SARS-CoV-2 Protein Nanoparticle Vaccines Formed In Situ From Lyophilized Lipids.

The receptor binding domain (RBD) of the SARS-CoV-2 Spike (S) glycoprotein is an appealing immunogen, but associated vaccine approaches must overcome the hapten-like nature of the compact protein and adapt to emerging variants with evolving RBD sequences. Here, a vaccine manufacturing methodology is proposed comprising a sterile-filtered freeze-dried lipid cake formulation that can be reconstituted with liquid proteins to instantaneously form liposome-displayed protein nanoparticles. Mannitol is used as a bulking agent and a small amount of Tween-80 surfactant is required to achieve reconstituted submicron particles that do not precipitate prior to usage. The lipid particles include an E. coli-derived monophosphoryl lipid A (EcML) for immunogenicity, and cobalt porphyrin-phospholipid (CoPoP) for antigen display. Reconstitution of the lipid cake with aqueous protein results in rapid conversion of the RBD into intact liposome-bound format prior to injection. Protein particles can readily be formed with sequent-divergent RBD proteins derived from the ancestral or Omicron strains. Immunization of mice elicits antibodies that neutralize respective viral strains. When K18-hACE2 transgenic mice are immunized and challenged with ancestral SARS-CoV-2 or the Omicron BA.5 variant, both liquid liposomes displaying the RBD and rapid reconstituted particles protect mice from infection, as measured by the viral load in the lungs and nasal turbinates.

A Multidrug Delivery Microrobot for the Synergistic Treatment of Cancer.

Multidrug combination therapy provides an effective strategy for malignant tumor treatment. This paper presents the development of a biodegradable microrobot for on-demand multidrug delivery. By combining magnetic targeting transportation with tumor therapy, it is hypothesized that loading multiple drugs on different regions of a single magnetic microrobot can enhance a synergistic effect for cancer treatment. The synergistic effect of using two drugs together is greater than that of using each drug separately. Here, a 3D-printed microrobot inspired by the fish structure with three hydrogel components: skeleton, head, and body structures is demonstrated. Made of iron oxide (Fe3 O4 ) nanoparticles embedded in poly(ethylene glycol) diacrylate (PEGDA), the skeleton can respond to magnetic fields for microrobot actuation and drug-targeted delivery. The drug storage structures, head, and body, made by biodegradable gelatin methacryloyl (GelMA) exhibit enzyme-responsive cargo release. The multidrug delivery microrobots carrying acetylsalicylic acid (ASA) and doxorubicin (DOX) in drug storage structures, respectively, exhibit the excellent synergistic effects of ASA and DOX by accelerating HeLa cell apoptosis and inhibiting HeLa cell metastasis. In vivo studies indicate that the microrobots improve the efficiency of tumor inhibition and induce a response to anti-angiogenesis. The versatile multidrug delivery microrobot conceptualized here provides a way for developing effective combination therapy for cancer.

Mid-term Efficacy and Safety of Drug-coated Balloon versus Nitinol Bare Metal Stent for Primary Lesions in Femoropopliteal Artery Disease.

OBJECTIVES: To compare drug-coated balloon (DCB) and bare metal stent (BMS) for primary lesions in femoropopliteal artery disease in Chinese population and to make subgroup analysis between the groups. METHODS: Patients with primary lesions who underwent BMS or DCB treatment of a single tertiary vascular center were included and followed up for 24 months. Clinical and anatomic status were reported using the criteria recommended by the Society for Vascular Surgery. The primary endpoint included primary patency, clinically target limb revascularization, composite safety endpoint and all-cause death over 24 months assessed by Kaplan-Meier. Secondary endpoints included technical success rate and stent-related complications. RESULTS: A total of 284 patients with 324 limbs were pooled into analysis and most of the baseline characteristics did not show significant difference. A total of 74 in BMS group and 71 in DCB group were claudicants while 83 in BMS group and 56 in DCB group suffered from chronic limb threatening ischemia (CLTI). The mean cumulative lesion length was 18.7 ± 9.8cm in BMS group while 17.2 ± 10.3cm in DCB group. Kaplan-Meier estimates of primary patency were 75.3% and 80.9% for BMS and DCB groups at 12 months while decreased to 63.9% and 70.2% at 24 months (log-rank P = 0.167), respectively. Freedom from clinically driven target limb revascularization was 86.8% and 92.7% for BMS and DCB groups at 12 months while dropped to 82.5% and 85.9% at 24 months (log-rank P = 0.342). Estimates of primary patency between BMS and DCB group did not show significant difference on lesions with poor runoff (58.8% vs. 67.3%, log-rank P = 0.127), severe calcification (64.5% vs. 69.4%, log-rank P = 0.525) and popliteal artery involvement (59.3% vs. 60.3%, log-rank P = 0.695) at 24 months. The overall survival (92.6% for BMS, 90.3% for DCB, log-rank P = 0.391) and freedom from composite safety endpoint (79.3% for BMS, 79.2% for DCB, log-rank P = 0.941) showed no significant difference at 24 months. CONCLUSIONS: Over the 24 month follow-up, BMS and DCB showed equivalent efficacy and safety outcomes for primary femoropopliteal artery disease, which indicated the reduction of permanent metallic implant insertion might be possible.

