Physiology and pathophysiology of aquaporins

Aquaporins (AQPs) are water channels that facilitate a rapid transport of water, across cell membranes. In some cases, these pores are also permeated by small solutes, particularly glycerol. Thirteen aquaporins (AQP0-12) have been identified so far in mammalian tissues. The disruption of the genes encoding aquaporins in transgenic mice has revealed their implication in physiological and pathophysiological processes, including renal water absorption, neural function, digestion, tumor angiogenesis, and reproduction. A subset of aquaporins that transport both water and glycerol, the 'aquaglyceroporins', regulate glycerol content in epidermal, fat and other tissues, and are involved in skin hydration, fat metabolism and gluconeogenesis. Better understanding of the exact mechanisms and regulation of aquaporins might be useful for designing potential drug targets against different metabolic disorders, such as stroke, glaucoma, brain edema, cancer, diabetes and obesity.Adipobiology 2010; 2: 9-22

Leptin Reduces the Expression and Increases the Phosphorylation of the Negative Regulators of GLUT4 Traffic TBC1D1 and TBC1D4 in Muscle of ob/ob Mice

Leptin improves insulin sensitivity in skeletal muscle. Our goal was to determine whether proteins controlling GLUT4 traffic are altered by leptin deficiency and in vivo leptin administration in skeletal muscle of wild type and ob/ob mice. Leptin-deficient ob/ob mice were divided in three groups: control, leptin-treated (1 mg/kg/d) and leptin pair-fed ob/ob mice. Microarray analysis revealed that 1,546 and 1,127 genes were regulated by leptin deficiency and leptin treatment, respectively. Among these, we identified 24 genes involved in intracellular vesicle-mediated transport in ob/ob mice. TBC1 domain family, member 1 (Tbc1d1), a negative regulator of GLUT4 translocation, was up-regulated (P = 0.001) in ob/ob mice as compared to wild types. Importantly, leptin treatment reduced the transcript levels of Tbc1d1 (P<0.001) and Tbc1d4 (P = 0.004) in the leptin-treated ob/ob as compared to pair-fed ob/ob animals. In addition, phosphorylation levels of TBC1D1 and TBC1D4 were enhanced in leptin-treated ob/ob as compared to control ob/ob (P = 0.015 and P = 0.023, respectively) and pair-fed ob/ob (P = 0.036 and P = 0.034, respectively) mice. Despite similar GLUT4 protein expression in wild type and ob/ob groups a different immunolocalization of this protein was evidenced in muscle sections. Leptin treatment increased GLUT4 immunoreactivity in gastrocnemius and extensor digitorum longus sections of leptin-treated ob/ob mice. Moreover, GLUT4 protein detected in immunoprecipitates from TBC1D4 was reduced by leptin replacement compared to control ob/ob (P = 0.013) and pair-fed ob/ob (P = 0.037) mice. Our findings suggest that leptin enhances the intracellular GLUT4 transport in skeletal muscle of ob/ob animals by reducing the expression and activity of the negative regulators of GLUT4 traffic TBC1D1 and TBC1D4

Estudio de la influencia de la osteopontina en el desarrollo de la obesidad y sus comorbilidades

In a first study we analysed the effect of the lack of osteopontin (OPN) on the development of obesity and hepatic steatosis induced by a high-fat diet (HFD) using OPN-KO mice. OPN expression was upregulated in epididymal white adipose tissue (EWAT) and liver in wild type (WT) mice under a HFD. OPN-KO mice had higher insulin sensitivity, lower body weight and fat mass with reduced adipose tissue extracellular matrix remodelling and reduced adipocyte size than WT mice under a HFD. Moreover, our data show for the first time that OPN-KO under a HFD mice display reduced fibrosis in adipose tissue and liver, as well as reduced oxidative stress in adipose tissue. OPN deficiency prevented hepatic steatosis. Furthermore, OPN-KO mice exhibited higher body temperature and improved brown adipose tissue (BAT) function. In a second study we analyzed the effect of sleeve gastrectomy (SG) on circulating levels of OPN and its expression in adipose tissue and liver in HFD-induced obese rats. Circulating OPN levels decreased with HFD feeding remaining unaltered after SG. The expression of Spp1 in EWAT and liver was not modified by SG. The global improvement of metabolism after SG appears not to involve changes in serum OPN concentrations as well as in EWAT and liver expression in rats. In a third study we compared the effect of Roux-en-Y gastric bypass (RYGB) and SG on plasma levels of OPN in humans. RYGB increased circulating OPN levels, while they remained unaltered after SG. The increase in OPN levels after RYGB could be related to the increased bone resorption in relation to its well-known effects on bone of this malabsorptive procedure in comparison to the merely restrictive SG. Therefore, our results suggest that OPN could be an attractive target for the treatment of obesity and associated pathologies

