Accepted: 12 June 2001Published online: 20 July 2001© Springer-Verlag 2001

Abstract Background and aims:Glucocorticoids are potent anti-in-flammatory drugs widely used in thetreatment of inflammatory boweldisease, but many patients do notbenefit from glucocorticoid therapy(glucocorticoid resistance) or needinappropriately high doses to retainremission (glucocorticoid dependen-cy). Because of the role of intestinalepithelial cells in inflammatory bow-el disease we examined glucocorti-coid receptor signaling and the effectof interleukin-1β as one of the mainproinflammatory cytokines in the in-testinal epithelial cell lines IEC-6and Caco-2. Methods: Dexametha-sone effects on transcriptional acti-vation was measured by reportergene assay using a construct contain-ing glucocorticoid-responsive ele-ments. The transrepressive effectwas monitored by a nuclear factor(NF) κB inducible reporter con-struct. In addition in IEC-6 cells im-muncytochemistry was used to mon-itor glucocorticoid receptor translo-cation. Results: Dexamethasone in-duced receptor-mediated reporter

gene transcription and receptortranslocation, while interleukin-1βsignificantly inhibited dexametha-sone effects. Dexamethasone inhibit-ed interleukin-1β induced, NF-κBdriven gene transcription only inIEC-6 and not in Caco-2 cells. How-ever, in Caco-2 cells glucocorticoidreceptor overexpression resulted in amarked decrease in NF-κB activityeven in absence of dexamethasone.Conclusions: These studies demon-strate that glucocorticoid receptordriven gene regulation in intestinalepithelial cells may contribute to theanti-inflammatory effects of gluco-corticoids in inflammatory boweldisease. Our data are consistent withthe notion that interleukin-1β pro-duced during inflammatory responseinduces steroid resistance, which is acommon clinical problem in treatingpatients with inflammatory boweldisease

Keywords Intestinal epithelial cell ·Glucocorticoid receptor ·Interleukin-1β · Chronic inflammatorybowel disease · Nuclear factor κB

Int J Colorectal Dis (2001) 16:377–383DOI 10.1007/s003840100331 O R I G I N A L A RT I C L E

Dirk RaddatzSzilvia TothHarald SchwörerGiuliano Ramadori

Glucocorticoid receptor signalingin the intestinal epithelial cell lines IEC-6and Caco-2: evidence of inhibitionby interleukin-1β

Introduction

The intestinal epithelium has an important role as ab-sorptive tissue allowing the transport of water, electro-lytes, and nutrients. It is also a barrier limiting systemiccontamination by micro-organisms residing in the lu-men of the gut. In the recent years it has become appar-ent that the intestinal epithelium itself plays an active

role in the immune response of the gut. Intestinal epi-thelial cells have several cytokine receptors [1] and pro-duce a number of cyto- and chemokines [2]. In the nor-mal mucosa the immune process is kept in a quiescentstate by accurately balanced mechanisms involving pro-and anti-inflammatory cytokines and chemokines. In-flammation such as in the case of inflammatory boweldisease (IBD) is associated with a disturbed balance of

D. Raddatz · S. Toth · H. SchwörerG. Ramadori (✉ )Department of Gastroenterology andEndocrinology, University of Göttingen,Robert-Koch-Strasse 40, 37075 Göttingen,Germanye-mail: gramado@med.uni-goettingen.deTel.: +49-551-396301Fax: +49-551-398596

these mechanisms and a loss of specific epithelial func-tions.