Chitosan and Whey Protein Bio-Inks for 3D and 4D Printing Applications with Particular Focus on Food Industry.

The application of chitosan (CS) and whey protein (WP) alone or in combination in 3D/4D printing has been well considered in previous studies. Although several excellent reviews on additive manufacturing discussed the properties and biomedical applications of CS and WP, there is a lack of a systemic review about CS and WP bio-inks for 3D/4D printing applications. Easily modified bio-ink with optimal printability is a key for additive manufacturing. CS, WP, and WP-CS complex hydrogel possess great potential in making bio-ink that can be broadly used for future 3D/4D printing, because CS is a functional polysaccharide with good biodegradability, biocompatibility, non-immunogenicity, and non-carcinogenicity, while CS-WP complex hydrogel has better printability and drug-delivery effectivity than WP hydrogel. The review summarizes the current advances of bio-ink preparation employing CS and/or WP to satisfy the requirements of 3D/4D printing and post-treatment of materials. The applications of CS/WP bio-ink mainly focus on 3D food printing with a few applications in cosmetics. The review also highlights the trends of CS/WP bio-inks as potential candidates in 4D printing. Some promising strategies for developing novel bio-inks based on CS and/or WP are introduced, aiming to provide new insights into the value-added development and commercial CS and WP utilization.

Hsp27 Responds to and Facilitates Enterovirus A71 Replication by Enhancing Viral Internal Ribosome Entry Site-Mediated Translation.

Enterovirus 71 (EV-A71) is a human pathogen that causes hand, foot, and mouth disease (HFMD) and fatal neurological diseases, and no effective treatment is available. Characterization of key host factors is important for understanding its pathogenesis and developing antiviral drugs. Here we report that Hsp27 is one of the most upregulated proteins in response to EV-A71 infection, as revealed by two-dimensional gel electrophoresis-based proteomics studies. Depletion of Hsp27 by small interfering RNA or CRISPR/Cas9-mediated knockout significantly inhibited viral replication, protein expression, and reproduction, while restoration of Hsp27 restored such virus activities. Furthermore, we show that Hsp27 plays a crucial role in regulating viral internal ribosome entry site (IRES) activities by two different mechanisms. Hsp27 markedly promoted 2Apro-mediated eukaryotic initiation factor 4G cleavage, an important process for selecting and initiating IRES-mediated translation. hnRNP A1 is a key IRES trans-acting factor (ITAF) for enhancing IRES-mediated translation. Surprisingly, knockout of Hsp27 differentially blocked hnRNP A1 but not FBP1 translocation from the nucleus to the cytoplasm and therefore abolished the hnRNP A1 interaction with IRES. Most importantly, the Hsp27 inhibitor 1,3,5-trihydroxy-13,13-dimethyl-2H-pyran [7,6-b] xanthone (TDP), a compound isolated from a traditional Chinese herb, significantly protected against cytopathic effects and inhibited EV-A71 infection. Collectively, our results demonstrate new functions of Hsp27 in facilitating virus infection and provide novel options for combating EV-A71 infection by targeting Hsp27.IMPORTANCE Outbreaks of infections with EV-A71, which causes hand, foot, and mouth disease, severe neurological disorders, and even death, have been repeatedly reported worldwide in recent decades and are a great public health problem for which no approved treatments are available. We show that Hsp27, a heat shock protein, supports EV-A71 infection in two distinct ways to promote viral IRES-dependent translation. A small-molecule Hsp27 inhibitor isolated from a traditional Chinese medicinal herb effectively reduces virus yields. Together, our findings demonstrate that Hsp27 plays an important role in EV-A71 infection and may serve as an antiviral target.

