An evaluation of a hepatotoxicity risk induced by the microplastic polymethyl methacrylate (PMMA) using HepG2/THP-1 co-culture model.

Inappropriate disposal of plastic wastes and their durability in nature cause uncontrolled accumulation of plastic in land/marine ecosystems, also causing destructive effects by bioaccumulating along the food chain. Microplastics may cause chronic inflammation in relation to their permanent structures, especially through oxidative stress and cytotoxic cellular damage, which could increase the risk of cancer development. The accumulation of microplastics in the liver is a major concern, and therefore, the identification of the mechanisms of their hepatotoxic effects is of great importance. Polymethyl methacrylate (PMMA) is a widely used thermoplastic. It has been determined that PMMA disrupts lipid metabolism in the liver in various aquatic organisms and causes reproductive and developmental toxicity. PMMA-induced hepatotoxic effects in humans have not yet been clarified. In our study, the toxic effects of PMMA (in the range of 3-10 µm) on the human liver were investigated using the HepG2/THP-1 macrophage co-culture model, which is a sensitive immune-mediated liver injury model. Cellular uptake of micro-sized PMMA in the cells was done by transmission electron microscopy. Determination of its effects on cell viability and inflammatory response, oxidative stress, along with gene and protein expression levels that play a role in the mechanism pathways underlying the effects were investigated. The results concluded that inflammation, oxidative stress, and disruptions in lipid metabolism should be the focus of attention as important underlying causes of PMMA-induced hepatotoxicity. Our study, which points out the potential adverse effects of microplastics on human health, supports the literature information on the subject.

Effects of polymethylmethacrylate nanoplastics on the polychaete Hediste diversicolor: Behavioural, regenerative, and biochemical responses.

Plastics, particularly microplastics (MPs) and nanoplastics (NPs), have been regarded as pollutants of emerging concern due to their effects on organisms and ecosystems, especially considering marine environments. However, in terms of NPs, there is still a knowledge gap regarding the effects of size and polymer on marine invertebrates, such as benthic organisms. Therefore, this study aimed to understand, regarding behavioural, physiological, and biochemical endpoints (neurotransmission, energy metabolism, antioxidant status, and oxidative damage), the effects of 50 nm waterborne polymethylmethacrylate (PMMA) NPs (0.5 to 500 µg/L) on the marine benthic polychaete Hediste diversicolor, a key species in estuarine and coastal ecosystems. Results demonstrated that worms exposed to PMMA NPs had a shorter burrowing time than control organisms. Nevertheless, worms exposed to PMMA NPs (0.5 and 500 µg/L) decreased cholinesterase activity. Energy metabolism was decreased at 50 and 500 µg/L, and glycogen content decreased at all concentrations of PMMA NPs. Enzymes related to the antioxidant defence system (superoxide dismutase and glutathione peroxidase) displayed increased activities in H. diversicolor specimens exposed to concentrations between 0.5 and 500 µg/L, which led to no damage at the cell membrane and protein levels. In this study, polychaetes also displayed a lower regenerative capacity when exposed to PMMA NPs. Overall, the data obtained in this study emphasize the potential consequences of PMMA NPs to benthic worms, particularly between 0.5 and 50 µg/L, with polychaetes exposed to 50 µg/L being the most impacted by the analysed NPs. However, since sediments are considered to be sinks and sources of plastics, further studies are needed to better understand the impacts of different sizes and polymers on marine organisms, particularly benthic species.

In vitro assessment of inflammatory skin potential of poly(methyl methacrylate) at non-cytotoxic concentrations.

