Jian-Min Shen

Luminescent/Magnetic Hybrid Nanoparticles with Folate-Conjugated Peptide Composites for Tumor-Targeted Drug Delivery

We developed a novel chitosan-based luminescent/magnetic hybrid nanoparticles with folate-conjugated tetrapeptide composites (CLMNPs-tetrapeptide-FA) by conjugation in situ. First, chitosan, CdTe quantum dots (QDs), and superparamagnetic iron oxide were directly gelled into ternary hybrid nanogels. Subsequently, tetrapeptides (GFFG and LGPV) and folate were conjugated orderly into the hybrid nanoparticles. The morphology, composition, and properties of the as-prepared copolymers have also been characterized and determined using TEM, EDX, XRD, FTIR spectra, DLS, fluorescence spectroscopy, VSM, and fluorescence microscopy imaging studies. The size range of the end product CLMNPs-tetrapeptide-FA copolymers was from 150 to 190 nm under simulated physiological environment. In vivo, the experimental results of magnetic accumulation showed that the copolymers could be trapped in the tumor tissue under magnetic guidance. Under the present experimental conditions, the loading efficiencies of CPT were approximately 8.6 wt % for CLMNPs-GFFG-FA and 1.1 wt % for CLMNPs-LGPV-FA, respectively. The CPT cumulative release under dialysis condition mainly occurred for the first 28 h, and could reach 55% at pH 5.3 and 46% at pH 7.4 from CPT-loaded CLMNPs-GFFG-FA, and 69% at pH 5.3 and 57% at pH 7.4 from CPT-loaded CLMNPs-LGPV-FA within 28 h, respectively. The hemolysis percentages (<2%) and coagulation properties of blank and CPT-loaded copolymers were within the scope of safe values. Compared to free CPT, the CPT-loaded CLMNPs-tetrapeptide-FA copolymers showed specific targeting to A549 cells in vitro. More than 75% viability in L02 cells were seen in CLMNPs-GFFG-FA and CLMNPs-LGPV-FA copolymer concentration of 500 μg/mL, respectively. It was found that the two kinds of copolymers were transported into the A549 cells by a folate-receptor-mediated endocytosis mechanism. These results indicate that the multifunctional CLMNPs-tetrapeptide-FA copolymers possess a moderate CPT loading efficiency, low cytotoxicity, and favorable biocompatibility, and are promising candidates for tumor-targeted drug delivery.

Metabolic signatures of esophageal cancer: NMR-based metabolomics and UHPLC-based focused metabolomics of blood serum

Focused metabolic profiling is a powerful tool for the determination of biomarkers. Here, a more global proton nuclear magnetic resonance ((1)H NMR)-based metabolomic approach coupled with a relative simple ultra high performance liquid chromatography (UHPLC)-based focused metabolomic approach was developed and compared to characterize the systemic metabolic disturbances underlying esophageal cancer (EC) and identify possible early biomarkers for clinical prognosis. Serum metabolic profiling of patients with EC (n=25) and healthy controls (n=25) was performed by using both (1)H NMR and UHPLC, and metabolite identification was achieved by multivariate statistical analysis. Using orthogonal projection to least squares discriminant analysis (OPLS-DA), we could distinguish EC patients from healthy controls. The predictive power of the model derived from the UHPLC-based focused metabolomics performed better in both sensitivity and specificity than the results from the NMR-based metabolomics, suggesting that the focused metabolomic technique may be of advantage in the future for the determination of biomarkers. Moreover, focused metabolic profiling is highly simple, accurate and specific, and should prove equally valuable in metabolomic research applications. A total of nineteen significantly altered metabolites were identified as the potential disease associated biomarkers. Significant changes in lipid metabolism, amino acid metabolism, glycolysis, ketogenesis, tricarboxylic acid (TCA) cycle and energy metabolism were observed in EC patients compared with the healthy controls. These results demonstrated that metabolic profiling of serum could be useful as a screening tool for early EC diagnosis and prognosis, and might enhance our understanding of the mechanisms involved in the tumor progression.

