Concentrations of 15-ketodihydro-PGF2α, cortisol, and progesterone in the plasma of healthy and pathologic newborn foals

Available online at www.sciencedirect.com

Theriogenology 72 (2009) 1032–1040 www.theriojournal.com

Concentrations of 15-ketodihydro-PGF2a, cortisol, and progesterone in the plasma of healthy and pathologic newborn foals S. Panzani a,*, M. Villani a,1, A. McGladdery b, M. Magri c, H. Kindahl d, G. Galeati e, P.A. Martino f, M.C. Veronesi a a

Department of Veterinary Clinical Sciences, Faculty of Veterinary Medicine, University of Milan, Via G. Celoria, 10 20133 Milan, Italy b Beaufort Cottage Stables, High Street, Newmarket, Suffolk CB8 8JS, United Kingdom c Clinica Veterinaria Spirano, Strada Provinciale Francesca, 24050 Spirano (BG), Italy d Department of Clinical Sciences, Faculty of Veterinary Medicine and Animal Science, Swedish University of Agricultural Sciences, P.O. Box 7054, SE-750 07 Uppsala, Sweden e Department of Veterinary Morphophysiology and Animal Production (DIMORFIPA), University of Bologna, Via Tolara di Sopra, 50 40064 Ozzano dell’Emilia (BO), Italy f Department of Animal Pathology, Hygiene and Veterinary Public Health, Faculty of Veterinary Medicine, University of Milan, Via G. Celoria, 10 20133 Milan, Italy Received 8 January 2009; received in revised form 8 June 2009; accepted 24 June 2009

Abstract Information regarding the plasma hormone profiles of prostaglandins (PGs), cortisol (C), and progesterone (P4) during pathologic processes in newborn foals is scarce. The aim of this study was to determine the plasma concentrations of these hormones in diseased foals (n = 40) and healthy at-term foals (n = 24) (Equus caballus) during the first 2 weeks of life. Blood samples were collected daily, before any treatment with nonsteroidal drugs in diseased foals, and plasma was analyzed by radioimmunoassay. 15-KetodihydroPGF2a (PGM) was consistently higher in diseased foals than in healthy foals, probably related to roles of PGs in completing organ maturation and/or the presence of oxidative stress or inflammation. Similar trends were observed for C and P4. In diseased newborns, only PGM was significantly higher in nonsurviving foals, although C showed a similar profile. When specific diseases were considered, the levels of PGM and C were lower in premature foals at 12 h of life, whereas the concentration of P4 was higher than in controls. The results of this study demonstrate the differences in plasma hormone levels between healthy and pathologic newborn foals, particularly during the first 2 d of life, probably reflecting the inability of diseased foals to cope with the transition between fetal and neonatal life. # 2009 Elsevier Inc. All rights reserved. Keywords: Cortisol; Newborn foals; Progesterone; Prostaglandins

1. Introduction Several hormones play important roles in the intrauterine and early extrauterine life of newborn * Corresponding author. Tel.: +39 02 50318149; fax: +39 02 50318148. E-mail address: [email protected] (S. Panzani). 1 Present address: Utrecht University, Department of Equine Sciences, Yalelaan 114, 3584 CM Utrecht, The Netherlands. 0093-691X/$ – see front matter # 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.theriogenology.2009.06.015

foals, as well as in human neonates [1]. Prostaglandins (PGs), cortisol (C), and progesterone (P4) are responsible for regulating the final maturational events of organs and systems. The activities of PGs, C, and P4 have been investigated in several species, including newborn foals; however, information regarding their plasma profiles during pathologic processes is scarce. At birth, one of the most important events is the closure of the ductus arteriosus [2]. In utero, PGs (PGE2

