BACKGROUND: Nowadays, companion and working dogs hold significant social and economic importance. Dry eye, also known as dry keratoconjunctivitis (KCS), a common disease in ophthalmology, can readily impact a dog\'s working capacity and lead to economic losses. Although there are several medications available for this disease, all of them only improve the symptoms on the surface of the eye, and they are irritating and not easy to use for long periods of time. Adipose-derived mesenchymal stem cells (ADMSC) are promising candidates for tissue regeneration and disease treatment. However, long-term in vitro passaging leads to stemness loss of ADMSC. Here, we aimed to use ADMSC overexpressing Secreted Protein Acidic and Rich in Cysteine (SPARC) to treat 0.25% benzalkonium chloride-treated dogs with dry eye to verify its efficacy. For in vitro validation, we induced corneal epithelial cell (HCECs) damage using 1 µg/mL benzalkonium chloride.
METHODS: Fifteen male crossbred dogs were randomly divided into five groups: normal, dry eye self-healing control, cyclosporine-treated, ADMSC-CMV-treated and ADMSC-OESPARC-treated. HCECs were divided into four groups: normal control group, untreated model group, ADMSC-CMV supernatant culture group and ADMSC-OESRARC supernatant culture group.
RESULTS: SPARC-modified ADMSC had the most significant effect on canine ocular surface inflammation, corneal injury, and tear recovery, and the addition of ADMSC-OESPARC cell supernatant also had a salvage effect on HCECs cellular damage, such as cell viability and cell proliferation ability. Moreover, analysis of the co-transcriptome sequencing data showed that SPARC could promote corneal epithelial cell repair by enhancing the in vitro viability, migration and proliferation and immunosuppression of ADMSC.
CONCLUSIONS: The in vitro cell test and in vivo model totally suggest that the combination of SPARC and ADMSC has a promising future in novel dry eye therapy.

Melatonin (Mel) is a phytohormone that plays a crucial role in various plant processes, including stress response. Despite numerous studies on the role of Mel in stress resistance, its significance in plants exposed to benzalkonium chloride (BAC) pollution remains unexplored. BAC, a common antiseptic, poses a threat to terrestrial plants due to its widespread use and inefficient removal, leading to elevated concentrations in the environment. This study investigated the impact of BAC (0.5 mg L-1) pollution on wild-type Col-0 and snat2 knockout mutant Arabidopsis lines, revealing reduced growth, altered water relations, and gas exchange parameters. On the other hand, exogenous Mel (100 μM) treatments mitigated BAC-induced phytotoxicity and increased the growth rate by 1.8-fold in Col-0 and 2-fold in snat2 plants. snat2 mutant seedlings had a suppressed carbon assimilation rate (A) under normal conditions, but BAC contamination led to further A repression by 71% and 48% in Col-0 and snat2 leaves, respectively. However, Mel treatment on stressed plants was successful in improving Fv/Fm and increased the total photosynthesis efficiency by regulating photochemical reactions. Excessive H2O2 accumulation in the guard cells of plants exposed to BAC pollution was detected by confocal microscopy. Mel treatments triggered almost all antioxidant enzyme activities (except POX) in both Arabidopsis lines under stress. This enhanced antioxidant activity, facilitated by foliar Mel application, contributed to the alleviation of oxidative damage, regulation of photosynthesis reactions, and promotion of plant growth in Arabidopsis. In addition to corroborating results observed in many agricultural plants regarding the development of tolerance to environmental stresses, this study provides novel insights into the action mechanisms of Mel under the emerging pollutant benzalkonium chloride.

Aquatic ecosystems and their communities are exposed to numerous stressors of various natures (chemical and physical), whose impacts are often poorly documented. In urban areas, the use of biocides such as dodecyldimethylbenzylammonium chloride (DDBAC) and their subsequent release in wastewater result in their transfer to urban aquatic ecosystems. DDBAC is known to be toxic to most aquatic organisms. Artificial light at night (ALAN) is another stressor that is increasing globally, especially in urban areas. ALAN may have a negative impact on photosynthetic cycles of periphytic biofilms, which in turn may result in changes in their metabolic functioning. Moreover, studies suggest that exposure to artificial light could increase the biocidal effect of DDBAC on biofilms. The present study investigates the individual and combined effects of DDBAC and/or ALAN on the functioning and structure of photosynthetic biofilms. We exposed biofilms in artificial channels to a nominal concentration of 30 mg.L-1 of DDBAC and/or ALAN for 10 days. ALAN modified DDBAC exposure, decreasing concentrations in the water but not accumulation in biofilms. DDBAC had negative impacts on biofilm functioning and structure. Photosynthetic activity was inhibited by > 90% after 2 days of exposure, compared to the controls, and did not recover over the duration of the experiment. Biofilm composition was also impacted, with a marked decrease in green algae and the disappearance of microfauna under DDBAC exposure. The integrity of algal cells was compromised where DDBAC exposure altered the chloroplasts and chlorophyll content. Impacts on autotrophs were also observed through a shift in lipid profiles, in particular a strong decrease in glycolipid content was noted. We found no significant interactive effect of ALAN and DDBAC on the studied endpoints.

