Development of Strong and Tough ß-TCP/PCL Composite Scaffolds with Interconnected Porosity by Digital Light Processing and Partial Infiltration.

Strong and tough ß-TCP/PCL composite scaffolds with interconnected porosity were developed by combining digital light processing and vacuum infiltration. The composite scaffolds were comprised of pure ß-TCP, ß-TCP matrix composite and PCL matrix composite. The porous ß-TCP/PCL composite scaffolds showed remarkable mechanical advantages compared with ceramic scaffolds with the same macroscopic pore structure (dense scaffolds). The composite scaffolds exhibited a significant increase in strain energy density and fracture energy density, though with similar compressive and flexural strengths. Moreover, the composite scaffolds had a much higher Weibull modulus and longer fatigue life than the dense scaffolds. It was revealed that the composite scaffolds with interconnected porosity possess comprehensive mechanical properties (high strength, excellent toughness, significant reliability and fatigue resistance), which suggests that they could replace the pure ceramic scaffolds for degradable bone substitutes, especially in complex stress environments.

Single-Center Retrospective Analysis of Paraneoplastic Syndromes with Peripheral Nerve Damage.

There are few clinical and electrophysiological studies on paraneoplastic neurological syndrome (PNS) with peripheral nerve damage, which brings great challenges to clinical identification and diagnosis. We analyzed the clinical and electrophysiological data of twenty-five confirmed PNS cases using peripheral nerve damage patients. The results showed the most common chief complaint was weakness (20/25, 80%), followed by numbness (13/25, 52%). Nineteen patients (76%) exhibited peripheral nervous system lesions prior to occult tumors, and the median time from symptom onset to the diagnosis of a tumor was 4 months. The electrophysiological results revealed a higher rate of abnormal amplitudes than latency or conduction velocity, especially in sensory nerves. Meanwhile, we found that, compared with patients >65 y, patients aged ≤65 y exhibited more chronic onset (p = 0.01) and longer disease duration (p = 0.01), more motor nerve involvements (p = 0.02), more amplitude involvement (p = 0.01), and higher rates of the inability to walk independently at presentation (p = 0.02). The present study construed that weakness and paresthesia are common symptoms in PNS with peripheral nerve damage in some areas, and the electrophysiological results mainly changed in amplitude. Tumor screening in young and middle-aged patients with peripheral neuropathy cannot be ignored.

Causal relationship among obesity and body fat distribution and epilepsy subtypes.

Objective: The observational studies indicate an association between obesity and epilepsy, but it is unclear whether such an association responds to causality. The objective of this study was to determine the causal relationship between obesity and fat distribution and epilepsy subtypes based on waist circumference, hip circumference (HP), waist-hip ratio (WHR), and body mass index (BMI). Methods: A two-sample Mendelian randomization study was conducted separately for the four indicators of obesity and epilepsy and its seven subtypes, with reverse Mendelian randomization and multivariate Mendelian randomization for significant outcomes. Results: A two-sample Mendelian randomized analysis informed us that waist circumference was a risk factor for juvenile myoclonic epilepsy (beta = 0.0299, P = 4.60 × 10-3). The increase in hip circumference increased the risk of juvenile myoclonic epilepsy and epilepsy, with effect values of 0.0283 (P = 2.01 × 10-3) and 0.0928 (P = 1.40 × 10-2), respectively. Furthermore, children with a higher BMI exhibit a higher risk of epilepsy (beta = 0.0148 P = 1.05 × 10-3). The reverse Mendelian randomization study revealed that childhood absence epilepsy increased its BMI (beta = 0.8980, P = 7.52 × 10-7), and juvenile myoclonic epilepsy increased its waist circumference (beta = 0.7322, P = 3.26 × 10-2). Multivariate Mendelian randomization revealed that an increase in hip circumference and waist-hip ratio increased the risk of juvenile myoclonic epilepsy, with an effect value of 0.1051 (P = 9.75 × 10-4) and 0.1430 (P = 3.99 × 10-3), respectively, while an increase in BMI and waist circumference instead decreased their risk, with effect values of -0.0951 (P = 3.14 × 10-2) and-0.0541 (P = 1.71 × 10-2). In contrast, multivariate Mendelian randomization for childhood absence epilepsy and epilepsy did not identify any independent risk factors. Significance: Our findings provide novel evidence in favor of obesity as a risk factor for epilepsy and waist circumference as a risk factor for juvenile myoclonic epilepsy. Increased hip circumference confers an elevated risk of juvenile myoclonic epilepsy and epilepsy (all documented cases), and a high BMI increases the risk of childhood absence epilepsy. With this, new insights are provided into the energy metabolism of epilepsy, which supports further nutritional interventions and the search for new therapeutic targets.

