RESUMO
Probiotics are considered to have a beneficial impact on humans, but in some cases, administration of live microorganisms might be risky. In the present study, immunomodulatory effects of different Escherichia coli strains and their super-natants were examined under different inflammatory conditions with living and heat-inactivated strains. HT-29 cells were incubated with E. coli strains (S2-G1, S2-G3, S2-G4 and S2-G8) and their supernatants with or without stimulation with tumor necrosis factor alpha (TNF-α) or interleukin (IL)-1ß. Quantification of IL-8 secretion and gene expression was performed by enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (PCR). IL-8 secretion by TNF-α- and IL-1ß-stimulated cells was attenuated by all four live strains. In contrast, heat inactivation resulted in an elevated IL-8 expression and secretion in unstimulated cells and did not maintain the anti-inflammatory effect of live bacteria in cytokine-stimulated cells. The supernatant of the live S2-G3 led to an elevated IL-8 secretion in unstimulated and IL-1ß-stimulated cells but not in TNF-α-stimulated cells. Live bacteria of all strains might induce an immunosuppressive effect after stimulation of HT-29 cells, whereas heat inactivation and the supernatant seem to induce an elevated immune response. These findings might have an impact depending on the indication and purpose of administration.
RESUMO
INTRODUCTION: Atopic dermatitis (AD) has been related to a deficiency of delta-6-desaturase, an enzyme responsible for the conversion of linoleic acid to gamma-linolenic acid (GLA). Evening primrose oil (EPO) contains high amounts of GLA. Therefore, this study investigated whether EPO supplementation results in an increase in plasma GLA and its metabolite dihomo-gamma-linolenic acid (DGLA) correlating with clinical improvement of AD, assessed by the SCORing Atopic Dermatitis (SCORAD) index. METHODS: The open study included 21 patients with AD. EPO (4-6 g) was administered daily for 12 weeks. Before treatment, and 4 and 12 weeks after initiation of EPO supplementation, objective SCORAD was assessed and plasma concentrations of GLA and DGLA were determined by gas chromatography. RESULTS: A significant increase in plasma GLA and DGLA levels and a decrease in the objective SCORAD were observed 4 and 12 weeks after initiation of EPO treatment. In the per-protocol population (n = 14), a significant inverse correlation between the changes in plasma GLA levels and SCORAD was found (P = 0.008). CONCLUSION: The clinical disease activity under EPO treatment correlates with the individual increase in plasma GLA levels. Thus, the results of this pilot study indicate that an increase in plasma GLA might be used as predictive parameter for responsiveness of AD to EPO therapy.
Assuntos
Ácido 8,11,14-Eicosatrienoico/sangue , Dermatite Atópica/tratamento farmacológico , Fármacos Dermatológicos/uso terapêutico , Ácidos Linoleicos/uso terapêutico , Óleos de Plantas/uso terapêutico , Ácido gama-Linolênico/sangue , Adolescente , Adulto , Criança , Pré-Escolar , Estudos de Coortes , Dermatite Atópica/sangue , Fármacos Dermatológicos/química , Feminino , Humanos , Ácidos Linoleicos/química , Masculino , Pessoa de Meia-Idade , Oenothera biennis , Óleos de Plantas/química , Estudos Prospectivos , Resultado do Tratamento , Adulto Jovem , Ácido gama-Linolênico/análise , Ácido gama-Linolênico/química , Ácido gama-Linolênico/uso terapêuticoRESUMO
PURPOSE: Preoperative radiochemotherapy is widely used in the treatment of locally advanced rectal cancer. The predictive value of response to neoadjuvant treatment remains uncertain. We retrospectively evaluated the impact of downstaging and tumor regression as prognostic factors and its influence on the ability to perform sphincter-sparing surgery. PATIENTS AND METHODS: A total of 72 consecutive patients with advanced rectal cancer were included in this retrospective analysis. All patients were treated with preoperative 5-fluorouracil-based chemotherapy and pelvic radiation with a total dose of 50.4 Gy followed by surgery 6 weeks later. RESULTS: A sphincter-preserving procedure could be performed on 42 patients, and in all 72 patients complete resection (R0) was achieved. A pathological complete response (ypT0, ypN0) was achieved in 8 (11%) patients. None of the patients showing a complete pathological response relapsed or died during the follow-up period. At a median follow-up of 28 months, 65 patients were alive, none of these patients had local recurrence and 15 patients had metastatic disease. Patients showing a complete pathological response had a significantly better 2-year disease-free survival compared to patients with ≥10% residual tumor cells (p = 0.024). Patients < 65 years showed a significantly better response rate, compared with those > 65 years of age (p = 0.036). Acute toxicity was moderate. CONCLUSION: Preoperative radiochemotherapy is an effective and safe treatment for patients with locally advanced rectal cancer. Pathological parameters after preoperative radiochemotherapy, including tumor regression grading, could be correlated with disease-free survival. The impact of tumor regression grading needs to be further validated in prospective clinical trials.
