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Mumps virus (MuV) is highly neurotropic and neurovirulent, hence, the neurovirulence of virus seeds used in the production of mumps vaccines must be tested. The previous neurovirulence evaluation method involves measuring the area of the cavity in the Lewis neonatal rat brain caused by MuV through paraffin sectioning and hematoxylin-eosin (HE) staining. However, the processes of paraffin sectioning and HE staining are time consuming and complicated. To solve this problem, in this study, a vibratome sectioning system was first deployed to evaluate MuV neurovirulence in the rat brain instead of paraffin sectioning and HE staining. The results showed that the vibratome sectioning method could assess the neurovirulence potential of MuV more objectively and efficiently. In addition, the effects of different MuV doses and the ages of the rats in days on this evaluation method were explored. The results indicate that MuV at no less than 10 50 % cell culture infective dose (CCID

Investigating the function of delicate mammalian eyes often requires chemical fixation, histological sectioning, immunohistochemistry (IHC) and

We performed a retrospective examination of spontaneous hepatocellular carcinomas (HCCs) (primary and metastatic tumors) in 14 captive prosimians brought to the Veterinary Medical Diagnostic Laboratory in North Carolina State University over a period of 11 years (2003 to 2014) to characterize the tumors. These animals are endangered primates; a better understanding of the main fatal neoplasms is crucial. In addition to the histologic evaluation, an immunohistochemical study was also performed, using a hepatocyte marker (hepatocyte paraffin 1 [HepPar-1]) and 2 cholangiocyte markers (keratin 7 [K7] and keratin 19 [K19]), in an attempt to identify a specific profile for HCCs with metastatic behavior. Six of the 14 HCCs had pulmonary metastases. The most frequent histopathological findings were a trabecular pattern (14/14, 100%), presence of multinucleated cells (12/14, 85.7%), and foci of extramedullary hematopoiesis (9/14, 64.3%). The mitotic count was significantly higher in the metastatic HCCs ( P < .05). HepPar-1 was detected in all primary and metastatic HCCs, with a strong intensity of staining. Labeling for K7 and K19 was positive in 12 HCCs (85.7%) and 1 HCC (7.1%), respectively. Contrary to the less aggressive HCCs, most of the metastatic HCCs (5/6) expressed K7 in more than 15% of cells. The percentage of K7-positive neoplastic hepatocytes was significantly higher in metastatic HCCs. This study suggests that K7 might be a prognostically relevant marker in HCCs of captive prosimians.

Stomatitis is inflammation of the oral mucosa. Angiopoietin-like protein 4 (ANGPTL4) has pleiotropic functions both anti-inflammatory and pro-inflammatory properties. In the present study, we tested whether there is a correlation between increased ANGPTL4 expression and inflammation in stomatitis mice and the mechanisms involved.In this study, the oral mucosa of mice was burned with 90% phenol and intraperitoneal injection of 5-fluorouracil to establish the model of stomatitis mice. The pathological changes of stomatitis mice were observed by HE staining of paraffin section. The expressions of cytokines and ANGPTL4 were detected by fluorescence quantitative PCR, and the protein levels of ANGPTL4 were detected by western blot. Compared with control group, the oral mucosal structure of model mice was damaged. The expression of ANGPTL4 were significantly increased concomitantly with elevated production of anti-inflammatory cytokine (peroxisome proliferator-activated receptor alpha) and pro-inflammatory cytokines [nuclear transcription factor-kappa B, interleukin-6 (IL-6), IL-1β, and tumor necrosis factor-α] in mice with stomatitis.This study suggests that ANGPTL4 may be a double-edged sword in multiple inflammatory responses in stomatitis mice.

Increasing global and domestic food trade and required logistics create uncertainties in food safety inspection due to uncertainties in food origins and extensive trade activities. Modern blockchain techniques have been developed to inform consumers of food origins but do not provide food safety information in many cases. A novel food safety tracking and modeling framework for quantifying toxic chemical levels in the food and the food origins was developed. By integrating chemicals' multimedia environment exchange, food web, and source tracking systems, the framework was implemented to identify short-chain chlorinated paraffin (SCCP) contamination of fresh hairtail fish sold by a Walmart supermarket in Xi'an, northwestern China, and sourced in Eastern China Sea coastal waters. The framework was shown to successfully predict SCCP level with a mean of 17.8 ng g

ObjectiveTo evaluate the safety and biocompatibility of bone wax as an implant material for medialisation laryngoplasty in a large animal model.MethodsThree Dorper-cross ewes underwent type I thyroplasty of the right vocal fold with bone wax. The animals were monitored for four weeks for general wellbeing. The animals were euthanised and the larynges harvested. Histological evaluation was performed to assess for adverse tissue reaction and biocompatibility.ResultsThe mean (± standard deviation) amount of bone wax implanted was 0.49 g (± 0.12 g). No adverse events were reported. Ex vivo vibration was present on high-speed imaging for all medialised vocal folds. Histology demonstrated implanted paraffin embedded within the thyroarytenoid muscle with no evidence of resorption, a minimal inflammatory infiltrate, and a thin fibrotic capsule.ConclusionThe results of this investigation suggest that bone wax may be a safe and efficacious implant material for medialisation laryngoplasty. Further studies are necessary to assess its long-term safety and efficacy.

Context Acanthamoeba is increasingly implicated in causing keratitis in patients wearing contact lens or ocular trauma and has a poor prognosis. Establishment of an animal model is critical to study the disease pathology, pathogenesis and to evaluate anti-amoebic drugs. Some studies have used contact lenses to establish Acanthamoeba keratitis (AK) in a mouse model, which is expensive and not very successful as lenses get dislodged. Objective To assess the feasibility of using parafilm (Bemis Company Inc., USA) as an alternative to contact lens for the establishment of AK in the mouse model. Methods Thirty-six Balb/c mice in three groups of six mice each for two strains of Acanthamoeba were used to induce AK. Three experimental approaches used were; i) Acanthamoeba impregnated contact lens, ii) Acanthamoeba impregnated parafilm and iii) scratching followed by inoculation of Acanthamoeba suspension. In all three models, tarsorrhaphy was performed. Infection was evaluated by clinical examination and also through microscopic examination of corneal scrapings and corneal sections. Results AK model was successfully established with parafilm whereas only one mouse developed AK with the use of contact lens and none with scratching and Acanthamoeba inoculation. Conclusion The use of parafilm is convenient, reliable and cheaper and can be considered an alternative to contact lenses to induce AK in a mouse model.

