Abstract
The Rab GTPase activating protein known as Akt substrate of 160 kDa (AS160 or TBC1D4) regulates insulin-stimulated glucose uptake in skeletal muscle, the heart, and white adipose tissue (WAT). A novel rat AS160-knockout (AS160-KO) was created with CRISPR/Cas9 technology. Because female AS160-KO versus wild type (WT) rats had not been previously evaluated, the primary objective of this study was to compare female AS160-KO rats with WT controls for multiple, important metabolism-related endpoints. Body mass and composition, physical activity, and energy expenditure were not different between genotypes. AS160-KO versus WT rats were glucose intolerant based on an oral glucose tolerance test (P<0.001) and insulin resistant based on a hyperinsulinemic-euglycemic clamp (HEC; P<0.001). Tissue glucose uptake during the HEC of female AS160-KO versus WT rats was: 1) significantly lower in epitrochlearis (P<0.05) and extensor digitorum longus (EDL; P<0.01) muscles of AS160-KO compared to WT rats; 2) not different in soleus, gastrocnemius or WAT; and 3) ~3-fold greater in the heart (P<0.05). GLUT4 protein content was reduced in AS160-KO versus WT rats in the epitrochlearis (P<0.05), EDL (P<0.05), gastrocnemius (P<0.05), soleus (P<0.05), WAT (P<0.05), and the heart (P<0.005). Insulin-stimulated glucose uptake by isolated epitrochlearis and soleus muscles was lower (P<0.001) in AS160-KO versus WT rats. Akt phosphorylation of insulin-stimulated tissues was not different between the genotypes. A secondary objective was to probe processes that might account for the genotype-related increase in myocardial glucose uptake, including glucose transporter protein abundance (GLUT1, GLUT4, GLUT8, SGLT1), hexokinase II protein abundance, and stimulation of the AMP-activated protein kinase (AMPK) pathway. None of these parameters differed between genotypes. Metabolic phenotyping in the current study revealed AS160 deficiency produced a profound glucoregulatory phenotype in female AS160-KO rats that was strikingly similar to the results previously reported in male AS160-KO rats.
Highlights
The Rab GTPase activating protein known as Akt substrate of 160 kDa is highly expressed by multiple tissues, including skeletal muscle, the heart, and white adipose tissue (WAT) [1,2,3,4,5]
The female AS160-KO versus wild type (WT) rats were glucose intolerant based on the Oral glucose tolerance test (OGTT) and insulin resistant based on GIR and glucose turnover rate (GTR) during the Hyperinsulinemic-euglycemic clamps (HEC)
GLUT4 glucose transporter protein abundance was substantially lower for female AS160-KO compared to WT rats in each of the skeletal muscles studied
Summary
The Rab GTPase activating protein known as Akt substrate of 160 kDa ( known as AS160 or TBC1D4) is highly expressed by multiple tissues, including skeletal muscle, the heart, and white adipose tissue (WAT) [1,2,3,4,5] These tissues are important sites for insulin-mediated glucose disposal, and phosphosite-specific phosphorylation of AS160 by Akt is crucial for insulinstimulated GLUT4 glucose transporter exocytosis and enhanced glucose transport. Other studies reported data for both male and female AS160-KO mice for some, but not all outcomes [3, 9]. Hyperinsulinemic-euglycemic clamps (HEC) have been performed only in male AS160-KO mice [4], and in vivo tissue-specific glucose uptake has been reported in male, but not female, mice [3, 9]. The only previous study of AS160-KO rats focused exclusively on male animals [5]
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