A New Bis(aquated) High Relaxivity Mn(II) Complex as an Alternative to Gd(III)-Based MRI Contrast Agent

Disclosed here are a piperazine, a pyridine, and two carboxylate groups containing pentadentate ligand H₂pmpa and its corresponding water-soluble Mn­(II) complex (<b>1</b>). DFT-based structural optimization implied that the complex had pentagonal bipyramidal geometry where the axial positions were occupied by two water molecules, and the equatorial plane was constituted by the ligand ON₃O donor set. Thus, a bis­(aquated) disc-like Mn­(II) complex has been synthesized. The complex showed higher stability compared with Mn­(II)–EDTA complex [log<i>K</i>_MnL = 14.29(3)] and showed a very high <i>r</i>₁ relaxivity value of 5.88 mM^–1 s^–1 at 1.41 T, 25 °C, and pH = 7.4. The relaxivity value remained almost unaffected by the pH of the medium in the range of 6–10. Although the presence of 200 equiv of fluoride and bicarbonate anions did not affect the relaxivity value appreciably, an increase in the value was noticed in the presence of phosphate anion due to slow tumbling of the complex. Cell viability measurements, as well as phantom MR images using clinical MRI imager, consolidated the possible candidature of complex <b>1</b> as a positive contrast agent

Kinetics and mechanism of interaction of Pt(II) complex with bio-active ligands and <i>in vitro</i> Pt(II)-sulfur adduct formation in aqueous medium: bio-activity and computational study

<p>Kinetics of interaction between [Pt(pic)(H₂O)₂](ClO₄)₂, <b>2</b> (where pic = 2-aminomethylpyridine) with the selected ligands DL-methionine (DL-meth) and DL-penicillamine (DL-pen) have been studied spectrophotometrically in aqueous medium separately as a function of [<b>2</b>] as well as [ligand], pH and temperature at constant ionic strength. The association equilibrium constants (<i>K</i>_E) for the outer sphere complex formation have been evaluated together with the rate constants for the two subsequent steps. Activation parameters (enthalpy of activation ΔH^≠ and entropy of activation ΔS^≠) were calculated from the Eyring equation. An associative mechanism of substitution is proposed for both reactions on the basis of the kinetic observations, evaluated activation parameters, and spectroscopic data. Structural optimizations, HOMO-LUMO energy calculation, and Natural Bond Orbital (NBO) analysis of <b>2</b>–<b>4</b> were carried out with Density Functional Theory. Bonding mode of thiol and thio-ether is confirmed by spectroscopic analyses and NBO calculation. Cytotoxic properties of <b>2</b>–<b>4</b> were explored on A549 carcinoma cell lines; DNA-binding properties of the complexes were also investigated by gel electrophoresis.</p

Protective effects of flavonoids in DSS-induced colitis in mice.

<p>Histopathological examination of the colon of mice exposed to DSS and/or flavones (Baicalein, Baicalin) is illustrated. The histology (inset shows higher magnification views) represents the prototypical injury to the colon induced by DSS. <i>Pxr^+/+</i>: wild-type mice; <i>Pxr^−/−</i>: <i>Pxr</i>-null mice; <i>hPXR</i>: humanized PXR mice. –DSS: DSS untreated mice; +DSS: DSS treated mice; single black arrow: vehicle exposed mucosa; double black arrow: flavonoids exposed mucosa. Scale bar, 50 µM.</p

Baicalein, in contrast to baicalin, induces PXR and Cdx2 mRNA.

<p>(A) Scrambled or Cdx2 siRNA (si-Cdx2) transfected LS174T colon cancer cells were exposed to 0.1% DMSO (vehicle), baicalein (25 µM) or baicalin (25 µM) for 48 hours. mRNA levels of PXR were quantified by RT-qPCR. (B, top panel) Representative western blot of PXR from the same experiment as in (A). (B, bottom panel) Absolute band intensity was quantified for each lanes of the western blot as in figure (B, top panel), using Image J software. (C) Scrambled or PXR shRNA (shPXR) transduced LS174T cells, were exposed to 0.1% DMSO (vehicle), baicalein (25 µM) or baicalin (25 µM) for 48 hours. Cdx2 mRNA levels were quantified by RT-qPCR. (D, top panel) Representative western blot of Cdx2 from the same experiment as in (C). (D, bottom panel) Absolute band intensity was quantified for each lanes of the western blot as in figure (D, top panel), using Image J software. Histogram, mean ± SEM. *<i>P</i><.05; ** <i>P</i><.01;*** <i>P</i><.001; ns, not significant.</p

Baicalein, in contrast to baicalin, docks within the PXR ligand binding domain and induces PXR and Cdx2 in LS174T colon carcinoma cells.