Amantadine is known to have a neuroprotective effect in many neurological diseases. This study aims at investigating the neuroprotective effect of amantadine in rats exposed to carbon monoxide (CO) poisoning.Rats were maintained under standard experimental laboratory conditions and randomized into 4 different groups of 7 each namely control, amantadine only, CO exposure, and amantadine + CO exposure. For immunohistochemical analysis, tissues taken from the prefrontal and hippocampal regions were taken into formalin and kept for at least one day. Afterward, the tissue was followed and blocked for paraffin blocking. N-Methyl D-Aspartate (NMDA) levels in homogenates were studied by the Enzyme-Linked Immunosorbent Assay (ELISA) method. Superoxide dismutase (SOD) and catalase (CAT) activities in the supernatants were studied with commercial kits. Nitric oxide (NO) and Asymmetric Dimethyl Arginine (ADMA) levels were studied by the ELISA method. Enzyme activity values were calculated by dividing the protein values in the supernatants and normalizing them.CAT, SOD, NMDA, ADMA, and NO levels were statistically significantly different between the groups (p0.05). According to post-hoc pairwise comparison test results, the values of the control and amantadine groups for CAT, SOD, NMDA, ADMA, and NO parameters were significantly higher than that of CO group. Similarly, values in the control and amantadine groups were considerably higher than values for the amantadine + CO group. NMDA values were significantly lower in group amantadine + CO than in CO group (p: 0.049).Apoptosis and endothelial damage after CO poisoning is a complex process, and amantadine administration has a limited contribution in preventing this process.

In this study, we aimed to investigate the effects of vitamin E on mouse adrenal glands in immobilization stress. Twenty-eight male, 10-week-old, BALB/C mice weighing 30-45 grams were divided into four groups. Mice were placed in a cage where no movement was allowed 6 hours/day for 7 days for immobilization stress. 10 ml/kg vitamin E was administered orogastrically 1 hour before immobilization stress in the vitamin E and stress+vitamin E group. At the end of the 7th day, all the animals were subjected to elevated-plus maze (anxiety) and forced swimming (depression) tests. Left adrenal glands were dissected for routine paraffin tissue embedding protocol. Adrenal sections were stained with hematoxylin-eosin and Azan. Malonaldehyde (MDA) levels were also measured in the adrenal tissues. Anxiety level (0.023), depression level (p=0.042) and MDA values (p=0.01) were significantly increased in the stress group. Histological sections of the stress group showed cortical atrophy, medullary hypertrophy, vascular dilation and hemorrhage. Azan staining revealed a thinned capsule and corticomedullary fibrosis in the stress group. Pathologies induced by immobilization stress were mostly reversed after vitamin E administration. The results suggested that vitamin E alleviates adverse effects of immobilization stress (oxidative, behavioral and histopathologic changes) in mice.

Reptarenaviruses cause boid inclusion body disease (BIBD), a poten- tially fatal disease, occurring in captive constrictor snakes boas and pythons worldwide. Classical BIBD, characterized by the formation of pathognomonic cyto- plasmic inclusion bodies (IBs), occurs mainly in boas, whereas in pythons, for example, reptarenavirus infection most often manifests as central nervous system signs with limited IB formation. The natural hosts of reptarenaviruses are unknown, although free-ranging/wild constrictor snakes are among the suspects. Here, we report BIBD with reptarenavirus infection in indigenous captive and wild boid snakes in Costa Rica using histology, immunohistology, transmission electron microscopy, and next-generation sequencing (NGS). The snakes studied represented diagnostic post- mortem cases of captive and wild-caught snakes since 1989. The results from NGS on archival paraffin blocks confirm that reptarenaviruses were already present in wild boa constrictors in Costa Rica in the 1980s. Continuous sequences that were de novo assembled from the low-quality RNA obtained from paraffin-embedded tissue allowed the identification of a distinct pair of reptarenavirus S and L segments in all studied animals; in most cases, reference assembly could recover almost complete segments. Sampling of three prospective cases in 2018 allowed an examination of fresh blood or tissues and resulted in the identification of additional reptarenavirus segments and hartmanivirus coinfection. Our results show that BIBD is not only a disease of captive snakes but also occurs in indigenous wild constrictor snakes in Costa Rica, suggesting boa constrictors to play a role in natural reptarenavirus circulation. Los reptarenavirus causan la enfermedad del cuerpo de inclusión de las boas (BIBD), una enfermedad potencialmente mortal que se produce en las serpientes constrictoras cautivas, boas y pitones, de las serpientes constrictoras en cautividad, las boas y las pitones, en todo el mundo. La BIBD clásica, caracterizada por la formación de cuerpos de inclusión cito de inclusión patognomónicos, se da principalmente en las boas, mientras que en las pitones, por ejemplo, la infección por reptarenavirus se manifiesta más a menudo como con una formación limitada de IB. Los huéspedes naturales de los reptarenavirus son desconocidos, aunque las serpientes constrictoras salvajes y en libertad están entre los sospechosos. En este caso, informamos de la existencia de de reptarenavirus en serpientes constrictoras autóctonas y silvestres de Costa Rica. Costa Rica utilizando histología, inmunohistología, microscopía electrónica de transmisión y secuenciación de próxima generación (NGS). Las serpientes estudiadas representaban casos de diagnóstico post de diagnóstico de serpientes cautivas y silvestres desde 1989. Los resultados de la NGS en de parafina de archivo confirman que los reptarenavirus ya estaban presentes en las boas en Costa Rica en la década de 1980. Las secuencias continuas que se ensamblaron de novo a partir del ARN de baja calidad obtenido de los tejidos embebidos en parafina, permitieron identificar un par de segmentos S y L del reptarenavirus en todos los animales estudiados. En la mayoría de los casos, el ensamblaje de referencia pudo recuperar segmentos casi completos. El muestreo de tres casos prospectivos en 2018 permitió un examen de sangre o tejidos y dio lugar a la identificación de segmentos adicionales de reptarenavirus y coinfección por hartmanivirus. Nuestros resultados muestran que la BIBD no es solo una enfermedad de las serpientes cautivas sino que también se da en las serpientes constrictoras silvestres de Costa Rica, lo que sugiere que las boas constrictoras desempeñan un papel en la circulación natural de reptarenavirus. Universidad Nacional, Costa Rica Escuela de Medicina Veterinaria

Chlorinated paraffins (CPs) are environmental pollutants of emerging concern. Long-chain CPs (LCCPs) are considered of lesser concern than other CPs in food due to their lower accumulation in most organisms. However, LCCPs have been shown to accumulate preferentially in birds. We used ultrahigh-performance liquid chromatography combined with electrospray ionization Orbitrap mass spectrometry (UPLC-ESI-Orbitrap MS) to analyze CPs (C10-26Cl4-12) in tissues of free-range hens, their feed, and local soils. Feed was found to be the main source of CP intake. The CP carbon chain length had little impact on their absorption. C18-CPs were excreted in preference to C13-CPs by laying. The metabolic elimination rates of CPs (0.2 μg/mL) estimated using chicken liver microsomes were in the order C12Cl6 (91%) > C12Cl8 (57%) > C18Cl6 (12%) > C18Cl8 (6%). CPs with longer carbon chains accumulated preferentially in muscle and adipose tissues, and the accumulation of specific carbon chain lengths was related to the content and composition of different CPs in the intake source.

