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Arsenical vesicants cause skin inflammation, blistering, and pain. The lack of appropriate animal models causes difficulty in defining their molecular pathogenesis. Here, Ptch1sup+/-/sup/C57BL/6 mice were employed to investigate the pathobiology of the arsenicals lewisite and phenylarsine oxide (PAO). Following lewisite or PAO challenge (24 h), the skin of animals becomes grayish-white, thick, leathery, and wrinkled with increased bi-fold thickness, Draize score, and necrotic patches. In histopathology, infiltrating leukocytes (macrophages and neutrophils), epidermal-dermal separation, edema, apoptotic cells, and disruption of tight and adherens junction proteins can be visualized. PCR arrays and nanoString analyses showed significant increases in cytokines/chemokines and other proinflammatory mediators. As hair follicles (HFs), which provide an immune-privileged environment, may affect immune cell trafficking and consequent inflammatory responses, we compared the pathogenesis of these chemicals in this model to that in Ptch1sup+/-/sup/SKH-1 hairless mice. Ptch1sup+/-/sup/SKH-1 mice have rudimentary, whereas Ptch1sup+/-/sup/C57BL/6 mice have well-developed HFs. Although no significant differences were observed in qualitative inflammatory responses between the two strains, levels of cytokines/chemokines differed. Importantly, the mechanism of inflammation was identical; both reactive oxygen species induction and consequent activation of unfolded protein response signaling were similar. These data reveal that the acute molecular pathogenesis of arsenicals in these two murine models is similar.

Efficacies of essential oils (EOs) of Vetiveria zizanioides (L.) Nash. (Poales: Poaceae) (VZ EO), Cananga odorata (Lam) Hook. F. & Thomson (Magnoliales: Annonaceae) (CO EO), and crude extract (CE) of Andrographis paniculata (Burm.F.) Wall ex. Nees (Lamiales: Acanthaceae) (AP CE), against laboratory (lab) and field strains of Culex quinquefasciatus Say were investigated. Irritant and repellent activities of individual and binary mixtures of plant extracts were compared with N,N-diethyl-m-toluamide (DEET) using an excito-repellency system. The irritant activity (direct tarsal contact), the mean percent escape response of VZ EO (91.67%, 83.33%), and CO EO (80%, 88.33%) were not significantly different compared with DEET (88.33%, 95%) against lab and field strains, respectively. Similarly, irritant responses in combinations (1:1 and 1:2, v:v) of either VZ EO or CO EO with AP CE were not significantly different from DEET against both strains (P > 0.001). The repellent activity (no tarsal contact), the mean percent escape response of VZ EO (68.33%), CO EO (61.67%), and VZ EO+AP CE (1:1, v:v) (81.67%) against lab strain and CO EO (85%) against field strain were not significantly different from that of DEET (P > 0.001). Interestingly, the greatest contact irritancy of VZ EO+AP CE (1:1, v:v) (96.67%) (P = 0.0026) and a stronger repellency response of CO EO (85%) (P = 0.0055) produced significantly different patterns of escape response compared with DEET against both lab and field strains, respectively. The EOs of VZ EO and CO EO or their mixture with AP CE showed potential as plant-based active ingredients for mosquito repellents. In addition, the major chemical constituents of VZ EO were β-vetivone (6.4%), khusimol (2.96%), and α-vetivone (2.94%) by gas chromatograpy–mass spectrometry.

The skin is an important barrier that protects against invasion by foreign substances, including irritants and harmful microorganisms, and holds water in the body. Washing the skin with cleansers and shampoos containing anionic surfactants, for example sodium dodecyl sulfate (SDS), is important for maintaining skin homeostasis. However, surfactants can cause dermatitis, cutaneous hypersensitivity (e.g., alloknesis), and pruritus in humans. Our previous studies revealed an alloknesis response in the skin with SDS-induced dermatitis in C57BL/6 mice. In addition, we found that alloknesis responses and afterdischarge responses following stimulation with light touch are related because they are observed contemporaneously. In this study, we used Hos:HR-1 hairless mice to establish a mouse model to evaluate long-term drug application for alloknesis responses. Alloknesis was observed in HR-1 mice with SDS-induced dermatitis. The mean number of c-Fos (a marker of neural activity) immunopositive neurons was increased in the lamina 1-2 (L1-2) spinal dorsal horn, but not in L3-4, of SDS-treated HR-1 mice compared to vehicle-treated mice. We also discovered that afterdischarge responses were observed in neurons in L1-2. There was also a correlation between the intensity of the afterdischarge responses and depth of the recording site. Thus, the following were suggested: 1) neurons that mediate these afterdischarge responses are located on the superficial layer of the spinal cord; 2) afterdischarge responses can be an index of alloknesis responses, and 3) the mouse model of SDS-induced dermatitis is an appropriate alloknesis model.

International audience; Chemotherapy is almost exclusively administered via the intravenous (IV) route, which has serious limitations (e.g., patient discomfort, long hospital stays, need for trained staff, high cost, catheter failures, infections). Therefore, the development of effective and less costly chemotherapy that is more comfortable for the patient would revolutionize cancer therapy. While subcutaneous (SC) administration has the potential to meet these criteria, it is extremely restrictive as it cannot be applied to most anticancer drugs, such as irritant or vesicant ones, for local toxicity reasons. Herein, we report a facile, general, and scalable approach for the SC administration of anticancer drugs through the design of well-defined hydrophilic polymer prodrugs. This was applied to the anticancer drug paclitaxel (Ptx) as a worst-case scenario due to its high hydrophobicity and vesicant properties (two factors promoting necrosis at the injection site). After a preliminary screening of well-established polymers used in nanomedicine, polyacrylamide (PAAm) was chosen as a hydrophilic polymer owing to its greater physicochemical, pharmacokinetic, and tumor accumulation properties. A small library of Ptx-based polymer prodrugs was designed by adjusting the nature of the linker (ester, diglycolate, and carbonate) and then evaluated in terms of rheological/viscosity properties in aqueous solutions, drug release kinetics in PBS and in murine plasma, cytotoxicity on two different cancer cell lines, acute local and systemic toxicity, pharmacokinetics and biodistribution, and finally their anticancer efficacy. We demonstrated that Ptx-PAAm polymer prodrugs could be safely injected subcutaneously without inducing local toxicity while outperforming Taxol, the commercial formulation of Ptx, thus opening the door to the safe transposition from IV to SC chemotherapy.

The present study aims to investigate the correlation between irritant toxicity variation and lectin content variation during the processing of Pinelliae Rhizoma products and to explore the feasibility of Western blot as a method for the detection of lectin. We processed Pinelliae Rhizoma Praeparatum Cum Alumine, Pinelliae Rhizoma Praeparatum, and Pinelliae Rhizoma Praeparatumcum Zingibere et Alumine to different degrees and then analyzed their irritant toxicity via Draize rabbit eye test. Western blot was employed to determine the lectin content in Pinelliae Rhizoma products processed with different methods. The correlation between toxicity variation and lectin content variation was then analyzed. Different decoction pieces of Pinelliae Rhizoma were collected for the determination of lectin content. The three processed products of Pinelliae Rhizoma showed gradually decreased toxicity and lectin content as the processing continued. The decreasing trend of lectin content was consistent with that of irritant toxicity during processing, which indicated that the change in lectin content could reflect the trend of irritant toxicity. No band of lectin appeared in the Western blot of processed products of Pinelliae Rhizoma, which suggested that western blotting can be used for the detection of toxic lectin in the processed products of Pinelliae Rhizoma. Lectin should not be detected in the Pinelliae Rhizoma products processed according to the methods in the Chinese Pharmacopoeia.

