Comparison of Effects of Aminosalicylic Acid, Glucocorticoids and Immunosuppressive Agents On The Expression of Multidrug-Resistant Genes in Ucerative Colitis

Objective: To compare the effects of aminosalicylic acid, glucocorticoids and immunosuppressive agents on the expression levels of multidrug-resistant genes in ucerative colitis (UC) patients, aiming to provide theoretical and therapeutic basis for the diagnosis, treatment and prevention of UC.Methods: Fresh specimen of colon mucosal tissues pathological mucosal tissues under endoscopy or postoperative pathological biopsy tissues were collected from 148 UC patients and then prepared for subsequent experiment.Immunohistochemical staining was conducted to detect the distribution site and pattern of P-glycoprotein （P-gp）. The effects of ASA glucocorticoids and immunosupresive agents on P-gp were statistically compared.The expression levels of Multidrug resistance gene （MDR1）mRNA before and after corresponding treatment were quantitatively measured by using RT-PCR. In addition, the effects of three agents upon MDR1 mRNA were also comparatively analyzed.Results: Administration of 5-aminosalicylic acid (5-ASA) drugs were not correlated with the expression of multidrug resistance genes in UC, whereas delivery of glucocorticoids and immunosuppressant drugs was positively correlated with the expression profile. The expression levels of MDR1 gene and its product P-gp were significantly up-regulated in patients who were nonresponsive to glucocorticoids and immunosuppressant agents.Conclusions: Overall, the findings in the present study demonstrate that 5-ASA exerts no effect upon the expression levels of MDR1 and its product P-gp in patients diagnosed with UC. Nevertheless, administration of glucocorticoids and immunosuppressant drugs can up-regulate the expression levels of MDR1.

Matrine Reverses Multidrug Resistance in Eca-109/VCR Cells Via Inhibiting PI3K/Akt/mTOR Signaling Pathway

Objective To investigate the effect of matrine on multidrug resistance of Eca-100/VCR cells. Methods Methyl thiazolyl tetrazolium assay was used to assess the cytotoxic activity of matrine to Eca-109/VCR cells and the sensitivity of cells to Vincristine (VCR). Then Eca-109/VCR cells were divided into four groups: control group, matrine group, VCR group and matrine combined with VCR group. Immunofluorescence staining was used to detect P-gp protein expression. Western blot was used to detect MRP1, P-gp, Beclin 1, p62(SQSTM1), LC3, p-mTOR, p-Akt and p-p70S6K protein expression. MDR1 mRNA expression was detected by RT-qPCR. Transmission electron microscopy was used to detect the ultra structural changes. Results Matrine could reverse the resistance of Eca-109/VCR cells to VCR with a reversal index of 3.52. Compared with the other groups, the expression of LC3-II, Beclin 1 increased while P-gp, MRP1, p62(SQSTM1), p-mTOR, p-Akt and p-p70S6K significantly decreased in matrine+VCR group (P<0.05). The expression of MDR1 mRNA decreased in Eca-109/VCR cells treated with matrine, VCR or their combination. The number of autophagic vacuoles in Eca-109/VCR cells increased after matrine treatment. Conclusion Matrine has anticancer and anti multidrug resistance functions in Eca-100/VCR cells, and it may be produced by promoting autophagy which mediated through PI3K/Akt/mTOR signaling pathway.

Paeoniflorin and albiflorin regulate P-gp-mediated aconitine and hypaconitine transport through an Madin Darby canine kidney-multi drug resistance protein 1 cell model