Glucocorticoids (GC) are the most potent drugs avail-able for the treatment of acute IBD attacks. GC action ismediated on the molecular level by intracellular glucocor-ticoid receptors (GRs). In quiescent state GRs are associ-ated with heat shock proteins which act as chaperones.Upon binding GC this complex dissociates and the acti-vated receptor translocates into the nucleus where it bindsto DNA. GRs regulate gene transcription by at least twomechanisms. First, they induce gene transcription (trans-activation) by binding to specific DNA motifs, so-calledglucocorticoid-responsive elements (GREs). The expres-sion of lipocortin 1, an inhibitor of phospholipase A2 isupregulated by this mechanism, resulting in a decrease inprostaglandin E2 synthesis in a number of cells includingepithelial cells [3]. Secondly, they may repress transcrip-tional activation by interfering with other transcriptionfactors (transrepression). There is evidence that GC sup-press immunity by repressing the transcripional activity ofnuclear factor (NF) κB [4], a transcription factor responsi-ble for the upregulation of many cytokine genes such asthat of interleukin (IL) 8 [5]. Recently Rogler et al. [6]have shown that NF-κB is activated in macrophages andepithelial cells in biopsy specimens taken from patientswith IBD. However, repression of NF-κB activity by GCis not a universal mechanism and seems to depend on thecell type and state of differentiation. The synthetic GCdexamethasone (Dex) does not inhibit NF-κB activationin rat mesangial cells [7]. There are only few data regard-ing the effect of GC upon the intestinal epithelium. GCstabilize epithelial function [8] and restore sodium absorp-tion in chronically inflamed epithelium [9]. Moreover GCsuppress IL-1β mediated monocyte-chemoattractant pro-tein 1 expression [10] and interferon-γ induced nitric ox-ide synthesis [11] in Caco-2 cells. The pleiotropic effectsof GC on morphology and function in the rat intestinalcell line IEC-6, a well established model of small intesti-nal crypt cells, have recently been well studied [12]. Thecolonic carcinoma cell line Caco-2 has been shown topossess characteristics of normal small intestinal epitheli-um [13]. In vitro it expresses a similar array of cytokinesas cultured primary epithelial cells do [14]. For these rea-sons IEC-6 and Caco-2 cells provide model systems tostudy the molecular mechanisms involved in GC actionwithin the intestinal epithelium.

About 20% of patients with IBD do not benefit fromGC therapy (GC resistance) or need very high GC dosesto maintain clinical remission (GC dependency) compli-cated by severe adverse reactions [15]. As alterations inGR function have been reported to play a role in otherinflammatory diseases such as asthma [16] and rheuma-toid arthritis [17], and cytokines may modulate GC sen-sitivity [18], a second aim of this study was to test thehypothesis that IL-1β, a major proinflammatory cyto-kine in IBD, affects GR signaling in intestinal epithelialcells.

Materials and methods

Plasmids

The plasmid pGRE-SEAP encodes the secreted alkaline phospha-tase reporter gene. It contains three tandem copies of GRE en-hancer fused to an herpes simplex virus thymidine kinase promo-tor. The plasmid pNFκB-SEAP contains a NFκB enhancer ele-ment in front of the SEAP reporter gene. Both vectors were ob-tained from Clontech (Palo Alto, Calif., USA). The expressionvector pRShGRα expresses the human GR under the control ofthe constitutively active Rous sarcoma virus promotor and was ob-tained from ATCC (Rockville, Md., USA).

Cell culture and transfection

IEC-6 cells were obtained from American Type Culture Collec-tion (Rockville, Md., USA) and were cultured in a 1:1 mixture ofDulbecco’s minimum essential medium and RPMI 1640 medium(Bio Whittacker, Verviers, Belgium) containing 10% fetal calf se-rum (FCS), 1% sodium pyruvate, 100 U/ml penicillin and100 U/ml streptomycin at 37°C in 5% CO2 atmosphere. Insulin(10–8 M) was freshly added to the cells every day. Medium waschanged every 3 days, and the cells were separated by trypsintreatment when they reached confluence. Caco-2 cells were ob-tained from DPZ (Braunschweig, Germany). Cells were grown inEMEM medium (Bio Whittacker) containing 20% FCS supple-mented with 100 U/ml each penicillin and streptomycin at 37°Cand 5% CO2.

The cells were plated in 24 well plates 48 h before transfec-tion (Nunc, Rushilde, Danmark) at a density of 30,000/well in2 ml medium. After 24 h medium was replaced by medium freeof endogenous steroids containing 0.5% bovine serum albuminand antibiotics at the above concentrations. The cells were trans-fected using the nonliposomal formulation FuGENE (Roche Mo-lecular Biochemicals, Mannheim, Germany). On the day oftransfection fresh medium was added. FuGENE was added to theplasmid DNA at a ratio of 3 µl/µg DNA. FuGENE was predilut-ed in 100 µl serum free medium and added dropwise to the con-centrated plasmid DNA. After 15 min at room temperature themixture was added to the cells. In transactivation studies 200 ngpGRE-SEAP/well were used for transient transfection. In trans-repression experiments the cells received 200 ng pNFκB-SEAP/well. In cotransfection experiments transfection was car-ried out with 200 ng of the respective reporter plasmid and 1 µgof the expression vector pRShGRα or an equal amount of Blue-script vector as control to equilibrate to an equal amount of plas-mid DNA/well.