Poly(methyl methacrylate) (PMMA) is considered an attractive substrate material for fabricating wearable skin sensors such as fitness bands and microfluidic devices. Despite its widespread use, inflammatory and allergic responses have been attributed to the use of this material. Therefore, the main objective of this study was to obtain a comprehensive understanding of potential biological effects triggered by PMMA at non-cytotoxic concentrations using in vitro models of NIH3T3 fibroblasts and reconstructed human epidermis (RhE). It was hypothesized that concentrations that do not reduce cell viability are sufficient to activate pathways of inflammatory processes in the skin. The study included cytotoxicity, cell metabolism, cytokine quantification, histopathological, and gene expression analyses. The NIH3T3 cell line was used as a testbed for screening cell toxicity levels associated with the concentration of PMMA with different molecular weights (MWs) (i.e., MW ~5,000 and ~15,000 g/mol). The lower MW of PMMA had a half-maximal inhibitory concentration (IC50 ) value of 5.7 mg/cm2 , indicating greater detrimental effects than the higher MW (IC50 = 14.0 mg/cm2 ). Non-cytotoxic concentrations of 3.0 mg/cm2 for MW ~15,000 g/mol and 0.9 mg/cm2 for MW ~5,000 g/mol) induced negative metabolic changes in NIH3T3 cells. Cell viability was severely reduced to 7% after the exposure to degradation by-products generated after thermal and photodegradation degradation of PMMA. PMMA at non-cytotoxic concentrations still induced overexpression of pro-inflammatory cytokines, chemokines, and growth factors (IL1B, CXCL10, CCL5, IL1R1, IL7, IL17A, VEGFA, FGF2, IFNG, IL15) on the RhE model. The inflammatory response was also supported by histopathological and gene expression analyses of PMMA-treated RhE, indicating tissue damage and gene overexpression. Results suggested that non-cytotoxic concentrations of PMMA (3.0 to 5.6 mg/cm2 for MW ~15,000 g/mol and 0.9 to 2.1 mg/cm2 for MW ~5,000 g/mol) were sufficient to negatively alter NIH3T3 cells metabolism and activate inflammatory events in the RhE skin.

UV-B radiation alleviated detrimental effects of polymethyl methacrylate microplastics on marine diatom Thalassiosira pseudonana.

Microplastics (MPs) in marine environments simultaneously affect microalgae with UV-B radiation, while their joint effecting mechanisms remain largely unknown. To fill this research gap, the joint effects of polymethyl methacrylate (PMMA) MPs and UV-B radiation (natural environments intensity) on the model marine diatom Thalassiosira pseudonana were investigated. Antagonism was found between the two factors with regards to population growth. Furthermore, we found more inhibited population growth and photosynthetic parameters when pre-treated with PMMA MPs compared to pre-treated with UV-B radiation before joint-treated by the two factors. Transcriptional analysis elucidated that UV-B radiation could alleviate the down-regulation of photosynthetic (PSII, cyt b6/f complex and photosynthetic electron transport) and chlorophyll biosynthesis genes caused by PMMA MPs. Besides, the genes encoding carbon fixation and metabolisms was up-regulated under UV-B radiation, which could provide extra energy for the enhanced anti-oxidative activities and DNA replication-repair processes. These consequences showed that the toxicity of PMMA MPs was comprehensively alleviated when T. pseudonana was jointed treated by UV-B radiation. Our results reveled the underlying molecular mechanisms of antagonistic effects between PMMA MPs and UV-B radiation. This study provides important information that environmental factors like UV-B radiation should be considered when accessing the ecological risks of MPs on marine organisms.

In vitro toxicity of naringin and berberine alone, and encapsulated within PMMA nanoparticles.

Phytochemical compounds, such as naringin and berberine, have been used for many years due to their antioxidant activities, and consequently, beneficial health effects. In this study, it was aimed to evaluate the antioxidant properties of naringin, berberine and poly(methylmethacrylate) (PMMA) nanoparticles (NPs) encapsulated with naringin or berberine and their possible cytotoxic, genotoxic, and apoptotic effects on mouse fibroblast (NIH/3 T3) and colon cancer (Caco-2) cells. According to the results of the study, it was found that the 2,2-diphenyl-1-picrylhydrazyl (DPPH) inhibition antioxidant activity of naringin, berberine, and naringin or berberine encapsulated PMMA NPs, was significantly increased at higher tested concentrations due to the antioxidant effects of naringin, berberine and naringin or berberine encapsulated PMMA NPs. As a result of the cytotoxicity assay, after 24-, 48- and 72-h of exposure, all of the studied compounds caused cytotoxic effects in both cell lines. Genotoxic effects of studied compounds were not registered at lower tested concentrations. Based on these data, polymeric nanoparticles encapsulated with naringin or berberine may contribute to new treatment approaches for cancer, but further in vivo and in vitro research is required.

