Sodium-dependent Vitamin C Transporter 2 Research Articles

Ascorbate Uptake and Retention by Breast Cancer Cell Lines and the Intracellular Distribution of Sodium-Dependent Vitamin C Transporter 2.

Ascorbate plays a vital role as a co-factor for a superfamily of enzymes, the 2-oxoglutarate dependent dioxygenases (2-OGDDs), which govern numerous pathways in cancer progression, including the hypoxic response and the epigenetic regulation of gene transcription. Ascorbate uptake into most cells is through active transport by the sodium-dependent vitamin C transporter 2 (SVCT2). The aims of this study were to determine the kinetics of ascorbate uptake and retention by breast cancer cell lines under various oxygen conditions, and to investigate the role of SVCT2 in mediating ascorbate uptake and intracellular trafficking. Human MDA-MB231 cells accumulated up to 5.1 nmol ascorbate/106 cells, human MCF7 cells 4.5 nmol/106 cells, and murine EO771 cells 26.7 nmol/106 cells. Intracellular ascorbate concentrations decreased rapidly after reaching maximum levels unless further ascorbate was supplied to the medium, and there was no difference in the rate of ascorbate loss under normoxia or hypoxia. SVCT2 was localised mainly to subcellular compartments, with the nucleus apparently containing the most SVCT2 protein, followed by the mitochondria. Much less SVCT2 staining was observed on the plasma membrane. Our data showed that careful management of the doses and incubation times with ascorbate in vitro allows for an approximation of in vivo conditions. The localisation of SVCT2 suggests that the distribution of ascorbate to intracellular compartments is closely aligned to the known function of ascorbate in supporting 2-OGDD enzymatic functions in the organelles and with supporting antioxidant protection in the mitochondria.

SVCT2/SLC23A2 is a sodium-dependent urate transporter: functional properties and practical application

Urate transporters play a pivotal role in urate handling in the human body, but the urate transporters identified to date do not account for all known molecular processes of urate handling, suggesting the presence of latent machineries. We recently showed that a urate transporter SLC2A12 is also a physiologically important exporter of ascorbate (the main form of vitamin C in the body) that would cooperate with an ascorbate importer, sodium-dependent vitamin C transporter 2 (SVCT2). Based on the dual functions of SLC2A12 and cooperativity between SLC2A12 and SVCT2, we hypothesized that SVCT2 might be able to transport urate. To test this proposal, we conducted cell-based analyses using SVCT2-expressing mammalian cells. The results demonstrated that SVCT2 is a novel urate transporter. Vitamin C inhibited SVCT2-mediated urate transport with a half-maximal inhibitory concentration of 36.59μM, suggesting that the urate transport activity may be sensitive to physiological ascorbate levels in blood. Similar results were obtained for mouse Svct2. Further, using SVCT2 as a sodium-dependent urate importer, we established a cell-based urate efflux assay that will be useful for identification of other novel urate exporters as well as functional characterization of nonsynonymous variants of already-identified urate exporters including ATP-binding cassette transporter G2. While more studies will be needed to elucidate the physiological impact of SVCT2-mediated urate transport, our findings deepen understanding of urate transport machineries.

Role of vitamin C and SVCT2 in neurogenesis.

Different studies have established the fundamental role of vitamin C in proliferation, differentiation, and neurogenesis in embryonic and adult brains, as well as in in vitro cell models. To fulfill these functions, the cells of the nervous system regulate the expression and sorting of sodium-dependent vitamin C transporter 2 (SVCT2), as well as the recycling of vitamin C between ascorbic acid (AA) and dehydroascorbic acid (DHA) via a bystander effect. SVCT2 is a transporter preferentially expressed in neurons and in neural precursor cells. In developmental stages, it is concentrated in the apical region of the radial glia, and in adult life, it is expressed preferentially in motor neurons of the cerebral cortex, starting on postnatal day 1. In neurogenic niches, SVCT2 is preferentially expressed in precursors with intermediate proliferation, where a scorbutic condition reduces neuronal differentiation. Vitamin C is a potent epigenetic regulator in stem cells; thus, it can induce the demethylation of DNA and histone H3K27m3 in the promoter region of genes involved in neurogenesis and differentiation, an effect mediated by Tet1 and Jmjd3 demethylases, respectively. In parallel, it has been shown that vitamin C induces the expression of stem cell-specific microRNA, including the Dlk1-Dio3 imprinting region and miR-143, which promotes stem cell self-renewal and suppresses de novo expression of the methyltransferase gene Dnmt3a. The epigenetic action of vitamin C has also been evaluated during gene reprogramming of human fibroblasts to induced pluripotent cells, where it has been shown that vitamin C substantially improves the efficiency and quality of reprogrammed cells. Thus, for a proper effect of vitamin C on neurogenesis and differentiation, its function as an enzymatic cofactor, modulator of gene expression and antioxidant is essential, as is proper recycling from DHA to AA by various supporting cells in the CNS.

