Fructose Transporter Research Articles

The roles of GLUT5 in cancer progression, metastasis, and drug resistance

Emerging evidence has suggested that high fructose intake, particularly from added sugars and processed foods, is associated with increased cancer risk and progression. The fructose intake is believed to be mediated by the abnormal expression of glucose transporter 5 (GLUT5), the specific fructose transporter in cancer cells. The GLUT5-regulated fructose metabolism has shown to greatly affect cancer progression, metastasis, and drug resistance. This review aims to synchronize the current knowledge to highlight the underlying mechanisms of those impacts and understand the therapeutic potential of GULT5. First, we review the fructose metabolism and its alteration in cancer cells by comparing with glucose metabolism. Subsequently, the key contributors or biological pathways involved in GLUT5-asosociated tumor growth, cancer metastasis, and drug resistance are discussed. The contributions of specific pathways, metabolites, and key enzymes from the fructose metabolism process are also covered, such as enhanced glycolysis for tumor growth, epithelial-mesenchymal transition and angiogenesis for cancer metastasis, and efflux pump expression and activation of survival pathways for cancer drug resistance. The detailed analysis of these mechanisms will allow further understanding of the therapeutic potential of GLUT 5-mediated fructose metabolism in cancer therapy. In particular, targeting GLUT 5 and its-associated processes in fructose metabolism may offer promising strategies for improving cancer treatment outcomes through dietary interventions, specific GLUT5 inhibitors, or in combination.

Exploring fructose metabolism as a potential therapeutic approach for pancreatic cancer.

Excessive fructose intake has been associated with the development and progression of pancreatic cancer. This study aimed to elucidate the relationship between fructose utilization and pancreatic cancer progression. Our findings revealed that pancreatic cancer cells have a high capacity to utilize fructose and are capable of converting glucose to fructose via the AKR1B1-mediated polyol pathway, in addition to uptake via the fructose transporter GLUT5. Fructose metabolism exacerbates pancreatic cancer proliferation by enhancing glycolysis and accelerating the production of key metabolites that regulate angiogenesis. However, pharmacological blockade of fructose metabolism has been shown to slow pancreatic cancer progression and synergistically enhance anti-tumor capabilities when combined with anti-angiogenic agents. Overall, targeting fructose metabolism may prove to be a promising therapeutic approach in the treatment of pancreatic cancer.

Intestinal fructose transporters GLUT5 and GLUT2 in children and adolescents with obesity and metabolic disorders

PurposeThe excessive fructose intake including high-fructose corn syrup (HFCS) may be responsible for increase of obesity occurrence. This study was designed to find potential differences in duodenal fructose transporters on mRNA and protein levels between obese and normal weight children and adolescents. Materials/methodsWe performed a cross-sectional study on a group of 106 hospitalized patients aged 12 to 18. Glucose transporter 2 (GLUT2) and glucose transporter 5 (GLUT5) mRNA as well as protein levels (ELISA and Western blot methods) were assessed in duodenal mucosa biopsies of the patients categorized as obese or normal weight. Additionally, the expression of the aforementioned transporters was analyzed in patients based on the presence of insulin resistance (IR) and metabolic syndrome (MS). ResultsIn children with obesity, increased duodenal protein levels of GLUT5 (Relative protein GLUT5 expression/ACTB) (0.027 ​± ​0.009 vs. 0.011 ​± ​0.006, p ​< ​0.05) but not GLUT2 as compared with the normal weight group, were revealed. No significant differences in duodenal relative GLUT2 and GLUT5 genes expression between the studied groups were found. There was no relationship between the presence of IR or MS and intestinal mRNA GLUT2 and GLUT5 as well as GLUT2 protein expression. ConclusionThe upregulation of the duodenal GLUT5 may contribute to obesity occurrence in children and adolescents.

Direct lactic acid production from raw jerusalem artichoke powder by Lacticaseibacillus paracasei NJ and the comparative transcriptomic profiling of inulin and monosaccharide utilization