Humans are exposed to

This study reports on the occurrence and distribution of short- and medium-chain chlorinated paraffins (SCCPs and MCCPs, respectively) in edible insects purchased from Asia and Europe. A total of 36 edible insect samples (n = 24 from Asia, n = 12 from Europe) authorized and prepared for human consumption were purchased and analyzed for SCCPs and MCCPs via gas chromatography and mass spectrometry. SCCPs were detected in 83% of all edible insect samples with an overall median ∑SCCP concentration of 8.7 ng/g dry weight (dw) and a range of

Background:The unique anatomy of the male reproductive organ reflects its complex function from sperm maturation to their storage for months until emission. Since light microscopy in two dimensions (2d) cannot sufficiently demonstrate its complex morphology, a comprehensive visualization is required to identify pathologic alterations in its entire anatomical context.Objectives:Aim of this study was to use three-dimensional (3d) light sheet fluorescence microscopy (LSFM) to visualize entire murine testes in 3d, label-free and at subcellular resolution, and to assign local autofluorescence to testicular and deferent structures.Materials and methods:Murine testes were fixed with four different fixatives and subsequently cleared with benzoic acid/benzyl benzoate. Hereafter, complete murine testes were scanned with LSFM with different fluorescence filter sets and subsequently embedded in paraffin for further conventional planar histology.Results:Autofluorescence signals of the murine reproductive organ allowed the unambiguous identification of the testicular anatomy from the seminiferous tubules to the vas deferens with their specific stratification independent of the used fixative. Blood vessels were visualized from the pampiniform plexus to the small capillaries of single tubules. Moreover, due to the specific intrinsic fluorescence properties of the efferent ducts and the epididymis, luminal caliber, the epithelial stratification and retronuclear cytoplasmic inclusions gave a unique insight into the interface of both morphological structures. Subsequent 2d histology confirmed the identified morphological structures.Discussion:LSFM analysis of the murine reproductive organ allows due to its intrinsic fluorescence a simple, label-free 3d assessment of its entire duct morphology, the epithelial composition, and the associated blood supply in its anatomical relation.Conclusion:LSFM provides the technical basis for comprehensive analyses of pathologically altered murine testes in its entirety by depicting specific autofluorescence. Thereby it facilitates mouse studies of testicular disease or their drug-related alterations in more detail potentially for clinical translation assessing human testicular biopsies.

Paraffin histology is one of the most important and commonly used laboratory techniques enabling the study of the microscopic structure of animal and plant tissues. This technique uses paraffin wax, which in liquid form impregnates fixed and dehydrated tissues and allows the preparation of thin sections when solidified in blocks. This protocol on good practices in paraffin histology of

Urea constitutes a physiological and presumably well-regulated constituent of tear fluid. Its lacrimal concentration is significantly decreased in dry eye disease. Urea homeostasis within the tear fluid may also depend on the expression of urea transporters. The present study reports on the expression patterns of urea transporter A (UT-A) in the cells and tissues of the ocular surface and the lacrimal glands.UT-A immunohistochemistry was performed on 5 µm paraffin sections of paraformaldehyde-fixed human, porcine, and murine corneas, eyelids, and lacrimal glands (n = 5 each).UT-A immunostaining was largely comparable in all three species. UT-A signals were detected in the corneal epithelium and endothelium, in the conjunctival epithelium, in the acinar cells and excretory ducts of the lacrimal gland, Meibomian gland, and in the glands of Moll and Zeis. The Meibomian glands and the glands of Zeis exhibited a marked UT-A-positive staining in the basal cells of the alveolar epithelia and in the ductal epithelia.UT-A shows comparable expression patterns to UT-B (previous study) at the ocular surface and in the lacrimal glands, as determined by immunohistochemistry. The presence of both urea transporters in the lacrimal functional unit suggests that they are essential for the normal function of the lacrimal system and the integrity of the tear film. Potential alterations in urea transporter expression might be associated with the significant reduction of urea found in the tear fluid of dry eye patients. They may thus play an important role in the pathogenesis of dry eye disease.

Short-chain chlorinated paraffins (SCCPs) have attracted attention because of their toxicological potential in humans and wildlife at environmentally relevant doses. However, limited information is available regarding mechanistic differences across species in terms of the biological pathways that are impacted by SCCP exposure. Here, a concentration-dependent reduced human transcriptome (RHT) approach was conducted to evaluate 15 SCCPs in HepG2 cells and compared with our previous results using a reduced zebrafish transcriptome (RZT) approach in zebrafish embryos (ZFEs). Generally, SCCPs induced a broader suite of biological pathways in ZFEs than HepG2 cells, and all of the 15 SCCPs were more potent in HepG2 cells compared to ZFEs. Despite these general differences, the transcriptional potency of SCCPs in both model systems showed a significant linear relationship (p = 0.0017, r2 = 0.57), and the average ratios of transcriptional potency for each SCCP in RZT to that in RHT were ∼100,000. C10H14Cl8 was the most potent SCCP, while C10H17Cl5 was the least potent in both ZFEs and HepG2 cells. An adverse outcome pathway network-based analysis demonstrated model-specific responses, such as xenobiotic metabolism that may be mediated by different nuclear receptor-mediated pathways between HepG2 cells (e.g., CAR and AhR activation) and ZFEs (e.g., PXR activation). Moreover, induced transcriptional changes in ZFEs associated with pathways and molecular initiating events (e.g., activation of nicotinic acetylcholine receptor) suggest that SCCPs may disrupt neural development processes. The cross-model comparison of concentration-dependent transcriptomics represents a promising approach to assess and prioritize SCCPs.

This study aimed to assess different approaches for bone healing evaluation on histological images and to introduce a new automatic evaluation method based on segmentation with distinct thresholds. We evaluated the hyperbaric oxygen therapy (HBO) effects on bone repair in type 1 diabetes mellitus rats. Twelve animals were divided into four groups (n = 3): non-diabetic, non-diabetic + HBO, diabetic, and diabetic + HBO. Diabetes was induced by intravenous administration of streptozotocin (50 mg/kg). Bone defects were created in femurs and HBO was immediately started at one session/day. After 7 days, the animals were euthanized, femurs were removed, demineralized, and embedded in paraffin. Histological sections were stained with hematoxylin and eosin (HE) and Mallory's trichrome (MT), and evaluated using three approaches: (1) conventional histomorphometric analysis (HE images) using a 144-point grid to quantify the bone matrix; (2) a semi-automatic method based on bone matrix segmentation to assess the bone matrix percentage (MT images); and (3) automatic approach, with the creation of a plug-in for ImageJ software. The time required to perform the analysis in each method was measured and subjected to Bland-Altman statistical analysis. All three methods were satisfactory for measuring bone formation and were not statistically different. The automatic approach reduced the working time compared to visual grid and semi-automated method (p .01). Although histological evaluation of bone healing was performed successfully using all three methods, the novel automatic approach significantly shortened the time required for analysis and had high accuracy.

