GLUT1 deficiency is a rare neurometabolic disorder that can be effectively treated with ketogenic diet. However, this condition is un- derdiagnosed due to its nonspecific, overlapping, and evolving symptoms with age. We retrospectively reviewed the clinical course of nine patients diagnosed with GLUT1 deficiency, based on SLC2A1 mutations and/or glucose concentration in cerebrospinal fluid. The patients included eight boys and one girl who initially presented with seizures (44%, 4/9) or delayed development (44%, 4/9) be- fore 2 years of age, except for one patient who presented with apnea as a neonate. Over the clinical course, all of the children devel- oped seizures of the mixed type, including absence seizures and generalized tonic–clonic seizures. About half (56%, 5/9) showed movement disorders such as ataxia, dystonia, or dyskinesia. We observed an evolution of phenotype over time, although this was not uniform across all patients. Only one child had microcephaly. In five patients, ketogenic diet was effective in reducing seizures and movement symptoms, and the patients exhibited subjective improvement in cognitive function. Diagnosing GLUT1 deficiency can be challenging due to the phenotypic variability and evolution. A high index of clinical suspicion in pediatric and even older patients with epilepsy or movement disorders is key to the early diagnosis and treatment, which can improve the patient’s quality of life.
Results: This study reports on a sibling pair with developmental delay, mental retardation, microcephaly, hypotonia, epilepsy, facial dysmorphism, ataxia and impaired speech. Chromosome analysis revealed a derivative chromosome 1 in both patients. FISH and MCB analysis showed two interstitial deletions at 1p34.2 and 1q44. SNP array and array-CGH analysis also determined the sizes of deletions detailed. The deleted region on 1p34.2 encompasses 33 genes, among which is GLUT1 gene (SLC2A1). However, the deleted region on 1q44 includes 59 genes and distal-proximal breakpoints were located in the ZNF672 gene and SMYD3 gene, respectively. Conclusion: Haploinsufficiency of GLUT1 leads to GLUT1 deficiency syndrome, consistent with the phenotype in patients of this study. Conversely, in the deleted region on 1q44, none of the genes are related to findings in these patients. Additionally, the results confirm previous reports on that corpus callosal development may depend on the critical gene(s) lying in 1q44 proximal to the SMYD3 gene.
On examination at age 6 years, head circumfer- ence was between the 3rd and 10th percentiles. The patient was restless, distractible, and impulsive, but cooperative and able to follow simple instructions. He had lower limb spasticity and hyperreflexia, mild dysarthria, truncal ataxia, intention tremor, poor coordination, and difficulty carrying out complex motor tasks. The diagnosis of Glut1 deficiency was confirmed by the finding of low CSF glucose concen- tration, reduced erythrocyte glucose uptake, and SLC2A1 analysis (table 1, patient 3).
with type 1 diabetes, in whom the ingestion of medium- chain triglycerides prevented the cognitive deficit in- duced by hypoglycemia by elevating blood levels of 3-hydroxybutyrate . Ketogenic diet (KD), which provides FFA as alternative fuel to carbohydrates for neuronal energy metabolism, has therefore a strong potential neuroprotective effect. The main indication of KD in children is the treatment of refractory epi- lepsy, but it is also the causal therapy of GLUT1 defi- ciency, a metabolic disorder characterized by epilepsy, developmental delay and movement disorders [14, 15]. In GLUT1 deficiency, neuroglycopenia that ensues as conse- quence of the impaired glucose transport across the blood-brain barrier [14, 15] is effectively improved by KD that provides ketone bodies as alternative energy source for the brain. In CHI, excessive insulin secretion not only induces severe neuroglycopenia, but also halts, by inhibit- ing gluconeogenesis, glycogenolysis and lipolysis, the use of other metabolic pathways that provide energetic sub- strates to the neurons. Other inherited metabolic dis- eases, such as mitochondrial fatty oxidation defects, share the same neurological risk of hypoglycemia be- cause of lack of ketones . Therefore, developing brain of patients with CHI is more vulnerable than other forms of hypoglycemia. Based on the similarities of brain metabolism perturbation shared by GLUT1 deficiency and CHI, we attempted to tackle neuroglyco- penic symptoms and outcome by administering KD in a patient with severe, drug-resistant form of CHI.