A feasible fermentation process for the production of lactic acid (LA), one of the most important food additives, from non-crop feedstocks would help alleviate the challenges of food safety. In this study, 215 g/L inulin-rich Jerusalem artichoke powder (JAP) were directly converted into LA without exogenous nitrogen addition under non-sterile conditions via selected Lacticaseibacillus paracasei NJ. The LA concentration, yield, and average productivity reached 144.08 g/L, 0.67 g/g, and 4.37 g/L/h, respectively, which can be scalable from 5 L to 50 L bioreactors. The viable counts of Lacticaseibacillus reached to 142.30–301.50 × 108 CFU/mL, allowing the products a promising probiotic in food additives. Transcriptional analysis revealed that strain NJ utilized inulin mainly via extracellular β-fructosidase and fructose transporters encoded by the fosRABCDXE operon, and the gene expression within the phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS), pyruvate metabolism, and fatty acid synthesis was significantly altered. For strain NJ grown on JAP, the expressions of genes related to amino acid biosynthesis and peptide/amino acid relevant ATP-binding cassette (ABC) transport system were upregulated to fit nitrogen limitation. We found the availability of L. paracasei NJ for high-yield lactic acid from raw JAP and explores the inner changes of gene expression by utilizing inulin, which provide both promising food additives and new perspectives on the understanding of inulin metabolism in Lacticaseibacillus.

Genome assembly and variant analysis of two Saccharomyces cerevisiae strains isolated from stingless bee pollen

Products from stingless bees are rich reservoirs of microbial diversity, including yeasts with fermentative potential. Previously, two Saccharomyces cerevisiae strains, JP14 and IP9, were isolated from Jataí (Tetragonisca angustula) and Iraí (Nannotrigona testaceicornis) bees, respectively, aiming at mead production. Both strains presented great osmotic and sulfite tolerance, and ethanol production, although they have a high free amino nitrogen demand. Herein, their genomes were sequenced, assembled, and annotated, and the variants were compared to the S. cerevisiae S288c reference strain. The final assembly of IP9 and JP14 presented high N50 and BUSCO scores, and more than 6430 protein-coding genes. Additionally, nQuire predicted the ploidy of IP9 as diploid, but the results were not enough to determine the ploidy of JP14. The mitochondrial genomes of IP9 and JP14 presented the same gene content as S288c but the genes were rearranged and fragmented in different patterns. Meanwhile, the genes with mutations of high impact (e.g., indels, gain of stop codon) for both yeasts were enriched for transmembrane transport, electron transfer, oxidoreductase, heme binding, fructose, mannose, and glucose transport, activities related to the respiratory chain and sugar metabolism. The IP9 strain presented copy number gains in genes related to sugar transport and cell morphogenesis; in JP14, genes were enriched for disaccharide metabolism and transport, response to reactive oxygen species, and polyamine transport. On the other hand, IP9 presented copy number losses related to disaccharide, thiamine, and aldehyde metabolism, while JP14 presented depletions related to disaccharide, oligosaccharide, asparagine, and aspartate metabolism. Notably, both strains presented a killer toxin gene, annotated from the assembling of unmapped reads, representing a potential mechanism for the control of other microorganisms population in the environment. Therefore, the annotated genomes of JP14 and IP9 presented a high selective pressure for sugar and nitrogen metabolism and stress response, consistent with their isolation source and fermentative properties.

Phenotypes of streptozotocin-induced gestational diabetes mellitus in mice.

Gestational diabetes mellitus (GDM) in human patients disrupts glucose metabolism post-pregnancy, affecting fetal development. Although obesity and genetic factors increase GDM risk, a lack of suitable models impedes a comprehensive understanding of its pathology. To address this, we administered streptozotocin (STZ, 75 mg/kg) to C57BL/6N mice for two days before pregnancy, establishing a convenient GDM model. Pregnant mice exposed to STZ (STZ-pregnant) were compared with STZ-injected virgin mice (STZ-virgin), citrate buffer-injected virgin mice (CB-virgin), and pregnant mice injected with citrate buffer (CB-pregnant). STZ-pregnant non-obese mice exhibited elevated blood glucose levels on gestational day 15.5 and impaired glucose tolerance. They also showed fewer normal fetuses compared to CB-pregnant mice. Additionally, STZ-pregnant mice had the highest plasma C-peptide levels, with decreased pancreatic islets or increased alpha cells compared to CB-pregnant mice. Kidneys isolated from STZ-pregnant mice did not display histological alterations or changes in gene expression for the principal glucose transporters (GLUT2 and SGLT2) and renal injury-associated markers. Notably, STZ-pregnant mice displayed decreased gene expression of insulin-receiving molecules (ISNR and IGFR1), indicating heightened insulin resistance. Liver histology in STZ-pregnant mice remained unchanged except for a pregnancy-related increase in lipid droplets within hepatocytes. Furthermore, the duodenum of STZ-pregnant mice exhibited increased gene expression of ligand-degradable IGFR2 and decreased expression of GLUT5 and GLUT12 (fructose and glucose transporters, respectively) compared to STZ-virgin mice. Thus, STZ-pregnant mice displayed GDM-like symptoms, including fetal abnormalities, while organs adapted to impaired glucose metabolism by altering glucose transport and insulin reception without histopathological changes. STZ-pregnant mice offer a novel model for studying mild onset non-obese GDM and species-specific differences in GDM features between humans and animals.

