Although a host of metabolic inhibitors are available for use in examining metabolic pathways, their specificities are often questionable. With molecular biologic techniques, however, metabolic pathways can be changed in quite specific ways, even in the intact animal. A protein (e.g., enzyme/carrier) can be overexpressed, expressed in a tissue that normally does not contain it, or eliminated in any particular cell type. Site-directed mutations allow a molecule to be dissected and particular component groups removed or altered so that their role in the molecule's functioning can be studied. Application of these techniques to the investigation of metabolic pathways and their regulation is just beginning (2.4). Recent examples involve the study of GLUT 1 and GLUT 4, which are responsible for transporting the bulk of the glucose from the blood, and investigating the role of GLUT 2 in glucose sensing in the insulin-secreting b cells of the pancreas. GLUTs 1 and 4 were altered in skeletal muscle and adipose tissue, respectively, (24) and the gene for GLUT 4 inactivated (25). In the case of GLUT 2, mice were created that did not express this transporter in the insulin-secreting cells of the pancreas ( 26). Brief comments about these studies show the new approaches fostered by molecular biologic techniques.
Transgenic mice were established that expressed high levels of human GLUT 1, and it was properly located in the muscle sarcolemma. The increase in the transporter created a 3- to 4-fold increase in glucose transport into specific tested muscles, confirming that GLUT 1 plays a major role in controlling glucose entry into resting muscle. Strangely, insulin did not increase the entry of glucose into the transgenic mice muscles despite the fact that GLUT 4 levels in these mice were the same as those of control mice. Possibly, the GLUT 1 levels are so high in the transgenic set that glucose is not limited any longer by transporter activity. Muscle glucose was 4- to 5-fold and glycogen 10-fold higher in the transgenic mice, even though they showed an 18 (fed) to 30% (fasted) decrease in plasma glucose concentration. Oral glucose loads did not increase plasma levels as much as in normal mice and glucose was more rapidly disposed of. Thus, changing the level of GLUT 1 transporters affected not only muscle metabolism but also whole animal responses.
Because increases in the insulin-responsive transporter GLUT 4 occur in human and rodent adipocytes and are associated with obesity, the question arises as to whether increasing the GLUT 4 levels in adipocytes plays a role in obesity. Transgenic mice were made that expressed human GLUT 4 in their adipocytes. Basal-level glucose transport into their adipocytes increased approximately 20-fold compared with controls, but insulin only stimulated glucose uptake by a factor of 2.5, rather than the 15-fold increase of controls. Again, the possible explanation is that glucose transport is so high in the transgenic adipocytes that the number of transporters activated by insulin makes little difference to the overall transport. While the fat cell size was unchanged in the transgenic mice, their number more than doubled, and body lipid nearly tripled, matching the increase in cell number. The results suggest that a specific increase in GLUT 4 in adipocytes plays a role in generating obesity.
Remarkably, mice created with an absence of GLUT 4 have nearly normal glucose homeostasis (25). Clearly, while GLUT 4 is critical to the handling of glucose in insulin-sensitive tissues, it does not appear to be essential for survival. Presumably, other mechanisms can substitute for the absence of GLUT 4.
The b cells of the pancreas first sense, then respond to, elevated blood glucose levels by secreting insulin. The glucose transporter in their cell membranes is GLUT 2; circumstantial evidence suggests that it may be involved in the glucose-sensing mechanism. However, studies in transgenic mice created to express a transforming ras protein in the b cells showed that these animals respond normally to a glucose load, even though their b cells do not express GLUT 2 ( 26). It is thus difficult to consider GLUT 2 part of the sensing apparatus.
One major difficulty in using transgenic and "knock-out" mice is that the induced changes occur early on in the developing animal. Thus, the changes observed can be due to the presence or absence of the trans-gene at the time of the laboratory measurements or the genetic change could have initiated a plethora of events that caused the observed phenotype. Techniques are being developed, however, to overcome this serious problem.
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Diabetes is a disease that affects the way your body uses food. Normally, your body converts sugars, starches and other foods into a form of sugar called glucose. Your body uses glucose for fuel. The cells receive the glucose through the bloodstream. They then use insulin a hormone made by the pancreas to absorb the glucose, convert it into energy, and either use it or store it for later use. Learn more...