Cosmetics containing rhododendrol (RD) were voluntarily recalled after incidents of leukoderma related to their use. Users reported using up to five different RD-containing products by layered application. In this study, we investigated the effects of layered application, formulations, and their components on the skin permeation of cosmetics containing RD. Experiments were designed to simulate actual in-use conditions, such as varying application volumes, physical mixing of formulations, sequence of cosmetics application and time interval between applications, to establish their effect on the skin permeation of RD. Milk and lotion RD-containing cosmetics (2%), 1% aqueous RD, and preparations of formulation components were applied as the first or second layers as finite doses of 10 or 20 µL/cm². Permeation experiments were performed through excised porcine ear skin using Franz diffusion cells with an effective diffusion area of 1.77 cm². Cosmetics applied by layered application exhibited lower skin permeation of RD compared with a single application despite having the same application dose. High initial volume (20 µL at 0 or 5 sec) did not exhibit any significant reduction in the permeation of RD. Formulations and their components caused varying reductions in RD permeation, probably due to changes in thermodynamic activity of the active component. Layered application, formulation components, application volume, time interval and sequence of application had significant influences on the skin permeation of the active component. Moreover, this study established a method of investigating the influence of formulations and their components on the skin permeation of actives after layered application.

Cosmetic ingredients are often complex mixtures from natural sources such as botanical extracts that might contain minute amounts of constituents with sensitizing potential. The sensitivity of in vitro skin sensitization test methods such as KeratinoSens^TM and h-CLAT for the detection of minute amounts of sensitizer in mixtures remains unclear. In this study, we assessed the detection sensitivity of the binary test battery comprising KeratinoSens^TM and h-CLAT for minute amounts of sensitizers by comparing the LLNA EC3 (estimated concentration of a substance expected to produce a stimulation index of 3) values to the minimum detection concentrations (MDCs) exceeding the positive criteria for each of the two in vitro test methods. 146 sensitizers with both sets of in vitro data and LLNA data were used. MDC values for KeratinoSens^TM and h-CLAT were calculated from exposure concentrations exceeding positive criteria for each in vitro test method (EC1.5 and minimum induction thresholds, respectively). The dilution rate used to expose culture medium was also considered. For 86% of analyzed sensitizers, the in vitro test methods showed MDC values lower than LLNA EC3 values, suggesting that the binary test battery with KeratinoSens^TM and h-CLAT have greater sensitivity for detection of minute amounts of sensitizer than LLNA. These results suggest the high applicability of KeratinoSens^TM and h-CLAT for detecting skin sensitizing constituents present in botanical extract.

Cosmetic ingredients often comprise complex mixtures, such as botanical extracts, which may contain skin sensitizing constituents. In our previous study for the sensitivity of the evaluations of skin sensitizing constituents in mixtures using the binary in vitro test battery with KeratinoSens^TM and h-CLAT, some sensitizers showed higher detection limits in in vitro test methods than in murine local lymph node assays (LLNA). Thus, to minimize the uncertainty associated with decreased sensitivity for these sensitizers, a risk assessment strategy was developed for mixtures with negative results from the binary test battery. Assuming that the no expected sensitization induction level of mixtures (mixture NESIL) can be derived for mixtures with negative in vitro test results, we assessed 146 sensitizers with in vitro and LLNA data according to the assumption of indeterminate constituents in mixtures. Finally, we calculated 95th percentile probabilities of mixture NESILs and derived dermal sensitization thresholds for mixtures (mixture DST) with negative in vitro test results of 6010 μg/cm². Feasibility studies indicated that this approach was practical for risk assessments of products in the cosmetic industry. This approach would be a novel risk assessment strategy for incorporating the DST approach and information from in vitro test methods.