BackgroundEvery evening, chimpanzees (Pan troglodytes) build a sleeping platform so called “nest” by intertwining branches of tree. Most of chimpanzees’ communities studied have a preference for tree species in which they nest. As female mosquitoes are feeding on the blood of their host at nighttime, chimpanzees may prevent being disturbed and bitten by mosquitoes by selecting tree species having properties to repel them.MethodsTo test the hypothesis that chimpanzees choose tree species for their aromatic properties, data related to 1,081 nesting trees built between 2017 and 2019 in the Sebitoli community of Kibale National Park (Uganda) were analysed. The 10 most used trees were compared to the 10 most common trees in the habitat that were not preferred for nesting. Leaves from the 20 trees species were collected and hydro-distillated to obtain essential oils and one of the by-products for behavioural bioassays against females of the African mosquito,Anopheles gambiae.ResultsSebitoli chimpanzees showed tree preferences: 10 species correspond to more than 80% of the nesting trees. Out of the essential oil obtained from the 10 nesting trees, 7 extracts for at least one concentration tested showed spatial repellency, 7 were irritant by contact and none were toxic. In the other hand, for the abundant trees in their habitat not used by chimpanzees, only 3 were repellent and 5 irritants.Discussion and conclusionThis study contributes to evidence that chimpanzees, to avoid annoying mosquitoes, may select their nesting trees according to their repellent properties (linked to chemical parameters), a potential inspiration for human health.

The sensory perception of irritant chemicals results in escape and repellency behavioral patterns in insects. Transient receptor potential channels are cation channels that function as sensor proteins for several types of signals, such as light, sound, temperature, taste, as well as chemical and physical stimuli; among these, the TRPA channel is widely conserved and activated by irritant chemicals. Certain plant-derived essential oils (EOs), produced by secondary metabolism, are mixtures of volatile compounds, which are used as repellents because they contain environmentally sustainable ingredients. Citronellal, which is present in citronella EO from Cymbopogon species, is a potentially viable insect repellent; however, the repellency capability against coleopteran beetles remains limited. We investigated the citronellal-derived repellency behavior for the red flour beetle Tribolium castaneum, in which TcTRPA1 and odorant receptor co-receptor (Orco) expressions were mediated by RNA interference. Area-preference tests showed dose-dependent repellency behavior for citronellal; additionally, both TcTRPA1 and TcOrco double-strand RNA (dsRNA) micro-injection showed clear TcTRPA1 and TcOrco transcript reductions, and only TcTRPA1 dsRNA treatment significantly impaired repellency behavior. The relative expression level of the TcTRPA1 transcripts, evaluated by quantitative reverse-transcription PCR (qRT-PCR), revealed dominant expression in the antennae, indicating the antennae-expressed TcTRPA1-mediated repellency behavior.

BACKGROUND Exploiting indoor-resting mosquitoes' innate behavioral responses to commonly used insecticide is crucial in vector control programs. Indoor residual spraying (IRS) and long-lasting insecticidal nets (LLINs) treated with pyrethroids have become widely used for controlling dengue fever vectors. The present study tested the effects of piperonyl butoxide (PBO) synergism and cuticular thickening on the contact irritancy response of field A. aegypti (Diptera: Culicidae) to deltamethrin in Taiwan and Thailand. RESULTS The escape response of field mosquitoes treated with PBO was significantly elicited, with an escape percentage increase between 2- and 10-fold. In addition, the escape time was significantly lower in PBO-pretreated mosquitoes compared with field mosquitoes treated with deltamethrin alone. PBO-pretreated mosquitoes from seven out of 11 field strains exhibited a knockdown percentage of 11.23-54.91%, significantly higher than that of mosquitoes in corresponding strains treated with deltamethrin only. The Annan, Zhongxi, Sanmin, and North strains exhibited weak knockdown responses (≤3.75%). The mortality of PBO-pretreated field mosquitoes increased 2- to 75-fold compared with those treated with deltamethrin alone (mortality: 0-6.70%). Furthermore, the effect of cuticular thickness on the escape response of field mosquitoes was significant, that is, the escape response marginally increased inversely to cuticular thickness. By contrast, cuticular thickness was not significantly associated with knockdown or mortality percentage. CONCLUSION Irritant behavior in mosquitoes was significantly elicited by PBO synergism. PBO incorporating deltamethrin IRS or LLINs may be effective for controlling dengue fever vectors. © 2021 Society of Chemical Industry.

Background Contact sensitizers may interfere with correct protein folding. Generation of un-/misfolded proteins can activate the IRE-1 or PERK signaling pathways initiating the unfolded protein response (UPR) and thereby determine inflammatory immune responses. We have analyzed the effect of sensitizers with different potencies on the induction of UPR activation/inhibition and the subsequent generation of a pro-inflammatory micromilieu in vitro as well as the effect of UPR modulation on the inflammatory response in the murine contact hypersensitivity (CHS) in vivo. Methods Semi-quantitative and quantitative PCR, fluorescence microscopy, ELISA, NF-κB activation and translocation assays, DC/keratinocyte co-culture assay, FACS, and in vivo CHS experiments were performed. Results Sensitizers and irritants activate IRE-1 and PERK in murine and human keratinocytes. Synergistic effects occur after combination of different weak sensitizers / addition of irritants. Moreover, tolerogenic dinitrothiocyanobenzene can be converted into a strong sensitizer by pre-activation of the UPR. Blocking UPR signaling results in decreased NF-κB activation and cytokine production in keratinocytes and in activation marker downregulation in a HaCaT/THP-1 co-culture. Interestingly, not only systemic but also topical application of UPR inhibitors abrogates CHS responses in vivo. Conclusion These observations highlight an important role of the UPR in determination of the inflammatory response in vitro and in vivo further underlining the importance of tissue stress and damage responses in the development of ACD and provide mechanistically based concepts as a basis for the development of new therapeutic approaches to treat allergic contact dermatitis.

The Expert Panel for Cosmetic Ingredient Safety (Panel) assessed the safety of 5 acyl sarcosines and 9 sarcosinate salts as used in cosmetics; all of these ingredients are reported to function in cosmetics as hair conditioning agents and most also can function as surfactants—cleansing agents. The ingredients reviewed in this assessment are composed of an amide comprising a fatty acyl residue and sarcosine and are either free acids or simple salts thereof. The Panel relied on relevant new data, including concentration of use, and considered data from the previous Panel report, such as the reaction of sarcosine with oxidizing materials possibly resulting in nitrosation and the formation of N-nitrososarcosine. The Panel concluded that these ingredients are safe as used in cosmetics when formulated to be non-irritating, but these ingredients should not be used in cosmetic products in which N-nitroso compounds may be formed.

The OptiSafe (OS) test is shelf-stable, macromolecular eye irritation test that does not include any animal ingredient or component ("vegan"). The purpose of this study was to evaluate the test's accuracy for an expanded application domain for both the original and recently updated OS method. This study involved the testing of additional ocular corrosives and previously excluded foaming agents ("surfactants") using both the original and updated OS methods and then combining these data with prior validation data for a total of 147 chemicals. Predictivity was evaluated by a statistical comparison of the OptiSafe predictions with historical in vivo "Draize" rabbit eye data for the same chemicals (from public databases). We report that for the detection of chemicals not requiring classification for eye irritation [Globally Harmonized System of Classification and Labeling of Chemicals (GHS) No Category], the accuracy, specificity, and sensitivity were 92.8%, 79.6%, and 100.0%, respectively, for the updated method; for the detection of chemicals inducing extreme eye damage/corrosion (GHS Category 1), the accuracy, specificity, and sensitivity were 79.4%, 71.8%, and 91.7%, respectively, for the updated method. Results indicate that both the original and updated methods have a high accuracy for the expanded application domain that included ocular corrosives and surfactants.