S. Panzani et al. / Theriogenology 72 (2009) 1032–1040

in particular), exert a vasodilatory effect to maintain its patency [3]; at birth, PGF2a causes constriction of the ductus arteriosus [4]. Some PGF2a metabolites, like 8-iso-PGF2a, are known as markers of oxidative stress in newborn babies affected by intracellular hypoxia; their levels decrease during chronic hypoxia and increase during acute hypoxia [5]. Prostaglandins also play an important role in renal physiology, as they are involved in hemodynamic, fluid, and electrolyte homeostasis [6,7]. The involvement of PGs in the hemodynamics of several organs has been evaluated in lambs and rats [8] and in preterm human newborns, who show higher urinary levels of PGF2a compared with that of at-term neonates. In full-term infants, a decrease in the PGE2/PGF2a ratio is accompanied by an increase in blood pressure and is associated with an increase in urinary osmolarity [9]. Blood levels of PGF and its metabolite 15-ketodihydroPGF2a (PGM) remain higher for a longer time than do PGE levels in full-term newborns. At 5 to 8 wk of life, the level of PGE is threefold higher in full-term infants than that in adults [10]. The role of PGs as potent mediators of the inflammatory process is well-established [11,12]. Alterations in plasma eicosanoid levels during sepsis and endotoxemia have been documented in adult horses [13], but there are no similar studies in foals. During the first few hours of life, high C levels represent a normal response to the stressors associated with labor, birth, and transition in newborn babies [14]. In fact, it is well known that both the initiation of parturition and the final fetal organ system maturation are strictly related to an increase in fetal C before delivery [15–17]. Preterm foals, who do not present this antenatal increase in fetal C, present signs of prematurity that often lead to death associated with multiorgan failure [18]. The C response in newborn babies is related to the presence of stressors represented by intrinsic and sensory stimuli. Intrinsic stimuli include glucose levels, oxygen saturation, blood pressure, and temperature. Sensory stimuli comprise movement, touch, and pain [14]. A recent study of newborns babies found that in neonates affected by sepsis syndrome or respiratory distress, basal circulating C concentrations were higher than that in normal infants [19]. Some studies have reported higher C levels in fetal lambs affected by hypoxemia compared with that in controls [20,21]. Another study found elevated C concentrations in newborn pigs affected by hypothermia [22]. Postnatal maladaptation, commonly found in newborn foals, is associated with poor stress responses and low C levels, often leading to hypotension [23].

1033

Others steroid hormones, like P4 and its metabolites, play integral roles in each step of horse pregnancy and may influence newborn physiology in the first days of life. Moreover, P4 represents a key precursor for the synthesis of neuroactive steroids and has been shown to have suppressive effects on brain function in newborn lambs [24]. In horses, plasma progestogen concentrations are high at birth and decrease by the end of the first day of life, falling to near zero at 2 d postpartum [25]. In premature and dysmature foals, progestogens remain persistently high, as in foals delivered by caesarean section or in maladjusted foals [25–27]. In maladjusted foals, progestogens decrease to near zero several days after birth during the recovery phase. In foals that did not survive, progestogens remained elevated until death [27]. The aim of this study was to investigate the hormones that might be involved in physiologic and pathologic processes during neonatal life. Plasma concentrations of PGM, C, and P4 in diseased foals and healthy at-term foals during the first 2 wk of life were evaluated as follows: (1) the plasma levels of each hormone were compared between control and pathologic foals; (2) the hormone profiles of pathologic newborns were compared between surviving and nonsurviving foals; and (3) PGM, C, and P4 plasma levels were evaluated relative to the specific diseases affecting the pathologic foals. 2. Materials and methods 2.1. Control group Twenty-four light-horses foals (Equus caballus), 14 females and 10 males, born by spontaneous delivery and housed at a private stud farm, were considered. All were full-term with birth weight ranging between 38 and 57 kg, normal size, coat, and fetlock joint extension. The Apgar index within 10 min of birth, the presence of suck and righting reflexes, the time to stand up (TSU) and to the first suck (TFS), and other physical and behavioral characteristics were used to assess foal maturity and viability; these were within the normal ranges [18,28]. Blood was collected from of each of the 24 foals via the jugular vein into heparinized tubes at 12 h from birth, then twice per day (between 7:00 and 9:00 a.m. and between 6:00 and 8:00 p.m.) from Day 1 to Day 7 and at Days 10 and 14 of age. 2.2. Pathologic group Forty diseased light-horses foals (Equus caballus) affected by various pathologies were enrolled. All were

