Placental Insulin Receptor Transiently Regulates Glucose Homeostasis in the Adult Mouse Offspring of Multiparous Dams

Abstract

In pregnancies complicated by maternal obesity and gestational diabetes mellitus, there is strong evidence to suggest that the insulin signaling pathway in the placenta may be impaired. This may have potential effects on the programming of the metabolic health in the offspring; however, a direct link between the placental insulin signaling pathway and the offspring health remains unknown. Here, we aimed to understand whether specific placental loss of the insulin receptor (InsR) has a lasting effect on the offspring health in mice. Obesity and glucose homeostasis were assessed in the adult mouse offspring on a normal chow diet (NCD) followed by a high-fat diet (HFD) challenge. Compared to their littermate controls, InsR KO^placenta offspring were born with normal body weight and pancreatic β-cell mass. Adult InsR KO^placenta mice exhibited normal glucose homeostasis on an NCD. Interestingly, under a HFD challenge, adult male InsR KO^placenta offspring demonstrated lower body weight and a mildly improved glucose homeostasis associated with parity. Together, our data show that placenta-specific insulin receptor deletion does not adversely affect offspring glucose homeostasis during adulthood. Rather, there may potentially be a mild and transient protective effect in the mouse offspring of multiparous dams under the condition of a diet-induced obesogenic challenge.

Keywords: fetal programming; gestational diabetes; multiparity; obesity; placental insulin receptor; type 2 diabetes.

Figure 1

Newborn offspring with placenta-specific InsR ablation presented with normal body weight and pancreatic β-cell mass. (A) Experimental schematic for generating InsR KO^placenta and littermate controls. (B) GFP reporter (Cyp19-cre; CAG^+/+) expressing endogenous green fluorescence (GFP) in a mouse embryonic E17.5 placenta section compared to the non-GFP control (CAG^+/+) (scale bars: 500 μm). (C) InsR mRNA expression in the E17.5 InsR KO^placenta compared to controls by quantitative reverse transcription polymerase chain reaction (qPCR) (n = 12, 13). (D) Histology of the E17.5 male control placentas (left) compared to InsR KO^placenta (right) (scale bars: 500 μm). Background of images was subtracted post-imaging. (E) Gross morphology of the male and (E’) female control (left) and the InsR KO^placenta (right) newborn pancreata. (F) Whole pancreatic sections (scale bars: 500 μm) of the male newborn control (left) and the InsR KO^placenta (right) mice immunostained for insulin-positive islets (green) and DAPI (blue). Single islet β-cells (scale bars: 50 μm) shown as insets. Single islet images were cropped post-imaging to highlight the single islet. (G) Beta cell mass for the male (n = 5) and (G’) female (n = 5) newborns. The values are reported as the means ± SEM, ** p <0.01. E: embryonic; P: postnatal; BW: body weight; BG: blood glucose; IPGTT: intraperitoneal glucose tolerance test; IPITT; intraperitoneal insulin tolerance test; NCD: normal chow diet.

Figure 2

Adult InsR KO^placenta mice displayed normal glucose homeostasis on a normal chow diet. (A) Body weight in all the males (n = 9, 10) and (A’) the female (n = 13) InsR KO^placenta and control mice on a normal chow diet. Body weight for the offspring presented as combined data from parity ≥ 3 and ≤2 litters. (B) IPGTT and AUC analysis (right) for the male (n = 4) and (B’) female (n = 4) mice from parity ≥ 3 on a normal chow diet at 9 weeks of age. (C) IPITT and AUC analysis (right) for the males (n = 4) and (C’) females (n = 4) from parity ≥ 3 at 11 weeks of age. Blood glucose values for IPITT are expressed as the baseline percentage of blood glucose. The values are reported as the means ± SEM, * p < 0.05.

Figure 3

Adult male InsR KO^placenta offspring demonstrated lower body weight under a high-fat diet challenge. (A) Schematic of experimental design for a high-fat diet (HFD). (B) Body weight monitored in all the males (n = 6, 7) and (B’) females (n = 8, 9) from parity ≥ 3 and ≤2 across 17 weeks on a HFD. (C) Non-fasting blood glucose levels for all the males (n = 6, 7) and (C’) females (n = 8, 9) from parity was ≥3 and ≤2 (measured across 17 weeks on a HFD). Measurements not included for T = 4, 8, 10, 14, and 18 weeks on a HFD due to phenotyping performed on these weeks. (D) Fasting blood glucose measured for the males (n = 4, 7) and (D’) females (n = 4, 9) from parity ≥ 3 and ≤2 at 4, 8, and 10 weeks on a HFD. (E) Fat mass for the males (n = 6, 7) and (E’) females (n = 8, 9) from parity ≥ 3 and ≤2 as assessed by EchoMRI at 18–19 weeks on a HFD. (F) Lean mass for the males (n = 6, 7) and (F’) females (n = 8, 9) from parity ≥ 3 and ≤2 as assessed by EchoMRI at 18–19 weeks on a HFD. (G) Pancreas weight for the males (n = 6, 7) and (G’) females (n = 8, 9) from parity ≥ 3 and ≤2 upon harvest at 19–20 weeks on a HFD. The values are reported as the means ± SEM, * p < 0.05. P: postnatal; BW: body weight; BG: blood glucose; IPGTT: intraperitoneal glucose tolerance test; IPITT; intraperitoneal insulin tolerance test; HFD: high-fat diet.

Figure 4

Adult male InsR KO^placenta mice from multiparous dams presented with a mild and transient improved glucose homeostasis on a high-fat diet challenge. (A) IPGTT and AUC analysis (right) for the male (n = 4) and (A’) female (n = 4) mice from parity ≥ 3 at 8 weeks on a HFD. Repeated measures two-way ANOVA revealed significant differences (p = 0.0332) between the InsR KO^placenta and control males at 8 weeks on a HFD. A separate unpaired t-test performed specifically for T = 60 min revealed p = 0.0593. (B) IPITT and AUC analysis (right) for the males (n = 4) and (B’) females (n = 4) from parity ≥ 3 at 14 weeks on a HFD. Blood glucose values for IPITT are expressed as the baseline percentage of blood glucose. Repeated measures two-way ANOVA analysis revealed a p-value of 0.0867 in the males at 14 weeks on a HFD. A separate unpaired t-test performed specifically for T = 30 and T = 60 min revealed p = 0.0843 and p = 0.0600, respectively. The values are reported as the means ± SEM, * p < 0.05.