Blue light promotes vitamin C-mediated ferroptosis of melanoma through specifically upregulating transporter SVCT2 and generating Fe2+

Vitamin C (VC)-based cancer therapy is a promising therapeutic approach for a variety of cancers due to its profound effects on redox reactions and metabolic pathways. However, high administration dosage of VC for necessary therapeutic efficacy for cancers increases the risk of overt side effects and limits its clinical use. Here, we show cutaneous blue light irradiation can specifically upregulate the sodium-dependent vitamin C transporter 2 (SVCT2) of the tumor and increase effectively the VC concentration at the tumor sites by an overall low dosage administration. In the mouse melanoma model, blue light stimulates the SVCT2 expression through the nuclear factor-kappa B (NF-κB) signaling pathway both in vitro and in vivo. The increased cellular VC together with Fe2+ generated by blue light simultaneously elevate cellular oxidative stress and trigger the ferroptosis of melanoma. With this revealed mechanism, the synergistic actions of blue light on the VC transporter and Fe2+ generation lead to a ca. 20-fold reduction in the administration dosage of VC with an effective melanoma elimination and prolonged survival. The work defines the killing mechanism of blue light on VC-based cancer therapy and provides a practical approach for promoting VC uptake. This light-assisted VC therapy is not only highly efficient for melanoma but also considerable for a broad clinical utility.

Research Note: Heat stress affects immune and oxidative stress indices of the immune organs of broilers by changing the expressions of adenosine triphosphate-binding cassette subfamily G member 2, sodium-dependent vitamin C transporter-2, and mitochondrial calcium uniporter

This study aimed to determine the mechanisms of heat-induced oxidative stress in the thymus and spleen of broilers. After 28 d, 30 broilers were randomly divided into the control (25°C ± 2°C; 24 h/d) and heat-stressed (36°C ± 2°C; 8 h/d) groups; the experiment lasted for 1 wk. The broilers in each group were euthanized, and some samples were collected and analyzed at 35 d. The results showed that the birds subjected to heat stress reduced the weight (P < 0.01) and the indices of thymus (P < 0.01), the activities of T-AOC (P < 0.01) and SOD (P < 0.05) of spleen, and levels of IL-10 (P < 0.05) and the GSH-PX (P < 0.05) in thymus and spleen, and increased the IL-6 content of thymus (P < 0.05), the MDA content (P < 0.01), and the reactive oxygen species (ROS) levels (P < 0.01) in thymus and spleen. Moreover, the expression of the IgG gene in the thymus and spleen of heat-stressed broilers was increased (P < 0.05); however, the expression of the IgM gene in the spleen was increased (P < 0.05), with no difference (P > 0.05) in the thymus of heat-stressed broilers compared with the control. Furthermore, the relative expression of adenosine triphosphate-binding cassette subfamily G member 2 (ABCG2) in the thymus and spleen both increased (P < 0.05). The sodium-dependent vitamin C transporter-2 (SVCT-2) (P < 0.01) and mitochondrial calcium uniporter (MCU) (P < 0.01) mRNA levels in the thymus of heat-stressed broilers increased, and the expression of ABCG2 (P < 0.05), SVCT-2 (P < 0.01), and MCU (P < 0.01) proteins in the thymus and spleen of heat-stressed broilers increased compared with the control group. This study confirmed that heat stress-induced oxidative stress in the immune organs of broilers, further reducing immune function.

