Rhodamine 123 (chloride)

Effect of hyperthermia on the action of cis-diamminedichloroplatinum(II), rhodamine 123(2) [tetrachloroplatinum(II)], rhodamine 123, and potassium tetrachloroplatinate in vitro and in vivo

Platinum rhodamine 123 [Pt(Rh-123)2] was synthesized in an effort to produce a new drug which would have the selective uptake into carcinoma cells of Rh-123 and the alkylating and radiosensitizing properties of the chloroplatinum moiety. Because both Rh-123 and cis-diamminedichloroplatinum(II) (CDDP) have been shown to become more cytotoxic at elevated temperatures, we tested the interactions between Pt(Rh-123)2 and hyperthermia both in EMT6 cells in vitro and in the Lewis lung carcinoma in vivo. In the EMT6 cells, CDDP was far more cytotoxic than Pt(Rh-123)2 at 37 degrees C, but its cytotoxicity was less enhanced by exposure of cells to the drug at 42 degrees C than was true for Pt(Rh-123)2 [about 2 logs of increased killing at 42 degrees C after exposure to 10 microM CDDP versus over 3 logs of increased killing at 42 degrees C after exposure to 500 microM Pt(Rh-123)2]. Both Rh-123 and K2PtCl4 also are more cytotoxic to EMT6 cells at 42 degrees C than at 37 degrees C, but the hyperthermic enhancement was far less striking. In the Lewis lung carcinoma, the growth delay produced by CDDP (8 mg/kg) increased by a factor of approximately 2.5 when the drug was given i.p. just prior to local heating of the s.c. thigh tumor to 43 degrees C for 30 min, but the growth delay produced by Pt(Rh-123)2 (100 mg/kg) given i.p. 1 h before local hyperthermia increased by a factor of 5. In contrast, K2PtCl4 and Rh-123 given i.p. produced very short growth delays at normal temperatures and these growth delays were not enhanced by hyperthermia. The effect of these drugs at 37 degrees C and 42 degrees C on the conformation of superhelical pBR322 DNA was also examined. Exposure to CDDP caused progressive alteration from the supercoiled to the linear form of the DNA over time. In contrast, Pt(Rh-123)2 apparently produced progressive degradation of the DNA. Hyperthermia did not alter the qualitative damage produced by the drugs but increased the rate at which the changes occurred. These results suggest both that Pt(Rh-123)2 probably has a different mechanism of action at the DNA than does CDDP and that Pt(Rh-123)2 may be a good drug to use with local hyperthermia and radiation.

Determination of Rhodamine 123 in cell lysate by HPLC with visible wavelength detection

Rhodamine 123 (R123) is widely used to quantify P-glycoprotein (P-GP) functional efflux activity in vitro. We developed a rapid and specific high-performance liquid chromatography (HPLC) method to quantify Rhodamine 123 for use in experimental cell culture studies. The R123 standards (2.5-250 ng/mL) and quality controls (QCs) (5, 75, 200 ng/mL) were prepared in cell lysis buffer consisting of 0.75% Triton 100X and 0.2% sodium chloride. The mobile phase consisted of acetonitrile, 1.5 mM tetrabutyl ammonium bromide in 20mM sodium acetate buffer (pH 4.0) (50:20:30) delivered at a rate of 1.0 mL/min. Samples (50 microl) were injected onto a C(18) reversed-phase HPLC column with detection at 500 nm. Analyte retention times were 1.4 and 4.3 min for R123 and internal standard (R6G), respectively. Intra- and inter-day coefficients of variation were < or = 4.2%. Samples were stable for at least three freeze-thaw cycles at room temperature for 24 and 48 h. This method was used to evaluate the functional activity of P-glycoprotein in renal tubule cell models including human kidney (HK-2), Madin-Darby canine kidney (MDCK) and multi-drug resistance gene-transfected MDCK cells (MDR1-MDCK).