Maternal recognition of pregnancy in llamas would be related to a still-unknown signal secreted by the embryo to extend the corpus luteum lifespan. In order to study llamas reproductive physiology and early pregnancy aspects, research on embryos between 8 and 12 days post-mating is essential. However, there is a lack of information on llama embryo paraffin embedding, a procedure that exhibits difficulties, mostly for embryo manipulation. This work presents an accessible, easy and rapid protocol for embedding llama blastocysts. Uterine flushings from four superovulated llamas were performed to recover embryos 8 days post-mating. Each embryo was fixed in 1 ml of 10 % neutral buffered formalin for 24 h, at 4 °C, and transferred to 70 % ethanol. Later, they were submitted to a series of baths: 500 µl PBS with 5 % bovine foetal serum solution (PBS+BFS) for 1 min; zona pellucida permeabilization with 500 µl PBS with 0.2 % Triton X-100 for 20 min; 500 µl PBS+BFS for 1 min; dehydration on baths with ascendent grades of ethanol (70 %, 96 %, and 100 % twice) for 10 min each; and bath of butanol for 5 min. Finally, embryos were embedded in a pellet of paraffin and afterwards in a paraffin block. Complete sections from 8 of 13 grade I and II embryos were obtained. In conclusion, by applying this method, good quality sections with preserved embryo morphology suitable for histological, histochemical or immunohistochemical approaches would be possible to acquire.

Chlorinated paraffins (CPs) are industrial chemicals produced in large quantities. Short-chain CPs (SCCPs) were classified as persistent organic pollutants under the Stockholm Convention in 2017. Medium-chain CPs (MCCPs) became candidate persistent organic pollutants in 2021. CPs are now ubiquitously found in the environment. Honey bees can be exposed to CPs during foraging, and this exposure subsequently results in the contamination of honey and other bee products along with colony food production and storage. Here, SCCP and MCCP concentrations in honey collected from Chinese apiaries in 2015 and 2021 were determined. Total CP concentrations in honey from 2021 to 2015 were comparable, but the ratio of MCCPs/SCCPs was higher in 2021 than in 2015. SCCP and MCCP congener group profiles in all honey samples were similar and dominated by C

Cytotoxicity in freshwater fishes induced by industrial effluents and dyes is a global issue. Trypan blue dye has many applications in different sectors, including laboratories and industries. This study determines to detect the cytotoxic effects of trypan blue dye in vivo. The objective of this study was to estimate the sub-lethal effects of azodye in fish. Cirrhinus mrigala, a freshwater fish, was exposed to three different grading concentrations of dye 5 mg/L, 10 mg/L, and 20 mg/L in a glass aquarium. Significant (p 0.05) decrease in the weight of fish was observed as 0.728 ± 0.14 g and 2.232 ± 0.24 g, respectively, in the trial groups exposed to 10 and 20 mg/L of dye in a week. After exposure to trypan blue dye, fishes were dissected to remove liver and kidney tissues. Histopathological assessments determined hepatotoxicity and nephrotoxicity induced by trypan blue through the paraffin wax method. This dye induces mild alterations in the liver such as congestion, hemolysis, dilated sinusoids, ruptured hepatocytes, vacuolization, edema of hepatocytes, necrosis, degeneration, aggregation, and inflammation. This dye not only alters liver tissue, also induces an acute level of tissue alterations in the kidneys, such as degeneration of epithelial cells of renal tubules, shrinkage of the glomerulus, congestion, reduced lumen, degeneration of glomerulus, absence of space of bowmen, glomerulonephritis, necrosis in hematopoietic interstitial tissues and glomerulus, reduced lumen, vacuolar degeneration of renal tubules, increased per tubular space. The current study concludes that trypan blue dye released even in small amounts is found to be associated with a high incidence of cytotoxicity. Such tissue alterations in this species could be used as biomarkers for azo dyes.

Background Dermanyssus gallinae, or poultry red mite (PRM), is an important ectoparasite in laying hen, having a significant effect on animal welfare and potentially causing economic loss. Testing novel control compounds typically involves in vitro methodologies before in vivo assessments. Historically, in vitro methods have involved PRM feeding on hen blood through a membrane. The use of hen blood requires multiple procedures (bleeds) to provide sufficient material, and the use of a larger species (e.g. goose) could serve as a refinement in the use of animals in research. Methods The in vitro feeding device used was that which currently employs a Parafilm™ M membrane (Bartley et al.: Int J Parasitol. 45:819–830, 2015). Adult female PMR were used to investigate any differences in mite feeding, egg laying and mortality when fed goose or hen blood. Effects on these parameters when PRM were fed through either the Parafilm™ M membrane or the Baudruche membrane alone or through a combination of the membrane with an overlaid polyester mesh were tested using goose blood. Results Poultry red mites fed equally well on goose or hen blood through the Parafilm™ M membrane, and there were no significant differences in mortality of PRM fed with either blood type. A significant increase (t test: t = 3.467, df = 4, P = 0.03) in the number of eggs laid per fed mite was observed when goose blood was used. A 70% increase in PRM feeding was observed when the mites were fed on goose blood through a Baudruche membrane compared to when they were fed goose blood through the Parafilm™ M membrane. The addition of an overlaid polyester mesh did not improve feeding rates. A significant increase (analysis of variance: F(3, 20) = 3.193, P = 0.04) in PRM egg laying was observed in mites fed on goose blood through the Baudruche membrane compared to those fed goose blood through the Parafilm™ M membrane. A mean of 1.22 (standard error of the mean ± 0.04) eggs per fed mite was obtained using the Baudruche feeding device compared to only 0.87 (SEM ± 0.3) eggs per fed mite using the Parafilm™ M device when neither was combined with a polyester mesh overlay. Conclusion The in vitro feeding of adult female PRM can be readily facilitated through the use of goose blood in feeding devices with the Baudruche membrane. Graphical Abstract

Studies on the biomagnification of short- and medium-chain chlorinated paraffins (SCCPs and MCCPs) in terrestrial ecosystems and their bioamplification during metamorphosis in insects and amphibians are scarce. Therefore, this study sought to characterize the occurrence and trophic dynamics of SCCPs and MCCPs in an insect-dominated terrestrial food web in an e-waste recycling site in South China. Median ∑SCCPs and ∑MCCPs concentrations in the organisms ranged from 2200 to 34 000 ng/g lipid weight and from 990 to 19 000 ng/g lipid weight, respectively. The homologue profiles of CPs in the predators were distinct from those in insects, presenting more short chain-high chlorinated congeners (C

Food consumption has been identified as a major pathway for human exposure to short-chain chlorinated paraffins (SCCPs) and medium-chain chlorinated paraffins (MCCPs), but evaluations of SCCP and MCCP intake from major dietary sources are limited. We used the sixth Chinese Total Diet Study to perform a comprehensive investigation of SCCPs and MCCPs in cereals, vegetables, potatoes, legumes, eggs, milk, meats, and aquatic foods from nine southern provinces. The geographical distribution of CP concentrations showed higher levels in Jiangsu, Hubei, and Zhejiang provinces. The CP concentrations in most animal-origin foods were higher than those in foods of plant origin. The total estimated daily intakes (EDIs) of SCCPs and MCCPs, with average values of 7.0 × 102 and 4.7 × 102 ng kg-1 day-1, respectively, were mostly contributed by cereals, vegetables, and meats. Risk assessment indicated the EDIs of CPs posed no significant risk to residents in South China.

