Bioaccumulation of functionalized polystyrene nanoplastics in sea cucumber Apostichopus japonicus (Selenka, 1867) and their toxic effects on oxidative stress, energy metabolism and mitochondrial pathway.

Micro/nano-plastics (M/NPs) are emerging contaminants in aquatic environment, however, little knowledge regarding the adverse effects of functionalized NPs has been documented so far. This study investigated the accumulation of different polystyrene nanoplastics (PS-NPs, i.e., plain PS, carboxyl-functional PS-COOH and amino-functional PS-NH2) at two particle sizes of 100 nm and 200 nm, and evaluated the impacts on oxidative stress, energy metabolism and mitochondrial pathway responses in intestine and respiratory tree of Apostichopus japonicus during the 20-d exposure experiment. The results showed that there were significant interactions of particle size and nanoplastic type on the accumulation of different PS-NPs. Exposure to NPs significantly increased the production of malondialdehyde, glutathione and reactive oxygen species, as well as the activities of antioxidant enzymes including glutathione reductase, superoxide dismutase and catalase, resulting in various degrees of oxidative damage in sea cucumber. The significant decrease in adenosine triphosphate content and increases in alkaline phosphatase and lactate dehydrogenase activities suggested that NPs impaired energy metabolism and modified their energy allocation. After 20-d exposure, the complex I, II and III activities in mitochondrial respiratory chain were significantly inhibited. Meanwhile, the Bax and Caspase-3 gene expression were significantly up-regulated, and Bacl-2 was down-regulated, indicating the toxicity on mitochondrial pathway of A. japonicus. The calculated IBR values elucidated the greater detriment to mitochondrial pathway than oxidative stress and energy metabolism. For 100 nm particle size, plain PS has stronger influence on all the biomarkers compared to PS-COOH/NH2, however, the opposite trends were observed in 200 nm PS-NPs. Furthermore, 100 nm PS-NPs were recognized to be more hazardous to sea cucumber than 200 nm microbeads. These findings provide new insights for understanding the differentiated toxic effects of functionalized NPs in marine invertebrates.

Adverse effects of dietary virgin (nano)microplastics on growth performance, immune response, and resistance to ammonia stress and pathogen challenge in juvenile sea cucumber Apostichopus japonicus (Selenka).

It has been well documented that micro- and nanoplastics are emerging pollutants in aquatic environments, and their potential toxic effects has attracted widespread concerns. Here, we evaluated the adverse effects of dietary polystyrene nanoplastics and microplastics (PS-N/MPs) on growth performance, oxidative stress induction, immune response, ammonia detoxification, and bacterial pathogen resistance of sea cucumber Apostichopus japonicus. After collection and acclimation, sea cucumbers were randomized into 3 groups (i.e., control, 100 nm PS-NPs and 20 µm PS-MPs at 100 mg kg-1 diet) for 60-day feeding experiment. Every group contained 360 sea cucumbers which were equally divided into 3 aquaria as biological triplicates. The results showed that the specific growth rate and final weight of the sea cucumbers fed with diets containing PS-N/MPs were significantly lower than those of control group. Dietary virgin PS-N/MPs significantly increased the reactive oxygen species production and malondialdehyde content in coelomic fluid, causing oxidative stress and damage to the growth and development of A. japonicus. During the experiment, 100 nm PS-NPs significantly induced the depletion in cellular and humoral immune parameters. The calculated IBR values based on multi-level biomarkers revealed the size-dependent toxic differences of PS-NPs > PS-MPs. The relative expression levels of GDH and GS mRNA showed first rise and then fall trends after exposure to ammonia, and 100 nm PS-NPs had a more profound impact on suppressing ammonia detoxification compared with 20 µm PS-MPs. Moreover, the expression of Hsp90, Hsp70, CL, TLR, and CASP2 genes were all down-regulated by ammonia exposure. Taken together of IBR results, ammonia stress test and pathogen challenge, we deduced that dietary 100 nm PS-NPs are more potentially hazardous than 20 µm PS-MPs. These findings provide valuable information for understanding the size-dependent toxic effects of PS-N/MPs and early risk warning on marine invertebrates.

Amphiphilic hyper-branched co-polymer nanoparticles for the controlled delivery of anti-tumor agents.

In this investigation, we have designed and synthesized an amphiphilic co-polymer with hyper-branched poly(amine-ester) and polylactide (HPAE-co-PLA) to generate nanoparticles (NPs). These have been used to encapsulate a highly active hydrophobic anti-tumor agent, 2-benzoylpyridine 4-ethyl-3-thiosemicarbazone (Bp4eT). Encapsulation in NPs was done in an effort to increase the anti-tumor activity of this agent by facilitating its delivery to tumor cells. We have also examined and optimized the formulation parameters of the NPs that alter their drug-loading capacity and their physical, chemical and biological properties. The resulting NPs exhibited high Bp4eT-loading capacity and substantial stability in aqueous solution. In vitro drug release studies demonstrated a controlled drug release profile with increased release at acidic pH. Anti-tumor proliferation assays showed that both free drug and drug-encapsulated NPs markedly inhibited tumor cell proliferation in a time- and concentration-dependent manner. Direct microscopic observation revealed that the fluorescent NPs were taken up by cells and localized, in part, in organelles consistent with lysosomes. These results demonstrate a feasible application of the amphiphilic hyper-branched co-polymer, HPAE-co-PLA, as nanocarriers for intracellular delivery of potent anti-tumor agents.