Towards A Design Of A Software-Defined Manufacturing System Based On A Systematic Literature Review For Enabling A Decentralised High-Rate Electrolyser Production

Hydrogen is critical for the transition to an environmentally sound and reliable energy supply. This transition requires large capacities of performant and cost-effective electrolysers. Although performant electrolysers already exist, they cannot yet be manufactured at a high rate in series production. The project H2Giga-FRHY is researching a reference factory for large-scale production of electrolysers, developing new production and testing modules. As an essential building block of the reference factory, a research group at Fraunhofer IPA is designing and implementing a comprehensive software-defined manufacturing system (SDMS), which supports the decentralized high-rate production of electrolysers and allows for far-reaching insights regarding high-rate capability, quality, and cost of products, processes, and technologies involved. For the SDMS implementation, different enterprise architecture (EA) approaches are considered and evaluated in the scope of a structured literature review with respect to criteria arising from the project context and related research questions. In this paper, an approach to designing a software-defined manufacturing system is described, and its necessity is based on the use case-specific criteria discussed

Size-dependent internalization of particles via the pathways of clathrin- and caveolae-mediated endocytosis.

Non-phagocytic eukaryotic cells can internalize particles <1 microm in size, encompassing pathogens, liposomes for drug delivery or lipoplexes applied in gene delivery. In the present study, we have investigated the effect of particle size on the pathway of entry and subsequent intracellular fate in non-phagocytic B16 cells, using a range of fluorescent latex beads of defined sizes (50-1000 nm). Our data reveal that particles as large as 500 nm were internalized by cells via an energy-dependent process. With an increase in size (50-500 nm), cholesterol depletion increased the efficiency of inhibition of uptake. The processing of the smaller particles was significantly perturbed upon microtubule disruption, while displaying a negligible effect on that of the 500 nm beads. Inhibitor and co-localization studies revealed that the mechanism by which the beads were internalized, and their subsequent intracellular routing, was strongly dependent on particle size. Internalization of microspheres with a diameter <200 nm involved clathrin-coated pits. With increasing size, a shift to a mechanism that relied on caveolae-mediated internalization became apparent, which became the predominant pathway of entry for particles of 500 nm in size. At these conditions, delivery to the lysosomes was no longer apparent. The data indicate that the size itself of (ligand-devoid) particles can determine the pathway of entry. The clathrin-mediated pathway of endocytosis shows an upper size limit for internalization of approx. 200 nm, and kinetic parameters may determine the almost exclusive internalization of such particles along this pathway rather than via caveolae

Efficient cationic lipid-mediated delivery of antisense oligonucleotides into eukaryotic cells: down-regulation of the corticotropin-releasing factor receptor

Oligonucleotides (ODNs) can be employed as effective gene-specific regulators. However, before ODNs can reach their targets, several physical barriers have to be overcome, as although ODNs may pass cell membranes, most become sequestered in endocytic compartments. Accordingly, sophisticated strategies are required for efficient delivery. Here we have employed a pyridinium-based synthetic amphiphile, called SAINT-2, which carries ODNs into cells in a highly efficient, essentially non-toxic and serum-insensitive manner. Intracellular delivery was examined by monitoring the trafficking of fluorescent ODNs and lipid, and by measuring the effect of specific antisense ODNs on target mRNA and protein levels of the receptor for the neuropeptide corticotropin-releasing factor (CRF-R), expressed in Chinese hamster ovary cells. ODN delivery is independent of lipoplex size, and fluorescently tagged ODNs readily acquire access to the nucleus, whereas the carrier itself remains sequestered in the endosomal-lysosomal pathway. While the release is independent of the presence of serum, it is not observed when serum proteins gain access within the lipoplex, and which likely stabilizes the lipoplex membrane. We propose that the amphiphile-dependent aggregate structure governs complex dissociation, and hence, the biological efficiency of ODNs. We demonstrate an essentially non-toxic and effective antisense-specific down-regulation of the CRF-R, both at the mRNA and protein level.