Arsenite modulates cardiac substrate preference by translocation of GLUT4, but not CD36, independent of mitogen-activated protein kinase signaling.

The protein thiol-modifying agent arsenite, a potent activator of stress signaling, was used to examine the involvement of MAPKs in the regulation of cardiac substrate uptake. Arsenite strongly induced p38 MAPK phosphorylation in isolated rat cardiac myocytes but also moderately enhanced phosphorylation of p42/44 ERK and p70 S6K. At the level of cardiomyocytic substrate use, arsenite enhanced glucose uptake dose dependently up to 5.1-fold but failed to stimulate long-chain fatty acid uptake. At the substrate transporter level, arsenite stimulated the translocation of GLUT4 to the sarcolemma but failed to recruit CD36 or FABPpm. Because arsenite did not influence the intrinsic activity of glucose transporters, GLUT4 translocation is entirely responsible for the selective increase in glucose uptake by arsenite. Moreover, the nonadditivity of arsenite-induced glucose uptake and insulin-induced glucose uptake indicates that arsenite recruits GLUT4 from insulin-responsive intracellular stores. Inhibitor studies with SB203580/SB202190, PD98059, and rapamycin indicate that activation of p38 MAPK, p42/44 ERK, and p70 S6K, respectively, are not involved in arsenite-induced glucose uptake. In addition, all these kinases do not play a role in regulation of cardiac glucose and long-chain fatty acid uptake by insulin. Hence, arsenite's selective stimulation of glucose uptake appears unrelated to its signaling actions, suggesting that arsenite acts via thiol modification of a putative intracellular protein target of arsenite within insulin-responsive GLUT4-containing stores. Because of arsenite's selective stimulation of cardiac glucose uptake, identification of this putative target of arsenite within the GLUT4-storage compartment may indicate whether it is a target for future strategies in prevention of diabetic cardiomyopathy.     

Skeletal muscle fatty acid transporter protein expression in type 2 diabetes patients compared with overweight, sedentary men and age-matched, endurance-trained cyclists

Aim: Membrane fatty acid transporters can modulate the balance between fatty acid uptake and subsequent storage and/or oxidation in muscle tissue. As such, skeletal muscle fatty acid transporter protein expression could play an important role in the etiology of insulin resistance and/or type 2 diabetes. Methods: In the present study, fatty acid translocase (FAT/CD36), plasma membrane‐bound fatty acid‐binding protein (FABPpm) and fatty acid transport protein 1 (FATP1) mRNA and protein expression were assessed in muscle tissue obtained from 10 sedentary, overweight type 2 diabetes patients (60 ± 2 years), 10 sedentary, weight‐matched normoglycemic controls (60 ± 2 years) and 10 age‐matched, endurance trained cyclists (57 ± 1 years). Results: Both FAT/CD36 and FATP1 mRNA and protein expression did not differ between groups. In contrast, FABPpm mRNA and protein expression were approx. 30–40% higher in the trained men compared with the diabetes patients (P < 0.01) and sedentary controls (P < 0.05). Conclusions: Skeletal muscle FAT/CD36, FABPpm and FATP1 mRNA and protein expression are not up‐ or downregulated in a sedentary and/or insulin resistant state. In contrast, FABPpm expression is upregulated in the endurance trained state and likely instrumental to allow greater fatty acid oxidation rates.     

Fluorophore-conjugated anti-CEA Antibody for the Intraoperative Imaging of Pancreatic and Colorectal Cancer

Introduction  Colorectal and pancreatic cancers together comprise the third and fourth most common causes of cancer-related death in the United States. In both of these cancers, complete detection of primary and metastatic lesions at the time of surgery is critical to optimal surgical resection and appropriate patient treatment. Materials and Methods  We have investigated the use of fluorophore-labeled anti-carcinoembryonic antigen (CEA) monoclonal antibody to aid in cancer visualization in nude mouse models of human colorectal and pancreatic cancer. Anti-CEA was conjugated with a green fluorophore. Subcutaneous, orthotopic primary and metastatic human pancreatic and colorectal tumors were easily visualized with fluorescence imaging after administration of conjugated anti-CEA. The fluorescence signal was detectable 30 min after systemic antibody delivery and remained present for 2 weeks, with minimal in vivo photobleaching after exposure to standard operating room lighting. Tumor resection techniques revealed improved ability to resect labeled tumor tissue under fluorescence guidance. Comparison of two different fluorophores revealed differences in dose–response and photobleaching in vivo. Conclusion  These results indicate that fluorophore-labeled anti-CEA offers a novel intraoperative imaging technique for enhanced visualization of tumors in colorectal and pancreatic cancer when CEA expression is present, and that the choice of fluorophore significantly affects the signal intensity in the labeled tumor. These data were presented at the Society for Surgery of the Alimentary Tract meeting as part of the Digestive Diseases Week, San Diego CA, May 21 2008. Sharmeela Kaushal and Michele K. McElroy shared authorship. Work supported in part by: Cancer Therapeutics Training Program (T32 CA121938) National Institutes of Health (CA109949-03) American Cancer Society (RSG-05-037-01-CCE).     