Silicosis is a worldwide serious occupational disease that is caused by inhalation of silica crystals. However, little is known about the pathogenesis mechanism of silicosis. We performed single-cell sequencing in bronchoalveolar lavage fluid (BALF) from mine workers with silicosis and their co-workers who did not develop silicosis, and found that the RAB20 deficiency in monocytes/macrophages was strongly linked to the development of silicosis. In the silicosis murine model, RAB20 knockout markedly enhanced the silica crystal-induced pulmonary interstitial fibrosis and respiratory dysfunction. Moreover, this process is strongly accompanied by IL-1β release and NLRP3 activation.

Purpose: Fluconazole is an effective anti-fungal drug. Due to the limitations of fluconazole, such as poor water solubility and consequently low ocular bioavailability, an optimized fluconazole nanoemulsion in-situ gel formulation (temperature-sensitive) was developed. Methods and Materials: To verify formulation’s safety for ophthalmic use, preparation was tested for potential ocular toxicity using a cell viability assay on retinal cells. The hen’s egg test-chorioallantoic membrane (HET-CAM), as a borderline test between in vivo and in vitro techniques, was chosen for investigating the irritation potential of the formulation. HET-CAM test was done by adding the formulation directly to the CAM surface and monitoring the vessels visually in terms of irritation reactions. Eye tolerance was determined using the modified Draize test. Results: Viability assay on retinal cells displayed that fluconazole nanoemulsion in-situ gel formulation was non-toxic and can be safely used in the eye at concentrations of 0.1% and 0.5%. HET-CAM and Draize tests revealed that optimized formulation of fluconazole did not result in any irritation and was considered non-irritant and well-tolerated for ocular use. Conclusion: Regarding to the findings of the three mentioned methods, fluconazole nanoemulsion in-situ gel formulation is harmless and as a proper and safe alternative, can be considered for ocular delivery of fluconazole in the future.

Introduction Electronic cigarettes (e-cigs) are currently used by millions of adults and adolescents worldwide. Major respiratory symptoms, such as coughing reported by e-cig users, including patients with e-cig, or vaping, product use-associated lung injury (EVALI), indicate e-cig constituent-induced sensory irritation. However, e-cig constituent-induced nociceptive activity in nasal and tracheal respiratory epithelia (RE) and neuronal activation in the trigeminal ganglia and brainstem nuclei, which receive airway chemosensory inputs have not been examined and compared. Comparisons of physiological responses between freebase nicotine and nicotine salts are also missing. Aims and Methods Event-related potential (ERP) was recorded electrophysiologically to assess mouse nasal and tracheal RE chemosensory responses to various flavorings, nicotine, including freebase and nicotine salts, e-liquid mixtures, and tussigenic stimuli. Also, mice were subjected to inhalation exposure to aerosol of a vanilla-flavored e-liquid or air (control), and the activated-trigeminal nociceptive neurons and brainstem neurons were examined using immunohistochemistry. Results Individual constituents and mixtures of e-liquids, capsaicin, and citric and acetic acids evoked significantly larger ERP in the nose than in the trachea with the exception of menthol. ERP responses to freebase nicotine were significantly larger than protonated nicotine. Four nicotine salts (benzoate, lactate, levulinate, and salicylate) induced similar responses. Compared with air-exposed mice, e-liquid aerosol-exposed mice showed a significant increase in numbers of activated trigeminal nociceptive neurons and brainstem neurons in the spinal trigeminal nucleus, paratrigeminal nucleus, and nucleus tractus solitarius. Conclusions E-liquid constituents region-dependently stimulate airway nociceptive chemosensory systems, and freebase nicotine is more potent than protonated nicotine. Implications Neural abnormalities have been implicated in the development of nasal and respiratory illnesses. The higher sensitivity of the nasal nociceptive chemosensory system to nicotine and flavorings may indicate a health risk for e-liquid aerosol-induced upper airway illnesses via neurogenic alteration and warrants further investigation.

The use of tobacco products is a major global public health issue, as it is the leading cause of preventable death worldwide. In addition, nicotine (NIC) is a key component of electronic and conventional cigarettes. Although nicotine’s addictive potential is well known, its health effects are not entirely understood. Thus, the main objective of the present study was to evaluate its toxicological profile both in vitro, at the level of three healthy cell lines, and in ovo, at the level of the chorioallantoic membrane. Five different concentrations of nicotine were used in keratinocytes, cardiomyocytes, and hepatocytes for the purpose of evaluating cell viability, cell morphology, and its impact on nuclei. Additionally, the hen’s egg test on the chorioallantoic membrane (HET-CAM) method was used to assess the biocompatibility and irritant potential of the chorioallantoic membrane. Across all cell lines studied, nicotine was proven to be significantly damaging to cell viability, with the highest concentration tested resulting in less than 2% viable cells. Moreover, the morphology of cells changed dramatically, with alterations in their shape and confluence. Nicotine-induced cell death appears to be apoptotic, based on its impact on the nucleus. In addition, nicotine was also found to have a very strong irritating effect on the chorioallantoic membrane. In conclusion, nicotine has an extremely strong toxicological profile, as demonstrated by the drastic reduction of cell viability and the induction of morphological changes and nuclear alterations associated with cellular apoptosis. Additionally, the HET-CAM method led to the observation of a strong irritating effect associated with nicotine.

Introduction: Repeated skin contact to detergents causes chronic irritant contact dermatitis (ICD) associated with itch sensation and eczema. However, the mechanisms of detergent-induced ICD are poorly understood. Here, we established a new murine model of detergent-induced ICD with H1-antihistamine-refractory itch. Methods: Ear skin of wild-type and mast cell-deficient mice on the C57BL/6 genetic background was treated with a detergent, sodium dodecyl/lauryl sulfate (SDS), daily for approximately 2 weeks with or without administration of an H1-antihistamine, fexofenadine. Skin inflammation, barrier dysfunction, and itching were analyzed. Quantitative PCR for earlobe gene expression and flow cytometry analysis for draining lymph node cells were conducted. Results: SDS treatment induced skin inflammation with ear swelling, increased transepidermal water loss, and hind-paw scratching behaviors in the wild-type and mast cell-deficient mice. The peak value of scratching bouts was retained for at least 48 h after the last SDS treatment. H1-antihistamine administration showed no or little reduction in the responses. SDS treatment upregulated gene expression for a Th2 cytokine IL-4 and Th17/Th22 cytokines, IL-17A, IL-17F, and IL-22, and increased cell numbers in draining lymph nodes of CD4+ T, CD8+ T, and γδT cells with enhanced expression of GATA3, RORγt, T-bet, or FOXP3 compared with untreated mice. Conclusions: The present study showed that SDS treatment of ear skin in C57BL/6 mice induces mast cell-independent skin inflammation with H1-antihistamine-refractory itch and suggested a possible Th cytokine- and/or lymphocyte-mediated regulation of the model. The model would be useful for elucidation of mechanisms for inflammation with H1-antihistamine-refractory itch in detergent-induced ICD.

