Multidrug Resistance Protein 2 Expression Research Articles

Consecutive baicalin treatment relieves its accumulation in rats with intrahepatic cholestasis by increasing MRP2 expression

Baicalin, an important flavonoid isolated from Scutellaria baicalensis Georgi, is a Chinese herb widely used in clinical practice. We previously reported the in vivo accumulation of baicalin in rats with intrahepatic cholestasis (IHC) after a single dose. However, the effects of the long-term administration of baicalin on its pharmacokinetics are unknown. Thus, we investigated the disposition of baicalin in normal rats and those with IHC after single and multiple consecutive administrations. In addition, we further investigated the effect of baicalin on multidrug resistance protein 2 (MRP2) in vivo to explore the underlying mechanism. In our study, the liquid chromatography-mass spectrometry (LC-MS) method established to determine baicalin concentrations in rat blood was simple, specific, and with linearity (R2 = 0.9980) in the range of 1.01–506.00 μg/mL. The relative standard deviations (RSD) for intra-day and inter-day precision were not more than 10.55%, and the intra-day and inter-day accuracies were 94.94%–109.13%. The recovery rate and stability were in line with the requirements of the quantitative analysis of biological samples as stated in the Chinese Pharmacopoeia (2020 Edition). Compared with that in normal rats, the Cmax and t1/2 increased significantly in EE-induced rats with IHC, whereas the clearance (CL) decreased after a single administration of baicalin. However, the area under the curve decreased, CL increased, and the t1/2 was shortened after the continuous administration of baicalin in the IHC rat model compared with the single administration of baicalin, and the pharmacokinetic characteristics were similar to those in normal rats. Moreover, MRP2 expression increased in rats with IHC with the continuous administration of baicalin. Continuous baicalin intervention could effectively reduce its accumulation in rats with IHC, and the mechanism may be attributed to its enhancement of MRP2 expression.

Sulforaphane has an additive anticancer effect to FOLFOX in highly metastatic human colon carcinoma cells.

Colorectal cancer (CRC) is one of the most common malignancies worldwide. Patients with CRC may need chemotherapy (CTx) in a neoadjuvant, adjuvant or palliative setting through the course of the disease. Unfortunately, its effect is limited by chemoresistance and chemotoxicity. Novel more effective and non‑toxic CTx regimens are needed to further improve CRC treatment outcomes. Thus, the present study was designed to test the hypothesis that non‑toxic sulforaphane (SF) is effective against CRC and has additive effects in combination with conventional 5‑fluorouracil, oxaliplatin and folinic acid (FOLFOX) CTx invitro. Highly metastatic human colon cancer cells, CX‑1, and fibroblasts were treated with FOLFOX ± SF. Cell viability was assessed using an MTT assay. The level of apoptosis and the expression of apoptotic proteins were measured by TUNEL assay and quantitative PCR analysis. Aldehyde dehydrogenase isoform 1 (ALDH1) and multidrug resistance protein 2 (MRP2) levels were evaluated. The ability of cells to form spheroids was measured in three‑dimensional cell culture. SF alone and in combination with FOLFOX effectively decreased the viability of the CX‑1 cells, promoted apoptosis within the CX‑1 cells, prevented cellular spheroid formation and decreased ALDH1 activity. However, SF promoted MRP2 expression and protein levels. In conclusion, SF together with conventional FOLFOX has additive anticancer effects against highly metastatic human CRC invitro.

Cigarette smoke extract combined with LPS down-regulates the expression of MRP2 in chronic pulmonary inflammation may be related to FXR

The transporter multidrug resistance protein 2 (MRP2) plays an important role in chronic pulmonary inflammation by transporting cigarette smoke and other related inflammatory mediators. However, it is not completely clear whether pulmonary inflammation caused by cigarette smoke extract (CSE) and lipopolysaccharide (LPS) is related to MRP2 and its signal factors. In this study, CSE combined with LPS was used to establish an inflammation model in vivo and in vitro. We found that compared with the control group, after CSE combined with LPS treatment, the expression of MRP2 in rat lung tissue in vivo and human alveolar cell line in vitro was down-regulated, while the expression of inflammatory factors was up-regulated. Through silencing and overexpression of FXR, it was found that silent FXR could down-regulate MRP2 and up-regulate the expression of inflammatory factors. On the contrary, overexpression of FXR could up-regulate MRP2 and down-regulate the expression of inflammatory factors. Our results show that CSE combined with LPS can down-regulate the expression of MRP2 under inflammatory conditions, and the down-regulation of MRP2 expression may be achieved partly through the FXR signal pathway.

