, but a similar small proportion of muscular arteries (Table). There was no abnormal precocious muscularization of arteries within the acinus; at each level more than 50% of arteries were nonmuscular. In both groups, the adventitial sheath was thin and normal. Miscellaneous findings. In both groups, pulmonary veins had normal size and structure. N o intravascular t h r o m b i were detected. Pulmonary vascular structure of male and female newborn rats was similar. DISCUSSION Our study demonstrates that prolonged gestational hypoxemia in the rat does not cause the pulmonary arterial structural changes characteristic of idiopathic P P H N in human neonates) ,3 There was no precociouS~extension of muscle within the acinus, medial hypertrophy of normally muscular preacinar arteries, or proliferation of adventitial connective tissue. That hypoxemia was present in the experimental group is indirectly confirmed by the elevation in hematocrit/s In the rat, gestational hypoxemia was not associated with growth retardation, as in the guinea pig.~6 Several previous studies in humans are consistent with our finding that gestational hypoxemia is not important in the pathogenesis of idiopathic P P H N . Polycythemia, produced by hypoxemia in the human fetus, 18 is usually not associated with idiopathic P P H N . 19,2~ The small pulmonary arteries of infants delivered to mothers residing at
high altitude (Leadville, Colorado, 10,150 feet, mean barometric pressure 525 m m Hg) have normal medial thickness at birth, although it is often increased by 3 weeks of age. 6 Infants delivered to mothers with hypoxemia associated with Eisenmenger syndrome can have polycythemia and right-to-left shunting through fetal channels that are corrected by partial exchange transfusion, 2t indicating that elevated pulmonary vascular resistance is caused by the increased hematocrit rather than by abnormal vessel structure. The findings of Goldberg et al. 15 probably represent vasoconstriction 13 rather than change in pulmonary arterial media. In that study of undistended vessels, only arteries larger than 50 /~m ~vere evaluated; these arteries would have been preacinar and thus normally muscular, because in our study all intraacinar arteries were less than 50 #m, although they had been distended before fixation. Vasoconstriction can contribute to idiopathic P P H N , 1,22 but does not represent the principal anatomic abnormality in fatal cases. 2,3 It has been suggested, but not proved, that P P H N is associated with fetal hypertension 23,24 or abnormalities in the prostaglandin 2528 or renin-angiotensin systems. 13 Inasmuch as fetal hypoxemia seems not to be the cause of the precocious muscularization associated with P P H N , further studies should concentrate on other mechanisms of pulmonary vascular control
Volume 108 N u m b e r 5, Part 1
We thank Karen Richter and Beth Little for technical assistance. REFERENCES 1. Geggel RL, Reid LM. The structural basis of PPHN. Clin Perinatol 1984;11:525-549. 2. Haworth SG, Reid L. Persistent fetal circulation: newly recognized structural features. J PEDIATR 1976;88:614-620. 3. Murphy JD, Rabinovitch M, Goldstein JD, Reid LM. The structural basis of persistent pulmonary hypertension of the newborn infant. J PEDIATR 1981;98:962-967. 4. James LS, Rowe RD. The pattern of response of pulmonary and systemic arterial pressures in newborn and older infants to short periods of hypoxia. J PEDJATR 1957;51:5-11. 5. Hughes JD, Rubin LJ. Relation between mixed venous oxygen tension and pulmonary vascular tone during normoxic, hyperoxic and hypoxic ventilation in dogs. Am J Cardiol 1984;54:1118-1123. 6. Naeye RL. Children at high altitude: pulmonary and renal abnormalities. Circ Res 1965;16:33-38. 7. Arias-Stella J, Saldana M. The terminal portion of the pulmonary arterial tree in people native to high altitudes. Circulation 1963;28:915-925. 8. Rabinovitch M, Gamble W, Nada AS, et al. Rat pulmonary circulation after chronic hypoxia; hemodynamic and structural features. Am J Physiol 1979;236:H818-27. 9. Siassi B, Goldberg S J, Emmanouilides GC, et al. Persistent pulmonary vascular obstruction in newborn infants. J PEDIATR 1971;78:610-615. 10. Soifer S J, Kaslow D, Heymann MA. Prolonged intrauterine hypoxia produces pulmonary hypertension in the newborn lamb [abst]. Pediatr Res 1983;17:336A. 11. Campbell AGM, Dawes GS, Fishman AP, Hyman A1. Pulmonary vasoconstriction and changes in heart rate during asphyxia in immature foetal lambs. J Physiol 1967;192:93110. 12. Campbell AGM, Cockburn F, Dawes GS, Milligan JE. Pulmonary vasoconstriction in asphyxia during cross-circulation between twin foetal lambs. J Physiol 1967;192:111121. 13. Drummond WH. Persistent pulmonary hypertension of the neonate (persistent fetal circulation syndrome). Adv Pediatr 1983;30:61-91. 14. Cohn HE, Sacks E, Heymann MA, Rudolph AM. Cardiovascular responses to hypoxemia and acidemia in fetal lambs. Am J Obstet Gynecol 1974;120:817-824.