Structural basis of vitamin C recognition and transport by mammalian SVCT1 transporter

Vitamin C (L-ascorbic acid) is an essential nutrient for human health, and its deficiency has long been known to cause scurvy. Sodium-dependent vitamin C transporters (SVCTs) are responsible for vitamin C uptake and tissue distribution in mammals. Here, we present cryogenic electron microscopy structures of mouse SVCT1 in both the apo and substrate-bound states. Mouse SVCT1 forms a homodimer with each protomer containing a core domain and a gate domain. The tightly packed extracellular interfaces between the core domain and gate domain stabilize the protein in an inward-open conformation for both the apo and substrate-bound structures. Vitamin C binds at the core domain of each subunit, and two potential sodium ions are identified near the binding site. The coordination of sodium ions by vitamin C explains their coupling transport. SVCTs probably deliver substrate through an elevator mechanism in combination with local structural arrangements. Altogether, our results reveal the molecular mechanism by which SVCTs recognize vitamin C and lay a foundation for further mechanistic studies on SVCT substrate transport.

Overcoming EGFR Resistance in Metastatic Colorectal Cancer Using Vitamin C: A Review.

Targeted monoclonal antibody therapy against Epidermal Growth Factor Receptor (EGFR) is a leading treatment modality against metastatic colorectal cancer (mCRC). However, with the emergence of KRAS and BRAF mutations, resistance was inevitable. Cells harboring these mutations overexpress Glucose Transporter 1 (GLUT1) and sodium-dependent vitamin C transporter 2 (SVCT2), which enables intracellular vitamin C transport, leading to reactive oxygen species generation and finally cell death. Therefore, high dose vitamin C is proposed to overcome this resistance. A comprehensive search strategy was adopted using Pubmed and MEDLINE databases (up to 11 August 2022). There are not enough randomized clinical trials to support its use in the clinical management of mCRC, except for a subgroup analysis from a phase III study. High dose vitamin C shows a promising role in overcoming EGFR resistance in mCRC with wild KRAS mutation with resistance to anti-epidermal growth factor inhibitors and in patients with KRAS and BRAF mutations.

Vitamin C transporter SVCT1 serves a physiological role as a urate importer: functional analyses and in vivo investigations

Uric acid, the end product of purine metabolism in humans, is crucial because of its anti-oxidant activity and a causal relationship with hyperuricemia and gout. Several physiologically important urate transporters regulate this water-soluble metabolite in the human body; however, the existence of latent transporters has been suggested in the literature. We focused on the Escherichia coli urate transporter YgfU, a nucleobase-ascorbate transporter (NAT) family member, to address this issue. Only SLC23A proteins are members of the NAT family in humans. Based on the amino acid sequence similarity to YgfU, we hypothesized that SLC23A1, also known as sodium-dependent vitamin C transporter 1 (SVCT1), might be a urate transporter. First, we identified human SVCT1 and mouse Svct1 as sodium-dependent low-affinity/high-capacity urate transporters using mammalian cell-based transport assays. Next, using the CRISPR-Cas9 system followed by the crossing of mice, we generated Svct1 knockout mice lacking both urate transporter 1 and uricase. In the hyperuricemic mice model, serum urate levels were lower than controls, suggesting that Svct1 disruption could reduce serum urate. Given that Svct1 physiologically functions as a renal vitamin C re-absorber, it could also be involved in urate re-uptake from urine, though additional studies are required to obtain deeper insights into the underlying mechanisms. Our findings regarding the dual-substrate specificity of SVCT1 expand the understanding of urate handling systems and functional evolutionary changes in NAT family proteins.

SVCT2-GLUT1-mediated ascorbic acid transport pathway in rat dental pulp and its effects during wound healing

