Postnatal effects of intrauterine treatment of the growth-restricted ovine fetus with intra-amniotic insulin-like growth factor-1

Abstract

Key points: Fetal growth restriction increases the risk of fetal and neonatal mortality and morbidity, and contributes to increased risk of chronic disease later in life. Intra-amniotic insulin-like growth factor-1 (IGF1) treatment of the growth-restricted ovine fetus improves fetal growth, but postnatal effects are unknown. Here we report that intra-amniotic IGF1 treatment of the growth-restricted ovine fetus alters size at birth and mechanisms of early postnatal growth in a sex-specific manner. We also show that maternal plasma C-type natriuretic peptide (CNP) products are related to fetal oxygenation and size at birth, and hence may be useful for non-invasive monitoring of fetal growth restriction. Intrauterine IGF1 treatment in late gestation is a potentially clinically relevant intervention that may ameliorate the postnatal complications of fetal growth restriction.

Abstract: Placental insufficiency-mediated fetal growth restriction (FGR) is associated with altered postnatal growth and metabolism, which are, in turn, associated with increased risk of adult disease. Intra-amniotic insulin-like growth factor-1 (IGF1) treatment of ovine FGR increases growth rate in late gestation, but the effects on postnatal growth and metabolism are unknown. We investigated the effects of intra-amniotic IGF1 administration to ovine fetuses with uteroplacental embolisation-induced FGR on phenotypical and physiological characteristics in the 2 weeks after birth. We measured early postnatal growth velocity, amino-terminal propeptide of C-type natriuretic peptide (NTproCNP), body composition, tissue-specific mRNA expression, and milk intake in singleton lambs treated weekly with 360 μg intra-amniotic IGF1 (FGRI; n = 13 females, 19 males) or saline (FGRS; n = 18 females, 12 males) during gestation, and in controls (CON; n = 15 females, 22 males). There was a strong positive correlation between maternal NTproCNP and fetal oxygenation, and size at birth in FGR lambs. FGR lambs were ∼20% lighter at birth and demonstrated accelerated postnatal growth velocity. IGF1 treatment did not alter perinatal mortality, partially abrogated the reduction in newborn size in females, but not males, and reduced accelerated growth in both sexes. IGF1-mediated upregulation of somatotrophic genes in males during the early postnatal period could suggest that treatment effects are associated with delayed axis maturation, whilst treatment outcomes in females may rely on the reprogramming of nutrient-dependent mechanisms of growth. These data suggest that the growth-restricted fetus is responsive to intra-amniotic intervention with IGF1, and that sex-specific somatotrophic effects persist in the early postnatal period.

Keywords: amino-terminal propeptide of C-type natriuretic peptide; fetal growth restriction; growth and metabolism; insulin-like growth factor-1; intrauterine intervention; somatotrophic maturation.

Figure 1. Characteristics of experimental groups and animal use

Animals utilised during the project according to experimental group and sex. Dashed boxes report exclusions, deaths and killing. Split boxes reporting postnatal deaths indicate the number of females (top left) and males (bottom right) for each experimental group: control (CON), saline‐treated fetal growth restricted (FGRS), and insulin‐like growth factor 1‐treated fetal growth restricted (FGRI). Dual X‐ray absorptiometry (DXA), biopsies, and milk intake were conducted in all groups. Circumstances constituting exclusion from the project and/or killing were: ^*pregnancy toxaemia, ^†mammary abscess n = 5, aspiration of rumen contents prior to intubation n = 1, twin pregnancy n = 2 and intrauterine infection n = 1; ^‡twin lambs n = 2, intestinal atresia n = 1; °preterm (<136 dGA) birth n = 1.

Figure 2. Fetal whole blood measures and maternal mean plasma N‐terminal pro‐C‐type natriuretic peptide

Data are means ± SEM. Whole blood measures of partial pressure of carbon dioxide ($P_{\mathrm{aC}{\mathrm{O}}_2}$, mmHg; A); partial pressure of oxygen ($P_{\mathrm{a}{\mathrm{O}}_2}$, mmHg; B); pH (C); glucose (mmol·L⁻¹; D); and) lactate (mmol·L⁻¹; E) in FGRS (grey, n = 20–34) and FGRI (black, n = 20–32) fetuses; and the correlation between mean maternal N‐terminal pro‐C‐type natriuretic peptide (NTproCNP, pmol·L⁻¹) during the embolisation period (103–107 dGA) and fetal $P_{\mathrm{a}{\mathrm{O}}_2}$ in growth‐restricted FGR (white, n = 2), FGRS (grey, n = 7), and FGRI (black, n = 7) fetuses (F). Symbols denote significant differences over time (RM ANOVA: i, P < 0.05).

