Evaluation of a Novel Thiol–Norbornene-Functionalized Gelatin Hydrogel for Bioprinting of Mesenchymal Stem Cells

Introduction: Three-dimensional bioprinting can be considered as an advancement of the classical tissue engineering concept. For bioprinting, cells have to be dispersed in hydrogels. Recently, a novel semi-synthetic thiolene hydrogel system based on norbornene-functionalized gelatin (GelNB) and thiolated gelatin (GelS) was described that resulted in the photoclick hydrogel GelNB/GelS. In this study, we evaluated the printability and biocompatibility of this hydrogel system towards adipose-tissue-derived mesenchymal stem cells (ASCs). Methods: GelNB/GelS was synthesized with three different crosslinking densities (low, medium and high), resulting in different mechanical properties with moduli of elasticity between 206 Pa and 1383 Pa. These hydrogels were tested for their biocompatibility towards ASCs in terms of their viability, proliferation and differentiation. The extrusion-based bioprinting of ASCs in GelNB/GelS-high was performed to manufacture three-dimensional cubic constructs. Results: All three hydrogels supported the viability, proliferation and chondrogenic differentiation of ASCs to a similar extent. The adipogenic differentiation of ASCs was better supported by the softer hydrogel (GelNB/GelS-low), whereas the osteogenic differentiation was more pronounced in the harder hydrogel (GelNB/GelS-high), indicating that the differentiation fate of ASCs can be influenced via the adaption of the mechanical properties of the GelNB/GelS system. After the ex vivo chondrogenic differentiation and subcutaneous implantation of the bioprinted construct into immunocompromised mice, the production of negatively charged sulfated proteoglycans could be observed with only minimal inflammatory signs in the implanted material. Conclusions: Our results indicate that the GelNB/GelS hydrogels are very well suited for the bioprinting of ASCs and may represent attractive hydrogels for subsequent in vivo tissue engineering applications.     

Optimized in-phase and opposed-phase MR imaging for accurate detection of small fat or water fractions: theoretical considerations and experimental application in emulsions

Object  

To optimize strategies and measurement parameters for quantification of small fat and water fractions (<10%) in mixtures of both components by 4-point in-phase and opposed-phase gradient-echo imaging and to compare theoretical results with in-vitro experiments using emulsions.     

Potential influence of Gadolinium contrast on image segmentation in MR-based attenuation correction with Dixon sequences in whole-body 18F-FDG PET/MR

Objective

To evaluate the influence of Gadolinium contrast agent on image segmentation in magnetic resonance (MR)-based attenuation correction (AC) with four-segment dual-echo time Dixon-sequences in whole-body [18F]-fluorodeoxyglucose positron emission tomography (PET)/MR imaging, and to analyze the consecutive effect on standardized uptake value (SUV).

Materials and methods

Hybrid imaging with an integrated PET/MR system was performed in 30 oncological patients. AC was based on MR imaging with a Dixon sequence with subsequent automated image segmentation. AC maps (µmaps) were acquired and reconstructed prior to (µmap_?gd) and after (µmap_+gd) Gd-contrast agent application. For quantification purposes, the SUV of organs and tumors based on both µmaps were compared.

Results

Tissue classification based on µmap_?gd was correct in 29/30 patients; based on µmap_+gd, the brain was falsely classified as fat in 12/30 patients with significant underestimation of SUV. In all cancerous lesions, tissue segmentation was correct. All concordant µmaps_?gd/+gd resulted in no significant difference in SUV.

Conclusion

In PET/MR, Gd-contrast agent potentially influences fat/water separation in Dixon-sequences of the head with above-average false tissue segmentation and an associated underestimation of SUV. Thus, MR-based AC should be acquired prior to Gd-contrast agent application. Additionally, integrating the MR-based AC maps into the reading-routine in PET/MR is recommended to avoid interpretation errors in cases where tissue segmentation fails.

     

Trans-Splicing Improvement by the Combined Application of Antisense Strategies

Spliceosome-mediated RNA trans-splicing has become an emergent tool for the repair of mutated pre-mRNAs in the treatment of genetic diseases. RNA trans-splicing molecules (RTMs) are designed to induce a specific trans-splicing reaction via a binding domain for a respective target pre-mRNA region. A previously established reporter-based screening system allows us to analyze the impact of various factors on the RTM trans-splicing efficiency in vitro. Using this system, we are further able to investigate the potential of antisense RNAs (AS RNAs), presuming to improve the trans-splicing efficiency of a selected RTM, specific for intron 102 of COL7A1. Mutations in the COL7A1 gene underlie the dystrophic subtype of the skin blistering disease epidermolysis bullosa (DEB). We have shown that co-transfections of the RTM and a selected AS RNA, interfering with competitive splicing elements on a COL7A1-minigene (COL7A1-MG), lead to a significant increase of the RNA trans-splicing efficiency. Thereby, accurate trans-splicing between the RTM and the COL7A1-MG is represented by the restoration of full-length green fluorescent protein GFP on mRNA and protein level. This mechanism can be crucial for the improvement of an RTM-mediated correction, especially in cases where a high trans-splicing efficiency is required.     

Pseudo-Starvation Driven Energy Expenditure Negatively Affects Ovarian Follicle Development

In the present investigation, we examined whether a change in whole body energy fluxes could affect ovarian follicular development, employing mice ectopically expressing uncoupling protein 1 in skeletal muscle (UCP1-TG). Female UCP1-TG and wild-type (WT) mice were dissected at the age of 12 weeks. Energy intake and expenditure, activity, body weight and length, and body composition were measured. Plasma insulin, glucose, leptin, plasma fibroblast growth factor 21 (FGF21) and plasma insulin-like growth factor 1 (IGF1) levels were analyzed and ovarian follicle and corpus luteum numbers were counted. IGF1 signaling was analyzed by immunohistochemical staining for the activation of insulin receptor substrate 1/2 (IRS1/2) and AKT. UCP1-TG female mice had increased energy expenditure, reduced body size, maintained adiposity, and decreased IGF1 concentrations compared to their WT littermates, while preantral and antral follicle numbers were reduced by 40% and 60%, respectively. Corpora lutea were absent in 40% of the ovaries of UCP1-TG mice. Phospho-IRS1, phospho-AKT -Ser473 and -Thr308 immunostaining was present in the granulosa cells of antral follicles in WT ovaries, but faint to absent in the antral follicles of UCP1-TG mice. In conclusion, the reduction in circulating IGF1 levels due to the ectopic expression of UCP1 is associated with reduced immunostaining of the IRS1-PI3/AKT pathway, which may negatively affect ovarian follicle development and ovulation.     

How to manage creativity time? Results from a social psychological time model lab experiment on individual creative and routine performance

Companies have long used various approaches for organizing their employees' time for creative and routine tasks in order to improve innovative performance. In this paper, we examine how work schedule autonomy affects individuals' creative and routine performance. We then evaluate non‐commissioned time models. Results of laboratory experiments with 233 participants reveal that while average routine performance is not affected by schedule autonomy, the effect of schedule autonomy on creative performance depends on the subject's impulsiveness. There is evidence of an inverse relationship between schedule autonomy and creative performance among subjects of low impulsiveness. Hence, our results indicate that the optimal management policy depends on the manager's focus on creative or routine performance and the types of employees the manager supervises. For routine performance, the creativity time model has no significant impact.