Time-temperature superposition and the controlling role of solvation in the viscoelastic properties of polyaniline thin films

Comprehensive exploration of the viscoelastic properties of polyaniline films exposed to aqueous perchloric acid has been made as a function of applied potential (E), temperature (T), and mechanical oscillation frequency (f = ω/2π) using high-frequency acoustic wave resonators. The outcomes are expressed in terms of storage and loss shear modulus signatures, G'(E, T, ω) and G″(E, T, ω). Surprisingly, these are barely sensitive to potential, through which both polymer charge and solvation are manipulated, and only modestly sensitive to temperature. In contrast, the response to timescale is dramatic. Using the principle of time-temperature superposition, G' and G″ at different temperatures and frequencies (time scales) can each be placed on master relaxation curves. Models developed for mechanical properties of bulk polymers at low frequency were applied to these thin film responses at high frequency. These include the Williams-Landel-Ferry model, the activation model, and the Rouse-Zimm model based, respectively, on concepts of free volume, thermal activation, and relaxation. Each of the models could be applied with physically reasonable outcomes in terms of the relevant parameters (thermal expansion coefficient, glass transition temperature, and activation enthalpy). G' and G″ values are correlated with solvent content. The enthalpy change for solvent entry is small, positive and relatively independent of polymer charge state, all of which contrast sharply with the behavior of thiophene-based conducting polymers in organic solvents.     

Intravenous RMP-7 increases delivery of ganciclovir into rat brain tumors and enhances the effects of herpes simplex virus thymidine kinase gene therapy

Herpes simplex virus thymidine kinase (HSV-tk) gene therapy for brain tumors depends on ganciclovir (GCV) and its transport across the blood-brain tumor barrier (BBTB). We examined whether RMP-7, the bradykinin analog and potent BBTB permeabilizer, could enhance the efficacy of GCV treatment of brain tumors by increasing the BBTB delivery of GCV. In vitro, a significant bystander cytocidal effect of GCV was shown in mixed HSV-tk-transduced (HSV-tk+) and control vector-transduced (HSV-tk-) C6 glioma cultures. A dose-dependent cytotoxic effect of GCV on untransformed C6 cells was also shown. In vivo, rats with 100% HSV-tk+ or 100% HSV-tk- intracerebral C6 gliomas were treated for 7 days with intravenous infusions of GCV alone or with GCV and RMP-7 (2.5 microg/kg/day). The growth of HSV-tk+ and HSV-tk- gliomas decreased with increasing doses of GCV. A high dosage (100 mg of GCV/kg/day) eradicated all HSV-tk- and HSV-tk+ tumors. An intermediate dosage (5 mg of GCV/kg/day) reduced the growth of HSV-tk- gliomas by 42% if given alone, and by 88% in combination with RMP-7. A low dosage (0.5 mg of GCV/kg/day) in combination with RMP-7 enhanced the regression of HSV-tk+ gliomas by 87% compared with GCV alone. Low-dose GCV was ineffective in HSV-tk- tumors. RMP-7 increased [3H] GCV tumoral uptake by 2.6- and 1.7-fold in the tumor center and periphery, respectively. We conclude that RMP-7 could be an important adjunctive treatment for suicide gene therapy of brain tumors, while an RMP-7/GCV combination may also have a significant antitumor effect in untransfected gliomas.     

Retinal cholinergic system: characterization of rat retinal acetyltransferases using specific inhibitors of choline- and carnitine-acetyltransferases