The changes of MRP2 expression in three kinds of pulmonary inflammation models: the downregulation occurred in cigarette smoke extract (CSE) stimulation group and CSE plus LPS stimulation group, unchanged in LPS stimulation group

The transporter multidrug resistance protein 2 (MRP2) can transport some tobacco carcinogens and plays an important role in the transport of mediators related to pulmonary inflammatory diseases. However, it is not fully understood whether the pulmonary inflammation caused by cigarette smoke extract (CSE) and lipopolysaccharide (LPS) is related to the regulation of MRP2. In this study, CSE and LPS were used alone and in combination as stimuli to induce pulmonary inflammation. In addition, the establishment of a pulmonary inflammation model was verified by animal experiments in vivo. We found that compared with those in the control group, the expression of MRP2 protein was downregulated and the expression of inflammatory cytokines was upregulated in pulmonary inflammation in the CSE group and the CSE combined with LPS group. However, there was almost no change in the expression of MRP2 stimulated by LPS alone. Our results show that CSE and CSE combined with LPS downregulate the expression of MRP2 under inflammatory conditions, while LPS has almost no effect on the expression of MRP2 under inflammatory conditions. The in vivo experimental results of CSE combined with LPS were consistent with the cellular results of CSE combined with LPS, which provides a model and basis for other studies of the role of MRP2 in pulmonary inflammation.

Uptake of gadoxetic acid in hepatobiliary phase magnetic resonance imaging and transporter expression in hypovascular hepatocellular nodules

AimsTo evaluate the association between contrast patterns on gadoxetic acid-enhanced hepatobiliary phase (HBP) MR images and transporter expression in surgically resected hypovascular hepatocellular nodules including early hepatocellular carcinomas (HCCs). MethodsForty-two hypovascular hepatic nodules and 43 hypervascular HCCs as a control were included in this retrospective study. Contrast of the nodules on HBP images was graded as hypo-, iso-, or hyperintense. Histopathological assessment was performed in the context of multistep hepatocarcinogenesis. Immunohistochemical staining of organic anion transporter 1B3 (OATP1B3) and multidrug resistance protein 2 (MRP2) was performed. Cramer’s coefficient was used to determine the linear relationship between contrast grades and transporter expression, and the Cochran-Armitage trend test was used to determine the relationship between transporter expression and progression of multistep hepatocarcinogenesis. ResultsModerate linear relationships between contrast grades and OATP1B3 expression were observed for both hypo- and hypervascular nodules. OATP1B3 expression was negatively correlated with the progression of multistep hepatocarcinogenesis. MRP2 expression was not associated with the contrast grades or histopathological results. ConclusionOATP1B3 expression was associated with contrast grades of hepatocellular nodules observed in HBP image of gadoxetic acid-enhanced MRI in the hypovascular hepatocellular nodules and was negatively correlated with hepatocarcinogenesis.

Multi-drug resistance protein 2 (MRP2) expression, adjuvant tamoxifen therapy, and risk of breast cancer recurrence: a Danish population-based nested case-control study