Ascorbic acid (AA; vitamin C) plays a crucial role in the biosynthesis and secretion of collagen to produce the organic matrix of hard tissues. Nevertheless, the detailed mechanism by which AA induces reparative dentinogenesis is still unknown. This study aimed to investigate the pathway and function of AA during wound healing in a rat pulpotomy model. Sodium-dependent vitamin C transporter (SVCT) 2 and glucose transporter (GLUT) 1 were detected in odontoblasts, endothelial cells, and nerve fibers in normal pulp tissues. SVCT2 and GLUT1 were also expressed in odontoblast-like cells in pulpotomized tissues of Wistar rats, and immunopositive cells of SVCT2 were significantly increased at 5 days after pulpotomy (p < 0.05). By contrast, osteogenic disorder Shionogi (ODS) rats, which cannot generate AA, also expressed SVCT2 and GLUT1 in normal and wound healing conditions. However, in ODS rats, when compared with the AA-addition group, the formation of dentin bridges in the AA-loss group was not evident, a layer of osteopontin was significantly increased beneath the wound surface (p < 0.05), and alpha smooth muscle actin at the odontoblast-like cells observed along this layer was significantly increased (p < 0.05), but not Nestin. Moreover, the amounts of type 1 collagen generated in the reparative dentin and beneath the wound healing site were significantly diminished (p < 0.05). Macrophages expressing CD68 and CD206 increased beneath the wound site. Hence, AA may be involved in odontoblast-like cell differentiation and anti-inflammatory response during dental pulp wound healing. Our results provide new insights into the function of AA through SVCT2 and GLUT1 in reparative dentinogenesis and may help in developing new therapeutic targets for dental pulpal disease.

Vitamin C Promotes Muscle Development Mediated by the Interaction of CSRP3 with MyoD and MyoG.

Previous studies have reported that vitamin C (VC), an essential nutrient, exerts beneficial effects on muscle health. However, the molecular mechanism involved in the VC-mediated regulation of muscle development is still unclear. The roles of VC in muscle development and the underlying molecular mechanisms were examined using cell and molecular biology, transcriptomics, proteomics, and animal experiments in this study. VC upregulated the expression of sodium-dependent vitamin C transporter 2 (SVCT2) and cysteine rich protein 3 (CSRP3). Additionally, VC promoted the differentiation of C2C12 cells and the repair of mouse muscle injury by upregulating the nuclear translocation of CSRP3, which subsequently interacted with MyoD and MyoG. This study provided a theoretical basis for elucidating the mechanism underlying the VC-mediated regulation of muscle development, as well as for developing animal nutritional supplements and therapeutic drugs for muscle diseases.

Glioblastoma Invasiveness and Collagen Secretion Are Enhanced by Vitamin C.

Aims: Glioblastoma (GB) is one of the most aggressive brain tumors. These tumors modify their metabolism, increasing the expression of glucose transporters, GLUTs, which incorporate glucose and the oxidized form of vitamin C, dehydroascorbic acid (DHA). We hypothesized that GB cells preferentially take up DHA, which is intracellularly reduced and compartmentalized into the endoplasmic reticulum (ER), promoting collagen biosynthesis and an aggressive phenotype. Results: Our results showed that GB cells take up DHA using GLUT1, while GLUT3 and sodium-dependent vitamin C transporter 2 (SVCT2) are preferably intracellular. Using a baculoviral system and reticulum-enriched extracts, we determined that SVCT2 is mainly located in the ER and corresponds to a short isoform. Ascorbic acid (AA) was compartmentalized, stimulating collagen IV secretion and increasing in vitro and in situ cell migration. Finally, orthotopic xenografts induced in immunocompetent guinea pigs showed that vitamin C deficiency retained collagen, reduced blood vessel invasion, and affected glomeruloid vasculature formation, all pathological conditions associated with malignancy. Innovation and Conclusion: We propose a functional role for vitamin C in GB development and progression. Vitamin C is incorporated into the ER of GB cells, where it favors the synthesis of collagen, thus impacting tumor development. Collagen secreted by tumor cells favors the formation of the glomeruloid vasculature and enhances perivascular invasion. Antioxid. Redox Signal. 37, 538-559.

Exploring the Pivotal Neurophysiologic and Therapeutic Potentials of Vitamin C in Glioma

Gliomas represent solely primary brain cancers of glial cell or neuroepithelial origin. Gliomas are still the most lethal human cancers despite modern innovations in both diagnostic techniques as well as therapeutic regimes. Gliomas have the lowest overall survival rate compared to other cancers 5 years after definitive diagnosis. The dietary intake of vitamin C has protective effect on glioma risk. Vitamin C is an essential compound that plays a vital role in the regulation of lysyl and prolyl hydroxylase activity. Neurons store high levels of vitamin C via sodium dependent-vitamin C transporters (SVCTs) to protect them from oxidative ischemia-reperfusion injury. Vitamin C is a water-soluble enzyme, typically seen as a powerful antioxidant in plants as well as animals. The key function of vitamin C is the inhibition of redox imbalance from reactive oxygen species produced via the stimulation of glutamate receptors. Gliomas absorb vitamin C primarily via its oxidized dehydroascorbate form by means of GLUT 1, 3, and 4 and its reduced form, ascorbate, by SVCT2. Vitamin C is able to preserve prosthetic metal ions like Fe2+ and Cu+ in their reduced forms in several enzymatic reactions as well as scavenge free radicals in order to safeguard tissues from oxidative damage. Therapeutic concentrations of vitamin C are able to trigger H2O2 generation in glioma. High-dose combination of vitamin C and radiation has a much more profound cytotoxic effect on primary glioblastoma multiforme cells compared to normal astrocytes. Control trials are needed to validate the use of vitamin C and standardization of the doses of vitamin C in the treatment of patients with glioma.