Flavonoids are considered as health-protecting food constituents. The testing of their biological effects is however hampered by their low oral absorption and complex metabolism. In order to investigate the direct effect(s) of unmetabolized flavonoid, a preparation in a biologically friendly solvent for intravenous administration is needed. Isorhamnetin, a natural flavonoid and a human metabolite of the most frequently tested flavonoid quercetin, has very low water solubility (<3.5 μg/mL). The aim of this study was to improve its solubility to enable intravenous administration and to test its pharmacokinetics in an animal model. By using polyvinylpyrrolidone (PVP10) and benzalkonium chloride, we were able to improve the solubility approximately 600 times to 2.1 mg/mL. This solution was then administered intravenously at a dose of 0.5 mg/kg of isorhamnetin to rats and its pharmacokinetics was analyzed. The pharmacokinetics of isorhamnetin corresponded to two compartmental model with a rapid initial distribution phase (t1/2α: 5.7 ± 4.3 min) and a slower elimination phase (t1/2β: 61 ± 47.5 min). Two sulfate metabolites were also identified. PVP10 and benzalkonium did not modify the properties of isorhamnetin (iron chelation and reduction, and cell penetration) substantially. In conclusion, the novel preparation reported in this study is suitable for future testing of isorhamnetin effects under in vivo conditions.

UNASSIGNED: Dry eye disease (DED) is a multifactorial ocular surface disease with a rising incidence. Therefore, it is urgent to construct a reliable and efficient drug delivery system for DED treatment.
UNASSIGNED: In this work, we loaded C-dots nanozyme into a thermosensitive in situ gel to create C-dots@Gel, presenting a promising composite ocular drug delivery system to manage DED.
UNASSIGNED: This composite ocular drug delivery system (C-dots@Gel) demonstrated the ability to enhance adherence to the corneal surface and extend the ocular surface retention time, thereby enhancing bioavailability. Furthermore, no discernible ocular surface irritation or systemic toxicity was observed. In the DED mouse model induced by benzalkonium chloride (BAC), it was verified that C-dots@Gel effectively mitigated DED by stabilizing the tear film, prolonging tear secretion, repairing corneal surface damage, and augmenting the population of conjunctival goblet cells.
UNASSIGNED: Compared to conventional dosage forms (C-dots), the C-dots@Gel could prolong exhibited enhanced retention time on the ocular surface and increased bioavailability, resulting in a satisfactory therapeutic outcome for DED.

Traditional experimental methodologies suffer from a few limitations in the toxicological evaluation of the preservatives added to eye drops. In this study, we overcame these limitations by using a microfluidic device. We developed a microfluidic system featuring a gradient concentration generator for preservative dosage control with microvalves and micropumps, automatically regulated by a programmable Arduino board. This system facilitated the simultaneous toxicological evaluation of human corneal epithelial cells against eight different concentrations of preservatives, allowing for quadruplicate experiments in a single run. In our study, the IC50 values for healthy eyes and those affected with dry eyes syndrome showed an approximately twofold difference. This variation is likely attributable to the duration for which the preservative remained in contact with corneal cells before being washed off by the medium, suggesting the significance of exposure time in the cytotoxic effect of preservatives. Our microfluidic system, automated by Arduino, simulated healthy and dry eye environments to study benzalkonium chloride toxicity and revealed significant differences in cell viability, with IC50 values of 0.0033% for healthy eyes and 0.0017% for dry eyes. In summary, we implemented the pinch-to-zoom feature of an electronic tablet in our microfluidic system, offering innovative alternatives for eye research.

Benzalkonium chloride (BAC) is a disinfectant with broad-spectrum antibacterial properties, yet despite its widespread use and detection in the environment, the effects of BAC exposure on microorganisms remain poorly documented. Herein, the impacts of BAC on a Pseudomonas aeruginosa strain Jade-X were systematically investigated. The results demonstrated that the minimum inhibitory concentration (MIC) of BAC against strain Jade-X was 64 mg L-1. Exposure to BAC concentrations of 8, 16, 32, and 64 mg L-1 significantly augmented biofilm formation by 2.03-, 2.43-, 2.96-, and 2.56-fold respectively. The swimming and twitching abilities, along with the virulence factor production, were inhibited. Consistently, quantitative reverse transcription PCR assays revealed significant downregulation of genes related to flagellate- and pili-mediated motilities (flgD, flgE, pilB, pilQ, and motB), as well as phzA and phzB genes involved in pyocyanin production. The results of disk diffusion and MIC assays demonstrated that BAC decreased the antibiotic susceptibility of ciprofloxacin, levofloxacin, norfloxacin, and tetracycline. Conversely, an opposite trend was observed for polymyxin B and ceftriaxone. Genomic analysis revealed that strain Jade-X harbored eleven resistance-nodulation-cell division efflux pumps, with mexCD-oprJ exhibiting significant upregulation while mexEF-oprN and mexGHI-opmD were downregulated. In addition, the quorum sensing-related regulators LasR and RhlR were also suppressed, implying that BAC might modulate the physiological and biochemical behaviors of strain Jade-X by attenuating the quorum sensing system. This study enhances our understanding of interactions between BAC and P. aeruginosa, providing valuable insights to guide the regulation and rational use of BAC.