Background: Adjuvant tamoxifen therapy approximately halves the risk of recurrence in estrogen receptor-positive (ER+) breast cancer patients, but many women respond insufficiently to therapy. Expression of multi-drug resistance protein 2 (MRP2) in breast cancer may potentiate tamoxifen resistance. Thus, we investigated the expression of MRP2 in breast cancer as a predictor of tamoxifen therapy effectiveness.Material and methods: We conducted a case-control study nested in the Danish Breast Cancer Group clinical database. The study included women aged 35–69 years diagnosed with stage l–lll breast cancer during 1985–2001, in Jutland, Denmark. We identified 541 recurrent breast cancers (cases) among women with estrogen receptor positive (ER+) disease treated with tamoxifen for at least 1 year (ER+/TAM+) and 300 cases among women with estrogen receptor-negative (ER−) disease, never treated with tamoxifen (ER−/TAM−). We matched one recurrence-free control to each recurrent case. We retrieved paraffin-embedded primary tumor tissue for all patients, and all available recurrent tumor tissue from pathology archives. MRP2 expression was evaluated using immunohistochemistry. We computed odds ratios (ORs) and 95% confidence intervals (95% CIs) associating MRP2 expression (positive vs. none) with breast cancer recurrence in conditional logistic regression models. We compared MRP2 expression in paired primary- and recurrent tumors.Results: MRP2 expression was more prevalent in the ER+/TAM + group, than in the ER−/TAM − group. No predictive utility of MRP2 for breast cancer recurrence was found in the ER+/TAM + group (ORadj = 0.96, 95% CI 0.70, 1.33). Further, no prognostic utility was found in the ER−/TAM − group (ORadj = 0.81, 95% CI 0.53, 1.23). MRP2 expression was not increased in recurrent versus primary tumors.Conclusions: MRP2 expression is neither a predictive marker of tamoxifen effectiveness nor a prognostic marker in breast cancer.

Dihydromyricetin reverses MRP2-mediated MDR and enhances anticancer activity induced by oxaliplatin in colorectal cancer cells.

Dihydromyricetin (DMY), extracted from the Chinese herbal medicine Ampelopsis grossedentata, possesses antitumor potential in different types of human cancer cells. Hence, its effects on drug resistance and molecular mechanisms in colorectal cancer (CRC) are still unknown. In our present study, we observed that DMY enhanced the chemosensitivity to oxaliplatin (OXA). DMY increased OXA-induced apoptosis and reduced 5(6)-carboxy-2',7'-dichlorofluorescein accumulation in OXA-resistant CRC HCT116/L-OHP cells. Our mechanistic study suggested that DMY treatment inhibited multidrug resistance protein 2 (MRP2) expression levels and promoter activity, indicating that DMY reduced not only MRP2 transcriptional and translational levels but also its function. Additional experiments indicated that the nuclear translocation of nuclear factor-erythroid 2 p45 related factor 2, a MRP2 regulator, was also inhibited by DMY. In summary, our study provided the first direct evidence that the inhibitory effects of DMY on MRP2 expression in OXA-resistant CRC cells were closely associated with the inhibition of nuclear factor-erythroid 2 p45 related factor 2 signaling. DMY could be a potential candidate for CRC chemotherapy.

SIRT1-Mediated FoxO1 Deacetylation Is Essential for Multidrug Resistance-Associated Protein 2 Expression in Tamoxifen-Resistant Breast Cancer Cells

Our previous studies have shown that multidrug resistance protein 2 (MRP2) is overexpressed in tamoxifen-resistant MCF-7 breast cancer cells (TAMR-MCF-7 cells) and forkhead box-containing protein, O subfamily1 (FoxO1), functions as a key regulator of multidrug resistance 1 (MDR1) gene transcription. This study aimed to investigate the role of FoxO1 in regulating MRP2 gene expression in TAMR-MCF-7 cells. The proximal promoter region of the human MRP2 gene contains four putative FoxO binding sites, and MRP2 gene transcription was stimulated by FoxO1 overexpression in MCF-7 cells. Subcellular fractionation and immunoblot analyses revealed that basal MRP2 expression and nuclear levels of FoxO1 were enhanced in TAMR-MCF-7 cells compared to MCF-7 cells and the enhanced MRP2 gene transcription was suppressed by FoxO1 siRNA. Because nuclear localization of FoxO1 is regulated by SIRT1 deacetylase, we were further interested in whether SIRT1 is involved in MRP2 expression. Overexpression of SIRT1 with FoxO1 potentiated the gene transcriptional activity of MRP2, and the basal activity and expression of SIRT1 was increased in TAMR-MCF-7 cells. In addition, SIRT1 inhibition reduced both the nuclear FoxO1 levels and MRP2 expression and enhanced cytotoxic effects of paclitaxel and doxorubicin in TAMR-MCF-7 cells. These results suggest that FoxO1 activation via SIRT1-mediated deacetylation is closely related with up-regulation of MRP2 in TAMR-MCF-7 cells.

THU0255 The Efficacy of Dmards Combination Therapy Using MTX and Low Dose Tacrolimus Depend on Modulation of MRP2 Expression.