Fructose overconsumption-induced reprogramming of microglia metabolism and function.

The overconsumption of dietary fructose has been proposed as a major culprit for the rise of many metabolic diseases in recent years, yet the relationship between a high fructose diet and neurological dysfunction remains to be explored. Although fructose metabolism mainly takes place in the liver and intestine, recent studies have shown that a hyperglycemic condition could induce fructose metabolism in the brain. Notably, microglia, which are tissue-resident macrophages (Mφs) that confer innate immunity in the brain, also express fructose transporters (GLUT5) and are capable of utilizing fructose as a carbon fuel. Together, these studies suggest the possibility that a high fructose diet can regulate the activation and inflammatory response of microglia by metabolic reprogramming, thereby altering the susceptibility of developing neurological dysfunction. In this review, the recent advances in the understanding of microglia metabolism and how it supports its functions will be summarized. The results from both in vivo and in vitro studies that have investigated the mechanistic link between fructose-induced metabolic reprogramming of microglia and its function will then be reviewed. Finally, areas of controversies and their associated implications, as well as directions that warrant future research will be highlighted.

Abstract 3610: Utilizing fructose metabolism to fuel anti-tumor immunity

Abstract Consumption of fructose in the form of high fructose corn syrup is elevated in the western diet, but it is not fully understood how fructose metabolism influences anti-tumor immunity. Fructose is canonically metabolized in the intestine and the liver and can shunt into glycolysis. In certain tumor types, fructose metabolism becomes deregulated, allowing tumor cells to metabolize fructose to power their biosynthetic processes. Similarly, reprogramming immune cells to be able to efficiently metabolize fructose may help us power immunotherapy to work in difficult to treat cancers, especially in individuals who have high fructose in their blood or in the tumor microenvironment (TME). Imparting effector T cells with the ability to use fructose for fuel can overcome one of the major metabolic limitations of the TME: limited nutrient supply. As T cells become activated, they alter their metabolism to support growth and proliferation. Activated CD8+ T cells upregulate their glycolysis which results in increased biosynthetic processes, such as synthesis of macromolecules and effector cytokines. However, tumor cells also consume substantial amounts of glucose, resulting in a glucose-depleted microenvironment, which could be one of the major barriers to immunotherapy working in solid tumors. Therefore, we engineered T cells to overexpress a fructose transporter, GLUT5, which is normally not expressed in the immune compartment at high levels. We hypothesized that increased fructose uptake by CD8+ T cells will ameliorate exhaustion caused by the glucose-low tumor microenvironment. Indeed, we determined that GLUT5 T cells are more efficient in killing in low glucose supplemented with fructose. We also observed that GLUT5 expression restores T cell glycolytic capacity under fructose to that of unexhausted T cells under glucose replete conditions. Similarly, GLUT5-expressing T cells proliferate faster in vitro and exhibit an improved ability to control B16 tumor growth in vivo. Additionally, combining GLUT5 cell infusion with checkpoint inhibition resulted in durable tumor control and prolonged mouse survival. Further, engineered CD19-targeted CAR T cells with expression of GLUT5 were more metabolically active and cytotoxic in low glucose conditions, mimicking the TME. In conclusion, GLUT5 expression in effector CD8+ and CAR T cells may allow immunotherapy to function in solid tumors under low glucose conditions. Citation Format: Tanya Schild, Rachel Chirayil, Lyric Haughton, Patrick Wallisch, Marjan Berishaj, David A. Scheinberg, Kayvan R. Keshari. Utilizing fructose metabolism to fuel anti-tumor immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3610.

Profiling Cell Heterogeneity and Fructose Transporter Expression in the Rat Nephron by Integrating Single-Cell and Microdissected Tubule Segment Transcriptomes.