This experiment aimed to investigate the feasibility of cytisine (CYT) in treating eye diseases with ocular topical application. An in vitro cytotoxicity test, a hen\'s egg test-chorioallantoic membrane (HET-CAM), and a mouse eye tolerance test were used to fully reveal the ocular safety profiles of CYT. For the efficacy evaluations, CYT\'s effects on cell wound healing, against H2O2-induced oxidative stress damages on cells, and on benzalkonium chloride (BAC)-induced dry eye disease (DED) in mice were evaluated. Results showed that CYT did not show any cytotoxicities at concentrations no higher than 250 μg/ml, while lipoic acid (α-LA) at 250 μg/ml and BAC at 1.25 μg/ml showed significant cytotoxicities within 48 h incubation. The HET-CAM and mouse eye tolerance test confirmed that 0.5% CYT eye drops demonstrated good safety characteristics. Efficacy evaluations showed that CTY significantly promoted cell migration and wound healing. CYT significantly improved cell survival against H2O2-induced oxidative stress damage by reversing the imbalance between the reactive oxygen species (ROS) and antioxidant defense mechanisms. The animal evaluation of the BAC-induced dry eye model revealed that CYT demonstrated a strong treatment effect, including reversing ocular surface damages, recovering corneal sensitivity, and inhibiting neovascularization; HMGB1/NF-κB signaling was involved in this DED treatment by CTY. In conclusion, CYT had strong experimental treatment efficacy against DED with good ocular safety profiles, and it might be a novel and promising drug for DED.

BACKGROUND: Benzalkonium chloride (BAC) is a quaternary ammonium compound (QAC), that can be found in a wide variety of household products-from disinfectants to medicaments and home fragrances-but also professional products. In pets, cats have long been reported as more sensitive than dogs to QACs; in fact, signs of irritation such as oral ulcerations, stomatitis and pharyngitis can be observed after contact with concentrations of 2% or lower. In a review of 245 cases of BAC exposure in cats, reported by the Veterinary Poisons Information Service (United Kingdom) only 1.2% of the cases died or were euthanized. Nevertheless, BAC toxidromes in cats can result in transitory CNS and respiratory distress, as well as severe mucosal and cutaneous lesions. Currently, only a few reports are available concerning BAC poisoning in this species.
METHODS: A 4 month-old kitten presented with severe glossitis, lameness in the hindlimbs and episodes of vomiting and diarrhoea. The cause was unknown until the owners reported use of a BAC-containing mould remover (5%) 4 days later. The patient developed severe oral burns requiring a pharyngeal tube for feeding and severe cutaneous chemical burns. The kitten was managed with supportive therapy and required hospitalization for 10 days. The symptoms disappeared completely 3 weeks after exposure.
CONCLUSIONS: BAC is a very common compound contained in several household and professional products but, to the best of our knowledge, no previous case had been reported in Italy. We hope that this report will help raise awareness on the hazards of BAC products for cats in both domestic and work contexts.

Visomitin eye drops are the first and, so far, the only drug based on SkQ1 - the mitochondria-targeted antioxidant 10-(6\'-plastoquinonyl) decyltriphenylphosphonium, developed in the laboratories of Moscow State University under the leadership of Academician V. P. Skulachev. SkQ1 is considered as a potential tool to combat the aging program. We have previously shown that it is able to prevent and/or suppress development of all manifestations of accelerated senescence in OXYS rats, including retinopathy, similar to the age-related macular degeneration (AMD). Here, we assessed the effect of Visomitin instillations on progression of the AMD-like pathology and p38 MAPK and ERK1/2 activity in the OXYS rat retina (from the age of 9 to 12 months). Wistar and OXYS rats treated with placebo (composition identical to Visomitin with the exception of SkQ1) were used as controls. Ophthalmological examination showed that in the OXYS rats receiving placebo, retinopathy progressed and severity of clinical manifestations did not differ from the intact OXYS rats. Visomitin suppressed progression of the AMD-like pathology in the OXYS rats and significantly improved structural and functional parameters of the retinal pigment epithelium cells and state of microcirculation in the choroid, which, presumably, contributed to preservation of photoreceptors, associative and ganglion neurons. It was found that the activity of p38 MAPK and ERK1/2 in the retina of 12-month-old OXYS rats is higher than that of the Wistar rats of the same age, as indicated by the increased content of phosphorylated forms of p38 MAPK and ERK1/2 and their target protein tau (at position T181 and S396). Visomitin decreased phosphorylation of p38 MAPK, ERK1/2, and tau indicating suppression of activity of these MAPK signaling cascades. Thus, Visomitin eye drops are able to suppress progression of the AMD-like pathology in the OXYS rats and their effect is associated with the decrease in activity of the MAPK signaling cascades.