As companies strive to develop artefacts intended for services instead of traditional sell-off, new challenges in the product development process arise to promote continuous improvement and increasing market profits. This creates a focus on product life-cycle components as companies then make life-cycle commitments, where they are responsible for the function availability during the extent of the life-cycle, i.e. functional products. One of these life-cycle components is manufacturing; therefore, companies search for new approaches of success during manufacturability evaluation already in engineering design. Efforts have been done to support early engineering design, as this phase sets constraints and opportunities for manufacturing. These efforts have turned into design for manufacturing methods and guidelines. A further step to improve the life-cycle focus during early engineering design is to reuse results and use experience from earlier projects. However, because results and experiences created during project work are often not documented for reuse, only remembered by some people, there is a need for design support. Knowledge engineering (KE) is a methodology for creating knowledge-based systems, e.g. systems that enable reuse of earlier results and make available both explicit and tacit corporate knowledge, enabling the automated generation and evaluation of new engineering design solutions during early product development. There are a variety of KE-approaches, such as knowledge-based engineering, case-based reasoning and programming, which have been used in research to develop design for manufacturing methods and applications. There are, however, opportunities for research where several approaches and their interdependencies, to create a transparent picture of how KE can be used to support engineering design, are investigated. The aim of the research presented in this thesis is to create new methods for design for manufacturing, by using several approaches of KE, and find the beneficial and less beneficial aspects of these methods in comparison to each other and earlier research. This thesis presents methods and applications for design for manufacturing using KE. KE has been employed in several ways, namely rule-based, rule-, programmingand finite element analysis (FEA)-based, and ruleand plan-based, which are tested and compared with each other. Results show that KE can be used to generate information about manufacturing in several ways. The rule-based way is suitable for supporting life-cycle commitments, as engineering design and manufacturing can be integrated with maintenance and performance predictions during early engineering design, though limited to the firing of production rules. The rule-, programmingand FEA-based way can be used to integrate computer-aided design tools and virtual manufacturing for non-linear stress and displacement analysis. This way may also bridge the gap between engineering designers and computational experts, even though this way requires a larger effort to program than the rule-based. The ruleand planbased way can enable design for manufacturing in two fashions – based on earlier manufacturing plans and based on rules. Because earlier manufacturing plans, together with programming algorithms, can handle knowledge that may be more intricate to capture as rules, as opposed to the time demanding routine work that is often automated by means of rules, several opportunities for designing for manufacturing exist.

Methods and Applications

Manipulation of Stem Cells Fates: The Master and Multifaceted Roles of Biophysical Cues of Biomaterials

The successful ex vivo expansion of T-cells in great numbers is the cornerstone of adoptive cell therapy. We aimed to achieve the most optimal T-cell expansion condition by comparing the expansion of T-cells at various seeding densities, IL-2 concentrations, and bead-to-cell ratios. we first expanded the peripheral blood mononuclear cells (PBMCs) of a healthy donor at a range of 20 to 500 IU/mL IL-2 concentrations, 125 × 103 to 1.5 × 106 cell/mL, and 1:10 to 10:1 B:C (Bead-to-cell) ratios and compared the results. We then expanded the PBMC of three healthy donors using the optimized conditions and examined the growth kinetics. On day 28, CD3, CD4, and CD8 expression of the cell populations were analyzed by flow cytometry. T-cells of the first donor showed greater expansion results in IL-2 concentrations higher than 50 IU/mL compared to 20 IU/mL (P = 0.02). A seeding density of 250 × 103 cell/mL was superior to higher or lower densities in expanding T-cells (P = 0.025). Also, we witnessed a direct correlation between the B:C ratio and T-cell expansion, in which, in 5:1 and 10:1 B:C ratios T-cell significantly expanded more than lower B:C ratios. The results of PBMC expansions of three healthy donors were similar in growth kinetics. In the optimized condition, 96–98% of the lymphocyte population expressed CD3. While the majority of these cells expressed CD8, the mean expression of CD4 in the donors was 19.3, 16.5, and 20.4%. Our methodology demonstrates an optimized culture condition for the production of large quantities of polyclonal T-cells, which could be useful for future clinical and research studies.

/pdf/optimizing-interleukin-2-concentration-seeding-density-and-1o8yovxqf8.pdf

Optimizing interleukin-2 concentration, seeding density and bead-to-cell ratio of T-cell expansion for adoptive immunotherapy

Summary This protocol entails a simple method for isolation, culturing, and in vitro differentiation of adult neural stem cells from the dentate gyrus in the hippocampus and the subventricular zone of adult mice. Cultured adult neural stem cells are an important in vitro model to investigate stem cell properties such as proliferation and differentiation and to expand the understanding of plasticity in the adult brain. For complete details on the use and execution of this protocol, please refer to Isaksen et al. (2020) .

/pdf/protocol-for-mouse-adult-neural-stem-cell-isolation-and-410rb5q90r.pdf

Protocol for mouse adult neural stem cell isolation and culture.

Neural stem cells (NSCs) can proliferate and differentiate into multiple cell types that constitute the nervous system. NSCs can be derived from developing fetuses, embryonic stem cells, or induced pluripotent stem cells. NSCs provide a good platform to screen drugs for neurodegenerative diseases and also have potential applications in regenerative medicine. Natural products have long been used as compounds to develop new drugs. In this review, natural products that control NSC fate and induce their differentiation into neurons or glia are discussed. These phytochemicals enable promising advances to be made in the treatment of neurodegenerative diseases.

/pdf/regulation-of-neural-stem-cell-fate-by-natural-products-3eq2pv9gsb.pdf

Regulation of Neural Stem Cell Fate by Natural Products.

Recently, the importance of surface nanotopography in the determination of stem cell fate and behavior has been revealed. In the current study, we generated polystyrene cell-culture dishes with an omnidirectional nanopore arrayed surface (ONAS) (diameter: 200 nm, depth: 500 nm, center-to-center distance: 500 nm) and investigated the effects of nanotopography on rat neural stem cells (NSCs). NSCs cultured on ONAS proliferated better than those on the flat surface when cell density was low and showed less spontaneous differentiation during proliferation in the presence of mitogens. Interestingly, NSCs cultured on ONAS at clonal density demonstrated a propensity to generate neurospheres, whereas those on the flat surface migrated out, proliferated as individuals, and spread out to attach to the surface. However, the differential patterns of proliferation were cell density-dependent since the distinct phenomena were lost when cell density was increased. ONAS modulated cytoskeletal reorganization and inhibited formation of focal adhesion, which is generally observed in NSCs grown on flat surfaces. ONAS appeared to reinforce NSC-NSC interaction, restricted individual cell migration and prohibited NSC attachment to the nanopore surface. These data demonstrate that ONAS maintains NSCs as undifferentiated while retaining multipotency and is a better topography for culturing low density NSCs.

/pdf/cell-density-dependent-differential-proliferation-of-neural-1as8mkd0c6.pdf

Cell density-dependent differential proliferation of neural stem cells on omnidirectional nanopore-arrayed surface.

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Cell density-dependent differential proliferation of neural stem cells on omnidirectional nanopore-arrayed surface.