Two rhodamine derivatives: 9-[2-(ethoxycarbonyl)phenyl]-3,6-bis-(ethylamino)-2,7-dimethylxanthylium chloride monohydrate and 3,6-diamino-9-[2-(methoxycarbonyl)-phenyl]xanthylium chloride trihydrate

The title compounds, C28H31N2O3(+)-Cl(-)-H2O (common name rhodamine-6g), (I), and C21H17N2O3(+)-Cl(-)-3H2O (common name rhodamine-123), (II), both have planar xanthene skeletons with a formal +1 charge on the amino N atoms delocalized through the pi-electron system so that the N-Csp(2) bond distances indicate significant double-bond character. The substituted planar phenyl groups make angles of 63.29 (8) and 87.96 (11) degrees with the xanthene planes in (I) and (II), respectively. In both molecules, the carbonyl bond vectors point toward the xanthene rings. The ethylamine groups in (I) are oriented similarly with their CH2-CH3 bond vectors pointing nearly perpendicular to the xanthene plane. The chloride ions and water molecules are disordered in both structures. In (I), the chloride ion and water molecule are disordered between two sites. One water and chloride alternately occupy the same site with occupancy factors of 0.5. The other 0.5-chloride and 0.5-water occupy two distinct positions separated by 0.747 (8) A. In (II), the chloride ion is disordered between three sites and one of the waters is disordered about two other sites. Both crystal structures are stabilized by hydrogen bonds involving the chloride ions, amino groups and water molecules, as well as by pi-pi stacking between xanthene planes.

Mitochondrial activity as an indicator of fish freshness

The current methods used to routinely assess freshness in the fishing industry reflect more a state of spoilage than a state of freshness. Mitochondria, the seat of cellular respiration, undergo profound changes in post mortem tissues. The objective of this study was to demonstrate that mitochondrial activity constitutes a putative early fish freshness marker. The structure of gilthead sea bream (Sparus aurata) muscle tissue was evaluated over time by transmission electron microscopy. Respiration was assessed in mitochondria isolated from sea bream fillets using oxygraphy. Membrane potential (ΔΨ_m) was determined by fluorescence (Rhodamine 123). Mitochondrial activity of fillets stored at +4 °C was studied for 6 days. Changes in mitochondrial cristae structure appeared from Day 3 highlighting the presence of dense granules. ΔΨ_m and mitochondrial activity were significantly disrupted in sea bream fillets after 96 h of storage at +4 °C. Mitochondrial activity constituted a reliable and early indicator of fish freshness.

Propofol protects PC12 cells from cobalt chloride-induced injury by mediating miR-134

Objective: Propofol (PRO) was reported to exert a neuroprotective effect by decreasing microRNA-134 (miR-134), a brain-specific miRNA, thus, the role of PRO against cobalt chloride (CoCl?)-induced injury in rat pheochromocytoma cells (PC12) via mediating miR-134 was explored.
Methods: CoCl?-induced PC12 cells treated with PRO were transfected with or without miR-134 negative control (NC)/ inhibitor/mimic, and the following detections were then performed using cell counting kit-8 (CCK-8), Annexin V-fluorescein isothiocyanate/propidium iodide (Annexin V-FITC/PI) and Hoechst 33258 staining. Autophagy was observed by transmission electron microscope (TEM). Mitochondrial membrane potential (MMP) was detected by Rhodamine-123 (Rh123) staining, and reactive oxygen species (ROS) by dichloro-dihydro-fluorescein diacetate (DCFH-DA) staining. Protein and gene expressions were measured by Western blotting and quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), respectively.
Results: PRO reversed the CoCl?-induced decrease in the PC12 cell viability and increased miR-134 in a dose-dependent manner. CoCl? increased LC3II/I ratio and Beclin-1 expression, but decreased p62 expression, which was abolished by PRO. In addition, an increased cell apoptosis rates triggered by CoCl? were reduced by PRO with the down-regulations of Bax and Caspase-3 and the up-regulation of Bcl-2. Furthermore, PRO decreased methylenedioxyamphetamine (MDA), nitric oxide (NO) and ROS in CoCl?-induced PC12 cells accompanying the increase in glutathione peroxidase (GSH-Px) and MMP. The effects of PRO on autophagy, apoptosis and oxidative stress in CoCl?-induced PC12 cell were reversed by miR-134 mimic.
Conclusion: PRO may mitigate CoCl?-induced autophagy in PC12 cells with decreased apoptosis and improved oxidative stress via mediating miR-134.