Serious eye damage and eye irritation have been authenticated to be significant human health issues in various fields such as ophthalmic pharmaceuticals. Due to the shortcomings of traditional animal testing methods, in silico methods have advanced to study eye toxicity. The models for predicting serious eye damage and eye irritation potential of compounds were developed using 2299 and 5214 compounds, respectively. The 40 global single models and 40 local models were developed by combining 5 molecular description methods and 4 machine learning methods. The 40 active learning models were developed by adopting uncertainty-based active learning strategies and taking local models as initial models. The 110 global consensus models based on 40 global single models were developed using a consensus strategy. Active learning models and global consensus models performed high prediction accuracy. The test accuracy of the best serious eye damage model and eye irritation model reached 0.972 and 0.959, respectively. The applicability domains for all models were calculated to verify the rationality of prediction effect. In addition, 8 structural alerts probably causing serious eye damage or eye irritation were sought out. The prediction models and structural alerts contributed to providing hazard identification and assessing chemical safety.

As a result of the efforts to introduce the principle of the 3Rs (replacement, reduction, and refinement) into animal testing, alternative in vitro skin corrosion test methods have been developed and standardized globally. However, alternative in vitro skin corrosion test methods have some limitations in terms of the use of humanely killed rats or commercial models and kits. The present study focused on the applicability of invertebrates as alternative in vivo skin models. Even though earthworm skin comprises the same biomolecules as human skin, the possibility of using earthworm skin as an alternative for skin testing remains unexplored. In this study, we developed a half-pipe tool for earthworm skin corrosion testing and optimized the test protocol. Subsequently, the applicability of the earthworm half-pipe assay for corrosion testing with six chemicals, including inorganic acids, organic acids, and alkalis, was investigated using stereomicroscopy and electron microscopy. It was observed that the specific concentrations for earthworm skin corrosion were lower than those for animal or in vitro tests. Therefore, the sensitivity of the earthworm half-pipe assay indicates that it could be useful as a screening tool before conducting in vivo animal tests or in vitro skin tests. This new method can contribute to research on alternative skin corrosion tests by reducing ethical issues, time, and cost while achieving effective results.

Exposure to vesicants, including sulfur mustard and nitrogen mustard, causes damage to the epithelia of the respiratory tract and the lung. With time, this progresses to chronic disease, most notably, pulmonary fibrosis. The pathogenic process involves persistent inflammation and the release of cytotoxic oxidants, cytokines, chemokines, and profibrotic growth factors, which leads to the collapse of lung architecture, with fibrotic involution of the lung parenchyma. At present, there are no effective treatments available to combat this pathological process. Recently, much interest has focused on nutraceuticals, substances derived from plants, herbs, and fruits, that exert pleiotropic effects on inflammatory cells and parenchymal cells that may be useful in reducing fibrogenesis. Some promising results have been obtained with nutraceuticals in experimental animal models of inflammation-driven fibrosis. This review summarizes the current knowledge on the putative preventive/therapeutic efficacy of nutraceuticals in progressive pulmonary fibrosis, with a focus on their activity against inflammatory reactions and profibrotic cell differentiation.

Background Preservatives represent one of the main causes of skin irritation and contact allergies. Aims To comprehensively evaluate the skin irritation potential of phenoxyethanol, methylparaben, propylparaben, imidazolidinyl urea, and DMDM hydantoin under regulatory acceptable concentrations. Methods A patch test and repeated open application test (ROAT) were applied to evaluate skin irritation in vivo. In vitro alternative methods consisting of the keratinocyte cytotoxicity assay, red blood cell (RBC) test, and hen's egg test-chorioallantoic membrane (HET-CAM) were performed to elucidate the mechanism of preservative-induced irritation responses. Results The patch test showed that all test substances showed a weak erythema response. Propylparaben had the highest occlusive irritancy potential in the patch test, owing to damage to the cell membrane. The two formaldehyde releasers showed noticeable skin irritation potential in the ROAT through their cytotoxicity to keratinocytes, while a visible response was observed after applying phenoxyethanol and the two parabens. No filtration was noticed in the in vivo tests, which might be attributed to the failure of subcutaneous vessel alteration by the preservatives. Conclusions Commonly used cosmetic preservatives have minor skin irritation potential with mild erythema reaction under practical use, especially formaldehyde releasers and propylparaben.

Occupational immune diseases are a serious public health burden and are often a result of exposure to low molecular weight (LMW) chemicals. The complete immunological mechanisms driving these responses are not fully understood which has made the classification of chemical allergens difficult. Antimicrobials are a large group of immunologically-diverse LMW agents. In these studies, mice were dermally exposed to representative antimicrobial chemicals (sensitizers: didecyldimethylammonium chloride (DDAC), ortho-phthalaldehyde (OPA), irritants: benzal-konium chloride (BAC), and adjuvant: triclosan (TCS)) and the mRNA expression of cytokines and cellular mediators was evaluated using real-time qPCR in various tissues over a 7-days period. All antimicrobials caused increases in the mRNA expression of the danger signals Tslp (skin), and S100a8 (skin, blood, lung). Expression of the TH2 cytokine Il4 peaked at different timepoints for the chemicals based on exposure duration. Unique expression profiles were identified for OPA (Il10 in lymph node, Il4 and Il13 in lung) and TCS (Tlr4 in skin). Additionally, all chemicals except OPA induced decreased expression of the cellular adhesion molecule Ecad. Overall, the results from these studies suggest that unique gene expression profiles are implicated following dermal exposure to various antimicrobial agents, warranting the need for additional studies. In order to advance the development of preventative and therapeutic strategies to combat immunological disease, underlying mechanisms of antimicrobial-induced immunomodulation must be fully understood. This understanding will aid in the development of more effective methods to screen for chemical toxicity, and may potentially lead to more effective treatment strategies for those suffering from immune diseases.

Inflammation is an innate immune response of the body against pathogens and other irritants. The NLRP3 (NACHT, LRR and PYD domains-containing protein 3) inflammasome is a major player in the inflammatory response against pathogenic microorganisms. In this study, we analyzed the relationship between the NLRP3 inflammasome and the influenza virus NS1 protein, which is involved in host immune escape. The canine influenza virus NS1 protein transcriptionally attenuated proinflammatory cytokines by inhibiting the nuclear factor-κB (NF-κB) activator. NS1 also directly interacted with NLRP3 and blocked ASC (Apoptosis-associated speck-like protein containing CARD) oligomerization, which deactivated the NLRP3 inflammasome. In addition, NS1 inhibited pro-caspase 1 cleavage into caspase-1, which prevents maturation of IL-1β and IL-18 from their respective precursors, eventually reducing the inflammatory response. Taken together, the influenza NS1 protein evades host immunity, and our findings provide a theoretical basis for the prevention and treatment of canine influenza.

To replace the Draize eye irritation test (OECD Test Guideline 404), several test methods based on reconstructed cornea-like epithelium (RhCE) have been developed and adopted in the OECD TG 492. The objective of this study was to stablish the experimental procedures and evaluate the performance assessment of QobuR-RhCE, an in-house RhCE model to be used for the evaluation of eye hazard. We define the essential structural, functional and procedural elements of the test method components to help assuring that the proposed test method is based on the same concepts as the validated reference methods. Performance assessment was evaluated in accordance with the revised performance standards for the assessment of proposed similar or modified in vitro reconstructed human cornea-like epithelium and the minimum list of reference chemicals was evaluated. As result, the proposed method scored 93.3% sensibility, 60% specificity, 76.7% accuracy and 96.7% within-laboratory reproducibility (WLR), providing a similar performance in comparison to the validated reference methods. Additionally, we describe a secondary endpoint based on Transepithelial Electrical Resistance (TEER) that could be of use to better discriminate between irritants and non-irritants. Taken together the results indicate that the QobuR-RhCE test method is an accurate screening tool that can be used as a standalone alternative to evaluate ocular irritation.

