3D Bioprinting of Neurovascular Tissue Modeling with Collagen-Based Low-Viscosity Composites.

In vitro neurovascular unit (NVU) models are valuable for investigating brain functions and developing drugs. However, it remains challenging to recapitulate the native architectural features and ultra-soft extracellular matrix (ECM) properties of the natural NVU. Cell-laden bioprinting is promising to prepare complex living tissues, but hard to balance the fidelity and cell growth. This study proposes a novel two-stage methodology for biomanufacturing functional 3D neurovascular constructs in vitro with low modulus of ECM. At the shaping stage, a low-viscosity alginate/collagen is printed through an embedded approach; at the culturing stage, the alginate is removed through targeted lysing. The low-viscosity and rapid crosslinking properties provide a printing resolution of ≈10 µm, and the lysis processing can decrease the hydrogels' modulus to ≈1 kPa and adjust the porosity of the microstructure, providing cells with an environment closing to the brain ECM. A 3D hollow coaxial neurovascular model is fabricated, in which the endothelial cells has expressed tight junction proteins and shown selective permeability, and the astrocytes outside of the endothelial layer are found to spread out with branches and directly interact with endothelial cells. The present study offers a promising modeling method for better understanding the NVU function and screening neuro-drugs.

High-Throughput and Untargeted Metabolic Profiling Revealed the Potential Effect and Mechanisms of Paeoniflorin in Young Asthmatic Rats.

Paeoniflorin (PF) is a multi-target monoterpenoid glycoside and possesses broad pharmacological functions, e.g., anti-inflammation, anti-depression, antitumor, abirritation, neuroprotection, antioxidant, and enhancing cognitive and learning ability. PF has gained a large amount of attention for its effect on asthma disease as the growth rate of asthma has increased in recent years. However, its mechanism of action on asthma is still unclear. In this study, we have explored the action mechanism of PF on asthma disease. Furthermore, high-throughput untargeted metabolic profiling was performed through ultraperformance liquid chromatography/electrospray ionization quadruple time-of-flight high-definition mass spectrometry (QA) UPLC-Q/TOF-MS combined with pattern recognition approaches and pathway analysis. A total of 20 potential biomarkers were discovered by UPLC/MS and urine metabolic profiling. The key pathways including the citrate cycle (the TCA cycle), pyrimidine metabolism, pentose phosphate pathway, tyrosine metabolism, and tryptophan metabolism were affected by PF. In conclusion, we have discovered metabolite biomarkers and revealed the therapeutic mechanism of PF based on liquid chromatography coupled with mass spectrometry untargeted metabolomics. The untargeted metabolomics combined with UPLC-MS is a useful tool for exploring the therapeutic mechanism and targets of PF in the treatment of asthma. Metabolomics combined with UPLC-MS is an integrated method to explore the metabolic mechanism of PF in the treatment of asthma rats and to reveal the potential targets, providing theoretical support for the study of the treatment of PF.

Circular RNA mmu_circ_0001295 from hypoxia pretreated adipose-derived mesenchymal stem cells (ADSCs) exosomes improves outcomes and inhibits sepsis-induced renal injury in a mouse model of sepsis.

Microvascular dysfunction causes mortality in the presence of sepsis and multi-organ failure. Previous studies have demonstrated that exogenous administration of exosomes from adipose-derived mesenchymal stem cells (ADSCs) protects against sepsis, improves organ function, decreases vascular leakage and increases survival. However, the underlying regulatory mechanism was largely unknown. Therefore, in this study, a mouse sepsis model based on cecal ligation and puncture (CLP) was constructed. Exosomes from various ADSCs were intravenously administered at 4 h post CLP. Treatment with ADSC exosomes (Exo), particularly those with hypoxic pretreatment (HExo), enhanced survival, suppressed renal vascular leakage and decreased kidney dysfunction in septic mice. HExo ameliorated sepsis-induced increases in chemokine and cytokine plasma levels. Furthermore, the HExo circRNA content, determined through next-generation sequencing, revealed abundant mmu_circ_0001295. Further studies demonstrated that downregulation of exosomal mmu_circ_0001295 suppressed the exosomes' protective effects against sepsis. HExo prevented microvascular dysfunction, thus potentially improving sepsis outcomes via mmu_circ_0001295 delivery. In summary, the data indicated that HExo elongate sepsis-induced renal injury through delivering mmu_circ_0001295.

Biofabrication of a Low Modulus Bioelectroprobe for Neurons to Grow Into.