Estudio de la influencia de la osteopontina en el desarrollo de la obesidad y sus comorbilidades

In a first study we analysed the effect of the lack of osteopontin (OPN) on the development of obesity and hepatic steatosis induced by a high-fat diet (HFD) using OPN-KO mice. OPN expression was upregulated in epididymal white adipose tissue (EWAT) and liver in wild type (WT) mice under a HFD. OPN-KO mice had higher insulin sensitivity, lower body weight and fat mass with reduced adipose tissue extracellular matrix remodelling and reduced adipocyte size than WT mice under a HFD. Moreover, our data show for the first time that OPN-KO under a HFD mice display reduced fibrosis in adipose tissue and liver, as well as reduced oxidative stress in adipose tissue. OPN deficiency prevented hepatic steatosis. Furthermore, OPN-KO mice exhibited higher body temperature and improved brown adipose tissue (BAT) function. In a second study we analyzed the effect of sleeve gastrectomy (SG) on circulating levels of OPN and its expression in adipose tissue and liver in HFD-induced obese rats. Circulating OPN levels decreased with HFD feeding remaining unaltered after SG. The expression of Spp1 in EWAT and liver was not modified by SG. The global improvement of metabolism after SG appears not to involve changes in serum OPN concentrations as well as in EWAT and liver expression in rats. In a third study we compared the effect of Roux-en-Y gastric bypass (RYGB) and SG on plasma levels of OPN in humans. RYGB increased circulating OPN levels, while they remained unaltered after SG. The increase in OPN levels after RYGB could be related to the increased bone resorption in relation to its well-known effects on bone of this malabsorptive procedure in comparison to the merely restrictive SG. Therefore, our results suggest that OPN could be an attractive target for the treatment of obesity and associated pathologies

Physiology and pathophysiology of aquaporins

Aquaporins (AQPs) are water channels that facilitate a rapid transport of water, across cell membranes. In some cases, these pores are also permeated by small solutes, particularly glycerol. Thirteen aquaporins (AQP0-12) have been identified so far in mammalian tissues. The disruption of the genes encoding aquaporins in transgenic mice has revealed their implication in physiological and pathophysiological processes, including renal water absorption, neural function, digestion, tumour angiogenesis, and reproduction. A subset of aquaporins that transport both water and glycerol, the ‘aquaglyceroporins’, regulate glycerol content in epidermal, fat and other tissues, and are involved in skin hydration, fat metabolism and gluconeogenesis. Better understanding of the exact mechanisms and regulation of aquaporins might be useful for designing potential drug targets against different metabolic disorders, such as stroke, glaucoma, brain ooedema, cancer, diabetes and obesity

Osteopontin deletion prevents the development of obesity and hepatic steatosis via impaired adipose tissue matrix remodeling and reduced inflammation and fibrosis in adipose tissue and liver in mice.

Osteopontin (OPN) is a multifunctional extracellular matrix (ECM) protein involved in multiple physiological processes. OPN expression is dramatically increased in visceral adipose tissue in obesity and the lack of OPN protects against the development of insulin resistance and inflammation in mice. We sought to unravel the potential mechanisms involved in the beneficial effects of the absence of OPN. We analyzed the effect of the lack of OPN in the development of obesity and hepatic steatosis induced by a high-fat diet (HFD) using OPN-KO mice. OPN expression was upregulated in epididymal white adipose tissue (EWAT) and liver in wild type (WT) mice with HFD. OPN-KO mice had higher insulin sensitivity, lower body weight and fat mass with reduced adipose tissue ECM remodeling and reduced adipocyte size than WT mice under a HFD. Reduced MMP2 and MMP9 activity was involved in the decreased ECM remodeling. Crown-like structure number in EWAT as well as F4/80-positive cells and Emr1 expression in EWAT and liver increased with HFD, while OPN-deficiency blunted the increase. Moreover, our data show for the first time that OPN-KO under a HFD mice display reduced fibrosis in adipose tissue and liver, as well as reduced oxidative stress in adipose tissue. Gene expression of collagens Col1a1, Col6a1 and Col6a3 in EWAT and liver, as well as the profibrotic cytokine Tgfb1 in EWAT were increased with HFD, while OPN-deficiency prevented this increase. OPN deficiency prevented hepatic steatosis via reduction in the expression of molecules involved in the onset of fat accumulation such as Pparg, Srebf1, Fasn, Mogat1, Dgat2 and Cidec. Furthermore, OPN-KO mice exhibited higher body temperature and improved BAT function. The present data reveal novel mechanisms of OPN in the development of obesity, pointing out the inhibition of OPN as a promising target for the treatment of obesity and fatty liver

Leptin reduces the expression and increases the phosphorylation of the negative regulators of GLUT4 traffic TBC1D1 and TBC1D4 in muscle of ob/ob mice