The diagnostic possibilities for refractory myo- clonic status epilepticus are metabolic enceph- alopathies (e.g., mitochondrial disease, multiple carboxylase deficiency, GLUT1 deficiency), stor- age diseases (including neuronopathic Gaucher disease, Tay-Sachs disease, Sandhoff disease, Canavan disease, and neuronal ceroid lipofuscino- sis [NCL]), CSF tetrahydrobiopterin deficiency, genetic syndromes such as Angelman syndrome and Aicardi-Goutie`res syndrome, a supratentorial
Intractable epilepsy is a common diagnosis among child neurology practitioners with medical man- agement remaining unsatisfactory in many cases. GLUT1 deficiency syndrome (GLUT1-DS) is a dis- order that should be considered in such situations. Evaluation by comparing serum to CSF glucose levels is a fast and relatively easy test, with hypoglycorrhachia being highly suggestive of GLUT1- DS. Furthermore, treatment with the ketogenic diet is well-established and can result in significant improvement in quality of life for these patients. The following case report outlines the presentation of one such patient and highlights common features that can be seen with GLUT1-DS. Of interest, she was found to have a spontaneous, novel mutation that has not been reported previously. Her case allows us to expand on the present literature and demonstrate the improvements that can be seen in a child with appropriate treatment. Neurology ® 2015;84:e111–e114
disorders. CoQ deficiency was identified both in muscle and fibroblasts (see Results). Because of this finding, CoQ supplementation (orally administered at 30 mg/Kg/day) was initiated. Ataxia improved dramatically after 6 months of therapy, and, upon reassessment after 4 years of CoQ treatment, ambulation remained essentially normal, with a mild residual reduction in velocity. Her nystagmus had also disappeared and her visual pursuit had normal- ized . However, mild dysmetria, dysarthria, myoclonic epilepsy and intellectual disability (perhaps refractory to CoQ), were present. During CoQ therapy, concomitant treatment with valproate and ethosuximide was given to control myoclonic epilepsy. No noticeable side-effects were observed when antiepileptic doses were raised to maintain therapeutic levels along the evolution of the disease due to the patient increasing weight. In order to further investigate these manifestations, a lumbar puncture was performed at 12 years of age, revealing diminished cerebrospinal glucose concentrations (Table 1). Plasma glucose concentration was normal. Diagnosis of GLUT1 deficiency (G1D) was established and a ketogenic diet (4:1 ratio, containing medium chain triglyceride oil) was initiated (CoQ treatment was then discontinued).
In childhood (as it is the case in adulthood), diagnosis approach is based on the presence or absence of progres- sive myoclonic epilepsy (PME). The causes of PME are dominated by lysosomal disorders, like in our study, (ceroide-lipofuscinosis, sialidosis, GM2 gangliosidosis, Gaucher disease type III, Niemann Pick type C), Lafora disease and mitochondriopathies [26-32]. In the absence of PME, treatable diseases (such as GLUT1 deficiency, creatine deficiency, porphyria and urea cycle disorders) should be considered [33-35]. Finally, other non treatable diseases (mainly mitochondriopathies) should be evoked. In our study, PME was the most prevalent epileptic syn- drome in childhood.