Implantable nerve electrodes, as a bridge between the brain and external devices, have been widely used in areas such as brain function exploration, neurological disease treatment and human-computer interaction. However, the mechanical properties mismatch between the electrode material and the brain tissue seriously affects the stability of electrode signal acquisition and the effectiveness of long-term service in vivo. In this study, a modified neuroelectrode was developed with conductive biomaterials. The electrode has good biocompatibility and a gradient microstructure suitable for cell growth. Compared with metal electrodes, bioelectrodes not only greatly reduced the elastic modulus (<10 kpa) but also increased the conductivity of the electrode by 200 times. Through acute electrophysiological analysis and a 12-week chronic in vivo experiment, the bioelectrode clearly recorded the rat's brain electrical signals, effectively avoided the generation of glial scars and induced neurons to move closer to the electrode. The new conductive biomaterial electrodes developed in this research make long-term implantation of cortical nerve electrodes possible.

3D Printing of Layered Gradient Pore Structure of Brain-like Tissue.

The pathological research and drug development of brain diseases require appropriate brain models. Given the complex, layered structure of the cerebral cortex, as well as the constraints on the medical ethics and the inaccuracy of animal models, it is necessary to construct a brain-like model in vitro. In this study, we designed and built integrated three-dimensional (3D) printing equipment for cell printing/culture, which can guarantee cell viability in the printing process and provide the equipment foundation for manufacturing the layered structures with gradient distribution of pore size. Based on this printing equipment, to achieve the purpose of printing the layered structures with multiple materials, we conducted research on the performance of bio-inks with different compositions and optimized the printing process. By extruding and stacking materials, we can print the layered structure with the uniform distribution of cells and the gradient distribution of pore sizes. Finally, we can accurately print a structure with 30 layers. The line width (resolution) of the printed monolayer structure was about 478 mm, the forming accuracy can reach 97.24%, and the viability of cells in the printed structure is as high as 94.5%.

Correction to: Effects of astrocyte on neuronal outgrowth in a layered 3D structure.

It was highlighted that the original article [1] contained an error in the Acknowledgments section.

Effects of astrocyte on neuronal outgrowth in a layered 3D structure.

BACKGROUND: Human brain models and pharmacological models of brain diseases are in high demand for drug screening because animal models have been found to be less than ideal for fully representing the human brain and are likely to fail during drug screening and testing; therefore, the construction of brain-like tissues is necessary. Due to the complexity of cortical tissue, the in vitro construction of brain-like tissue models has been restricted to mostly two-dimensional (2D) models and, on a limited scale, three-dimensional (3D) models. METHODS: In this study, 3D tissue blocks encapsulating neurons and astrocytes were constructed and cultured in vitro to mimic the cortex of the brain and to investigate the effects of astrocytes on the growth of neurons in a 3D culture. RESULTS: The results indicated that such methodology can provide a 3D culture environment suitable for neurons and astrocytes to live and function. When both cells were evenly mixed and cultured in a 3D manner, the astrocytes, which showed better outgrowth and a higher proliferation rate, benefited more than the neurons. On the other hand, the neurons benefited, showing longer axons and a denser network of dendrites, when they were accompanied by astrocytes at a certain distance. CONCLUSION: In conclusion, astrocytes stimulated the outgrowth of neurons in a 3D culture environment in vitro. Regardless, the spatial relationship between both types of cells should be controlled. Thus, culturing cells in a 3D manner is necessary to investigate the correlations between them. This study provides a foundation for biofabricating 3D neurons' cultures to allow for a deeper insight into the relationship between astrocytes or other glial cells and neurons in a 3D culture that is similar to the natural environment of the brain.

In vitro model of the glial scar.

The trauma of central nervous system (CNS) can lead to glial scar, and it can limit the regeneration of neurons at the injured area, which is considered to be a major factor affecting the functional recovery of patients with CNS injury. At present, the study of the glial scar model in vitro is still limited to two-dimensional culture, and the state of the scar in vivo cannot be well mimicked. Therefore, we use a collagen gel and astrocytes to construct a three-dimensional (3D) model in vitro to mimic natural glial scar tissue. The effects of concentration changes of astrocytes on cell morphology, proliferation, and tissue performance were investigated. After 8 days of culture in vitro, the results showed that the tissue model contracted, with a measured shrinkage rate of 4.5%, and the compressive elastic modulus increased to nearly 4 times. Moreover, the astrocytes of the 3D tissue model have the ability of proliferation, hyperplasia, and formation of scar clusters. It indicates that the model we constructed has the characteristics of glial scar tissue to some extent and can provide an in vitro model for the research of glial scar and brain diseases.