Microplastics are one of the main environmental pollutants in marine ecosystems, and their presence in seawater is a consequence of the widespread use of plastic materials in modern commodities. This wide usage of plastics includes the employment of microspheres in common personal care products, which end up being ultimately released into the aquatic compartment. Known ecotoxicological effects of microplastics favoured the search for technologically viable and environmentally safer alternatives, such as paraffin wax microparticles, whose ecotoxicological risks have not been entirely characterized. To address this gap, the present study exposed mussels (Mytilus sp.) for 96 h to three densities (5 mg/L, 20 mg/L and 80 mg/L) of four size ranges (100–300 μm, 300–500 μm, 400–850 μm, and 800–1200 μm) of paraffin wax particles. Toxicological endpoints were the activities of four enzymes involved in key cellular processes, including antioxidant defence (catalase (CAT), glutathione reductase (GRed) and glutathione peroxidase (GPx)) and phase II metabolism (glutathione S-transferases (GSTs)), as well as lipid content and fatty acid profiles of the digestive gland. Significant interactions between the presence/absence of food and paraffin particle density were found, as food sometimes worked as a confounding factor in the analysed biomarkers. Despite this uncertainty, some overall patterns emerged. In general, smaller paraffin wax particles (100–300 μm) caused little effects on the activity of the four enzymes tested, whereas larger particles (800–1200 μm) caused significant effects on almost all biomarkers. CAT activity was enhanced in animals exposed to larger paraffin particles, whilst GPx activity was depressed; GRed activity was not affected by the exposure to paraffin particles. The activity of GSTs was enhanced, but only in one tested condition. No effects were observed in terms of the total lipid content and fatty acids of exposed animals. Overall, data obtained in this work suggest that, at densities of paraffin wax particles comparable to the levels found in the environment for microplastic beads, no toxicity is expected to occur in the tested mussel species, by measuring the here-assessed toxicological endpoints., This study received financial support from CESAM (UID/AMB/50017/2019) and from FCT/MEC through national funds, and the co-funding by the ERDF, within the PT2020 Partnership Agreement and Compete 2020.

Short chain chlorinated paraffins (SCCPs) are emerging persistent organic pollutants of great concern due to their ubiquitous distribution in the environment. However, little information is available on the biotransformation of SCCPs in organisms. In this study, a chlorinated decane: 1, 2, 5, 5, 6, 9, 10-heptachlorodecanes (HeptaCDs) was subjected to in vitro metabolism by human and chicken liver microsomes at environmentally relevant concentration. Using ultra-performance liquid chromatography-Q-Exactive Orbitrap mass spectrometry, two metabolites: monohydroxylated hexachlorodecane (HO-HexCD) and monohydroxy heptachlorodecane (HO-HeptaCD) were detected in human liver microsomal assays, while only one metabolite (HO-HexCD) was identified in chicken liver microsomal assays. The formation of HO-HexCD was fitted to a Michaelis-Menten model for chicken liver microsomes with a V

Short-chain chlorinated paraffins (SCCPs), a class of ubiquitous pollutants, are considered to be embryotoxic and teratogenic. However, little is known regarding the bioactivity and mechanisms at environmentally relevant concentrations at the embryonic period. Here, a concentration-dependent reduced transcriptomic approach was used to evaluate the environmental dose ( 60%) (p = 0.038), and the chlorine content of SCCP congeners affected the bioactivity associated with genotoxic pathways. The concentration-dependent reduced transcriptomic approach significantly improved the understanding of the ecological risk of environmental contaminants at early life stages.

For three-dimensional observation of unstained bio-specimens using X-ray microscopy with computed tomography (CT), one main problem has been low contrast in X-ray absorption. Here we introduce paraffin-mediated contrast enhancement to visualize biopsy samples of mouse kidney using a laboratory-based X-tray microscope. Unlike conventional heavy-atom staining, paraffin-mediated contrast enhancement uses solid paraffin as a negative contrast medium to replace water in the sample. The medium replacement from water to paraffin effectively lowers the absorption of low-energy X-rays by the medium, which eventually enhances the absorption contrast between the medium and tissue. In this work, paraffin-mediated contrast enhancement with 8 keV laboratory X-rays was used to visualize cylindrical renal biopsies with diameters of about 0.5 mm. As a result, reconstructed CT images from 19.4 h of data collection achieved cellular-level resolutions in all directions, which provided 3D structures of renal corpuscles from a normal mouse and from a disease model mouse. These two structures with and without disease allowed a volumetric analysis showing substantial volume differences in glomerular subregions. Notably, this nondestructive method presents CT opacities reflecting elemental composition and density of unstained tissues, thereby allowing more unbiased interpretation on their biological structures.

Here we describe a multiplex chromogenic immunohistochemistry platform to stain and analyze two markers in paraffin tissue sections from mouse or human. The basis of the protocol is a series of stripping and re-probing steps with subsequent image analysis, which allows the user to perform multiplex imaging in a reliable and affordable manner. Here, we describe specific usage to assess the levels of PD-L1 in tumor-associated macrophages. We have used different antibodies and assessed this protocol for up to five consecutive antibodies per slide. For complete details on the use and execution of this protocol, please refer to Orgaz et al. (2020).

This study was performed to evaluate the presence of chlorinated paraffins (CPs) in complementary baby food from different European countries, to assess portion-based dietary intake of CPs, and to characterize possible risks to children arising from CPs uptake. In total, 86 baby food samples from 22 countries were grouped into eight food-type categories considering the WHO classification. In six out of eight sample categories, predominance of medium-chain CPs (MCCPs) concentrations over short-chain CPs (SCCPs) was observed and, contrary to expectations, samples that contained ingredients of animal origin (e.g., meat or fish) did not display elevated CPs levels. The median ∑

International audience; Beeswax is intended for use in the beekeeping sector but also in the agro-food, pharmaceutical or cosmetics sectors. The adulteration of beeswax is an emerging issue that was reported lately at several occasions in the scientific literature. This issue tends to become more frequent and global, but its exact extent is not accurately defined. The present study aims to assess the current situation in Belgium through a nationwide survey. Randomized beeswax samples originating from Belgian beekeepers (N = 98) and commercial suppliers (N = 9) were analysed with a Fourier transform infrared spectroscopy (FTIR) coupled with Attenuated Total Reflectance (ATR) accessory (FTIR-ATR spectroscopy) for adulteration. The survey revealed a frequency of 9.2% and 33.3% of adulteration in beekeepers beeswax samples (9 samples out of 98: 2 with paraffin and 7 with stearin/stearic acid) and commercial beeswax samples (3 samples out of 9: all adulterated with stearin/stearic acid), respectively. The analysed samples were adulterated with various percentages of paraffin (12 to 78.8%) and stearin/stearic acid (1.2 to 20.8%). This survey indicates that in the beekeepers samples, beeswax adulteration was more frequent in comb foundation and crude beeswax than in comb wax. With the example of this nationwide survey conducted in Belgium, this study shows the emergence of the issue and the urgent need for action to safeguard the health of both honey bees health and humans, in particular with the setting of a proper regulation legal framework and a specific routine analytical testing of commercial beeswax to ensure beeswax quality.