Aconitine and hypaconitine are the main active ingredients of Radix Aconiti, paeoniflorin and albiflorin are the primary components of Radix Paeoniae Alba. Both Radix Aconiti and Radix Paeoniae Alba are herbs that are commonly used in traditional Chinese medicine. This study sought to explore the mechanistic transport of aconitine and hypaconitine across MDCK-MDR1 cells and to assess the effect of paeoniflorin and albiflorin on aconitine and hypaconitine transmembrane transport as a potential attenuation mechanism. Drug cytotoxicity was tested via the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, and transport studies were performed in both directions. The effects of drugs on P-gp ATPase activity, P-gp efflux function, MDR1 mRNA and P-gp expression were evaluated in MDCK-MDR1 cells. Aconitine and hypaconitine treatment with the verapamil could significantly decrease the efflux rate (ER). The ER of aconitine and hypaconitine were significantly increased with the coadministration of paeoniflorin and albiflorin, suggesting that paeoniflorin and albiflorin can promote the efflux of these two alkaloids. Aconitine and hypaconitine can induce P-gp enzymatic activity, inhibit P-gp-mediated efflux, and downregulate the expression of P-gp protein to produce cytotoxic effects. When treatment in combination with paeoniflorin and albiflorin, it could stimulated P-gp ATPase activity, increasing mRNA expression, enhance P-gp efflux function, and upregulate P-gp protein expression.

First Sequencing of Caprine Mdr1 (Abcb1) mRNA Due to Suspected Neurological Adverse Drug Reaction in a Thuringian Goat Following Extra-Label Use of Doramectin

The multidrug resistance gene MDR1 encodes for an efflux transporter called P-glycoprotein (P-gp). In the canine Mdr1 gene, a nonsense mutation was identified in certain dog breeds causing increased drug sensitivity to various P-gp substrates such as antiparasitic macrocyclic lactones. Symptoms of neurologic toxicity include ataxia, depression, salivation, tremor, apparent blindness, and mydriasis. In the current report, a Thuringian goat developed similar neurological signs after treatment with doramectin, a compound from the macrocyclic lactone class. Therefore, Mdr1 might be defective in this individual goat. For diagnostic purposes, sequencing of the complete mRNA transcript coding for caprine Mdr1 was performed to investigate a potential missense mutation. The Mdr1 transcripts of two related goats without drug sensitivity were also sequenced to allow differential diagnosis and were compared to the suspected drug-sensitive goat. The only position where the Mdr1 sequence from the suspected drug-sensitive goat differed was in the 3′-untranslated region, being a heterozygous single nucleotide polymorphism c.3875C&gt;A. It can be suspected that this variant affects the expression level, stability, or translation efficiency of the Mdr1 mRNA transcript and therefore might be associated with the suspected drug sensitivity. To clarify this, further studies are needed, particularly investigating the Mdr1 mRNA and protein expression levels from brain material of affected goats. In conclusion, Mdr1 variants may exist not only in dogs, but also in individual goats. The current report provides the first Mdr1 mRNA transcript sequence of a goat and therefore represents the basis for more detailed Mdr1 sequence and expression analyses.

The Intestinal Efflux Transporter Inhibition Activity of Xanthones from Mangosteen Pericarp: An In Silico, In Vitro and Ex Vivo Approach

The capacity of α-mangostin (α-MG) and β-mangostin (β-MG) from mangosteen pericarp on P-glycoprotein (Pgp) in silico, in vitro, and ex vivo was investigated in this study. Screening with the ADMET Predictor™ program predicted the two compounds to be both a Pgp inhibitor and Pgp substrate. The compounds tended to interact with Pgp and inhibit Pgp ATPase activity. Additionally, bidirectional transport on Caco-2 cell monolayers demonstrated a significantly lower efflux ratio than that of the control (α-(44.68) and β-(46.08) MG versus the control (66.26); p < 0.05) indicating an inhibitory effect on Pgp activity. Test compounds additionally revealed a downregulation of MDR1 mRNA expression. Moreover, an ex vivo absorptive transport in everted mouse ileum confirmed the previous results that α-MG had a Pgp affinity inhibitor, leading to an increase in absorption of the Pgp substrate in the serosal side. In conclusion, α- and β-MG have the capability to inhibit Pgp and they also alter Pgp expression, which makes them possible candidates for reducing multidrug resistance. Additionally, they influence the bioavailability and transport of Pgp substrate drugs.