Effects of hydrogen peroxide on endothelial function in three-dimensional hydrogel vascular model and regulation mechanism of polar protein Par3.

Three-dimensional (3D) cell cultures better reflect the function of endothelial cells (ECs) than two-dimensional (2D) cultures. In recent years, studies have found that ECs cultured in a 3D luminal structure can mimic the biological characteristics and phenotypes of vascular ECs, thus making it more suitable for endothelial dysfunction research. In this study, we used a 3D model and 2D tissue culture polystyrene (TCP) to investigate the effects of cell polarity on hydrogen peroxide (H2O2)-induced endothelial dysfunction and its related mechanisms. We observed the cell morphology, oxidative stress, and barrier and endothelial function of human umbilical vein ECs (HUVECs) in 3D and 2D cultures. We then used Illumina to detect the differentially expressed genes (DEGs) in the 3D-cultured HUVEC with and without H2O2stimulation, using clusterProfiler for Gene Ontology function enrichment analysis and Kyoto Encyclopaedia of Genes and Genomes pathway enrichment analysis of DEGs. Finally, we explored the role and mechanism of polar protein partitioning defective protein 3 (Par3) in the regulation of ECs. ECs were inoculated into the 3D hydrogel channel; after stimulation with H2O2, the morphology of HUVECs changed, the boundary was blurred, the expression of intercellular junction proteins decreased, and the barrier function of the EC layer was damaged. 3D culture increased the oxidative stress response of cells stimulated by H2O2compared to 2D TCPs. The polarity-related protein Par3 and cell division control protein 42 were screened using bioinformatics analysis, and western blotting was used to verify the results. Par3 knockdown significantly suppressed claudin1 (CLDN1) and vascular endothelial cadherin. These results suggest that the polar protein Par3 can protect H2O2-induced vascular ECs from damage by regulating CLDN1 and VE-cadherin.

A comparative study unraveling the effects of TNF-α stimulation on endothelial cells between 2D and 3D culture.

Endothelial cell (EC) dysfunction is an important predictor of and contributor to the pathobiology of cardiovascular diseases. However, most in vitro studies are performed using monolayer cultures of ECs on 2D tissue polystyrene plates (TCPs), which cannot reflect the physiological characteristics of cells in vivo. Here, we used 2D TCPs and a 3D culture model to investigate the effects of dimensionality and cardiovascular risk factors in regulating endothelial dysfunction. Cell morphology, oxidative stress, inflammatory cytokines and endothelial function were investigated in human umbilical vein endothelial cells (HUVECs) cultured in 2D/3D. The differentially expressed genes in 2D/3D-cultured HUVECs were analysed using Enrichr, Cytoscape and STRING services. Finally, we validated the proteins of interest and confirmed their relevance to TNF-α and the culture microenvironment. Compared with 2D TCPs, 3D culture increased TNF-α-stimulated oxidative stress and the inflammatory response and changed the mediators secreted by ECs. In addition, the functional characteristics, important pathways and key proteins were determined by bioinformatics analysis. Furthermore, we found that some key proteins, notably ACE, CD40, Sirt1 and Sirt6, represent a critical link between endothelial dysfunction and dimensionality, and these proteins were screened by bioinformatics analysis and verified by western blotting. Our observations provide insight into the interdependence between endothelial dysfunction and the complex microenvironment, which enhances our understanding of endothelial biology or provides a therapeutic strategy for cardiovascular-related diseases.

Gelatin-based perfusable, endothelial carotid artery model for the study of atherosclerosis.