Porcine circovirus type 2 (PCV-2) is of great economic significance to porcine industry worldwide. PCV-2 variants and genotypes, alternating world over, are the etiological agent of several clinical syndromes such as porcine dermatitis and nephropathy syndrome (PDNS), post-weaning multi-systemic wasting syndrome (PMWS) and others in pigs. This study is reporting an atypical manifestation of PDNS in twelve grower pigs, 3- to - 4.5 months age and either sex, died of the disease, with predominant lesions of nephropathy and no obvious clinical lesions in skin. Necropsy examination of pigs showed lesions of petechial -to- ecchymotic hemorrhages in the kidneys and in the right auricular musculature of the hearts. Microscopic lesions in HE sections of the kidneys showed acute glomerulonephritis, interstitial nephritis, and vasculitis, but the skin morphology and architecture remained unaltered in contrast to the pathognomonic lesions of PDNS described in the literature. Other syndromic associations of PDNS in these cases included-perimyocarditis, interstitial pneumonia, depleted lymphoid tissues, tonsillitis, enteritis, and meningo-encephalitis. The lesional sites in duplicate paraffin tissue sections of kidneys, heart, lungs, spleen, lymph nodes, intestine, and brain demonstrated PCV-2 antigen in the cytoplasm of cells as highlighted by the intense immunolabeling on IHC staining. The PCV-2 positive organs reconfirmed by PCR, targeting ORF2 gene, which yielded 481bp size of products. The sequencing results of 481bp products on phylogenetic analysis showed 94% similarity with that of PCV-2 sequences in the database that grouped into PCV2d-2 genotype. The present report confirms, probably for the first time, the atypical PDNS cases due to PCV2d-2 genotype in naturally affected grower pigs of India.

As new persistent organic pollutants, short chain chlorinated paraffins (SCCPs) have recently received particular attention. However, knowledge on their bioaccumulation and trophic magnification in marine organisms from East China Sea (ECS) is still scare. In this study, we investigated the concentrations of SCCPs in seawater (n = 15) and marine organisms (n = 88) collected from ECS. The total concentrations of SCCPs (∑SCCPs) ranged from 12.5 to 242 ng/L in seawater and from 12.8 to 1819 ng/g wet weight (ww) in organisms. C10–11 SCCPs and Cl5–7 SCCPs were the predominant homologues in all samples, with the mean proportions of 70 ± 6.5% and 80 ± 7.8% in seawater, as well as 52–77% and 61–84% in marine organisms, respectively. The logarithm bioaccumulation factor (log BAF) values of ∑SCCPs were in the range of 2.04–3.79 in zooplankton, fish, shrimp, crab, shellfish, snail, and cephalopod. The log BAF values of SCCP homologues (1.33–4.75) increased significantly with the increase of their logarithm octanol–water partition coefficients (log KOW) values, indicating that hydrophobicity is the major factor controlling the bioaccumulation of SCCPs. The trophic magnification factor (TMF) value of ∑SCCPs was calculated to be 3.98, indicating the potential trophic magnification of SCCPs in this marine food web from ECS. A slightly increasing trend was observed between the TMF values of SCCP homologues and their log KOW values. Overall, for the first time, this study systematically examined the bioaccumulation and trophic magnification of SCCPs in the marine food web from East China Sea.

This study reports on concentrations of short- and medium-chain chlorinated paraffins (SCCPs and MCCPs, respectively) in a wide range of food samples (n = 211) purchased in Belgium during 2020. Samples were analysed by gas chromatography-mass spectrometry (GC-MS) and quantified using chlorine content calibration. ESCCPs were present above LOQ in 25% of samples with an overall range of

Microfibers are one of the most abundant microplastic particle types found in the environment, where they cause negative impacts on organisms and possibly on human health. Microfibers should be included in a wide range of laboratory studies; however, microfibers for scientific studies are not commercially available. Current methods to make microfibers generally create particles with large size ranges and poor precision, and efficient production of particles ≤100 µm is difficult. Laboratory studies of the biological and toxicological effects and chemical interactions of microfibers require uniform, small microfibers in sufficient numbers for environmentally relevant experiments. We developed a novel fiber embedding technique and modified a seminal cryomicrotomy method to produce precise microfibers in quantities suitable for environmentally relevant concentrations. Polyethylene terephthalate (PET) and nylon fibers were strategically wound onto a spindle, embedded in paraffin wax, and sectioned using a standard paraffin microtome. After processing with a suitable organic solvent to remove the wax, microfiber size distributions were assessed. The small microfibers (10-42 µm) were accurate to the target lengths with excellent precision and a production rate ≥13.5 times higher than previous methods. As a proof of application, three lengths of manufactured PET fibers were stained with Nile red and exposed to eastern oyster larvae (Crassostrea virginica) for 24 h. Larvae ingested the smaller fiber lengths (14 and 28 µm), and the Nile red-stained fibers were visible and distinguishable in the guts of the larvae. This experiment was the first to demonstrate ingestion of plastic particles other than microspheres by oyster larvae. The present method facilitates the use of small microfibers in laboratory experiments, allowing for a more complete understanding of microplastic effects in the environment. Environ Toxicol Chem 2022;41:944-953. © 2021 SETAC.

Chlorinated paraffins (CPs) are chemicals with high production volumes that can accumulate at high levels in general populations. The pharmacokinetics of CPs as pollutants is unknown, and there is no evidence that the medium chain chlorinated paraffins (MCCPs) and long chain chlorinated paraffins (LCCPs) are safe replacements for short chain chlorinated paraffins (SCCPs). In this study, SCCPs, MCCPs, and LCCPs were first in vivo and in vitro exposed to rat and liver microsomes, respectively. Toxicokinetics of these compounds were assessed and used to establish the corresponding physiologically based pharmacokinetic (PBPK) models in rats. More than 90% of ingested CPs were deposited in the liver and fat, and the compounds were extremely resistant to metabolism and mostly eliminated via biliary excretion. Then, humans' external and internal exposures to CPs were investigated for one year in Shenzhen, South China. The results were used to calibrate the key parameters for the establishment of a PBPK model in humans. In the PBPK models of rats and humans, the rate of biliary excretion had the greatest influence on the accumulated levels and half-lives of CPs. The body half-lives of human were estimated to be 5.1, 1.2, and 0.6 years for SCCPs, MCCPs, and LCCPs, respectively, suggesting the high accumulation of SCCPs in humans compared to other CPs.