Responses of individual and combined polystyrene and polymethyl methacrylate nanoplastics on hormonal content, fluorescence/photochemistry of chlorophylls and ROS scavenging capacity in Lemna minor under arsenic-induced oxidative stress.

Nanoplastics alter the adverse impacts of hazardous contaminants such as heavy metals by changing their adsorption and accumulation. Few findings are available on the interaction between nanoplastic and heavy metals in plants. However, there is no report on the mechanisms for removing metal stress-mediated oxidative damage by the combination treatments of nanoplastics. To address this lack of information, polystyrene nanoplastic (PS, 100 mg L-1) and polymethyl methacrylate (PMMA, 100 mg L-1) were hydroponically applied to Lemna minor exposed to arsenate (As, 100 µM) for 7 days. PS or PMMA caused a reduction in the contents of N, P, K, Ca, Mg and Mn, but the improved contents were detected in the presence of PS or PMMA plus As stress. The hormone contents (auxin, gibberellic acid, cytokinin, salicylic acid and jasmonic acid) reduced by stress were re-arranged through PS or PMMA applications. Based on chlorophyll efficiency, fluorescence kinetics and performance of PSII, the impaired photosynthesis by As stress was improved via PS or PMMA applications. This alleviation did not continue under the combined form of PS and PMMA in As-applied plants. All analyzed antioxidant activity (superoxide dismutase (SOD), catalase (CAT), peroxidase (POX), ascorbate peroxidase (APX), glutathione reductase (GR), glutathione S-transferase (GST), glutathione peroxidase (GPX), monodehydroascorbate reductase (MDHAR) and dehydroascorbate reductase (DHAR)) decreased or unchanged under As, PS or PMMA. Due to the inactivation of the defense system, L. minor had high levels of hydrogen peroxide (H2O2) and thiobarbituric acid reactive substances (TBARS), showing lipid peroxidation. After As toxicity, induvial applications of PS or PMMA indicated the activated enzyme capacity (SOD, POX, GST and GPX) and upregulated AsA/DHA, GSH/GSSG and redox state of GSH, which facilitated the removal of radical accumulation. The efficiency of the antioxidant system in As + PS + PMMA-applied L. minor was not enough to remove damage induced by As stress; hereby, TBARS and H2O2 contents were similar to the As-treated group. Our findings from alone or combined application of PS and PMMA provide new information to advance the tolerance mechanism against As exposure in L. minor.

Understanding the biological impact of organic pollutants absorbed by nanoplastics.

Many organisms are consuming food contaminated with micro- and nanoparticles of plastics, some of which absorb persistent organic pollutants (POPs) from the environment and acting as carrier vectors for increasing the bioavailability in living organisms. We recently reported that polymethylmethacrylate (PMMA) nanoparticles at low concentrations are not toxic to animal models tested. In this study, the toxicity of diphenylamine (DPA) incorporated PMMA nanoparticles are assessed using barnacle larvae as a model organism. The absorption capacity of DPA from water for commercially available virgin PMMA microparticles is relatively low (0.14 wt%) during a 48 h period, which did not induce exposure toxicity to barnacle nauplii. Thus, PMMA nanoparticles encapsulated with high concentrations of DPA (DPA-enc-PMMA) were prepared through a reported precipitation method to achieve 40% loading of DPA inside the particles. Toxicity of DPA-enc-PMMA nanoparticles were tested using freshly spawned acorn barnacle nauplii. The observed mortality of nauplii from DPA-enc-PMMA exposure was compared to the values obtained from pure DPA exposure in water. The mortality among the exposed acorn barnacle nauplii did not exceed 50% even at a high concentration of DPA inside the PMMA nanoparticles. The results suggest that the slow release of pollutants from polymer nanoparticles may not induce significant toxicity to the organism living in a dynamic environment. The impact of long-term exposure of DPA absorbed plastic nanoparticles need to be investigated in the future.

