Self-Assembled Fibrinogen Scaffolds Support Cocultivation of Human Dermal Fibroblasts and HaCaT Keratinocytes.

Self-assembled fibrinogen scaffolds are highly attractive biomaterials to mimic native blood clots. To explore their potential for wound healing, we studied the interaction of cocultures of human dermal fibroblasts (HDFs) and HaCaT keratinocytes with nanofibrous, planar, and physisorbed fibrinogen. Cell viability analysis indicated that the growth of HDFs and HaCaTs was supported by all fibrinogen topographies until 14 days, either in mono- or coculture. Using scanning electron microscopy and cytoskeletal staining, we observed that the native morphology of both cell types was preserved on all topographies. Expression of the marker proteins vimentin and cytokeratin-14 showed that the native phenotype of fibroblasts and undifferentiated keratinocytes, respectively, was maintained. HDFs displayed their characteristic wound healing phenotype, characterized by expression of fibronectin. Finally, to mimic the multilayered microenvironment of skin, we established successive cocultures of both cells, for which we found consistently high metabolic activities. SEM analysis revealed that HaCaTs arranged into a confluent top layer after 14 days, while fluorescent labeling confirmed the presence of both cells in the layered structure after 6 days. In conclusion, all fibrinogen topographies successfully supported the cocultivation of fibroblasts and keratinocytes, with fibrinogen nanofibers being particularly attractive for skin regeneration due to their biomimetic porous architecture and the technical possibility to be detached from an underlying substrate.

Inhibition of PHLPP1/2 phosphatases rescues pancreatic ß-cells in diabetes.

Pancreatic ß-cell failure is the key pathogenic element of the complex metabolic deterioration in type 2 diabetes (T2D); its underlying pathomechanism is still elusive. Here, we identify pleckstrin homology domain leucine-rich repeat protein phosphatases 1 and 2 (PHLPP1/2) as phosphatases whose upregulation leads to ß-cell failure in diabetes. PHLPP levels are highly elevated in metabolically stressed human and rodent diabetic ß-cells. Sustained hyper-activation of mechanistic target of rapamycin complex 1 (mTORC1) is the primary mechanism of the PHLPP upregulation linking chronic metabolic stress to ultimate ß-cell death. PHLPPs directly dephosphorylate and regulate activities of ß-cell survival-dependent kinases AKT and MST1, constituting a regulatory triangle loop to control ß-cell apoptosis. Genetic inhibition of PHLPPs markedly improves ß-cell survival and function in experimental models of diabetes in vitro, in vivo, and in primary human T2D islets. Our study presents PHLPPs as targets for functional regenerative therapy of pancreatic ß cells in diabetes.

Self-assembled fibrinogen nanofibers support fibroblast adhesion and prevent E. coli infiltration.

Fibrinogen nanofibers hold great potential for wound healing applications since they mimic the native blood clot architecture and offer important binding sites to support fibroblast adhesion and migration. Recently, we introduced a new method of salt-induced self-assembly to prepare nanofibrous fibrinogen scaffolds. Here, we present our results on the mechanical properties of these scaffolds and their interaction with 3T3 fibroblasts and E. coli bacteria, which we used as model systems for wound healing. Hydrated, nanofibrous fibrinogen scaffolds showed a Young's modulus of 1.3 MPa, which is close to the range of native fibrin. 3T3 fibroblasts adhered and proliferated well on nanofibrous and planar fibrinogen up to 72 h with a less pronounced actin cytoskeleton on nanofibers in comparison to planar fibrinogen. Fibroblasts on nanofibers were smaller with many short filopodia while larger cells with few long filopodia were found on planar fibrinogen. Live cell tracking revealed higher migration velocities on nanofibers in comparison to planar fibrinogen. The growth of E. coli bacteria on nanofibrous fibrinogen was significantly reduced as compared to agar controls with no bacteria migrating through the nanofibers. In summary, we conclude that self-assembled fibrinogen nanofibers could become highly attractive as future scaffolds for wound healing applications.

Principles of Fibrinogen Fiber Assembly In Vitro.

Fibrinogen nanofibers hold great potential for applications in wound healing and personalized regenerative medicine due to their ability to mimic the native blood clot architecture. Although versatile strategies exist to induce fibrillogenesis of fibrinogen in vitro, little is known about the underlying mechanisms and the associated length scales. Therefore, in this manuscript the current state of research on fibrinogen fibrillogenesis in vitro is reviewed. For the first time, the manifold factors leading to the assembly of fibrinogen molecules into fibers are categorized considering three main groups: substrate interactions, denaturing and non-denaturing buffer conditions. Based on the meta-analysis in the review it is concluded that the assembly of fibrinogen is driven by several mechanisms across different length scales. In these processes, certain buffer conditions, in particular the presence of salts, play a predominant role during fibrinogen self-assembly compared to the surface chemistry of the substrate material. Yet, to tailor fibrous fibrinogen scaffolds with defined structure-function-relationships for future tissue engineering applications, it still needs to be understood which particular role each of these factors plays during fiber assembly. Therefore, the future combination of experimental and simulation studies is proposed to understand the intermolecular interactions of fibrinogen, which induce the assembly of soluble fibrinogen into solid fibers.

Iron-Doping of Copper Oxide Nanoparticles Lowers Their Toxic Potential on C6 Glioma Cells.