Single-cell RNA sequencing (scRNAseq) is a crucial tool in kidney research. These technologies cluster cells based on transcriptome similarity, irrespective of the anatomical location and order within the nephron. Thus, a transcriptome cluster may obscure the heterogeneity of the cell population within a nephron segment. Elevated dietary fructose leads to salt-sensitive hypertension, in part, through fructose reabsorption in the proximal tubule (PT). However, the organization of the four known fructose transporters in apical PTs (SGLT4, SGLT5, GLUT5, and NaGLT1) remains poorly understood. We hypothesized that cells within each subsegment of the proximal tubule exhibit complex, heterogeneous fructose transporter expression patterns. To test this hypothesis, we analyzed rat kidney transcriptomes and proteomes from publicly available scRNAseq and tubule microdissection databases. We found that microdissected PT-S1 segments consist of 81% ± 12% cells with scRNAseq-derived transcriptional characteristics of S1, whereas PT-S2 express a mixture of 18% ± 9% S1, 58% ± 8% S2, and 19% ± 5% S3 transcripts, and PT-S3 consists of 75% ± 9% S3 transcripts. The expression of all four fructose transporters was detectable in all three PT segments, but key fructose transporters SGLT5 and GLUT5 progressively increased from S1 to S3, and both were significantly upregulated in S3 vs. S1/S2 (Slc5a10: 1.9 log2FC, p < 1 × 10-299; Scl2a5: 1.4 log2FC, p < 4 × 10-105). A similar distribution was found in human kidneys. These data suggest that S3 is the primary site of fructose reabsorption in both humans and rats. Finally, because of the multiple scRNAseq transcriptional phenotypes found in each segment, our findings also imply that anatomical labels applied to scRNAseq clusters may be misleading.

Functional Characterization of CsSWEET5a, a Cucumber Hexose Transporter That Mediates the Hexose Supply for Pollen Development and Rescues Male Fertility in Arabidopsis.

Pollen cells require large amounts of sugars from the anther to support their development, which is critical for plant sexual reproduction and crop yield. Sugars Will Eventually be Exported Transporters (SWEETs) have been shown to play an important role in the apoplasmic unloading of sugars from anther tissues into symplasmically isolated developing pollen cells and thereby affect the sugar supply for pollen development. However, among the 17 CsSWEET genes identified in the cucumber (Cucumis sativus L.) genome, the CsSWEET gene involved in this process has not been identified. Here, a member of the SWEET gene family, CsSWEET5a, was identified and characterized. The quantitative real-time PCR and β-glucuronidase expression analysis revealed that CsSWEET5a is highly expressed in the anthers and pollen cells of male cucumber flowers from the microsporocyte stage (stage 9) to the mature pollen stage (stage 12). Its subcellular localization indicated that the CsSWEET5a protein is localized to the plasma membrane. The heterologous expression assays in yeast demonstrated that CsSWEET5a encodes a hexose transporter that can complement both glucose and fructose transport deficiencies. CsSWEET5a can significantly rescue the pollen viability and fertility of atsweet8 mutant Arabidopsis plants. The possible role of CsSWEET5a in supplying hexose to developing pollen cells via the apoplast is also discussed.

Role of Carbohydrate response element-binding protein in mediating dexamethasone-induced glucose transporter 5 expression in Caco-2BBE cells and during the developmental phase in mice

ABSTRACT Glucose transporter 5 (GLUT5), the main fructose transporter in mammals, is primarily responsible for absorbing dietary fructose in the small intestine. The expression of this intestinal gene significantly increases in response to developmental and dietary cues that reach the glucocorticoid receptor and carbohydrate response element-binding protein (ChREBP), respectively. Our study demonstrates that ChREBP is involved in the dexamethasone (Dex)-induced expression of GLUT5 in Caco-2BBE cells and the small intestine of both wild-type and ChREBP-knockout mice. Dex, a glucocorticoid, demonstrated an increase in GLUT5 mRNA levels in a dose- and time-dependent manner. While the overexpression of ChREBP moderately increased GLUT5 expression, its synergistic increase in the presence of Dex was noteworthy, whereas the suppression of ChREBP significantly reduced Dex-induced GLUT5 expression. Dex did not increase ChREBP protein levels but facilitated its nuclear translocation, thereby increasing the activity of the GLUT5 promoter. In vivo experiments conducted on 14-day-old mice pups treated with Dex for three days revealed that only wild-type mice (not ChREBP-knockout mice) exhibited Dex-mediated Glut5 gene induction, which further supports the role of ChREBP in regulating GLUT5 expression. Collectively, our results provide insights into the molecular mechanisms involved in the regulation of GLUT5 expression in response to developmental and dietary signals mediated by glucocorticoids and ChREBP. General significance: The transcription factor ChREBP is important for Dex-mediated Glut5 gene expression in the small intestine.

