Cystine/Glutamate Antiporter (xCT) Is Required for Chief Cell Plasticity After Gastric Injury

Abstract

Background & Aims Many differentiated epithelial cell types are able to reprogram in response to tissue damage. Although reprogramming represents an important physiological response to injury, the regulation of cellular plasticity is not well understood. Damage to the gastric epithelium initiates reprogramming of zymogenic chief cells into a metaplastic cell lineage known as spasmolytic polypeptide-expressing metaplasia (SPEM). The present study seeks to identify the role of xCT, a cystine/glutamate antiporter, in chief cell reprogramming after gastric injury. We hypothesize that xCT-dependent reactive oxygen species (ROS) detoxification is required for the reprogramming of chief cells into SPEM. Methods Sulfasalazine (an xCT inhibitor) and small interfering RNA knockdown were used to target xCT on metaplastic cells in vitro. Sulfasalazine-treated wild-type mice and xCT knockout mice were analyzed. L635 or DMP-777 treatment was used to chemically induce acute gastric damage. The anti-inflammatory metabolites of sulfasalazine (sulfapyridine and mesalazine) were used as controls. Normal gastric lineages, metaplastic markers, autophagy, proliferation, xCT activity, ROS, and apoptosis were assessed. Results xCT was up-regulated early as chief cells transitioned into SPEM. Inhibition of xCT or small interfering RNA knockdown blocked cystine uptake and decreased glutathione production by metaplastic cells and prevented ROS detoxification and proliferation. Moreover, xCT activity was required for chief cell reprogramming into SPEM after gastric injury in vivo. Chief cells from xCT-deficient mice showed decreased autophagy, mucus granule formation and proliferation, as well as increased levels of ROS and apoptosis compared with wild-type mice. On the other hand, the anti-inflammatory metabolites of sulfasalazine did not affect SPEM development. Conclusions The results presented here suggest that maintaining redox balance is crucial for progression through the reprogramming process and that xCT-mediated cystine uptake is required for chief cell plasticity and ROS detoxification.

Decrease in MiR-148a Expression During Initiation of Chief Cell Transdifferentiation

“…The expression of miR-148a was significantly down-regulated in ImChief cells treated with the inhibitor (Figure 5 B ). Known chief cell and SPEM cell marker expression was examined by quantitative reverse transcription polymerase chain reaction (PCR), and ImChief cells treated with miR-148a inhibitor showed significant up-regulation of the variant 9 splice isoform of Cd44 ( Cd44v9 ), an early marker of SPEM,18, 19 although Tff2 expression was not detected (Figure 5C). The expression of Sox9 and Clusterin ( Clu ), other SPEM-associated markers, was increased, but not significantly (Figure 5 C ).…”

“…Chief cell transdifferentiation occurs through coordinated stepwise events, and the process of transdifferentiation can be arrested at different steps. Our group recently showed that inhibition of the cystine transporter xCT by sulfasalazine treatment or xCT knockout arrests chief cell transdifferentiation process and prevents development of SPEM in mouse models of acute oxyntic atrophy 19 . It was also shown that xCT deficiency blocks chief cell transdifferentiation process at a distinct step where the initiating step of Mist1 loss in chief cell reprogramming is not affected, but up-regulation of CD44v9 and autophagy that occur during chief cell transdifferentiation are blocked.…”

“…Acute oxyntic atrophy induces an early loss of the chief cell maturation-specifying transcription factor, Mist1,15, 16, 17 and up-regulation of the specific splice variant of CD44, CD44 variant 9 (CD44v9) 17, 18, 19. CD44v9 is an activator and a stabilizer of xCT, a cystine transporter, which promotes adaptation to reactive oxygen species and cellular stress 18, 19, 20. The increased cellular stress is associated with an increase in autophagy, which is necessary for breaking down the zymogen granules 21 .…”

“…The increased cellular stress is associated with an increase in autophagy, which is necessary for breaking down the zymogen granules 21 . Importantly, the process of transdifferentiation can be arrested at different stages,17, 19, 21 suggesting that chief cell transdifferentiation occurs through a set of stepwise events that are coordinated and maintained in a defined order.…”

Coping With the Stress of Metaplasia

“…In this issue, 2 groups seek to elucidate some of the metabolic changes that characterize the progression to metaplasia. On one hand, Meyer et al 1 highlight the zymogenic chief cell’s capacity to handle intracellular reactive oxygen species (ROS) as a crucial licensing event during metaplastic reprogramming. Various independent lines of evidence had pointed to the reprogramming of chief cells into a population of proliferative, metaplastic cells as driving epithelial regeneration following glandular injury 2, 3, 4.…”