Copper oxide nanoparticles (CuO-NPs) are well known for their cytotoxicity which in part has been attributed to the release of copper ions from CuO-NPs. As iron-doping has been reported to reduce the susceptibility of CuO-NPs to dissolution, we have compared pure CuO-NPs and CuO-NPs that had been doped with 10% iron (CuO-Fe-NPs) for copper release and for their toxic potential on C6 glioma cells. Physicochemical characterization revealed that dimercaptosuccinate (DMSA)-coated CuO-NPs and CuO-Fe-NPs did not differ in their size or zeta potential. However, the redox activity and liberation of copper ions from CuO-Fe-NPs was substantially slower compared to that from CuO-NPs, as demonstrated by cyclic voltammetry and by the photometric quantification of the copper ion-bathocuproine complex, respectively. Exposure of C6 cells to these NPs caused an almost identical cellular copper accumulation and each of the two types of NPs induced ROS production and cell toxicity. However, the time- and concentration-dependent loss in cell viability was more severe for cells that had been treated with CuO-NPs compared to cells exposed to CuO-Fe-NPs. Copper accumulation and toxicity after exposure to either CuO-NPs or CuO-Fe-NPs was prevented in the presence of copper chelators, while neutralization of the lysosomal pH by bafilomycin A1 prevented toxicity without affecting cellular copper accumulation or ROS production. These data demonstrate that iron-doping does not affect cellular accumulation of CuO-NPs and suggests that the intracellular liberation of copper ions from CuO-NPs is slowed by the iron doping, which in turn lowers the cell toxic potential of iron-doped CuO-NPs.

Uptake of Intact Copper Oxide Nanoparticles Causes Acute Toxicity in Cultured Glial Cells.

Copper oxide nanoparticles (CuO-NPs) dispersions are known for their high cell toxic potential but contaminating copper ions in such dispersions are a major hurdle in the investigation of specific nanoparticle-mediated toxicity. In order to distinguish between the adverse effects exhibited by CuO-NPs and/or by contaminating ionic copper, the membrane-impermeable copper chelator bathocuproine disulfonate (BCS) was added in a low molar ratio (20% of the total copper applied) in order to chelate the copper ions that had been released extracellularly from the CuO-NPs before or during the incubation. Physicochemical characterization of synthesized CuO-NPs revealed that the presence of this low concentration of BCS did not alter the size or zeta potential of the CuO-NPs. Application of CuO-NPs to C6 glioma cells and primary astrocytes induced a concentration- and temperature-dependent copper accumulation which was accompanied by a severe loss in cell viability. The adverse consequences of the CuO-NP application were not affected by the presence of 20% BCS, while the copper accumulation and cell toxicity observed after application of ionic copper were significantly lowered in the presence of BCS. These results demonstrate that for the experimental conditions applied the adverse consequences of an exposure of cultured glial cells to dispersions of CuO-NPs are mediated by accumulated NPs and not caused by the uptake of contaminating copper ions.

Nutritional Mobile Applications for CKD Patients: Systematic Review.

INTRODUCTION: Mobile health applications offer the potential to help people living with chronic kidney disease (CKD) manage diet-related challenges. This systematic review examined CKD dietary mobile app interventions; specifically, app characteristics, feasibility, and effectiveness in changing user behavior, as well as user satisfaction. METHODS: This review was reported in accordance with PRISMA guidelines. We searched scholarly databases, as well as the gray literature, for all randomized controlled trials, observational studies, needs assessments, and pilot testing/studies/trials focused on the development or evaluation of CKD dietary mobile app interventions. The characteristics, user satisfaction with, usability/feasibility, and effectiveness in changing dietary behavior of the mobile application were summarized using descriptive statistics and in a narrative manner. RESULTS: Thirteen full-text studies were included, of which 11 were single center, with a mean sample size of 23. Of the 7 studies that measured usability/feasibility, all found at least some aspects of the application feasible/useful. Of the 5 studies that reported an evaluation of changes in behavior/diet related to self-management, all reported some positive change. CONCLUSION: According to current studies, nutritional apps show promise in CKD self-management.

Uptake and Toxicity of Copper Oxide Nanoparticles in C6 Glioma Cells.

Copper oxide nanoparticles (CuO-NPs) are frequently used for many technical applications, but are also known for their cell toxic potential. In order to investigate a potential use of CuO-NPs as a therapeutic drug for glioma treatment, we have investigated the consequences of an application of CuO-NPs on the cellular copper content and cell viability of C6 glioma cells. CuO-NPs were synthesized by a wet-chemical method and were coated with dimercaptosuccinic acid and bovine serum albumin to improve colloidal stability in physiological media. Application of these protein-coated nanoparticles (pCuO-NPs) to C6 cells caused a strong time-, concentration- and temperature-dependent copper accumulation and severe cell death. The observed loss in cellular MTT-reduction capacity, the loss in cellular LDH activity and the increase in the number of propidium iodide-positive cells correlated well with the specific cellular copper content. C6 glioma cells were less vulnerable to pCuO-NPs compared to primary astrocytes and toxicity of pCuO-NPs to C6 cells was only observed for incubation conditions that increased specific cellular copper contents above 20 nmol copper per mg protein. Both cellular copper accumulation as well as the pCuO-NP-induced toxicity in C6 cells were prevented by application of copper chelators, but not by endocytosis inhibitors, suggesting that liberation of copper ions from the pCuO-NPs is the first step leading to the observed toxicity of pCuO-NP-treated glioma cells.