Protein quality control is essential for cellular survival. Failure to eliminate pathogenic proteins leads to their intracellular accumulation in the form of protein aggregates. Autophagy can recognize protein aggregates and degrade them in lysosomes. However, some aggregates escape the autophagic surveillance. Here we analyse the autophagic degradation of different types of aggregates of synphilin-1, a protein often found in pathogenic protein inclusions. We show that small synphilin-1 aggregates and large aggresomes are differentially targeted by constitutive and inducible autophagy. Furthermore, we identify a region in synphilin-1, necessary for its own basal and inducible aggrephagy and sufficient for the degradation of other pro-aggregating proteins. Although the presence of this peptide is sufficient for basal aggrephagy, inducible aggrephagy requires its ubiquitination, which diminishes protein mobility on the surface of the aggregate and favours the recruitment and assembly of the protein complexes required for autophagosome formation. Our study reveals different mechanisms for cells to cope with aggregate proteins via autophagy and supports the idea that autophagic susceptibility of prone-to-aggregate proteins may not depend on the nature of the aggregating proteins per se, but on their dynamic properties in the aggregate.

3-dimensional (3D) in vitro models were developed in order to mimic the complexity of real organ/tissue in a dish. They offer new possibilities to model biological processes in more physiologically relevant ways which can be applied to a myriad of applications including drug development, toxicity screening and regenerative medicine. Hydrogels are the most relevant tissue-like matrices to support the development of 3D in vitro models since they are in many ways akin to the native extracellular matrix (ECM). For the purpose of further improving matrix relevance or to impart specific functionalities, composite hydrogels have attracted increasing attention. These could incorporate drugs to control cell fates, additional ECM elements to improve mechanical properties, biomolecules to improve biological activities or any combinations of the above. In this Review, recent developments in using composite hydrogels laden with cells as biomimetic tissue-or organ-like constructs, and as matrices for multi-cell type organoid cultures are highlighted. The latest composite hydrogel systems that contain nanomaterials, biological factors, and combinations of biopolymers (e.g., proteins and polysaccharide), such as Interpenetrating Networks (IPNs) and Soft Network Composites (SNCs) are also presented. While promising, challenges remain. These will be discussed in light of future perspectives toward encompassing diverse composite hydrogel platforms for an improved organ environment in vitro.

No abstract

Background Hydroxyapatite (HA) [Ca 5 (PO4) 3 (OH)] is a naturally occurring calcium phosphate which makes up 60–70% of the dry weight of human bones. Nano-scale HA particles are increasingly being used as carriers for controlled and targeted delivery of bioactive agents like drugs, proteins, and nucleic acids due to their high porosity, negative charge, and biodegradability. Purpose Although much effort has been devoted to understanding the delivery kinetics and effects of the payloads in such carriers, a thorough understanding of the influence of the carriers themselves is lacking. Methods HA particles (300 µg/mL) were administered to primary human dermal fibroblasts (HDFs). The uptake and intracellular localization of the particles were determined by flow cytometry, confocal imaging, and transmission electron microscopy (TEM). Immunological assays and PCR were performed to determine the levels of pro-inflammatory cytokines and collagens in cell lysates and media supernatant. Results The current study explores the effects of poly-dispersed HA particles on primary HDFs as a model system. The majority of the particles were determined to range between 150 and 200 nm in diameter. Upon exposure to HA suspensions, primary HDFs internalized the particles by endocytosis within 6 hours of exposure, showing maximum uptake at 72 hours following which the particles were exocytosed by 168 hours. This correlated to reduced secretion of various pro-inflammatory and pro-collagenic cytokines. Biochemical analysis further revealed a reduction in Type I collagen expression and secretion. Conclusion HA particles have an immune-modulatory effect on dermal fibroblasts and reduce collagen production, which may impact the integrity of the extracellular matrix (ECM). This study demonstrates the need to consider the secondary effects of particulate carriers like HA, beyond basic cytotoxicity, in the specific tissue environment where the intended function is to be realized.

Bone exhibits piezoelectric properties. Thus, electrical stimulations such as pulsed electromagnetic fields (PEMFs) and stimuli-responsive piezoelectric properties of scaffolds have been investigated separately to evaluate their efficacy in supporting osteogenesis. However, current understanding of cells responding under the combined influence of PEMF and piezoelectric properties in scaffolds is still lacking. Therefore, in this study, we fabricated piezoelectric scaffolds by functionalization of polycaprolactone-tricalcium phosphate (PCL-TCP) films with a polyvinylidene fluoride (PVDF) coating that is self-polarized by a modified breath-figure technique. The osteoinductive properties of these PVDF-coated PCL-TCP films on MC3T3-E1 cells were studied under the stimulation of PEMF. Piezoelectric and ferroelectric characterization demonstrated that scaffolds with piezoelectric coefficient d33 = −1.2 pC/N were obtained at a powder dissolution temperature of 100 °C and coating relative humidity (RH) of 56%. DNA quantification showed that cell proliferation was significantly enhanced by PEMF as low as 0.6 mT and 50 Hz. Hydroxyapatite staining showed that cell mineralization was significantly enhanced by incorporation of PVDF coating. Gene expression study showed that the combination of PEMF and PVDF coating promoted late osteogenic gene expression marker most significantly. Collectively, our results suggest that the synergistic effects of PEMF and piezoelectric scaffolds on osteogenesis provide a promising alternative strategy for electrically augmented osteoinduction. The piezoelectric response of PVDF by PEMF, which could provide mechanical strain, is particularly interesting as it could deliver local mechanical stimulation to osteogenic cells using PEMF.