Epigallocatechin-3-Gallate Reduces Cytotoxic Effects Caused by Dental Monomers: A Hypothesis.

Resin monomers from dental composite materials leached due to incomplete polymerization or biodegradation may cause contact allergies and damage dental pulp. The cytotoxicity of dental resin monomers is due to a disturbance of intracellular redox equilibrium, characterized by an overproduction of reactive oxygen species (ROS) and depletion of reduced glutathione (GSH). Oxidative stress caused by dental resin monomers leads to the disturbance of vital cell functions and induction of cell apoptosis in affected cells. The nuclear factor-erythroid 2-related factor 2 (Nrf2) pathway plays a key role in the cellular defense system against oxidative and electrophilic stress. Epigallocatechin-3-gallate (EGCG) can activate the Nrf2 pathway and induce expression of a multitude of antioxidants and phase II enzymes that can restore redox homeostasis. Therefore, here, we tested the hypothesis that EGCG-mediated protection against resin monomer cytotoxicity is mediated by activation of the Nrf2 pathway. This study will help to elucidate the mechanism of resin monomer cytotoxicity and provide information that will be helpful in improving the biocompatibility of dental resin materials.

Cloud point extraction-flame atomic absorption spectrometry method for preconcentration and determination of trace cadmium in water samples.

A method based on cloud point extraction (CPE) separation/preconcentration of trace cadmium (Cd) as a prior step to its determination by flame atomic absorption spectrometry has been developed. Cadmium reacted with 8-hydroxyquinoline to form hydrophobic chelates, which were extracted into the micelles of nonionic surfactant oligoethylene glycol monoalkyl ether (Genapol X-080) in an alkaline medium. Octanol was used to depress the cloud point of Genapol X-080 in the extraction process. The chemical variables that affect the CPE, such as pH of complexation reaction, amount of chelating agent, Genapol X-080 and octanol were evaluated and optimized. Under optimized conditions, linearity was obeyed in the range of 10-500 µg/L, with the correlation coefficient of 0.9993. For 5 mL of sample solution, the enhancement factor was about 20. The limit of detection and limit of quantification of the method were 0.21 and 0.63 µg/L, respectively. The relative standard deviations (n = 6) was 3.2% for a solution containing 100 µg/L of Cd. The accuracy of the preconcentration system was evaluated by recovery measurements on spiked water samples. Recoveries of spiked samples varied in the range of 94.1-103.8%.

Synthesis and evaluation of folate-mediated targeting and poly (ß-amino ester)-mediated pH-responsive delivery system of riccardin D based on the O-carboxymethylated chitosan micelles.

This study aimed to combine the active targeting function of folate (FA) receptor-mediated endocytosis with the pH-responsive drug delivery of poly (ethylene glycol)-grafted-poly (-amino ester) copolymers (PEG-PAE) in cancer targeting therapy. Herein, O-carboxymethylated chitosan (OCMC) was grafted with hydrophobic deoxycholic acid (DOCA). Further, PEG-PAE and FA-conjugated DOCA modified OCMC were synthesized to develop the potential cancer-targeted carrier (PEG-PAE-DOMC-FA), for which the structure was investigated by 1H NMR and FTIR. Then riccardin D (RD) was successfully loaded for tumor-targeted drug delivery. The particle size, zeta potential, encapsulating efficiencies, and loading content profiles of PEG-PAE-DOMC-FA/RD showed a strong dependence on the environmental pH values. The cumulative release of PEG-PAE-DOMC-FA/RD at pH 5.0 (90.63 %) was higher than pH 7.4 (51.12 %), which also indicated the pH sensitivity. Moreover, a lower IC50 and higher coumarin-6 uptake were found because of the folate-receptor-mediated endocytosis. In pharmacokinetic study, PEG-PAE-DOMC-FA/RD significantly improved the mean retention time (MRT) and AUC(0-∞) from 7.89 h and 36.1 mg/L·h of control group to 10.03 h and 123.8 mg/L·h. In the xenograft mice model, stronger antitumor efficacy and lower toxicity were confirmed. In conclusion, the multi-functional micelles could be considered as a promising vehicle for delivering hydrophobic drugs to tumors.