BackgroundMethotrexate (MTX) is an anchor drug for the treatment of rheumatoid arthritis (RA) because of efficacy and safety. Although MTX is used in monotherapy generally, combination therapy with biologics or...

In Vitro and In Vivo Prostate Cancer Metastasis and Chemoresistance Can Be Modulated by Expression of either CD44 or CD147

CD44 and CD147 are associated with cancer metastasis and progression. Our purpose in the study was to investigate the effects of down-regulation of CD44 or CD147 on the metastatic ability of prostate cancer (CaP) cells, their docetaxel (DTX) responsiveness and potential mechanisms involved in vitro and in vivo. CD44 and CD147 were knocked down (KD) in PC-3M-luc CaP cells using short hairpin RNA (shRNA). Expression of CD44, CD147, MRP2 (multi-drug resistance protein-2) and MCT4 (monocarboxylate tranporter-4) was evaluated using immunofluorescence and Western blotting. The DTX dose-response and proliferation was measured by MTT and colony assays, respectively. The invasive potential was assessed using a matrigel chamber assay. Signal transduction proteins in PI3K/Akt and MAPK/Erk pathways were assessed by Western blotting. An in vivo subcutaneous (s.c.) xenograft model was established to assess CaP tumorigenecity, lymph node metastases and DTX response. Our results indicated that KD of CD44 or CD147 decreased MCT4 and MRP2 expression, reduced CaP proliferation and invasive potential and enhanced DTX sensitivity; and KD of CD44 or CD147 down-regulated p-Akt and p-Erk, the main signal modulators associated with cell growth and survival. In vivo, CD44 or CD147-KD PC-3M-luc xenografts displayed suppressed tumor growth with increased DTX responsiveness compared to control xenografts. Both CD44 and CD147 enhance metastatic capacity and chemoresistance of CaP cells, potentially mediated by activation of the PI3K and MAPK pathways. Selective targeting of CD44/CD147 alone or combined with DTX may limit CaP metastasis and increase chemosensitivity, with promise for future CaP treatment.

Hepatic expression of multidrug resistance protein 2 in biliary atresia

BackgroundBiliary atresia (BA) is an idiopathic inflammatory obliterative cholangiopathy of neonates, leading to progressive biliary cirrhosis. Hepatoportoenterostomy (Kasai procedure) can cure jaundice in 30% to 80% of patients. Postoperative clearance of jaundice is one of the most important factors influencing long-term outcomes of BA patients. Multidrug resistance protein 2 (MRP2) is one of the canalicular export pumps located in hepatocytes; it exports organic anions and their conjugates (e.g., bilirubin) into bile canaliculus. Although MRP2 is an essential transporter for the excretion of bilirubin, its role in the clinical course of BA patients is unclear. The present study investigated the relationship between hepatic MRP2 expression and clinical course in BA patients, with particular emphasis in curing jaundice after hepatoportoenterostomy.ResultsNo significant differences in hepatic MRP2 expression level were observed between BA and controls groups. There was no correlation between MRP2 expression and age at time of surgery in BA and control groups. In BA patients, MRP2 expression level in the jaundice and jaundice-free group did not differ significantly (2.0 × 10-4 vs 3.1 × 10-4, p = 0.094). Although the serum level of total bilirubin just before surgery did not correlate with MRP2 expression level (rs = 0.031, p = 0.914), the serum level of total bilirubin measured at 2 weeks (rs = -0.569, p = 0.034) and 4 weeks after surgery (rs = -0.620, p = 0.018) were significantly correlated with MRP2 expression level. Furthermore, MRP2 expression level was inversely correlated with ratio of change in serum total bilirubin level over 4 weeks (rs = -0.676, p = 0.008), which represents the serum bilirubin level measured at 4 weeks after surgery divided by value just before surgery. There was no correlation between expression level of MRP2 and nuclear receptors, such as retinoid × receptor α, farnesoid × receptor, pregnane × receptor, or constitutive androstane receptor.ConclusionsHepatic MRP2 expression level was associated with postoperative clearance of jaundice in BA patients, at least within 1 month after hepatoportoenterostomy. This finding suggests that not only morphological appearance of the liver tissue but also the biological status of hepatocytes is important for BA pathophysiology.

