Evaluation of tire tread particle toxicity to fish using rainbow trout cell lines.

Tire and road wear particles (TRWP) resulting from tire abrasion while driving raise concerns due to their potential contribution to aquatic toxicity. Our study aimed to assess cryogenically milled tire tread (CMTT) particle toxicity, used as a proxy for TRWP, and associated chemicals to fish using two Rainbow Trout (Oncorhynchus mykiss) cell lines representing the gill (RTgill-W1) and the intestinal (RTgutGC) epithelium. CMTT toxicity was evaluated through several exposure pathways, including direct contact, leaching, and digestion, while also assessing the impact of particle aging. Following OECD TG249, cell viability was assessed after 24 h acute exposure using a multiple-endpoint assay indicative of cell metabolic activity, membrane integrity and lysosome integrity. In vitro EC50 values for the fish cell lines exceeded river TRWP concentrations (2.02 g/L and 4.65 g/L for RTgill-W1 and RTgutGC cell lines, respectively), and were similar to in vivo LC50 values estimated at 6 g/L. Although toxicity was mainly driven by the leaching of tire-associated chemicals, the presence of the particles contributed to the overall toxicity by inducing a continuous leaching, highlighting the importance of considering combined exposure scenarios. Aging and digestion conditions were also found to mediate CMTT toxicity. Thermooxidation resulted in a decreased chemical leaching and toxicity, while in vitro digestion under mimicked gastrointestinal conditions increased leaching and toxicity. Specific chemicals, especially Zn, 2-mercaptobenzothiazole, 1,3-diphenylguanidine, and N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine (6PPD) were identified as contributors to the overall toxicity. Although 6PPD-quinone was detected in CMTT digestate, cytotoxicity assays with RTgill-W1 and RTgutGC cell lines showed no toxicity up to 6 mg/L, supporting the notion of a specific mode of action of this chemical. This study provides insights into the toxicological mechanisms induced by tire particles and their associated chemicals and can help in the evaluation of potential risks to aquatic life associated with TRWP.

Estrogenic, Genotoxic, and Antibacterial Effects of Chemicals from Cryogenically Milled Tire Tread.

Tire and road wear particles (TRWP) contain complex mixtures of chemicals and release them to the environment, and potential toxic effects of these chemicals still need to be characterized. We used a standardized surrogate for TRWP, cryogenically milled tire tread (CMTT), to isolate and evaluate effects of tire-associated chemicals. We examined organic chemical mixtures extracted and leached from CMTT for the toxicity endpoints genotoxicity, estrogenicity, and inhibition of bacterial luminescence. The bioassays were performed after chromatographic separation on high-performance thin-layer chromatography (HPTLC) plates. Extracts of CMTT were active in all three HPTLC bioassays with two estrogenic zones, two genotoxic zones, and two zones inhibiting bacterial luminescence. Extracts of CMTT artificially aged with thermooxidation were equally bioactive in each HPTLC bioassay. Two types of aqueous leachates of unaged CMTT, simulating either digestion by fish or contact with sediment and water, contained estrogenic chemicals and inhibitors of bacterial luminescence with similar profiles to those of CMTT extracts. Of 11 tested tire-associated chemicals, two were estrogenic, three were genotoxic, and several inhibited bacterial luminescence. 1,3-Diphenylguanidine, transformation products of N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine, and benzothiazoles were especially implicated through comparison to HPTLC retention factors in the CMTT samples. Other bioactive bands in CMTT samples did not correspond to any target chemicals. Tire particles clearly contain and can leach complex mixtures of toxic chemicals to the environment. Although some known chemicals contribute to estrogenic, genotoxic, and antibacterial hazards, unidentified toxic chemicals are still present and deserve further investigation. Overall, our study expands the understanding of potential adverse effects from tire particles and helps improve the link between those effects and the responsible chemicals. Environ Toxicol Chem 2024;43:1962-1972. © 2024 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Moving targets, long-lived infrastructure, and increasing needs for integration and adaptation in water management: an illustration from Switzerland.

Switzerland provides an example of successful management of water infrastructure and water resources that was accomplished largely without integration across sectors. Limitations in this approach have become apparent; decisions that were formerly based only on technical and economic feasibility must now incorporate broader objectives such as ecological impact. In addition, current and emerging challenges relate to increasingly complex problems that are likely to demand more integrated approaches. If such integration is to be of benefit, it must be possible to redirect resources across sectors, and the synergies derived from integration must outweigh the additional cost of increased complexity.