Cytoplasmic fatty acid-binding protein facilitates fatty acid utilization by skeletal muscle

The intracellular transport of long-chain fatty acids in muscle cells is facilitated to a great extent by heart-type cytoplasmic fatty acid-binding protein (H-FABP). By virtue of the marked affinity of this 14.5-kDa protein for fatty acids, H-FABP dramatically increases their concentration in the aqueous cytoplasm by non-covalent binding, thereby facilitating both the transition of fatty acids from membranes to the aqueous space and their diffusional transport from membranes (e.g. sarcolemma) to other cellular compartments (e.g. mitochondria). Striking features are the relative abundance of H-FABP in muscle, especially in oxidative muscle fibres, and the modulation of the muscular H-FABP content in concert with the modulation of other proteins and enzymes involved in fatty acid handling and utilization. Newer studies with mice carrying a homozygous or heterozygous deletion of the H-FABP gene show that, in comparison with wild-type mice, hindlimb muscles from heterozygous animals have a markedly lowered (-66%) H-FABP content but unaltered palmitate uptake rate, while in hindlimb muscles from homozygous animals (no H-FABP present) palmitate uptake was reduced by 45%. These findings indicate that H-FABP is present in relative excess and plays a substantial, but merely permissive role in fatty acid uptake by skeletal muscles.     

Contribution of FAT/CD36 to the regulation of skeletal muscle fatty acid oxidation: an overview

Long chain fatty acids (LCFAs) are an important substrate for ATP production within the skeletal muscle. The process of LCFA delivery from adipose tissue to muscle mitochondria involves many regulatory steps. Recently, it has been recognized that LCFA oxidation is not only dependent on LCFA delivery to the muscle, but also on regulatory steps within the muscle. Increasing selected fatty acid binding proteins/transporters on the plasma membrane facilitates a very rapid LCFA increase into the muscle, independent of any changes in LCFA delivery to the muscle. Such a mechanism of LCFA transporter translocation is activated by muscle contraction. Intramuscular triacylglycerols may also be hydrolysed to provide fatty acids for mitochondrial oxidation, particularly during exercise, when hormone-sensitive lipase and other enzymes are activated. Mitochondrial LCFA entry is also highly regulated. This however does not involve only the malonyl CoA carnitine palmitoyltransferase-I (CPTI) axis. Exercise-induced fatty acid entry into mitochondria is also regulated by at least one of the proteins (FAT/CD36) that also regulates plasma membrane fatty acid transport. Among individuals, differences in mitochondrial fatty acid oxidation appear to be correlated with the content of mitochondrial CPTI and FAT/CD36. This paper provides a brief overview of mechanisms that regulate LCFA uptake and oxidation in skeletal muscle during exercise and in obesity. We focus largely on our own work on FAT/CD36, which contributes to regulating, in a coordinated fashion, LCFA uptake across the plasma membrane and the mitochondrial membrane. Very little is known about the roles of FATP1-6 on fatty acid transport in skeletal muscle.     

Staging of ampullary and pancreatic carcinoma: comparison between endosonography and surgery

Background: The aim of the study was to update our previously published data on the clinical TNM staging of ampullary and pancreatic carcinoma by endosonography. Methods: Endosonography was performed in 70 patients with pancreatic cancer and in 32 patients with ampullary carcinoma. TNM staging was carried out before surgery and compared with findings of histology and/or surgery. Results: Endosonography was accurate in staging the depth of tumor invasion. Early-stage carcinomas could be distinguished from advanced cancers. Nonresectability was accurately assessed on the basis of vascular involvement using real-time ultrasound. Tumor compression due to peritumoral pancreatitis and direct tumor invasion into the base of the mesocolon could not be diagnosed by endosonography. The overall accuracy in tumor staging for pancreatic and ampullary carcinomas was 83.6% and 84.4%, respectively. Endosonography was accurate in diagnosing regional lymph node metastases but not accurate in defining nonmetastatic lymphadenopathy and distant metastases. Conclusion: Endosonography was accurate in staging tumor stage and lymph node metastases. Minimally invasive methods of resection for superficial ampullary cancers should be based on endosonography staging. (Gastrointest Endosc 1996;44:706-13.)     

Signalling components involved in contraction-inducible substrate uptake into cardiac myocytes

Glucose and long-chain fatty acids (LCFA) are two major substrates used by heart and skeletal muscle to support contractile activity. In quiescent cardiac myocytes a substantial portion of the glucose transporter GLUT4 and the putative LCFA transporter fatty acid translocase (FAT)/CD36 are stored in intracellular compartments. Induction of cellular contraction by electrical stimulation results in enhanced uptake of both glucose and LCFA through translocation of GLUT4 and FAT/CD36 respectively to the sarcolemma. The involvement of protein kinase A, AMP-activated protein kinase (AMPK), protein kinase C (PKC) isoforms and the extracellular signal-regulated kinases was evaluated in cardiac myocytes as candidate signalling enzymes involved in recruiting these transporters in response to contraction. The collected evidence excluded the involvement of PKA and implicated an important role for AMPK and for one (or more) PKC isoform(s) in contraction-induced translocation of both GLUT4 and FAT/CD36. The unravelling of further components along this contraction pathway can provide valuable information on the coordinated regulation of the uptake of glucose and of LCFA by an increase in mechanical activity of heart and skeletal muscle.