Resveratrol inhibits genistein-induced multi-drug resistance protein 2 (MRP2) expression in HepG2 cells

The interactions between various dietary cancer chemopreventive phytochemicals in drug transporter functions are not well studied. In this study, the effects of genistein and resveratrol on the multidrug resistance protein 2 (MRP2) expression and the underlying molecular mechanisms were investigated using HepG2-C3 cells that are stably transfected with a construct containing human MRP2 promoter region conjugated with luciferase reporter gene. A 3-fold induction of MRP2 luciferase activity was observed after genistein (50μM) treatment to HepG2-C3 cells, but was diminished by the resveratrol (50μM) cotreatment. This observation was further validated by Western blot analysis and RT-PCR analysis as resveratrol also inhibited genistein-induced MRP2 protein synthesis and mRNA expression. Immunofluorescence study revealed that genistein-induced formation of MRP2 vacuoles was dramatically reduced by resveratrol. The binding affinity between retinoid X receptor alpha (RXRα) and MRP2 promoter was examined by DNA–protein pull-down assay. The results showed that resveratrol inhibited the genistein-induced binding of RXRα to the promoter sequence of MRP2 gene, and this mechanism could potentially contribute to the inhibition of genistein-induced MRP2 expression by resveratrol. Taken together, our present study suggests that naturally occurring phytochemicals can potentially interfere with each other’s regulatory function on the cancer chemoprevention-related genes through a competitive mechanism.

Role of multidrug resistance protein 2 (MRP2) in chemoresistance and clinical outcome in oesophageal squamous cell carcinoma

Background:Although multidrug resistance protein 2 (MRP2) confers chemoresistance in some cancer types, its implication on oesophageal squamous cell carcinoma (ESCC) remains unclear.Methods:We evaluated MRP2 expression by immunohistochemistry and RT–PCR using 81 resected specimens from ESCC patients who did or did not receive neo-adjuvant chemotherapy (NACT), including 5-fluorouracil, doxorubicin, and cisplatin (CDDP). Correlation between MRP2 expression and response to chemotherapy was also examined in 42 pre-therapeutic biopsy samples and eight ESCC cell lines.Results:MRP2-positive immunostaining was more frequently observed in ESCCs with NACT than in those without NACT (27.3 vs 5.4%). The MRP2-positive patients showed poorer prognosis than MRP2-negative patients (5-year survival rate, 25.6 vs 55.7%). Concordantly, ESCC with NACT showed 2.1-fold higher mRNA expression of MRP2 than those without NACT (P=0.0350). In pre-therapeutic biopsy samples of patients with NACT, non-responders showed 2.9-fold higher mRNA expression of MRP2 than responders (P=0.0035). Among the panel of ESCC cell lines, TE14 showed the highest MRP2 mRNA expression along with the strongest resistance to CDDP. Inhibition of MRP2 expression by small-interfering RNA reduced chemoresistance to CDDP.Conclusion:Our data suggested that MRP2 is one of molecules, which regulate the sensitivity to chemotherapy including CDDP in advanced ESCC patients.

HDAC inhibitors downregulate MRP2 expression in multidrug resistant cancer cells: Implication for chemosensitization

Although histone deacetylase (HDAC) inhibitors are emerging as a promising class of cancer chemotherapeutic agents, their effects on multidrug resistance (MDR) are poorly understood. In this study, we investigated whether HDAC inhibitors overcome MDR phenotype. HDAC inhibitors suppress the growth of both MDR positive cancer cells KBV20C and its parental cells KB with similar potencies. In parallel, histone acetylation and p21WAF1 expression by the HDAC inhibitors were similarly increased in both cell types, indicating that these HDAC inhibitors are poor substrates of ABC drug transporters and effective in MDR cancer cells. In addition, multidrug resistance protein 2 (MRP2) expression is selectively attenuated by HDAC inhibitors, especially SAHA and TSA, in KBV20C cells, whereas MDR1 and BCRP expressions are not affected. This downregulation of MRP2 contributes to increase in paclitaxel-induced G2/M arrest and apoptosis, which might be due to intracellular accumulation of paclitaxel. Collectively, our data provide a molecular rationale for the application of HDAC inhibitors to overcome MDR in cancer cells.