The lipid phosphatase activity of PTEN is believed to play the major anti-cancer function, since the inhibition of PIP3-dependent phosphorylation of AKT impacts on a plethora of downstream pathways that control cell proliferation, apoptosis and protein synthesis besides glucose uptake . Besides the lipid-phosphatase activity, PTEN possesses also a tyrosine and serine/ threonine phosphatase activity . Yet, the role of the protein-phosphatase activity of PTEN in cancer is largely neglected, also because very few protein substrates involved in the malignant phenotype have been identified so far. PTEN was shown to influence cell migration by dephosphorylating FAK (Focal Adhesion Kinase) , chemoresistance by dephosphorylating the non-receptor Tyr kinase SRC , and nuclear transcription by dephosphorylating CREB (cAMP responsive-element- binding protein) . More recently, it has been reported that PTEN can dephosphorylate the insulin receptor substrate-1, thus dumping the insulin and Insulin Growth Factor signals that also impinge on glucose metabolism and cell proliferation . Here we show for the first time that PTEN physically interacts with and dephosphorylates AKT. So far, the oncosuppressor function of PTEN has been attributed mainly to its lipid phosphatase activity that antagonizes the activation of the AKT pathway. Our data indicate that PTEN regulates this pathway also through its protein phosphatase activity. In fact, the G129E mutant that lacks the lipid phosphatase activity while retaining the protein phosphatase activity  could reduce the level of Trh308-phospho-AKT in the OVCAR-3 cells, which express an active PI3KC1 and an inactive Y155S PTEN mutant, and in the homogenate of FTC-133 cells, which are PTEN null and express constitutively phospho-AKT. The lowest level of phospho-AKT was achieved when the wt PTEN was ectopically expressed in OVCAR-3 cells, consistent with its dual (lipid and protein) phosphatase action in the two steps of the PI3K-AKT pathway, namely at PIP3 level and directly on the Thr308-phospho-AKT. By contrast, the C124S, lacking both the lipid and the protein phosphatase activities , and the K128_R130del PTEN mutants were unable to reduce the level of phospho- AKT. The K128_R130del PTEN mutant was first isolated from A2780 cells . This mutation involves the exon 5 in the gene, which codes for the phosphatase domain of the protein. Therefore, not surprisingly, in A2780 cells AKT is highly phosphorylated (Figure 1C) and GLUT1 can be found (at least partly) on the plasmamembrane. In spite of this fact, however, the uptake of glucose in
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Objectives: Diabetic nephropathy (DN) is one of the chronic microangiopathic complications of type 2 diabetes (T2DM) and has become the most frequent cause of end-stage renal disease. The XbaI polymorphism in the glucose transporter (GLUT1) has been suggested in the development of DN. We examined the association between XbaI polymorphism of GLUT1 and susceptibility to T2DM and development of DN. Methods: The study included 227 T2DM patients divided into 107 without DN (DM − DN) and 120 with DN (DM + DN), in addition to 100 apparently healthy controls. Genotyping was done by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP). Results: The GLUT1 XbaI T allele was associated with increased susceptibility to T2DM, when comparing the healthy controls to the whole diabetic group, odds ratio (OR) = 1.899, 95% confidence interval (CI) (1.149 - 3.136), p = 0.011. This association was also significant be- tween healthy controls and DM − DN OR = 1.997 (1.079 - 3.699), p = 0.026 as well as between healthy controls and DM + DN OR = 1.818 (1.016 - 3.253), p = 0.042. However there was no signifi- cant association of XbaI polymorphism with DN when comparing DM − DN to DM + DN OR = 0.910 (0.474 - 1.747), p = 0.777. Conclusion: XbaI T allele is associated with increased susceptibility to T2DM, but not to development of DN. Further studies are needed to replicate such findings.
Among the few downregulated proteins observed following SVIP transfection (Supplemental Table 1), we found the glucose transporter 1 (GLUT1), also known as SLC2A1, a glucose transporter that constitutes a rate-limiting step of glucose metabolism and whose overexpression in tumors is associated with increased aerobic glycolysis (16, 34). Western blot confirmed its loss in SVIP-transfected BB30- HNC (Figure 4G), BHY (Supplemental Figure 5A), and Ca-Ski (Supplemental Figure 5A) cells, but we also found decreased levels of its transcript (Figure 4G and Supplemental Figure 5A), so a compensatory mechanism associated with the emergence of mitochondrial metabolism proteins upon SVIP restoration might occur instead of being a direct effect of SVIP on its protein degradation. In this regard, we found that SVIP promoter CpG island hypermethylation was also associated with the downregulation of the GLUT1 transcript in the Sanger panel of cancer cell lines (15) (Figure 4H), being more significant for the 4 tumor types more methylated at the SVIP locus (Figure 4H). SVIP downregulation was also asso- ciated with low levels of the GLUT1 transcript in the same panel of cell lines (Supplemental Figure 5B). SVIP methylation–associated silencing is also associated with high levels of expression of the glycolytic enzyme hexokinase 1 and low levels of the uncoupling mitochondrial enzyme UCP2 (Supplemental Figure 5C). Overall, the picture that emerges indicates that SVIP reintroduction caused an enrichment of mitochondrial metabolism compared with SVIP epigenetically deficient cancer cells.
