Meiotic activity in orthotopic xenografts derived from human postpubertal testicular tissue.

Grafting of frozen-thawed testicular tissue has been suggested as a novel fertility preservation method for patients undergoing gonadotoxic treatments. However, this technique still needs further optimization before any clinical application. So far, grafting of human testicular tissue has only been performed to the back skin of nude mice and has shown spermatogonial stem-cell survival and occasionally differentiation up to primary spermatocytes. In this study, orthotopic grafting to mouse testes was evaluated as an alternative, and the effect of freezing and the donor's age was studied.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe

Directed evolution of the substrate specificity of dialkylglycine decarboxylase

Dialkylglycine decarboxylase (DGD) is an unusual pyridoxal phosphate dependent enzyme that catalyzes decarboxylation in the first and transamination in the second half-reaction of its ping-pong catalytic cycle. Directed evolution was employed to alter the substrate specificity of DGD from 2-aminoisobutyrate (AIB) to 1-aminocyclohexane-1-carboxylate (AC6C). Four rounds of directed evolution led to the identification of several mutants, with clones in the final rounds containing five persistent mutations. The best clones show ~2.5-fold decrease in K(M) and ~2-fold increase in k(cat), giving a modest ~5-fold increase in catalytic efficiency for AC6C. Additional rounds of directed evolution did not improve catalytic activity toward AC6C. Only one (S306F) of the five persistent mutations is close to the active site. S306F was observed in all 33 clones except one, and the mutation is shown to stabilize the enzyme toward denaturation. The other four persistent mutations are near the surface of the enzyme. The S306F mutation and the distal mutations all have significant effects on the kinetic parameters for AIB and AC6C. Molecular dynamics simulations suggest that the mutations alter the conformational landscape of the enzyme, favoring a more open active site conformation that facilitates the reactivity of the larger substrate. We speculate that the small increases in k(cat)/K(M) for AC6C are due to two constraints. The first is the mechanistic requirement for catalyzing oxidative decarboxylation via a concerted decarboxylation/proton transfer transition state. The second is that DGD must catalyze transamination at the same active site in the second half-reaction of the ping-pong catalytic cycle

A New Fuzzy Multi-criteria Decision Making Approach: Extended Hierarchical Fuzzy Axiomatic Design Approach with Risk Factors

In recent years, Axiomatic Design (AD) has been widely used as a multi criteria decision making approach. AD approach compares the design objects and system capabilities in a framework and then selects the best alternative based on these comparisons. Some researchers then include fuzziness in the AD approach which helps to evaluate alternatives in fuzzy environments. The main advantage of fuzzy AD approach is the ability to evaluate both crisp and fuzzy values at the same time during decision process. However, these approaches are not appropriate for hierarchical decision problems. Therefore, these are extended to solve the hierarchical decision problems and Hierarchical Fuzzy Axiomatic Design Approach (HFAD) is presented. In this study, HFAD is extended to include risk factors for the first time in literature and a new approach called RFAD is proposed. Moreover, the application of the new approach is shown on a real world supplier selection problem and the results are compared to the other widely used decision making approaches in literature. © Springer International Publishing Switzerland 2014