Mass fluxes and spatial trends of xenobiotics in the waters of the city of Halle, Germany.

The behaviour and the effects of xenobiotics including pharmaceuticals and fragrances in the environment are widely unknown. In order to improve our knowledge, field investigations and modelling approaches for the entire area of the city of Halle/Saale, Germany, were performed. The distribution of the concentration values and mass fluxes are exemplified using indicators such as Bisphenol A, t-Nonylphenol, Carbamacepine, Galaxolide, Tonalide, Gadolinium and isotopes. Concentrations at a magnitude of ng/L to microg/L were found ubiquitously in the ground and surface waters. Using the concentration values, the impact of the city concerning the indicators was not always evident. Only the assessment of the mass fluxes shows significant urban impacts along the city passage. The calculation of the mass fluxes shows increasing values for all investigated xenobiotics during the city passage; only Bisphenol A stagnates. A balance model of water and indicator mass fluxes was built up for the entire city area.

Who is chasing whom? A call for a more integrated approach to reduce the load of micro-pollutants in the environment.

One of the key questions arising from the presence of micro-pollutants in surface-, ground-, and drinking water is whether they pose a risk to human and ecosystem health. In our laboratories we have identified a number of biological effects by several pharmaceuticals and personal care products (PPCPs) on human, animal and/or plant cells at different levels of biological organisation. In part, these effects occur at concentrations even below those reported in drinking water. Even though it is often still difficult to fully deduce the role of some of these effects on the whole organism or population level as well as after chronic exposure, the effects observed illustrate that the input of micro-pollutants into the environment must be avoided or as far as possible reduced. Much effort has already been devoted to improved treatment of sewage and raw drinking water. A comprehensive protection from aquatic micro-pollutants, however, cannot reside in water treatment technology alone. Instead, all components of the life cycle of these chemicals must be put to the table to turn around the current trend of increasing environmental loads. The goal of this report is to illustrate why a more comprehensive way of risk assessment is needed and what this should include.

Conductive composite fibres from reduced graphene oxide and polypyrrole nanoparticles.

Continuous composite fibres composed of polypyrrole (PPy) nanoparticles and reduced graphene oxide (rGO) at different mass ratios were fabricated using a single step wet-spinning approach. The electrical conductivity of the composite fibres increased significantly with the addition of rGO. The mechanical properties of the composite fibres also improved by the addition of rGO sheets compared to fibres containing only PPy. The ultimate tensile strength of the fibres increased with the proportion of rGO mass present. The elongation at break was greatest for the composite fibre containing equal mass ratios of PPy nanoparticles and rGO sheets. L929 fibroblasts seeded onto fibres showed no reduction in cell viability. To further assess toxicity, cells were exposed to media that had been used to extract any aqueous-soluble leachates from developed fibre. Overall, these composite fibres show promising mechanical and electrical properties while not significantly impeding cell growth, opening up a wide range of potential applications including nerve and muscle regeneration studies.

Biophysical and biochemical properties of a binary lipid mixture for DNA transfection.

The phase and miscibility behavior of a triple-chain phosphatidylcholine (TPHPC) and a single-chain surfactant (CTAB) were investigated in aqueous dispersions and in monolayers at the air/water interface. CTAB can be incorporated in the TPHPC monolayer because of its complementary molecule shape and reduces the tilt angle of TPHPC. The type of phases and the phase sequence (L2 - LS) are the same in the pure TPHPC monolayer and in the TPHPC/CTAB (80:20 mol:mol) mixture. No indication of any ordering of adsorbed DNA was observed. In the aqueous dispersions, TPHPC exhibits an inverted hexagonal phase above the chain melting. The addition of 30 mol% CTAB leads to the appearance of a lamellar Lalpha phase. The binding of DNA to the mixture is obvious but this is accompanied by a separation of the two lipids what is supported by monolayer experiments. The system has no long-term stability. The main reason seems to be not only the stronger interaction of DNA with CTAB, but also especially the unexpected weak interaction between CTAB and TPHPC. The transfection efficiency is lower compared with lipofectamine. The main disadvantage of this system is the cytotoxicity of CTAB, which could not be lowered by incorporation of CTAB in the TPHPC bilayer.

Slit-lamp fundus microscopy without contact lens and with laser attachment.