1034

S. Panzani et al. / Theriogenology 72 (2009) 1032–1040

hospitalized in a neonatal intensive care unit (NICU) during the first 2 wk of life. At admission, after the clinical exam, blood samples were collected from all of the newborns for hematologic and biochemical analyses. Three samples collected 30 min apart were used for blood cultures. From the day of admission until discharge or death, animals were monitored constantly by clinical, laboratory, and instrumental examinations, as indicated. Blood samples were collected from each foal twice per day (between 7:00 and 9:00 a.m. and between 6:00 and 8:00 p.m.). If treatment with nonsteroidal drugs was needed, blood was collected before the administration of the drug. The main diseases affecting the foals were prematurity/dysmaturity, hypoxic-ischemic encephalopathy (HIE), and septicemia. A foal was defined as premature if it was born before 320 d gestation [29]. Dysmaturity describes foals that demonstrated some signs of physical immaturity despite having a ‘‘normal’’ gestational age [30]. Hypoxic-ischemic encephalopathy refers to noninfectious syndromes of newborns less than 3 d of age, characterized by signs of central nervous system disease [31]. Septicemia was indicated by a modified sepsis score > 11 [32] and/or a positive blood culture. In addition to these main diseases considered individually, other less severe pathologies affecting the digestive tract and the respiratory and locomotory systems were considered together in the group ‘‘others.’’ Each foal was included in one of these groups, according to the main affecting disease. 2.3. 15-Ketodihydro-PGF2a assay The profile of PGF2a was monitored by measuring its main initial plasma metabolite, PGM, by radioimmunoassay (RIA) [33]. Duplicate 0.2 mL samples of unextracted plasma were assayed. Before the addition of antibody and radioactive tracer, 0.3 mL 0.25% bovine gamma globulin in buffer was added, and the tubes were heat-treated for 30 min at 45 8C. In the case of a high value (>800 pmol/L), the analysis was repeated with 50 mL unextracted plasma and 0.45 mL 0.25% bovine gamma globulin in buffer. The antibody cross-reacted with 15-keto-PGF2a (16%), 13,14-dihydro-PGF2a (4%), and 15-ketodihydro-PGE2 (1.7%). Other tested prostaglandins crossreacted < 0.1%. The detection limit of the assay using 0.2 mL plasma was 75 pmol/L. The intra-assay and interassay coefficients of variation were 8.5% and 14%, respectively.

2.4. Cortisol and progesterone assays Steroid hormone concentrations were measured by validated RIAs as previously described [34,35]. The sensitivities were 4.3 pg/tube for the C assay and 2.7 pg/tube for the P4 assay. The intra-assay and interassay coefficients of variation were 5.4% and 8.6%, respectively, for C and 6.2% and 9.7% for P4. Crossreactions of other steroids with antiserum raised against P4 were progesterone (100%), 11a-hydroxyprogesterone (90.9%), 20a-hydroxyprogesterone (1.5%), 17ahydroxyprogesterone (1.5%), 5a-pregnan-3-20-dione (2.5%), 20a-hydroxy-4-pregnen-3-one (0.9%) and pregnenolone (<0.01%). Cross-reactions of various steroids with antiserum raised against C were cortisol (100%), cortisone (20.4%), 11a-deoxycortisol (49.8%), and corticosterone (1.13%). The results are expressed as nanograms per milliliter (ng/mL). 2.5. Statistical analysis Diseased and control foals were age-matched to compare the hormone profiles between the groups. The mean values for each hormone at each sampling time in control and diseased foals were compared by the t-test for independent samples. The same test was used to analyze hormone concentrations in surviving and dead foals within the pathologic group. One-way analysis of variance followed by a least significant difference test for multiple comparisons were used to compare hormone profiles in foals among the disease groups (prematurity/dysmaturity, HIE, septicemia, others). Statistical significance was set at P < 0.05. 3. Results 3.1. Clinical results All of the 24 control foals were born at 338  8.9 d gestation. In these foals, the mean Apgar index was 9.4  1, the mean TSU was 61.5  41.7 min, and the mean TFS was 94  51.7 min. No diseases or abnormalities were detected in any of these foals during the 2 wk of observation. At admission, the diseased foals were of different ages. Twelve foals were hospitalized within 12 h of birth, nine at 1 d, ten at 2 d, three at 3 d, one at 4 d, two at 5 d, two between 8 and 10 d, and one between 11 and 14 d of life. Six foals were classified as premature/dysmature, 14