Leptin improves insulin sensitivity in skeletal muscle. Our goal was to determine whether proteins controlling GLUT4 traffic are altered by leptin deficiency and in vivo leptin administration in skeletal muscle of wild type and ob/ob mice. Leptin-deficient ob/ob mice were divided in three groups: control, leptin-treated (1 mg/kg/d) and leptin pair-fed ob/ob mice. Microarray analysis revealed that 1,546 and 1,127 genes were regulated by leptin deficiency and leptin treatment, respectively. Among these, we identified 24 genes involved in intracellular vesicle-mediated transport in ob/ob mice. TBC1 domain family, member 1 (Tbc1d1), a negative regulator of GLUT4 translocation, was up-regulated (P = 0.001) in ob/ob mice as compared to wild types. Importantly, leptin treatment reduced the transcript levels of Tbc1d1 (P<0.001) and Tbc1d4 (P = 0.004) in the leptin-treated ob/ob as compared to pair-fed ob/ob animals. In addition, phosphorylation levels of TBC1D1 and TBC1D4 were enhanced in leptin-treated ob/ob as compared to control ob/ob (P = 0.015 and P = 0.023, respectively) and pair-fed ob/ob (P = 0.036 and P = 0.034, respectively) mice. Despite similar GLUT4 protein expression in wild type and ob/ob groups a different immunolocalization of this protein was evidenced in muscle sections. Leptin treatment increased GLUT4 immunoreactivity in gastrocnemius and extensor digitorum longus sections of leptin-treated ob/ob mice. Moreover, GLUT4 protein detected in immunoprecipitates from TBC1D4 was reduced by leptin replacement compared to control ob/ob (P = 0.013) and pair-fed ob/ob (P = 0.037) mice. Our findings suggest that leptin enhances the intracellular GLUT4 transport in skeletal muscle of ob/ob animals by reducing the expression and activity of the negative regulators of GLUT4 traffic TBC1D1 and TBC1D4

Effect of leptin on the intracellular location of GLUT4 protein in skeletal muscle.

<p>Representative images of GLUT4 immunostaining in sections of the gastrocnemius, extensor digitorum longus (EDL) and soleus of wild type (WT), control <i>ob/ob</i> (OB), pair-fed <i>ob/ob</i> (PF) and leptin-treated <i>ob/ob</i> (LE) mice. Larger-magnification images within black frames of immunohistochemical staining for GLUT4 in gastrocnemius, EDL and soleus sections of leptin-treated <i>ob/ob</i> mice are shown. Arrows indicate punctuate distribution of GLUT4 vesicles in plasma membrane and cytosol of muscle sections. Scale bars = 50 µm.</p

Genes involved in vesicle-mediated transport altered by leptin in the gastrocnemius muscle of <i>ob/ob</i> mice.

<p>Differential expression of genes is indicated as fold changes with respect to the wild type group presenting only the genes which were significantly different (<i>P</i><0.05) between the leptin-treated and the control <i>ob/ob</i> groups. Ratio: fold change value for leptin-treated <i>vs</i> the <i>ob/ob</i> group.</p

Effect of leptin on the regulation of the negative regulators of GLUT4 translocation, TBC1D1 and TBC1D4.

<p>A: Representative phospho-Akt substrate (PAS) immunoblot of gastrocnemius lysate of wild type (WT), control <i>ob/ob</i> (OB), pair-fed <i>ob/ob</i> (PF) and leptin-treated <i>ob/ob</i> (LE) mice. The arrow indicates the band that may correspond to TBC1D1/4. B: Representative blots of immunoprecipitation and immunoblotting experiments. Equal amounts of protein (250 µg) from lysates of the gastrocnemius of WT, OB, PF and LE mice were incubated with TBC1D1 or TBC1D4 antibodies. Immunoprecipitated (IP) samples of TBC1D1 and TBC1D4 were resolved by SDS-PAGE and membranes were immunoblotted (WB) using PAS and GLUT4 antibodies. Membranes of immunoprecipitates probed with PAS and GLUT4 antibodies were stripped and re-probed with the corresponding total antibody TBC1D1 and TBC1D4. The signals from PAS-TBC1D1 and GLUT4-TBC1D1 and PAS-TBC1D4 and GLUT4-TBC1D4 were related to total TBC1D1 and TBC1D4, respectively (n = 6 per group). Leptin increases the phosphorylation levels of TBC1D1 (C) and TBC1D4 (D) in the gastrocnemius of leptin-treated <i>ob/ob</i> mice. GLUT4 co-precipitates with intracellular TBC1D1 (E) and TBC1D4 (F) proteins. Values are presented as means±SEM. * <i>P</i><0.05 by one-way ANOVA followed by LSD tests.</p