Gullstrand's fundus microscopy is adapted for slit-lamp microscopy without a contact lens, equipped by a wide-aperture argon laser beam. The z axis magnification of the image is used to advantage for vitrectomy. The mobility of the device permits vitrectomy on a fixed eye.

Argon laser surgery of the iris, optimized by contact lenses.

Convex-surfaced contact lenses increase the power of the laser optically for surgery on the iris. Their power-increasing faculty becomes obvious by their power of image magnification. When linked by functions, lens power, laser power, and magnification increases. The laser power, which is multiplied by the lenses, contains a safety feature for the tissues outside the focus of the laser beam as the beam increases in angle. Safety and power is gained by the appropriate use of power-multiplying contact lenses for argon laser surgery.

Simplified photogrammetry of the optic disc.

Simplified photogrammetry of the optic disc depends on axis-correct fundus photography. The evaluation can be initiated by a time-based method of photogrammetry that proves presence or absence of change of the object. The degree of change can be simply measured by a telemetric device that permits one to evaluate the third dimension from stereophotographs.

Stereochronoscopy of the optic disc with stereoscopic cameras.

Goldmann's chronoscopy, a form of time-based photogrammetry of the optic disc, can be accomplished by repeated simultaneous stereophotography. The variations in centration and image orientation are nulled by azimuthal rotation and fusion of two pairs of stereophotographs taken at separate times.

Ability of 16 priority PAHs to be directly cytotoxic to a cell line from the rainbow trout gill.

Sixteen polycyclic aromatic hydrocarbons (PAHs) were screened for their ability to be directly cytotoxic to a cell line from the rainbow trout gill, RTgill-W1. Exposure times of 2 h or less were sufficient for direct cytotoxicity to be detected, which appeared to be caused by a common mechanism, the general perturbation of membranes. This was judged by the similarity of results obtained for three fluorescent indicator dyes, alamar Blue, 5-carboxyfluorescein diacetate acetoxymethyl ester (CFDA-AM) and neutral red. Among the 16 PAHs tested, just two- and three-ring PAHs were found to be directly cytotoxic. These were naphthalene approximately = acenaphthylene approximately = acenaphthene > fluorene approximately = phenanthrene. The results suggest that water solubility and lipophilicity are the critical properties determining the direct cytotoxicity of PAHs and that they do so by influencing PAH accumulation in membranes. Only naphthalene was effective at concentrations well below its water solubility limit. Therefore, direct cytotoxicity is likely to be most environmentally relevant only with naphthalene.

Ability of 16 priority PAHs to be photocytotoxic to a cell line from the rainbow trout gill.

Sixteen polycyclic aromatic hydrocarbons (PAHs) were screened for their ability to be photocytotoxic to a cell line from the rainbow trout gill, RTgill-W1. PAHs could be divided into one of three groups: incapable of being photocytotoxic, able to be both photocytotoxic and directly cytotoxic, or capable of being only photocytotoxic. Photocytotoxicity was distinct from direct cytotoxicity in that EC50 values were lower with the neutral red assay immediately after the PAH/UV treatment than with alamar Blue or CFDA-AM, indicating a more specific action on lysosomes. As well, in photocytotoxicity but not in direct cytotoxicity, the three assays showed increased impairment 24 h after treatment. Most PAHs were found to be strictly photocytotoxic; however, only six compounds were photocytotoxic at concentrations theoretically achievable in water. When photocytotoxic PAHs were ranked relative to fluoranthene to establish fluoranthene equivalent factors (FEFs), benzo[a]pyrene and benzo[g,h,i]perylene were found to be most potent. However, when the water solubility of each compound was taken into account in order to calculate the potential environmental photocytotoxic potency (PEPP), fluoranthene and pyrene appeared to have the most potential to impact fish through photocytotoxicity.

Methodology for demonstrating and measuring the photocytotoxicity of fluoranthene to fish cells in culture.