BACKGROUND: Carotid artery geometry is important for recapitulating a pathophysiological microenvironment to study wall shear stress (WSS)-induced endothelial dysfunction in atherosclerosis. Endothelial cells (ECs) cultured with hydrogel have been shown to exhibit in vivo-like behaviours. However, to date, studies using hydrogel culture have not fully recapitulated the 3D geometry and blood flow patterns of real-life healthy or diseased carotid arteries. In this study, we developed a gelatin-patterned, endothelialized carotid artery model to study the endothelium response to WSS. RESULTS: Two representative regions were selected based on the computational fluid dynamics on the TF-shaped carotid artery: Region ECA (external carotid artery) and Region CS (carotid sinus). Progressive elongation and alignment of the ECs in the flow direction were observed in Region ECA after 8, 16 and 24 h. However, the F-actin cytoskeleton remained disorganized in Region CS after 24 h. Further investigation revealed that expression of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) was greatly increased in Region CS relative to that in Region ECA. The physiological WSS in the carotid artery system was found to stimulate nitric oxide (NO) and prostacyclin (PGI2) release and inhibit endothelin-1 (ET-1) release after 24-h perfusion experiments. The effective permeability (E.P) of fluorescein isothiocyanate (FITC)-dextran 40 kDa in Regions ECA and CS was monitored, and it was found that the turbulence WSS value (in Region CS) was less than 0.4 Pa, and there was a significant increase in the E.P relative to that in Region ECA, in which laminar WSS value was 1.56 Pa. The tight junction protein (ZO-1) production was shown that the low WSS in Region CS induced ZO-1-level downregulation compared with that in Region ECA. CONCLUSIONS: The results suggested that the gelatin-based perfusable, endothelial carotid artery model can be effective for studying the pathogenesis of atherosclerosis by which flow dynamics control the endothelium layer function in vitro.

A study of cryogenic tissue-engineered liver slices in calcium alginate gel for drug testing.

To address issues such as transportation and the time-consuming nature of tissue-engineered liver for use as an effective drug metabolism and toxicity testing model, "ready-to-use" cryogenic tissue-engineered liver needs to be studied. The research developed a cryogenic tissue-engineered liver slice (TELS), which comprised of HepG2 cells and calcium alginate gel. Cell viability and liver-specific functions were examined after different cryopreservation and recovery culture times. Then, cryogenic TELSs were used as a drug-testing model and treated with Gefitinib. Cryogenic TELSs were stored at -80 °C to ensure high cell viability. During recovery in culture, the cells in the cryogenic TELS were evenly distributed, massively proliferated, and then formed spheroid-like aggregates from day 1 to day 13. The liver-specific functions in the cryogenic TELS were closely related to cryopreservation time and cell proliferation. As a reproducible drug-testing model, the cryogenic TELS showed an obvious drug reaction after treatment with the Gefitinib. The present study shows that the cryopreservation techniques can be used in drug-testing models.

Fabrication of circular microfluidic network in enzymatically-crosslinked gelatin hydrogel.

It is a huge challenge to engineer vascular networks in vital organ tissue engineering. Although the incorporation of artificial microfluidic network into thick tissue-engineered constructs has shown great promise, most of the existing microfluidic strategies are limited to generate rectangle cross-sectional channels rather than circular vessels in soft hydrogels. Here we present a facile approach to fabricate branched microfluidic network with circular cross-sections in gelatin hydrogels by combining micromolding and enzymatically-crosslinking mechanism. Partially crosslinked hydrogel slides with predefined semi-circular channels were molded, assembled and in situ fully crosslinked to form a seamless and circular microfluidic network. The bonding strength of the resultant gelatin hydrogels was investigated. The morphology and the dimension of the resultant circular channels were characterized using scanning electron microscopy (SEM) and micro-computerized tomography (µCT). Computational fluid dynamic simulation shows that the fabrication error had little effect on the distribution of flow field but affected the maximum velocity in comparison with designed models. The microfluidic gelatin hydrogel facilitates the attachment and spreading of human umbilical endothelial cells (HUVECs) to form a uniform endothelialized layer around the circular channel surface, which successfully exhibited barrier functions. The presented method might provide a simple way to fabricate circular microfluidic networks in biologically-relevant hydrogels to advance various applications of in vitro tissue models, organ-on-a-chip systems and tissue engineering.