Assuntos
Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/radioterapia , Adenocarcinoma/cirurgia , Carcinoma de Células em Anel de Sinete/tratamento farmacológico , Carcinoma de Células em Anel de Sinete/radioterapia , Carcinoma de Células em Anel de Sinete/cirurgia , Carcinoma/tratamento farmacológico , Carcinoma/radioterapia , Carcinoma/cirurgia , Terapia Neoadjuvante , Neoplasias Retais/tratamento farmacológico , Neoplasias Retais/radioterapia , Neoplasias Retais/cirurgia , Adenocarcinoma/patologia , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Antimetabólitos Antineoplásicos/administração & dosagem , Carcinoma/patologia , Carcinoma de Células em Anel de Sinete/patologia , Terapia Combinada , Intervalo Livre de Doença , Fracionamento da Dose de Radiação , Feminino , Fluoruracila/administração & dosagem , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Dosagem Radioterapêutica , Neoplasias Retais/patologia , Reto/patologia , Reto/cirurgia , Estudos RetrospectivosRESUMO
Interactions of acutely administered grapefruit juice (GFJ) with cytochrome P450 isoform 3A4 (CYP3A4) and P-glycoprotein (Pgp) function are well established. In this study, we investigated in vitro the effect of repeated administration of GFJ and its major constituents (the flavonoid naringin, its aglycone naringenin and the furanocoumarin bergamottin) on mRNA expression of MDR1 and CYP3A4 in LS180 cells. Since the bergamottin content is higher in the peel than in the fruit, we compared GFJ containing peel (GFJP+) with juice without any peel extract (GFJP-). GFJP- (1%) showed no significant effect on MDR1 and CYP3A4 mRNA expression, whereas 1% GFJP+ increased expression of MDR1 3.7-fold (P<0.01) and CYP3A4 2.3-fold (P<0.05). Of the tested constituents, only 10 microM bergamottin and 200 microM naringenin induced MDR1 mRNA levels 2.9- and 4.0-fold, respectively (P<0.01 for both), and CYP3A4 mRNA levels 3.2- and 15.6-fold (P<0.01 for both), respectively. Western blot analysis and rhodamine 123 uptake experiments partly confirmed these findings on the protein and the functional level. In summary, GFJ containing no peel extract may have a lower potential for interactions with CYP3A4 or P-glycoprotein.
Assuntos
Bebidas/efeitos adversos , Citrus paradisi/química , Subfamília B de Transportador de Cassetes de Ligação de ATP , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Linhagem Celular Tumoral , Citocromo P-450 CYP3A , Relação Dose-Resposta a Droga , Esquema de Medicação , Interações Medicamentosas , Frutas/química , Humanos , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
Budesonide, a glucocorticoid with a high first-pass metabolism, is used for the oral treatment of inflammatory bowel disease. Cytochrome P450 3A4 (CYP3A4) is an enzyme involved in the metabolism of numerous drugs, including budesonide. Since inhibition or induction of CYP3A4 is often the cause of drug-drug interactions we analyzed how budesonide affects the activity and expression of this enzyme. CYP3A4 activity was assessed by the metabolism of a luminogenic substrate (luciferin-benzylether) using recombinant human CYP3A4 protein. We observed no inhibition of the metabolism in the presence of budesonide at concentrations up to 25 microM. Induction experiments in human LS180 colon carcinoma cells showed an increased expression of CYP3A4 mRNA after budesonide treatment. Transactivation assays revealed that budesonide activates the CYP3A4 promoter via the pregnane X receptor (PXR). In mice, oral budesonide administration (25mg/kg) for 4 days induced the murine homolog Cyp3a11 in the intestine 3-fold, whereas liver expression was notably less influenced. In knockout mice devoid of PXR, budesonide-mediated inductions were reduced compared to wild-type mice. In conclusion, we could demonstrate that budesonide is not an efficient inhibitor but rather an inducer of CYP3A via a PXR-mediated mechanism. In vivo, however, oral budesonide administration to mice showed only modest gene induction, which occurred mainly in the intestine. Therefore, the risk for budesonide-mediated drug interactions seems to be low but cannot be ruled out entirely.