Chitosan-based luminescent/magnetic hybrid nanogels for insulin delivery, cell imaging, and antidiabetic research of dietary supplements

In this work, the chitosan-based luminescent/magnetic (CLM) nanomaterials were synthesized by direct gelation of chitosan, CdTe and superparamagnetic iron oxide into the hybrid nanogels. The morphology, sizes and properties of the nanogels prepared with different chitosan/QD/MNP ratios and under different processing parameters were researched. Fluorescence microscopy, FTIR spectra and TEM images confirmed the success of the preparation of the CLM hybrid nanogels. Spherical CLM hybrid nanogels with appropriate average sizes (<160 nm) were used for insulin loading. The actual loading amount of insulin was approximately 40.1mg/g. Human normal hepatocytes L02 cell line was used to explore the effects of additives, such as mangiferin (MF), (-)-epigallocatechin gallate (EGCG), and (-)-epicatechin gallate (ECG) on the insulin-receptor-mediated cellular uptake using insulin-loaded CLM (ICLM) hybrid nanogels. Above 80% of viability of L02 cells were watched at a nanogels concentration of 500 μg/mL whatever the additives existed or not. The study discovered that the fluorescent signals of the ICLM hybrid nanogels in L02 cells were more intense in the presence of MF, EGCG and ECG in medium than in the absence of these components, respectively. These results demonstrate that MF, EGCG and ECG are potentially able to enhance targeting combination of insulin with L02 cells and improve insulin sensitivity in L02 cells. The hybrid nanogels designed as a targeting carrier can potentially offer an approach for integration of insulin delivery, cell imaging, and antidiabetic investigation of dietary supplements.

Toxicity and biodistribution of aqueous synthesized ZnS and ZnO quantum dots in mice

Abstract In the present study, ZnS and ZnO quantum dots (QDs) were synthesized via an all-aqueous process with polyethylene glycol (PEG) chains on their surface, and their toxicity as well as biodistribution were evaluated. No haemolysis occurred at a high concentration of 1600 µg/mL in vitro haemolytic assay, which demonstrated that the QDs-PEG displayed good blood compatibility. Following intravenous administration at 2, 6, and 20 mg/kg of the QDs-PEG in mice, the biodistribution, excretion and biocompatibility were characterized at 1 h, 24 h and 7 days, respectively. Quantitative analysis results indicated that the biodistribution trend of ZnS QDs-PEG was similar to that of ZnO QDs-PEG. The QDs-PEG were mainly trapped in the lung and liver, and almost removed from blood within 1 h. QDs-PEG were primarily excreted in faeces at the 2 and 6 mg/kg doses. Coefficients, haematology, blood biochemistry and histopathology results indicated that the QDs-PEG were safe and biocompatible.

Graphene oxide–Fe3O4 nanocomposite for combination of dual-drug chemotherapy with photothermal therapy

Multi-modal strategies may enhance anticancer efficacy by overcoming tumor drug resistance due to different anticancer pathways and mechanisms. Herein, a novel method of synthesizing ultra-fine graphene oxide (uGO)–magnetic nanoparticle (MNP) composites is presented. This composite is fabricated by combination of a simple and effective chemical deposition with further oxidation of iron ions on a carboxylated uGO base, followed by coating oleic acid on MNPs. Two anticancer drugs, camptothecin (CPT) and methotrexate (MTX), are separately bound to uGO sheets and the carboxyl terminals of uGO on the hybrid, forming a superparamagnetic & dual drug-loaded MTX@uGO–COOH@MNP@OA@CPT nanocomposite. The size of the composite is approximately 80 nm by DLS. The entrapment efficiencies of MNPs, CPT, and MTX reach approximately 458 mg g−1, 682 mg g−1, and 896 mg g−1, respectively. In vitro release and apoptotic assay results show that the nanocomposite can cause the apoptosis and death of HepG2 cells by preferentially releasing drugs to the tumor microenvironment. The tumor inhibitory rate of 73.9% in S-180 sarcoma-bearing Balb/c mice suggests that the combination of nanocomposite-mediated dual drug synergic chemotherapy with photothermal therapy has remarkable therapeutic potential against drug-resistant tumors.

