Clinical trial: modulation of human placental multidrug resistance proteins in cholestasis of pregnancy by ursodeoxycholic acid.
ABSTRACT The effects of ursodeoxycholic acid on human placental bile acids and bilirubin transporters in intrahepatic cholestasis of pregnancy are still undefined.
To evaluate whether ursodeoxycholic acid affects MRP2, MRP3 and MRP4 expression in the placenta.
Forty-three pregnant women were enrolled; fourteen subjects had physiological pregnancies. Intrahepatic cholestasis of pregnancy patients were divided into two groups: (i) 13 received ursodeoxycholic acid (20 mg/kg/day) and (ii) 16 untreated. Total bile acid and bilirubin in serum and cord blood were determined in each subject. Multidrug resistance proteins expression (immunoblot, quantitative real-time PCR) was evaluated in placentas collected at delivery. anova test was used for statistical analysis of data.
Ursodeoxycholic acid administration significantly improved maternal serum bile acid and cord blood bilirubin and bile acid levels. MRP2 protein and RNA expression was significantly increased in placentas from treated patients compared to controls (P < 0.001 and P < 0.01, respectively). MRP3 protein expression was not significantly different between the groups while RNA expression was significantly decreased in treated patients (P < 0.01). MRP4 did not show significant differences between the groups.
Ursodeoxycholic acid administration induces placental MRP2 expression, and reduces bilirubin and bile acid levels in cord blood.
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ABSTRACT: Sperm capacitation has been largely associated with an increase in cAMP, although its relevance in the underlying mechanisms of this maturation process remains elusive. Increasing evidence shows that the extrusion of cAMP through multidrug resistance associated protein 4 (MRP4), regulates cell homeostasis not only in physiological but also in pathophysiological situations and studies from our laboratory strongly support this assumption. In the present work we sought to establish the role of cAMP efflux in the regulation of sperm capacitation. Sperm capacitation was performed in vitro by exposing bovine spermatozoa to bicarbonate 40 and 70 mM; cAMP; probenecid (a MRPs general inhibitor) and an adenosine type 1 receptor (A1 adenosine receptor) selective antagonist (DPCPX). Capacitation was assessed by chlortetracycline assay and lysophosphatidylcholine-induced acrosome reaction assessed by PSA-FITC staining. Intracellular and extracellular cAMP was measured by radiobinding the regulatory subunit of PKA under the same experimental conditions. MRP4 was detected by Western blot and immunohistochemistry assays. Results showed that the inhibition of soluble adenylyl cyclase significantly inhibited bicarbonate-induced sperm capacitation. Furthermore, in the presence of 40 and 70 mM bicarbonate bovine spermatozoa synthesized and extruded cAMP. Interestingly, in the absence of IBMX (a PDEs inhibitor) cAMP efflux still operated in sperm cells, suggesting that cAMP extrusion would be a physiological process in the spermatozoa complementary to the action of PDE. Blockade of MRPs by probenecid abolished the efflux of the cyclic nucleotide resulting not only in the accumulation of intracellular cAMP but also in the inhibition of bicarbonate-induced sperm capacitation. The effect of probenecid was abolished by exposing sperm cells to cAMP. The high affinity efflux pump for cAMP, MRP4 was expressed in bovine spermatozoa and localized to the mid-piece of the tail as previously reported for soluble adenylyl cyclase and A1 adenosine receptor. Additionally, blockade of A1 adenosine receptor abolished not only bicarbonate-induced sperm capacitation but also that stimulated by cAMP. Present findings strongly support that cAMP efflux, presumably through MRP4, and the activation of A1 adenosine receptor regulate some events associated to bicarbonate-induced sperm capacitation, and further suggest a paracrine and/or autocrine role for cAMP.Molecular Human Reproduction 08/2013; · 4.54 Impact Factor
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ABSTRACT: During pregnancy, the disposition of various drugs is altered due to changes in physiological condition, maternal gastrointestinal absorption, gastric secretion and motility. A fixed dose combination of antiretrovirals is commonly prescribed for the treatment of HIV infection. There is a need to understand the pharmacokinetics and placental transfer of efavirenz-emtricitabine-tenofovir in fixed dose combination during pregnancy. The pharmacokinetics and placental transfer of efavirenz-emtricitabine-tenofovir fixed dose combination was evaluated in timed pregnant and non-pregnant Sprague-Dawley rats at 30, 10, 15 mg/kg p.o., respectively. The plasma, placental tissue, amniotic fluid and fetal tissue concentrations were measured using high performance liquid chromatography combined with tandem mass spectrometric detector (LC-MS/MS). To summarize, the pharmacokinetic profile of efavirenz remained similar in the pregnant and non-pregnant rats. However, a considerable difference in the pharmacokinetics of emtricitabine and tenofovir was observed in pregnant and non-pregnant rats. Efavirenz and emtricitabine showed appreciable placental, amniotic fluid and fetal exposure compared with tenofovir. The present study suggests that a profound impact on antiretroviral pharmacokinetics was observed during pregnancy and there is a need to monitor the exposure levels of each drug when administered as a fixed dose combination during pregnancy. Further studies to explore the pharmacokinetic parameters of fixed dose antiretrovirals during the preclinical stage in a timed-pregnancy rat model are required. Such studies can help in the development of safe and effective medications with a reduced risk of perinatal transmission of HIV-1 infection.Biopharmaceutics & Drug Disposition 05/2012; 33(5):265-77. · 2.09 Impact Factor
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ABSTRACT: Ursodeoxycholic acid (UDCA) administration in intrahepatic cholestasis of pregnancy (ICP) induces bile acids (BA) efflux from the foetal compartment, but the molecular basis of this transplacental transport is only partially defined. To determine if placental breast cancer resistance protein (BCRP), able to transport BA, is regulated by UDCA in ICP. 32 pregnant women with ICP (14 untreated, 34.9±5.17 years; 18 treated with UDCA - 25 mg/Kg/day, 32.7±4.62 years,) and 12 healthy controls (33.4±3.32 years) agreed to participate in the study. Placentas were obtained at delivery and processed for membrane extraction. BCRP protein expression was evaluated by immunoblotting techniques and chemiluminescence quantified with a luminograph measuring emitted photons; mRNA expression with real time PCR. Statistical differences between groups were evaluated by ANOVA with Dunn's Multiple Comparison test. BCRP was expressed only on the apical membrane of the syncytiotrophoblast. A significant difference was observed among the three groups both for mRNA (ANOVA, p = 0.0074) and protein (ANOVA, p<0.0001) expression. BCRP expression was similar in controls and in the untreated ICP group. UDCA induced a significant increase in placental BCRP mRNA and protein expression compared to controls (350.7±106.3 vs 100±18.68% of controls, p<0.05 and 397.8±56.02 vs 100±11.44% of controls, p<0.001, respectively) and untreated ICP (90.29±17.59% of controls, p<0.05 and 155.0±13.87%, p<0.01). Our results confirm that BCRP is expressed only on the apical membrane of the syncytiotrophoblast and show that ICP treatment with high dose UDCA significantly upregulates placental BCRP expression favouring BA efflux from the foetal compartment.PLoS ONE 01/2013; 8(5):e64101. · 3.73 Impact Factor