Assuntos
Budesonida/farmacologia , Citocromo P-450 CYP3A/biossíntese , Indução Enzimática/efeitos dos fármacos , Glucocorticoides/farmacologia , Proteínas de Membrana/biossíntese , Receptores de Esteroides/fisiologia , Animais , Linhagem Celular Tumoral , Citocromo P-450 CYP3A/genética , Interações Medicamentosas , Humanos , Mucosa Intestinal/metabolismo , Fígado/metabolismo , Masculino , Camundongos , Camundongos Knockout , Receptor de Pregnano X , Regiões Promotoras Genéticas , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Esteroides/genéticaRESUMO
AIMS: Efflux transporters such as breast cancer resistance protein (BCRP/ABCG2) and P-glycoprotein (Pgp; MDR1/ABCB1) are protecting the enterocytes from potentially toxic compounds. Both transporters have been reported to be downregulated in patients with active ulcerative colitis (UC). The aim of this study was to evaluate transporter expression in both unaffected and inflamed mucosa of patients with active UC, in drug-naïve and treated patients with UC and compare the results with transporter expression in healthy subjects. METHODS: Transporter expression was determined with real-time RT-PCR (TaqMan) in inflamed and unaffected mucosa of newly diagnosed (n = 12) and therapy-refractory (n = 11) patients with UC. Expression levels were compared with UC patients in remission (n = 11) and control subjects (n = 26). BCRP and Pgp expression was evaluated by immunohistochemistry. RESULTS: Compared with unaffected mucosa, BCRP expression was significantly reduced in inflamed mucosa of newly diagnosed drug-naïve patients with UC (expression reduced to 30%) as well as in patients not responding to treatment (reduced to 25%) with either 5-aminosalicylates (n = 7) or prednisone (n = 4). Unaffected mucosa of UC patients showed comparable transporter expression to unaffected mucosa of control subjects. MDR1 expression depicts a similar pattern. Protein staining for Pgp and BCRP was significantly reduced in the inflamed mucosa of patients with active UC. CONCLUSIONS: Expression of both efflux transporters BCRP and MDR1 is reduced, but only in inflamed tissue of patients with active UC. Transporter expression in unaffected mucosa of patients with active UC is comparable to healthy controls. The data suggest that the inflammatory process is responsible for the reduced levels. A major role in the pathogenesis of UC is unlikely.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/genética , Colite Ulcerativa/genética , Proteínas de Neoplasias/genética , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Adulto , Idoso , Feminino , Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , RecidivaRESUMO
Multidrug resistance protein 2 (MRP2/Mrp2) is a transporter that can influence the absorption, distribution, and elimination of many drugs. Mrp2 knockout mice are being used to study Mrp2 functions in vivo, including pharmacokinetics of drugs. To assess possible species-specific differences between human MRP2 and mouse Mrp2, we generated polarized cell lines expressing mouse Mrp2 and used these to investigate transport of clinically important agents. We also tested the ability of other drugs to modulate MRP2/Mrp2-mediated transport, a phenomenon that can lead to drug-drug interactions. In MDCK cells stably expressing human MRP2 or mouse Mrp2, saquinavir and docetaxel were more efficiently transported by mouse Mrp2, whereas vinblastine was transported better by human MRP2. MRP2/Mrp2-mediated transepithelial transport of several drugs could be stimulated by probenecid and sulfanitran, but stimulation was often more pronounced for human MRP2 than for mouse Mrp2. Interestingly, for some drugs the MRP2 modulator sulfinpyrazone had opposite effects on both transporters, stimulating human MRP2 and inhibiting mouse Mrp2 activity. In vesicular transport studies, transport of estradiol-17beta-glucuronide by mouse Mrp2 showed homotropic cooperativity, as previously described for human MRP2. The MRP2 modulators again showed differential effects on estradiol-17beta-glucuronide transport, most notably with sulfinpyrazone stimulating human MRP2 and profoundly inhibiting mouse Mrp2 activity. In conclusion, although human and mouse MRP2/Mrp2 have largely overlapping substrate specificities, there are important species differences in the transport efficiency of MRP2 substrates and in the modulation of transport by other compounds. These differences should be taken into account when results obtained in mice are extrapolated to humans.