Chemotherapy causes intestinal mucositis, which includes villous atrophy and altered mucosal barrier function. However, there is an uncertainty regarding how the reduced small-intestinal surface area affects the mucosal permeability of the small marker probe mannitol (MW 188), and how the mucosa responds to luminal irritants after chemotherapy. The aims in this study were to determine (i) the relationship between chemotherapy-induced villus atrophy and the intestinal permeability of mannitol and (ii) how the mucosa regulate this permeability in response to luminal ethanol and sodium dodecyl sulfate (SDS). This was investigated by treating rats with a single intraperitoneal dose of doxorubicin, irinotecan, or 5-fluorouracil. After 72 h, jejunum was single-pass perfused and mannitol permeability determined at baseline and after 15 min luminal exposure to 15% ethanol or 5 mg/mL SDS. Tissue samples for morphological analyses were sampled from the perfused segment. All three chemotherapeutics caused a similar 30% reduction in villus length. Mannitol permeability increased with irinotecan (1.3-fold) and 5-fluorouracil (2.5-fold) and was reduced with doxorubicin (0.5-fold), suggesting that it is not epithelial surface area alone that regulates intestinal permeability to mannitol. There was no additional increase in mannitol permeability induced by luminal ethanol or SDS in the chemotherapy-treated rats compared to controls, which may be related to the relatively high basal permeability of mannitol compared to other common low-permeability probes. We therefore suggest that future studies should focus on elucidating the complex interplay between chemotherapy in combination with luminal irritants on the intestinal permeability of other probes.

The use of chemical warfare agents is prohibited but they have been used in recent Middle Eastern conflicts. Their accidental exposure (e.g. arsenical lewisite) is also known and causes extensive painful cutaneous injury. However, their molecular pathogenesis is not understood. Here, we demonstrate that a nexus of stress granules (SGs), integrated stress, and RNA binding proteins (RBPs) Roquin and Reganse-1 play a key role. Lewisite and its prototype phenylarsine oxide (PAO) induce SG assembly in skin keratinocytes soon after exposure, which associate with various RBPs and translation-related proteins. SG disassembly was detected several hours after exposure. The dynamics of SG assembly-disassembly associates with the chemical insult and cell damage. Enhanced Roquin and Regnase-1 expression occurs when Roquin was recruited to SGs and Regnase-1 to the ribosome while in the disassembling SGs their expression is decreased with consequent induction of inflammatory mediators. SG-targeted protein translational control is regulated by the phosphorylation-dependent activation of eukaryotic initiation factors 2α (eIF2α). Treatment with integrated stress response inhibitor (ISRIB), which blocks eIF2α phosphorylation, impacted SG assembly dynamics. Topical application of ISRIB attenuated the inflammation and tissue disruption in PAO-challenged mice. Thus, the dynamic regulation of these pathways provides underpinning to cutaneous injury and identify translational therapeutic approach for these and similar debilitating chemicals.

Chemical disinfection of water provides significant public health benefits. However, disinfectants like chlorine can react with naturally occurring materials in the water to form disinfection byproducts (DBPs). Natural levels of iodine have been reported to be too low in some source waters to account for the levels of iodinated DBPs detected. Iodinated X-ray contrast media (ICM) have been identified as a potential source of iodine. The toxicological impact of ICM present in source water at the time of disinfection has not been fully investigated. Iopamidol, iohexol, iopromide, and diatrizoate are among the ICM most frequently detected in water. In this study, source water containing one of these four ICM was chlorinated; non-chlorinated ICM-containing water samples served as controls. Reactions were conducted at an ICM concentration of 5 µM and a chlorine dose of 100 µM over 72 hr. Water concentrates (20,000-fold) were prepared by XAD-resin/ethyl acetate extraction and DMSO solvent exchange. We used the MatTek® reconstituted human epithelial skin irritation model to evaluate the water concentrates and also assessed the dermal irritation and sensitization potential of these concentrates using the LLNA:BrdU ELISA in BALB/c mice. None of the water concentrates tested (2500X) resulted in a skin irritant response in the MatTek® skin irritation model. Likewise, none of the concentrates (2500X, 1250X, 625X, 312.5X, 156.25X) produced a skin irritation response in mice: erythema was minimal; the maximum increase in ear thickness was less than 25%. Importantly, none of the concentrates produced a positive threshold response for allergic skin sensitization at any concentration tested in the LLNA:BrdU ELISA. We conclude that concentrates of water disinfected in the presence of four different ICM did not cause significant skin irritation or effects consistent with skin sensitization at the concentrations tested., The authors thank Mr. Carey Copeland, Ms. Debbie Andrews, Mr. Tony McDonald, and Mr. Yusupha M. Sey for expert technical support. C.P. acknowledges support of Marie Curie IOF grant agreement 274379 funded by the European Union 7th R&D Framework Programme (FP7/2007-2013) and grant RYC2020-028901-I funded by MCIN/AEI/10.13039/501100011033 and “ESF investing in your future” . Drs. C. Weitekamp and YH Kim are acknowledged for their thoughtful and critical review of this manuscript.

This study developed an air-liquid interface (ALI) corneal model using explants bovine eyes for ocular toxicity assessment of ten chemicals and seven hair straightening mixtures. It was successfully maintained physiologically viable and normal for six days. Both eye damage (GHS cat. 1) and irritating (GHS cat. 2) chemicals induced corneal injury in our model. However, cat. 2 irritants triggered moderate damage when compared to cat. 1 agents, which induced a marked cytotoxicity profile. The mixtures were also able to trigger viability reduction associated with histopathological changes in the corneal tissues, especially when the exposure was via aerosol particles. Thus, the chemical exposure microenvironment simulation seemed to provide more reliable toxicological data. Moreover, mixture-induced corneal damage correlated with increased ROS levels, suggesting a close correlation between tissue death and oxidative stress. Besides mixtures showing the potential to induce moderate/mild ocular toxicity, we could verify that the corneal tissue damage showed reversibility due to the recovery from the injury after exposure to some of the mixtures. Hence, our ex vivo corneal model seems to be a simple and cost-effective approach for future studies related to further investigating the reversibility of damage in the cornea triggered by chemicals and their mixtures.

In vitro models are available as alternatives for the Draize eye irritation test. However, most of the alternative models are not quantitative nor designed to evaluate the effects of chemicals on the corneal barrier such as those encountered in pharmaceutical and cosmetic products. The objective of the present study was to investigate tissue electrical resistance to provide sensitive in vitro testing of tissue alteration caused by chemicals in pharmaceutical and cosmetic formulations as a potential eye irritation testing approach. The experimental protocols for effective tissue resistance measurements were examined using two in vitro eye models: porcine cornea and EpiCorneal. In these models, a test chemical was applied to the cornea or EpiCorneal tissue for 1 min, and tissue resistances/conductances were measured at 1-60 min after the application. The changes in conductance of the tissues after exposure to the chemicals were shown to provide quantitative evaluations to the influence of the chemicals. A correlation was found between the two in vitro models. The results suggest that these models can provide quantitative in vitro assessments of eye-irritating chemicals.

