Abstract:The purpose of this study was to investigate the correlation between mRNA and protein levels for P-glycoprotein (P-gp) expressed in various cell lines to validate the estimation of P-gp activity from its mRNA levels. P-gp expression levels in various cell monolayers, normal, P-gp-induced, P-gp-highly induced, (multidrug resistance, MDR) MDR1-knockdown (A2-2) and MDR1-knockdown (B2-2) Caco-2 cells and MDCKII/MDR1 cells, were quantified by real-time quantitative polymerase chain reaction (PCR) and western blot a… Show more
“…2c, d). In line with published data for OATP4C1 (Taghikhani 2019;Mikkaichi et al 2004;Shirasaka 2009), a band with a molecular weight of 115 kDa was detected in the MDCK-OATP4C1 single-transfected as well as in the MDCK-OATP4C1-P-gp double-transfected cell line (Fig. 2c).…”
Section: Characterization Of the Double-transfected Mdck-oatp4c1-p-gpsupporting
confidence: 90%
“…For P-gp, a band with a molecular mass of 170 kDa was detected in MDCK-P-gp and MDCK-OATP4C1-P-gp cells (Fig. 2 d) (Shirasaka 2009 ). Fig.…”
Elevated plasma concentrations of the uremic toxin asymmetric dimethylarginine (ADMA) and low plasma concentrations of l-homoarginine are independently associated with cardiovascular events and mortality. Key enzymes involved in the homeostasis of both arginine derivatives are expressed in proximal tubule cells of the kidney. To get access to these enzymes, transport proteins are important. One of the transporters mediating the transport of ADMA and l-homoarginine is the solute carrier superfamily (SLC) member OATP4C1, located in the basolateral membrane of proximal tubule cells. To gain insights into the role of export pumps in the transport of both substances, we established a double-transfected MDCK cell line expressing OATP4C1 and the export pump P-glycoprotein (P-gp). Using MDCK cell monolayers, we demonstrated in time-dependent and concentration-dependent vectorial transport experiments that ADMA and l-homoarginine are transported from the basolateral to the apical compartment of MDCK-OATP4C1-P-gp cells with significantly higher transport rates compared to single-transfected MDCK-OATP4C1, MDCK-P-gp and MDCK-VC (control) cells (e.g. transport ratio MDCK-OATP4C1-P-gp/MDCK-VC: for 50 µM ADMA = 2.0-fold, for 50 µM l-homoarginine = 3.4-fold). These results indicate that both OATP4C1 and P-gp transport the arginine derivatives ADMA and l-homoarginine and are, therefore, important for the homoeostasis of both substances.
“…2c, d). In line with published data for OATP4C1 (Taghikhani 2019;Mikkaichi et al 2004;Shirasaka 2009), a band with a molecular weight of 115 kDa was detected in the MDCK-OATP4C1 single-transfected as well as in the MDCK-OATP4C1-P-gp double-transfected cell line (Fig. 2c).…”
Section: Characterization Of the Double-transfected Mdck-oatp4c1-p-gpsupporting
confidence: 90%
“…For P-gp, a band with a molecular mass of 170 kDa was detected in MDCK-P-gp and MDCK-OATP4C1-P-gp cells (Fig. 2 d) (Shirasaka 2009 ). Fig.…”
Elevated plasma concentrations of the uremic toxin asymmetric dimethylarginine (ADMA) and low plasma concentrations of l-homoarginine are independently associated with cardiovascular events and mortality. Key enzymes involved in the homeostasis of both arginine derivatives are expressed in proximal tubule cells of the kidney. To get access to these enzymes, transport proteins are important. One of the transporters mediating the transport of ADMA and l-homoarginine is the solute carrier superfamily (SLC) member OATP4C1, located in the basolateral membrane of proximal tubule cells. To gain insights into the role of export pumps in the transport of both substances, we established a double-transfected MDCK cell line expressing OATP4C1 and the export pump P-glycoprotein (P-gp). Using MDCK cell monolayers, we demonstrated in time-dependent and concentration-dependent vectorial transport experiments that ADMA and l-homoarginine are transported from the basolateral to the apical compartment of MDCK-OATP4C1-P-gp cells with significantly higher transport rates compared to single-transfected MDCK-OATP4C1, MDCK-P-gp and MDCK-VC (control) cells (e.g. transport ratio MDCK-OATP4C1-P-gp/MDCK-VC: for 50 µM ADMA = 2.0-fold, for 50 µM l-homoarginine = 3.4-fold). These results indicate that both OATP4C1 and P-gp transport the arginine derivatives ADMA and l-homoarginine and are, therefore, important for the homoeostasis of both substances.
“…The transport of Rh2s in parental MDCKII cells was used as a negative control because the cells have very low expression of P-gp (Shirasaka et al, 2009). As expected, the efflux ratio of Rh2s was much lower in parental MDCKII cells compared with MDR1-MDCKII cells at 2 M (28.0 versus 2.4).…”
ABSTRACT:The development of 20(S)-ginsenoside Rh2 (Rh2s) as a chemoprevention agent is limited by its low oral bioavailability. The goals of this study were to determine the mechanisms responsible for its poor oral absorption and to improve its bioavailability by overcoming the barrier to its absorption. Comprehensive studies were conducted using the following models: 1) monolayers of Caco-2, parental, and multidrug resistance gene ( at 20 mg/kg) was substantially increased by P-gp inhibitor to 33.18 and 27.14%, respectively. As expected, deletion or inhibition of P-gp significantly increased absorption and steady-state plasma concentration of Rh2s in a mouse intestinal perfusion model. In conclusion, Rh2s is a good substrate of P-gp, and inhibition of P-gp can significantly enhance its oral bioavailability.
“…It has been shown that ABC-transporter mRNA levels in cell lines do not always predict protein expression [ 57 ] or in vitro drug sensitivity [ 58 ]. Furthermore ABC-transporter expression or function in human tumor samples does not necessarily predict in vivo chemosensitivity [ 59 , 60 ].…”
Drug resistance (DR) is the major limiting factor in the successful treatment of systemic neoplasia with cytotoxic chemotherapy. DR can be either intrinsic or acquired, and although the development and clinical implications are different, the underlying mechanisms are likely to be similar. Most causes for DR are pharmacodynamic in nature, result from adaptations within the tumor cell and include reduced drug uptake, increased drug efflux, changes in drug metabolism or drug target, increased capacity to repair drug-induced DNA damage or increased resistance to apoptosis. The role of active drug efflux transporters, and those of the ABC-transporter family in particular, have been studied extensively in human oncology and to a lesser extent in veterinary medicine. Methods reported to assess ABC-transporter status include detection of the actual protein (Western blot, immunohistochemistry), mRNA or ABC-transporter function. The three major ABC-transporters associated with DR in human oncology are ABCB1 or P-gp, ABCC1 or MRP1, and ABCG2 or BCRP, and have been demonstrated in canine cell lines, healthy dogs and dogs with cancer. Although this supports a causative role for these ABC-transporters in DR cytotoxic agents in the dog, the relative contribution to the clinical phenotype of DR in canine cancer remains an area of debate and requires further prospective studies.
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