Inhibiting ferroptosis in brain microvascular endothelial cells: A potential strategy to mitigate polystyrene nanoplastics‒induced blood‒brain barrier dysfunction.

Polystyrene nanoplastics (PS-NPs), a group of ubiquitous pollutants, may injure the central nervous system through the blood‒brain barrier (BBB). However, whether exposure to PS-NPs contributes to BBB disruption and the underlying mechanisms are still unclear. In vivo, we found that PS-NPs (25 mg/kg BW) could significantly increase BBB permeability in mice and downregulate the distribution of the tight junction-associated protein zona occludens 1 (ZO-1) in brain microvascular endothelial cells (BMECs). Using an in vitro BBB model, exposure to PS-NPs significantly reduced the transendothelial electrical resistance and altered ZO-1 expression and distribution in a dose-dependent manner. RNA-seq analysis and functional investigations were used to investigate the molecular pathways involved in the response to PS-NPs. The results revealed that the ferroptosis and glutathione metabolism signaling pathways were related to the disruption of the BBB model caused by the PS-NPs. PS-NPs treatment promoted ferroptosis in bEnd.3 cells by inducing disordered glutathione metabolism in addition to Fe2+ and lipid peroxide accumulation, while suppressing ferroptosis with ferrostatin-1 (Fer-1) suppressed ferroptosis-related changes in bEnd.3 cells subjected to PS-NPs. Importantly, Fer-1 alleviated the decrease in ZO-1 expression in bEnd.3 cells and the exacerbation of BBB damage induced by PS-NPs. Collectively, our findings suggest that inhibiting ferroptosis in BMECs may serve as a potential therapeutic target against BBB disruption induced by PS-NPs exposure.

Highly efficient mesophyll protoplast isolation and PEG-mediated transient gene expression for rapid and large-scale gene characterization in cassava (Manihot esculenta Crantz).

BACKGROUND: Cassava (Manihot esculenta Crantz) is a major crop extensively cultivated in the tropics as both an important source of calories and a promising source for biofuel production. Although stable gene expression have been used for transgenic breeding and gene function study, a quick, easy and large-scale transformation platform has been in urgent need for gene functional characterization, especially after the cassava full genome was sequenced. METHODS: Fully expanded leaves from in vitro plantlets of Manihot esculenta were used to optimize the concentrations of cellulase R-10 and macerozyme R-10 for obtaining protoplasts with the highest yield and viability. Then, the optimum conditions (PEG4000 concentration and transfection time) were determined for cassava protoplast transient gene expression. In addition, the reliability of the established protocol was confirmed for subcellular protein localization. RESULTS: In this work we optimized the main influencing factors and developed an efficient mesophyll protoplast isolation and PEG-mediated transient gene expression in cassava. The suitable enzyme digestion system was established with the combination of 1.6% cellulase R-10 and 0.8% macerozyme R-10 for 16 h of digestion in the dark at 25 °C, resulting in the high yield (4.4 × 107 protoplasts/g FW) and vitality (92.6%) of mesophyll protoplasts. The maximum transfection efficiency (70.8%) was obtained with the incubation of the protoplasts/vector DNA mixture with 25% PEG4000 for 10 min. We validated the applicability of the system for studying the subcellular localization of MeSTP7 (an H+/monosaccharide cotransporter) with our transient expression protocol and a heterologous Arabidopsis transient gene expression system. CONCLUSION: We optimized the main influencing factors and developed an efficient mesophyll protoplast isolation and transient gene expression in cassava, which will facilitate large-scale characterization of genes and pathways in cassava.

A pH-neutral bioactive glass coated 3D-printed porous Ti6Al4V scaffold with enhanced osseointegration.

Osseointegration is vital for the success of non-degradable implants like those made of titanium alloys. In order to promote osseointegration, implants are made porous, providing space for bone ingrowth. Despite extensive optimization of the pore geometry and porosity, bone ingrowth into implants is still marginal; further modification to promote bone ingrowth as well as osseointegration becomes paramount. In this study, a pH neutral bioactive glass with the composition of 10.8% P2O5-54.2% SiO2-35% CaO (mol%; hereinafter referred to as PSC) was successfully coated on 3D-printed porous Ti6Al4V scaffolds using an in situ sol-gel method. This PSC coating is strongly bonded to the substrate and quickly induces the formation of hydroxyapatite on the scaffold surface upon contact with body fluid. In vitro, the PSC-coated Ti6Al4V scaffolds showed superior biocompatibility, cell proliferation promotion, cell adhesion, osteogenic differentiation and mineralization compared to their bare counterparts, implying better osseointegration. In vivo experiments confirmed this expectation; after being implanted, the coated scaffolds had more bone ingrowth and osseointegration, and consequently, higher push-out strength was achieved, proving the validity of the proposed concept in this study. In conclusion, PSC coating on 3D-printed porous Ti6Al4V scaffolds can improve osteogenesis, bone ingrowth, and osseointegration. Together with the versatility of this in situ sol-gel coating method, titanium alloy implants with better biological performances may be developed for immediate clinical applications.

Modified Medial Collateral Ligament Indentation Technique in Total Knee Arthroplasty with Severe Type II Valgus Deformity.

OBJECTIVE: To explore the feasibility and clinical efficacy of a modified medial collateral ligament indentation technique in total knee arthroplasty (TKA) with severe type II valgus deformity. METHODS: Consecutive patients with Krackow type II valgus deformity >20° who underwent a primary unilateral TKA between May 2008 and June 2017 were studied retrospectively. A medial collateral ligament indentation technique was performed in 20 patients (MCLI group), and 23 patients received the routine lateral structures release technique (LSR group). Radiological parameters, such as the valgus angle (VA), and functional outcomes including the use of constraint implants, Knee Society Score (KSS), Knee Society Function score (KSF), and thickness of the polyethylene insert were compared between the two groups. RESULTS: A total of 43 consecutive patients had a minimum 2-year follow-up. The preoperative VA was comparable between the MCLI (23.5° ± 5.8°) and LSR groups (21.3° ± 3.2°, P = 0.134), as was the postoperative VA (1.1° ± 2.1° and 2.5° ± 3.0°, respectively, P = 0.084). The mean KSS and KSF scores in the MCLI group were 30.2 ± 4.8 and 38.8 ± 4.8, respectively, before surgery, and they increased to 91.3 ± 2.6 and 86.5 ± 2.4 at the last follow-up. The scores in the LSR group were 31.5 ± 7.5 and 36.5 ± 7.8 before surgery and 92.4 ± 3.5 and 88.5 ± 3.6 at the last follow-up. While no statistically significant differences in pre- or postoperative functional scores were found between the two groups, the MCLI group had thinner polyethylene inserts (9.5 ± 1.1 mm vs 12.9 ± 1.5 mm) and less use of constrained condylar inserts (15% vs 69.6%). During follow-up, the MCLI group had fewer complications. CONCLUSION: A modified MCLI technique can achieve good outcomes in TKA with type II valgus deformity of >20°. It can maintain a normal joint line level, reduce the use of constrained condylar knee prostheses, and is a reliable choice for severe genu valgum.