Interaction between CD44 and hyaluronate induces chemoresistance in non-small cell lung cancer cell

CD44s is a principle hyaluronate (HA) receptor and has been reported to play an important role in cancer cell invasion and metastasis. The aim of our study is to determine if the interaction between HA and CD44s influences in vitro chemosensitivity of non-small cell lung cancer (NSCLC). NSCLC cell line, H322 cells, transfected with the CD44s gene (H322/CD44s) cultured on HA coated plates were more resistant to cisplatin (CDDP) than that on bovine serum albumin. Multidrug resistance protein2 (MRP2) expression was induced in H322/CD44s cells cultured on HA. MRP2 inhibitor, MK571, not only suppressed MRP2 expression but also reversed CDDP resistance. These results suggest that the interaction between CD44s and HA play a pivotal role in acquired resistance to CDDP in NSCLC and MRP2 could be involved in this potential mechanism.

Impaired expression of hepatic multidrug resistance protein 2 is associated with posthepatectomy hyperbilirubinemia in patients with biliary cancer

Hyperbilirubinemia is a critical complication following hepatectomy for biliary cancer. Hepatic multidrug resistance protein 2 (MRP2), a bilirubin transporter, is shown to be down-regulated by acute biliary obstruction in rats. However, little is known about the effect of chronic obstruction by malignancy on the MRP2 expression in patients or the association of MRP2 expression with posthepatectomy hyperbilirubinemia. The MRP2 expression before hepatectomy was determined by immunostaining and Western blotting in patients with biliary cancer. To directly determine the effect of chronic bile duct obstruction on the MRP2 expression, the expression levels were compared between the cholestatic and noncholestatic lobes in each of seven patients. In another 39 patients, the correlation of the MRP2 expression of the anticipated remnant liver with the posthepatectomy severe hyperbilirubinemia, defined as a serum total bilirubin concentration>or=200 micromol/l, was evaluated. The MRP2 staining in the cholestatic lobes was weak and not restricted to the canalicular membrane, unlike the noncholestatic lobes. The expression levels in the cholestatic lobes were 45% of those in the noncholestatic lobes. Postoperative maximum bilirubin levels were significantly correlated with MRP2 expression of the anticipated remnant liver. The MRP2 expression had been already impaired before hepatectomy in all patients who eventually developed severe hyperbilirubinemia. Decreased MRP2 expression, caused by biliary obstruction due to cancer, is a possible risk factor for posthepatectomy severe hyperbilirubinemia.

Identification and Functional Analysis of Two Novel Mutations in the Multidrug Resistance Protein 2 Gene in Israeli Patients with Dubin-Johnson Syndrome

Dubin-Johnson syndrome (DJS) is an inherited disorder characterized by conjugated hyperbilirubinemia and is caused by a deficiency of the multidrug resistance protein 2 (MRP2) located in the apical membrane of hepatocytes. The aim of this study was to identify the mutations in two previously characterized clusters of patients with Dubin-Johnson syndrome among Iranian and Moroccan Jews and determine the consequence of the mutations on MRP2 expression and function by expression studies. All 32 exons and adjacent regions of the MRP2 gene were screened by polymerase chain reaction and DNA sequencing. Two novel mutations were identified in exon 25. One mutation, 3517A-->T, predicting a I1173F substitution, was found in 22 homozygous Iranian Jewish DJS patients from 13 unrelated families and a second mutation, 3449G-->A, predicting a R1150H substitution, was found in 5 homozygous Moroccan Jewish DJS patients from 4 unrelated families. Use of four intragenic dimorphisms and haplotype analyses disclosed a specific founder effect for each mutation. The mutations were introduced into an MRP2 expression vector by site-directed mutagenesis, transfected into HEK-293 cells, and analyzed by a fluorescence transport assay, immunoblot, and immunocytochemistry. Continuous measurement of probenecid-sensitive carboxyfluorescein efflux revealed that both mutations impaired the transport activity of MRP2. Immunoblot analysis and immunocytochemistry showed that MRP2 (R1150H) matured properly and localized at the plasma membrane of transfected cells. In contrast, expression of MRP2 (I1173F) was low and mislocated to the endoplasmic reticulum of the transfected cells. These findings provide an explanation for the DJS phenotype in these two patient groups. Furthermore, the close localization of the two mutations identify this region of MRP2 as important for both activity and processing of the protein.

Hepatic transport of bile salts.

