Prevention of Hepatic Steatosis and Hepatic Insulin Resistance by Knockdown of cAMP Response Element-Binding Protein

Abstract

Summary In patients with poorly controlled type 2 diabetes mellitus (T2DM), hepatic insulin resistance and increased gluconeogenesis contributes to fasting and postprandial hyperglycemia. Since CREB is a key regulator of gluconeogenic gene expression, we hypothesized that decreasing hepatic CREB expression would reduce fasting hyperglycemia in rodent models of T2DM. In order to test this hypothesis, we used a CREB-specific antisense oligonucleotide (ASO) to knock down CREB expression in liver. CREB ASO treatment dramatically reduced fasting plasma glucose concentrations in ZDF rats, ob/ob mice and a STZ-treated high-fat fed rat model of T2DM. Surprisingly, CREB ASO treatment also decreased plasma cholesterol and triglyceride concentrations, as well as hepatic triglyceride content due to decreases in hepatic lipogenesis. These results suggest that CREB is an attractive therapeutic target for correcting both hepatic insulin resistance and dyslipidemia associated with NAFLD and T2DM by down regulation of both lipogenic and gluconeogenic gene expression.

PKA phosphorylation couples hepatic inositol-requiring enzyme 1α to glucagon signaling in glucose metabolism

“…S10B), accompanied by a considerable reduction of XBP1 splicing. This finding clearly demonstrates that the obesity-associated increase of hepatic IRE1α phosphorylation indeed primarily stemmed from dysregulation of PKA activity (33), rather than from classical ER stress resulting from increased workload of protein folding. Conceivably, in the face of obesity, this PKA-dependent hyperactivation of IRE1α may arise largely from hyperglucagonemia, which plays a critical part in type 2 diabetes (26,35).…”

“…5A), suggesting simultaneous hyperactivation of PKA and PERK or PKR kinases under obesity-associated metabolic ER stress (20,33,34). Surprisingly, this hyperactivation state of IRE1α in db/db livers, as detected by prominently elevated Ser 724 phosphorylation, was not associated with increased Xbp1 mRNA splicing (Fig.…”

cAMP-responsive Element-binding Protein (CREB)-regulated Transcription Coactivator 2 (CRTC2) Promotes Glucagon Clearance and Hepatic Amino Acid Catabolism to Regulate Glucose Homeostasis

“…Similarly, in T2DM, fasting hyperglycemia is attributable to increased endogenous glucose production (11,12), much of which results from hepatic gluconeogenesis (13). Surprisingly, however, expression of G6Pase and PEPCK correlates poorly with rates of HGP in diabetic human and rodent livers (14), even though inhibition of CREB and FoxO1 can correct this defect (15,16). These results suggest that pathologic induction of gluconeogenesis during T2D is attributable to mechanisms other than just transcription of these prototypical, fasting-induced gene products.…”

“…CRTC2 and control ASO solutions were prepared in normal saline and injected intraperitoneally twice a week at a dose of 37.5 mg/kg body weight for 4 weeks to achieve maximal knockdown. Delivery of ASO by this method has been shown to result in target knockdown in liver, white adipose tissue, kidney, and macrophages (15,16). The T1D model was induced with a 65 mg/kg intraperitoneal streptozotocin injection into SD rats fed normal chow following 4 weeks of ASO injections.…”

A Role for Mitochondrial Phosphoenolpyruvate Carboxykinase (PEPCK-M) in the Regulation of Hepatic Gluconeogenesis

“…Antisense oligonucleotides (ASOs) stabilized with a 2Ј-O-(2-methoxy)-ethyl modification and a phosphorothioate backbone have been widely used to acutely silence genes in liver, adipose, and proximal tubules but not skeletal muscle or pancreas of healthy, regular chow-fed adult male Sprague-Dawley rats as well as in humans (20). Twice weekly intraperitoneal injections were given for 4 weeks of 25 mg/kg of PEPCK-M ASO (PCK2 ASO ; ISIS421062:5Ј-GCACGCGCTCTTCCAGGGCC-3) as compared with a control ASO (Con ASO ; ISIS141923:5Ј-CCTTCCCTGAAGGTTC-CTCC-3Ј) (21,22).…”