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Abstract: Background and aims: Glycogen storage disease type Ib (GSD1b) is a rare metabolic and immune disorder caused by a deficiency in the glucose‐6‐phosphate transporter (G6PT) and characterized by impaired glucose homeostasis, myeloid dysfunction, and long‐term risk of hepatocellular adenomas. Despite maximal therapy, based on a strict diet and on granulocyte colony‐stimulating factor treatment, long‐term severe complications still develop. Understanding the pathophysiology of GSD1b is a prerequisite to develop new therapeutic strategies and depends on the availability of animal models. The G6PT‐KO mouse mimics the human disease but is very fragile and rarely survives weaning. We generated a conditional G6PT‐deficient mouse as an alternative model for studying the long‐term pathophysiology of the disease. We utilized this conditional mouse to develop an inducible G6PT‐KO model to allow temporally regulated G6PT deletion by the administration of tamoxifen (TM). Methods: We generated a conditional G6PT‐deficient mouse utilizing the CRElox strategy. Histology, histochemistry, and phenotype analyses were performed at different times after TM‐induced G6PT inactivation. Neutrophils and monocytes were isolated and analyzed for functional activity with standard techniques. Results: The G6PT‐inducible KO mice display the expected disturbances of G6P metabolism and myeloid dysfunctions of the human disorder, even though with a milder intensity. Conclusions: TM‐induced inactivation of G6PT in these mice leads to a phenotype which mimics that of human GSD1b patients. The conditional mice we have generated represent an excellent tool to study the tissue‐specific role of the G6PT gene and the mechanism of long‐term complications in GSD1b. [ABSTRACT FROM AUTHOR]

Here, we report the identification of three novel missense mutations in the calsequestrin-1 ( CASQ1) gene in four patients with tubular aggregate myopathy. These CASQ1 mutations affect conserved amino acids in position 44 (p.(Asp44Asn)), 103 (p.(Gly103Asp)), and 385 (p.(Ile385Thr)). Functional studies, based on turbidity and dynamic light scattering measurements at increasing Ca2+ concentrations, showed a reduced Ca2+-dependent aggregation for the CASQ1 protein containing p.Asp44Asn and p.Gly103Asp mutations and a slight increase in Ca2+-dependent aggregation for the p.Ile385Thr. Accordingly, limited trypsin proteolysis assay showed that p.Asp44Asn and p.Gly103Asp were more susceptible to trypsin cleavage in the presence of Ca2+ in comparison with WT and p.Ile385Thr. Analysis of single muscle fibers of a patient carrying the p.Gly103Asp mutation showed a significant reduction in response to caffeine stimulation, compared with normal control fibers. Expression of CASQ1 mutations in eukaryotic cells revealed a reduced ability of all these CASQ1 mutants to store Ca2+ and a reduced inhibitory effect of p.Ile385Thr and p.Asp44Asn on store operated Ca2+ entry. These results widen the spectrum of skeletal muscle diseases associated with CASQ1 and indicate that these mutations affect properties critical for correct Ca2+ handling in skeletal muscle fibers. [ABSTRACT FROM AUTHOR]

Loss-of-function mutations in the gene encoding GLUT10 are responsible for arterial tortuosity syndrome (ATS), a rare connective tissue disorder. In this study GLUT10-mediated dehydroascorbic acid (DAA) transport was investigated, supposing its involvement in the pathomechanism. GLUT10 protein produced by in vitro translation and incorporated into liposomes efficiently transported DAA. Silencing of GLUT10 decreased DAA transport in immortalized human fibroblasts whose plasma membrane was selectively permeabilized. Similarly, the transport of DAA through endomembranes was markedly reduced in fibroblasts from ATS patients. Re-expression of GLUT10 in patients' fibroblasts restored DAA transport activity. The present results demonstrate that GLUT10 is a DAA transporter and DAA transport is diminished in the endomembranes of fibroblasts from ATS patients. [ABSTRACT FROM AUTHOR]

The P2×7 receptor ( P2×7r) is expressed in innate immune cells (e.g. monocyte/macrophages), playing a key role in IL-1β release. Since innate immune activation and IL-1β release seem to be implicated in Behçet's disease ( BD), a systemic immune-inflammatory disorder of unknown origin, we hypothesized that P2×7r is involved in the pathogenesis of the disease. Monocytes were isolated from 18 BD patients and 17 healthy matched controls. In BD monocytes, an increased P2×7r expression and Ca2+ permeability induced by the selective P2×7r agonist 2′-3′-O-(4-benzoylbenzoyl) ATP ( Bz ATP) was observed. Moreover, IL-1β release from LPS-primed monocytes stimulated with Bz ATP was markedly higher in BD patients than in controls. TNF-α-incubated monocytes from healthy subjects almost reproduced the findings observed in BD patients, as demonstrated by the increase in P2×7r expression and Bz ATP-induced Ca2+ intake. Our results provide evidence that in BD monocytes both the expression and function of the P2×7r are increased compared with healthy controls, as the possible result, at least in part, of a positive modulating effect of TNF-α on the receptor. These data indicate P2×7r as a new potential therapeutic target for the control of BD, further supporting the rationale for the use of anti- TNF-α drugs in the treatment of the disease. [ABSTRACT FROM AUTHOR]

Abstract: Effect of 5–100μM epigallocatechin gallate (EGCG) on hepatic glucose 6-phosphatase (G6Pase) system was investigated. EGCG inhibited G6Pase in intact but not in permeabilized rat liver microsomes, suggesting the interference with the transport. However, EGCG did not hinder microsomal glucose 6-phosphate (G6P) uptake. Instead, it increased the accumulation of radioactivity after the addition of [14C]G6P, presumably due to a slower release of [14C]glucose, the product of luminal hydrolysis. Indeed, EGCG was found to inhibit microsomal glucose efflux. Since G6Pase activity is depressed by glucose in a concentration-dependent manner, we concluded that EGCG inhibits G6Pase through an elevated luminal glucose level. [Copyright &y& Elsevier]