Introduction: Pregestational maternal obesity (MO) is associated with adverse cardio-metabolic outcome in the offspring compared to normal pregestational maternal weight (MN). One of the early stages contributing to cardiovascular disease is insulin desensitization. Endoplasmic reticulum (ER) stress contributes to obesity-associated insulin desensitization via the activation of inositol-requiring enzyme 1 alpha (IRE1α), which inhibits insulin receptor substrate 1 (IRS1) – a key protein in the insulin signaling pathway – by activating c-Jun N-terminal kinase (JNK). Our previous studies showed that human umbilical vein endothelial cells (HUVECs) from MO (HUVECs-MO) show insulin desensitization (i.e. a lower insulin-induced nitric oxide (NO) generation) and ER stress, whose reduction improved the insulin response. However, the mechanisms for ER stress-associated insulin-desensitization in HUVECs-MO is not known. We hypothesize that ER stress-associated insulin desensitization in HUVECs-MO is mediated by IRE1α, which is not observed in MN.
Methods: HUVECs were isolated from MN (HUVECs-MN) or MO deliveries at the Hospital Clínico UC-CHRISTUS, treated with siRNA against IRE1α or pre-incubated (8h) with or without tunicamycin (ER stress inducer), KIRA6 (IRE1α inhibitor), SP600125 (JNK inhibitor), and subsequently with insulin (20 min). The activation state of IRE1α, JNK, and IRS1 was evaluated by Western blot and the NO levels in the absence or presence of L-NAME (NOS inhibitor) were assayed using the DAF-FM probe.
Results: Insulin did not increase NO generation in HUVECs-MO, but did in HUVECs-MN (P=0.002). Basal IRE1α/JNK (p=0.017) activity and IRS1 inhibition (p=0.031) were increased in HUVECs-MO compared to MN. Both knockdown and inhibition of IRE1α reduced JNK activation (p=0.004) and IRS1 inhibition (p=0.024). IRE1α inhibition increased insulin-induced NO generation in HUVECs-MO (p=0.030).
Conclusion: MO-associated ER stress impairs insulin-induced NO generation in HUVECs through the activation of IRE1α. This study contributes to a better understanding of the mechanisms involved in the MO-associated insulin desensitization in fetoplacental endothelium.
Marcelo Farías– Assistant professor, Department of Obstetrics, Division of Obstetrics and Gynaecology, Faculty of Medicine, Pontificia Universidad Católica de Chile
Luis Sobrevia– Professor, Department of Obstetrics, Division of Obstetrics and Gynaecology, Faculty of Medicine, Pontificia Universidad Católica de Chile