A Prunus avium L. Infusion Inhibits Sugar Uptake and Counteracts Oxidative Stress-Induced Stimulation of Glucose Uptake by Intestinal Epithelial (Caco-2) Cells.

Sweet cherry (Prunus avium L.) is among the most valued fruits due to its organoleptic properties and nutritional worth. Cherry stems are rich in bioactive compounds, known for their anti-inflammatory and antioxidant properties. Innumerable studies have indicated that some bioactive compounds can modulate sugar absorption in the small intestine. In this study, the phenolic profile of a cherry stem infusion was investigated, as well as its capacity to modulate intestinal glucose and fructose transport in Caco-2 cells. Long-term (24 h) exposure to cherry stem infusion (25%, v/v) significantly reduced glucose (3H-DG) and fructose (14C-FRU) apical uptake, reduced the apical-to-basolateral Papp to 3H-DG, and decreased mRNA expression levels of the sugar transporters SGLT1, GLUT2 and GLUT5. Oxidative stress (induced by tert-butyl hydroperoxide) caused an increase in 3H-DG uptake, which was abolished by the cherry stem infusion. These findings suggest that cherry stem infusion can reduce the intestinal absorption of both glucose and fructose by decreasing the gene expression of their membrane transporters. Moreover, this infusion also appears to be able to counteract the stimulatory effect of oxidative stress upon glucose intestinal uptake. Therefore, it can be a potentially useful compound for controlling hyperglycemia, especially in the presence of increased intestinal oxidative stress levels.

Molecular Cloning, Expression Analysis, and Functional Analysis of Nine IbSWEETs in Ipomoea batatas (L.) Lam

Sugar Will Eventually be Exported Transporter (SWEET) genes play an important regulatory role in plants’ growth and development, stress response, and sugar metabolism, but there are few reports on the role of SWEET proteins in sweet potato. In this study, nine IbSWEET genes were obtained via PCR amplification from the cDNA of sweet potato. Phylogenetic analysis showed that nine IbSWEETs separately belong to four clades (Clade I~IV) and contain two MtN3/saliva domains or PQ-loop superfamily and six~seven transmembrane domains. Protein interaction prediction showed that seven SWEETs interact with other proteins, and SWEETs interact with each other (SWEET1 and SWEET12; SWEET2 and SWEET17) to form heterodimers. qRT-PCR analysis showed that IbSWEETs were tissue-specific, and IbSWEET1b was highly expressed during root growth and development. In addition to high expression in leaves, IbSWEET15 was also highly expressed during root expansion, and IbSWEET7, 10a, 10b, and 12 showed higher expression in the leaves. The expression of SWEETs showed a significant positive/negative correlation with the content of soluble sugar and starch in storage roots. Under abiotic stress treatment, IbSWEET7 showed a strong response to PEG treatment, while IbSWEET10a, 10b, and 12 responded significantly to 4 °C treatment and, also, at 1 h after ABA, to NaCl treatment. A yeast mutant complementation assay showed that IbSWEET7 had fructose, mannose, and glucose transport activity; IbSWEET15 had glucose transport activity and weaker sucrose transport activity; and all nine IbSWEETs could transport 2-deoxyglucose. These results provide a basis for further elucidating the functions of SWEET genes and promoting molecular breeding in sweet potato.

IMMU-23. FRUCTOSE METABOLISM IN MICROGLIA PROMOTES GLIOBLASTOMA GROWTH

Abstract Glioblastoma (GBM) is the most common brain malignancy in adults and has a dismal prognosis. Advances in standard-of-care treatments for GBM have not kept pace with those for other malignancies, in part due to the inefficacy of immunotherapies in treating GBM. The tumor microenvironment (TME) of GBM is heavily populated with pro-tumorigenic tumor-associated macrophages (TAMs) and lacks robust infiltration of T cells. Given that the number of TAMs in the TME correlates with poor prognosis, understanding the mechanisms by which they thrive is essential for discovering new therapies. Bulk RNASeq data from mouse tumors indicates that the fructose transporter, SLC2A5, is highly expressed on TAMs relative to peripheral myeloid cells. Furthermore, human glioblastoma single cell RNASeq (scRNASeq) data indicates that SLC2A5 is expressed specifically on microglia, the tissue resident macrophage of the brain. To assess the necessity of SLC2A5 in microglia, we orthotopically implanted CT2A or QPP4 glioblastoma cells into wild-type mice and mice lacking SLC2A5 (SLC2A5 KO mice). We observed a significant increase in median survival in SLC2A5 KO mice compared to wild-type mice. This indicates a necessity of fructose metabolism in microglia to promote tumorigenesis. Upon ex vivo analysis of the immune compartment of the TME, we observed that microglial cells from tumors in SLC2A5 KO mice have higher expression of MHC-II and PDL1 molecules on the cell surface compared to those from wild-type mice. Additionally, the CD4:CD8 T cell ratio in tumors from SLC2A5 KO mice was lower than that in tumors from wild-type mice. These data suggest that fructose metabolism in microglia facilitates an immunosuppressive phenotype that promotes GBM progression.

