Identification of Molecular targets to Improve Glucose Homeostasis
The incidence of Type 2 (non-insulin-dependent) diabetes mellitus is growing at an astronomical rate, as millions of people are diagnosed with this profound metabolic disorder every year. Skeletal muscle is quantitatively the most important tissue involved in maintaining glucose homeostasis under insulin-stimulated conditions, and is a major site of insulin resistance in Type 2 diabetic patients.
Glucose transport is the rate-limiting step for whole body glucose uptake and can be activated in skeletal muscle by two separate and distinct signaling pathways, one stimulated by insulin and the second by muscle contractions. The molecular signaling mechanisms by which insulin and exercise (muscle contraction) increase glucose transport and gene expression in skeletal muscle are largely undefined.
Insulin signaling and Type 2 Diabetes Mellitus:
Skeletal muscle is quantitatively the most important tissue involved in maintaining glucose homeostasis. Glucose transport and metabolism, protein synthesis and gene expression are all regulated by activation of the insulin signaling pathway (Fig. 1).
This signal transduction pathway is engaged upon insulin binding to the insulin receptor b-subunit, followed by autophosphorylation of the b-subunit, leading to activation of down-stream signaling events. Insulin-receptor substrates (IRS) are regulatory docking proteins that associate with the insulin receptor and play a central role in the selection and differentiation of the insulin signal toward further metabolic or mitogenic events.
The downstream substrate of IRS-1, phosphatidylinositol 3-kinase (PI 3-kinase) is essential for insulin-stimulated glucose transport. We have shown in human skeletal muscle, insulin-stimulated glucose transport, mediated primarily by the glucose transporter GLUT4, is inhibited following exposure to PI 3-kinase inhibitors (Diabetologia 40:1172-1177, 1997). Furthermore, we have provided the first evidence that early defects in the insulin signaling-transduction cascade i.e., defective phosphorylation of the IRS-1 and reduced activity of PI 3-kinase (Diabetes 49:284-292, 2000) are coupled to impaired insulin-stimulated GLUT4 translocation and glucose transport (Diabetes 49:647-654, 2000) in skeletal muscle from diabetic patients.
Importantly, we now have evidence that these same steps in the insulin signaling cascade are impaired in skeletal muscle from glucose intolerant (IGT) relatives of Type 2 diabetic patients (Diabetes 50:2770-2778, 2001), providing the first molecular evidence that there are genetic lesions directly in the insulin signaling cascade that precede overt Type 2 diabetes.
AMPK: A novel molecular target to bypass defective insulin signal transduction in skeletal muscle
Type 2 diabetes mellitus is one of the major causes of death and disability, due to the complications accompanying this disease (Diabetologia 43:821-835, 2000). Thus, development of effective intervention strategies is essential in order to prevent and manage Type 2 diabetes. Physical exercise profoundly enhances substrate utilization and insulin sensitivity, which in turn lowers blood glucose and lipid levels. We have shown that muscle contraction increases protein expression and function of important components in the insulin signaling pathway (Proc Natl Acad Sci USA 97:38-43, 2000), pharmacological intervention with compounds designed to mimic the "exercise-response" may be efficacious in the management of metabolic abnormalities associated with Type 2 diabetes.
AMPK is considered a "master switch" in the regulation of key proteins in metabolic pathways known to control hepatic fatty acid oxidation and ketogenesis, lipogenesis and triglyceride synthesis, adipocyte lipolysis, modulation of insulin secretion from the pancreatic b-cells, and skeletal muscle fatty acid oxidation (Am J Physiol 277:E1-E10, 1999). We have identified AMP-activated protein kinase (AMPK) as an important mediator of muscle contraction-induced glucose transport and a target for pharmacological intervention to treat altered glucose homeostasis associated with Type 2 diabetes and obesity (Diabetologia 44:2180-2186, 2001; Diabetologia 45:56-65, 2002).
AMPK is a heterotrimeric protein, composed of one catalytic (a) and two non-catalytic (b and g) subunits and is activated by cellular stress associated with ATP depletion (Am J Physiol 277:E1-E10, 1999). One such compound is 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR).
AICAR is an adenosine analog that can be taken up into intact hepatocytes, adipocytes and skeletal muscle and is phosphorylated to form 5-aminoimidazole-4-carboxamide ribonucleotide (ZMP), the monophosphorylated derivative that mimics the effects of AMP on AMPK without affecting ATP or ADP levels (FASEB J 9:541-546, 1995). We have shown AICAR-treatment leads to a striking normalization of blood glucose levels and glucose tolerance in KKAy-CETP and ob/ob mice after in vivo treatment.
These changes in glucose homeostasis are accompanied by increased protein expression of GLUT4 and hexokinase II due to increased MEF2 DNA binding activity (Diabetologia 45:56-65, 2002). Since AMPK appears to be a molecular component of the exercise signaling pathway to glucose transport, we hypothesize that activation of AMPK may be a molecular target involved in the regulation of glucose homeostasis via insulin-independent mechanisms.
Our central hypothesis is that alterations in signal transduction to glucose transport should have a profound effect on whole body glucose homeostasis. The overall aim of our work is to validate novel pathways in the regulation of glucose transport and metabolism:
- To identify differentially expressed genes in skeletal muscle from Type 2 diabetic patients.
- To identify the intracellular signaling mechanisms linking the biochemical and mechanical events of muscle contraction with increased glucose metabolism and gene expression.
- To examine the kinetics of trafficking mechanisms that lead to these changes in cell surface GLUT4 in diabetic versus control muscle.
Diabetes is a life-long condition. If left untreated, diabetes can lead to severe medical complications, which include heart disease, stroke, kidney disease, blindness, nerve damage, and severe infections leading to gangrene and foot and leg amputations. Maintaining glucose homeostasis may help prevent these complications.
By identifying the molecular mechanisms controlling insulin sensitivity it will be possible to develop pharmacological and physiological (exercise and diet) intervention strategies aimed to improve glucose homeostasis. However, diabetes is a heterogeneous metabolic disorder, consequently, a multitude of intervention strategies need to be applied to achieve good glucose control.
The long-term goal is to identify and characterize new molecular targets that control glucose uptake and metabolism in skeletal muscle. Intervention at the level of these targets may control and prevent insulin resistance in Type 2 diabetes.