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absence of progesterone receptor, and triple- negative phenotype. GLUT1 expression was also an independent prognostic factor of poor- er overall survival and disease-free survival . Masin M’ s research provided evidence that GLUT3 is strongly up-regulated during epi- thelial-mesenchymal transition and contributes to glucose uptake in lung tumor cells . Similar results have been observed by Kocdor M. A that GLUT-3 expression is increased in Estrogen-induced breast carcinogenesis . It was reported that GLUT1 and GLUT3 expres- sion is upregulated by hypoxia-inducible factor- 1α (HIF-1α) which is induced by low oxygen con- Table 2. Comparison of clinicopathologic characteristics in
glucose (2 μCi; Perkin Elmer Biosciences). Oocytes were exposed to the uptake solution for 10 minutes. The solution was then rapidly aspirated and replaced with 1 mL of ice cold phosphate-buﬀered saline (150 mM NaCl, 10 mM sodium phosphate; pH 7.4) containing 0.1 mM phloretin (Sigma, Australia). Each group of 5 oocytes was solubilized in 250 μL of 2.5% sodium dodecyl sulphate overnight on a rocking platform. Ultima Gold scintillant (2.0 mL) (Perkin Elmer, USA) was added and counts were measured for 2 minutes in a liquid scintillation counter (TRI CARB 2900-TR, Perkin Elmer). Raw counts per minute were converted to nanomoles per minute (for a detailed account of the GLUT1 uptake assay, please see reference 14), and values were plotted against 3-O- Methyl- D -glucose concentration and data ﬁt with the Michaelis-Menten equation using GraphPad Prism (Graph Pad software, La Jolla, CA). Calibration curves were constructed using known volumes of radioactive tracer in the uptake solu- tion in the range of 3-O-Methyl- D -glucose concentrations.
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GLUT4 mRNA expression is largely restricted to brown and white adipose tissue, skeletal muscle, and heart. Glucose uptake via GLUT4 in these tissues is used for very different purposes related to the tissue’s function. GLUT4 gene expression is also a matter to up and down regulation depending on the physiologic state of the organism. Changes in GLUT4 gene expression are observed in physiologic states of altered glucose homeostasis. In general, GLUT4 mRNA expression is reduced in severe insulin deficiency such as STZ-induced diabetes and nutritional deprivations such as starvation (18,19). These changes in steady-state GLUT4 mRNA levels are tissue specific. For example, changes in GLUT4 mRNA expression occur much more rapidly in adipose tissue than skeletal muscle (20). Chronic fasting markedly reduces GLUT4 mRNA levels in adipose tissue, but has little or no effect on GLUT4 mRNA in skeletal muscle (21). These tissue- specific physiologic adaptations are consistent with the tissue-specific fates of GLUT4- dependent glucose uptake in these tissues. Specifically, muscle must retain its pool of GLUT4 so that it can call on GLUT4 to respond to exercise and muscle contraction. Changes in steady state levels of GLUT4 mRNA could potentially be the result of
Febrile seizure (FS) is the most common type of childhood seizure which occurs in 2-5% of neurologically healthy children. FS is defined as a seizure associated with a febrile illness in the absence of central nervous system infections or acute electrolyte abnormalities in 6-60 months old children without previous afebrile seizures. FS is further classified as simple and complex types. Complex FS is defined as a seizure lasting more than 15 minutes, recurring within 24 hours or focal seizure (1). Iron deficiency is one of the most frequent micronutrient deficiencies that affect at least one
The 3 ′ end of cod GLUT1 was obtained using a combination of genome walking and 3 ′ RLM-RACE. Attempts to clone the remaining 3 ′ sequences by 3 ′ RACE at this point were unsuccessful. This was not surprising since northern blot analysis indicated that the mRNA for cod GLUT1 was approximately 4.6·kb, leaving about 3.7·kb of unknown 3 ′ sequence. Therefore, genome walking was chosen as it would break up the long 3 ′ UTR into more reasonable fragments based upon the presence of restriction enzyme sites within. GenomeWalker libraries for DraI, EcoRV, MscI, MslI, PvuII, SspI and StuI were constructed using the Universal GenomeWalker Kit (BD Biosciences Clontech, Palo Alto, CA, USA) according to the manufacturer’s protocol. The first walk was performed using Primer 7 and the GenomeWalker Adaptor Primer 1 (AP1). The reaction was diluted 1/50 and 1· µ l used as the template for nested PCR using Primer 8 and the GenomeWalker Nested Adaptor Primer 2 (AP2). Both the primary and nested PCR amplifications were performed at 94 ° C for 10·s, 72 ° C for 3·min for 7 cycles, followed by 94 ° C for 10·s, 67 ° C for 30·s, 68 ° C for 3·min for 32 cycles, using Elongase Enzyme Mix (Invitrogen, Burlington, ON, Canada). Bands were obtained for MslI, DraI, SspI, PvuII and StuI, the largest of which was the 1.6·kb band for StuI. Although this band contained a predicted intron (191·bp), the remaining 218·bp of coding sequence and 1070·bp of the 3 ′ UTR were resolved using GENSCAN.