PSIV-14 Effects of Maternal Nutrient Restriction During Late Gestation on Expelled Cotyledonary Mrna Expression in Primiparous Beef Females

Abstract To determine the effects of late gestational nutrient restriction on expelled cotyledonary mRNA expression, fall-calving Hereford × Simmental-Angus heifers [single-sired; BW: 451 ± 28 (SD) kg; BCS: 5.4 ± 0.7] bred to a single sire were allocated by fetal sex and expected calving date to receive either 100% (control; CON; n = 12) or 70% (nutrient restricted; NR; n = 11) of NASEM metabolizable energy and metabolizable protein requirements for maintenance, pregnancy, and growth. Beginning on d 160 of gestation, heifers were individually fed chopped sorghum sudan hay (1.74 Mcal ME/kg, 6.69% CP, 72.0% NDF; DM basis) and based on individual intakes, supplemented to meet targeted nutritional planes. Calving was observed for all females and placentas were collected at natural expulsion. Immediately after collection (average: 4.5 h post-calving; range: 2.3 to 12.1 h post-calving), 2 representative cotyledons on the ipsilateral side were cleaned with phosphate buffered saline, and cotyledonary tissue was flash frozen. Total RNA was extracted, and RT-qPCR was performed for mRNA expression of glucose transporters (SLC2A1, SLC2A3, SLC2A4), a fructose transporter (SLC2A5), amino acid transporters (SLC1A1, SLC3A2, SLC7A1, SLC7A5, SLC7A8, SLC38A2), angiogenic factors/receptors (VEGFA, NOS3, FLT1, KDR, GUCY1B3), and pregnancy-associated glycoprotein 2 (PAG2). Relative mRNA expression was analyzed with nutritional plane, treatment initiation date, time to cotyledon sampling, and calf sex (when P &amp;lt; 0.25) as fixed effects. We have previously reported that total uterine blood flow during late gestation was not affected (P ≥ 0.20) by nutritional plane or the nutritional plane × day of gestation interaction. Post-calving, NR dams were 64 kg BW less (P &amp;lt; 0.001) and 2.0 BCS less (P &amp;lt; 0.001), but calf birth weight was not affected (P = 0.72) by nutritional plane. Contralateral placental dry weight tended to be less (P = 0.07) in NR dams, but total placental weight was not affected (P = 0.86) by late gestational nutritional plane. Cotyledonary relative mRNA expression of SLC2A3 and SLC38A2 was greater (P ≤ 0.05) and relative expression of SLC2A1, SLC2A4, and NOS3 tended to be greater (P ≤ 0.07) in NR dams compared with CON. Late gestational nutritional plane did not affect (P ≥ 0.13) cotyledonary relative mRNA expression of FLT1, GUCY1B3, KDR, PAG2, SLC1A1, SLC2A5, SLC3A2, SLC7A1, SLC7A5, SLC7A8, and VEGFA. In summary, first parity beef females who experienced late gestational nutrient restriction maintained uterine blood flow and placental mass and had 4 nutrient transporters and 1 angiogenic factor upregulated in cotyledons, all which likely contributed to their conservation of fetal growth.

GLUT5, GLUT7, and GLUT11 expression and Bcl-2/Bax ratio on Breast Cancer Cell Line MCF-7 Treated with Fructose and Glucose.