238  135.6*97  28.2** 405  196.2538  379.6 10  6.4 10  6.5** 29  22.9 25  16.8 0.1  0.2 0 1.5  2.7 0.1  0.1 133  71.2* 447  452.9 14  12.8 18  21.8 0 0.9  2.4 P4

C

561  438.3 614  537.9 18  12 29  19.7 1.2  2.8** 7.7  6.8

Note: Differences between groups within each sampling time are represented by *P < 0.05 or **P < 0.01.

11  45.1* 411  333.5 13  11.9 27  26.7 0.1  0.1 1.4  3 217  272.5 376  347.9 13  6.5 18  17.9 0.2  0.4 1.8  4.1 403  436.8* 618  808 14  8.1* 32  30 0.5  0.8 2  4.4

6d 5d 4d 3d 2d

991  786.6 142  1065.7 17  9.3* 44  35.5 2  2.9** 8.5  6.1 PGM

Pathologic foals had higher P4 plasma concentrations compared with that of normal foals at 12 h (8.6 ng/ mL vs. 2 ng/mL) and at 1 and 2 d of life (7.7 ng/mL vs. 1.2 ng/mL and 4.1 ng/mL vs. 0.5 ng/mL, respectively; P < 0.01) (Table 1). No other significant differences in circulating P4 levels were detected (Figs. 5 and 6).

Control foals Pathologic foals Control foals Pathologic foals Control foals Pathologic foals

3.4. Plasma progesterone concentrations

1d

Plasma C concentrations were higher in diseased foals than in normal foals at 12 h (44 ng/mL vs. 17 ng/ mL), 2 and 3 d (32 ng/mL vs. 15 ng/mL and 32 ng/mL vs. 13 ng/mL, respectively; P < 0.05), and at 7 d of life (29 ng/mL vs. 9 ng/mL; P < 0.01) (Table 1). No other significant differences in plasma C levels were detected among the various groups (Figs. 3 and 4).

12 h

3.3. Plasma cortisol concentrations

Table 1 Plasma hormone levels of each hormone in control and pathologic foals at all sampling times.

Disease foals had higher levels of PGM compared with that of normal foals at Days 2 (648 pmol/L vs. 266 pmol/L), 5 (411 pmol/L vs. 111 pmol/L), 6 (447 pmol/L vs. 133 pmol/L), 7 (405 pmol/L vs. 238 pmol/L; P < 0.05), and 10 (538 pmol/L vs. 97 pmol/L; P < 0.01) (Table 1). In addition, in the pathologic group, nonsurviving foals had significantly (P < 0.05) higher PGM levels compared with that of surviving newborns at 2 and 3 d of life (1069 pmol/L vs. 508 pmol/L and 1261 pmol/L vs. 417 pmol/L, respectively) (Fig. 1). When the specific class of disease was considered, foals with HIE showed higher PGM concentrations than did foals with other diagnoses (prematurity/dysmaturity, septicemia, and other diseases) at Days 5 (841 pmol/L vs. 176 pmol/L, 395 pmol/L, and 224 pmol/L, respectively; P < 0.01) and 6 (1043 pmol/L vs. 162 pmol/L, 545 pmol/L, and 132 pmol/L, respectively; P < 0.05) (Fig. 2).