Methodology was developed for quantifying the photocytotoxicity of fluoranthene to a gill cell line from rainbow trout for future use in screening polycyclic aromatic hydrocarbons for their relative photocytotoxicity to fish. Solubilization in a modified culture medium was achieved with and without foetal bovine serum (FBS) and with and without dimethyl sulfoxide (DMSO). FBS caused most of the fluoranthene to remain in solution and blocked photocytotoxicity if present during UV irradiation. DMSO had little effect on fluoranthene distribution in cell cultures but caused cells to be slightly more sensitive to the phototoxicity of fluoranthene. The indicator dyes alamar Blue() and 5-carboxyfluorescein diacetate acetoxymethyl ester were used to quantify cytotoxicity in two different ways-singly in two separate assays, and mixed together in a novel single assay, which saved time and material. With UV irradiation for 2 hr at a photon fluence rate of either 1.4 mumol UV-B/m(2)/sec (UV-A:UV-B, 1.5) or 1.1 mumol UV-B/m(2)/sec (UV-A:UV-B, 9.7), both dyes indicated increasing loss of viability with increasing doses of fluoranthene. EC(50) values ranged from 18 to 44 ng/ml (89-217 nM), with the alamar Blue assay being slightly more sensitive.

Ability of fractionated petroleum refinery effluent to elicit cyto- and photocytotoxic responses and to induce 7-ethoxyresorufin-O-deethylase activity in fish cell lines.

The ability of fractionated petroleum refinery effluent to cause cellular responses in fish cell lines was evaluated. The cellular responses, which included direct and indirect cytotoxicity, photocytotoxicity and induction of 7-ethoxyresorufin-O-deethylase (EROD) activity, may potentially be linked to sublethal effects observed in effluent-exposed fish and fish larvae. In order to be able to quantify cellular responses rapidly, microtitre plates were used along with fluorescent probes. For the quantification of cyto- and photocytotoxicity, the fluorescent probes were alamar Blue and carboxyfluorescein diacetate acetoxymethyl ester (CFDA-AM), which monitor metabolic activity and cell membrane integrity, respectively. EROD activity was measured as the rate of conversion by EROD of the substrate 7-ethoxyresorufin to its fluorescent product, resorufin. Effluent from an Ontario refinery was fractionated into aqueous and particulate phase. As well, a solid phase extract (SPE) was used to prepare concentrated effluent for testing in the cell lines. The effluent was able to elicit all of the responses of interest although significant cyto- and photocytotoxicity required effluent equivalent concentrations above 100% effluent and could only be revealed upon exposure of cells to the SPE concentrated effluent. Based on their retention on C18, the cytotoxicants are likely to be non-polar to moderately polar chemicals. The presence of polar compounds affecting cellular metabolism was indicated by the responses of exposed cells to a 90% aqueous phase effluent. In contrast to cyto- and photocytotoxicity, EROD induction occurred at effluent equivalent concentrations well below 100% effluent and was elicited by the SPE and the particulate fraction thereby suggesting that most EROD-inducers were particle-bound. Among other applications, the described techniques could help to determine the source of causative agents of sublethal effects in the refining process.

Polycyclic aromatic hydrocarbons as inducers of cytochrome P4501A enzyme activity in the rainbow trout liver cell line, RTL-W1, and in primary cultures of rainbow trout hepatocytes.

In order to investigate cell-specific differences in the response of in vitro models to environmental toxicants, we compared the capacity of nine polycyclic aromatic hydrocarbons (PAHs) to induce cytochrome P4501A (CYPIA) in primary rainbow trout (Oncorhynchus mykiss) hepatocytes and a rainbow trout liver cell line, RTL-W1. Induction of CYPIA was estimated from the catalytic activity of 7-ethoxyresorufin-O-deethylase (EROD) and compared by median effective concentration (EC50) values, induction spans, and benzo[a]pyrene induction equivalency factors for inducing PAHs. The influence of culture conditions was investigated with respect to the presence or absence of serum and varying exposure times. Both in vitro systems lead to an identical classification of the PAHs in noninducing (anthracene, fluoranthene, phenanthrene, and pyrene) and inducing compounds with a similar ranking of inducing PAHs. Mean EC50 values in RTL-W1 cells were, respectively, 343 and 266 nM for benzo[a]anthracene, 57 and 92 nM for BaP, 134 and 283 nM for benzo[b]fluoranthene, 455 and 270 nM for chrysene, and 98 and 116 nM for 3-methylcholanthrene. Compared to primary hepatocytes, the RTL-W1 cell line was more sensitive in its EROD response to the presence or absence of serum and to the increase in exposure time, which led to higher EC50 values.

Transient induction of 7-ethoxyresorufin-O-deethylase (EROD) activity by medium change in the rainbow trout liver cell line, RTL-W1.