Fructose and glucose are types of sugars commonly found in the diet that have been linked to cancer development. Glucose transporters (GLUTs) are facilitating the uptake of these hexoses. Expression of GLUT5 is higher in cancer cells than in healthy tissue. GLUT7 and GLUT11 facilitate the transport of glucose and fructose; however, their expression in breast cancer has not been extensively studied. The Bcl-2 family has been known as a regulator of the cell's survival and death. Here, we investigated the effect of the fructose-glucose combination in MCF-7 breast cancer cells on the viability, migration, and expression of GLUT5, GLUT7, GLUT11, and Bcl-2/Bax ratio. Breast cancer cells MCF-7 were treated with fructose, glucose, and combinations of fructose:glucose (75%:25%, 50%:50%, 25%:75%). Cell viability was assessed using an MTT test. Cell migration was examined with a wound-healing assay. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was performed to evaluate the mRNA expression of GLUT5, GLUT7, GLUT11, and Bcl-2/Bax. The viability and migration of MCF-7 breast cancer cells elevated when treated with a combination of fructose and glucose, and glucose alone, compared to fructose alone. The expression levels of GLUT5 and GLUT7 were highest in combination of fructose:glucose (75%:25%). Conversely, the expression of GLUT11 was consistently low across all treated media. The highest Bcl-2/Bax ratio was shown in fructose:glucose combination (25%:75%). The viability, migration, and Bcl-2/Bax ratio are enhanced in the combination media with higher glucose. In contrast, when the fructose composition was higher in the media, expression of GLUT5 and GLUT7 increased.

Viability, Migration Rate, and mRNA Expression of GLUT5, GLUT7, GLUT11 in WiDr Colorectal Cancer Cell Line

BACKGROUND: Insufficient glucose levels in colorectal cancer (CRC) patients leads to a condition where fructose might become an alternative source for cells proliferation, but the role of fructose or fructose-glucose combinations in development of CRC has not been elucidated well. In this study, the effect of fructose-glucose variations on viability, migration, and glucose transporter (GLUT)5, GLUT7, GLUT11 mRNA expressions in WiDr CRC cell line were examined.METHODS: Cells were treated with varying ratios of fructose-glucose (F100%; F75%:G25%; F50%:G50%; F25%:G75%; G100%; F: Fructose, G: Glucose). Untreated cells (F0:G0) were used as cell control. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was used for cell viability test, scratch assay was used to examine the cell migration, and quantitative polymerase chain reaction (qPCR) was performed to examine mRNA expressions. Data were analyzed using one-way analysis of variance (ANOVA) and followed with Tukey's post-hoc test, with p&lt;0.05 consideres as significant.RESULTS: Fructose-glucose combinations and glucose 100% significantly increased the cell viability compared to control (p&lt;0.05). All treatment groups showed a significant increase in cell migration compared to control (p=0.000). Only GLUT7 and GLUT11 expressions in the G100% group were significantly different compared to the control (p=0.000). GLUT7 and GLUT11 expressions were also significantly different in F100% and F50%:G50% treatments compared to G100% (p=0.000).CONCLUSION: Taken together, fructose might play important role in cell migration. However, in cell viability, combination with glucose could increase fructose's effect. Fructose might not affect the mRNA expressions of GLUT5, GLUT7 and GLUT11.KEYWORDS: GLUT5, GLUT7, GLUT11, fructose transporter, colorectal cancer, WiDr

Altered expression of proteins involved in metabolism in LGMDR1 muscle is lost in cell culture conditions

BackgroundLimb-girdle muscular dystrophy R1 calpain 3-related (LGMDR1) is an autosomal recessive muscular dystrophy due to mutations in the CAPN3 gene. While the pathophysiology of this disease has not been clearly established yet, Wnt and mTOR signaling pathways impairment in LGMDR1 muscles has been reported.ResultsA reduction in Akt phosphorylation ratio and upregulated expression of proteins implicated in glycolysis (HK-II) and in fructose and lactate transport (GLUT5 and MCT1) in LGMDR1 muscle was observed. In vitro analysis to establish mitochondrial and glycolytic functions of primary cultures were performed, however, no differences between control and patients were observed. Additionally, gene expression analysis showed a lack of correlation between primary myoblasts/myotubes and LGMDR1 muscle while skin fibroblasts and CD56− cells showed a slightly better correlation with muscle. FRZB gene was upregulated in all the analyzed cell types (except in myoblasts).ConclusionsProteins implicated in metabolism are deregulated in LGMDR1 patients’ muscle. Obtained results evidence the limited usefulness of primary myoblasts/myotubes for LGMDR1 gene expression and metabolic studies. However, since FRZB is the only gene that showed upregulation in all the analyzed cell types it is suggested its role as a key regulator of the pathophysiology of the LGMDR1 muscle fiber. The Wnt signaling pathway inactivation, secondary to FRZB upregulation, and GLUT5 overexpression may participate in the impaired adipogenesis in LGMD1R patients.