Choline acetyltransferase catalyzes the synthesis of acetylcholine from choline and acetylcoenzyme A (ACoA) in both nervous and non-nervous tissues. Carnitine acetyltransferase occurs in several tissues and transfers acetyl groups from ACoA to carnitine forming acetylcarnitine and exhibits weak choline acetyltransferase activity. Several haloacetylcholines and haloacetylcarnitines were synthesized to develop selective inhibitors of choline acetyltransferase and carnitine acetyltransferase. Acetylcholine is a transmitter for some presynaptic neurons and/or amacrine cells in retina. Selective inhibitors of choline acetyltransferase and carnitine acetyltransferase were used in the evaluation of choline acetyltransferase and carnitine acetyltransferase activities in the rat retina. Choline acetyltransferase and carnitine acetyltransferase activities were assayed by transferring of [14C]acetyl group from [14C]ACoA to choline or carnitine and estimating [14C]-acetylcholine or [14C]acetylcarnitine. This study gave the following results: (a) Bromoacetylcholine (BrACh) was a selective inhibitor of purified choline acetyltransferase (I50, 2.2 microM); (b) (R)-bromoacetylcarnitine [(R)-BrACa] was more potent for inhibiting purified carnitine acetyltransferase (I50, 4 microM) than purified choline acetyltransferase (I50, 46 microM); (c) Rat retinal sonicate gave choline acetyltransferase activity of 98 +/- 6 nmol of ACh formed/mg/10 min. When the carnitine acetyltransferase was completely inhibited by (R)-BrACa, the activity for choline acetyltransferase decreased to 47 +/- 1 nmol, and this decrease was possibly due to the formation of some [14C]acetylcholine by carnitine acetyltransferase. The net retinal choline acetyltransferase activity was 51 nmol acetylcholine/mg protein/10 min; (d) Rat retinal sonicate contained carnitine acetyltransferase activity of 102 +/- 7 nmol acetylcarnitine formed/mg protein/10 min. This was not altered by inhibition of choline acetyltransferase with BrACh. This means that choline acetyltransferase did not use carnitine as a substrate. Choline acetyltransferase and carnitine acetyltransferase activities did not change after dialysis of retinal sonicates at 4 degrees C for 24 hrs. These observations suggest that BrACh and (R)-BrACa are useful for assessing the correct values for choline acetyltransferase and carnitine acetyltransferase activities in retinal tissues.     

Influence of nicotine and cotinine on retinal phospholipase A2 and its significance to macular function

The macula is a constituent of the sensory retina that is necessary for sharp contrast and color vision. A significant relationship has been found between tobacco smoking and age-related macular degeneration. Opsin, rhodopsin and phospholipase A2 (PLA2) are located in excitable membranes of retina which are enriched with polyunsaturated fatty acids (PUFA). A question may arise as to whether nicotine and its major metabolite cotinine influence PLA2 so that arachidonic acid (AA) and proinflammatory prostaglandins (PG) are produced in the retina. Therefore, the effects of nicotine and cotinine on the retinal PLA2 were studied. PLA2 activity of rat retinal sonicates was assayed using 1-palmitoyl-2[1-14C]arachidonyl-Phosphatidylethanolamine (PE, 2.2 nmol) as a substrate in Tris buffer (10 mM, pH 7.4) at 37 degrees C with and without nicotine or cotinine in the assay medium. These studies gave the following results: (1) Rat retinal PLA2 activity was 4.2+/-0.8 pmol PE hydrolyzed/100 ng protein/hr. (2) Nicotine in low concentrations (1-150 nM) activated PLA2 (EC50 5 nM). (3) Cotinine also activated PLA2 (EC50 300 nM). (4) Only high concentrations of nicotine (> 1.0 microM) and cotinine (> 25 microM) exhibit inhibition of PLA2. (5) All three known PLA2 inhibitors, mepacrine, 4-bromophenacyl bromide and bromoacetylcholine bromide (IC50: 0.5mM, 88 microM, 30 mM, respectively) inhibited retinal PLA2 activity. These observations suggest that polyunsaturated fatty acids are cleaved, and arachidonic acid, the precursor for prostaglandins and related pro-inflammatory mediators, is liberated by nicotine and cotinine. Oxidative stress (reduced levels of antioxidants), vascular insufficiency, as well as activation of PLA2 by nicotine and cotinine may contribute for retinal degeneration in smokers during aging.