In transactivation experiments cells were treated with decreas-ing concentrations (10–6 M–10–9 mol/l) of dexamethasone (Sigma,Munich, Germany), RU-486 (Exelgyn, Paris, France) 10–6 mol/land/or IL-1β (Roche, Mannheim, Germany) 5–500 U/ml 24 hposttransfection. Medium for determination of SEAP activity wascollected 24 h after stimulation. In transrepression experiments IL-1β at different concentrations (5–500 U/ml) was added to the me-dium 24 h post transfection. The effect of GC was investigated bysimultaneous incubation with Dex (1 µmol/l). Controls received anequal amount of vehicle substances ethanol (never exceeding0.01%) or phosphate-buffered solution (PBS). Six hours afterstimulation the medium was collected for determination of SEAPactivity. Alkaline phosphatase activity was measured in a chemo-luminescence based assay in the culture medium samples accord-ing to the manufacturers protocol (SEAP assay kit, Roche). Che-moluminescence signals were counted in a Wallac scintillationcounter (Wallac, Uppsala, Sweden) with the coincidence circuitturned off. Data are expressed as proportional increase over con-trol. Data are shown as means + SEM of three experiments. Allmeasurements were carried out in duplicate.

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Immuncytochemistry

Immuncytochemistry was carried out as described before [19]with some modifications. Briefly, IEC-6 cells were seeded ontolab-Tek Permanox 8-chamber slides (Nunc, Narparville, USA) at10,000 cells/well in 0.5 ml complete culture medium. After 24 hmedium was replaced by medium containing 10% albumin insteadof FCS and IL-1β (500 U/ml) was added as indicated. After fur-ther 24 h, cells were treated with Dex (1 µmol/l) dissolved in etha-nol (Sigma, Munich, Germany) for 1 h. Controls received the ve-hicle substance ethanol (0.01%) alone.

After removal of the medium cells were washed twice in PBSpH 7.4, at +37°C and fixed in 3% paraformaldehyde for 40 min at+37°C. After two washes in PBS cells were permeabilized for ex-actly 10 min in 0.3% Triton X-100 dissolved in PBS. Unspecificbinding sites were blocked by exposure of the cells to PBS/0.2%gelatin for 15 min. Thereafter IEC-6 cells were incubated over-night with the monoclonal antibody BuGR2 (Affinity Bioreagents,Neshanic Station, USA; 10 µg/ml) diluted 1:200, followed bythree washes for 5 min each. The second fluorescein-labeled anti-mouse-IgG antibody (Sigma, Munich, Germany) was then appliedat +37°C for 1 h diluted 1:200. Finally three washing steps werecarried out in PBS/0.2% gelatin, two washes in PBS, and threerinses in tap water. Slides were then mounted in Fluoromount(Southern Biotechnology, Birmingham, Ala., USA) and subjectedto confocal laser scanning microscopy using a Zeiss LSM laserscanning confocal microscope (Zeiss, Göttingen, Germany). Incontrol experiments primary antibody was replaced by normalmouse serum. This experiment was carried out twice.

Results

GR transactivation

To characterize the transactivating capability of gluco-corticoids in intestinal epithelial cells the activation ofGR in IEC-6 and Caco-2 cells was monitored using aGRE-containing reporter plasmid. GR activation resultedin a maximal 5.2±0.4-fold induction of the reporter geneproduct SEAP over basal in IEC-6 and a 3.8±0.8-fold in-duction in Caco-2 cells in the presence of Dex(1 µmol/l). GR transactivation by Dex was shown to beconcentration dependent in both cell lines (Fig. 1). Theeffect of Dex was antagonized by the competitive GRantagonist RU-486. RU-486 by itself had no effect onGR activation (Fig. 1, conditions 6, 7)

GR repression of NF-κB activity

To study the transrepressive effect of GC in IEC-6 andCaco-2 cells we studied whether GR also mediates tran-scriptional repression of the NF-κB pathway. IEC-6 andCaco-2 cells were transiently transfected with pNFκB-SEAP. IL-1β induced a concentration- dependent in-crease in NF-κB mediated reporter gene activity with amaximal stimulatory effect of 7.3±2.3-fold increase at aconcentration of 500 U/ml in IEC-6 and 8.6±0.2-fold in-crease in Caco-2 (Fig. 2). In Caco-2 cells Dex did notsignificantly affect NF-κB stimulation induced by IL-1β.However, in IEC-6 cells Dex significantly decreased IL-