Investigation of mechanical properties and low-temperature degradation of dental 3Y-TZP ceramics fabricated by stereolithography in combination with microwave sintering.

The purpose of this study was to obtain dental ceramic materials with excellent mechanical properties and high resistance to low temperature degradation (LTD) via stereolithography (SLA) in combination with microwave sintering (MWS). The results have shown that the unaged MWS-1425 °C 3Y-TZP ceramics with uniform microstructure have high density up to 99.04% and excellent mechanical properties (Vickers hardness 14.07 GPa, fracture toughness 4.32 MPa m1/2, flexural strength 947.87 MPa). After 50 h of LTD, the m-phase content of MWS 3Y-TZP ceramics accounts for only 10.3%. In addition, the surface roughness increases by only 1.3 nm, the degraded depth is less than 5 µm, and the flexural strength exceeds 900 MPa. This exhibits the high resistance to LTD and excellent mechanical properties of dental 3Y-TZP ceramics can be obtained.

Electroactive, Antibacterial, and Biodegradable Poly(lactic acid) Nanofibrous Air Filters for Healthcare.

Poly(lactic acid) (PLA)-based nanofibrous membranes (NFMs) hold great potential in the field of biodegradable filters for air purification but are largely limited by the relatively low electret properties and high susceptibility to bacteria. Herein, we disclosed a facile approach to the fabrication of electroactive and antibacterial PLA NFMs impregnated with a highly dielectric photocatalyst. In particular, the microwave-assisted doping (MAD) protocol was employed to yield Zn-doped titanium dioxide (Zn-TIO), featuring the well-defined anatase phase, a uniform size of ∼65 nm, and decreased band gap (3.0 eV). The incorporation of Zn-TIO (2, 6, and 10 wt %) into PLA gave rise to a significant refinement of the electrospun nanofibers, decreasing from the highest diameter of 581 nm for pure PLA to the lowest value of 264 nm. More importantly, dramatical improvements in the dielectric constants, surface potential, and electret properties were simultaneously achieved for the composite NFMs, as exemplified by a nearly 94% increase in surface potential for 3-day-aged PLA/Zn-TIO (90/10) compared with that of pure PLA. The well regulation of morphological features and promotion of electroactivity contributed to a distinct increase in the air filtration performance, as demonstrated by 98.7% filtration of PM0.3 with the highest quality factor of 0.032 Pa-1 at the airflow velocity of 32 L/min for PLA/Zn-TIO (94/6), largely surpassing pure PLA (89.4%, 0.011 Pa-1). Benefiting from the effective generation of reactive radicals and gradual release of Zn2+ by Zn-TIO, the electroactive PLA NFMs were ready to profoundly inactivate Escherichia coli and Staphylococcus epidermidis. The exceptional combination of remarkable electret properties and excellent antibacterial performance makes the PLA membrane filters promising for healthcare.

Promotion of bone formation and antibacterial properties of titanium coated with porous Si/Ag-doped titanium dioxide.

Implant materials are mainly used to repair and replace defects in human hard tissue (bones and teeth). Titanium (Ti) and Ti alloys are widely used as implant materials because of their good mechanical properties and biocompatibilities, but they do not have the ability to induce new bone formation and have no antibacterial properties. Through surface modification, Ti and its alloys have certain osteogenic and antibacterial properties such that Ti implants can meet clinical needs and ensure integration between Ti implants and bone tissue, and this is currently an active research area. In this study, bioactive Si and Ag were introduced onto a Ti surface by plasma oxidation. The surface morphology, structure, elemental composition and valence, surface roughness, hydrophilicity and other physical and chemical properties of the coating were characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), a profiler and a contact angle meter (CA). Adhesion and extensions of osteoblasts on the surface of the material were observed by scanning electron microscopy, and mineralization of osteoblasts on the surface of the material were observed by alizarin red staining. The antibacterial properties of the material were tested by culturing Staphylococcus aureus on the surface of the material. The osteogenic properties of Ti implants with porous Si/Ag TiO2 (TCP-SA) coatings were evaluated with in vivo experiments in rats. The results showed that Si and Ag were successfully introduced onto the Ti surface by plasma oxidation, and doping with Si and Ag did not change the surface morphology of the coating. The osteoblasts showed good adhesion and extension on the surfaces of Si/Ag coated samples, and the porous Si/Ag TiO2 coating promoted cell proliferation and mineralization. The bacterial experiments showed that the porous TiO2 coatings containing Si/Ag had certain antibacterial properties. The animal experiments showed that Si/Ag-coated Ti implants promoted integration between the implants and the surrounding bone. It was concluded that the porous Si/Ag TiO2 coating on the Ti surface had good osteogenic and antibacterial properties and provides an optimal strategy for improving the osteogenic and antibacterial properties of Ti implants.

