Up-regulation and cytoprotective role of epithelial multidrug resistance-associated protein 1 in inflammatory bowel disease.

MRP1 (multidrug resistance-associated protein 1) is well known for its role in providing multidrug resistance to cancer cells. In addition, MRP1 has been associated with both pro- and anti-inflammatory functions in nonmalignant cells. The pro-inflammatory function is evident from the fact that MRP1 is a high affinity transporter for cysteinyl-leukotriene C4 (LTC4), a lipid mediator of inflammation. It remains unexplained, however, why the absence of Mrp1 leads to increased intestinal epithelial damage in mice treated with dextran-sodium sulfate, a model for inflammatory bowel disease (IBD). We found that MRP1 expression is induced in the inflamed intestine of IBD patients, e.g. Crohn disease and ulcerative colitis. Increased MRP1 expression was detected at the basolateral membrane of intestinal epithelial cells. To study a putative role for MRP1 in protecting epithelial cells against inflammatory cues, we manipulated MRP1 levels in human epithelial DLD-1 cells and exposed these cells to cytokines and anti-Fas. Inhibition of MRP1 (by MK571 or RNA interference) resulted in increased cytokine- and anti-Fas-induced apoptosis of DLD-1 cells. Opposite effects, e.g. protection of DLD-1 cells against cytokine- and anti-Fas-induced apoptosis, were observed after recombinant MRP1 overexpression. Inhibition of LTC4 synthesis reduced anti-Fas-induced apoptosis when MRP1 function was blocked, suggesting that LTC4 is the pro-apoptotic compound exported by epithelial MRP1 during inflammation. These data show that MRP1 protects intestinal epithelial cells against inflammation-induced apoptotic cell death and provides a functional role for MRP1 in the inflamed intestinal epithelium of IBD patients.

Decreased P-glycoprotein (P-gp/MDR1) expression in inflamed human intestinal epithelium is independent of PXR protein levels.

BACKGROUND: Altered P-glycoprotein expression (P-gp/MDR1) and/or function may contribute to the pathogenesis of gastrointestinal inflammatory disorders. Low intestinal mRNA levels of the pregnane X receptor (PXR) have been linked to low MDR1 mRNA levels in patients with ulcerative colitis (UC). Here we compared intestinal MDR1 mRNA and protein expression in uninflamed and inflamed intestinal epithelium (IE) of patients with gastrointestinal inflammatory disorders to healthy controls. METHODS: Intestinal mucosal biopsies were obtained from patients with Crohn's disease (CD, n = 20), UC (n = 10), diverticulitis (n = 3), collagenous colitis (n = 3), and healthy controls (n = 10). MDR1, iNOS, MRP1, CYP3A4, and PXR expression was determined using real-time reverse-transcriptase polymerase chain reaction (RT-PCR), Western blotting, and/or immunohistochemistry. Furthermore, MDR1 expression was determined in human intestinal biopsies and the human colon carcinoma cell line DLD-1 after exposure to cytokines (TNF-alpha, IFN-gamma, and/or IL-1beta). RESULTS: MDR1 mRNA levels in uninflamed colon of UC patients were comparable to healthy controls, while they were slightly decreased in ileum and slightly increased in colon of CD patients. MDR1 expression, however, was strongly decreased in inflamed IE of CD, UC, collagenous colitis, and diverticulitis patients. A cytokine-dependent decrease of MDR1 expression was observed in human intestinal biopsies, but not in DLD-1 cells. Remarkably, PXR protein levels were equal in uninflamed and inflamed tissue of CD and UC patients despite low PXR mRNA levels in inflamed tissue. CONCLUSIONS: MDR1 expression is strongly decreased in inflamed IE of patients with gastrointestinal disorders and this is independent of PXR protein levels. Low MDR1 levels may aggravate intestinal inflammation.

High expression of MDR1, MRP1, and MRP3 in the hepatic progenitor cell compartment and hepatocytes in severe human liver disease.

An increase in bile ductular structures is observed in diverse human liver diseases. These structures harbour the progenitor cell compartment of the liver. Since ATP-binding cassette (ABC) transporters may have a cytoprotective role in liver disease, an immunohistochemical study was performed on human liver specimens from patients with primary biliary cirrhosis (PBC), chronic hepatitis C virus (HCV) infection, submassive cell necrosis, and normal liver. The expression of MDR1, MDR3, BSEP, MRP1, MRP2, and MRP3 was determined using specific antibodies. Dilution series were constructed to determine the critical staining level in order to estimate the factor of up-regulation. In normal liver, hepatocytes showed canalicular staining for MDR3, BSEP, and MRP2. MDR1 stained the canalicular membrane of hepatocytes as well as that of cholangiocytes. MRP3 showed low immunoreactivity of bile duct epithelial cells and centrilobular hepatocytes only. Normal liver showed no immunoreactivity for MRP1. In diseased liver, the expression of MDR3, BSEP, and MRP2 was relatively stable. In PBC, HCV, and submassive necrosis, the expression levels of MDR1, MRP1, and MRP3 were increased. The strongest immunoreactivity was seen after submassive necrosis, where remaining islands of hepatocytes showed strong canalicular staining for MDR1 and MRP3. Regenerating bile ductules at the interface of portal tracts and necrotic areas stained intensely for MDR1, MRP1, and MRP3. In conclusion, MDR1, MRP1, and MRP3 are up-regulated in hepatocytes in severe human liver disease. Strong MDR1, MRP1, and MRP3 reactivity is seen in regenerating human bile ductules.