Cytochrome P4501A (CYP1A) metabolizes a wide array of lipophilic xenobiotics. In fish liver, CYP1A is constitutively expressed at low levels, but xenobiotics can strongly induce CYP1A expression via a receptor-mediated pathway. While induction of hepatic CYP1A in teleosts by xenobiotics is well investigated, very little is known on the regulation of constitutive CYP1A expression and its induction by factors other than xenobiotics. In the present study we show that in the rainbow trout liver cell line, RTL-W1, CYP1A-catalyzed 7-ethoxyresorufin-O-deethylase (EROD) activity can be induced by a change of the culture medium, in the absence of xenobiotics. The increase in cellular EROD levels is of transient nature. Experiments with cell incubation solutions supplemented with various medium components indicate that photooxidized tryptophan is the agent causing the increase of EROD activity after medium change.

Moisture in the granules.

This article offers manufacturers who process a broad range of plastics tips on how to handle polymers that are sensitive to moisture. It covers detection and drying procedures that will help ensure good-quality mouldings are produced.

Effects of clofibric acid alone and in combination with 17ß-estradiol on mRNA abundance in primary hepatocytes isolated from rainbow trout.

Clofibric acid (CA) is the active substance of lipid lowering drugs. It is resistant to degradation, polar in nature, and has been found ubiquitously in the aquatic environment. Though CA is classified as a peroxisomal proliferator in rodents and is considered as a potential endocrine disruptor, little information exists on the effects of CA in aquatic organisms, such as fish. In the present study, we examined the mRNA levels of peroxisome proliferator- and estrogen-sensitive genes on the exposure of primary rainbow trout (Oncorhynchus mykiss) hepatocytes to CA alone and in combination with the natural female sex hormone, 17ß-estradiol (E2). Our results demonstrate that rainbow trout hepatocytes are relatively refractory to the effects of CA on the PPAR signaling pathway and lipid metabolism. Moreover, CA did not show recognizable estrogenic activity, but after the induction of vitellogenesis by E2, CA significantly reduced vitellogenin (VTG) mRNA abundance. Apparently, the indirect repression of VTG transcription, independent of estrogen receptors, occurred. The mechanism is not yet clearly understood but may involve disruption of the stabilization of VTG mRNA known to be induced by E2.

Engineering a multimodal nerve conduit for repair of injured peripheral nerve.

Injury to nerve tissue in the peripheral nervous system (PNS) results in long-term impairment of limb function, dysaesthesia and pain, often with associated psychological effects. Whilst minor injuries can be left to regenerate without intervention and short gaps up to 2 cm can be sutured, larger or more severe injuries commonly require autogenous nerve grafts harvested from elsewhere in the body (usually sensory nerves). Functional recovery is often suboptimal and associated with loss of sensation from the tissue innervated by the harvested nerve. The challenges that persist with nerve repair have resulted in development of nerve guides or conduits from non-neural biological tissues and various polymers to improve the prognosis for the repair of damaged nerves in the PNS. This study describes the design and fabrication of a multimodal controlled pore size nerve regeneration conduit using polylactic acid (PLA) and (PLA):poly(lactic-co-glycolic) acid (PLGA) fibers within a neurotrophin-enriched alginate hydrogel. The nerve repair conduit design consists of two types of PLGA fibers selected specifically for promotion of axonal outgrowth and Schwann cell growth (75:25 for axons; 85:15 for Schwann cells). These aligned fibers are contained within the lumen of a knitted PLA sheath coated with electrospun PLA nanofibers to control pore size. The PLGA guidance fibers within the nerve repair conduit lumen are supported within an alginate hydrogel impregnated with neurotrophic factors (NT-3 or BDNF with LIF, SMDF and MGF-1) to provide neuroprotection, stimulation of axonal growth and Schwann cell migration. The conduit was used to promote repair of transected sciatic nerve in rats over a period of 4 weeks. Over this period, it was observed that over-grooming and self-mutilation (autotomy) of the limb implanted with the conduit was significantly reduced in rats implanted with the full-configuration conduit compared to rats implanted with conduits containing only an alginate hydrogel. This indicates return of some feeling to the limb via the fully-configured conduit. Immunohistochemical analysis of the implanted conduits removed from the rats after the four-week implantation period confirmed the presence of myelinated axons within the conduit and distal to the site of implantation, further supporting that the conduit promoted nerve repair over this period of time. This study describes the design considerations and fabrication of a novel multicomponent, multimodal bio-engineered synthetic conduit for peripheral nerve repair.