Hepatotoxicity induced by ZnO quantum dots in mice

ZnO quantum dots (QDs) with unique optical properties are potential useful tools for biological labeling and biosensing. With the increasing use of ZnO QDs, the toxicity evaluation of ZnO QDs is urgent. In this study, the hepatotoxicity including serum aminotransferases (ALT and AST), antioxidant enzymes (CAT, GSH-Px and SOD), lipid peroxidation and ultrastructure were evaluated after consecutive intravenous injection of ZnO QDs and ZnO QDs–PEG for 7 days in mice. Both ZnO QDs and ZnO QDs–PEG did not affect the coefficient of liver and the levels of serum aminotransferases. The antioxidant enzymes and lipid peroxidation had significant change after injecting 5 mg kg−1 ZnO QDs in 24 h, but all of these parameters returned to control levels in 28 days. ZnO QDs–PEG had a less harmful effect on antioxidant enzymes and malondialdehyde than ZnO QDs at the same dose. According to the results of hepatocyte ultrastructure, both ZnO QDs and ZnO QDs–PEG were located in the mitochondrion and induced nuclear malformation in 24 h. The ultrastructure of hepatocyte was as normal as of the control group in 28 days and ZnO QDs were mainly trapped in the mitochondrion while ZnO QDs–PEG mainly accumulated in the lysosomes. These findings would be helpful for wide use of quantum dots based bioimaging and biomedical applications in the future.

Heterogeneous dimer peptide-conjugated polylysine dendrimer-Fe 3 O 4 composite as a novel nanoscale molecular probe for early diagnosis and therapy in hepatocellular carcinoma

A novel nanoscale molecular probe is formulated in order to reduce toxicity and side effects of antitumor drug doxorubicin (DOX) in normal tissues and to enhance the detection sensitivity during early imaging diagnosis. The mechanism involves a specific targeting of Arg-Gly-Asp peptide (RGD)-GX1 heterogeneous dimer peptide-conjugated dendrigraft poly-l-lysine (DGL)–magnetic nanoparticle (MNP) composite by αvβ3-integrin/vasculature endothelium receptor-mediated synergetic effect. The physicochemical properties of the nanoprobe were characterized by using transmission electron microscope, Fourier transform infrared spectroscopy, X-ray diffraction, dynamic light scattering (DLS), and vibrating sample magnetometer. The average diameter of the resulting MNP–DGL–RGD-GX1–DOX nanoparticles (NPs) was ~150−160 nm by DLS under simulate physiological medium. In the present experimental system, the loading amount of DOX on NPs accounted for 414.4 mg/g for MNP–DGL–RGD-GX1–DOX. The results of cytotoxicity, flow cytometry, and cellular uptake consistently indicated that the MNP–DGL–RGD-GX1–DOX NPs were inclined to target HepG2 cells in selected three kinds of cells. In vitro exploration of molecular mechanism revealed that cell apoptosis was associated with the overexpression of Fas protein and the significant activation of caspase-3. In vivo magnetic resonance imaging and biodistribution study showed that the MNP–DGL–RGD-GX1–DOX formulation had high affinity to the tumor tissue, leading to more aggregation of NPs in the tumor. In vivo antitumor efficacy research verified that MNP–DGL–RGD-GX1–DOX NPs possessed significant antitumor activity and the tumor inhibitory rate reached 78.5%. These results suggested that NPs could be promising in application to early diagnosis and therapy in hepatocellular carcinoma as a specific nanoprobe.

Hepatotoxicity assessment of Mn-doped ZnS quantum dots after repeated administration in mice.