Emergency Management in a Dental Clinic During the Coronavirus Disease 2019 (COVID-19) Epidemic in Beijing.

BACKGROUND: In mid-March 2020, the World Health Organization declared that COVID-19 was to be characterised as a pandemic. The purpose of this article is to recommend emergency management procedures for dental clinics during this public health emergency. MATERIALS AND METHODS: We have implemented a series of emergency management measures to prevent cross-infection in our dental clinic during the COVID-19 pandemic, including personnel scheduling, division of the clinic into functional areas, limitation or delay of non-emergency patients, staff protection and infection controls, clinical environmental disinfection, and the use of online consultation services, among others. RESULTS: Due to public health policy and dental emergency management, the number of dental visitors to our clinic dropped sharply, and no COVID-19 suspected cases or high-risk patients received treatment. There have been no reports of infection of dental staff or patients during dental treatment in China to date. CONCLUSION: These public health policies and dental emergency management measures were effective in controlling cross-infection of COVID-19 in the dental clinic. PRACTICAL IMPLICATIONS: We share control measures for COVID-19, and hope that they will be helpful for dental professionals worldwide to continue to provide dental care in a safe and orderly manner.

Effects of nanoplastics on energy metabolism in the oriental river prawn (Macrobrachium nipponense).

Nanoplastics are common pollutants in aquatic environments and have attracted widespread research attention. However, few studies focus on the effects of nanoplastic exposure on energy metabolism in crustaceans. Accordingly, we exposed juvenile oriental river prawns (Macrobrachium nipponense) to different concentrations of 75-nm polystyrene nanoplastics (0, 5, 10, 20, and 40 mg/L) for 7, 14, 21, or 28 days. Thereafter, the effects of nanoplastic exposure on metabolite content, energy metabolism-related enzyme activity, and gene expression were evaluated. Our results showed that (1) with increasing nanoplastic concentration and exposure time, the survival rate decreased, while weight gain rate and molting number increased and then decreased; glycogen, triglyceride, and total cholesterol content all declined while lactic acid content increased with higher exposure to nanoplastic concentrations; (2) the activities of acetyl-CoA carboxylase (ACC), hexokinase (HK), carnitine palmitoyl transferase-1, pyruvate kinase (PK), lipase, and fatty acid synthase tended to decrease, while the activity of lactate dehydrogenase (LDH) increased. In particular, the activity of 6-phosphofructokinase exposed to 5 mg/L nanoplastics increased significantly (P < 0.05). (3) Expression of the metabolism-related genes 6-phosphate glucokinase (G-6-Pase), HK, PK, ACC, Acetyl-CoA-binding protein (ACBP), CPT-1, and fatty-acid-binding protein 10 (FABP 10) increased and then decreased, while expression of the LDH gene showed an upward trend. These results indicate that nanoplastics affect growth, enzyme activity, and the gene expression of energy metabolism in M. nipponense, and that high concentrations of nanoplastics have a negative impact on energy metabolism.

Development of an adverse outcome pathway for nanoplastic toxicity in Daphnia pulex using proteomics.

Nanoplastics are a growing environmental and public health concern. However, the toxic mechanisms of nanoplastics are poorly understood. Here, we evaluated the effects of spherical polystyrene nanoplastics on reproduction of Daphnia pulex and analyzed the proteome of whole animals followed by molecular and biochemical analyses for the development of an adverse outcome pathway (AOP) for these contaminants of emerging concern. Animals were exposed to polystyrene nanoplastics (0, 0.1, 0.5, 1 and 2 mg/L) via water for 21 days. Nanoplastics negatively impacted cumulative offspring production. A total of 327 differentially expressed proteins (DEPs) were identified in response to nanoplastics which were further validated from gene expression and enzyme activity data. Based on these results, we propose an AOP for nanoplastics, including radical oxygen species production and oxidative stress as the molecular initiating event (MIE); followed by changes in specific signaling pathways (Jak-STAT, mTOR and FoxO) and in the metabolism of glutathione, protein, lipids, and molting proteins; with an end result of growth inhibition and decrease reproductive output. This study serves as a foundation for the development of a mechanistic understanding of nanoplastic toxicity in crustaceans and perhaps other aquatic organisms.

