Molecular pharmacology
-
Molecular pharmacology · Jul 2006
Comparative StudyHigh accumulation of platinum-DNA adducts in strial marginal cells of the cochlea is an early event in cisplatin but not carboplatin ototoxicity.
Ototoxicity is a typical dose-limiting side effect of cancer chemotherapy with cisplatin but much less so with carboplatin. To elucidate the underlying molecular pathological mechanisms, we have measured the formation and persistence of drug-induced DNA adducts in the nuclei of inner ear cells of guinea pigs after short-term exposure to either cisplatin or carboplatin using immunofluorescence staining and quantitative image analysis. After application of carboplatin, all cells of the cochlea exhibited a similar burden of guanine-guanine intrastrand cross-links in DNA. ⋯ Because cisplatin ototoxicity is often attributed to oxidative stress mediated by the generation of radical oxygen species (ROS), we have measured in parallel the levels of the lead DNA oxidation product 8-oxoguanine (8-oxoG) in cochlear cryosections. Compared with basal levels in untreated control cochleas, no additional formation of 8-oxoG was detectable up to 48 h after cisplatin treatment in the DNA of either inner-ear cell type. This suggests that the generation of ROS may be a secondary event in cisplatin ototoxicity.
-
Molecular pharmacology · Jul 2006
Subtype specificity of scorpion beta-toxin Tz1 interaction with voltage-gated sodium channels is determined by the pore loop of domain 3.
Voltage-gated sodium (Nav) channels are modulated by a variety of specific neurotoxins. Scorpion beta-toxins affect the voltage-dependence of channel gating: In their presence, Nav channels activate at subthreshold membrane voltages. Previous mutagenesis studies have revealed that the beta-toxin Css4 interacts with the extracellular linker between segments 3 and 4 in domain 2 of Nav channels with the effect to trap this voltage sensor in an open position (Neuron 21: 919-931, 1998 ). ⋯ Analysis of channel chimeras in which whole domains of Nav1.2 were inserted into a Nav1.4 background revealed that the Nav1.2 phenotype was not conferred to Nav1.4 by domain 2 but by domain 3. The interaction epitope could be narrowed down to residues Glu1251, Lys1252, and His1257 located in the C-terminal pore loop in domain 3. The receptor site for beta-toxin interaction with Nav channels thus spans domains 2 and 3, where the pore loop in domain 3 specifies the pharmacological properties of individual neuronal Nav channel types.
-
Molecular pharmacology · Mar 2006
Nitric oxide-dependent reduction in soluble guanylate cyclase functionality accounts for early lipopolysaccharide-induced changes in vascular reactivity.
We investigated the role of soluble guanylate cyclase in lipopolysaccharide-induced hyporesponsiveness to phenylephrine. The effects of phenylephrine on the blood pressure of female Wistar rats were evaluated at 2, 8, and 24 h after lipopolysaccharide injection (12.5 mg/kg i.p.). Vasoconstrictive responses to phenylephrine were reduced 40 to 50% in all time periods. ⋯ Guanylate cyclase protein levels were lower than controls in lungs harvested from rats injected 8 h earlier and were back to normal values in lungs of rats injected 24 h earlier with lipopolysaccharide. Thus, data indicate that there is a temporal window of 8 h after lipopolysaccharide injection in which soluble guanylate cyclase is not functional and that this loss of function is NO-dependent. Thus, the putative use of soluble guanylate cyclase inhibitors in the treatment of endotoxemia may be beneficial mainly at early stages of this condition.
-
Molecular pharmacology · Mar 2006
Evidence for a multivalent interaction of symmetrical, N-linked, lidocaine dimers with voltage-gated Na+ channels.
The interaction of symmetrical lidocaine dimers with voltage-gated Na+ channels (VGSCs) was examined using a FLIPR membrane potential assay and voltage-clamp. The dimers, in which the tertiary amines of the lidocaine moieties are linked by an alkylene chain (two to six methylene units), inhibited VGSC activator-evoked depolarization of cells heterologously-expressing rat (r) Na(v)1.2a, human (h) Na(v)1.5, and rNa(v)1.8, with potencies 10- to 100-fold higher than lidocaine (compound 1). The rank order of potency (C4 (compound 4) > C3 (compound 3) > or = C2 (compound 2) = C5 (compound 5) = C6 (compound 6) > compound 1) was similar at each VGSC. ⋯ The observation that both the potency and dissociation rate of the dimers was dependent upon linker length is consistent with a multivalent interaction at VGSCs. hNa(v)1.5 VGSCs did not recover from inhibition by compound 4. However, "chase" with free local anesthetic site inhibitors increased the rate of dissociation of compound 4. Together, these data support the hypothesis that compound 4 simultaneously occupies two binding sites on VGSCs, both of which can be bound by known local anesthetic site inhibitors.
-
Molecular pharmacology · Mar 2006
Janus kinase-signal transducer and activator of transcription mediates phosphatidic acid-induced interleukin (IL)-1beta and IL-6 production.
We have found previously that phosphatidic acid (PA) can induce inflammatory mediators such as cytokines, which implies that PA plays a role in inflammatory response. In the present study, we provide evidence of the PA-mediated activation of the Janus tyrosine kinase (JAK)-signal transducer and activator of transcription (STAT) signaling pathway, which results in the production of interleukin (IL)-1beta and IL-6. PA elicited the rapid phosphorylations of JAK2 and STAT1/3, and the subsequent nuclear translocation. ⋯ The knockdown of JAK2 in macrophages by small interfering RNA significantly attenuated PA-induced IL-1beta and IL-6 production. In addition, JAK2 inhibitor suppressed PA-induced Akt phosphorylation, and the Akt inhibitor 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002) blocked GAS activation (GAS contains a promoter that responds to PA), suggesting that PA-mediated JAK2 activation leads to phosphatidylinositol 3-kinase/Akt phosphorylation and STAT activation, and the subsequent translocation of STAT to the nucleus. Together, our data demonstrate that PA-activated macrophages produce IL-1beta and IL-6 and that these processes require the activation of the JAK2-STAT1/3 or JAK2-Akt-STAT signaling pathways.