The vectorial secretion of bile salts from blood into bile is a major driving force for bile formation. The basolateral hepatocyte membrane extracts bile salts from sinusoidal blood via Na(+)-dependent and Na(+)-independent membrane transporters. Na(+)-dependent uptake of bile salts is mediated by the Na(+)-taurocholate co-transporting polypeptide, a 51-kDa protein that is exclusively expressed in hepatocytes. Na(+)-independent uptake of bile salts is mediated by the organic anion transporting polypeptides, a superfamily of multispecific bile salt and amphipathic substrate transporters. Within the hepatocyte, bile salts are bound to cytosolic proteins and traverse the cell mainly by diffusion. Transport across the canalicular membrane is the rate-limiting step in overall hepatocellular bile salt excretion and is mediated by the bile salt export pump (BSEP), a homologue of the P-glycoproteins or multidrug resistance gene products. BSEP is a vulnerable target for inhibition by estrogen metabolites, drugs such as cyclosporine A, and abnormal bile salt metabolites, all of which can cause retention of bile salts and consequently intrahepatic cholestasis. Canalicular efflux of divalent sulfated or glucuronidated bile salts is mediated by the multidrug resistance protein 2 (MRP2), which is strongly decreased in cholestasis. Decreased MRP2 expression leads to compensatory increases in the basolateral expression of MRP1 and MRP3, which mediate the sinusoidal efflux of divalent bile salt conjugates and other organic anions. Thus, the hepatocyte can regulate expression levels of individual bile salt transporters during cholestasis to evade hepatotoxic injury.

Dexamethasone- and osmolarity-dependent expression of the multidrug-resistance protein 2 in cultured rat hepatocytes1

Expression of the conjugate export pump multidrug-resistance protein 2 (MRP2) in liver is regulated by endotoxin and anti-tumour agents. This paper reports on the effects of dexamethasone and osmolarity on MRP2 expression. MRP2 expression was studied at the protein, mRNA, immunocytochemical and functional levels in cultured rat hepatocytes. Protein and mRNA expression of MRP2 in rat hepatocytes 24 and 48 h after isolation were largely dependent on the presence of dexamethasone (100 nmol/l) in the culture medium. MRP2 was localized at the pseudocanalicular membrane and increased expression of MRP2 was accompanied by a widening of the pseudocanaliculi. In presence of dexamethasone, hypo-osmolarity (205 mosmol/l) led to a strong induction of MRP2 mRNA and protein, whereas expression was decreased by hyperosmolarity (405 mosmol/l). Also, a decay of MRP2 protein and mRNA following dexamethasone withdrawal was osmosensitive. Expression of dipeptidylpeptidase IV, another canalicular protein, was unaffected by dexamethasone and osmolarity. It is concluded that glucocorticoids are strong inducers of MRP2 in liver. Besides short-term carrier insertion/retrieval, osmoregulation of MRP2 also involves a long-term effect on MRP2 expression.

Dexamethasone- and osmolarity-dependent expression of the multidrug-resistance protein 2 in cultured rat hepatocytes1

Expression of the conjugate export pump multidrug-resistance protein 2 (MRP2) in liver is regulated by endotoxin and anti-tumour agents. This paper reports on the effects of dexamethasone and osmolarity on MRP2 expression. MRP2 expression was studied at the protein, mRNA, immunocytochemical and functional levels in cultured rat hepatocytes. Protein and mRNA expression of MRP2 in rat hepatocytes 24 and 48 h after isolation were largely dependent on the presence of dexamethasone (100 nmol/l) in the culture medium. MRP2 was localized at the pseudocanalicular membrane and increased expression of MRP2 was accompanied by a widening of the pseudocanaliculi. In presence of dexamethasone, hypo-osmolarity (205 mosmol/l) led to a strong induction of MRP2 mRNA and protein, whereas expression was decreased by hyperosmolarity (405 mosmol/l). Also, a decay of MRP2 protein and mRNA following dexamethasone withdrawal was osmosensitive. Expression of dipeptidylpeptidase IV, another canalicular protein, was unaffected by dexamethasone and osmolarity. It is concluded that glucocorticoids are strong inducers of MRP2 in liver. Besides short-term carrier insertion/retrieval, osmoregulation of MRP2 also involves a long-term effect on MRP2 expression.