<p>(A) Molecular docking of baicalein and baicalin to PXR ligand binding domain (LBD). Figure (i) represent molecular docking of baicalein to PXR LBD. The 2D schematic diagrams shown in (ii, baicalein) and (iii, baicalin) were generated using the LIGX module of MOE program. The binding site residues are colored by their nature, with hydrophobic residues in green, polar residues in purple and charged residues highlighted with bold contours. Blue spheres and contours indicate matching regions between ligand and receptors. Hydrogen bond interactions are shown by green and blue arrows for side chain and main chain interactions, respectively. (B) LS174T cells were exposed to 0.1% DMSO (vehicle) and baicalein (25 µM) or baicalin (25 µM) for 48 hours and total RNA was isolated for PXR mRNA expression analysis by real-time quantitative (RT-qPCR). (C & D) LS174T cells were exposed to different concentrations of baicalein or baicalin as illustrated, for 48 hours and total RNA isolated for Cdx2 mRNA expression analysis by (C) semi-quantitative polymerase chain reaction and by (D) RT-qPCR. β-actin was used as internal control. (E, top panel) Representative western blot of Cdx2 from the same experiment as in (C). (E, bottom panel) Absolute band intensity was quantified for each lanes of the western blot as in figure (E, top panel), using Image J software. Histogram, mean ± SEM. *<i>P</i><.01; ** <i>P</i><.001; ns, not significant.</p

Flavonoids induce Cdx2 mRNA and protein regardless of <i>Pxr</i> genotype in murine intestinal mucosa.

<p>(A & B) <i>Pxr</i>^+/+ and <i>Pxr</i>^−/− mice (DSS untreated) exposed to flavones (baicalein, baicalin), as represented in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036075#pone-0036075-g003" target="_blank">Figure 3</a>, were evaluated for (A) Cdx2 mRNA expression by RT-qPCR and (B, top panel) Cdx2 protein abundance by western blot. (B, bottom panel) Absolute band intensity was quantified for each lanes of the western blot as in figure (B, top panel), using Image J software. (C & D) <i>Pxr</i>^+/+ mice (DSS treated) pre-exposed to Inh1 and subsequently treated with flavones (baicalein, baicalin), as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036075#pone-0036075-g003" target="_blank">Figure 3</a>, were evaluated for (C) histological score and (D) Cdx2 mRNA expression (analyzed by RT-qPCR) in colon isolated on day 9 of DSS treatment. (E & F) <i>Pxr</i>^+/+ mice were treated with flavonoids (baicalein, baicalin) and mRNA expressions of <i>Mdr1</i> and <i>Cyp3a11</i> were analyzed in colon and liver. Flavonoids (baicalein, baicalin) have no effect on induction of (E) <i>Mdr1</i> and (F) <i>Cyp3a11</i> mRNA in <i>Pxr</i>^+/+ mice liver; however, they robustly induce gene expression in the colon. Inh1 abolishes baicalin’s effect on induction of <i>Mdr1</i> and <i>Cyp3a11</i> mRNA in colon, as assessed by RT-qPCR. Histogram, mean ± SEM. * <i>P</i><.05; ** <i>P</i><.01; ns, not significant.</p

Cdx2 binds to two specific endogenous binding sites on the proximal PXR promoter.

<p>(A) Schematic details of the sites (blue marker) that were identified with a high match [matrix scores (≥0.85)], represent potential functional Cdx2 binding sites on PXR promoter. (Note: three potential Cdx1 binding sites, which are also identified on analysis, were excluded from consideration). The <i>in silico</i> analysis identified a total of 19 potential Cdx1/2 binding sites, of which, we verified endogenous binding for two sites, BS1 and BS2 by (C) Chromatin Immunoprecipitation (ChIP) assays. BS, Cdx2 binding site. ChIP assays were performed with Cdx2 and non-specific IgG antibodies and Cdx2 binding was assessed on PXR promoter (BS1 and BS2), PXR intron (negative control) and Furin promoter (positive control) regions. (D) Pol II ChIP assays were performed, using chromatin extracted from LS174T cells that were exposed to scrambled or Cdx2 siRNA (si-Cdx2). Pol II occupancy was assessed on PXR proximal promoter (TATA box) region, using Pol II and non-specific IgG antibodies as a control. (E) Cdx2 transactivation assay was performed in 293T cells, transfected with ∼6 kb PXR promoter reporter constructs (with or without deletion and mutations, as shown in the figure) and Cdx2 expression plasmid. Data expressed as RLU. RLU, relative light unit. Histogram, mean ± SD.</p