7d

3.2. 15-Ketodihydro-PGF2a

265  220.8 690  612.3 15  8* 32  28.9 0.5  0.8** 4.1  6.3

8 to 10 d

11 to 14 d

foals showed signs of HIE, septicemia was diagnosed in 9 foals, and other diseases (such as diarrhea, bladder rupture, pneumonia, etc.) were found in the remaining 11 newborns. Among the diseased foals, 11 newborns died. Of these, three were premature/dysmature, five were septicemic, two showed signs of HIE, and one with tendon contracture and pneumonia was euthanized for economic reasons.

1035 93  28.7 506  293.1 7  2.6 20  8.3 0.1 0

S. Panzani et al. / Theriogenology 72 (2009) 1032–1040

1036

S. Panzani et al. / Theriogenology 72 (2009) 1032–1040

Fig. 1. 15-Ketodihydro-PGF2a plasma concentrations in control (n = 24) and in surviving and nonsurviving pathologic foals (mean + SD). Numbers of foals involved are indicated within bars. Differences between surviving and nonsurviving foals are represented by the asterisk (*P < 0.05).

Fig. 2. 15-Ketodihydro-PGF2a plasma concentrations in different classes of disease (mean + SD). Numbers of foals involved are indicated within bars. a,bP < 0.05; A,BP < 0.01.

4. Discussion This study investigated the plasma profiles of three hormones involved in physiologic and pathologic processes during neonatal life in healthy and pathologic foals. The control group consisted of normal, healthy neonatal foals, and the pathologic group consisted of spontaneous diseased foals admitted to a NICU and was more heterogeneous in age than the control group. To allow comparisons between the groups, the pathologic foals were grouped according to age at admission. Most of the pathologic foals were hospitalized within the first 2 d of life (78%), highlighting that this period represents the most challenging for the newborn [36].

Among the reasons for admission, HIE was the most prevalent disease detected (35%), followed by septicemia (23%) and prematurity/dysmaturity (15%). The overall mortality rate was 27%. The principal causes of death were septicemia (45%) and prematurity/ dysmaturity (27%), which together caused 73% of deaths. This is in agreement with studies reporting that generalized infection is the major cause of mortality in newborn foals [37]. Statistical analysis revealed significant age-dependent differences in hormone plasma profiles between the normal and diseased groups. 15-KetodihydroPGF2a levels were consistently higher in pathologic foals than that in control foals, with significant

S. Panzani et al. / Theriogenology 72 (2009) 1032–1040

1037

Fig. 3. Cortisol plasma concentrations in control (n = 24) and in surviving and nonsurviving pathologic foals (mean + SD). Numbers of foals involved are indicated within bars.

Fig. 4. Cortisol plasma concentrations in different classes of disease (mean + SD). Numbers of foals involved are indicated within bars.

Fig. 5. Progesterone plasma concentrations in control (n = 24) and in surviving and nonsurviving pathologic foals (mean + SD). Numbers of foals involved are indicated within bars.

1038

S. Panzani et al. / Theriogenology 72 (2009) 1032–1040

Fig. 6. Progesterone plasma concentrations in different classes of disease (mean + SD). Numbers of foals involved are indicated within bars.