The approximation of euglycemia is the most effective means of preventing diabetic complications, which is achieved through effective insulin delivery. Recent reports indicate that insulin phenolic preservatives, which are found in all commercial insulin formulations, are cytotoxic, pro-inflammatory and induce secondary fibrosis. Therefore, we hypothesize that these preservatives induce an inflammatory response at the site of insulin infusion leading to diminished glycemic control and adverse pharmacokinetic outcomes. Insulin degradation by inflammatory cell proteases was quantitated following protease treatment in vitro. A modified murine air pouch model was utilized to evaluate the relative inflammatory responses following infusions of saline, insulin preservatives, and insulin, utilizing the adjuvant irritant thioglycolate. Blood glucose levels were monitored in diabetic mice with and without air pouch irritation. A pharmacokinetic analysis evaluated insulin effectiveness for diabetic mice between these two conditions. Inflammatory cells are significantly present in insulin preservative-induced inflammation, which effects diminished blood glucose control by both insulin uptake and degradation. Insulin containing these preservatives resulted in similar degrees of inflammation as observed with the irritant thioglycolate. These studies imply that the preservative agents found in commercial insulin formulations induce an intense localized inflammatory reaction. This inflammatory reaction may be responsible for the premature failure of insulin infusion devices. Future studies directed at reducing this inflammatory reaction may prove to be an important step in extending the lifespan of insulin infusion devices.

There is a global trend towards the development of physiologically relevant in vitro skin models to reduce or replace animal testing in the evaluation of therapeutic drug candidates. However, only commercial reconstructed human epidermis models (RHEm) have undergone formal validation. Although these commercial models are suitable for a wide range of applications, they are costly, lack flexibility, and the protocols used to generate them are not transparent. In this study, we present an open-source full-thickness skin model (FTSm) and assess its potential for drug testing. The FTSm was developed using endogenous extracellular matrix to recreate the dermal compartment, avoiding animal-derived hydrogels. An RHEm based on an open-source protocol was evaluated in parallel. The integrity of the skin barrier was analyzed by challenging the surface with detergents and measuring cell viability as well as by trans-epithelial electrical resistance (TEER) measurements. Skin irritation studies were performed based on OECD guidelines and complemented with an evaluation of the impact on the skin barrier by TEER measurement. The permeation of a dye through the developed models and a commercial membrane (Strat-M®) was compared using Franz diffusion cells and an infinite dose approach. The FTSm demonstrated structural and barrier properties comparable to native human skin. Although the RHEm showed a better performance in drug testing, the FTSm presented better barrier properties than commercial models as reported in the literature. These skin models can be a valuable contribution to accelerating the development and dissemination of alternatives to animal testing, avoiding the limitations of commercial models.

Fluorouracil (5-FU) is a widely used chemotherapeutic agent in various malignant tumors. However, intestinal toxicity is considered the irritant unavoidable adverse effect during the course therapy. The aim of the current study was to screen the effect of a new selective histamine receptor 1 blocker and platelet-activating factor (PAF) blocker on 5-FU induced intestinal toxicity. Five groups (6 rats each) of adult male rats (Wistar) were arranged as follows: (1) control group that was treated with carboxymethylcellulose, (2) a group that received rupatadine (higher dose) only, (3) a group that received 5-FU and (4) and (5) groups that received 5-FU plus lower or higher dose rupatadine, respectively. At end of the experiment, we determined intestinal malondialdehyde (MDA), glutathione reduced (GSH), nitric oxide (NO), tumor necrosis factor (TNF-α), interleukin 1β, 6, 10 (IL-1β, IL-6, IL-10), PAF, histamine, myeloperoxidase, cysteine-aspartic acid protease-3 (caspase-3), and nuclear factor kappa B (NF-κB) as well as the histological analysis. 5-FU injection caused marked elevation of MDA, NO, TNF-α, IL-1β, IL-6, PAF, histamine, myeloperoxidase, caspase-3, and NF-κB expressions. The intoxicated animals showed deficient GSH and IL-10 along with significant loss of villi, disorganized crypts, and inflammatory cell infiltration. Rupatadine pretreatment reduced the previously mentioned parameters, preserved a nearly normal intestinal mucosa picture with replenished GSH and elevated IL-10. In conclusion, rupatadine is a dual histamine receptor 1, and a PAF blocker could reduce 5-FU-induced oxidative damage, inflammation, apoptosis, and ulceration of the intestinal epithelium. Rupatadine may be a valuable modality to decrease 5-FU induced intestinal mucositis.

Oleanolic acid (OA) is a pentacyclic triterpenoid, abundantly found in plants of the Oleaceae family, and is well known for its beneficial pharmacological activities. Previously, we reported the inhibitory effect of OA on mast cell-mediated allergic inflammation. In this study, we investigated the effects of OA on atopic dermatitis (AD)-like skin lesions and its underlying mechanism of action. We evaluated the inhibitory effect of OA on AD-like responses and the possible mechanisms using a 1-chloro-2,4-dinitrochlorobenzene (DNCB)-induced AD animal model and tumor necrosis factor (TNF)-α/interferon (IFN)-γ-stimulated HaCaT keratinocytes. We found that OA has anti-atopic effects, including histological alterations, on DNCB-induced AD-like lesions in mice. Moreover, it suppressed the expression of Th2 type cytokines and chemokines in the AD mouse model and TNF-α/IFN-γ-induced HaCaT keratinocytes by blocking the activation of serine-threonine kinase Akt, nuclear factor-κB, and the signal transducer and activator of transcription 1. The results demonstrate that OA inhibits AD-like symptoms and regulates the inflammatory mediators, therefore, it may be used as an effective and attractive therapeutic agent for allergic disorders, such as AD. Moreover, the findings of this study provide novel insights into the potential pharmacological targets of OA for treating AD.

The Expert Panel for Cosmetic Ingredient Safety (Panel) reviewed the safety of inorganic hydroxides, which function in cosmetics primarily as pH adjusters. Representatives from the cosmetic industry have indicated these ingredients are used in depilating and hair waving/straightening formulations to raise pH values. The Panel considered relevant data related to these ingredients. The Panel concluded that these inorganic hydroxides are safe in hair straighteners and depilatories under conditions of recommended use; users should minimize skin contact. These ingredients are safe for all other present practices of use and concentration described in this safety assessment when formulated to be nonirritating.

Exposure to sensitizer has been suggested to be hazardous to human health, evaluation the sensitization of sensitizer is particularly important and urgently needed. Dendritic cells (DCs) exert an irreplaceable function in immunity, and the T cell receptor (TCR) repertoire is key to ensuring immune response to foreign antigens. We hypothesized that a co-culture model of human monocyte-derived dendritic cells (Mo-DCs) and T cells could be employed to evaluate the sensitization of DNCB. An experimental model of DNCB-induced sensitization in rat was employed to examine alterations of cluster of differentiation CD103+ DCs and T cells. A co-cultured model of Mo-DCs and T cells was developed in vitro to assess the sensitization of DNCB through the phenotypic and functional alterations of Mo-DCs, as well as the TCR repertoire. We found that the CD103+ DCs phenotype and T-helper (Th) cells polarization altered in sensitization rats. In vitro, phenotypic alteration of Mo-DCs caused by DNCB were consistent with in vivo results, antigen uptake capacity of Mo-DCs diminished and capacity of Mo-DCs to prime T cell increased. Clones of the TCR repertoire and the diversity of TCR repertoire were enhanced, changes were noted in the usage of variable, joining, and variable-joining gene combinations. DNCB exposure potentiated alterations and characteristics of Mo-DCs and the TCR repertoire in a co-culture model. Such changes provided innovative ideas for evaluating sensitization of DNCB.