Figure 3. Maternal mean plasma N‐terminal pro‐C‐type natriuretic peptide and offspring size at birth

Data are means ± SEM. The correlation between mean maternal N‐terminal pro‐C‐type natriuretic peptide (NTproCNP) during the embolisation period (103–107 dGA) and birthweight (A), crown–rump length (B), and hock–toe length (C) in FGRS (grey, n = 7), and FGRI (black, n = 7) lambs.

Figure 4. Early postnatal growth in females

Data are means ± SEM. A, bodyweight (WT); B, crown–rump length (CRL); C, chest circumference; D, abdominal circumference (Abdo); E, hock–toe length (HT); F, hindlimb length (HL); G, forelimb length (FL); H, biparietal diameter (BPD). CON (white, n = 15), FGRS (grey, n = 16–18), FGRI (black, n = 13). Roman numerals denote the significant difference between experimental groups (RM ANOVA: ⁱ P < 0.05; ⁱⁱ P < 0.01; ⁱⁱⁱtime × experimental group interaction P < 0.05), symbols refer to post hoc P values for differences between experimental groups at each time point (ANOVA: CON vs. FGRS: ^* P < 0.05; CON vs. FGRI: ^† P < 0.05).

Figure 5. Early postnatal growth in males

Data are means ± SEM. A, bodyweight (WT); B, crown–rump length (CRL); C, chest circumference; D, abdominal circumference (Abdo); E, hock—toe length (HT); F, hindlimb length (HL); G, forelimb length (FL); H, biparietal diameter (BPD). CON (white, n = 21–22), FGRS (grey, n = 13), FGRI (black, n = 18‐19). Symbols denote significant difference between experimental groups (RM ANOVA: ⁱ P < 0.05; ⁱⁱ P < 0.01), and refer to post hoc P values for differences between experimental groups at each time point (ANOVA: CON vs. FGRS: ^* P < 0.05; CON vs. FGRI: ^† P < 0.05).

Figure 6. Plasma IGF1 characteristics at birth

Plasma concentrations of IGF1 in females (A) and males (B), and their correlation with weight at birth (C and D, respectively). Female CON (white) n = 13, FGRS (grey) n = 13, FGRI (black) n = 12, and male CON (white) n = 15, FGRS (grey) n = 11 and FGRI (black) n = 9 lambs. Data are means ± SEM. Symbols refer to post hoc P values for differences between experimental groups (ANOVA: CON vs. FGRS: ^* P < 0.05).

Figure 7. Plasma hormone and metabolite concentrations over the first 2 weeks after birth in females

Plasma concentrations of glucose (A) insulin (B), non‐esterified fatty acids (NEFA; F), urea (G) and lactate (H), and the change in insulin over the first (C) and second (D) weeks, and the natural log transformation (ln) of the glucose to insulin (G:I) ratio in the first 2 weeks after birth (E). CON (white) n = 14, FGRS (grey) n = 11–14 and FGRI (black) n = 13 lambs. Data are means ± SEM. Roman numerals denote the significant difference between experimental groups (RM ANOVA: ⁱ P < 0.05, ⁱⁱⁱtime × experimental group interaction P < 0.05). Symbols denote post hoc P values for differences between experimental groups at each time point (ANOVA: CON vs. FGRS: ^* P < 0.05; CON vs. FGRI: ^† P < 0.05; ^‡ P < 0.05 FGRS vs. FGRI).

Figure 8. Plasma hormone and metabolite concentrations over the first 2 weeks after birth in males

Plasma concentrations of glucose (A), insulin (B), non‐esterified fatty acids (NEFA; F), urea (G) and lactate (H), and the change in insulin over the first (C) and second (D) weeks, and the natural log transformation (ln) of the glucose to insulin (G:I) ratio in the first 2  weeks after birth (E). CON (white) n = 16, FGRS (grey) n = 11 and FGRI (black) n = 10–11 lambs. Data are means ± SEM. Roman numerals denote significant difference between experimental groups (RM ANOVA: ⁱⁱⁱtime × experimental group interaction P < 0.05). Symbols denote post hoc P values for differences between experimental groups at each time point (ANOVA: CON vs. FGRS: ^* P < 0.05; CON vs. FGRI: ^† P < 0.05).