Enhanced or reduced fetal growth induced by embryo transfer into smaller or larger breeds alters post-natal growth and metabolism in pre-weaning horses

Abstract

In equids, placentation is diffuse and nutrient supply to the fetus is determined by uterine size. This correlates with maternal size and affects intra-uterine development and subsequent post-natal growth, as well as insulin sensitivity in the newborn. Long-term effects remain to be described. In this study, fetal growth was enhanced or restricted through ET using pony (P), saddlebred (S) and draft (D) horses. Control P-P (n = 21) and S-S (n = 28) pregnancies were obtained by AI. Enhanced and restricted pregnancies were obtained by transferring P or S embryos into D mares (P-D, n = 6 and S-D, n = 8) or S embryos into P mares (S-P, n = 6), respectively. Control and experimental foals were raised by their dams and recipient mothers, respectively. Weight gain, growth hormones and glucose homeostasis were investigated in the foals from birth to weaning. Fetal growth was enhanced in P-D and these foals remained consistently heavier, with reduced T3 concentrations until weaning compared to P-P. P-D had lower fasting glucose from days 30 to 200 and higher insulin secretion than P-P after IVGTT on day 3. Euglycemic clamps in the immediate post-weaning period revealed no difference in insulin sensitivity between P-D and P-P. Fetal growth was restricted in S-P and these foals remained consistently lighter until weaning compared to S-D, with elevated T3 concentrations in the newborn compared to S-S. S-P exhibited higher fasting glycemia than S-S and S-D from days 30 to 200. They had higher maximum increment in plasma glucose than S-D after IVGTT on day 3 and clamps on day 200 demonstrated higher insulin sensitivity compared to S-D. Neither the restricted nor the enhanced fetal environment affected IGF-1 concentrations. Thus, enhanced and restricted fetal and post-natal environments had combined effects that persisted until weaning. They induced different adaptive responses in post-natal glucose metabolism: an early insulin-resistance was induced in enhanced P-D, while S-P developed increased insulin sensitivity.

Figure 1. Establishment of control and experimental pregnancies by artificial insemination (AI) and embryo transfer (ET), respectively.

Figure 2. Mares' parameters from the 5^th gestational month to weaning in the five groups.

A: body weight. B: body scores. C: plasma NEFA. D: plasma leptin. (P-P: Pony in Pony (•), P-D: Pony in Draft (○), S-P: Saddlebred in Pony (▾), S-S: Saddlebred in Saddlebred (▪), S-D: Saddlebred in Draft (Δ)). Curves are presented as medians and interquartile ranges.

Figure 3. Foals' body weights from birth to weaning in the five groups.

A: P-P (•) vs S-S (▪). B: P-P (•) vs P-D (○). C: S-P (▾) vs S-S (▪) vs S-D (Δ) (P-P: Pony in Pony, P-D: Pony in Draft, S-P: Saddlebred in Pony, S-S: Saddlebred in Saddlebred, S-D: Saddlebred in Draft). Curves are presented as medians and interquartile ranges. The median values between the asterisks differ significantly from each other (F1-LD-F1 model followed by Mann-Whitney or Kruskal-Wallis test, p<0.05). In graph C, median values under the lower and upper dotted lines between asterisks significantly differ between S-P and S-S and between S-P and S-D, respectively. NB: Different scales were used for A, B and C in order to show the differences.

Figure 4. Foals' plasma IGF-1 levels from birth to weaning in the five groups.

A: P-P (full yellow) vs S-S (full green). B: P-P (full yellow) vs P-D (chequered blue). C: S-P (striped pink) vs S-S (full green) vs S-D (chequered red) (P-P: Pony in Pony, P-D: Pony in Draft, S-P: Saddlebred in Pony, S-S: Saddlebred in Saddlebred, S-D: Saddlebred in Draft). Curves are presented as medians and interquartile ranges. The median values under the asterisks differ significantly from each other (F1-LD-F1 model followed by Mann-Whitney or Kruskal-Wallis test, p<0.05).

Figure 5. Foals' plasma T₃ levels from birth to weaning in the five groups.

A: P-P (full yellow) vs S-S (full green). B: P-P (full yellow) vs P-D (chequered blue). C: S-P (striped pink) vs S-S (full green) vs S-D (chequered red) (P-P: Pony in Pony, P-D: Pony in Draft, S-P: Saddlebred in Pony, S-S: Saddlebred in Saddlebred, S-D: Saddlebred in Draft). Curves are presented as medians and interquartile ranges and the scale on the y-axis is semi-logarithmic. The median values under the asterisks differ significantly from each other (F1-LD-F1 model followed by Mann-Whitney or Kruskal-Wallis test, p<0.05.

Figure 6. Foals' plasma T₄ levels from birth to weaning in the five groups.

A: P-P (full yellow) vs S-S (full green). B: P-P (full yellow) vs P-D (chequered blue). C: S-P (striped pink) vs S-S (full green) vs S-D (chequered red) (P-P: Pony in Pony, P-D: Pony in Draft, S-P: Saddlebred in Pony, S-S: Saddlebred in Saddlebred, S-D: Saddlebred in Draft). Curves are presented as medians and interquartile ranges and the scale on the y-axis is semi-logarithmic. The median values under the asterisks differ significantly from each other (F1-LD-F1 model followed by Mann-Whitney or Kruskal-Wallis test, p<0.05).

Figure 7. Foals' fasting glucose from birth to weaning in the five groups.

A: P-P (•) vs S-S (▪). B: P-P (•) vs P-D (○). C: S-P (▾) vs S-S (▪) vs S-D (Δ) (P-P: Pony in Pony, P-D: Pony in Draft, S-P: Saddlebred in Pony, S-S: Saddlebred in Saddlebred, S-D: Saddlebred in Draft). Curves are presented as medians and interquartile ranges. The median values under the asterisks differ significantly from each other (F1-LD-F1 model followed by Mann-Whitney or Kruskal-Wallis test, p<0.05). In graph C, median values under the simple and double asterisks significantly differ between S-P and S-S and between S-P and S-D, respectively.

Figure 8. Changes in the plasma concentrations of glucose and insulin in response to glucose bolus in the five groups.

A: Glycemia (A1) and insulinemia (A2) in P-P (•) vs P-D (○). B: Glycemia (B1) and insulinemia (B2) in S-P (▾) vs S-S (▪) vs S-D (Δ). C: Area under the curve for glucose (C1) and insulin (C2) in P-P (full yellow), P-D (chequered blue), S-P (striped pink), S-S (full green) and S-D (chequered red) (P-P: Pony in Pony, P-D: Pony in Draft, S-P: Saddlebred in Pony, S-S: Saddlebred in Saddlebred, S-D: Saddlebred in Draft). Curves are presented as medians and interquartile ranges. The median values under the asterisks differ significantly from each other (F1-LD-F1 model followed by Mann-Whitney or Kruskal-Wallis test, p<0.05). In graph B2, the asterisk indicates a significant difference between S-P and S-S.