Polystyrene nanoplastic induces oxidative stress, immune defense, and glycometabolism change in Daphnia pulex: Application of transcriptome profiling in risk assessment of nanoplastics.

Aquatic environments are generally contaminated with nanoplastic material. As a result, molecular mechanisms for sensitive species like Daphnia are needed, given that mechanistic nanoplastic toxicity is largely unknown. Here, global transcriptome sequencing (RNA-Seq) was performed on D. pulex neonates to quantitatively measure the expression level of transcripts. A total of 208 differentially expressed genes (DEGs) were detected in response to nanoplastic exposure for 96 h, with 107 being up-regulated and 101 down-regulated. The gene functions and pathways for oxidative stress, immune defense, and glycometabolism were identified. In this study, D. pulex neonates provide some molecular insights into nanoplastic toxicity. However, more studies on DEGs are needed to better understand the underlying mechanisms that result as a response to nanoplastic toxicity in aquatic organisms.

Cu-Co Co-Doped Microporous Coating on Titanium with Osteogenic and Antibacterial Properties.

Titanium (Ti) and its alloys are widely used in bone surgery by virtue of their excellent mechanical properties and good biocompatibility; however, complications such as loosening and sinking have been reported post-implantation. Herein we deposited a copper-cobalt (Cu-Co) co-doped titanium dioxide (TUO) coating on the surface of Ti implants by microarc oxidation. The osteogenic and antimicrobial properties of the coating were evaluated by in vitro experiments, and we also assessed ß-catenin expression levels on different sample surfaces. Our results revealed that the coating promoted the adhesion, proliferation, and differentiation of MG63 osteoblasts, and TUO coating promoted ß-catenin expression; moreover, the proliferation of Staphylococcus aureus was inhibited. To summarize, we report that Cu-Co co-doping can enhance the osteogenic and antibacterial activities of orthopedic Ti implants, leading to potentially improved clinical performance.

Significant elevation of B cells at the acute stage in enterovirus 71-infected children with central nervous system involvement.

A feature of the large outbreak of human enterovirus 71 (EV71)-associated hand-foot-and-mouth (HFMD) disease in China in 2008 was that severe cases presented with encephalitis. This study was performed to evaluate the immunophenotypic characteristics of patients with neurological involvement. Twenty-one patients with encephalitis and 14 with uncomplicated HFMD were recruited. Age-matched healthy volunteers were enrolled as controls. Peripheral lymphocyte subsets were analyzed by use of 3-colour flow cytometry, and the quantitative determination of plasma immunoglobulin (Ig) levels was also monitored. Comparisons between severe and mild cases demonstrated significant elevations of B cells and IgG levels and corresponding general decreases in natural killer (NK) cells and T lymphocytes in severe cases at the acute stage of infection (p < 0.01 for all). During the convalescent phase, rapid recoveries of B cells and IgG to the normal levels were observed, which appeared to be accompanied by an increase in EV71-specific neutralizing antibody titres. In summary, our data demonstrate that elevated B cells and IgG might be associated with neurological manifestations in EV71 infection.

Effects of nanoplastics at predicted environmental concentration on Daphnia pulex after exposure through multiple generations.

The biological effects of nanoplastics are a growing concern. However, most studies have focused on exposure to high concentrations or short-term exposure. The potential effects of exposure to low environmental nanoplastic concentrations over the long-term and across multiple generations remain unclear. In the present study, Daphnia pulex was exposed over three 21-day generations to a typical environmental nanoplastic concentration (1 µg/L) and the effects were investigated at physiological (growth and reproduction), gene transcription and enzyme activity levels. Chronic exposure did not affect the survival or body length of D. pulex, whereas the growth rate and reproduction were influenced in the F2 generation. Molecular responses indicated that environmental nanoplastic concentrations can modulate the response of antioxidant defenses, vitellogenin synthesis, development, and energy production in the F0-F1 generations, and prolongation resulted in inhibitory effects on antioxidant responses in F2 individuals. Some recovery was observed in the recovery group, but reproduction and stress defenses were significantly induced. Taken together, these results suggest that D. pulex recovery from chronic exposure to nanoplastic may take several generations, and that nanoplastics have potent long-term toxic effects on D. pulex. The findings highlight the importance of multigenerational and chronic biological evaluations to assess risks of emerging pollution.

