Endoplasmic reticulum stress regulates inflammation and insulin resistance in skeletal muscle from pregnant women
Introduction
Gestational diabetes mellitus (GDM), defined as glucose intolerance with first onset during pregnancy, affects up to 18% of all pregnant women (Coustan et al., 2010). Advancing maternal age, ethnicity and obesity are known risk factors for GDM (Ferrara, 2007). GDM is associated with pregnancy complications and long-term adverse health problems for both mother and infant (Perkins et al., 2007, Al-Khalifah et al., 2012, Mitanchez, 2010). Pregnancy complications associated with GDM include fetal macrosomia, shoulder dystocia and higher rates of Caesarean delivery. Neonatal complications include jaundice, respiratory distress and hypoglycaemia. The long-term health risks for mothers and their infants include the development of cardiovascular diseases, obesity, type 2 diabetes and certain cancers later in life (Dabelea et al., 2000). With the growing rates of maternal obesity and GDM in developed countries, they are of significant public health concern with enormous long-term financial costs to the health care system and the community (Hossain et al., 2007).
Peripheral insulin resistance (Cani et al., 2007, Friedman et al., 2008, Borst, 2004, Catalano et al., 2003, Friedman et al., 1999, Kuhl, 1991) is a central feature of pregnancies complicated by maternal obesity and/or GDM. Insulin sensitivity is about 56% less in GDM pregnancies (Kuhl, 1991) and 47% less in obese normal glucose tolerant (NGT) pregnant women when compared to lean pregnant NGT women (Catalano et al., 1999). This decrease in insulin sensitivity is caused by multiple defects in the insulin signalling pathway in skeletal muscle of GDM and obese women (Friedman et al., 1999, Colomiere et al., 2009, Shao et al., 2002, Barbour et al., 2006). This pronounced peripheral insulin resistance increases nutrient availability to the fetus, which in turn increases the risk of fetal adiposity (Lain and Catalano, 2007) and also the development of childhood obesity and other metabolic dysfunctions later on in life (Dabelea et al., 2000).
Inflammation and endotoxemia are also key features of GDM and obese pregnancies (Lappas, 2014a, Lappas, 2014b, Lappas, 2014c, Lappas, 2014d, Radaelli et al., 2003, Basu et al., 2011, Sobel et al., 1992, Dhurandhar et al., 1997). We have previously shown that sterile inflammation and infections by bacteria or viruses can induce inflammation and insulin resistance in tissues from pregnant women (Lappas, 2014a, Lappas, 2014b, Lappas, 2014c, Lappas, 2014d, Lappas, 2015, Liong and Lappas, 2015a, Liong and Lappas, 2015b). There is, however, a paucity of studies that have investigated the mechanisms involved in peripheral insulin resistance and inflammation in skeletal muscle.
There is now increasing evidence that the inositol requiring enzyme 1α (IRE1α) arm of the endoplasmic reticulum (ER) stress response plays a key role in regulating inflammation and insulin resistance. Various endogenous and exogenous cellular insults such as viral infection, inflammation and environmental toxins are known to trigger the ER stress response (Shen et al., 2004) resulting in the induction of pro-inflammatory cytokines, chemokines and other mediators of the immune response in gestational and non-gestational tissues (Liong and Lappas, 2015a, Liong and Lappas, 2015b, Kim et al., 2013, Gargalovic et al., 2006, Martinon et al., 2010, Liong and Lappas, 2014). ER stress has also been implicated to be responsible for the development of peripheral insulin resistance, obesity and type 2 diabetes (Ozcan et al., 2004, Ozcan et al., 2006. Indeed, we have previously described the IRE1α arm of the ER stress response is increased in maternal adipose tissue from pregnancies complicated by GDM and obesity (Liong and Lappas, 2015a, Liong and Lappas, 2015b).
To our knowledge there have been no studies on the role of the IRE1α pathway of the ER stress response in skeletal muscle in the context of obesity in pregnancy and/or GDM. We hypothesise that (i) the IRE1α arm of the ER stress response is increased in skeletal muscle in pregnancies complicated by maternal obesity and/or GDM; (ii) inflammation and infection activate the IRE1α arm of the ER stress response in skeletal muscle; and (iii) inhibition of inflammation- and infection-induced ER stress decreases inflammation, activates the insulin signalling pathway and increases glucose uptake. Inflammation and insulin resistance, tissues were obtained from normal glucose tolerant (NGT) women and stimulated with either bacterial lipopolysaccharide (LPS; a toll-like receptor (TLR)4 ligand), polyinosinic-polycytidylic acid (poly(I:C); a TLR3 ligand), or pro-inflammatory cytokines (i.e. IL-1β, TNF-α) to generate models of obese and GDM-like state. To inhibit ER stress, we used (i) the ER stress inhibitor tauroursodeoxycholic acid (TUDCA), which is an endogenous bile acid derivative that acts as a potent chemical chaperone that inhibits the ER stress response (Ozcan et al., 2006, Miller et al., 2007); and (ii) siRNA specific knockdown of ER stress components IRE1α and GRP78 in primary human skeletal muscle cells.
Section snippets
Tissue collection and preparation
Human skeletal muscle tissue (pyramidalis) was obtained (with the Research Ethics Committee of Mercy Health approval) from consenting women who delivered healthy, singleton infants at term (>37 weeks gestation). Indications for Caesarean section were breech presentation and/or previous Caesarean section. Women with any underlying medical conditions such as pre-existing diabetes, asthma, polycystic ovarian syndrome, preeclampsia and macrovascular complications were excluded from this study.
ER stress is increased in skeletal muscle tissue from obese pregnant women
Skeletal muscle tissue was obtained from 12 NGT women at the time of term Caesarean section (in the absence of labour). Women were grouped as being either lean (n = 6 patients) or obese (n = 6 patients) based on pre-pregnancy BMI. Demographic data is summarised in Table 1. Maternal age, gestational age at delivery, and fetal birth weight did not differ between NGT obese versus lean women. As expected, maternal BMI (both pre-pregnancy and at delivery) was higher in obese compared to lean women.
Discussion
This study, for the first time, has demonstrated that the IRE1α arm of the ER stress pathway is increased in skeletal muscle tissue from obese pregnant women and women with GDM. Specifically, we found the expression of ER stress proteins IRE1α and XBP-1s, but not GRP78, were elevated in skeletal muscle of obese NGT women when compared to lean NGT women. Further, in lean GDM women, IRE1α, GRP78 and XBP-1s were also higher compared to lean NGT women. On the other hand, in obese women, only IRE1α
Disclosure statement
The authors have nothing to disclose.
Funding
Associate Professor Martha Lappas is supported by a Career Development Fellowship by the National Health and Medical Research Council (NHMRC; grant no. 1047025). Dr. Stella Liong is a recipient of the Glyn White Research Fellowship by the Royal Australian and New Zealand College of Obstetricians and Gynaecologists (RANZCOG) Research Foundation. This work was funded by the Norman Beischer Medical Research Foundation.
Acknowledgements
The following are gratefully acknowledged: Gillian Barker (Obstetrics, Nutrition and Endocrinology Group, Department of Obstetrics and Gynaecology, University of Melbourne) for her technical assistance; the clinical Research Midwives Genevieve Christophers, Gabrielle Pell and Rachel Murdoch for sample collection; and the Obstetrics and Midwifery staff of the Mercy Hospital for Women for their co-operation.
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