Sulfur mustard (SM) is a notorious, bifunctional alkylating vesicant that was first used in warfare during World War I in 1917 and since then has been deployed in numerous skirmishes with its most recent documented use being during the Middle Eastern conflicts. Apart from its use in combat and terrorist activities, continual threat of accidental exposure from old stockpiles and improperly discarded munitions is ever present, especially to the innocent and unassuming civilian populations. SM can cause devastating injuries, depending on the dosage of SM exposure, route of exposure, as well as the physiological conditions of the individuals exposed. The most common routes of exposure are ocular, dermal, and exposure to the lungs and respiratory tissues through inhalation. Eyes are the most susceptible organ to SM-induced toxicities owing to their high moisture content and rapidly dividing cells. Additionally, ocular injury causes the most expeditious disablement of individuals even upon whole-body exposures. Therefore, it is imperative to understand the mechanisms underlying SM-induced ocular toxicity and design therapeutic interventions to prevent/mitigate ocular injuries. Ocular SM exposure may cause a wide range of symptoms such as inflammation, lacrimation, itching, dryness, photophobia, edema of the cornea/sclera/retina/iris, conjunctivitis, degradation of the corneal layer, fusion of two or more ocular layers, neovascularization, fibrosis, and temporary or permanent structural damage to one or more ocular layers. These symptoms may lead to vision impairments, resulting in partial or complete blindness that may be permanent. The highly toxic and exceedingly notorious nature of SM makes it a highly regulated chemical, requiring very expensive licensing, security, and safety requirements; thus, the more easily accessible analogue, nitrogen mustard (NM) that mimics SM-induced toxicity and injuries is employed in plethora of studies conducted in different animal models and culture systems. This review provides a comprehensive account of the injuries and symptoms that occur upon ocular SM exposures in human patients as well as studies in animal (in vivo, ex vivo) and cell (in vitro) models of SM and NM ocular exposures. Special emphasis has been laid on highlighting the strengths and lacunae in the research as well as the possible unexplored avenues of mechanisms underlying mustard-induced ocular injury that can be explored in future research endeavors. Furthermore, development of therapeutic interventions and targets of interest in the ocular system exposed to SM and NM, based on studies in human patients as well as in vivo, ex vivo, and in vitro models has been discussed in great depth, providing a valuable knowledge database to delineate pathways associated with vesicant-induced toxicity, and strategies/diagnostic tools against SM-induced toxicity.

Introduction Extravasation injuries are one of the most feared complications of intravenous drug administration. The most common drugs associated with extravasation injury include chemotherapy agents and contrast media. Natural course of vesicant extravasation is discomfort, pain, swelling, inflammation, and ultimately skin ulceration. While diligence is the principle approach in prevention, immediate bed-side measures are as important in controlling the extent of tissue damage. Various options, either medical or interventional are next steps in treatment of the condition including antidotes, volume dilution, flushing, suction, hyperbaric oxygen therapy, and surgery. Materials and methods 12 male Wistar albino rats were divided into two groups; one group received fat injections following subdermal doxorubicin infiltration in their right thighs, while other group received saline injection following subdermal doxorubicin infiltration in their right thighs for dilution. Left thighs of both groups were left untreated following subdermal doxorubicin infiltration. Total area of necrosis, as well as resultant epidermal thicknesses were assessed. Histological analyses were conducted using modified Verhofstad scoring system for comparison. Results Mean necrotic area was significantly smaller in the fat injection group compared to other groups. Median Verhofstad score was lesser in the fat injection group as well. Median epidermal thickness, on the other hand, was greater in the fat injection group. Conclusion Injection of fat grafts following vesicant extravasation might be beneficial in preventing the progression of tissue damage, if employed early.

The Expert Panel for Cosmetic Ingredient Safety (Panel) assessed the safety of Phosphoric Acid and its salts (31 ingredients), which are reported to function as buffering agents, corrosion inhibitors, chelating agents, and pH adjusters in cosmetic products. The Panel reviewed data relating to the safety of these ingredients and concluded that Phosphoric Acid and its salts are safe in the present practices of use and concentration in cosmetics when formulated to be nonirritating.

The Expert Panel for Cosmetic Ingredient Safety (Panel) reopened the safety assessment of Sodium Sulfate, a cosmetic ingredient that is an inorganic salt reported to function in cosmetics as a viscosity increasing agent—aqueous. The Panel reviewed the relevant new data for the ingredient, including frequency of use and concentration of use, and considered data from the previous Panel assessment. The Panel concluded that Sodium Sulfate is safe in cosmetics in the present practices of use and concentrations described in this safety assessment when formulated to be nonirritating.

NF-erythroid 2–related factor 2 (Nrf2) is a major transcription factor to protect cells against reactive oxygen species (ROS) and reactive toxicants. Meanwhile, Nrf2 can inhibit contact dermatitis through redox-dependent and -independent pathways. However, the underlying mechanisms of how Nrf2 mediates irritant contact dermatitis (ICD) are still unclear. In this article, we elucidated the role of Nrf2 in 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced acute ICD. Our study demonstrated that the ear thickness, redness, swelling, and neutrophil infiltration were significantly increased, accompanied by increased expression of inflammatory cytokines (IL-1α, IL-1β, IL-6, etc.) and decreased expression of antioxidant genes (HO-1 and NQO1) in Nrf2 knockout mice. Moreover, ERK phosphorylation was elevated in mouse embryonic fibroblasts (MEFs) from Nrf2 knockout mouse. Inhibition of ERK significantly alleviated TPA-induced cutaneous inflammation and ROS accumulation in MEFs derived from mouse. Conversely, ROS scavenging inhibited the ERK activation and TPA-induced inflammation in MEFs. Taken together, the findings illustrate the key role of the Nrf2/ROS/ERK signaling pathway in TPA-induced acute ICD.

The toxic effects resulting from inhalation exposure depend on both the concentration (C) of the inhaled substance and the exposure duration (t), including the assumptions that the exposure-limiting toxic effect is linearly linked with the accumulated C × t (inhaled dose), and detoxification or compensatory responses diminishing this dose are negligible. This interrelationship applies for both constant and fluctuating concentrations and is usually expressed by the toxic load equation Cn × t = constant effect (k). The toxic load exponent ‘n’ is derived from both C- and t-dependent exponents with Cb2×tb3 = k with n = b2/b3. This model is taken as a fundamental basis for assessing the acute hazard posed by atmospheric releases of noxious substances, whether deliberate or accidental. Despite its universal use, especially for inhaled irritants, the toxicological significance of this mathematical construct is still discussed controversially. With n = 1 this equation is called Haber’s rule. The underlying assumption is that the exposure-based calculated and the actually inhaled Cb2×tb3 are identical. Unlike the calculated dose, the latter is dependent on the test species and its t-dependent change in respiratory minute volume (MV). The retention patterns of inhaled irritant vapors may differ in obligate nasal breathing rodents and oronasally breathing humans as well. Thus, due to the interdependence of n on both C, t and k, this mathematical construct generates a bioassay-specific ‘n’ which can hardly be considered as human-equivalent, especially following exposure to sensory irritants known to elicit reflex-related changes in MV. The C- and t-dependent impact on Cn × t = k was analyzed with the sensory irritant n-butyl monoisocyanate and compared with t-dependent changes elicited by highly, moderately, and poorly water-soluble sensory irritants ammonia, toluene diisocyanate, and phosgene, respectively. This comparison reveals that n depends on several factors: In cases where MV is instantly and plateau-like depressed with onset of exposure, n appears to be most dependent on Cb2 × MV whereas for a similar slower time-dependent response n becomes more dependent on MV × tb3. For any ensuing risk characterization that focuses on acute non-lethal threshold Cb2 × tb3’s, the sensory irritation-related depression in MV must be known to arrive at meaningful conclusions. In summary, both Cn- and t-dependent dosimetry-related pitfalls may occur in acute bioassays on rodents following inhalation exposure to irritants. These must be identified and dealt with judiciously prior to translation to apparently similar human exposures. By default, extrapolations from one duration to another should start with that Cn × t eliciting the least depression in MV with n = 1.