differences at Days 2, 5, 6, and 7 and between Days 8 and 10. In normal foals, the high PGM levels present during the first days of life could be related to their roles in completing organ maturation. However, within 10 d of life, the PGM concentrations in normal foals decreased to basal levels [38], whereas PGM levels remained high (over 500 pmol/L) in the pathologic group, even at the end of the observation period. It is possible that the profile observed in the diseased foals was related to a maturational delay or the presence of oxidative stress or inflammation due to the pathologic condition. A similar trend was observed for C plasma concentrations. Cortisol levels were higher in pathologic foals than in control foals, with significant differences at 12 h and Days 2, 3, and 7. In normal foals, the plasma C level presents a low mean level at 12 h of life (17 ng/mL), in agreement with previous reports [38–40]. The higher C concentrations in diseased foals could have been due to the presence of stressors that influence the hypothalamus-pituitaryadrenal axis and induce the production of C. The plasma profile for P4 was similar to those of PGM and C. Whereas P4 levels decreased rapidly to basal levels in normal foals [25], higher concentrations of this hormone were detected in pathologic newborns, with statistically significant differences at 12 h and at 1 and 2 d of age. Although there were no statistical differences found in the following days, pathologic foals showed a trend of elevated P4 plasma concentrations until 7 d of life. Previous studies reported similar results for progestogens [25–27]. On the basis of the high cross-reactivities of our antibody, we could suppose that our results reflect the levels of both P4 and its main metabolites. When we compared the

plasma profiles of surviving and nonsurviving pathologic foals, only their PGM profiles differed significantly, probably because of the relative small number of foals involved. Nonsurviving foals had higher plasma concentrations of PGM at 2 and 3 d of life. However, the C plasma concentrations were consistently higher in nonsurviving foals, which could be explained by great stress preceding the exitus. When we evaluated the plasma profiles of pathologic foals according to disease, PGM was significantly higher in foals affected by HIE on Days 5 and 6. Unfortunately, the small number of subjects studied precluded us from detecting significant differences in all hormone concentrations when analyzed according to specific disease. The PGM profile of hypoxic foals could be explained by the role of some PGF2a metabolites as markers of oxidative stress in newborns affected by intracellular hypoxia [5]. Although not significantly different, the concentration of PGM tended to be lower in premature/dysmature foals at 12 h of age. Considering the role of PGM in the maturational processes of the neonate, low levels during the first hours of life could be related to the multisystemic immaturity of a premature newborn. Similarly, even if not significantly different compared with that of foals affected by other diseases, the low concentrations of C at 12 h and during the first day from birth in premature/dysmature foals could be related to the absence of the increase in fetal C that normally precedes delivery. Without this predelivery surge in C, neonates show signs of prematurity often leading to death associated with multiorgan failure [18]. Premature/dysmature foals had higher P4 concentrations at 12 h of life compared with that of foals affected by other diseases, as has been reported for progestogens [25–27].

S. Panzani et al. / Theriogenology 72 (2009) 1032–1040

In conclusion, these data demonstrate that there are distinct differences in plasma hormone profiles between healthy and pathologic newborn foals. Pathologic foals have higher concentrations of PGM, C, and P4 than those of normal, healthy foals. The differences in hormone concentrations are most marked during the first 2 d of life and could reflect the pathologic foals’ limited ability to cope with the transition between fetal and neonatal life. Alternatively, theses differences could reflect the specific pathologic condition. The virtual absence of significant differences in hormone concentrations between surviving and nonsurviving pathologic foals, even though the few animals involved could have affected this result, suggests that hormone evaluation alone is not suitable for prognosis of survival in newborn foals. Unfortunately, the small number of foals in the specific disease subgroups prohibited us from detecting significant relationships between different hormone concentrations and the most common neonatal pathologies. References [1] Saeed SA, Aftab O, Khawar T, Zaigham MA, Hansraj N, Rasheed H, Mernon SJ. New prospects in the control of arachidonic acid metabolism in the fetus and the neonate. J Med Sci 2003;3(3):192–208. [2] Ivey KN, Srivastava D. The paradoxical patent ductus arteriosus. J Clin Invest 2006;116(1):2863–5. [3] Smith GC. The pharmacology of the ductus arteriosus. Pharm Res 1998;50:35–58. [4] Elliott RB, Starling MB. The effect of prostaglandin F2a in the closure of the ductus arteriosus. Prostaglandins 1972;2(5):399– 403. [5] Reznichenko IuG, Reznichenko GI, Ventskovskyi BM. Prostaglandins content in the blood of newborns with various types of hypoxia. Fiziol Zh 1993;39(2–3):72–6. [6] Friedman Z, Demers LM. Prostaglandin synthetase in the human neonatal kidney. Pediatr Res 1980;14(3):190–3. [7] Benzoni D, Vincent M, Betend B, Sassard J. Urinary escretion of prostaglandins and electrolytes in developing children. Kidney Int 1981;20:386–8. [8] Pace-Asciak CR. Prostaglandin biosynthesis and catabolism in several organs of developing fetal and neonatal animals. In: Coceani F, Olley PM, editors. Advances in Prostaglandin and Thromboxane Research, Volume4. Raven Press; 1978. p. 45–59. [9] Joppich R, Scherer B, Weber PC. Renal prostaglandins: relationship to the development of blood pressure and concentrating capacity in pre-term and full term healthy infants. Eur J Pediatr 1979;132(4):253–9. [10] Mitchell MD, Lucas A, Etches PC, Brunt JD, Turnbull AC. Plasma prostaglandin levels during early neonatal life following term and pre-term delivery. Prostaglandins 1978;16(2): 319–26. [11] Higgins AJ, Lees P. The acute inflammatory process, arachidonic acid metabolism and the mode of action of anti-inlammatory drugs. Equine Vet J 1984;16:163–5.

