Superhydrophilic nanocomposite adsorbents modified via nitrogen-rich phosphonate-functionalized ionic liquid linkers: enhanced phosphopeptide enrichment and phosphoproteome analysis of tau phosphorylation in the hippocampal lysate of Alzheimer's transgenic mice.

In this study, new graphene-based IMAC nanocomposites for phosphopeptide enrichment were prepared according to the guideline of our new design strategy. Superhydrophilic polyethyleneimine (PEI) was introduced, to which a phosphonate-functionalized ionic liquid (PFIL) was covalently bound, to form superhydrophilic and cationic surface layers with high densities of nitrogen atoms, phosphonate functional groups, and high-loading metal ions. Due to the combined features of superhydrophilicity, flexibility, highly dense metal binding sites, large surface area and excellent size-exclusion effect, the fabricated nanocomposite G@mSiO2@PEI-PFIL-Ti4+ exhibits superior detection sensitivity to enrich phosphopeptides (tryptic ß-casein digest, 0.1 fmol), and extraordinary enrichment specificity to enrich phosphopeptides from a digest mixture of ß-casein and bovine serum albumin (BSA) (molar ratio, 1 : 12 000). The excellent size-exclusion effect was also observed, and 27 endogenous phosphopeptides were identified in human saliva. All these results could be attributed to the unique superhydrophilic nanocomposite structure with a high density of a cationic linker modified with phosphonate functionality. Moreover, G@mSiO2@PEI-PFIL-Ti4+ adsorbents were used to extract phosphopeptides from the tryptic digests of hippocampal lysates for quantitative phosphoproteome analysis. The preliminary results indicate that 1649 phosphoproteins, 3286 phosphopeptides and 4075 phosphorylation sites were identified. A total of 13 Alzheimer's disease (AD)-related phosphopeptides within tau proteins were detected with a wide coverage from p-Thr111 to p-Ser404, in which the amounts of some phoshopeptides at certain sites in AD transgenic mice were found statistically higher than those in wild type littermates. Besides, phosphorylated neurofilament heavy chains, a potential biomarker for amyotrophic lateral sclerosis and traumatic brain injury, were also identified. Finally, the adsorbent was applied to human cerebrospinal fluid (CSF) and blood samples. 5 unique phosphopeptides of neuroendocrine specific VGF were identified in the CSF, while many phosphopeptides originated from the nervous system were found in the blood sample. All these results suggest that our new IMAC materials exhibit unbiased enrichment ability with superior detection sensitivity and specificity, allowing the global phosphoproteome analysis of complicated biological samples more convincible and indicating the potential use in disease diagnosis.

One-step preparation of carbonaceous spheres rich in phosphate groups via hydrothermal carbonization for effective phosphopeptides enrichment.

A green strategy was developed to prepare carbonaceous spheres rich in phosphoric acid groups on the surface with D-Glucose 6-phosphate sodium salt (called G6PNa2) as a sole carbon source through one-step hydrothermal carbonization method. The method is simple and facile and meets the standards of green chemistry as water is the sole solvent employed. Following the hydrothermal carbonization synthesis, the carbonaceous spheres were further functionalized with Ti4+. The main factors including reaction temperature, reaction time, and concentration of G6PNa2 were systematically studied in order to obtain the desirable morphology and the optimum phosphopeptides enrichment, for the resulting Ti4+ functionalized carbonaceous spheres (CS-Ti4+). The performance evaluation of the CS-Ti4+ prepared under the optimum conditions demonstrated excellent selectivity (1:1000), low detection limit (1 fmol) and high recovery rate (85%) towards phosphopeptides. Furthermore, 24 low-abundance phosphopeptides were captured from human saliva using CS-Ti4+, indicating its great potential in mass spectrometry-based phosphoproteome studies.

Alendronate-Decorated Nanoparticles as Bone-Targeted Alendronate Carriers for Potential Osteoporosis Treatment.

Osteoporosis is a skeletal disorder characterized by a low bone mass and density. Alendronate (Alen), a second-generation bisphosphonate drug, was indicated as the first-line regimen for the treatment of osteoporosis. However, the use of Alen has been limited due to its low bioavailability and gastrointestinal side effects. Herein, Alen-decorated nanoparticles were prepared through ionic cross-linking between poly (lactic-co-glycolic acid), ß-cyclodextrin-modified chitosan (PLGA-CS-CD), and Alen-modified alginate (ALG-Alen) for Alen loading and bone-targeted delivery. Alen was selected as a therapeutic drug and a bone-targeting ligand. The nanoparticles have negatively charged surfaces, and sustained release of Alen from the nanoparticles can be observed. Cytotoxicity detected using cell counting kit-8 (CCK-8) assay and lactate dehydrogenase release test on MC3T3 cells showed that the nanoparticles had good cytocompatibility. A hemolysis test showed that the hemolysis ratios of nanoparticles were <5%, indicating that the nanoparticles had no significant hemolysis effect. Moreover, the Alen-decorated nanoparticles exhibited enhanced binding affinity to the hydroxyapatite (HAp) disks compared with that of nanoparticles without Alen modification. Thus, the Alen-decorated nanoparticles might be developed as promising bone-targeted carriers for the treatment of osteoporosis.

PLGA-PEG Nanoparticles Facilitate In Vivo Anti-Alzheimer's Effects of Fucoxanthin, a Marine Carotenoid Derived from Edible Brown Algae.

The marine natural product fucoxanthin has been reported previously to produce anti-Alzheimer's disease (AD) neuroprotective effects in vitro and in vivo. Fucoxanthin was also demonstrated to be safe in preclinical and small population clinical studies, but the low bioavailability of fucoxanthin in the central nervous system (CNS) has limited its clinical applications. To overcome this, poly lactic-co-glycolic acid-block-polyethylene glycol loaded fucoxanthin (PLGA-PEG-Fuc) nanoparticles with diameter at around 200 nm and negative charge were synthesized and suggested to penetrate into the CNS. Loaded fucoxanthin could be liberated from PLGA-PEG nanoparticles by sustained released in the physiological environment. PLGA-PEG-Fuc nanoparticles were shown to significantly inhibit the formation of Aß fibrils and oligomers. Moreover, these nanoparticles were taken up by both neurons and microglia, leading to the reduction of Aß oligomers-induced neurotoxicity in vitro. Most importantly, intravenous injection of PLGA-PEG-Fuc nanoparticles prevented cognitive impairments in Aß oligomers-induced AD mice with greater efficacy than free fucoxanthin, possibly via acting on Nrf2 and NF-κB signaling pathways. These results altogether suggest that PLGA-PEG nanoparticles can enhance the bioavailability of fucoxanthin and potentiate its efficacy for the treatment of AD, thus potentially enabling its future use for AD therapy.