Ref. Accumulation/  Efflux Model Rho 123 Detection Study Aim Results
[97] LS-180
(48-well plate)		 Fluorometry Investigate 7 drugs according to their potencies to up-regulate P-gp activity. Verapamil enhanced Rho 123 accumulation factor.
P-gp activity increased after 48 h of pre-treatment with each test compound.
[98] MCF-7/ADR
(24-well plate)		 Fluorometry Investigate the effect of quercetin on P-gp. Intracellular accumulation of Rho 123 is:
4-fold higher in MCF-7 than in MCF-7/ADR;
2-fold higher in MCF-7/ADR cells pre-incubated with quercetin (10 µM, 24h.
[99] KB-V1 Flow Cytometry Investigate whether Kuguacin J from Momordica charantia leaves modules P-gp. Kuguacin J increased Rho 123 accumulation factor more than 2.5-fold.
[100] L5178-MDR1 Flow Cytometry Investigate whether TBN is a P-gp modulator. Accumulation factor of Rho 123 increased:
10-fold in presence of verapamil; 100-fold in presence of TBN.
[101] PBMEC
(96-well plate)		 Fluorometry Examine P-gp functionality in the immortalized PBMEC/C1-2 cell line. Verapamil enhance uptake of Rho 123 in PBMEC/C1-2;
After knock-down P-gp, Rho 123 uptake increased.
[102] Caco-2 Flow Cytometry Investigate in vitro the effect of duloxetine (10µM) on P-gp function. Accumulation factor of Rho 123 increased 130% in presence of duloxetine.
[103] Caco-2
(96-well plate)		 Flow Cytometry Investigate whether the piperidine alkaloid, lobeline, could inhibit P-gp and reverse P-gp dependent resistance. Rho 123 fluorescence in Caco-2 cells is positively correlated with increase of lobeline concentrations.
[104] LLC-PK1
& L-MDR1		 Flow Cytometry Investigate the effect of 7 antipsychotic drugs on P-gp activity in LLC-PK1 and L-MDR1 cells using Rho 123 as P-gp probe. Intracellular fluorescence of Rho 123 was 3.1-fold lower in L-MDR1.
PSC833 enhanced the accumulation factor of Rho 123 in L-MDR1.
Antipsychotic drugs decreased Rho 123 accumulation factor in L-MDR1.
Ref. Bi-directional Transport Rho 123 Detection Study Aim Results
[105] HBMEC
& Caco-2		 Fluorometry Investigate whether venlafaxine and desvenlafaxine are inducers of P-gp. Venlafaxin (1-50 µM) reduced Papp (A-B) of Rho 123 across HBMEC and Caco-2 by respectively 30% and 25%.
Desvenlafaxin (1-50 µM) had no effect on Rho 123 transport.
[75] Rat intestine membrane in Ussing Chamber Fluorometry Investigate the potential of a novel polymer (Thiolated PEG-PEI) to inhibit P-gp by evaluating their influence on transport of Rho 123. Papp (A-B) of Rho123 enhanced:
2-fold after verapamil addition (100 µM);
3.3-fold after thiolated polymer addition (0.5%, w/v).
R(B-A/A-B) of Rho 123 was reduced to 1.0, 1.4 and 2.0 after thiolated PEG-PEI applied at 0.1%, 0.25% and 0. 5% (w/v).
[106] Rat gastro-intestinal segments in Ussing Chamber Spectro-fluorimetry Investigate the influence of distinct pharmaceutical excipients on melatonin absorption. Rho 123 was used as positive control. Transport of Rho 123 in:
A-B direction: 1052.3±192.6 pg/mL; B-A direction: 3126.5±319.2 pg/mL
Melatonin transport in A-B direction was higher than that of Rho 123 and similar to that in B-A direction.
[107] Caco-2
(12-well plate)		 Fluorometry Understand the transport profiles of astilbin and taxifolin in Caco-2 cell model and their effect on P-gp. R(B-A/A-B) of Rho 123 ≈ 4.62;
After addition of astilbin or taxifolin (10-1000 µM):
R(B-A/A-B) varied from 4.29 to 4.88
After verapamil addition (100µM): R(B-A/A-B) ≈ 1.22
After pre-incubation with astilbin or taxifolin (1-100 µM) for 24 or 36 h: transport of Rho 123 in B-A direction increased significantly (p<0.05)
[108] hCMEC/D3
(12-well plate)		 Fluorometry Examine P-gp function in hCMEC/D3 cells to determine their suitability as in vitro human BBB model for identifying P-gp. Papp (A-B) of Rho123 enhanced:
67% by tariquidar (300 nM) and 55% by vinblastine (22 µM).
Papp (B-A) of Rho 123 was not altered.
[109] Rat small intestine everted sac HPLC-FL Investigate the effect of docosahexaenoic acid (DHA) on intestinal P-gp using Rho 123 as P-gp substrate. Transport of Rho 123 in A-B direction:
increased from 4.34±0.69 to 74.83±21.7 pmol/min/6cm with PSC833;
maintained in presence of DHA at concentration range (10-1000 µM).
[110] Rat small intestine everted sac Spectro-fluorimetry
HPLC-FL		 Investigate the effects of 50% ethyl acetate extracts of GFJ and OJ on the transport activity of P-gp in rat small intestine. Verapamil at 300 µM (p<0.01) increased Rho 123 efflux.
GFJ (p<0.05) and OJ (p<0.05) decreased Rho 123 efflux.
Ref. In vivo animal model Rho 123 Detection Study Aim Effects on pharmacokinetics of Rho 123
[83] Male Wistar Rats Fluorometry Investigate the effect of cyclosporin A on nifedipine efflux and pharmacokinetic characteristics. Rho 123 was used as positive control. Pre-treated rats with cyclosporin A were administered with Rho123 (0.2 mg/Kg/mL, IV). In comparison to untreated animals, Rho 123 presented:
t1/2 decreased by 19.2%; Cmax increased by 64.7%; AUC0-t increased by 26.5%; CLT decreased by 24.6%
Results for nifedipine were comparable to those from Rho 123.
[74] Male Sprague- Dawley Rats Fluorometry Provide a proof-of-principle for a delivery system based on thiolated chitosan in vivo in rats. AUC0-12 increased 217% by the delivery system based on thiolated chitosan.
Table 4: In vitro and in vivo assays using Rhodamine 123 (Rho 123) as a probe substrate and their characteristics. AUC, area under the curve; BCE, brain endothelial cell; FL, fluorescence; GFJ, grapefruit juice; HBMEC, human brain endothelial cells; hCMEC/D3, immortalized human BEC cell line; KB-V1, multidrug resistant cervical carcinoma cell line; LLC-PK1, porcine kidney tubular cell line; L-MDR1, porcine kidney tubular cell line transfected with human MDR1 gene; LS-180, human colorectal adencarcinoma cell line; L5178-MDR1, Mouse T-cell lymphoma drug resistant cell line transfected with human MDR1 gene; MCF-7/ADR, breast cancer cell line overexpressing P-gp; OJ, oranje juice; Papp, apparent permeability; R(B-A/A-B), efflux ratio given by the ratio between apparent permeability in efflux (B-A) and absorption (A-B) directions; TBN, N-tylosil-1-a-amino-(3-bromophenyl)-methyl-2-naphthol; tmax, time to reach maximum concentration (Cmax).
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