Effects of nanoplastics on zebrafish embryo-larval stages: A case study with polystyrene (PS) and polymethylmethacrylate (PMMA) particles.

Plastic production has been rising consistently in the last 30 years and with it, the presence of plastic particles in the environment. A decrease in size often increases the bioavailability and reactivity of the particles. In this study the impact of polystyrene (PS; 22 nm) and polymethylmethacrylate (PMMA; 32 nm) nanoparticles on zebrafish embryo-larval stages was assessed by studying mortality, hatching, morphological features, and biochemical endpoints (associated with neurotransmission, antioxidant status and oxidative damage, and energy metabolism) after 96 h exposure, and swimming behavior after 120 h exposure. Organisms exposed to PMMA nanoparticles exhibited higher mortality and pericardial edema than those exposed to PS nanoparticles but displayed less effects on swimming behavior. Biochemical endpoints were altered to a higher degree in organisms exposed to PS nanoparticles (acetylcholinesterase, glutathione S-transferase and catalase activities) but higher peroxidative damage was found after exposure to lower concentrations of PMMA nanoparticles. Both types of nanoparticles affected energy metabolism with higher levels of glycogen found in animals exposed to PS nanoparticles. The use of integrated biomarker response index confirmed that PS nanoparticles had a higher impact on biochemical endpoints of zebrafish.

Polymethyl methacrylate resin for provisional restoration affects rat macrophage function in in vitro conditions.

PURPOSE: This study evaluated the cytotoxic effects of polymethyl methacrylate resin extracts on rat macrophage viability in in vitro conditions. METHODS: Prepared test specimens were immersed in 5 mL of artificial saliva and incubated for 24, 48, and 72 h at 37°C. The cytotoxicity of the obtained solutions of extracted resins, used as a stock solution (100%) and diluted with Roswell Park Memorial Institute (RPMI) medium to obtain the working solutions (50, 40, 30, 20, 10, and 5%), was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. RESULTS: No dose-dependent toxic activity in macrophage culture was detected for the three types of extracts obtained after 24, 48, and 72 h of material extraction. The shortest extraction of material was found to be completely non-toxic, and the 20% concentration of this extract obtained caused a significant increase in cell ability to metabolize MTT. Extracts obtained after 72 h of extraction showed the highest cytotoxic potential of 50%, 40% and 30%, and extracts obtained after 48 and 72 h of extraction at concentrations of 5% and 10% had a proliferative effect on the macrophage cell line. CONCLUSION: This study demonstrated that the highest cytotoxic effect was observed in cells exposed to the highest concentrations (50, 40, and 30%) of the extracts that were extracted for 72 h.

The hormetic dose-risks of polymethyl methacrylate nanoplastics on chlorophyll a fluorescence transient, lipid composition and antioxidant system in Lactuca sativa.

Nanoplastic pollution has become an increasing problem due to over-consumption and degradation in ecosystems. A little is known about ecological toxicity and the potential risks of nanoplastics on plants. To better comprehend the hormetic effects of nanoplastics, the experimental design was conducted on the impacts of polymethyl methacrylate (PMMA) on water status, growth, gas exchange, chlorophyll a fluorescence transient, reactive oxygen species (ROS) content (both content and fluorescence visualization), lipid peroxidation and antioxidant capacity (comparatively between leaves and roots). For this purpose, PMMA (10, 20, 50 and 100 mg L-1) was hydroponically applied to Lactuca sativa for 15 days(d). PMMA exposure resulted a decline in the growth, water content and osmotic potential. As based on assimilation rate (A), stomatal conductance (gs), and intercellular CO2 concentrations (Ci), the decreased stomatal limitation (Ls) and, A/Ci and increased intrinsic mesophyll efficiency proved low carboxylation efficiency showing impaired photosynthesis as a non-stomatal limitation. PMMA toxicity increased the trapping fluxes and absorption with a decrease in electron transport fluxes caused the disruption in reaction centers of photosystems. The leaves and roots had a similar effect against PMMA toxicity, with increased superoxide dismutase (SOD) activity. Although, catalase (CAT) and peroxidase (POX) of leaves increased under 10 mg L-1 PMMA, these defense activities failed to prevent radicals from attacking. Compared to the leaves, the lettuce roots showed an intriguing result for AsA-GSH cycle against PMMA exposure. In the roots, the lowest PMMA application provided the high ascorbate/dehydroascorbate (AsA/DHA), GSH/GSSG and the pool of AsA/glutathione (GSH) and non-suppressed GSH redox state. Also, 10 mg L-1 PMMA helped remove high hydrogen peroxide (H2O2) by both glutathione peroxidase (GPX) and glutathione S-transferase (GST). Since this improvement in the antioxidant system could not be continued in roots after higher applications than 20 mg L-1 PMMA, TBARS (Thiobarbituric acid-reactive substances), indicating the level of lipid peroxidation, and H2O2 increased. Our findings obtained from PMMA-applied lettuce provide new information to advance the tolerance mechanism against nanoplastic pollution.