Due to its excellent properties such as ultraviolet obscuration, chemical stability and small particle size, nano-titanium dioxide (nano-TiO₂) is widely used, particularly in sunblock products. The skin is therefore a chief route for exposure. Studies have found that oral or respiratory exposure to nano-TiO₂ has an adverse impact on the cardiovascular system. The relationship between dermal exposure to nano-TiO₂ and cardiovascular system toxicity, particularly the causative mechanisms, remain unclear. In this study, we used Balb/c mice to evaluate cardiovascular toxicity from nano-TiO₂ dermal exposure, and the underlying mechanisms associated with the human umbilical vein endothelial cells (HUVECs) were explored. Our results showed that nano-TiO₂ treatment resulted in an obvious increase in reactive oxygen species and 8-hydroxy-2’-deoxyguanosine, indicating the appearance of oxidative stress. Moreover, the levels of inflammatory biomarkers, such as immunoglobulin E, soluble intercellular adhesion molecule-1, interleukin-8, and hypersensitive C-reactive protein, also increased. Exposing HUVECs to nano-TiO₂ led to a decline in cell vitality, and an increase in caspase-3 levels, suggesting that nano-TiO₂ exposure caused cytotoxicity and even cell apoptosis. Interestingly, neutralizing oxidative stress by administering Vitamin E was shown to reduce the inflammatory response and cytotoxicity. Our findings suggest that nano-TiO₂ can injure the cardiovascular system via dermal exposure, and does this via oxidative stress-induced inflammation and cytotoxicity. Vitamin E treatment may be a strategy to mitigate the damage.

Galectin-3 (Gal-3) is a multifunctional carbohydrate-binding protein associated with cell migration, cell proliferation, cell adhesion, and cell-cell interaction in tumor cells. It has been implied to be involved in the tumor progression and chemoresistance of epithelial ovarian cancer (EOC). However, it is unclear whether the Gal-3-mediated regulation on the EOC chemosensitivity is associated with hypoxia or mitochondrial dysfunction. In the present study, we examined the regulation by Gal-3 overexpression on cisplatin-sensitivity or cisplatin-resistance in EOC cells in vitro. We manipulated Gal-3 via plasmid transfection and RNA interference in the cisplatin-resistant EOC cells, and re-evaluated the sensitivity of the cisplatin-resistant EOC cells to cisplatin, with CCK-8 assay, colony forming assay, apoptosis analysis and mitochondrial function examination. Results demonstrated that galectin-3 overexpression downregulated the cisplatin sensitivity in EOC OVCAR-3 cell clone, resulting in an upregulated growth and a reduced apoptosis in the cisplatin-treated OVCAR-3 cells. On the other hand, the Gal-3 knockdown with Gal-3-specific siRNA transfection aggravated cisplatin-induced apoptosis in OVCAR-3 cells. In conclusion, Galectin-3 reduces the sensitivity of ovarian cancer cells to cisplatin via regulating cisplatin-induced mitochondrial dysfunction. Galectin-3 knockdown inhibits the chemo-resistance of EOC cells. It implies that Galectin-3 might be a potential target to overcome the chemo-resistance in EOC cells.

Cisplatin (CP) is an anti-cancer drug used for treatment of solid tumors, but the major adverse effect is drug-induced nephrotoxicity. The current study aimed to determine biomarkers that might predict nephrotoxicity induced by CP using serum or urinary proton nuclear magnetic resonance (¹H NMR) spectral data in male Sprague-Dawley (S-D) rats. CP (0, 0.5 or 5 mg/kg) was intraperitoneally (i.p.) administered for single dose. Animals were sacrificed 2 days (D2) or 8 days (D8) after administration of CP in order to perform analysis of serum biochemistries and histopathologic examination. Urine samples were collected every 24 hr from pre-treatment to sacrifice. Serum and urinary ¹H NMR spectral data revealed apparent differential clustering between control and CP-treated groups as evidenced by principal component analysis (PCA) and orthogonal projections to latent structures-discriminant analysis (OPLS-DA) in global and targeted profiling. The concentrations of endogenous serum metabolites, alanine, betaine, glucose, glutamine, lactate, and leucine were significantly increased on D2. Urinary concentrations of alanine, glucose, glycine, guanidinoacetate, acetate, and lactate were significantly elevated on D2 or D8, whereas concentrations of urinary metabolites, citrate and hippurate were significantly decreased on D2 or D8. The correlation of serum and urinary ¹H NMR OPLS-DA with serum biochemistry and renal histopathologic changes suggests that ¹H NMR urinalysis may be used to reliably predict or screen CP-induced nephrotoxicity. Data suggest that these altered endogenous metabolites might serve as specific biomarkers for CP-induced nephrotoxicity.