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Fig. 1 Effect of dexamethasone (Dex) on transcriptional activa-tion in IEC-6 and Caco-2 cells. IEC-6 and Caco-2 cells weretransfected with the SEAP reporter plasmid pGRE-SEAP. Thecells were incubated with Dex and the GC antagonist RU-486 for24 h at concentrations as indicated. Reporter gene activation wascalculated in relation to the level of IEC-6 and Caco-2 cells treatedwith the vehicle substance ethanol alone. Data are shown as means+SEM for n=3. All experiments were carried out in duplicate

Fig. 2 IL-1β mediated NFκB activation and repressive effect ofdexamethasone (Dex). IEC-6 and Caco-2 cells were transientlytransfected with the NF-κB inducible SEAP reporter plasmidpNFκB-SEAP. Twenty-four hours posttransfection cells werestimulated with IL-1β (5–500 U/ml) alone or with Dex (1 µmol/l)for 6 h. Data are shown as increase relative to controls which re-ceived the Dex vehicle substance ethanol and PBS instead of IL-1β. Relative reporter gene activities are shown as means +SEMfor n=3. All experiments were carried out in duplicate

1β induced NFκB activity even at high IL-1β concentra-tions (Fig. 2).

GR overexpression in Caco-2 cells

Since Caco-2 cells obviously have GR which allowedtransactivation but seemed to fail to mediate transcrip-tional repression of the NF-κB pathway, we examinedwhether this apparently partial GC resistance is over-come by GR overexpression. When Caco-2 cells werecotransfected with pGRE-SEAP and pRShGRα Dex in-cubation (1 µmol/l) resulted in a 29.5±10.6-fold increasein reporter gene activity. Overexpression of GR without

stimulation with Dex had no effect on reporter gene ac-tivity compared to a mock-transfected control (Fig. 3).

When GR was overexpressed in Caco-2 together withpNF-κB-SEAP, spontaneous NF-κB activity was re-duced to 56±3% (Fig. 4, condition 1). Incubation withDex (1 µmol/l) resulted in a further decrease in NF-κBactivity to 46±4% of control (Fig. 4, condition 2). Whenstimulated with IL-1β (500 U/ml), NF-κB activity inCaco-2 cells was increased but did not largely excedelevels of the mock transfected control under basal condi-tions. In contrast to cells which do not overexpress GR, asimultaneous incubation with Dex (1 µmol/l) resulted ina marked decrease in IL-1β-stimulated NF-κB activity(Fig. 4, condition 4).

Effect of IL-1β on GR-transactivation

To test the hypothesis that the proinflammatory cytokineIL-1β inhibits GC-mediated transactivation pGRE-SEAPtransfected cells were coincubated with IL-1β and Dex(1 µmol/l) for 24 h. IL-1β concentration dependently re-pressed the stimulating effect of Dex on transactivationin both IEC-6 and Caco-2 cells (Fig. 5). IL-1β impairedGRE activation also under the condition of GR overex-pression in Caco-2 cells (Fig. 3, condition 3). IL-1β didnot affect the basal reporter gene transcription (data notshown).

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Fig. 3 Effect of dexamethasone (Dex) and IL-1β on transcription-al activation in Caco-2 cells overexpressing glucocorticoid recep-tor using the expression vector pRShGRα. Cells were stimulatedwith Dex (1 µmol/l) in the absence and presence of IL-1β(500 U/ml). Transcriptional activation was calculated relative toreporter gene activity obtained from unstimulated cells mocktransfected with Bluescript vector (transcriptional activation=1).Relative reporter gene activities are shown as means +SEM forn=3. All measurements were performed in duplicate

Fig. 4 IL-1β mediated NFκB activation and repressive effect ofdexamethasone (Dex) in Caco-2 cells overexpressing glucocorti-coid receptor utilizing the expression vector pRShGRα. Twenty-four hours posttransfection Caco-2 cells were stimulated with IL-1β 500 U/ml and/or Dex (1 µmol/l) for 6 h. Transcriptional activa-tion/repression was calculated relative to reporter gene activity ob-tained from unstimulated cells, mock transfected with Bluescriptvector (transcriptional activation=1). Relative reporter gene activi-ties are shown as means +SEM for n=3. All experiments were car-ried out in duplicate