Polymethylmethacrylate nanoplastics can cause developmental malformations in early life stages of Xenopus laevis.

Polymethylmethacrylate (PMMA) production has increased almost 20% over the last years. With its release into the aquatic environment, its breakdown or degradation to nano dimensions (nanoplastics-NPLs) due to biological and physical/mechanical action is, theoretically, anticipated. The occurrence of PMMA-NPLs in aquatic ecosystems may thus cause adverse effects particularly to early life stages of amphibians, which may be in contact with PMMA-NPLs suspended in the water column or deposited in upper layers of the sediments. Accordingly, this work aimed at assessing the effects of PMMA-NPLs to aquatic early life stages of the model anuran species Xenopus laevis. To attain this objective, two types of toxicity assays were carried out by exposing embryos [Nieuwkoop and Faber (NF) stage 8-11] or tadpoles (NF 45) to three concentrations of PMMA-NPLs (1, 100 and 1000 µg/L): i) 96-h embryo teratogenicity assay, where survival, malformation, and total body length (BL) of embryos were assessed; and ii) 48-h feeding rate assay, where survival, feeding (FR), malformations and growth rates (body weight-BW and BL) of tadpoles were evaluated. PMMA-NPLs exposure had no significant effects on mortality, malformations of X. laevis embryos but BL was lower at 1000 µg PMMA-NPLs/L. In tadpoles, no effects on survival or FR were observed after exposure to PMMA-NPLs, but significant changes occured in BW and BL. Moreover, anatomical changes in the abdominal region (externalization of the gut) were observed in 62.5% of the tadpoles exposed to 1000 µg PMMA-NPLs/L. Despite the lack of knowledge regarding the environmental levels of NPLs, it is expected that sediments constitute a sink for these contaminants, where they can become available for organisms that, like tadpoles, feed on the organic matter at the surface of sediments. Considering the continuous release and subsequent accumulation of PMMA, the malformations obtained in the feeding assays suggest that, in the future, these nano-polymers may constitute a risk for aquatic life stages of amphibians.

Interactive effects of polymethyl methacrylate (PMMA) microplastics and salinity variation on a marine diatom Phaeodactylum tricornutum.

Until now, knowledge about the interactive effects of microplastics and environmental factors on primary producers is quite limited. In this work, a marine diatom (Phaeodactylum tricornutum) was exposed to polymethyl methacrylate (PMMA) microplastics at different salinities (25, 35, and 45‰) for 10 days in order to study their interactive effects. Results showed that growth of P. tricornutum was negatively affected by PMMA microplastics and salinity variation with a minimum EC50 value of 91.75 mg L-1. Photosynthetic activity of P. tricornutum was also inhibited by the two factors, and their interactive effects on chlorophyll fluorescence parameters (Fv/Fm and ΦPSII) were significant. In the algal cells, soluble protein accumulated, activities of two antioxidant enzymes changed, and malondialdehyde (MDA) content increased when this diatom was exposed to the microplastics at different salinities. These data would help to evaluate the risks of microplastics to primary producers under different environmental factors.

Single and joint toxicity of polymethyl methacrylate microplastics and As (V) on rapeseed (Brassia campestris L.).

