Original Contribution
Low vitamin C and increased oxidative stress and cell death in mice that lack the sodium-dependent vitamin C transporter SVCT2

https://doi.org/10.1016/j.freeradbiomed.2010.06.008Get rights and content

Abstract

The sodium-dependent vitamin C transporter (SVCT2) is responsible for the transport of vitamin C into cells in multiple organs, from either the blood or the cerebrospinal fluid. Mice null for SVCT2 (SVCT2−/−) do not survive past birth but the cause of death has not yet been ascertained. After mating of SVCT2+/− males and SVCT2+/− females, fewer SVCT2−/− and SVCT2+/− progeny were observed than would be expected according to Mendelian ratios. Vitamin C levels in SVCT2−/−, SVCT2+/−, and SVCT2+/+ were genotype-dependent. SVCT2−/− fetuses had significantly lower vitamin C levels than littermates in placenta, cortex, and lung, but not in liver (the site of vitamin C synthesis). Low vitamin C levels in placenta and cortex were associated with elevations in several markers of oxidative stress: malondialdehyde, isoketals, F2-isoprostanes, and F4-neuroprostanes. Oxidative stress was not elevated in fetal SVCT2−/− lung tissue despite low vitamin C levels. In addition to the expected severe hemorrhage in cortex, we also found hemorrhage in the brain stem, which was accompanied by cell loss. We found evidence of increased apoptosis in SVCT2−/− mice and disruption of the basement membrane in fetal brain. Together these data show that SVCT2 is critical for maintaining vitamin C levels in fetal and placental tissues and that the lack of SVCT2, and the resulting low vitamin C levels, results in fetal death and, in SVCT2−/− mice that survive the gestation period, in oxidative stress and cell death.

Section snippets

SVCT2+/− mice

These mice were provided by Dr. Robert Nussbaum. Originally on the 129/SvEvTac background, they were backcrossed more than 10 generations to C57BL/6 mice to place them on this background. Animals were housed in breeding pairs in tub cages in a temperature- and humidity-controlled vivarium. Mice were kept on a 12:12-h light:dark cycle with lights on at 6:00 AM. Mice had free access to food and water for the duration of the experiment. Deionized water was supplemented with 0.33 g/L ascorbic acid

Genotype distribution among the litters

Using data from nine litters the genotype ratios were 12 (−/−):34 (+/−):31 (+/+). A χ2 test against the expected 1:2:1 ratio distribution was significant (χ2 = 10.428, p < 0.01, df = 2). Given 31 SVCT2+/+ mice, if Mendelian ratios were followed, then we would expect 31 SVCT2−/− and 62 SVCT2+/− mice. Separate χ2 tests revealed that both of these genotypes were underrepresented in the sample (χ2 > 11.64, p < 0.001).

Placenta

A gene-dosage effect was seen in VC levels such that for each additional copy of the SVCT2 a

Discussion

In contrast to a previous report [6], we found fewer than expected SVCT2−/− and SVCT2+/− mice in litters delivered by cesarean section, indicating that a smaller proportion of these fetuses survive until birth. This result suggests that lack of SVCT2, and therefore lack of VC in several major organs, contributes to fetal death. In both this and the earlier study [6] SVCT2+/− dams were supplemented with VC in the drinking water; however, the amounts given varied. In this study mice received 0.33 g

Acknowledgments

This work was supported by NIH Grant NS057674-03 to J.M.M. and NIH Grant DK59637 to the Lipid, Lipoprotein, and Atherosclerosis Core of the Vanderbilt Mouse Metabolic Phenotype Centers. The authors thank Lewis F. Pennock, Jr., for the photography of fetal brains.

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