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F e t a l h y p o x e m i a re n e o n a t a l h y p e r t e n s i o n -.
15. Goldberg S J, Levy RA, Siassi B, Betten J. The effects o f maternal hypoxia and hyperoxia upon the neonatal pulmonary vasculature. Pediatrics 1971;48:528-533. 16. Murphy JD, Aronovitz M J, Reid LM. Effects of chronic in-utero hypoxia on the newborn guinea pig. Pediatr Res (in press). 17. Davies G, Reid L. Growth of the alveoli and pulmonary arteries in childhood. Thorax 1970;25:669-681. 18. Walker J, Turnbull EPN. Haemoglobin and red cells in the human foetus and their relation to the oxygen content of the blood in the vessels of the umbilical cord. Lancet 1953;2:312318. 19. Drummond WH, Peckham G J, Fox WW. The clinical profile of the newborn with persistent pulmonary hypertension. Clin Pediatr 1977;16:335-341. 20. Levin DL, Heymann MA, Kitterman JA, et al. Persistent pulmonary hypertension of the newborn infant. J PED1ATR 1976;89:626-630. 21. Mukhtar AI, Halliday HL. Eisenmenger syndrome in pregnancy: a possible cause of neonatal polycythemia and persistent fetal circulation. Obstet Gynecol 1982;60:651-652. 22. Gersony WM, Morishima HO, Daniel SS, et al. The hemodynamic effects of intrauterine hypoxia: an experimental model in newborn lambs. J PED~ATR 1976;89:631-635. 23. Levin DL, Hyman AI, Heyrnann MA, Rudolph AM. Fetal hypertension and the development of increased pulmonary vascular smooth muscle: a possible mechanism for persistent pulmonary hypertension of the newborn infant. J PEDIATR 1978;92:265-269. 24. Ruiz U, Piasecki G J, Balogh K, et al. An experimental model for fetal pulmonary hypertension. Am J Surg 1972;123:468471. 25. Levin DL, Mills L J, Weinberg AG. Hemodynamic, pulmonary vascular, and myocardial abnormalities secondary to pharmacologic constriction of the fetal dnctus arteriosus. Circulation 1979;60:360-364. 26. Harker LC, Kirkpatrick SE, Friedman WF, Bloor CM. Effects of indomethacin on fetal rat lungs: a possible cause of persistent fetal circulation (PFC). Pediatr Res 1981 ;15:147151. 27. Levin DL, Fixler DE, Morriss FC, Tyson J. Morphologic analysis of the pulmonary vascular bed in infants exposed in utero to prostaglandin inhibitors. J PEDIATR 1978;92:478483. 28. Perkin RM, Levin DL, Clark R. Serum salicylate levels and right-to-left ductus shunts in newborn infants with persistent pulmonary hypertension. J PEDIATR 1980;96:721-726.