Most microplastics and arsenic (As) have been released into farmland via industrial and agricultural activities, posing a potential threat to crop growth and food safety. Thus far, few studies have focused on the phytoxicity of microplastics and As to leafy vegetable. In this study, we evaluated the single and combined toxicological effects of polymethyl methacrylate (PMMA) and As(V) on rapeseed (Brassia campestris L.). Single treatments of two sizes of PMMA particles, namely PMMA nano-plastics (PMMANPs) and PMMA micro-plastics (PMMAMPs) and As(V) significantly (P < 0.05) inhibited the germination index (GI) of rapeseed. The IC50 indicates that PMMANPs were more toxic than PMMAMPs. Combine-pollution experiments demonstrated that the GI, biomass, root length, and sprout length of the rapeseed under the combined treatment were lower than those subjected to As(V) or PMMANPs single treatment. Analysis of variance showed that the interaction effects of PMMANPs and As(V) for GI and root length were significant, and there was synergistic interaction between PMMANPs and As(V) on rapeseed germination. PMMANPs promoted the accumulation of As in sprouts under high As(V) concentrations (40 and 60 mg/L). The activities of lipase in rapeseed generally increased under single and combined treatments of As(V) and PMMANPs, and while α-amylase activities first increased and then decreased with the increase of PMMANPs. It appears that the combined stress of microplastics and As(V) exhibited synergistic interaction on the growth of rapeseed.

Postpolymerization of a 3D-printed denture base polymer: Impact of post-curing methods on surface characteristics, flexural strength, and cytotoxicity.

OBJECTIVE: This study investigated the influence of postpolymerization of a three-dimensional (3D) printed denture base polymer. The effect of post-curing methods on surface characteristics, flexural strength, and cytotoxicity was evaluated. METHODS: A total of 172 specimens were additively manufactured using one denture base material (V-Print dentbase, VOCO) and further post-cured by different light-curing devices, including Otoflash G171 (OF), Labolight DUO (LL), PCU LED (PCU), and LC-3DPrintbox (PB), respectively. Polymethyl methacrylate resin (PalaExpress Ultra) was used as a reference (REF). Afterward, surface topography was observed using scanning electron microscopy, and surface roughness was measured (n = 6). Furthermore, flexural strength was tested (n = 20). Cytotoxicity was evaluated by the extract and direct contact tests. The data were analyzed using the Kolmogorov-Smirnov test and one-way ANOVA followed by Tukey's multiple comparisons and Kruskal-Wallis tests (p < 0.05). RESULTS: The different post-curing methods applied did not significantly influence surface topography and roughness (Ra). Meanwhile, specimens post-cured by PCU (162.3 ± 44.16 MPa) and PB (171.2 ± 34.41 MPa) showed significantly higher flexural strength than those post-cured by OF (131.3 ± 32.87 MPa) and REF (131.2 ± 19.19 MPa), respectively. Additionally, various post-curing methods effectively decreased the cytotoxic effects of 3D-printed denture base polymer. CONCLUSIONS: Different post-curing methods did not significantly alter the Ra values of the 3D-printed denture base material. However, flexural strength was significantly affected by the postpolymerization methods, which might be attributed to the different wavelengths of post-curing devices. In addition, various postpolymerization methods reduced the cytotoxic effects of the 3D-printed denture base polymer. CLINICAL SIGNIFICANCE: Flexural strength of additively manufactured denture bases depends on the postpolymerization strategy. Therefore, an appropriate post-curing method is required to optimize the flexural strength of 3D-printed denture materials.

Short-term exposure to polymethylmethacrylate nanoplastics alters muscle antioxidant response, development and growth in Sparus aurata.

Polymethylmethacrylate (PMMA) plastic fragments have been found abundant in the environment, but the knowledge regarding its effects on the physiology of aquatic animals is still poorly studied. Here the short-term (96 h) effects of waterborne exposure to PMMA nanoplastics (PMMA-NPs) on the muscle of gilthead sea bream (Sparus aurata) fingerlings was evaluated at a concentration range that includes 0.001 up to 10 mg/L. The expression of key transcripts related to cell stress, tissue repair, immune response, antioxidant status and muscle development, together with several biochemical endpoints and metabolic parameters. Results indicate that exposure to PMMA-NPs elicit mildly antioxidant responses, enhanced the acetylcholinesterase (AChE) activity, and inhibited key regulators of muscle development (growth hormone receptors ghr-1/ghr-2 and myostatin, mstn-1 transcripts). However, no effects on pro-inflammatory cytokines (interleukin 1ß, il1ß and tumor necrosis factor α, tnfα) expression nor on the levels of energetic substrates (glucose, triglycerides and cholesterol) were found.

