Enzymatic and Immunologic Identification of Succinic Semialdehyde Dehydrogenase in Rat and Human Neural and Nonneural Tissues

Abstract: We have identified succinic semialdehyde dehydrogenase protein in rat and human neural and nonneural tissues. Tissue localization was determined by enzymatic assay and by western immunoblotting using polyclonal antibodies raised in rabbit against the purified rat brain protein. Although brain shows the highest level of succinic semialdehyde dehydrogenase activity, substantial amounts of enzyme activity occur in mammalian liver, pituitary, heart, and ovary. We further demonstrate the absence of succinic semialdehyde dehydrogenase enzyme activity and protein in brain, liver, and kidney tissue samples from an individual affected with succinic semialdehyde dehydrogenase deficiency, thereby verifying the specificity of our antibodies.     

Estimating processing time of stream benthic samples

Multiple regression analysis was used to generate equations relating time required to sort and tabulate benthic invertebrates to size characteristics of benthic samples collected from a 5th-order river and a 3rd-order woodland stream. Number of invertebrates in a sample was the primary determinant of sample sorting time. Amount of detritus, occurrence of filamentous algae (Cladophora) and source stream contributed significant but minor additional variability to estimates (cumulativeR ² = 0.95). When number of animals was excluded as a variable, amount of detritus and presence/absence ofCladophora alone could be used to predict sorting time (R ² = 0.81). A typical sample containingCladophora required 29% longer to sort than samples containing equivalent amounts of organic material but noCladophora. The influence of sampler size and subsampling on processing and sorting time are considered. A general equation was derived to provide guidelines for selecting a size of stream sampler that subsequently minimizes total processing time required to estimate density of benthos with acceptable, constant precision. Overall sample processing and sorting time are reduced by using a smaller sampler or by subsampling only if benthic densities of animals are high. Use of regression equations to anticipate processing and sorting time required for a particular sampling program permits development of more efficient designs.     

Novel inhibitors of leukocyte transendothelial migration

Leukocyte transendothelial migration is one of the most important step in launching an inflammatory immune response and chronic inflammation can lead to devastating diseases. Leukocyte migration inhibitors are considered as promising and potentially effective therapeutic agents to treat inflammatory and auto-immune disorders. In this study, based on previous trioxotetrahydropyrimidin based integrin inhibitors that suboptimally blocked leukocyte adhesion, twelve molecules with a modified scaffold were designed, synthesized, and tested in vitro for their capacity to block the transendothelial migration of immune cells. One of the molecules, namely, methyl 4-((2-(tert-butyl)-6-((2,4,6-trioxotetrahydropyrimidin-5(2H)-ylidene) methyl) phenoxy) methyl) benzoate, (compound 12), completely blocked leukocyte transendothelial migration, without any toxic effects on immune or endothelial cells (IC₅₀ = 2.4 µM). In vivo, compound 12 exhibited significant therapeutic effects in inflammatory bowel disease (IBD)/Crohn’s disease, multiple sclerosis, fatty liver disease, and rheumatoid arthritis models. A detailed acute and chronic toxicity profile of the lead compound in vivo did not reveal any toxic effects. Such a type of molecule might therefore provide a unique starting point for designing a novel class of leukocyte transmigration blocking agents with broad therapeutic applications in inflammatory and auto-immune pathologies.