In Ukraine Globally Harmonized System of classification of chemicals has not been implemented yet. In this article we analyze differences between GHS/CLP classification systems and Hygienic Classification of Pesticides by the Degree of Hazard currently in force in Ukraine in respect of approach and criteria for classification of effects on skin. As a case study, we conducted in silico modelling of herbicide imazamox using skin irritation/corrosion modules of ToxTree. The prediction of ToxTree was “Not Corrosive to skin”. Then skin irritation and skin corrosion in vitro tests (OECD TGs 439, 431) were conducted. Classification of this substance based on in vitro and in vivo results according to GHS/CLP was the same, while it was not possible based on in vitro results to assign certain hazard class of Ukrainian classification due to difference in its and GHS/CLP criteria. However, ongoing process of harmonization of Ukrainian legislation with EU will give opportunity not only use alternative methods, but also adopt most recent advances and incorporate data from non-animal methods directly into classification criteria.

The use of lower cut-off values/concentration limits for the calculation of mixture classification in UN GHS/EU CLP versus the previous regulatory scheme (EU Dangerous Preparations Directive, DPD), has resulted in an increased number of classifications in the highest eye hazard category. Herein, a semi-quantitative categorisation of severity of eye effects, following accidental human exposures to detergents, was compared to the classification category of the products. Three schemes were evaluated: EU DPD; EU CLP (based on all available data and information, including weight of evidence); and EU CLP (based entirely on the calculation method). As reported by four EU Poison Centres, the vast majority of exposures had caused minor or no symptoms. Classification was a poor predictor of effects in man subjected to accidental exposure. Note however that this is also because effects are not only driven by the intrinsic hazard (as reflected in the classification), but also by the exposure conditions and mitigation (i.e. rinsing). EU CLP classification using all available data and information was more predictive of medically relevant symptoms than the EU CLP calculation method. The latter led to a poorer differentiation between irritating products versus products potentially causing serious eye damage.

Irritant Contact Dermatitis (ICD) is the most common occupational skin disorder. During ICD, keratinocytes initiate the inflammatory cascade by producing cytokines including IL-6. This laboratory previously reported that IL-6 deficiency exacerbates skin inflammation during ICD, yet the role of the IL-6Rα in keratinocyte function has yet to be elucidated. To investigate how IL-6Rα function in keratinocytes influences the inflammatory response during ICD, keratinocyte-specific IL-6Rα KO (IL6raΔker) and WT mice were exposed to two well-known occupational irritants; JP-8 jet fuel, and benzalkonium chloride (BKC), or acetone control for three days. Dermatitis lesions were collected and flow cytometric and immunohistochemical analyses revealed that IL6raΔker skin displayed increased populations of CD11b+CD45+ and F4/80+ cells respectively relative to WT. However, IL6raΔker mouse skin contained reduced numbers of γδ T cells relative to WT. Furthermore, IL6raΔker skin expressed increased levels of pro-inflammatory cytokines including IL-1β, IL-22, and CCL4 but decreased levels of anti-inflammatory cytokines IL-4 and IL-10. These results indicate that epidermal keratinocyte IL-6Rα function modulates epidermal hyperplasia, immune cell infiltration into skin and cytokine expression during ICD and suggests that the previously reported protective effect of IL-6 during ICD might be mediated primarily by keratinocyte derived IL-6Rα.

Researchers from the Ramazzini Institute have reported that lifespan dosing of rats with aspartame treatment is associated with an increased overall incidence of malignant tumors, including leukemias/lymphomas, transitional cell carcinomas of the renal pelvis/ureter, and malignant schwannomas of the peripheral nerves. Other carcinogenicity studies conducted on aspartame have shown no such carcinogenic potential in any organ system. Additional data to assess the carcinogenic potential of aspartame, especially in relation to the publications of the Ramazzini Institute, were obtained from a third-party histological evaluation of tissues from a carcinogenicity study previously conducted to assess the potential for aspartame to induce tumors of the brain. The results of this histological evaluation provide no evidence of a tumorigenic effect of aspartame in any organ group, including those organs/tissues reportedly affected in the Ramazzini Institute's studies. The only effects identified were an increased incidence of renal pelvic mineralization and renal pelvic hyperplasia secondary to the irritant properties of the mineralization process. The toxicological significance of these particular findings is widely considered minimal. There is no evidence that aspartame is carcinogenic in rats, at least to doses of 4 g/kg body weight/day administered over a 2-year period.

Objective To evaluate the efficacy of the non-steroidal anti-inflammatory drug, meloxicam, in alleviating pain and inflammation and on production-related variables in a model of sterile acute inflammation in sheep. Methods Groups of 12 mature Merino ewes received 0, 0.5, 1.0 or 1.5 mg/kg meloxicam subcutaneously 90 min before injection of 0.1 mL turpentine subcutaneously on the anterior aspect of the proximal phalanx of a forelimb. Pain- and inflammation-related variables were assessed at -18, 3, 6, 9, 12, 24, 48 and 72 h relative to meloxicam administration. Daily feed intake and body weight change 7 days later were also assessed. Pain-related variables measured were weight borne on each forelimb, lameness score, time each forelimb was raised in a 20-s interval and tolerance to a noxious mechanical stimulus. Inflammation-related variables measured were skin temperature, limb circumference, body temperature, plasma haptoglobin concentration and peripheral blood leucocyte parameters. Results Meloxicam was effective in improving all pain-related variables. A dose-dependent response was seen between 0 and 1.0 mg/kg, with no additional benefit provided by 1.5 mg/kg. At a dose rate of 1.0 mg/kg, meloxicam improved weight borne on the turpentine-treated limb by 14%, reduced the time the treated limb was held in a non-weight-bearing posture by 46%, reduced the lameness score by 58% and improved tolerance to pressure by 52%. No significant effects of meloxicam on inflammatory variables or appetite were observed. Conclusions Using a validated pain model, the data suggested that 1.0 mg/kg meloxicam provided significant analgesic benefits to sheep.

Sulfur mustard (HD) is a highly toxic vesicant and is prohibited by the Organisation for the Prohibition of Chemical Weapons (OPCW). HD can modify human serum albumin (HSA) to generate hydroxyethylthioethyl (HETE) adducts, which could be utilized as biomarkers for verifying HD exposure in forensic analysis. Here, five amino acid adducts generated from pronase digestion of HD-exposed human serum albumin (HD-HSA) in plasma were selected as biomarkers to retrospectively detect HD exposure. HD-HSA was precipitated from plasma with acetone, digested by pronase, derivatized with propionic anhydride (PA), and analysed with ultra-high performance liquid chromatography coupled with triple quadrupole mass spectrometry (UHPLC-TQ MS). The limits of detection (LODs) and limits of quantification (LOQs) of the HD exposure concentrations were evaluated as 1.00 ng/mL at S/N≥3 and 3.00 ng/mL at S/N≥10, respectively, which are approximately 60 times lower than those of the reported method. The approach shows good linearity (R