Cytotoxicity and cell response of preosteoblast in calcium sulfate-augmented PMMA bone cement.

Poly(methyl methacrylate) (PMMA) has been widely used in orthopedic applications, but bone ingrowth and toxic monomer release are drawback of this material. Particle reinforcement with osteoconductive substitute, such as calcium sulfate (CaSO4), is one of the solutions used to modify PMMA bone cement. The current study investigated the mechanical, chemical and biological properties of CaSO4-augmented bone cement. Mechanical strength was measured by a material testing machine. The concentration of methyl methacrylate (MMA) monomer from the various formulations of PMMA mixed with CaSO4was measured by ultra-performance liquid chromatography (UPLC). CCK-8 assay and ALP assay were performed to evaluate cytotoxicity of released MMA monomer and cell differentiation. The attachment of cells to CaSO4-augmented bone cement discs was observed by confocal and scanning electron microscopy, and surface topography was also evaluated by atomic force microscopy. The results revealed that increased CaSO4weight ratios led to compromised mechanical strength and increased MMA monomer release. Cell density and cell differentiation on CaSO4-augmented bone cement discs were decreased at CaSO4weight ratios above 10%. In addition, the presence of micropores on the surface and surface roughness were both increased for PMMA composite discs containing higher levels of CaSO4. These results demonstrated that fewer MC3T3-E1 cells on the surface of CaSO4-PMMA composites was correlated to increased MMA monomer release, micropore number and surface roughness. In summary, the augmentation of a higher proportion of CaSO4(>10 wt. %) to PMMA did not promote the biological properties of traditional PMMA bone cement.

Is the toxicity of nanosized polymethylmethacrylate particles dependent on the exposure route and food items?

The environmental effects of nanoplastics-NPLs have been addressed mainly through short-term exposures to a few types of polymers, neglecting other NPLs that are economically relevant like polymethylmethacrylate - PMMA. This work aimed to assess long-term effects of PMMA-NPLs on the marine primary consumer Brachionus plicatilis, evaluating the influence of different exposure routes (waterborne, foodborne and both) and food items (Nannochloropsis gaditana and Tetraselmis chuii). Rotifers were 21 days exposed to: a) control, with clean medium and food-CTR; b) contaminated medium (8.1 mg PMMA-NPLs/L) and clean algae-MC; c) clean medium and contaminated algae (pre-incubated for 96 h on 8.1 mg PMMA-NPLs/L)-AC; and, d) contaminated medium and algae-MC/AC. Mortality (lx), total number of organisms (TN), fecundity (mx), populational growth rate (r), generational time (gt), and feeding rates were assessed. Effects on r and mx were found after 21 days. Organisms from AC had higher r than MC. MC/AC organisms performed better than control in all endpoints. Overall organisms fed with N. gaditana had higher TN, mx and r than those fed with T. chuii. In the AC treatments, rotifers fed with N. gaditana had higher mx. Results highlight that exposure route and food type may modulate NPLs' effects, supporting the need for standardization of assays.

Assessment of Flexural Strength and Cytotoxicity of Heat Cure Denture Base Resin Modified with Titanium Dioxide Nanoparticles: An In Vitro Study.