Assuntos
Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Animais , Transporte Biológico/fisiologia , Linhagem Celular , Cães , Células Epiteliais/metabolismo , Humanos , Insetos , Camundongos , Camundongos Endogâmicos BALB C , Proteína 2 Associada à Farmacorresistência Múltipla , Preparações Farmacêuticas/metabolismo , Especificidade da Espécie , Especificidade por SubstratoRESUMO
BACKGROUND/AIMS: The expression of transporters involved in bile acid homeostasis is differentially regulated during obstructive cholestasis. Since the drug efflux transporter breast cancer resistance protein (BCRP) is known to transport bile acids, we investigated whether duodenal BCRP expression could be altered during cholestasis. METHODS: Using real-time RT-PCR analysis we determined mRNA expression levels in duodenal tissue of 19 cholestatic patients. Expression levels were compared to 14 healthy subjects. BCRP protein staining was determined in biopsies of 6 cholestatic and 6 healthy subjects by immunohistochemistry. RESULTS: We found that in patients with obstructive cholestasis mean duodenal BCRP mRNA levels were significantly reduced to 53% and mean protein staining was reduced to 57%. CONCLUSIONS: BCRP, a transporter for bile acids and numerous drugs, appears to be down-regulated in the human duodenum during cholestasis. The clinical impact of these results has to be investigated in further studies.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Colestase/metabolismo , Duodeno/metabolismo , Proteínas de Neoplasias/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/análise , Transportadores de Cassetes de Ligação de ATP/genética , Adulto , Fatores Etários , Idoso , Ácidos e Sais Biliares/metabolismo , Ductos Biliares/fisiopatologia , Índice de Massa Corporal , Colestase/genética , Colestase/fisiopatologia , Regulação para Baixo , Duodeno/química , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/análise , Proteínas de Neoplasias/genética , RNA Mensageiro/análise , RNA Mensageiro/metabolismoRESUMO
BACKGROUND: There is growing clinical interest in thalidomide for the treatment of various disorders due to its anti-inflammatory, immunomodulatory, and anti-angiogenic properties. In numerous clinical trials thalidomide is used as an adjunct to standard therapy. Therefore, clinicians should be aware of all possible drug-drug interactions that might occur with this drug. P-glycoprotein (P-gp), a drug efflux transporter that is expressed in many tissues, is the cause of several drug-drug interactions. P-gp induction or inhibition can lead to ineffective therapy or side-effects. In this study, we investigated thalidomide's potential to cause drug-drug interactions on the level of P-gp. METHODS: LS180 cells were incubated with thalidomide for 72 h in order to determine P-gp induction using real-time RT-PCR. A human leukaemia cell line over-expressing MDR1 (CCRF-CEM/MDR1) was used to measure uptake of rhodamine 123, a P-gp substrate, in the presence of thalidomide. Dose-dependent and bi-directional transport of thalidomide through Caco-2 cell monolayers was performed to assess site-directed permeability. Transport rates were determined using HPLC including chiral separation of the thalidomide enantiomers. RESULTS: Thalidomide did not induce P-gp expression in LS180 cells. The uptake of rhodamine 123 in CCRF cells over-expressing MDR1 was not influenced by co-incubation with thalidomide. The transport through Caco-2 monolayers was linear and the permeability was similar for both directions. No differences between the thalidomide enantiomers were observed. CONCLUSIONS: Our study indicates that thalidomide is neither a substrate, nor an inhibitor or an inducer of P-gp. Therefore, P-gp-related drug-drug interactions with thalidomide are not likely.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Imunossupressores/metabolismo , Talidomida/metabolismo , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Transporte Biológico , Células CACO-2 , Interações Medicamentosas , Resistência a Múltiplos Medicamentos , Humanos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Rodamina 123/metabolismoRESUMO
Human breast cancer resistance protein (BCRP/ABCG2) is an ABC-transporter that is present on the luminal membrane of intestinal epithelial cells and restricts absorption of anticancer drugs such as methotrexate, topotecan, mitoxantrone, and doxorubicin. The exact anatomic distribution of BCRP along the gastrointestinal (GI) tract, however, has not been determined before. The aim of this study was, therefore to investigate BCRP mRNA expression pattern along the GI tract in 14 healthy subjects. Furthermore, BCRP duodenal mRNA expression was compared with MDR1/ABCB1 mRNA. Additionally, BCRP mRNA expression was investigated in two human intestinal cell lines (Caco-2 and LS180). Since previous animal studies have suggested sex specific differences in BCRP expression, we analyzed intestinal BCRP expression with respect to sex. Biopsies were taken from different gut segments (duodenum, terminal ileum and ascending, transverse, descending and sigmoid colon). Gene expression was assessed by quantitative real-time PCR (Taqman). BCRP mRNA expression was maximal in the duodenum and decreased continuously down to the rectum (terminal ileum 93.7%, ascending colon 75.8%, transverse colon 66.6%, descending colon 62.8%, and sigmoid colon 50.1% compared to duodenum, respectively). BCRP expression in the duodenum was comparable to MDR1/ABCB1 gene expression. Caco-2 cells showed a comparable expression of BCRP as human duodenal tissue. Gender specific differences in BCRP expression were not observed. These findings represent the first systematic site-specific analysis of BCRP expression along the GI tract. This information might be helpful to develop target strategies for orally administered anticancer drugs.