Short-chain chlorinated paraffins (SCCPs) are frequently detected in environmental matrices and human tissues. It was hypothesized that SCCPs might interact with the peroxisome proliferator-activated receptor α (PPARα). In the present study, an in vitro, dual-luciferase reporter gene assay and in silico molecular docking analysis were employed together to study the interactions between SCCPs congeners and PPARα. Expressions of genes downstream in pathways activated by PPARα in liver of rats exposed to 1, 10, or 100 mg/kg bm/d of C10−13-CPs (56.5% Cl) for 28 days were examined to confirm activation potencies of SCCPs toward PPARα signaling. Effects of exposure to C10−13-CPs (56.5% Cl) on fatty acid metabolism in rat liver were also explored via a pseudo-targeted metabolomics strategy. Our results showed that C10−13-CPs (56.5% Cl) caused a dose-dependent greater expression of luciferase activity of rat PPARα. Molecular docking modeling revealed that SCCPs had a strong capacity to bind with PPARα only through hydrophobic interactions and the binding affinity was dependent on the degree of chlorination in SCCPs congeners. In livers of male rats, exposure to 100 mg/kg bm/d of C10−13-CPs (56.5% Cl) resulted in up-regulated expressions of 11 genes that are downstream in the PPARα-activated pathway and regulate catabolism of fatty acid. Consistently, accelerated fatty acid oxidation was observed mainly characterized by lesser concentrations of ∑fatty acids in livers of rats. Overall, these results demonstrated, for the first time, that SCCPs could activate rat PPARα signaling and thereby disrupt metabolism of fatty acid in livers of male rats.

Most emitters of short chain chlorinated paraffins (SCCPs) in China are located in eastern China, posing potential risks to the marine environment and food web. Here we employed a comprehensive atmospheric transport model combined with multiple environmental compartment exchange modules and a marine food web model to simulate levels and risks of SCCPs in the marine environment and fish in the Yellow Sea (YS), East China Sea (ECS), and South China Sea (SCS). Results unveiled a decreasing SCCP level in seawaters and sediments towards offshore. The modeled SCCP total (dry + wet) loadings to the three seawater bodies ranged from 0.0013 to 0.1635 mg/m2/season and gaseous diffusive deposition ranged from 43 to 4443 kg/month. The meteorological factors and secondary emission contributing to seasonal changes in SCCPs were also discussed. A tagging technique was used to trace origins of SCCPs, demonstrating that source proximity contributes most in SCCP contamination to these seawater bodies. Modeled SCCP levels in 5 marine fish in the YS, ECS, and SCS ranged from 23 to 111 ng/g. Our results showed the current SCCP levels in the marine environment and fish did not pose exposure risks to fish consumers for different age groups and genders. However, if consumed fish were harvested and imported from more seriously contaminated seawaters by SCCPs, the estimated dietary intake (EDI) would considerably increase.

Short-chain chlorinated paraffins (SCCPs) are particularly prone to environmental dispersal through long range atmospheric transport. Consequently, they have been detected in biota and environmental matrices at both the North Pole and South Pole. This study shows the first detection of SCCPs in southern hemisphere humpback whales feeding in Antarctic waters. Blubber of specimens stranded along the Australian coastline was analysed and SCCPs were detected in 7 out of 9 individuals. Levels of SCCPs detected in this study were generally low with concentrations up to only 46 ng/g lw. These results were significantly lower than those detected in Northern Hemisphere odontocetes from previous studies, although no reported burdens in northern hemisphere baleen whales are available for comparison. Both the highest level and lowest ( C13. Further investigation is needed in order to evaluate the presence and distribution of SCCPs in the remote Antarctica ecosystem, and delineate longer term environmental consequences of recent inclusion of SCCPs under Annex A of the Stockholm Convention, securing their phase out in ratifying nations.

Short-, medium-, and long-chain chlorinated paraffins (SCCPs, MCCPs, and LCCPs) have a wide range of physical-chemical properties, indicating their varying bioaccumulation tendencies in marine and terrestrial ecosystems. However, there are few empirical data to reveal such bioaccumulation tendencies. In this study, we analyzed SCCPs, MCCPs, and LCCPs in samples from 18 species at both low and high trophic levels of marine and terrestrial ecosystems from the Scandinavian region collected during the past decade. These included fish, seabirds, marine mammals, and terrestrial birds and mammals. SCCPs, MCCPs, and LCCPs were present in all the species, with concentrations ranging from 26-1500, 30-1600, 6.0-1200 ng/g lipid, respectively. Although MCCPs and SCCPs predominated in most species, many terrestrial species had generally higher concentrations of LCCPs than marine species. Terrestrial raptors in particular accumulated higher concentrations of LCCPs, including C 24/25 - which are predominant among very-long-chain components. LCCP concentrations were highest and predominated (55% of total CPs) in peregrine falcons in this study, which is the first report where concentrations of LCCPs surpass those of SCCPs and MCCPs in wildlife. The results also indicate biomagnification of SCCPs, MCCPs, and LCCPs in both marine and terrestrial food chains, but in-depth studies of specific food webs are needed.

Chlorinated paraffins (CPs) are ubiquitous environmental pollutants that are lipophilic and can accumulate in the food chain. Animal-derived products are predominant contributors to human CP exposure. CPs in animal feed might accumulate in domestic animals through dietary exposure, leading to potential contamination of animal-derived food products and human health risks. However, information on the presence of CPs in animal feed materials is scarce. In this study, 16 animal feed material samples were collected in China in 2016. Thirteen of the samples were of animal origin and three were of plant origin. The concentrations and carbon and chlorine congener group profiles of shortchain chlorinated paraffins (SCCPs) and medium-chain chlorinated paraffins (MCCPs) in these animal feed materials were investigated. The concentrations of SCCPs were higher than those of MCCPs in all of the samples. The SCCP concentration range was 120 to 1700 ng/g (mean 640 ng/g), and the MCCP concentration range was 6.4 to 260 ng/g (mean 78 ng/g). Fish meal had the highest SCCP and MCCP concentrations. The lowest SCCP and MCCP concentrations were detected in peanut meal and whey powder, respectively. The concentrations of SCCPs and MCCPs varied among the types of animal feed materials. SCCP and MCCP concentrations also varied among samples of the same type of animal feed material. Relatively high concentrations of SCCPs and MCCPs were detected in feed materials of animal origin. The predominant congener groups in the animal feed materials were C10-11Cl6-7 for SCCPs and C14Cl7 for MCCPs. The carbon and chlorine congener group profiles of SCCPs indicated that SCCP contamination in the animal feed materials might arise from commercial CP mixtures.