AIMS: To assess the effect of titanium dioxide nanoparticles (NPs) on flexural strength and cytotoxicity of heat-cured polymethyl methacrylate (PMMA) resins. METHODOLOGY: Sixty-four rectangular and 12 circular specimens were fabricated from metal dies to test flexural strength and cytotoxicity, respectively. The rectangular specimens were grouped into four (16 specimens each)-control group (Group 1), Group 2 with 3% TiO2, Group 3 with 5% TiO2, and Group 4 with 7% TiO2. They were tested for flexural strength using universal testing machine. The circular specimens were grouped into two (six specimens each)-control group and the test group which included the group which showed a highest flexural strength. They were tested for cytotoxicity using MTT assay. The analysis of variance (ANOVA) test was used to analyze the mean flexural strength of each group and Tukey's post hoc test, for pairwise group comparison (p <0.05). An independent sample t-test was used to analyze the cytotoxicity between the groups (p <0.05). RESULT: The study showed that there was a significant decrease in the flexural strength from the control group (mean: 298.95), which reduced as the concentration of TiO2 increased. However, the toxicity reduced considerably from 24 hours to 7 days in both groups while the test group showed better cell viability (%) than the control group. CONCLUSION: The flexural strength of heat cure acrylic resin (modified and unmodified) was much higher than the recommended flexural strength for these resins. On adding TiO2 NPs, flexural strength decreased when compared to the control group. However, with 3% TiO2 NPs, there was no significant decrease in flexural strength as compared to conventional resins. TiO2 NP-modified heat cure acrylic resin showed less toxicity on day 1 and even lesser toxicity after 7 days indicating that it is biocompatible. CLINICAL SIGNIFICANCE: TiO2 NPs incorporated at 3% concentration in denture base resin had less cytotoxicity and adequate flexural strength, to be used as a promising alternative to conventional denture base resin.

Waterborne exposure of gilthead seabream (Sparus aurata) to polymethylmethacrylate nanoplastics causes effects at cellular and molecular levels.

This study evaluated the effect of a short-term exposure to 45 nm polymethylmethacrylate nanoplastics (PMMA-NPs) on the gilthead seabream (Sparus aurata), by assessing biomarkers at different levels of biological organization in liver and plasma. Fish were exposed via water to PMMA-NPs (0, 0.001, 0.01, 0.1, 1 and 10 mg L-1) and sampled after 24 and 96 h. Results showed a general up-regulation of mRNA levels of key genes associated with lipid metabolism (e.g. apolipoprotein A1 and retinoid X receptor). Together with the modulation of the lipid pathway genes we also found a global increase in cholesterol and triglycerides in plasma. Antioxidant-related genes (e.g. glutathione peroxidase 1) were also up-regulated after 24 h of exposure, but their expression levels returned to control afterwards. Total antioxidant capacity (TAC) was increased throughout the experiment, however at 96 h the antioxidant capacity became less efficient, reflected by an increase in the total oxidative status (TOS). Concomitantly, we found an increase in the erythrocytic nuclear abnormalities (ENAs) throughout the trial. Altogether, PMMA-NPs activated the organism's antioxidant defenses and induced alterations in lipid metabolism pathways and genotoxicity in the blood cells of gilthead seabream.

Polymethylmethacrylate nanoplastics effects on the freshwater cnidarian Hydra viridissima.

The current understanding of nanoplastics (NPLs) toxicity to freshwater biota, especially the potential toxic effects of polymethylmethacrylate (PMMA), remains limited. Thus, the present work intended to add knowledge about the ecotoxicity of ∼40 nm PMMA-NPLs focusing on lethality, morphology, feeding and regeneration capacity of the freshwater cnidarian Hydra viridissima, after an exposure period of 96 h. Results showed that high concentrations of PMMA-NPLs can impair the survival of H. viridissima, with an estimated 96 h-LC50 of 84.0 mg PMMA-NPLs/L. Several morphological alterations were detected at concentrations below 40 PMMA-NPLs mg/L, namely partial or total loss of tentacles, which, however, did not induce significant alterations on the feeding rates. Morphological alterations not previously reported in the literature were also found after the 96 h exposure, such as double or elbow-like tentacles. Exposure to 40 mg PMMA-NPLs/L significantly impacted hydra regeneration, with organisms exposed to PMMA-NPLs presenting significant slower regeneration rates comparatively to controls, but with no impacts on the feeding rates. Overall, this work highlights the need to assess the effects of NPLs in freshwater biota. Hydra viridissima species was sensitive in a wide range of endpoints showing its value as a biological model to study the effects of small plastic particles.