1039

[12] Bos CL, Rickel DJ, Ritsema T, Peppelenbosch MP, Versteeg HH. Prostanoids and prostanoid receptors in signal transduction. Int J Bioch Cell Biol 2004;36:1187–205. [13] Ward DS, Fessier JF, Bottoms GD, Turek J. Equine endotoxemia: cardiovascular, eicosanoid, hemaatologic, blood chemical, and plasma enzyme alterations. Am J Vet Res 1987;48(7): 1150–6. [14] Elverson CA, Wilson ME. Cortisol: circadian rhythm and response to a stressor. Newborn and Infant Nursing Reviews 2005;5:159–69. [15] Liggins GC. The role of cortisol in preparing the fetus for birth. Reprod Fertil Dev 1994;6:141–50. [16] Fowden AL. Endocrine regulation of fetal growth. Reprod Fertil Dev 1995;7(3):351–63. [17] Ousey JC. Hormone profile and treatments in the late pregnant mare. Vet Clin North Am Equine Practice 2006;22:727–47. [18] Rossdale PD, Ousey JC, Silver M, Fowden A. Studies on equine prematurity 6: guidelines for assessment of foal maturity. Equine Vet J 1984;16(4):300–2. [19] Soliman AT, Taman KH, Rizk MM, Nasr IS, Alrimawy H, Hamido MS. Circulating adrenocorticotropic hormone (ACTH) and cortisol concentrations in normal, appropriate-for-gestational-age newborns versus those with sepsis and respiratory distress: cortisol response to low-dose and standard-dose ACTH tests. Metabolism 2004;53:209–14. [20] Gardner DS, Fletcher AJW, Fowden AL, Giussani DA. Plasma adrenocorticotropin and cortisol concentrations during acute hypoxemia after a reversible period of adverse intrauterine conditions in the ovine fetus during late gestation. Endocrinology 2001;142:589–98. [21] Gardner DS, Fletcher AJW, Bloomfield MR, Fowden AL, Giussani DA. Effects of prevailing hypoxemia, acidaemia or hypoglycaemia upon the cardiovascular, endocrine and metabolic responses to acute hypoxaemia in the ovine fetus. J Physiol 2002;540:351–66. [22] Thoresen M, Satas S, Loberg EM, Whitelaw A, Acolet D, Lindgren C, et al. Twenty-four hours of mild hypothermia in unsedated newborn pigs starting after a severe global hypoxicischemic insult is not neuroprotective. Pediatr Res 2001;50: 405–11. [23] O’Connor SJ, Gardner DS, Ousey JC, Holdstock N, Rossdale P, Edwards CMB, et al. Development of baroflex and endocrine responses to hypotensive stress in newborn foals and lambs. Eur J Physiol 2005;450:298–306. [24] Billiards SS, Walker DW, Canny BJ, Hirst JJ. Endotoxin increases sleep and brain allopregnanolone concentrations in newborn lambs. Pediatr Res 2002;52:892–9. [25] Rossdale PD, Ousey JC, McGladdery AJ, Prandi S, Holdstock N, Grainger L, Houghton E. A retrospective study of increased plasma progestagen concentrations in compromised neonatal foals. Reprod Fertil Dev 1995;7:567–75. [26] Houghton E, Holtamn D, Grainger L, Voller BE, Rossdale PD, Ousey JC. Plasma progestagen concentrations in the normal and dysmature newborn foal. J Reprod Fertil Suppl 1991;44: 609–17. [27] Rossdale PD, Ousey JC, Cottrill CM, Chavatte P, Allen WR, McGladdery AJ. Effects of placental pathology and mammary secretion calcium concentrations and on neonatal adrenocortical function in the horse. J Reprod Fert Suppl 1991;44:579–90. [28] Koterba AM. Physical examination. In: Koterba AM, Drummond WH, Kosch PC, editors. Equine Clinical Neonatology. Lea & Febiger; 1990. p. 71–86.

