Extracurricular laboratory:new discovery of (S)-Bis(3,5-bis(trifluoromethyl)phenyl)(pyrrolidin-2-yl)methanol

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Arctic ground squirrel resist peroxynitrite-mediated cell death in response to oxygen glucose deprivation

Cerebral ischemia-reperfusion (I/R) injury initiates a cascade of events, generating nitric oxide (NO) and superoxide(O2?-) to form peroxynitrite (ONOO-), a potent oxidant. Arctic ground squirrels (AGS; Urocitellus parryii) show high tolerance to I/R injury. However, the underlying mechanism remains elusive. We hypothesize that tolerance to I/R modeled in an acute hippocampal slice preparation in AGS is modulated by reduced oxidative and nitrative stress. Hippocampal slices (400 mum) from rat and AGS were subjected to oxygen glucose deprivation (OGD) using a novel microperfusion technique. Slices were exposed to NO, O2- donors with and without OGD; pretreatment with inhibitors of NO, O2- and ONOO- followed by OGD. Perfusates collected every 15 min were analyzed for LDH release, a marker of cell death. 3-nitrotyrosine (3NT) and 4-hydroxynonenal (4HNE) were measured to assess oxidative and nitrative stress. Results show that NO/O2- alone is not sufficient to cause ischemic-like cell death, but with OGD enhances cell death more in rat than in AGS. A NOS inhibitor, SOD mimetic and ONOO- inhibitor attenuates OGD injury in rat but has no effect in AGS. Rats also show a higher level of 3NT and 4HNE with OGD than AGS suggesting the greater level of injury in rat is via formation of ONOO-.

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Metal catalyst and ligand design,
Ligand Template Strategies for Catalyst Encapsulation – NCBI

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In vivo cultivation of tumor cells in hollow fibers

Advancement of potential anti-cancer agents from ‘discovery’ in an in vitro screen to pre-clinical development requires a demonstration of in vivo efficacy in one or more animal models of neoplastic disease. Most such models require considerable materials in terms of laboratory animals and test compound as well as substantial amounts of time (and cost) to determine whether a given experimental agent or series of agents have even minimal anti-tumor activity. The present study was initiated to assess the feasibility of employing an alternate methodology for preliminary in vivo evaluations of therapeutic efficacy. Results of experimentation to date demonstrate that a hollow fiber encapsulation/implantation methodology provides quantitative indices of drug efficacy with minimum expenditure of time and materials. Following further pharmacologic calibrations, the hollow fiber technique is anticipated (a) to identify compounds having moderate to prominent anti-cancer activity and (b) to facilitate the identification of sensitive tumor cell line ‘targets’ and optimal or near-optimal treatment regimens for subsequent testing using standard in vivo solid tumor models. The potential suitability of this methodology is demonstrated with several standard anti-neoplastic agents.

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Metal catalyst and ligand design,
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Chemical-induced inhibition of blue light-mediated seedling development caused by disruption of upstream signal transduction involving cryptochromes in Arabidopsis thaliana

Plants have a remarkable ability to perceive and respond to various wavelengths of light and initiate regulation of different cascades of light signaling and molecular components. While the perception of red light and the mechanisms of its signaling involving phytochromes are largely known, knowledge of the mechanisms of blue light signaling is still limited. Chemical genetics involves the use of diverse small active or synthetic molecules to evaluate biological processes. By combining chemicals and analyzing the effects they have on plant morphology, we identified a chemical, 3-bromo-7-nitroindazole (3B7N), that promotes hypocotyl elongation of wild-type Arabidopsis only under continuous blue light. Further evaluation with loss-of-function mutants confirmed that 3B7N inhibits photomorphogenesis through cryptochrome-mediated light signaling. Microarray analysis demonstrated that the effect of 3B7N treatment on gene expression in cry1cry2 is considerably smaller than that in the wild type, indicating that 3B7N specifically interrupts cryptochrome function in the control of seedling development in a light-dependent manner. We demonstrated that 3B7N directly binds to CRY1 protein using an in vitro binding assay. These results suggest that 3B7N is a novel chemical that directly inhibits plant cryptochrome function by physical binding. The application of 3B7N can be used on other plants to study further the blue light mechanism and the genetic control of cryptochromes in the growth and development of plant species.

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Metal catalyst and ligand design,
Ligand Template Strategies for Catalyst Encapsulation – NCBI

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Isoform specific inhibition for treatment of pain and reduction of anesthetic threshold

Several lines of evidence have shown a role for the nitric oxide (NO)/cyclic guanosine monophosphate (cGMP) signaling pathway in the development of spinal hyperalgesia. However, the roles of effectors for cGMP are not fully understood in the processing of pain in the spinal cord. cGMP-dependent protein kinase (PKG) Ialpha but not PKGIbeta was localized in the neuronal bodies and processes, and was distributed primarily in the superficial laminae of the spinal cord. Intrathecal administration of an inhibitor of PKGIalpha, Rp-8-[(4-Chlorophenyl)thio]-cGMPS triethylamine, produces significant antinociception. Moreover, PKGIalpha protein expression was dramatically increased in the lumbar spinal cord after noxious stimulation. This upregulation of PKGIalpha expression was completely blocked not only by a neuronal NO synthase inhibitor, and a soluble guanylate cyclase inhibitor, but also by an N-methyl-D-aspartate (NMDA) receptor antagonist, MK-801. Noxious stimulation not only initially activates but also later upregulates PKGIalpha expression in the superficial laminae via an NMDA-NO-cGMP signaling pathway, suggesting that PKGIalpha plays an important role in the central mechanism of inflammatory hyperalgesia in the spinal cord.

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Metal catalyst and ligand design,
Ligand Template Strategies for Catalyst Encapsulation – NCBI