Fig. 5 IL-1β inhibition of transcriptional activation by glucocor-ticoids in IEC-6 and Caco-2 cells. Cells were stimulated withdexamethasone (Dex; 1 µmol/l) in the presence of IL-1β(5–500 U/ml) for 24 h. Transcriptional activation was calculatedrelative to reporter gene activity obtained from cells incubatedwithout Dex and IL-1β. Data shown are means +SEM from n=3independent experiments carried out in duplicate

Effect of IL-1β on GR-immunostaining

By means of immunostaining the shuttling of GR fromcytoplasm to the nucleus upon GC activation can be vi-sualized. To answer the question in which subcellularcompartment IL-1β interferes with GR signaling we per-formed immuncytochemistry with the monoclonal anti-GR antibody BuGR2. The nuclear staining was neglegi-ble in IEC-6 controls grown in steroid-free medium,while a cytoplasmatic staining was clearly detected(Fig. 6). A marked increase in nuclear signal was ob-served upon Dex administration. This effect was blockedby pretreatment with IL-1β 500 U/ml. IL-1β treatmentby itself did not affect the staining intensity or subcellu-lar distribution.

Discussion

GR is an ubiquitously expressed transcription factor. Inthe rat its distribution in the colon is well characterized[20]. In humans there is functional evidence for the pres-ence of GR in the intestine since GC causes an increasein sodium absorption in normal and inflamed mucosa[21]. However, the molecular mechanisms involved ingene regulation by GC in the intestinal mucosa and espe-

cially in the epithelium have been poorly understood. Bytransfection with a GRE-containing reporter gene wedemonstrated that Dex causes a transcriptional activationin IEC-6 and Caco-2 intestinal cell lines in a concentra-tion-dependent manner. This effect was antagonized byRU-486, a GR-antagonist, indicating that this effect wasspecifically mediated by GR.

We observed that IL-1β, one of the main cytokines inIBD, activates the transcription factor NF-κB in a dose-dependent manner in both cell-lines, illustrating that epi-thelial cells are part of the inflammatory process in IBD.NF-κB was found to be activated in biopsy samples andmononuclear cells from the lamina propria [22]. Sincegene regulation of many proinflammatory mediatorssuch as cyto- and chemokines, enzymes, and adhesionmolecules is driven by this trancription factor, it is an at-tractive candidate for playing a central role in pathogen-esis of IBD [5]. Neurath et al. [23] have reported that in-hibition of NF-κB by an antisense oligonucleotide di-rected against NF-κB mRNA improves experimental co-litis in mice. Therefore NF-κB inactivation offers a valu-able possibility for therapy of IBD.

Two mechanism have been proposed to explain GR-mediated inhibition of NF-κB. First GR can physicallyinteract with p65 a NF-κB subunit. Secondly, GR canupregulate IκBα, an cytoplasmic inhibitor of NF-κB[24]. We found that Dex inhibits NF-κB activation in theepithelial cell-line IEC-6. This observation is in accor-dance with a number of previous clinical observations.Schreiber et al. [22] have reported that NF-κB binding toDNA is increased in mucosal biopsy specimens from pa-tients with Crohn’s disease during acute inflammation.This increased DNA binding proved reversible by GCtreatment. Thiele et al. [25] demonstrated by immunohis-tochemistry that gut specimens from patients withCrohn’s disease under GC therapy have significantlylower nuclear NF-kB p65 levels than specimens fromuntreated patients. This was the case for epithelial, endo-thelial, and mononuclear cells. However, a parallel IκBαinduction was present only in vascular endothelial cells,suggesting cell specificity in the mode of NF-κB inhibi-tion by GC.

In Caco-2 cells Dex failed to inhibit IL-1β inductionof NF-κB activity despite a regular GR transactivation,reflecting a partial GC refractoriness. A correspondingobservation was made by Ardite et al. [26] in vivo inves-tigating NF-κB activity in biopsies from patients withIBD. One patient out of 15, who did not respond to GCtreatment showed a persisting increase in NF-κB bindingto DNA. Moreover, Bantel et al. [27] showed an abnor-mal activation of NF-κB in colonic epithelial cells in pa-tients with steroid resistant IBD by means of immunohis-tochemistry.

