Inhibition of intestinal ascorbic acid uptake by lipopolysaccharide is mediated via transcriptional mechanisms

Abstract

Ascorbic acid (AA) accumulation in intestinal epithelial cells is an active transport process mainly mediated by two sodium-dependent vitamin C transporters (SVCT-1 and SVCT-2). To date, little is known about the effect of gut microbiota generated lipopolysaccharide (LPS) on intestinal absorption of water-soluble vitamins. Therefore, the objective of this study was to investigate the effects of bacterially-derived LPS on AA homeostasis in enterocytes using Caco-2 cells, mouse intestine and intestinal enteroids models. Pre-treating Caco-2 cells and mice with LPS led to a significant decrease in carrier-mediated AA uptake. This inhibition was associated with a significant reduction in SVCT-1 and SVCT-2 protein, mRNA, and hnRNA expression. Furthermore, pre-treating enteroids with LPS also led to a marked decrease in SVCT-1 and SVCT-2 protein and mRNA expression. Inhibition of SVCT-1 and SVCT-2 occurred at least in part at the transcriptional level as promoter activity of SLC23A1 and SLC23A2 was attenuated following LPS treatment. Subsequently, we examined the protein and mRNA expression levels of HNF1α and Sp1 transcription factors, which are needed for basal SLC23A1 and SLC23A2 promoter activity, and found that they were significantly decreased in the LPS treated Caco-2 cells and mouse jejunum; this was reflected on level of the observed reduction in the interaction of these transcription factors with their respective promoters in Caco-2 cells treated with LPS. Our findings indicate that LPS inhibits intestinal carrier- mediated AA uptake by down regulating the expression of both vitamin C transporters and transcriptional regulation of SLC23A1 and SLC23A2 genes.

Liver Injury, Endotoxemia, and Their Relationship to Intestinal Microbiota Composition in Alcohol‐Preferring Rats

“…Thus, EtOH exposure can possibly enhance lipid synthesis via acetaldehyde and then acetate, and concomitant lipid accumulation possibly results from a deficit of vitamins, such as thiamine (vitamin B 1 ), riboflavin (vitamin B 2 ), and pantothenic acid (vitamin B 5 ). Vitamin deficiencies are commonly found in patients with AUD (Chen and Schnabl, ; Said, ), and chronic EtOH consumption inhibits carrier‐mediated intestinal vitamin uptake, possibly through IM‐produced LPS (Lakhan and Said, ; Subramanian et al., ). These findings suggest that pathological changes associated with chronic EtOH consumption (e.g., inflammation and liver pathology) may be, in part, a consequence of changes in the IM.…”

Tumor necrosis factor alpha reduces intestinal vitamin C uptake: a role for NF-κB-mediated signaling

“…In vitro and in vivo 14 C-AA uptake analysis. For in vitro studies, the post-confluent monolayers of Caco-2 cells were treated with either 20 ng/ml of TNF-␣ or TNF-␣ plus 100 nM celastrol (40) to perform 14 C-AA uptake (44,46). After 48 h, cells were incubated for 30 min in Krebs-Ringer (K-R) buffer at 37°C in the presence of 32 M 14 C-AA and prepared to determine the radioactivity content as described before (44,46).…”

“…For in vitro studies, the post-confluent monolayers of Caco-2 cells were treated with either 20 ng/ml of TNF-␣ or TNF-␣ plus 100 nM celastrol (40) to perform 14 C-AA uptake (44,46). After 48 h, cells were incubated for 30 min in Krebs-Ringer (K-R) buffer at 37°C in the presence of 32 M 14 C-AA and prepared to determine the radioactivity content as described before (44,46). For in vivo studies, we used the adult Gulo knockout (KO) mice, which cannot synthesize vitamin C endogenously like humans because of a lack of L-gulono-␥-lactone oxidase (Gulo), an enzyme involved in the terminal step of L-ascorbic acid biosynthesis, and 33 mg of AA was provided in 100 ml of drinking water to meet the mice body requirement.…”

“…Although the mechanisms responsible for this dysregulation remain to be defined, the effects of proinflammatory cytokines on the AA absorptive processes can play a role. Notably, intestinal inflammation and systemic bacterial infection are associated with an increase in the level of tumor necrosis factor-␣ (TNF-␣) in the intestinal mucosa and blood (22,42,46,50). TNF-␣ is one of the important secreted cytokines, and it plays key roles in initiating and propagating the inflammatory response in the host (4,42).…”

How to Determine the Role of the Microbiome in Drug Disposition

“…The parameters governing absorption typically lie in the chemical properties of the drug itself and how the host may actively import, or export, the molecule. Early microbiome studies hinted that drug transport may be influenced by the microbiome in a composition dependent manner (Hooper et al, 2001), and multiple animal studies have demonstrated that the microbiome modulates drug absorption (J-K Kim et al, 2018;Subramanian et al, 2018). In this section we will highlight methods for examining the influence of microbes on drug transport.…”