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assessed by spectrophotometry at 260/280 nM wavelength (36). 20 m g of RNA were denatured in formaldehyde and size-fractionated in 1% agarose gels where integrity and relative amounts of RNA were checked by ultraviolet shadowing. The RNA was transferred to “Ny- tran” nylon membranes by capillarity (Schleicher and Schuell Inc., Keene, NH). mRNA from LLC-PK1 cells was also measured on dot blots according to the manufacturer’s instructions (Schleicher and Schuell Inc.). 20 m g of RNA was dissolved in 100 m l of DEPC water with 200 m l of 6.15 M formaldehyde plus 5 3 SSC (1 3 SSC is 150 mM NaCl and 15 mM Na-citrate, pH 7.0), and then blotted using vac- uum on Nytran nylon membranes (37), rinsed under vacuum with 5 3 SSC, dried, and baked (80 8 C, 2 h) under vacuum. The RNA on the membranes was hybridized (38) at high stringency (50% formamide, 2 3 Denhardt’s solution, 1% SDS, 5 3 SSC, and salmon sperm DNA, 100 m g/ml, at 42 8 C) to porcine GLUT1 cDNA (21, 39) and then to rat 28S rRNA (40), both labeled with 32 P-ATP to a sp act of 0.5–1 3 10 9
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in PCa as in liver or lung cancer . However, the correlation between PCa and glycolysis remains largely unknown. Additionally, differ- ences in terms of the expression of key enzymes involved in glycolysis between PCa and adja- cent normal tissue and the relationship among these key enzymes have not been fully eluci- dated. Here, we investigated the expression of five glycolytic enzymes by immunohistochemis- try and found that all enzymes were more highly expressed in PCa tissues than in the adjacent normal tissues. Importantly, the key glycoly- tic enzymes, HK2 and PKM2, were positively stained in almost all 90 cases of PCa tiss- ues, while GLUT1, PFKFB3, and PFKFB4 were positive in approximately 60% of the cases. Table 2. Spearman’s correlation assay of the correlation between proteins expression in PCa tissue PCa tissue GLUT1 staining HK2 staining PFKFB3 staining PFKFB4 staining PKM2 staining
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The NIC transgene expresses both Neu and Cre recombinase; therefore, we used Cre expression as an in- dicator of transgene expression. Strong, nuclear Cre ex- pression was detected in all tumors examined from the NIC-GLUT1 +/+ mice (Fig. 2a), and these tumors also expressed GLUT1 (Fig. 2b). Both Cre recombinase and GLUT1 were detected in tumors from the NIC- GLUT1 F/+ mouse (Fig. 2d and e). In the tumors from the NIC-GLUT1 F/F mouse, we detected Cre recombin- ase, but not epithelial expression of GLUT1 (Fig. 2g and h). GLUT1 was detected in immune cells within the mammary lymph node from the NIC-GLUT1 F/F mouse (Fig. 2h, inset). Histologically, all tumors arising in the NIC mice were consistent with those previously described in this model and in the MMTV-Neu mam- mary tumor model (Fig. 2c, f, and i) . The resolution of immunostaining is not sufficient to determine the ex- tent of GLUT1 staining in the plasma membrane versus the cytoplasm of tumor cells, although it does appear that some GLUT1 may be cytosolic. Together, these data suggest that GLUT1 is required for the early stages of Neu-induced mammary tumorigenesis, and that loss of only a single copy of Slc2a1 is sufficient to prevent the majority of Neu-induced mammary tumors.
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4. Atrah HI, Davidson RJL (1988) Iron deficiency in pernicious anaemia: a neglected diagnosis. Postgrad Med J 64:110-111] 5. Remacha AF, Sarda` MP, Canals C, Queralto` JM et al (2013) Combined cobalamin and iron deficiency anemia: a diagnostic approach using a model based on age and homocysteine assessment. Ann Hematol 92:527-531. doi:10.1007/s00277-012- 1634-8