Determination of tissue-specific interaction between vitamin C and vitamin E in vivo using senescence marker protein-30 knockout mice as a vitamin C synthesis deficiency model.

Vitamin E (α-tocopherol; VE) is known to be regenerated from VE radicals by vitamin C (L-ascorbic acid; VC) in vitro. However, their in vivo interaction in various tissues is still unclear. Therefore, we alternatively examined the in vivo interaction of VC and VE by measurement of their concentrations in various tissues of senescence marker protein-30 (SMP30) knockout (KO) mice as a VC synthesis deficiency model. Male SMP30-KO mice were divided into four groups (VC+/VE+, VC+/VE-, VC-/VE+ and VC-/VE-), fed diets with or without 500 mg/kg VE and given water with or without 1·5 g/l VC ad libitum. Then, VC and VE concentrations in the plasma and various tissues were determined. Further, gene expression levels of transporters associated with VC and VE, such as α-tocopherol transfer protein (α-TTP) and sodium-dependent vitamin C transporters (SVCTs), were examined. These results showed that the VE levels in the VC-depleted (VC-/VE+) group were significantly lower than those in the VC+/VE+ group in the liver and heart; the VC levels in the VE-depleted (VC+/VE-) group were significantly lower than those in the VC+/VE+ group in the kidneys. The α-TTP gene expression in the liver and kidneys was decreased by VC and/or VE depletion. Moreover, SVCT1 gene expression in the liver was decreased by both VC and VE depletion. In conclusion, these results indicate that VC spares VE mainly in the liver and heart and that VE spares VC in the kidneys of SMP30-KO mice. Thus, interaction between VC and VE is likely to be tissue specific.

Calsyntenin-3 interacts with the sodium-dependent vitamin C transporter-2 to regulate vitamin C uptake

Ascorbic acid (AA) uptake in neurons occurs via a Na+-dependent carrier-mediated process mediated by the sodium-dependent vitamin C transporter-2 (SVCT2). Relatively little information is available concerning the network of interacting proteins that support human (h)SVCT2 trafficking and cell surface expression in neuronal cells. Here we identified the synaptogenic adhesion protein, calsyntenin-3 (CLSTN3) as an hSVCT2 interacting protein from yeast two-hybrid (Y2H) screening of a human adult brain cDNA library. This interaction was confirmed by co-immunoprecipitation, mammalian two-hybrid (M2H), and co-localization in human cell lines. Co-expression of hCLSTN3 with hSVCT2 in SH-SY5Y cells led to a marked increase in AA uptake. Reciprocally, siRNA targeting hCLSTN3 inhibited AA uptake. In the J20 mouse model of Alzheimer's disease (AD), mouse (m)SVCT2 and mCLSTN3 expression levels in hippocampus were decreased. Similarly, expression levels of hSVCT2 and hCLSTN3 were markedly decreased in hippocampal samples from AD patients. These findings establish CLSTN3 as a novel hSVCT2 interactor in neuronal cells with potential pathophysiological significance.