The Influence and Mechanism of Paeoniflorin and Albiflorin on Strychnine and Brucine Transport Through Madin-Darby Canine Kidney/Multidrug Resistance1 Cells In Vitro Model

This research sought to study the influence and potentialmechanism of paeoniflorin and albiflorin on strychnine and brucine transport in MDCK-MDR1 cells regulated by P-gp. Cytotoxicity of drugs was tested by MTT assay, and the transport studies were performed in both directions in MDCK-MDR1 cells. The influence of drugs on P-gp ATPase, and the efflux function of P-gp, the expression levels of P-gp and MDR1 mRNA were also estimated. Strychnine and brucine showed well absorption, and the main transport mechanism might be passive diffusion. Verapamil could significantly decrease the efflux rate (ER) of strychnine and brucine, while the ER of strychnine and brucine was increased significantly when co-administrated with paeoniflorin or albiflorin, indicating that paeoniflorin and albiflorin could promote the efflux of these two alkaloids. Strychnine and brucine could activate the activity of P-gp ATPase, suppress the efflux function of P-gp, but have no significant effect on the expression of P-gp. In addition, strychnine could upregulate the expression of MDR1 mRNA. Paeoniflorin and albiflorin could increase the transmembrane transport of strychnine and brucine mediated by P-gp when co-administrated with strychnine or brucine via stimulating the activity of P-gp ATPase, enhancing the efflux function of P-g, increasing the expression levels of MDR1 mRNA and P-gp.

Transport Oligonucleotides—A Novel System for Intracellular Delivery of Antisense Therapeutics

Biological activity of antisense oligonucleotides (asON), especially those with a neutral backbone, is often attenuated by poor cellular accumulation. In the present proof-of-concept study, we propose a novel delivery system for asONs which implies the delivery of modified antisense oligonucleotides by so-called transport oligonucleotides (tON), which are oligodeoxyribonucleotides complementary to asON conjugated with hydrophobic dodecyl moieties. Two types of tONs, bearing at the 5′-end up to three dodecyl residues attached through non-nucleotide inserts (TD series) or anchored directly to internucleotidic phosphate (TP series), were synthesized. tONs with three dodecyl residues efficiently delivered asON to cells without any signs of cytotoxicity and provided a transfection efficacy comparable to that achieved using Lipofectamine 2000. We found that, in the case of tON with three dodecyl residues, some tON/asON duplexes were excreted from the cells within extracellular vesicles at late stages of transfection. We confirmed the high efficacy of the novel and demonstrated that MDR1 mRNA targeted asON delivered by tON with three dodecyl residues significantly reduced the level of P-glycoprotein and increased the sensitivity of KB-8-5 human carcinoma cells to vinblastine. The obtained results demonstrate the efficacy of lipophilic oligonucleotide carriers and shows they are potentially capable of intracellular delivery of any kind of antisense oligonucleotides.

Study on the Administration Method of Combining Tetrandrine with Adriamycin to Reverse Drug-Resistance of Leukemia

Increasing drug efflux pumps (P-gp) on the cell membrane due to the overexpression of MDR1 gene in tumor cells is the main mechanism of multidrug resistance of cancer cells. P-gp belongs to the ATP-binding cassette family and functions as a transmembrane efflux pump that translocates chemotherapy drugs from an intracellular to an extracellular domain; thus, it is impossible to achieve efficient drug accumulation in tumor cells. Tetrandrine (TTD) has been shown in both in vitro and in animal experiments to reverse MDR by reducing the expression of MDR1 mRNA and P-gp. However, the effect of administration of TTD one week before chemotherapy is not ideal in the current clinical trials. In view of the previous in vitro and animal experiments have confirmed the effect of TTD in reversing the drug resistance of leukemia, this study returned to the cell experiment, selected K562 and K562/ADM cell lines as the research object, through different ADM combined with TTD administration sequences, to detect the inhibitory effect of different administration sequences on cell proliferation and whether it affected the intracellular ADM concentration, P- gp ATPase activity, MDR1 mRNA and P-gp expression levels, comparing the effect of TTD combined with ADM different administration sequences on the reversal of MDR in K562/ADM cells. We found that TTD can significantly antagonize P-gp-mediated ADM resistance by competitively binding P-gp and down-regulating P-gp expression and we, therefore, conclude that the co-administration of TTD, as a drug resistance reversal agent with ADM may be a better administration in the clinic.