Our present findings demonstrate that IL-1β not onlycounteracts GC effects on NF-κB repression but also in-hibits GC driven transactivation both in IEC-6 and Caco-

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Fig. 6 Glucocorticoid receptor (GR) immunostaining of IEC-6cells grown on multichamber slides. Preincubation with and with-out IL-1β (500U/ml) for 24 h was carried out as indicated. Afterincubation with or without 1 µmol/l dexamethasone (Dex) for afurther 1 h at +37°C, cells were processed as described above. Inthe absence of hormone, staining is mostly cytoplasmic (upper leftpanel). A marked increase in nuclear immunofluorescence wasobserved after dexamethasone addition (upper right panel; bar10 µm). After IL-1β preincubation no GR translocation was ob-served (lower panels). Representative visual fields from one oftwo experiments are shown

2 cells. These observations raise the question of the mo-lecular mechanisms involved in steroid refractoriness. Inperipheral blood T cells from patients with steroid resis-tant ulcerative colitis GC fail to inhibit lectin-inducedproliferation [28]. One possible explanation is an in-creased multidrug resistance expression as proposed byFarrell et al. [29]. However, other data suggest direct al-terations in GR homeostasis. Schottelius et al. [30] foundan increased GR expression and decreased receptor af-finity in peripheral blood mononuclear cells from pa-tients with IBD. Although patients were not steroid resis-tant, these observations illustrate the systemic characterof inflammation in IBD. Rogler et al. [31] found thatspecific GC binding is reduced in colonic mucosal biop-sies, hypothesizing that a decreased protection againstNF-κB contributes to the chronic inflammation in IBD.

Another possible explanation for steroid refractor-iness in the case of ulcerative colitis may also be an in-creased expression of the GR isotype GRβ, which hasbeen reported to be an dominant negative inhibitor ofGR [32]. The significance of these phenomena is pres-ently unknown. However, they suggest the possibility ofheterogeneity in the pathogenesis of steroid refractor-iness in IBD. In the context of our cell culture model,partial steroid refractoriness of Caco-2 cells were re-stored by GR overexpression, suggesting that the amountof available GR is limiting for transrepression of NF-κBby GC. Therefore GR expression may be a positive pre-dictor for steroid responsiveness in IBD.

IL-1β inhibition of GR signaling seems to be causedby an inhibition of GR translocation to the nucleus asshown by GR immunostaining in IEC-6 cells. Althougha quantitative analysis of GR staining was not carriedout, there was no visible difference between IL-1β treat-ed cells and controls, suggesting that this inhibition oftranscriptional activation was not due to downregulationof GR protein. The fact that transcriptional activationfrom an exogenous GR is also inhibited by IL-1β further

suggests a mechanism independent of the regulation ofthe GR expression. It seems more likely that an imbal-ance in GR and NF-κB is responsible for inhibition ofGR signaling. A high quantity of activated NF-κB in-duced by inflammatory cytokines such as IL-1β mayoverwhelm GR, thereby inhibiting its function and al-lowing unhindered transcription of proinflammatorygenes. Alternatively, apart from direct protein-protein in-teraction one possible explanation is a competition ofGR and proinflammatory transcription factors for acti-vating cofactors. GR overexpression suppresses NF-κBactivity without the need of GC stimulation. In contrast,transactivation in Caco-2 cells is strictly hormone-de-pendent. GR overexpression therefore offers the possi-bility of an anti-inflammatory therapy without GC typi-cal side effects which have been widely attributed to thetransactivating capacity of GC.

Conclusion

These data suggest that the proinflammatory cytokineIL-1β produced during inflammation in IBD induces GCresistance in epithelial cells. Intestinal epithelial cells arepotential targets of GC hormone action and should be in-cluded into the concept of anti-inflammation by GC inthe treatment of IBD. The expression level of GR seemsto be a crucial point in effectiveness of both transactiva-tion and transrepression by GC. In view of the problemof GR resistance and various attempts of alternative anti-inflammatory therapies of IBD, further attention must bedrawn to mechanisms involved in GR regulation andfunction. This may reveal new concepts of improved GCtherapy in IBD which may also include gene therapy byGR overexpression.

Acknowledgements This work was supported by the DeutscheForschungsgemeinschaft Graduiertenkolleg 335

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