Background Objective of our work was to develop a sequential double nonfluorescent immunostaining method which allows the selective identification of myelinated motor fibers in paraffin-embedded samples of peripheral nerves. Motor recovery after a nerve gap-lesion repaired by artificial nerve-guides ("conduits") is often less complete and slower than sensory recovery. The mechanism for this is not fully understood. New method Incubation in sheep polyclonal choline acetyltransferase antibody (Abcam 18,736) at dilution of 1:150 was followed by incubation in mouse monoclonal anti-myelin basic protein antibody (Abcam 62,631) at a dilution of 1:5000. Counterstaining was performed with hematoxylin QS (Vector Labs H-3404). Results Immunostaining of choline acetyltransferase and myelin basic protein can be combined together and results show a good contrast between the light brown of the choline acetyltransferase reaction product and the green of myelin basic protein reaction product. Cell nuclei are stained blue. This new protocol retains the advantages of paraffin embedded sections such as (i) having a relatively simple methodology, (ii) years-long storage life, and (iii) easy sharing among laboratories. Comparison with existing method. This specific combinatorial protocol has never been used before on paraffin embedded sections. It has been named "reciprocal nerve staining" (RNS). Conclusions Routine combination of choline acetyltransferase and myelin basic protein immunostaining provides a highly specific, highly contrasted paraffin-embedded sections where optical differentiation of myelinated motor fibers is easy and straightforward. This method will likely simplify and speed-up the routine histological study of nerve regeneration and will contribute a better identification of the nerve motor component.

The bone is a dynamic and metabolically active organ in which growth and resorption of the osteochondral matrix is orchestrated by osteoblasts and osteoclasts. For decalcified paraffin-embedded specimens, decalcifying agents alter the staining intensity, and excess decalcification interferes with bone staining. Robust bone staining methods independent of the decalcification conditions and animal species are lacking. In this study, we have developed a novel polychrome staining method, named JFRL staining, which stains the components of osteochondral tissue in different colors. With this staining we could visualize the hyaline cartilage as blue by alcian blue, osteoid as red by picrosirius red, and mineralized bone as green by picro-light green SF or picro-naphthol green B and easily distinguished osteoblasts, osteocytes, and osteoclasts. In mineralized bone, this staining revealed the obvious lamellar structures and woven bone. Notably, this staining was independent of the decalcification conditions and experimental animal species examined. To verify the usefulness of JFRL staining, we observed cotton rat tail which has shorter length and shows a false autotomy. The caudal vertebrae were normally developed via endochondral ossification without a fracture plane. At 6 months of age, the number of chondrocytes declined and the hypertrophic zone was absent at the epiphyseal plate, which might reflect the shorter tail. In conclusion, JFRL staining is the first method to simultaneously distinguish osteochondral matrix and bone cells in one section regardless of decalcifying conditions. This robust staining will provide new information for a wide number of biomedical fields, including bone development, physiology, and pathology.

For the most complex artificial chlorinated environmental contaminants, much less is known for medium-chain CPs than short-chain CPs. In this research, the spatial distributions of MCCPs and SCCPs in farmland soil and maize leaves near a CP production facility were found marginally influenced by seasonal winds. The levels of ∑MCCPs and ∑SCCPs were in the ranges of

Although all cells contain iron, most histochemical methods fail to reveal the presence of iron within many cells of the central nervous system (CNS), particularly neurons. Previously, a sensitive method was developed that limited the extraction of iron in paraffin sections, and this method revealed staining within neurons. However, the staining was often too robust making it difficult to discern discrete intracellular structures. In 1970, a study incorporated acetone in an iron histochemical procedure to facilitate the demarcation of staining features. In the present study, both acetone and limits to iron extraction were included in a simplified staining procedure. This procedure was applied to paraffin sections of CNS tissue from CISD2 deficient and littermate control mice. Discrete nuclear and cytoplasmic staining features were detected in all mice. Although widely present in neurons, punctate cytoplasmic staining was particularly prominent in large neurons within the hindbrain. Evaluation of extended depth of focus images, from serial focal planes, revealed numerous stained cytoplasmic structures. Additionally, the simplified staining procedure was applied to paraffin sections from Alzheimer’s disease and control cases. Despite suboptimal processing conditions compared to mouse tissue, discrete staining of cytoplasmic structures was revealed in some neurons, although many other neurons had nondescript staining features. In addition, initial findings revealed iron deposited within some vessels from patients with Alzheimer’s disease. In summary, since paraffin sections are commonly used for histological preparations, this simplified histochemical procedure could facilitate the study of iron in various CNS conditions by revealing staining details often missed by other procedures.

International audience; Reliable human health risk assessment associated with chlorinated paraffins (CPs) exposure is limited by the lack of data on the fate of this complex family of contaminants. To gain knowledge on the accumulation and distribution of CPs in biota after ingestion, laying hens were dietary exposed to technical mixtures of short- (SCCPs), medium- (MCCPs), or long-chain (LCCPs) CPs of various chlorine contents during 91 days, at 200 ng/g of feed, each. Adipose tissue, liver, muscle and serum were collected at the steady-state, along with excreta. All C10-C36 CPs were detected in liver. However, differences were observed in CP distribution: LCCPs high %Cl were retained in the liver; LCCPs low %Cl circulated through the serum and were distributed in the different compartments, but were mostly excreted through the eggs; SCCPs and MCCPs were found in all tissues at similar levels. Finally, a mass balance indicated a potential for biotransformation.

Urban herring gulls (Larus argentatus) are exposed to contaminants from aquatic, terrestrial and anthropogenic sources. We aim to assess if differences in urbanisation affect ecological niche and contaminant concentrations in female herring gulls. Furthermore, we investigated maternal transfer from mothers to eggs for all the target compounds, including chlorinated paraffins (CPs) and cyclic volatile methyl siloxane (cVMSs), which to our knowledge have not been assessed in herring gulls previously. We compare concentrations of legacy and emerging contaminants and metals in blood and eggs between two herring gull colonies located 51 km apart, in the urban influenced Norwegian Oslofjord. While both colonies are within an urbanised area, the inner fjord is more so, as it is surrounded by Oslo, the capital and largest city in Norway Stable isotopes of carbon and nitrogen indicated a more marine ecological niche in the outer than the inner fjord colony, although with overlap. Persistent organic pollutant (POP) concentrations were similar in the inner and outer fjord colonies, while the short-chained chlorinated paraffins (SCCP), which are recently added to the Stockholm convention and contaminants of emerging concern (CECs) varied, with higher concentrations of SCCP and the cVMS decamethylcyclopentasiloxane (D5) in females and eggs of the inner fjord colony. Per- and polyfluorinated substances (PFAS) concentrations were higher in the outer fjord colony, likely linked to releases from a point-source (airport and waste management facility with open access to food waste). In blood, chlorinated paraffins contributed most the total lipophilic contaminants (inner: 78%, outer: 56%), while polychlorinated biphenyls (PCBs) were the most abundant lipophilic contaminants in eggs (inner: 62%, outer: 46%). Dechloranes and brominated flame retardants (BFRs) were detected in few samples. Maternal transfer, assessed by egg to blood ratios, of cVMSs were similar to the POPs with mean log ratio 0.39 (D5), while it was lower for SCCPs, with log ratios-0.77. Our results indicate comparable POP exposure of the herring gulls in the inner and outer Oslofjord, likely due to overlap in ecological niches between the colonies and wide distribution of POPs. The differences between the colonies in concentrations of PFAS, cVMS and CPs shows that point source exposures and urban influence may be more important than ecological niche for these compounds.