Liver Glucokinase and Lipid Metabolism

Control of energy metabolism is crucial for optimal functioning of organs and tissues. Amongst all nutrients, glucose is the principal energy source for most cells and, therefore, minimum blood glucose levels must be guaranteed. Alterations in glycaemia can lead to hyperglycaemic states (producing protein glycosylation and toxicity in glucose-sensitive cells) or hypoglycaemic states (that can affect brain function), both harmful. Therefore, mechanisms must exist to keep glycaemia in a narrow physiological range (4-8 mM) independently of the nutritional state. To achieve control of blood glucose levels, our body has a complex, interorgan signaling system using nutrients (glucose, lipids, amino acids), hormones (insulin, glucagon, ghrelin, etc.) and the autonomic nervous system. In response to these signals, organs and tissues (mainly intestine, endocrine pancreas, liver, skeletal muscle, adipose tissue, brain and adrenal glands) adapt their function to energetic requirements. The liver plays a pivotal role in the maintenance of glucose homeostasis by continuously adapting its metabolism to energetic needs. In the fed state, when blood glucose levels are high and there is insulin, liver takes-up part glucose to replenish glycogen stores. Besides, when glucose stores are full, the liver has the capacity to synthesize lipids de novo from glucose for-long term energy storage. Lipids are packaged in very low-density lipoprotein (VLDL) particles and then transported to the adipose tissue. Conversely during starvation, when glycaemia falls and glucagon increases, the liver produces glucose to maintain circulating glucose levels by breaking down glycogen stores or by synthesizing glucose de novo through gluconeogenesis. Gluconeogenesis, as an energy-consuming pathway, is linked to -oxidation of fatty acids (fuel supplier pathway)..

Liver GlucokinaseA456V Induces Potent Hypoglycemia without Dyslipidemia through a Paradoxical Induction of the Catalytic Subunit of Glucose-6-Phosphatase

Recent reports point out the importance of the complex GK-GKRP in controlling glucose and lipid homeostasis. Several GK mutations affect GKRP binding, resulting in permanent activation of the enzyme. We hypothesize that hepatic overexpression of a mutated form of GK, GKA456V, described in a patient with persistent hyperinsulinemic hypoglycemia of infancy (PHHI) and could provide a model to study the consequences of GK-GKRP deregulation in vivo. GKA456V was overexpressed in the liver of streptozotocin diabetic mice. Metabolite profiling in serum and liver extracts, together with changes in key components of glucose and lipid homeostasis, were analyzed and compared to GK wild-type transfected livers. Cell compartmentalization of the mutant but not the wild-type GK was clearly affected in vivo, demonstrating impaired GKRP regulation. GKA456V overexpression markedly reduced blood glucose in the absence of dyslipidemia, in contrast to wild-type GK-overexpressing mice. Evidence in glucose utilization did not correlate with increased glycogen nor lactate levels in the liver. PEPCK mRNA was not affected, whereas the mRNA for the catalytic subunit of glucose-6-phosphatase was upregulated ~4 folds in the liver of GKA456V-treated animals, suggesting that glucose cycling was stimulated. Our results provide new insights into the complex GK regulatory network and validate liver-specific GK activation as a strategy for diabetes therapy

Specific Jak3 Downregulation in Lymphocytes Impairs γc Cytokine Signal Transduction and Alleviates Antigen-driven Inflammation In Vivo

Jak3, one of the four members comprising the Jak family of cytosolic tyrosine kinases, has emerged as a promising target for nontoxic immunotherapies. Although a number of Jak inhibitors has already demonstrated efficacy, they suffer from secondary effects apparently associated to their pan-Jak activity. However, whether selective Jak3 inhibition would afford therapeutic efficacy remains unclear. To address this question we have investigated the immunosuppressive potential of selective Jak3 intervention in lymphocytes using RNA interference (RNAi) technology in vitro and in vivo. Using synthetic small interference RNA (siRNA) sequences we achieved successful transfections into human and mouse primary T lymphocytes. We found that Jak3 knockdown was sufficient to impair not only interleukin-2 (IL-2) and T cell receptor (TCR)-mediated cell activation in vitro, but also antigen-triggereds welling, inflammatory cell infiltration, and proinflammatory cytokine raise in vivo. Furthermore, Jak1 (which mediates gamma c cytokine signaling in conjunction with Jak3) cosilencing did not provide higher potency to the aforementioned immunosuppressant effects. Our data provides direct evidences indicating that Jak3 protein plays an important role in gamma c cytokine and antigen-mediated T cell activation and modulates Th1-mediated inflammatory disorders, all in all highlighting its potential as a target in immunosuppressive therapies

Specific Jak3 Downregulation in Lymphocytes Impairs γc Cytokine Signal Transduction and Alleviates Antigen-driven Inflammation In Vivo

Jak3, one of the four members comprising the Jak family of cytosolic tyrosine kinases, has emerged as a promising target for nontoxic immunotherapies. Although a number of Jak inhibitors has already demonstrated efficacy, they suffer from secondary effects apparently associated to their pan-Jak activity. However, whether selective Jak3 inhibition would afford therapeutic efficacy remains unclear. To address this question we have investigated the immunosuppressive potential of selective Jak3 intervention in lymphocytes using RNA interference (RNAi) technology in vitro and in vivo. Using synthetic small interference RNA (siRNA) sequences we achieved successful transfections into human and mouse primary T lymphocytes. We found that Jak3 knockdown was sufficient to impair not only interleukin-2 (IL-2) and T cell receptor (TCR)-mediated cell activation in vitro, but also antigen-triggereds welling, inflammatory cell infiltration, and proinflammatory cytokine raise in vivo. Furthermore, Jak1 (which mediates gamma c cytokine signaling in conjunction with Jak3) cosilencing did not provide higher potency to the aforementioned immunosuppressant effects. Our data provides direct evidences indicating that Jak3 protein plays an important role in gamma c cytokine and antigen-mediated T cell activation and modulates Th1-mediated inflammatory disorders, all in all highlighting its potential as a target in immunosuppressive therapies

Liver Glucokinase A456V induces potent hypoglycemia without dyslipidemia through a paradoxical induction of the catalytic subunit of glucose-6-phosphatase

Recent reports point out the importance of the complex GK-GKRP in controlling glucose and lipid homeostasis. Several GK mutations affect GKRP binding, resulting in permanent activation of the enzyme. We hypothesize that hepatic overexpression of a mutated form of GK, GKA456V, described in a patient with persistent hyperinsulinemic hypoglycemia of infancy (PHHI) and could provide a model to study the consequences of GK-GKRP deregulation in vivo. GKA456V was overexpressed in the liver of streptozotocin diabetic mice. Metabolite profiling in serum and liver extracts, together with changes in key components of glucose and lipid homeostasis, were analyzed and compared to GK wild-type transfected livers. Cell compartmentalization of the mutant but not the wild-type GK was clearly affected in vivo, demonstrating impaired GKRP regulation. GKA456V overexpression markedly reduced blood glucose in the absence of dyslipidemia, in contrast to wild-type GK-overexpressing mice. Evidence in glucose utilization did not correlate with increased glycogen nor lactate levels in the liver. PEPCK mRNA was not affected, whereas the mRNA for the catalytic subunit of glucose-6-phosphatase was upregulated ~4 folds in the liver of GKA456V-treated animals, suggesting that glucose cycling was stimulated. Our results provide new insights into the complex GK regulatory network and validate liver-specific GK activation as a strategy for diabetes therapy