Abstract 044: Knocking Out Sodium Glucose-Linked Transporter 5 Prevents Fructose-Induced Salt-Sensitive Hypertension

We showed: 1) a 20% fructose high-salt (3.7% NaCl; FHS) but not a 20% glucose high-salt diet increases systolic blood pressure (SBP) in a sex independent manner; 2) on normal salt, 20% fructose augments the ability of angiotensin II (Ang-II) to increase proximal tubule superoxide (O 2 - ); and 3) rat proximal tubule brush borders express sodium glucose-linked transporters (SGLT) 4 and SGLT5, which transport fructose. However, whether FHS enhances Ang II-stimulated proximal tubule O 2 - , its role in FISSH and the roles of SGLT4 and 5 are unknown. We hypothesized that fructose transport mediated by SGLT5 but not SGLT4 is required for FHS to enhance Ang II-stimulated proximal tubule O 2 - in males and females, and this is required for fructose-induced salt-sensitive hypertension (FISSH). We measured SBP in wild-type, SGLT4 knockout, and SGLT5 knockout male and female rats on a Sprague Dawley background fed FHS for 7 days. We also measured basal and Ang II-stimulated (37 nM) O 2 - in proximal tubule suspensions from these rats using the lucigenin assay. FHS increased SBP in wild type male rats 11 ± 2 mmHg (from 131 ± 4 to 142 ± 5 mmHg; n = 5, p &lt; 0.01) and in female rats by 21 ± 5 mmHg (from 114 ± 4 to 130 ± 3 mmHg; n = 8; p &lt; 0.01), not significantly different from males. FHS increased SBP in SGLT4 males by 22 ± 3 mmHg (from 126 ± 5 to 149 ± 6 mmHg; n = 8, p &lt; 0.01) and in females 21 ± 5 mmHg (from 126 ± 3 to 138 ± 2 mmHg; n = 8; p &lt; 0.01), not significantly different from males. In SGLT5 males SBP was 128 ± 6 on day 0 and 129 ± 8 mmHg on day 7 (n = 7). In SGLT5 females SBP was 114 ± 4 on day 0 and 117 ± 5 mmHg on day 7 (n = 7). Ang-II increased O 2 - by 11 ± 3 relative light units (RLU)/μg/s from 53 ± 9 to 64 ± 8 RLU/μg/s (n = 11, p &lt; 0.01) in tubules from wild type males fed FHS for 7 days and by 10±3 RLU (n = 5; p &lt; 0.02) in females fed FHS for 7 days. Ang-II increased O 2 - by 8 ± 2 RLU/μg/s in proximal tubules from FHS-fed SGLT4 males (n = 6, p &lt; 0.02) and by 8 ± 2 in FHS-fed females (n = 6, p &lt; 0.01). Ang-II did not significantly stimulate O 2 - in proximal tubules from FHS-fed SGTL5 male or female rats (Δ 3±7 and 0±3 RLU/μg/s, n = 5 and 3, respectively). We conclude that transport by SGLT 5 but not SGLT4 is required for FHS to augment Ang II-stimulated proximal tubule O 2 - in males and females, and that the increase in proximal tubule oxidative stress is required for FISSH.

Determinants of sugar-induced influx in the mammalian fructose transporter GLUT5

In mammals, glucose transporters (GLUT) control organism-wide blood-glucose homeostasis. In human, this is accomplished by 14 different GLUT isoforms, that transport glucose and other monosaccharides with varying substrate preferences and kinetics. Nevertheless, there is little difference between the sugar-coordinating residues in the GLUT proteins and even the malarial Plasmodium falciparum transporter PfHT1, which is uniquely able to transport a wide range of different sugars. PfHT1 was captured in an intermediate ‘occluded’ state, revealing how the extracellular gating helix TM7b has moved to break and occlude the sugar-binding site. Sequence difference and kinetics indicated that the TM7b gating helix dynamics and interactions likely evolved to enable substrate promiscuity in PfHT1, rather than the sugar-binding site itself. It was unclear, however, if the TM7b structural transitions observed in PfHT1 would be similar in the other GLUT proteins. Here, using enhanced sampling molecular dynamics simulations, we show that the fructose transporter GLUT5 spontaneously transitions through an occluded state that closely resembles PfHT1. The coordination of D-fructose lowers the energetic barriers between the outward- and inward-facing states, and the observed binding mode for D-fructose is consistent with biochemical analysis. Rather than a substrate-binding site that achieves strict specificity by having a high affinity for the substrate, we conclude GLUT proteins have allosterically coupled sugar binding with an extracellular gate that forms the high-affinity transition-state instead. This substrate-coupling pathway presumably enables the catalysis of fast sugar flux at physiological relevant blood-glucose concentrations.