1040

S. Panzani et al. / Theriogenology 72 (2009) 1032–1040

[29] Rossdale PD. Clinical view of disturbances in equine foetal maturation. Equine Vet J Suppl 1993;14:3–7. [30] Lester GD. Maturity of the neonatal foal. Vet Clin North Am Equine Practice 2005;21(2):333–55. [31] Vaala WE, House JK. Perinatal adaptation, asphyxia, and resuscitation. In: Smith BP, editor. Large Animal Internal Medicine. Mosby; 2002. p. 266–76. [32] Paradis MR. Update on neonatal septicemia. Vet Clin North Am Equine Practice 1994;10:109–35. [33] Kindahl H, Knudsen O, Madej A, Edqvist LE. Progesterone, prostaglandin F2a, PMSG and oestrone sulphate during early pregnancy in the mare. J Reprod Fertil Suppl 1982;32: 353–9. [34] Tamanini C, Giordano N, Chiesa F, Seren E. Plasma cortisol variations induced in the stallion by mating. Acta Endocrinol 1983;102:447–50. [35] Bono G, Cairoli F, Tamanini C, Abrate L. Progesterone, estrogen, LH, FSH and PRL concentrations in plasma during the estrous cycle in goat. Reprod Nutr Dev 1983;23:217–22.

[36] Losinger WC, Traub-Dargatz JL, Sampath RK, Morley PS. Operation-management factors associated with early-postnatal mortality of US foals. Prev Vet Med 2000;47(3):157–75. [37] Peek SF, Darien BJ, Semrad SD, McGuirk S, Lien L, Riseberg A, Marques F, Slack JA, Coombs D. A prospective study of neonatal septicaemia and factors influencing survival. 50th Annual Convention of the American Association of Equine Practitioners, Denver, 2004. [38] Panzani S, Villani M, Govoni N, Kindahl H, Faustini M, Romano G, Veronesi MC. 15-Ketodihydro-PGF2a and cortisol plasma concentrations in newborn foals after spontaneous or oxytocininduced parturition. Theriogenology 2009;71(5):768–74. [39] Rossdale PD, Silver M, Ellis L, Frauenfelder H. Response to adrenal cortex to tetracosactrin (ACTH1-24) in the premature and full-term foal. J Reprod Fertil Suppl 1982;32:545–53. [40] Silver M, Fowden AL, Knox J, Ousey J, Cash R, Rossdale PD. Relationship between circulating tri-iodothyronine and cortisol in the perinatal period in the foal. J Reprod Fertil Suppl 1991;44:619–26.