Effects of exposure to waterborne polystyrene microspheres on lipid metabolism in the hepatopancreas of juvenile redclaw crayfish, Cherax quadricarinatus.

Previous research has identified microplastics as new environmental pollutants that are widely distributed in a variety of environments, including aquaculture environments. However, the potential hazard of microplastics to aquaculture animals, especially toward lipid metabolism involved with the survival and growth of aquatic animal, has not yet been investigated. In the present study, redclaw crayfish (Cherax quadricarinatus) were exposed to different concentrations of 200 nm-sized polystyrene microspheres (0, 0.5, and 5 mg/L) for 21 days, to investigate the effects of microplastics on lipid metabolism. After ingestion, the microplastics were distributed in the intestines and hepatopancreas, and appeared to inhibit the growth of Cherax quadricarinatus. Subsequently, the lipid levels in the hepatopancreas and hemolymph was detected, and found that after 21 days of exposure, the lipid content and free fatty acids in the hepatopancreas and hemolymph decreased significantly, and total cholesterol and triglycerides levels increased significantly in the hemolymph. This might have been caused by insufficient intake of exogenous fat. A significant decrease in lipase activity also supported this view. The activity of lipoprotein lipase related to lipolysis in the hepatopancreas increased significantly, while the activity of fatty acid synthase related to fat synthesis increased, and the activity of acetyl-CoA carboxylase decreased. These results indicated disturbed lipid metabolism in the hepatopancreas. The significant increase of lipid transport-related low-density lipoprotein indicated that the lipolytic capacity was higher than the lipid synthesis capacity. The expression levels of fatty acid metabolism-related genes FAD6 and FABP decreased significantly, indicating that the fatty acid utilization ability of hepatopancreas cells was inhibited, which was consistent with the results of enzyme activities. Thus, microplastics represent a potential hazard to redclaw crayfish, at least on lipid metabolism. This study provided basic data on the ecotoxicological effects of microplastics on crustaceans.

Polystyrene nanoplastic induces ROS production and affects the MAPK-HIF-1/NFkB-mediated antioxidant system in Daphnia pulex.

Recently, research on the biological effects of nanoplastics has grown exponentially. However, studies on the effects of nanoplastics on freshwater organisms and the mechanisms of the biological effects of nanoplastics are limited. In this study, the content of reactive oxygen species (ROS), gene and protein expression in the MAPK-HIF-1/NFkB pathway, and antioxidant gene expressions and enzyme activities were measured in Daphnia pulex exposed to polystyrene nanoplastic. In addition, the full-length extracellular signal-regulated kinases (ERK) gene, which plays an important role in the MAPK pathway, was cloned in D. pulex, and the amino acid sequence, function domain, and phylogenetic tree were analyzed. The results show that nanoplastic caused the overproduction of ROS along with other dose-dependent effects. Low nanoplastic concentrations (0.1 and/or 0.5 mg/L) significantly increased the expressions of genes of the MAPK pathway (ERK; p38 mitogen-activated protein kinases, p38; c-Jun amino-terminal kinases, JNK; and protein kinase B, AKT), HIF-1 pathway (prolyl hydroxylasedomain, PHD; vascular endothelial growth factor, VEGF; glucose transporter, GLUT; pyruvate kinase M, PKM; hypoxia-inducible factor 1, HIF1), and CuZn superoxide dismutase (SOD) along with the activity of glutathione-S-transferase. As the nanoplastic concentration increased, these indicators were significantly suppressed. The protein expression ratio of ERK, JNK, AKT, HIF1α, and NFkBp65 (nuclear transcription factor-kB p65) as well as the phosphorylation of ERK and NFkBp65 were increased in a dose-dependent manner. The activities of other antioxidant enzymes (catalase, total SOD, and CuZn SOD) were significantly decreased upon exposure to nanoplastic. Combined with our previous work, these results suggest that polystyrene nanoplastic causes the overproduction of ROS and activates the downstream pathway, resulting in inhibited growth, development, and reproduction. The present study fosters a better understanding of the biological effects of nanoplastics on zooplankton.