Doped ZnS quantum dots (QDs) have a longer dopant emission lifetime and potentially lower cytotoxicity compared to other doped QDs. The liver is the key organ for clearance and detoxification of xenobiotics by phagocytosis and metabolism. The present study was designed to synthesize and evaluate the hepatotoxicity of Mn-doped ZnS QDs and their polyethylene glycol-coated counterparts (1 mg/kg and 5 mg/kg) in mice. The results demonstrated that daily injection of Mn-doped ZnS QDs and polyethylene glycol-coated QDs via tail vein for 7 days did not influence body weight, relative liver weight, serum aminotransferases (alanine aminotransferase and aspartate aminotransferase), the levels of antioxidant enzymes (catalase, glutathione peroxidase, and superoxide dismutase), or malondialdehyde in the liver. Analysis of hepatocyte ultrastructure showed that Mn-doped ZnS QDs and polyethylene glycol-coated QDs mainly accumulated in mitochondria at 24 hours after repeated intravenous injection. No damage to cell nuclei or mitochondria was observed with either of the QDs. Our results indicate that Mn-doped ZnS QDs did not cause obvious damage to the liver. This study will assist in the development of Mn-doped ZnS QDs-based bioimaging and biomedical applications in the future.

Metabolomic investigation of Arthus reaction in a rat model using proton nuclear magnetic resonance (1H NMR) spectroscopy and rapid resolution liquid chromatography (RRLC)

Arthus reaction (AR), a type of unconventional immune complex-mediated inflammation, is likely accompanied by alterations in circulating metabolites. Here, a proton nuclear magnetic resonance ((1)H NMR) spectroscopy method coupled with a rapid resolution liquid chromatography (RRLC) method was developed to evaluate the systemic metabolic consequences of AR and characterize metabolic aberrations. Serum and urine samples from AR rats and normal controls were compared to determine whether there were significant alterations associated with AR. The partial least squares discriminant analysis (PLS-DA) models of metabolomic results demonstrated good intergroup separations between AR rats and normal controls. Multivariate statistical analysis revealed significant alterations in the levels of 34 metabolites, which were termed as the disease-associated biomarkers. Differential metabolites identified from the metabolomic analysis suggested that AR caused dysfunctions of kidney and liver accompanied with changes in widespread metabolic pathways including the tricarboxylic acid (TCA) cycle, gut microbiota metabolism, lipids and cell membranes metabolism, glucose metabolism, fatty acid β-oxidation, amino acids metabolism and ketogenesis. This study assessed and provided important metabolomic variations in serum and urine associated with AR and, therefore, demonstrated metabolomics as a powerful approach for the complete elucidation of the underlying pathophysiologic mechanisms of AR.

pH-Sensitive nanoparticles as smart carriers for selective intracellular drug delivery to tumor

ABSTRACT Herein, a smart pH‐sensitive nanoparticle (DGL‐PEG‐Tat‐KK‐DMA‐DOX) was prepared to achieve the selective intracellular drug delivery. In this nanoparticle, a PEG‐grafted cell penetrating peptide (PEG‐Tat‐KK) was designed and acted as the cell penetrating segment. By introducing the pH‐sensitive amide bonds between the peptide and blocking agent (2,3‐dimethylmaleic anhydride, DMA), the controllable moiety (PEG‐Tat‐KK‐DMA) endowed the nanoparticle with a charge‐switchable shell and temporarily blocked penetrating function, thus improving the specific internalization. Besides, dendrigraft poly‐L‐lysine (DGL) used as the skeleton can greatly improve the drug loading because of the highly dendritic framework. Under the stimuli of acidic pH, this nanoparticle exhibited a remarkable charge‐switchable property. The drug release showed an expected behavior with little release in the neutral pH media but relatively fast release in the acidic media. The in vitro experiments revealed that the cellular uptake and cytotoxicity were significantly enhanced after the pH was decreased. In vivo biodistribution and antitumor research indicated that the nanoparticle had noteworthy specificity and antitumor efficacy with a tumor inhibition rate of 79.7%. These results verified this nanoparticle could efficiently improve the selective intracellular delivery and possessed a great potential in tumor treatment.