A lipidomic screen of palmitate-treated MIN6 β-cells links sphingolipid metabolites with endoplasmic reticulum (ER) stress and impaired protein trafficking

Abstract

Saturated fatty acids promote lipotoxic ER (endoplasmic reticulum) stress in pancreatic β-cells in association with Type 2 diabetes. To address the underlying mechanisms we employed MS in a comprehensive lipidomic screen of MIN6 β-cells treated for 48 h with palmitate. Both the overall mass and the degree of saturation of major neutral lipids and phospholipids were only modestly increased by palmitate. The mass of GlcCer (glucosylceramide) was augmented by 70% under these conditions, without any significant alteration in the amounts of either ceramide or sphingomyelin. However, flux into ceramide (measured by [3H]serine incorporation) was augmented by chronic palmitate, and inhibition of ceramide synthesis decreased both ER stress and apoptosis. ER-to-Golgi protein trafficking was also reduced by palmitate pre-treatment, but was overcome by overexpression of GlcCer synthase. This was accompanied by increased conversion of ceramide into GlcCer, and reduced ER stress and apoptosis, but no change in phospholipid desaturation. Sphingolipid alterations due to palmitate were not secondary to ER stress since they were neither reproduced by pharmacological ER stressors nor overcome using the chemical chaperone phenylbutyric acid. In conclusion, alterations in sphingolipid, rather than phospholipid, metabolism are more likely to be implicated in the defective protein trafficking and enhanced ER stress and apoptosis of lipotoxic β-cells.

Heterogeneity of fatty acid metabolism in breast cancer cells underlies differential sensitivity to palmitate‐induced apoptosis

“…Exposure to elevated levels of the saturated FA palmitate induces apoptosis in a range of cells including 3T3 fibroblasts (Kamili et al ., 2015), peripheral blood mononuclear cells (RostamiRad et al ., 2018), human cardiac progenitor cells (Leonardini et al ., 2017), pancreatic β cells (Boslem et al ., 2011; Luo et al ., 2017), macrophages (Kim et al ., 2017), and hepatocytes (Penke et al ., 2017). Palmitate also impairs MDA‐MB‐231 cell proliferation and activates apoptosis (Baumann et al ., 2016; Hardy et al ., 2000, 2003; Kourtidis et al ., 2009; Wu et al ., 2017) via a number of mechanisms, including ER stress (Baumann et al ., 2016; Boslem et al ., 2011), impaired autophagy (RostamiRad et al ., 2018; Wu et al ., 2017), altered NAD metabolism (Penke et al ., 2017), and ceramide synthesis (Luo et al ., 2017).…”

“…Palmitate also impairs MDA‐MB‐231 cell proliferation and activates apoptosis (Baumann et al ., 2016; Hardy et al ., 2000, 2003; Kourtidis et al ., 2009; Wu et al ., 2017) via a number of mechanisms, including ER stress (Baumann et al ., 2016; Boslem et al ., 2011), impaired autophagy (RostamiRad et al ., 2018; Wu et al ., 2017), altered NAD metabolism (Penke et al ., 2017), and ceramide synthesis (Luo et al ., 2017). Importantly, many of the deleterious effects of palmitate on cellular function are mitigated by cotreatment with other FAs, in particular the monounsaturated FA oleate (Colvin et al ., 2017; Kim et al ., 2017; Penke et al ., 2017; Sargsyan et al ., 2016), which itself is pro‐proliferative and activates phosphoinositide 3‐kinase signaling (Hardy et al ., 2000).…”

Oxidative and endoplasmic reticulum stress in β-cell dysfunction in diabetes

“…Palmitate and the unsaturated fatty acid oleate also affect Ca 2C handling in the ER leading to protein misfolding and ER stress apparently independently of oxidative stress (Cnop et al 2010), although oleate causes much milder ER stress than palmitate (Cunha et al 2008). Other reports indicate that palmitate rapidly increases the saturated lipid content of the ER, which leads to compromised ER morphology and integrity resulting in ER stress, and interestingly, NEFA exposure causes ER stress in yeast demonstrating a fundamental mechanism not restricted to mammalian cells (Borradaile et al 2006, Pineau & Ferreira 2010, Boslem et al 2011. Desaturation of palmitate with stearoyl coenzyme A desaturase 1 (SCD1) reduces palmitateinduced cell death, whereas deficiency increases apoptosis (Busch et al 2005, Thorn et al 2010, Green & Olson 2011.…”

Sphingolipid regulators of cellular dysfunction in Type 2 diabetes mellitus: a systems overview

“…The ER is important in cellular function and has been shown to contribute to the insulin resistance that leads to the development of T2DM [34]. Studies have implicated ceramide as both a causative agent and a product of ER stress [35,36]. Roles for specific sphingolipid species and isoforms of the enzymes of sphingolipid metabolism are presently emerging in diabetes and other contexts, and have been thoroughly reviewed elsewhere [37,38].…”

“…Similarly, ceramide production was shown to promote ER stress in β-cells through mechanisms that are still being elucidated [56,57]. Recently, altered membrane dynamics have been proposed as a key mechanism by which this occurs.…”