Limited Association Between Ascorbate Concentrations and Vitamin C Transporters in Renal Cell Carcinoma Cells and Clinical Samples

Maintenance of whole-body ascorbate levels and distribution is mediated via sodium-dependent vitamin C transporters (SVCTs). The kidney is one of a few organs that express both SVCT1 and SVCT2. Recent evidence suggests that accumulation of ascorbate may be different in tumour compared to normal tissue, but data on SVCT levels in tumours is sparse. The role of the two SVCT isoforms in ascorbate uptake in renal cell carcinoma (RCC) was investigated in vitro and in clinical samples. In three human RCC cell lines, we investigated SVCT protein levels and cellular location in response to ascorbate supplementation and withdrawal. In clinical RCC samples (n=114), SVCT patterns of staining and protein levels were analysed and compared to ascorbate levels. In cell culture, transporter levels and cellular location were not modified by ascorbate availability at any time up to 8h, although basal SVCT2 levels governed maximal ascorbate accumulation. In clinical samples, SVCT1 protein levels in papillary RCC (pRCC) were similar to matched normal renal cortex, but were increased in clear-cell RCC (ccRCC). Native SVCT2 (72 kDa) was significantly decreased in both pRCC and ccRCC tissues compared to cortex (p<0.01), whereas a modified form of SVCT2 (100 kDa) was significantly increased (p<0.001). There was no association between the transporters (SVCT1, native or modified SVCT2) and ascorbate concentrations in either normal or tumour tissues. SVCT1 and SVCT2 displayed diffuse cytoplasmic staining in both pRCC and ccRCC tumour cells, with cortex showing distinct membrane staining for SVCT1. We observed a re-distribution of ascorbate transporters in tumour tissue compared to normal cortex and a shift from native to modified SVCT2 in cell culture and clinical samples. Data presented here show that SVCT protein levels do not appear to predict intracellular ascorbate accumulation in RCC.

Exploring the effectiveness of in ovo feeding of vitamin C based on the embryonic vitamin C synthesis and absorption in broiler chickens

BackgroundMany researches about in ovo feeding (IOF) of vitamin C (VC) are gradually carried out to explore physiological development in chicken, but little studies focus on VC synthesis capacity of the embryo itself, the selection of injection site and the effectiveness of IOF of VC. This study aims to explore the above problems.ResultsKidney and yolk sac were the main organs for VC synthesis and L-gulonolactone oxidase (GLO) expression was lower during pre-hatch development than that during post-hatch development. Sodium-dependent vitamin C transporter 1 (SVCT1) expression was increased continuously in yolk sac from embryonic age 19 (E19) to post-hatch day 1 (D1) and in intestine (duodenum, jejunum and ileum) from E17 to D1. Plasma VC content was higher at D1 than that at D21 and D42. IOF of VC significantly reduced GLO expression in liver, kidney and yolk sac as well as SVCT1 expression in duodenum, jejunum and ileum, but increased the VC content in plasma, brain, kidney and liver. In addition, IOF of VC obviously reduced the embryonic morality and increased the hatchability under heat stress.ConclusionsThis study suggested that IOF of VC at E11 in yolk was effective for embryonic VC supplementation. These findings provide a theoretical reference about the method of embryonic VC supplementation and effective methodology on embryonic VC nutrition in broiler chickens.

Vitamin C Enhances Antiviral Functions of Lung Epithelial Cells.

Vitamin C is well documented to have antiviral functions; however, there is limited information about its effect on airway epithelial cells—the first cells to encounter infections. Here, we examined the effect of vitamin C on human bronchial epithelium transformed with Ad12-SV40 2B (BEAS-2B) cells, and observed that sodium-dependent vitamin C transporter 2 (SVCT2) was the primary vitamin C transporter. Transcriptomic analysis revealed that treating BEAS-2B cells with vitamin C led to a significant upregulation of several metabolic pathways and interferon-stimulated genes (ISGs) along with a downregulation of pathways involved in lung injury and inflammation. Remarkably, vitamin C also enhanced the expression of the viral-sensing receptors retinoic acid-inducible gene 1 (RIG-1) and melanoma differentiation-associated protein 5 (MDA-5), which was confirmed at the protein and functional levels. In addition, the lungs of l-gulono-γ-lactone oxidase knockout (GULO-KO) mice also displayed a marked decrease in these genes compared to wild-type controls. Collectively, our findings indicate that vitamin C acts at multiple levels to exert its antiviral and protective functions in the lungs.

Effect of Lipopolysaccharide and TNFα on Neuronal Ascorbic Acid Uptake.

Vitamin C (ascorbic acid: AA) uptake in neurons occurs via the sodium-dependent vitamin C transporter-2 (SVCT2), which is highly expressed in the central nervous system (CNS). During chronic neuroinflammation or infection, CNS levels of lipopolysaccharide (LPS) and LPS-induced tumor necrosis factor-α (TNFα) are increased. Elevated levels of LPS and TNFα have been associated with neurodegenerative diseases together with reduced levels of AA. However, little is known about the impacts of LPS and TNFα on neuronal AA uptake. The objective of this study was to examine the effect of LPS and TNFα on SVCT2 expression and function using in vitro and in vivo approaches. Treatment of SH-SY5Y cells with either LPS or TNFα inhibited AA uptake. This reduced uptake was associated with a significant decrease in SVCT2 protein and mRNA levels. In vivo exposure to LPS or TNFα also decreased SVCT2 protein and mRNA levels in mouse brains. Both LPS and TNFα decreased SLC23A2 promoter activity. Further, the inhibitory effect of LPS on a minimal SLC23A2 promoter was attenuated when either the binding site for the transcription factor Sp1 was mutated or cells were treated with the NF-κB inhibitor, celastrol. We conclude that inflammatory signals suppress AA uptake by impairing SLC23A2 transcription through opposing regulation of Sp1 and NF-κB factors.

Alterations on vitamin C synthesis and transportation and egg deposition induced by dietary vitamin C supplementation in Hy-Line Brown layer model.

In ovo feeding of vitamin C (VC) has positive effects on the growth performance, immune and antioxidant function in poultry, which indicates that increasing VC content in eggs may be of benefit. This study was to investigate the effects of dietary VC supplementation on VC synthesis and transportation and egg deposition. In Exp. 1, in order to select a suitable animal model, VC content was detected in different eggs from different layer species. Vitamin C content was lower in ISA Brown breeder eggs and Hy-Line Brown layer eggs (P < 0.05) then in Arbor Acres breeder eggs. In Exp. 2, a total of 24 Hy-Line Brown layers (42-week-old) were randomly divided into 3 treatments with 8 replicates and fed a basal diet with VC at 0, 200 and 400 mg/kg. Sodium-dependent VC transporter 1 and 2 (SVCT1 and SVCT2) expressions were higher in ileum than in duodenum and jejunum (P < 0.05). SVCT1 expression was higher but SVCT2 expression was lower in the magnum than in the ovary (P < 0.05). L-Gulonolactone oxidase (GLO) and SVCT1 expressions were higher but SVCT2 was lower in the kidney than in the liver (P < 0.05). Dietary VC supplementation at 400 mg/kg increased SVCT1 expression in duodenum, ovary and magnum, but decreased GLO and SVCT1 expression in liver (P < 0.05). Dietary VC supplementation at 200 and 400 mg/kg increased SVCT2 expression in duodenum, but decreased GLO and SVCT1 expression in kidney and SVCT2 expression in liver (P < 0.05). Dietary VC supplementation promoted VC absorption in duodenum and jejunum, but reduced endogenous VC synthesis in liver and kidney. Although dietary VC supplementation enhanced VC transportation in ovary and magnum, it did not increase VC deposition in produced eggs.

Mutation SVCT2 promotes cell proliferation, invasion and migration in colorectal cancer.

The sodium-dependent vitamin C transporter 2 (SVCT2) surface glycoprotein regulates ascorbate accumulation in the plasma, often resulting in the induction of cancer cell death. Therefore, high expression of this gene associates with increased overall survival in several cancers. However, in colorectal cancer (CRC), high (likely mutated) SVCT2 expression relates to poor overall survival, and its functional significance has not been studied. Thus, we hypothesize that mutant SVCT2 expression could affect CRC patient survival. According to biological databases, SVCT2 has been found to be mutated frequently, and SVCT2 E264K has a particularly high pathogenic score (0.98), compared to other SVCT2 mutant sites, in CRC patients. Interestingly, our results reveal expression of SVCT2 E264K in many CRC tissues and cells. Also, we found wild-type SVCT2 expression to be largely localized to the cytoplasm and membrane, while SVCT2 E264K was restricted to the cytoplasm. We further found that SVCT2 E264K overexpression increases cell growth. By contrast, SVCT2 E264K knockdown significantly reduced cell proliferation and promoted cell apoptosis, resulting in inhibition of cell invasion and migration. Taken together, SVCT2 E264K plays a critical role in proliferation in CRC. Our results suggest that SVCT2 E264K could be a promising novel therapeutic target in CRC.